WO2010117425A1 - Certain substituted pyrimidines, pharmaceutical compositions thereof, and methods for their use - Google Patents

Certain substituted pyrimidines, pharmaceutical compositions thereof, and methods for their use Download PDF

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WO2010117425A1
WO2010117425A1 PCT/US2010/000958 US2010000958W WO2010117425A1 WO 2010117425 A1 WO2010117425 A1 WO 2010117425A1 US 2010000958 W US2010000958 W US 2010000958W WO 2010117425 A1 WO2010117425 A1 WO 2010117425A1
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optionally substituted
compound
pharmaceutically acceptable
acceptable salt
alkyl
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PCT/US2010/000958
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French (fr)
Inventor
Jiandong Shi
Arthur Taveras
Ryan Van De Water
Ryan Lamer
Kevin Hong
Marco Biamonte
Jianhua Chao
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Biogen Idec Ma Inc.
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Publication of WO2010117425A1 publication Critical patent/WO2010117425A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P19/00—Drugs for skeletal disorders
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D519/00—Heterocyclic compounds containing more than one system of two or more relevant hetero rings condensed among themselves or condensed with a common carbocyclic ring system not provided for in groups C07D453/00 or C07D455/00

Definitions

  • Hsp90s are ubiquitous chaperone proteins that are involved in folding, activation and assembly of a wide range of proteins, including key proteins involved in signal transduction, cell cycle control and transcriptional regulation.
  • Hsp90 chaperone proteins are associated with important signaling proteins, such as steroid hormone receptors and protein kinases, including, e.g., Raf-1 , AKT, KIT, EGFR, v-Src family kinases, Cdk4, and ErbB-2.
  • Hsp70 e.g., Hsp70, p60/Hop/Sti1 , Hip, Bag1 , Hsp40/Hdj2/Hsj1 , immunophilins, p23, and p50, may assist Hsp90 in its function.
  • co-chaperones e.g., Hsp70, p60/Hop/Sti1 , Hip, Bag1 , Hsp40/Hdj2/Hsj1 , immunophilins, p23, and p50, may assist Hsp90 in its function.
  • Hsp90 possesses a binding pocket at its N-terminus. This pocket is highly conserved and has weak homology to the ATP-binding site of DNA gyrase. Further, ATP and ADP have both been shown to bind this pocket with low affinity and to have weak ATPase activity. In vitro and in vivo studies have demonstrated that occupancy of this N-terminal pocket by ansamycins and other Hsp90 inhibitors alters Hsp90 function and inhibits protein folding. At high concentrations, ansamycins and other Hsp90 inhibitors have been shown to prevent binding of protein substrates to Hsp90. Hsp90 inhibitors, e.g. ansamycins, have also been demonstrated to inhibit the ATP-dependent release of chaperone-associated protein substrates. In either event, the substrates are degraded by an ubiquitin-dependent process in the proteasome.
  • Hsp90 is required for the stability, folding, and activity of specific "client” proteins that are involved in tumor cell signaling such as HER-2, ER, EGFR, cKIT, BRaf. Inhibition of Hsp90 results in client protein degradation leading to tumor cell stasis or death.
  • Hsp90 inhibitors described herein can be used to inhibit multiple tumor signaling pathways, enhance the activity of these agents and lead to greater antitumor activity.
  • Hsp90 inhibitors have also been implicated in a wide variety of other utilities, such as, anti-inflammation agents, anti-infectious disease agents, agents for treating autoimmunity, agents for treating stroke, ischemia, multiple sclerosis, cardiac disorders, central nervous system related disorders and agents useful in promoting nerve regeneration. Overlapping somewhat with the above, there are reports in the literature that fibrogenic disorders also may be treatable with Hsp90 inhibitors.
  • Fibrogenic disorders include, but are not limited to, connective tissue diseases, such as scleroderma (or systemic sclerosis), polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders, including liver cirrhosis, keloid formation, interstitial nephritis, and pulmonary fibrosis.
  • connective tissue diseases such as scleroderma (or systemic sclerosis), polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders, including liver cirrhosis, keloid formation, interstitial nephritis, and pulmonary fibrosis.
  • Medications for the treatment of systemic lupus erythematosis are limited, and no new medication for SLE has been approved in the past 30 years.
  • the mainstay of therapy continues to be corticosteroids for flares of moderate to severe disease.
  • Nonsteroidal anti-inflammatory drugs NSAIDs
  • antimalarials corticosteroids
  • other immunosuppressive agents such as cyclophosphamide, azathioprine or 6-mercaptopurine, methotrexate (MTX), and mycophenolate mofetil (MMF) are used alone or in combination for the many manifestations of this disease.
  • MTX methotrexate
  • MMF mycophenolate mofetil
  • Azathioprine, cyclophosphamide, MTX, and MMF are common immunosuppressives for subsets of SLE patients exhibiting chronic active or relapsing disease activity who are not responsive to antimalarials and prednisone alone.
  • belimumab anti-BAFF monoclonal antibody
  • RA rheumatoid arthritis
  • NSAIDs non-steroidal anti-inflammatory drugs
  • glucocorticoids glucocorticoids
  • DMARDs disease modifying anti-rheumatic drugs
  • MTX methotrexate
  • newer biologic agents that target specific pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF- ⁇ ) and interleukin-1 (IL-1) or anti-CD20 and antagonists of CD80/CD86.
  • NSAIDs treat inflammation and pain but do not modify the course of disease and are associated with well known complications (gastrointestinal ulceration, exacerbation of hypertension, headache and depression, and exacerbation of underlying renal disease).
  • Glucocorticoids treat acute symptomatic joint flares, inflammation, and pain when NSAIDs alone are inadequate.
  • DMARDs are the next line of treatment (sulfasalazine, hydroxychloroquine, chloroquine, gold salts, D-penicillamine, and MTX).
  • MTX in combination with NSAIDs has become the mainstay of initial DMARD therapy because MTX has proven efficacy, treats synovitis, and slows progression of structural damage.
  • many patients either have an inadequate response to initial MTX therapy or lose responsiveness over time.
  • biologic agents have been shown to be beneficial.
  • examples include etanercept (soluble TNF receptor fusion protein), infliximab (chimeric anti-TNF- ⁇ ), adalimumab (humanized anti-TNF- ⁇ ), rituximab (chimeric anti-CD20), abatacept (human CTLA4-Fc) and tocilizumab (humanized anti-IL-6R).
  • etanercept soluble TNF receptor fusion protein
  • infliximab chimeric anti-TNF- ⁇
  • adalimumab humanized anti-TNF- ⁇
  • rituximab chimeric anti-CD20
  • abatacept human CTLA4-Fc
  • tocilizumab humanized anti-IL-6R
  • R 1 is selected from hydrogen, halogen, -OR 11 , -SR 12 , amino, and optionally substituted lower alkyl;
  • R 5 and R 6 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; or
  • R 7 is selected from a bond, optionally substituted alkylene, -C(O)-, -C(S)-, -S(O)-, and -SO 2 -;
  • R 8 is selected from alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which is optionally substituted;
  • R 11 and R 12 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted;
  • R 21 and R 22 are independently selected from hydrogen, alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl, each of which, except for hydrogen, is optionally substituted, or
  • R 23 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
  • R 24 is selected from alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
  • R 90 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and wherein from 1 to 5 hydrogen atoms in the compound of Formula I are optionally replaced with deuterium, and provided that when X is an optionally substituted alkylene chain of the formula (CR a R b ) n , n is 1 , the (CR a R b ) unit is replaced by -N(R a )-, and R 7 is -CH 2 -, then R 8 is not 3,5-dimethyl-4-methoxy-pyridin-2- yi- [0017] Also provided is a pharmaceutical composition comprising one or more pharmaceutically acceptable excipients and at least one compound or pharmaceutically acceptable salt thereof described herein.
  • Also provided is a method of treating an individual having an Hsp90 mediated disorder comprising administering to said individual at least one compound or pharmaceutically acceptable salt described herein or a pharmaceutical composition described herein.
  • Figure 1 shows the effect of Compound A on client proteins in MCF7 cells.
  • Figure 2 shows the effect of Compound A on client proteins in N87 cells.
  • Figure 3 shows that Compound A down modulates CD4 expression on human blood T cells.
  • Figure 4 shows that in a mouse macrophage cell line Compound A decreased phosphorylation of MEK1/2, ERK1/2, and JNK1/2 in response to LPS.
  • Figure 5 summarizes Compound A tumor efficacy studies.
  • Figure 6 shows the antitumor activity of Compound A in the high HER-2 N87 gastric carcinoma xenograft model.
  • Figure 7 shows the antitumor activity of Compound A in the high HER-2 BT474 breast carcinoma xenograft model.
  • Figure 8 shows the antitumor activity of Compound A in the HT29 colon carcinoma xenograft model.
  • FIG. 9 shows that Compound A inhibits LPS-induced systemic TNF- ⁇ release in DBA/1 mice.
  • Figure 10 shows the efficacy of orally administered Compound A in a rat collagen- induced arthritis model.
  • a "pharmaceutically acceptable salt” may be prepared for any compound having a functionality capable of forming a salt, for example, an acid or base functionality.
  • Pharmaceutically acceptable salts may be derived from organic or inorganic acids and bases.
  • Compounds that contain one or more basic functional groups, e.g., amino or alkylamino, are capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable organic and inorganic acids.
  • These salts can be prepared in situ during the final isolation and purification of the compounds, or by separately reacting a purified compound in its free base form with a suitable organic or inorganic acid, and isolating the salt thus formed.
  • Suitable acid salts include acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2- hydroxyethanesulfonate, lactate, maleate, malonate, methanesulfonate, 2-napthalenesulfonate, nicotinate, nitrate, oxalate, palmoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate,
  • compositions that contain one or more acidic functional groups are capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable bases.
  • pharmaceutically acceptable salts refers to the relatively non-toxic, inorganic and organic base addition salts of compounds described herein. These salts can likewise be prepared in situ during the final isolation and purification of the compounds, or by separately reacting the purified compound in its free acid form with a suitable base, such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary or tertiary amine.
  • suitable base such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary or tertiary amine.
  • alkali or alkaline earth salts include the lithium, sodium, potassium, calcium, magnesium, and aluminum salts and the like.
  • Illustrative examples of some of the bases that can be used include sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate, N + (C ⁇ alkyl) 4 , and the like.
  • Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine and the like.
  • salts wherein one or more basic nitrogen-containing groups are quarternized. Water or oil-soluble or dispersible products may be obtained by such quaternization. See, for example, Berge et al., supra.
  • alkyl refers to an optionally substituted straight- chain, or optionally substituted branched-chain saturated hydrocarbon radical having from one to thirty carbons, such as from one to twelve carbons.
  • alkyl radicals include methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, tert-butyl, tert-amyl, pentyl, hexyl, heptyl, octyl and the like.
  • a "lower alkyl” is an alkyl having from one to six carbons.
  • alkylene refers to an optionally substituted straight-chain, or optionally substituted branched-chain saturated hydrocarbon radical having from one to thirty carbons, such as from one to twelve carbons, and having two points of attachment. Alkylene groups will usually have from 1 to 20 carbon atoms, for example 1 to 8 carbon atoms, such as from 1 to 6 carbon atoms. For example, C 1 alkylene is a methylene group (-CH 2 -).
  • a "lower alkylene” is an alkylene having from one to six carbons.
  • alkenyl refers to an optionally substituted straight- chain, or optionally substituted branched-chain hydrocarbon radical having one or more carbon-carbon double-bonds and having from two to thirty carbon atoms, such as two to eighteen carbons.
  • alkenyl radicals include ethenyl, propenyl, butenyl, 1 ,3-butadienyl and the like.
  • a “lower alkenyl” refers to an alkenyl having from two to six carbons.
  • alkynyl refers to an optionally substituted straight- chain or optionally substituted branched-chain hydrocarbon radical having one or more carbon-carbon triple-bonds and having from two to thirty carbon atoms, such as from two to twelve carbon atoms, from two to six carbon atoms as well as those having from two to four carbon atoms.
  • alkynyl radicals include ethynyl, 2-propynyl, 2-butynyl, 1 ,3-butadiynyl and the like.
  • a “lower alkynyl” refers to an alkynyl having from two to six carbons.
  • carbon chain embraces any alkyl, alkenyl, or alkynyl group, which is linear, cyclic, or any combination thereof. If the chain is part of a linker and that linker comprises one or more rings as part of the core backbone, for purposes of calculating chain length, the "chain” only includes those carbon atoms that compose the bottom or top of a given ring and not both, and where the top and bottom of the ring(s) are not equivalent in length, the shorter distance shall be used in determining the chain length.
  • membered ring can embrace any cyclic structure, including aryl, heteroaryl, alicyclic, heterocyclic and polycyclic fused ring systems as described below.
  • membered is meant to denote the number of skeletal atoms that constitute the ring.
  • pyridine, pyran, and pyhmidine are six-membered rings and pyrrole, tetrahydrofuran, and thiophene are five-membered rings.
  • optionally substituted ring can embrace any any cyclic structure, including optionally substituted aryl, optionally substituted heteroaryl, optionally substituted alicyclic, optionally substituted heterocyclic and optionally substituted polycyclic fused ring systems as described below.
  • aryl alone or in combination, refers to an optionally substituted aromatic hydrocarbon radical of six to twenty ring atoms, and includes mono-aromatic rings and fused aromatic rings.
  • a fused aromatic ring radical contains from two to four fused rings where the ring of attachment is an aromatic ring, and the other individual rings within the fused ring may be aromatic, heteroaromatic, alicyclic or heterocyclic.
  • aryl includes mono-aromatic ring and fused aromatic rings containing from six to twelve carbon atoms, as well as those containing from six to ten carbon atoms.
  • aryl groups include, without limitation, phenyl, naphthyl, anthryl, chrysenyl, and benzopyrenyl ring systems.
  • heteroaryl refers to optionally substituted aromatic radicals containing from five to twenty skeletal ring atoms and where one or more of the ring atoms is a heteroatom such as, for example, oxygen, nitrogen, sulfur, selenium and phosphorus.
  • heteroaryl includes optionally substituted mono-heteroaryl radicals and fused heteroaryl radicals having at least one heteroatom (e.g., quinoline, benzothiazole).
  • a fused heteroaryl radical may contain from two to four fused rings and where the ring of attachment is a heteroaromatic ring, the other individual rings within the fused ring system may be aromatic, heteroaromatic, alicyclic or heterocyclic.
  • heteroaryl also includes mono- heteroaryls or fused heteroaryls having from five to twelve skeletal ring atoms, as well as those having from five to ten skeletal ring atoms.
  • heteroaryls include, without limitation, furanyl, benzofuranyl, chromenyl, pyridyl, pyrrolyl, indolyl, quinolinyl, pyridyl-N-oxide, pyrimidyl, pyrazinyl, imidazolyl, pyrazolyl, oxazolyl, isoxazolyl, benzothiozolyl, benzimidazolyl, benzoxazolyl, benzothiadiazolyl, benzoxadiazolyl, benzotriazolyl, quinolinyl, isoquinolinyl, indolyl, purinyl, indolizinyl, thienyl and the like and their oxides.
  • alicyclic alone or in combination, refers to an optionally substituted saturated or unsaturated nonaromatic hydrocarbon ring system, including both monocyclic and multicyclic ring systems, including fused ring systems, containing from three to twenty ring atoms.
  • Monocyclic alicylic groups typically have from 3 to about 8 carbon ring atoms or from 3 to about 7 carbon ring atoms.
  • Multicyclic alicyclic groups may have 2 or 3 fused rings or contain bridged or caged groups.
  • a fused alicyclic radical contains from two to four fused rings where the ring of attachment is an alicyclic ring, and the other individual rings within the fused ring may be aromatic, heteroaromatic, alicyclic or heterocyclic.
  • Alicyclic substituents may be pendant from a substituted nitrogen or carbon atom, or a substituted carbon atom that may have two substituents may have an alicyclic group, which is attached as a spiro group.
  • alicylic groups include cyclopropyl, cyclopropenyl, cyclobutyl, cyclopentyl, or cyclohexyl as well as bridged or caged saturated ring groups such as norbornyl or adamantyl.
  • heterocyclic refers to optionally substituted saturated or unsaturated nonaromatic ring radicals containing from five to twenty ring atoms where one or more of the ring atoms are heteroatoms such as, for example, oxygen, nitrogen, sulfur, and phosphorus.
  • heterocyclic includes mono-heterocyclic and fused heterocyclic ring radicals, as well as spiro-heterocyclic ring radicals.
  • a fused heterocyclic radical may contain from two to four fused rings where the attaching ring is a heterocyclic, and the other individual rings within the fused heterocyclic radical may be aromatic, heteroaromatic, alicyclic or heterocyclic.
  • heterocyclic also includes mono-heterocyclic and fused heterocyclic radicals having from five to twelve skeletal ring atoms, as well as those having from five to ten skeletal ring atoms.
  • heterocyclics include without limitation, tetrahydrofuranyl, benzodiazepinyl, tetrahydroindazolyl, dihydroquinolinyl, and the like.
  • arylalkyl refers to an alkyl radical as defined above in which one H atom is replaced by an aryl radical as defined above, each of which may be optionally substituted.
  • arylalkyl groups include benzyl, 2-phenylethyl and the like.
  • heteroarylalkyl refers to an alkyl radical as defined above in which one H atom is replaced by a heteroaryl radical as defined above, each of which may be optionally substituted.
  • alicyclicalkyl refers to an alkyl radical as defined above in which one H atom is replaced by an alicyclic radical as defined above, each of which may be optionally substituted.
  • heterocyclicalkyl refers to an alkyl radical as defined above in which one H atom is replaced by a heterocyclic radical as defined above, each of which may be optionally substituted.
  • alkoxy alone or in combination, refers to an alkyl ether radical, alkyl-O-, wherein the term alkyl is defined as above and wherein the alkyl group may be optionally substituted.
  • alkoxy radicals include methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, iso-butoxy, sec- butoxy, tert-butoxy and the like.
  • a "lower alkoxy” is an alkoxy having from one to six carbons.
  • aryloxy refers to an aryl ether radical wherein the term aryl is defined as above and wherein the aryl group may be optionally substituted.
  • aryloxy radicals include phenoxy and the like.
  • alkylthio alone or in combination, refers to an alkyl thio radical, alkyl-S-, wherein the term alkyl is as defined above.
  • arylthio refers to an aryl thio radical, aryl-S-, wherein the term aryl is as defined above.
  • heteroarylthio refers to the group heteroaryl-S-, wherein the term heteroaryl is as defined above.
  • acyl refers to a radical -C(O)R where R includes alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl or heteroarylalkyl wherein the alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl or heteroarylalkyl groups may be optionally substituted.
  • acyloxy refers to the ester group -OC(O)R, where R is H, alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl, or heteroarylalkyl wherein the alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl or heteroarylalkyl may be optionally substituted.
  • carboxymers refers to -C(O)OR where R is alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, heteroarylalkyl or arylalkyl, wherein each is optionally substituted.
  • BOC refers to -C(O)Otbutyl
  • R and R' are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl and heteroarylalkyl, each of which, other than the hydrogen, being optionally substituted.
  • halogen refers to F, Cl, Br and I.
  • haloalkyl, haloalkenyl, haloalkynyl and haloalkyloxy include alkyl, alkenyl, alkynyl and alkoxy structures, as described above, that are substituted with one or more fluorines, chlorines, bromines or iodines, or with combinations thereof.
  • perhaloalkyl, perhaloalkyloxy and perhaloacyl refer to alkyl, alkyloxy and acyl radicals as described above, wherein all the H atoms are substituted with fluorines, chlorines, bromines or iodines, or combinations thereof.
  • Perhaloalkyl, perhaloalkyloxy,” and perhaloacyl are species, respectively, of "haloalkyl,” “haloalkyloxy,” and “haloacyl”.
  • amino refers to the group -NH 2 .
  • alkylamino refers to the group -NHR where R is independently selected from alkyl, arylalkyl, and heteroarylalkyl.
  • dialkylamino refers to the group -NRR' where R and R' are independent alkyl, arylalkyl, and heteroarylalkyl.
  • aminocarbonyl refers to the group -C(O)NRR' where R and R' are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, alicyclic, heteroaryl, heterocyclic, hydroxy, alkoxy, amino, alkylamino, dialkylamino, and carboxamido, and where R and R' are optionally joined together with the nitrogen bound thereto to form a heterocyclic or substituted heterocyclic group.
  • R and R' are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, alicyclic, heteroaryl, heterocyclic, hydroxy, alkoxy, amino, alkylamino, dialkylamino, and carboxamido, and where R and R' are optionally joined together with the nitrogen bound thereto to form a heterocyclic or substituted heterocyclic group.
  • R and R' are independently selected from hydrogen, alkyl, alkenyl, alkynyl, ary
  • aryl optionally mono- or di- substituted with an alkyl means that the alkyl may but need not be present, or either one alkyl or two may be present, and the description includes situations where the aryl is substituted with one or two alkyls and situations where the aryl is not substituted with an alkyl.
  • Optionally substituted groups may be substituted or unsubstituted.
  • optionally substituted alkyl, alkylene, alkenyl and alkynyl refer to alkyl, alkylene, alkenyl or alkynyl radicals, as defined herein, that may be optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) independently selected from alkyl, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted by one, two, or three groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three alkyl groups,
  • ), -C( E)N(R,)S(O) 2 R
  • ), -C ⁇ N, -C( E)OR,, and halogen, wherein each E is independently O, NR 9 or S; each R 9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, wherein each E is independently O,
  • optionally substituted aryl refers to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted by one, two, or three groups independently chosen from alkoxy and hydroxy, hetero
  • optionally substituted aryl refers to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, heteroaryl optional
  • aryl including “optionally substituted phenyl”, “optionally substituted alicyclic”, “optionally substituted heteroaryl”, and “optionally substituted heterocyclic” refer to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, heteroaryl optional
  • aryl including “optionally substituted phenyl”, “optionally substituted alicyclic”, “optionally substituted heteroaryl”, and “optionally substituted heterocyclic” refer to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three al
  • substitution can occur on any atom of the alkyl, alkylene, alkenyl, alkynyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, alicyclic, alicyclicalkyl, heterocyclic, and heterocyclicalkyl groups. It will be understood by those skilled in the art, with respect to any group containing one or more substituents, that such groups are not intended to introduce any substitution or substitution patterns that are sterically impractical, synthetically non-feasible and/or inherently unstable.
  • Optionally substituted alicyclic and optionally substituted heterocyclic may additionally be substituted with oxo, thiono, imino, oxime or hydrazone, on a saturated carbon of their respective ring system.
  • Optionally substituted heteroaryl wherein the heteroatom(s) is/are nitrogen may additionally be substituted by oxo on the nitrogen atom of the ring system.
  • substituted amino refers to the group -NHR d or -NR d R e wherein R d is selected from hydroxy, optionally substituted alkoxy, optionally substituted alkyl, optionally substituted alicyclic, optionally substituted acyl, optionally substituted carbamoyl, aminocarbonyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclic, optionally substituted alkoxycarbonyl, and sulfonyl, and R e is selected from optionally substituted alkyl, optionally substituted alicyclic, optionally substituted aryl, optionally substituted heteroaryl, and optionally substituted heterocyclic, and wherein substituted alkyl, substituted alicyclic, substituted aryl, substituted heteroaryl, and substituted heterocyclic are as described herein.
  • substituted amino also refers to N-oxides of the groups -NHR d , and NR d R e each as described above.
  • N-oxides can be prepared by treatment of the corresponding amino group with, for example, hydrogen peroxide or m-chloroperoxybenzoic acid.
  • the person skilled in the art is familiar with reaction conditions for carrying out the N-oxidation.
  • pyridine-1 -oxy also means "pyhdine-N-oxy.”
  • aminocarbonyl refers to the group -CONR b R c , where R b is selected from hydrogen, optionally substituted alkyl, optionally substituted alicyclic, optionally substituted heterocyclic, optionally substituted aryl, and optionally substituted heteroaryl; and R c is independently selected from hydrogen and optionally substituted alkyl; or R b and R c taken together with the nitrogen to which they are bound, form an optionally substituted 5- to 7-membered nitrogen-containing heterocyclic which optionally includes 1 or 2 additional heteroatoms selected from O, N, and S in the heterocyclic; wherein substituted alkyl, substituted alicyclic, substituted heterocyclic, substituted aryl, and substituted heteroaryl are as described herein.
  • Some of the compounds described herein may contain one or more chiral centers and therefore may exist in enantiomeric and diastereomeric forms.
  • the term "compound” is intended to cover all isomers per se, as well as mixtures of cis and trans isomers, mixtures of diastereomers and racemic mixtures of enantiomers (optical isomers) as well. Further, it is possible using well known techniques to separate the various forms, and some embodiments described herein may feature purified or enriched species of a given enantiomer or diastereomer.
  • a "pharmaceutical composition” refers to a mixture of one or more of the compounds or pharmaceutically acceptable salts thereof, described herein, with one or more pharmaceutically acceptable carriers.
  • the purpose of a pharmaceutical composition is to facilitate administration of a compound to an organism.
  • pharmaceutically acceptable carrier means a pharmaceutically-acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting the subject agent from one organ, or portion of the body, to another organ, or portion of the body.
  • a pharmaceutically-acceptable material such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting the subject agent from one organ, or portion of the body, to another organ, or portion of the body.
  • Each carrier must be “acceptable” in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient.
  • materials which can serve as pharmaceutically-acceptable carriers include: (1) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydrox
  • sulfonyl refers to the groups: -S(O 2 )-OH, -S(O 2 )-(optionally substituted alkyl), -
  • a "therapeutically effective amount” means an amount which is capable of providing a therapeutic effect.
  • the specific dose of substance administered to obtain therapeutic effect will, of course, be determined by the particular circumstances surrounding the case, including, for example, the specific substance administered, the route of administration, the condition being treated, and the individual being treated.
  • a typical daily dose (administered in single or divided doses) will contain a dosage level of from about 0.01 mg/kg to about 50-100 mg/kg of body weight of the active substance.
  • daily doses generally will be from about 0.05 mg/kg to about 20 mg/kg and ideally from about 0.1 mg/kg to about 10 mg/kg.
  • Factors such as clearance rate, half-life and maximum tolerated dose (MTD) have yet to be determined but one of ordinary skill in the art can determine these using standard procedures.
  • the therapeutic effect is the inhibition, to some extent, of the growth of cells characteristic of a proliferative disorder, e.g., a cancer.
  • a therapeutic effect will also normally, but need not, relieve to some extent one or more of the symptoms other than cell growth or size of cell mass.
  • a therapeutic effect may include, for example, one or more of 1) a reduction in the number of cells; 2) a reduction in cell size; 3) inhibition (i.e., slowing to some extent, or even stopping) of cell infiltration into peripheral organs, e.g., in the instance of cancer metastasis; 4) inhibition (i.e., slowing to some extent, or even stopping) of tumor metastasis; 5) inhibition, to some extent, of cell growth; and/or 6) relieving to some extent one or more of the symptoms associated with the disorder.
  • the therapeutic effect is the decrease, to some extent, in the metabolic rate of the tumor, as measured, e.g,. by 18 F-glucose PET scan.
  • IC 50 refers to an amount, concentration or dosage of a particular test compound that achieves a 50% inhibition of a maximal response in an assay that measures such response.
  • the "IC 50 " value of a compound can be greater for normal cells than for cells exhibiting a proliferative disorder, e.g., cancer cells. The value depends on the assay used.
  • a "standard” is meant a positive or negative control.
  • a negative control in the context of HER2 expression levels is, e.g., a sample possessing an amount of HER2 protein that correlates with a normal cell.
  • a negative control may also include a sample that contains no HER2 protein.
  • a positive control does contain HER2 protein, for example, of an amount that correlates with overexpression as found in proliferative disorders, e.g., cancers.
  • the controls may be from cell or tissue samples, or else contain purified ligand (or absent ligand), immobilized or otherwise. In some embodiments, one or more of the controls may be in the form of a diagnostic "dipstick.”
  • selective targeting is meant affecting one type of cell to a greater extent than another, e.g., in the case of cells with high as opposed to relatively low or normal HER2 levels.
  • R 1 is selected from hydrogen, halogen, -OR 11 , -SR 12 , amino, and optionally substituted lower alkyl; or
  • R 5 and R 6 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; or
  • R 7 is selected from a bond, optionally substituted alkylene, -C(O)-, -C(S)-, -S(O)-, and -SO 2 -;
  • R 8 is selected from alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which is optionally substituted;
  • R 11 and R 12 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted;
  • R 23 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
  • R 24 is selected from alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
  • R 90 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and wherein from 1 to 5 hydrogen atoms in the compound of Formula I are optionally replaced with deuterium, and provided that when X is an optionally substituted alkylene chain of the formula (CR a R b ) n , n is 1 , the (CR a R b ) unit is replaced by -N(R 3 )-, and R 7 is -CH 2 - then R 8 is not 3,5-dimethyl-4-methoxy-pyridin-2-yl.
  • X is selected from -O-, CH 2 , -CH 2 N(R a )-, and -N(R a )-, or X is a bond.
  • R 1 is halogen selected from fluoro, chloro, and bromo. In some embodiments, R 1 is chloro.
  • X is a bond and R 1 and R 3 , taken together with the atoms to which they are attached, form an optionally substituted ring of 3-8 ring atoms wherein one of the 3-8 ring atoms optionally is O or S.
  • R 21 is hydrogen
  • R 22 is hydrogen
  • R 3 is selected from hydrogen, optionally substituted alkyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, and halogen.
  • R 3 is lower alkyl. In some embodiments, R 3 is methyl. In some embodiments, R 3 is f-butyl.
  • R 3 is alkyl substituted with one or two groups selected from
  • R 31 is selected from hydrogen, alkyl, trialkylsilyl, and -C(O)R 40 wherein R 40 is selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl, and heteroaryl
  • -S(O) n R 32 or -OS(O) 2 R 32 wherein R 32 is selected from alkyl, phenyl, and -NR 51 R 52 wherein R 51 and R 52 are independently selected from hydrogen and alkyl, and n is 1 or 2
  • -NR 33 R 34 wherein R 33 and R 34 are independently selected from hydrogen, lower alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, -C(O)R 60 wherein R 60 is selected from hydrogen, alkyl, aryl, and amino, and -S(O) 2 -alkyl or R 33 and R ,34 taken together with the nitrogen to which they are bound form an optionally substituted heterocyclic or optionally substituted heteroaryl ring; -
  • R 3 is alkyl substituted with one or two groups selected from
  • R 31 is selected from hydrogen, alkyl, and -C(O)R 40 wherein R 40 is selected from hydrogen, alkyl, and aryl;
  • -OS(O) 2 R 32 wherein R 32 is selected from alkyl and phenyl; -NR 33 R 34 wherein R 33 and R 34 are independently selected from hydrogen, lower alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, -C(O)R 60 wherein R 60 is selected from hydrogen, alkyl, aryl, and amino, and -S(O) 2 -alkyl or R 33 and R 34 taken together with the nitrogen to which they are bound form an optionally substituted heterocyclic or optionally substituted heteroaryl ring; -C(O)YR 35 wherein Y is selected from -NR 36 and O and wherein R 35 and R 36 are independently selected from hydrogen, alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, alicyclic, heterocyclic, aryl, and heteroaryl; and azido.
  • R 3 is selected from -CH 2 -OH, -CH 2 OCH 3 , -CH 2 CH 2 -OH, -
  • R 3 is alkyl substituted with -NR 33 R 34 wherein R 33 and R 34 optionally taken together with the nitrogen to which they are bound form an optionally substituted heterocyclic or optionally substituted heteroaryl ring selected from isoindolinyl, triazolyl, and piperidinyl.
  • the optional substitutents on the isoindolinyl or triazolyl rings are independently selected from oxo and -CH 2 N(CH 3 J 2 .
  • R 3 is alkyl substituted with -C(O)YR 35 wherein Y is -NR 36 and wherein R 35 and R 36 are independently selected from hydrogen and alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy. [00103] In some embodiments, R 3 is selected from -CH 2 C(O)NH 2 , -CH 2 C(O)NHCH 3 ,
  • R 3 is alkyl substituted with -C(O)YR 35 wherein Y is -NR 36 and wherein R 35 and R 36 join together to form an optionally substituted heterocyclic ring.
  • the optionally substituted heterocyclic ring is selected from morpholinyl, azetidinyl, pyrrolidinyl, and piperidinyl, each of which is optionally substituted.
  • the substituents on the morpholinyl, azetidinyl, pyrrolidinyl, and piperidinyl rings are selected from -CH 2 OH, -
  • R 3 is alkyl substituted -C(O)YR 35 wherein Y is -NR 36 and wherein
  • R and R are independently selected from hydrogen, optionally substituted alicyclic ring, and optionally substituted heterocyclic ring.
  • R 35 and R 36 are independently selected from hydrogen, optionally substituted cyclopropanyl, and optionally substituted oxetanyl.
  • R 3 is selected from optionally substituted phenyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, andoptionally substituted heteroarylalkyl.
  • R is selected from:
  • R 4 is selected from hydrogen and lower alkyl. In some embodiments, R 4 is hydrogen.
  • R 4 taken together with the carbon to which they are attached, form a piperidinyl ring optionally substituted with one or two groups selected from acyl and arylalkyl.
  • R 3 and R 4 taken together with the carbon to which they are attached, form a piperidinyl ring substituted with -C(O)CH 3 or benzyl.
  • R 3 and R 4 taken together with the carbon to which they are attached, form a piperidinyl ring wherein the nitrogen of the piperidinyl ring is substituted with -C(O)CH 3 or benzyl.
  • R 5 and R 6 are independently selected from hydrogen and optionally substituted lower alkyl.
  • R 5 is hydrogen.
  • R 5 is -
  • R 6 is hydrogen
  • R 7 is a bond or optionally substituted alkylene. In some embodiments, R 7 is optionally substituted lower alkylene. In some embodiments, R 7 is -CH 2 -. In some embodiments, R 7 is -CH(CH 3 )-.
  • R 8 is selected from alicyclic, aryl, heterocyclic, and heteroaryl, each of which is optionally substituted.
  • R 8 is phenyl, pyridinyl, pyrimidinyl, pyridazinyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, benzothiazolyl, pyrazolyl, thiazolyl, tetrahydroisoquinolinyl, imidazolyl, or benzodioxolyl, each of which is optionally substituted.
  • R 8 is selected from:
  • R 8 is selected from:
  • R is
  • R 8 is optionally substituted lower alkyl.
  • R 8 from 1 to 5 hydrogen atoms of R 8 are optionally replaced with
  • R 1 is chloro;
  • R 2 is -NH 2 ;
  • R 3 is optionally substituted alkyl;
  • R 4 is hydrogen;
  • R 5 and R 6 are hydrogen;
  • R 7 is -CH 2 -;
  • R 8 is selected from optionally substituted aryl and optionally substituted heteroaryl.
  • the stereocenter to which R 3 and R 4 are attached is of the S- configuration.
  • the stereocenter to which R 3 and R 4 are attached is of the R- configuration.
  • the stereocenter to which R 5 and R 6 are attached is of the S- configu ration.
  • the stereocenter to which R 5 and R 6 are attached is of the R- configuration.
  • R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , and R 8 are as described for compounds of Formula I.
  • R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , and R 8 are as described for compounds of Formula I.
  • R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , and R 8 are as described for compounds of Formula I.
  • the compounds and pharmaceutically acceptable salts described herein may be synthesized by various methods known in the art. See, e.g., U.S. Patent No. 7,138,402 and WO 2008/093075, each of which is incorporated herein by reference to the extent that it refers to preparation of compounds described therein.
  • the chemical reagents are available commercially, e.g., from Aldrich Chemical Co., Milwaukee, Wis., USA. Otherwise their preparation is facile and known to one of ordinary skill in the art, or it is referenced or described herein.
  • conventional protecting groups may be necessary to prevent certain functional groups from undergoing undesired reactions.
  • Suitable protecting groups for various functional groups as well as suitable conditions for protecting and deprotecting particular functional groups are well known in the art. For example, numerous protecting groups are described in T. W. Greene and P. G. M. Wuts, Protecting Groups in Organic Synthesis, Third Edition, Wiley, New York, 1999, and references cited therein.
  • stereoisomers i.e., as individual enantiomers or diastereomers, or as stereoisomer-enriched mixtures. All such stereoisomers (and enriched mixtures) are included within the scope of this invention, unless otherwise indicated. Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active starting materials or stereoselective reagents well-known in the art. Alternatively, racemic mixtures of such compounds can be separated using, for example, chiral column chromatography, chiral resolving agents and the like.
  • Compounds of Formula I also include crystalline and amorphous forms of those compounds, including, for example, polymorphs, pseudopolymorphs, solvates (including hydrates), unsolvated polymorphs (including anhydrates), conformational polymorphs, and amorphous forms of the compounds, as well as mixtures thereof.
  • Crystal form may be used interchangeably herein, and are meant to include all crystalline and amorphous forms of the compound, including, for example, polymorphs, pseudopolymorphs, solvates (including hydrates), unsolvated polymorphs (including anhydrates), conformational polymorphs, and amorphous forms, as well as mixtures thereof, unless a particular crystalline or amorphous form is referred to.
  • pharmaceutically acceptable salts of compounds of Formula I also include crystalline and amorphous forms of those salts, including, for example, polymorphs, pseudopolymorphs, solvates (including hydrates), unsolvated polymorphs (including anhydrates), conformational polymorphs, and amorphous forms of the salts, as well as mixtures thereof.
  • crystalline forms may be prepared using methods known to the skilled artisan.
  • solvates may be prepared using standard low molecular weight solvents using methods known to the skilled artisan. See, e.g., Polymorphism in Pharmaceutical Solids, ed. Harry G. Brittan, Vol. 95, Marcel Dekker, Inc., New York, 1999 and more particularly, pages 202-208 which describe how hydrates and solvates of both organic and inorganic compounds are routinely prepared.
  • Compounds of Formula I also include different enriched isotopic forms, e.g., compounds enriched in the content of 2 H, 3 H, 11 C, 13 C and/or 14 C.
  • the compounds are deuterated.
  • Such deuterated forms can be made by the procedure described in U.S. Patent Nos. 5,846,514 and 6,334,997. As described in U.S. Patent Nos. 5,846,514 and 6,334,997, deuteration may improve the efficacy and increase the duration of action of drugs.
  • Deuterium substituted compounds can be synthesized using various methods such as described in: Dean, Dennis C; Editor. Recent Advances in the Synthesis and Applications of Radiolabeled Compounds for Drug Discovery and Development. [In: Curr. Pharm. Des., 2000; 6(10)] 2000, 110 pp.; Kabalka, George W.; Varma, Rajender S. The Synthesis of Radiolabeled Compounds via Organometallic Intermediates, Tetrahedron, 1989, 45(21), 6601-21 ; and Evans, E. Anthony. Synthesis of Radiolabeled Compounds, J. Radioanal. Chem., 1981 , 64(1-2), 9-32.
  • Scheme 1 includes three methods to prepare compounds of Formula I and also shows that compounds of Formula I can be further elaborated at, for example R 3 , to generate additional species. It should be understood that other methods can be used to prepare compounds of Formula I as well.
  • Scheme 2
  • Compounds of Formula I can be synthesized from compounds of Formula 1.1 according to Scheme 2.
  • the compound of Formula 1.1 is, for example, reacted with phosphoryl chloride and benzyltriethylammonium chloride to yield a compound of Formula 1.2.
  • the compound of Formula 1.2 is then alkylated using electrophiles such as LG-R 7 -R 8 , where LG is a leaving group, in a solvent such as dimethylsulfoxide or dimethylformamide optionally in the presence of a base such as sodium hydride or potassium carbonate.
  • Leaving groups include, but are not limited to, e.g., mesylate, halogen, triflate, tosylate, etc.
  • the compounds of Formula 1.1 can be synthesized from aldehydes of Formula 2.1 and ketones of Formula 3.1 according to Scheme 3. Accordingly, a compound of Formula 2.1 wherein R 3 is alkyl, is brominated with, for example 5,5-dibromobarbituric acid and hydrogen bromide in a solvent such as dichloromethane, to yield a compound of Formula 2.2. The brominated aldehyde of Formula 2.2 is then condensed with an amino substituted pyrimidine to yield a compound of Formula 2.3 which upon reduction results in a compound of Formula 1.1. [00141] Compounds of Formula 1.1 can also be synthesized from ketones of Formula 3.1 according to Scheme 3.
  • a compound of Formula 3.1 wherein R 3 and R 4 are each independently alkyl or taken together with the carbon to which they are attached form a heterocyclic, is reacted with a diester, for example ethyl malonate, to yield a compound of Formula 3.2.
  • a diester for example ethyl malonate
  • Michael addition of nitromethane to the compound of Formula 3.2, followed by reduction of the nitro group to an amine results in a spontaneous cyclization to an ⁇ -carboxy- ⁇ -lactam of Formula 3.4.
  • the compound of Formula 3.4 is activated with, for example, triethyloxonium tetrafluoroborate in a solvent such as dichloromethane or with P 2 S 5 to produce a compound of Formula 3.5 which is then treated with guanidine to yield a pyrrolopyrimidine-4-one of Formula 1.1.
  • the lactam is then activated with Et 3 O + BF 4 " or P 2 Ss and treated with guanidine to yield the compound of Formula 1.1. Protection of the NH 2 group with Ac 2 O, deoxychlorination with POCI 3 and Bn(Et 3 ) 2 NCI, and deprotection with, for example, HCI yield the 4-chloro-6,7-dihydro- pyrrolo[2,3-d]pyrimidine scaffold. Finally, alkylation with 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine hydrochloride in dimethysulfoxide with sodium hydride provides the compound of Formula I.
  • Compounds of Formula I can also be synthesized from compounds of Formula 4.1 according to Scheme 5.
  • the compound of Formula 4.1 is, for example, alkylated using electrophiles such as LG-R 7 -R 8 , where LG is a leaving group, in a solvent such as dimethylsulfoxide or dimethylformamide. Leaving groups include, but are not limited to, e.g., mesylate, halogen, triflate, tosylate, etc.
  • LG-R 7 -R 8 can be, for example, a halide substituted arylalkyl.
  • the compounds of formula 4.1 can be synthesized from amino substituted pyrimidines of
  • the amino group of the pyrimidine of Formula 5.1 is optionally protected with, for example tert-butoxycarbonyl, by reacting the compound with di-tert-butyl dicarbonate under basic conditions.
  • the protected amino pyrimidine of Formula 5.2 is then alkylated with an allyl halide, for example methyl 4-bromocrotonate.
  • an allyl halide for example methyl 4-bromocrotonate.
  • the protected amino is recommended with some allyl halides (e.g., methyl bromocrotonate), but is unnecessary with others (e.g., allyl bromide).
  • the 5-position is brominated with, for example, NBS or Br 2 to set up the compound for radical cyclization.
  • the radical cyclization can be mediated by hydrogen donors such as Bu 3 SnH or TMS 3 SiH inc onjunction with a radical initiator such as AIBN, Vazo 88, or Et 3 B/O 2 .
  • a radical initiator such as AIBN, Vazo 88, or Et 3 B/O 2 .
  • the resulting enantiomers may optionally be separated by chiral HPLC or enzymatic resolution.
  • the amine protecting group is then removed with for example, trifluoroacetic acid in dichloromethane, to yield the compound of Formula 4.1.
  • Scheme 7 An example of producing a compound of Formula I by proceeding through the synthesis outlined in Scheme 6 is shown in Scheme 7.
  • the synthesis begins with a 6-chloropyrimidine-2,4-diamine of Formula 5.1a.
  • the amino group of the 6-chloropyrimidine-2,4-diamine is protected with tert- butoxycarbonyl under basic conditions using potassium tert-butoxide.
  • the Boc-protected NH is then alkylated with methyl 4-bromocrotonate.
  • the 5-position is brominated with NBS followed by a radical cyclization mediated by TMS 3 SiH and Vazo88.
  • the two enantiomers of the resulting racemic bicyclic scaffold are separated by chiral HPLC followed by cleavage of the Boc group with trifluoroacetic acid in dichloromethane.
  • the 7-position of the pyrrolopyrimidine is then alkylated in dimethylsulfoxide with a chloro substituted arylalkyl and potassium tert-butoxide to yield an ester of Formula I.
  • the protecting groups are then cleaved with H 2 SO 4 , and the hydroxy group is converted to a chlorine atom with, for example, POCI 3 .
  • Alkylation with a chloride substituted arylalkyl yields a compound of Formula I.
  • Formula 7.1 with an alkyne, followed by hydrolysis of the triple bond, to yield an ⁇ , ⁇ -unsaturated ketone of Formula 9.2.
  • the ⁇ , ⁇ -unsaturated ketone of Formula 9.2 is then reduced with, for example, a hindered borohydride such as lithium tri-sec-butylborohydride, to yield a compound of Formula I.
  • a hindered borohydride such as lithium tri-sec-butylborohydride
  • an ⁇ , ⁇ -unsaturated ketone of Formula 9.2a is obtained from the 2-amino, 4-chloro pyrrolopyrimidine by Sonagashira coupling with an alkyne, followed by hydrolysis of the triple bond.
  • the ⁇ , ⁇ -unsaturated ketone of Formula 9.2a is then reduced with lithium tri-sec-butylborohydride (LiBH(sec-Bu) 3 ) to yield a compound of Formula I.
  • the compounds of Formula I can be synthesized by other methods as well.
  • One example of another method is shown in Scheme 13.
  • the five membered ring of the compound of Formula I is constructed by starting with a 4- aminopyrimidine of Formula 11.1 and forcing the nucelophilic 4-amino group to displace an appropriately positioned leaving group (LG), such as mesylate or an epoxide. If the leaving group is an epoxide then, as shown in Scheme 14, the ring formation results in a primary alcohol.
  • LG leaving group
  • LG is a leaving group and Nuc is a nucleophile.
  • Leaving groups include, but are not limited to, e.g., mesylate, halogen, hydroxyl substituents derivatized by the Mitsunobu reaction, triflate, tosylate, etc.
  • Nucleophiles include, but are not limited to, e.g., halides, cyanide, azide, heterocycles, hydrides, etc.
  • the substitution reactions can be carried out in solvents such as acetone, tetrahydrofuran, dimethylsulfoxide, dimethylacetamide, etc. and in the presence of bases such as potassium carbonate. Following substitution, further reactions such as oxidation and coupling reactions can then be performed.
  • R 3 substituent can be elaborated by a variety of other reactions known to those skilled in the art such as cyclizations, reductions, acyl transfer reactions, sulfonation, etc. Examples of reactions that may be used to elaborate the R 3 substitutent are shown and described in Schemes 16-24 below.
  • an OH substituted R 3 moiety can displace a group on the 4- position of the pyrimidine ring to form an additional ring.
  • the displacement of the group on the 4-position can be accomplished by heating a compound of Formual 12.1 in the presence of sodium hydroxide as shown in Scheme 17.
  • Amides can be generated from an ester substituted R i3 substituent by conversion of the ester group into an amide group as shown in Scheme 18.
  • a wide variety of conditions can be employed for this conversion.
  • the ester can be hydrolyzed, activated (e.g., with pentafluorophenyl ester), and treated with ammonia as shown in Scheme 19.
  • the ester group can be directly converted to the primary amide by heating with ammonia or by heating with an ammonia equivalent such as HCONH 2 /tBuOK.
  • a synthesis analogous to that shown in Scheme 20 can be used. Specifically, the ester can be hydrolyzed, activated (e.g., with pentafluorophenyl ester), and then treated with an amine other than NH 3 .
  • Primary amides can be also generated from a cyano substituted R 3 substitent.
  • the cyano substituted R 3 substitent can be generated from a compound of Formula 14.1 by nucleophilic substitution with a CN reagent, such as potassium cyanide.
  • the cyano compound of Formula 14.2 can then be treated with, for example, H 2 CVNaOH, to yield a primary amide of Formula I.
  • An example of the route outlined in Scheme 19 starting with a 2-amino-4-chloro-6,7- dihydropyrrolopyrimidine is shown in Scheme 22.
  • the OH group of the R 3 substituent is activated by reaction with methanesulfonyl chloride which is then displaced by potassium cyanide.
  • the cyano compound is treated with H 2 O 2 /NaOH to yield a primary amide of Formula I.
  • the R 3 substituent can be activated and then substituted. Further, the R 3 substituent can by cyclized or reduced and then acylated. As a representative example, a hydroxy group can be converted into a leaving group such as a mesylate, then displaced with a nucleophile such as azide as Shown in Scheme 24. The azide can then be reacted with a terminal alkyne to give a [2+3] dipolar cycloaddition or reduced to an amine that can be further acylated. [00158] The compounds and pharmaceutically acceptable salts described herein exhibit acceptable stability towards acids, bases, and amino acids. For example, the compounds and pharmaceutically acceptable salts described herein exhibited stability towards acids.
  • the stability of the compounds and pharmaceutically acceptable salts described herein in acidic medium can be tested by incubating the compound or pharmaceutically acceptable salt in a 5M solution of methanesulfonic acid in DMSO/Water 1 :1 at about 40 0 C. Certain of the compounds described herein displayed a half-life greater than 100 h when tested under such conditions. [00159] Furthermore, compounds and pharmaceutically acceptable salts described herein exhibit stability with regard to reactions with bases such as lysine, proteins, and the like. The stability of the compounds and pharmaceutically acceptable salts described herein with bases such as lysine can be tested by incubating the compound or pharmaceutically acceptable salt in a 10 mM solution of lysine in DMSO/Water 1 :1 at about 40 0 C. Certain of the compounds described herein displayed a half-life greater than 10O h when tested under such conditions.
  • disorders that are mediated by Hsp90.
  • disorders include, but are not limited to, inflammatory diseases, infections, autoimmune disorders, stroke, ischemia, cardiac disorder, neurological disorders, fibrogenic disorders, proliferative disorders, and metabolic diseases.
  • Fibrogenic disorders include but are not limited to connective tissue diseases, such as scleroderma (or systemic sclerosis), polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders, including liver cirrhosis, keloid formation, interstitial nephritis, and pulmonary fibrosis.
  • connective tissue diseases such as scleroderma (or systemic sclerosis), polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders, including liver cirrhosis, keloid formation, interstitial nephritis, and pulmonary fibrosis.
  • Proliferative disorders include cancer, psoriasis and benign prostatic hyperplasia (BPH).
  • BPH benign prostatic hyperplasia
  • Cancer includes, for example, malignant tumors, e.g., non-solid tumors such as leukemia, multiple myeloma or lymphoma, and also solid tumours, for example, oesophageal cancer, myeloma, hepatocellular, pancreatic, cervical cancer, ewings tumour, neuroblastoma, kaposis sarcoma, ovarian cancer, endometrial cancer, colorectal cancer, prostate cancer, bladder cancer, melanoma, head and neck cancer, renal cancer, bile duct cancer, bone cancer, neuronal cancer, testicular cancer, ovarian cancer.
  • non-solid tumors such as leukemia, multiple myeloma or lymphoma
  • solid tumours for example, oesophageal cancer, myeloma, hepatocellular, pancreatic, cervical cancer, ewings tumour, neuroblastoma, kaposis sarcoma, ovarian cancer, endo
  • Additional forms of cancer include, for example, breast, ovary, lung (including small cell lung cancer, non-small cell lung cancer and bronchioalveolar cancer), colon, rectum, prostate, bile duct, bone, bladder, head and neck, kidney, liver, gastrointestinal tissue, oesophagus, pancreas, skin, testes, thyroid, uterus, cervix, vulva or other tissues, as well as leukemias and lymphomas including CLL and CML, tumors of the central and peripheral nervous system, and other tumor types such as melanoma, multiple myeloma, fibrosarcoma and osteosarcoma, and malignant brain tumors.
  • the cancer to be treated is identified by the expression or lack of expression of at least one molecular marker known to correlate with either or both of 1) dependency on Hsp90 for cancer cell growth and/or survival, and 2) susceptibility to treatment by administration of an Hsp90 inhibitor disclosed herein.
  • a cancer biopsy may be screened and identified as HER- 2 positive and that patent may then be selected for treatment by administration of a chemical entity disclosed herein.
  • the molecular marker is chosen from HER-2, estrogen receptor, progesterone receptor, androgen receptor, EGF receptor, B Raf, AKT, and NF-kB.
  • Hsp90-mediated disorders include multiple sclerosis, lupus, rheumatoid arthritis and irritable bowel syndrome CJD, Huntington's disease and Alzheimer's disease.
  • pharmaceutical compositions comprising one or more pharmaceutically acceptable carriers and at least one compound or pharmaceutically acceptable salt thereof, described herein.
  • the compounds or salts thereof may be administered either alone or in combination with pharmaceutically acceptable carriers in a pharmaceutical composition, according to standard pharmaceutical practice.
  • the compounds and pharmaceutically acceptable salts can be administered orally or parenterally, including the intravenous, intramuscular, intraperitoneal, subcutaneous, rectal and topical routes of administration.
  • the pharmaceutical compositions described herein can be administered locally to the area in need of treatment.
  • This may be achieved by, for example, but not limited to, local infusion during surgery, topical application, e.g., cream, ointment, injection, catheter, or implant, said implant made, e.g., out of a porous, non-porous, or gelatinous material, including membranes, such as sialastic membranes, or fibers.
  • the administration can also be by direct injection at the site (or former site) of a tumor or neoplastic or pre-neoplastic tissue.
  • the compounds or pharmaceutically acceptable salts described herein can be delivered in a vesicle, e.g., a liposome (see, for example, Langer, Science 1990, 249,1527 1533; Treat et al., Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Bernstein and Fidler, Ed., Liss, N.Y., pp. 353 365, 1989).
  • a liposome see, for example, Langer, Science 1990, 249,1527 1533; Treat et al., Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Bernstein and Fidler, Ed., Liss, N.Y., pp. 353 365, 1989).
  • compositions described herein can also be delivered in a controlled release system.
  • a pump may be used (see, Sefton, 1987, CRC Crit. Ref. Biomed. Eng. 14:201 ; Buchwald et al. Surgery, 1980 88, 507; Saudek et al. N. Engl. J. Med. 1989, 321 , (574).
  • a controlled release system can be placed in proximity of the therapeutic target. (See, Goodson, Medical Applications of Controlled Release, 1984, Vol. 2, pp. 115 138).
  • compositions described herein can also contain the active ingredient in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs.
  • Compositions intended for oral use may be prepared according to any method known in the art for the manufacture of pharmaceutical compositions, and such compositions may contain one or more agents selected from sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient in admixture with one or more pharmaceutically acceptable carriers which are suitable for the manufacture of tablets.
  • These pharmaceutically acceptable carriers may be, for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as microcrystalline cellulose, sodium crosscarmellose, corn starch, or alginic acid; binding agents, for example starch, gelatin, polyvinyl-pyrrolidone or acacia, and lubricating agents, for example, magnesium stearate, stearic acid or talc.
  • the tablets may be un-coated or coated by known techniques to mask the taste of the drug or delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.
  • Formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water soluble carrier such as polyethyleneglycol or an oil medium, for example peanut oil, liquid paraffin, or olive oil.
  • an inert solid diluent for example, calcium carbonate, calcium phosphate or kaolin
  • water soluble carrier such as polyethyleneglycol or an oil medium, for example peanut oil, liquid paraffin, or olive oil.
  • Aqueous suspensions contain the active material in admixture with one or more pharmaceutically acceptable carriers suitable for the manufacture of aqueous suspensions.
  • Such carriers are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethyl-cellulose, sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be a naturally-occurring phosphatide, for example lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadecaethylene-oxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and a hexitol such as polyoxyethylene sorbitol monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene sorbit
  • the aqueous suspensions may also contain one or more preservatives, for example ethyl, or n-propyl p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents, such as sucrose, saccharin or aspartame.
  • Oily suspensions may be formulated by suspending the active ingredient in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in mineral oil such as liquid paraffin.
  • the oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an anti-oxidant such as butylated hydroxyanisol or alpha-tocopherol.
  • Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already mentioned above. Additional substances, for example sweetening, flavoring and coloring agents, may also be present. These compositions may be preserved by the addition of an anti-oxidant such as ascorbic acid.
  • the pharmaceutical compositions described herein may also be in the form of an oil-in- water emulsion.
  • the oily phase may be a vegetable oil, for example olive oil or arachis oil, or a mineral oil, for example liquid paraffin, or mixtures of these.
  • Suitable emulsifying agents may be naturally- occurring phosphatides, for example soybean lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monooleate.
  • the emulsions may also contain sweetening agents, flavoring agents, preservatives and antioxidants.
  • Syrups and elixirs may be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative, flavoring and coloring agents and antioxidant.
  • sweetening agents for example glycerol, propylene glycol, sorbitol or sucrose.
  • Such formulations may also contain a demulcent, a preservative, flavoring and coloring agents and antioxidant.
  • compositions may be in the form of a sterile injectable aqueous solution.
  • acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution.
  • the sterile injectable preparation may also be a sterile injectable oil-in-water microemulsion where the active ingredient is dissolved in the oily phase.
  • the active ingredient may be first dissolved in a mixture of soybean oil and lecithin.
  • the oil solution may then be introduced into a water and glycerol mixture and processed to form a microemulsion.
  • the injectable solutions or microemulsions may be introduced into a patient's bloodstream by local bolus injection. Alternatively, it may be advantageous to administer the solution or microemulsion in such a way as to maintain a constant circulating concentration of the instant compound. In order to maintain such a constant concentration, a continuous intravenous delivery device may be utilized.
  • the pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleagenous suspension for intramuscular and subcutaneous administration.
  • This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above.
  • the sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in 1 ,3-butanediol.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • compositions described herein may also be administered in the form of suppositories for rectal administration of the drug.
  • a suitable nonirritating pharmaceutically acceptable carrier which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug.
  • suitable nonirritating pharmaceutically acceptable carrier include cocoa butter, glycerinated gelatin, hydrogenated vegetable oils, mixtures of polyethylene glycols of various molecular weights and fatty acid esters of polyethylene glycol.
  • creams, ointments, jellies, solutions or suspensions, etc., containing a compound pharmaceutically acceptable salts described herein can be used.
  • topical application can include mouth washes and gargles.
  • the compounds and pharmaceutically acceptable salts described herein can be administered in intranasal form via topical use of suitable intranasal vehicles and delivery devices, or via transdermal routes, using those forms of transdermal skin patches well known to those of ordinary skill in the art.
  • the dosage administration will, of course, be continuous rather than intermittent throughout the dosage regimen.
  • the compounds and pharmaceutically acceptable salts described herein may also be used in conjunction with other well known therapeutic agents that are selected for their particular usefulness against the condition that is being treated.
  • the compounds and pharmaceutically acceptable salts described herein may be useful in combination with at least one additional anti-cancer and/or cytotoxic agents.
  • the compounds and pharmaceutically acceptable salts described herein may also be useful in combination with other inhibitors of parts of the signaling pathway that links cell surface growth factor receptors to nuclear signals initiating cellular proliferation.
  • Such known anti-cancer and/or cytotoxic agents that may be used in combination with the compounds and pharmaceutically acceptable salts described herein include:
  • antiproliferative/antineoplastic drugs and combinations thereof as used in medical oncology, such as alkylating agents (for example cis-platin, oxaliplatin, carboplatin, cyclophosphamide, nitrogen mustard, melphalan, chlorambucil, busulphan, temozolamide and nitrosoureas); antimetabolites (for example gemcitabine and antifolates such as fluoropyrimidines like 5-fluorouracil and tegafur, raltitrexed, methotrexate, cytosine arabinoside, and hydroxyurea); antitumour antibiotics (for example anthracyclines like adriamycin, bleomycin, doxorubicin, daunomycin, epirubicin, idarubicin, mitomycinC, dactinomycin and mithramycin); antimitotic agents (for example vinca alkaloids like vincristine, vinblastine
  • cytostatic agents such as antioestrogens (for example tamoxifen, fulvestrant, toremifene, raloxifene, droloxifene and iodoxyfene), antiandrogens (for example bicalutamide, flutamide, nilutamide and cyproterone acetate), LHRH antagonists or LHRH agonists (for example goserelin, leuprorelin and buserelin), progestogens (for example megestrol acetate), aromatase inhibitors (for example as anastrozole, letrozole, vorazole and exemestane) and inhibitors of 5a-reductase such as finasteride;
  • antioestrogens for example tamoxifen, fulvestrant, toremifene, raloxifene, droloxifene and iodoxyfene
  • antiandrogens for example
  • anti-invasion agents for example c-Src kinase family inhibitors like 4-(6-chloro- 2,3methylenedioxyanilino)-7-[2-(4-methylpiperazin-l-yl)ethoxy]-5-tetrahydropyran-4yloxyquinazoline (AZD0530; International Patent Application WO 01/94341), N-(2- chloro-6-methylphenyl)-2- ⁇ 6-[4-(2- hydroxyethyl)piperazin-l-yl]-2-methylpyrimidin-4ylamino ⁇ thiazole-5-carboxamide (dasatinib, BMS-354825; J. Med.
  • anti-invasion agents for example c-Src kinase family inhibitors like 4-(6-chloro- 2,3methylenedioxyanilino)-7-[2-(4-methylpiperazin-l-yl)ethoxy]-5-tetrahydropyran-4yloxyqui
  • inhibitors of growth factor function include growth factor antibodies and growth factor receptor antibodies (for example the anti-erbB2 antibody trastuzumab [HerceptinTM], the anti-EGFR antibody panitumumab, the anti-erbB 1 antibody cetuximab [Erbitux, C225] and any growth factor or growth factor receptor antibodies disclosed by Stem et al. Critical reviews in oncology/haematology, 2005, Vol.
  • inhibitors also include tyrosine kinase inhibitors, for example inhibitors of the epidermal growth factor family (for example EGFR family tyrosine kinase inhibitors such as W-fS-cft/oro ⁇ -fluorophenyO ⁇ -methoxy-e-fS-morpholinopropoxyJquinazolin ⁇ -amine (gefitinib, ZD1839), N-(3-ethynylphenyl)-6,7-bis(2-methoxyethoxy)quinazolin-4-amine (erlotinib, OSI-774) and 6-acrylamido-N-(3-chloro-4-fluorophenyl)-7-(3- morpholinopropoxy)-quinazolin-4-amine (Cl 1033), erbB2 tyrosine kinase inhibitors such as lapatinib); inhibitors of the hepatocyte growth factor family; inhibitors of the hepatocyte growth factor family
  • antiangiogenic agents such as those which inhibit the effects of vascular endothelial growth factor, [for example the anti-vascular endothelial cell growth factor antibody bevacizumab (AvastinTM) and for example, a VEGF receptor tyrosine kinase inhibitor such as vandetanib(ZD6474), vatalanib (PTK787), sunitinib (SU1 I248), axitinib (AG-013736), pazopanib (GW 786034) and 4 ⁇ 4-fluoro-2- methylindol-5-yloxy)-6-methoxy-7-(3pyrrolidin-l-ylpropoxy)quinazoline (AZD2I7I; Example 240 within WO 00/47212), compounds such as those disclosed in International Patent Applications W097/22596, WO 97/30035, WO 97/32856 and WO 98/13354 and compounds that work by other mechanisms (for example l)
  • vascular damaging agents such as Combretastatin A4 and compounds disclosed in International Patent Applications WO 99/02166, WO 00/40529, WO 00/41669, WO 01/92224, WO 02/04434 and WO 02/08213;
  • an endothelin receptor antagonist for example zibotentan (ZD4054) or atrasentan;
  • antisense therapies for example those which are directed to the targets listed above, such as ISIS 2503, an anti-ras antisense;
  • (ix) gene therapy approaches including for example approaches to replace aberrant genes such as aberrant p53 or aberrant BRCAI or BRCA2, GDEPT (gene-directed enzyme pro-drug therapy) approaches such as those using cytosine deaminase, thymidine kinase or a bacterial nitroreductase enzyme and approaches to increase patient tolerance to chemotherapy or radiotherapy such as multidrug resistance gene therapy; and
  • GDEPT gene-directed enzyme pro-drug therapy
  • (x) immunotherapy approaches including for example ex-vivo and in-vivo approaches to increase the immunogenicity of patient tumour cells, such as transfection with cytokines such as interleukin 2, interleukin 4 or granulocyte-macrophage colony stimulating factor, approaches to decrease T-cell anergy, approaches using transfected immune cells such as cytokine-transfected dendritic cells, approaches using cytokine-transfected tumour cell lines and approaches using anti-idiotypic antibodies.
  • cytokines such as interleukin 2, interleukin 4 or granulocyte-macrophage colony stimulating factor
  • the compound or pharmaceutically acceptable salt is administered in combination with one or more agents chosen from pacliataxel, bortezomib, dacarbazine, gemcitabine, trastuzumab, bevacizumab, capecitabine, docetaxel, erlotinib, aromatase inhibitors, such as AROMASINTM (exemestane), and estrogen receptor inhibitors, such as FASLODEXTM (fulvestrant).
  • the compounds and pharmaceutically acceptable salts described herein may be useful in combination with at least one additional anti-inflammatory and/or immuno-modulatory agent.
  • Such known additional anti-inflammatory and/or immuno-modulatory agents that may be used in combination with the compounds and pharmaceutically acceptable salts described herein include:
  • TNF-blocking agents such as etanercept, infliximab, and adalimumab
  • anti-B cell therapeutics such as rituximab
  • nonsteroidal anti-inflammatory drugs such as diclofenac, etodolac, fenoprofen, flurbiprofen, ibuprofen, indomethacin, ketoprofen, meclofenamate, nabumetone, naproxen, oxaprozin, piroxicam, sulindac and tolmetin;
  • COX-2 inhibitors such as celecoxib and meloxicam
  • DMARDs disease modifying anti-rheumatic drugs
  • DMARDs disease modifying anti-rheumatic drugs
  • sulfasalazine hydroxychloroquine, chloroquine, gold salts, D-penicillamine, and methotrexate
  • anti-malarials such as hydroxychloroquine, chloroquine, and quinacrine
  • corticosteroids such as prednisone, betamethasone, dexamethasone, hydrocortisone, methylprednisolone, and triamcinolone;
  • the compound or pharmaceutically acceptable salt is administered in combination with one or more agents chosen from hydroxychloroquine and methotrexate.
  • the daily dosage will normally be determined by the prescribing physician with the dosage generally varying according to the age, weight, and response of the individual patient, as well as the severity of the patient's symptoms.
  • a suitable amount of compound is administered to a mammal undergoing treatment for cancer, for example, breast cancer.
  • Administration typically occurs in an amount of between about 0.01 mg/kg of body weight to about 100 mg/kg of body weight per day (administered in single or divided doses), such as at least about 0.1 mg/kg of body weight per day.
  • a particular therapeutic dosage can include, e.g., from about 0.01 mg to about 1000 mg of compound, such as including, e g , from about 1 mg to about 1000 mg
  • the quantity of active compound in a unit dose of preparation may be varied or adjusted from about 0 1 mg to 1000 mg, such as from about 1 mg to 300 mg, for example 10 mg to 200 mg, according to the particular application
  • the amount administered will vary depending on the particular IC 50 value of the compound used and the judgment of the attending clinician taking into consideration factors such as health, weight, and age In combinational applications in which the compound is not the sole active ingredient, it may be possible to administer lesser amounts of compound and still have therapeutic or prophylactic effect
  • the pharmaceutical preparation is in unit dosage form
  • the preparation is subdivided into unit doses containing appropriate quantities of the active component, e g , an effective amount to achieve the desired purpose
  • the actual dosage employed may be varied depending upon the requirements of the patient and the severity of the condition being treated Determination of the proper dosage for a particular situation is within the skill of the art Generally, treatment is initiated with smaller dosages which are less than the optimum dose of the compound Thereafter, the dosage is increased by small amounts until the optimum effect under the circumstances is reached For convenience, the total daily dosage may be divided and administered in portions during the day if desired
  • the amount and frequency of administration of the compounds and pharmaceutically acceptable salts described herein, and if applicable other chemotherapeutic agents and/or radiation therapy will be regulated according to the judgment of the attending clinician (physician) considering such factors as age, condition and size of the patient as well as severity of the disease being treated
  • the chemotherapeutic agent and/or radiation therapy can be administered according to therapeutic protocols well known in the art It will be apparent to those skilled in the art that the administration of the chemotherapeutic agent and/or radiation therapy can be varied depending on the disease being treated and the known effects of the chemotherapeutic agent and/or radiation therapy on that disease Also, in accordance with the knowledge of the skilled clinician, the therapeutic protocols (e g , dosage amounts and times of administration) can be varied in view of the observed effects of the administered therapeutic agents ( ⁇ e , antineoplastic agent or radiation) on the patient, and in view of the observed responses of the disease to the administered therapeutic agents
  • the compounds and pharmaceutically acceptable salts described herein need not be administered in the same pharmaceutical composition as a chemotherapeutic agent, and may, because of different physical and chemical characteristics, be administered by a different route
  • the compounds/compositions may be administered orally to generate and maintain good blood levels thereof, while the chemotherapeutic agent may be administered intravenously
  • the determination of the mode of administration and the advisability of administration, where possible, in the same pharmaceutical composition, is well within the knowledge of the skilled clinician
  • the initial administration can be made according to established protocols known in the art, and then, based upon the observed effects, the dosage, modes of administration and times of administration can be modified by the skilled clinician.
  • the compounds and pharmaceutically acceptable salts described herein may be administered concurrently (e.g., simultaneously, essentially simultaneously or within the same treatment protocol) or sequentially, depending upon the nature of the proliferative disease, the condition of the patient, and the actual choice of chemotherapeutic agent and/or radiation to be administered in conjunction (i.e., within a single treatment protocol) with the compound/composition.
  • the compound/composition and the chemotherapeutic agent and/or radiation need not be administered simultaneously or essentially simultaneously, and the initial order of administration of the compound/composition, and the chemotherapeutic agent and/or radiation, may not be important.
  • the compounds and pharmaceutically acceptable salts described herein may be administered first followed by the administration of the chemotherapeutic agent and/or radiation; or the chemotherapeutic agent and/or radiation may be administered first followed by the administration of the compounds and pharmaceutically acceptable salts described herein. This alternate administration may be repeated during a single treatment protocol.
  • the determination of the order of administration, and the number of repetitions of administration of each therapeutic agent during a treatment protocol is well within the knowledge of the skilled physician after evaluation of the disease being treated and the condition of the patient.
  • the chemotherapeutic agent and/or radiation may be administered first, and then the treatment continued with the administration of the compounds and pharmaceutically acceptable salts described herein followed, where determined advantageous, by the administration of the chemotherapeutic agent and/or radiation, and so on until the treatment protocol is complete.
  • the practicing physician can modify each protocol for the administration of a compound/composition for treatment according to the individual patient's needs, as the treatment proceeds.
  • Hsp90 competitive binding assays and functional assays can be performed as known in the art by substituting in the compounds and pharmaceutically acceptable salts described herein. Chiosis et al. Chemistry & Biology 2001 , 8, 289 299, describe some of the known ways in which this can be done.
  • competition binding assays using, e.g., geldanamycin or 17-AAG as a competitive binding inhibitor of Hsp90 can be used to determine relative Hsp90 affinity of the compounds and pharmaceutically acceptable salts described herein by immobilizing the compound or salt of interest or other competitive inhibitor on a gel or solid matrix, preincubating Hsp90 with the other inhibitor, passing the preincubated mix over the gel or matrix, and then measuring the amount of Hsp90 that retains or does not retain on the gel or matrix.
  • competition binding assays using a geldanamycin fluorescent probe e.g., reduced FITC- geldanamycin, as a competitive binding inhibitor of Hsp90 or an Hsp90 paralog
  • a geldanamycin fluorescent probe e.g., reduced FITC- geldanamycin
  • competition binding assays using a geldanamycin fluorescent probe can be be performed by pre-incubating Hsp90 or the Hsp90 paralog with the geldanamycin fluorescent probe, adding the compound or salt of interest, incubating the reaction with gentle shaking, and then measuring the fluorescence.
  • Maximum signal and background controls contain no compound or no Hsp90 (or Hsp90 paralog), respectively.
  • Downstream effects can also be evaluated based on the known effect of Hsp90 inhibition on function and stability of various steroid receptors and signaling proteins including, e.g., Raf1 and HER2.
  • Compounds and pharmaceutically acceptable salts described herein induce dose-dependent degradation of these molecules, which can be measured using standard techniques.
  • Hsp90 inhibition can be determined with a cell based assay that quantifies the expression level of HER2 in tumor cells treated with the compound or salt of interest. Inhibition of Hsp90 also results in up- regulation of Hsp90 and related chaperone proteins that can similarly be measured.
  • Antiproliferative activity on various cancer cell lines can also be measured, as can morphological and functional differentiation related to Hsp90 inhibition.
  • the tumor cell growth inhibitory activity of the compound or salt of interest may be determined by treating tumor cells with various concentrations of the compound or salt of interest followed by measurement of cell viability.
  • Indirect techniques include nucleic acid hybridization and amplification using, e.g., polymerase chain reaction (PCR). These techniques are known to the person of skill and are discussed, e.g., in Sambrook, Fritsch & Maniatis Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y., 1989; Ausubel, et al.
  • HER2 expression in breast cancer cells can be determined with the use of an immunohistochemical assay, such as the Dako HercepTM test (Dako Corp., Carpinteria, Calif.)-
  • the HercepTM test is an antibody staining assay designed to detect HER2 overexpression in tumor tissue specimens. This particular assay grades HER2 expression into four levels: 0, 1 , 2, and 3, with level 3 representing the highest level of HER2 expression.
  • Accurate quantitation can be enhanced by employing an Automated Cellular Imaging System (ACIS) as described, e.g., by Press, M. et al. Modern Pathology 2000, 13, 225A.
  • ACIS Automated Cellular Imaging System
  • Antibodies polyclonal or monoclonal, can be purchased from a variety of commercial suppliers, or may be manufactured using well-known methods, e.g., as described in Harlow et al. Antibodies: A Laboratory Manual, 2nd ed; Cold Spring Harbor Laboratory, Cold Spring Harbor, N. Y., 1988.
  • HER2 overexpression can also be determined at the nucleic acid level since there is a reported high correlation between overexpression of the HER2 protein and amplification of the gene that codes for it.
  • One way to test this is by using RT-PCR.
  • the genomic and cDNA sequences for HER2 are known.
  • Specific DNA primers can be generated using standard, well-known techniques, and can then be used to amplify template already present in the cell. An example of this is described in Kurokawa, H. et al. Cancer Res. 2000, 60, 5887 5894.
  • PCR can be standardized such that quantitative differences are observed as between normal and abnormal cells, e.g, cancerous and noncancerous cells.
  • Well known methods employing, e.g., densitometry can be used to quantitate and/or compare nucleic acid levels amplified using PCR.
  • FISH fluorescent in situ hybridization
  • other assays can be used, e.g., Northern and/or Southern blotting.
  • FISH fluorescent in situ hybridization
  • these rely on nucleic acid hybridization between the HER2 gene or mRNA and a corresponding nucleic acid probe that can be designed in the same or a similar way as for PCR primers, above. See, e.g., Mitchell M S, and Press M. F. Oncol., Suppl. 1999, 12, 108 116.
  • this nucleic acid probe can be conjugated to a fluorescent molecule, e.g., fluorescein and/or rhodamine, that does not interfere with hybridization, and which fluorescence can later be measured following hybridization.
  • lmmuno and nucleic acid detection can also be directed against proteins other than
  • Hsp90 and HER2 which proteins are nevertheless affected in response to Hsp90 inhibition.
  • In vitro and in vivo assays are also available to determine the anti-inflammatory and immunomodulatory activities of the compounds and pharmaceutically acceptable salts described herein. For example, the stability and function of newly synthesized and activated/phosphorylated forms of a proximal signaling molecule in the T cell receptor activation pathway, the Src family kinase p56 fck , is dependent upon Hsp90, with Hsp90 inhibition resulting in p56"* degradation and subsequent internalization of CD4.
  • the ability of the compound or salt of interest to inhibit human CD4 + T cell proliferation in response to an allogeneic non-T cell stimulus and the ability of the compound or salt of interest to down modulate T cell surface CD4 can be measured.
  • the ability of the compound or salt of interest to inhibit lipopolysaccharide (LPS)-induced activation of mitogen-activated protein kinase (MAPK) pathways can be determined.
  • the autoclave was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 80 0 C while stirring for 3 hours. The reaction was cooled to room temperature and diluted with water (20OmL), and extracted with a mixture of dichloromethane: isopropanol (95:5, 300mLx3). The organic layer was filtered over a celite pad, concentrated, and the solid was collected, washed with cold methanol (5mLx3), and dried over high vacuum pump to give the title compound as a white solid (16.0 g, purity 85.0%, yield 63.0%).
  • reaction mixture was warmed to room temperature, allowed to stir for 30 min, poured into ice-water (10OmL), washed with sat. NH 4 CI solution, dried over sodium sulfate, and evaporated.
  • the aqueous layer was purified by reverse- phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA). The desired fractions were neutralized with sat. NaHCO 3 solution, extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (400 mg, purity 99.6%, yield 40.0%).
  • reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI, and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white powder (60.0 mg, purity 99.5%, yield 70.0%).
  • reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI, and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white powder (60.0 mg, purity 99.5%, yield 70.0%).
  • reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI, and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA).
  • desired fractions were neutralized with sat. NaHCO 3 , extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (20.0 mg, purity 99.6%, yield 20.0%).
  • reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI solution, and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA).
  • desired fractions were neutralized with sat. NaHCO 3 solution, extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (20.0 mg, purity 99.6%, yield 20.0%).
  • the reaction mixture was stirred at room temperature overnight, evaporated, diluted with water (10OmL), adjusted to pH 4.0 with 2N HCI.
  • the impurities were washed with dichloromethane (30mLx3).
  • the aqueous layer was purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1 % TFA).
  • the desired fractions were neutralized with NaHCO 3 , extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (70.0 mg, purity 99.5%, yield 50.0%).
  • the reaction mixture was stirred at room temperature overnight, evaporated, diluted with water (10OmL), and adjusted to pH 4.0 with 2N HCI.
  • the impurities were washed with dichloromethane (30mLx3).
  • the aqueous layer was purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA).
  • the desired fractions were neutralized with aqueous NaHCO 3 , extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (55.0 mg, purity 99.5%, yield 43.0%).
  • the crude was diluted with water (3OmL), adjusted to pH 4.0 with 2N HCI and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA).
  • the desired fractions were neutralized with NaHCO 3 , extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (15.0 mg, purity 99.6%, yield 24.0%).
  • the reaction mixture was diluted with water (60ml), and extracted with dichloromethane: isopropanol (85:15, 20ml_x3). The organic layer was evaporated and treated with trifluoroacetic acid (0.110 mL, 1.43 mol) overnight. The reaction mixture was evaporated, diluted with water (20 mL), adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA). The desired fractions were neutralized with NaHCO 3 , extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (18.0 mg, purity 99.6%, yield 28.0%).
  • reaction mixture was warmed to room temperature, allowed to stir for 1hr, carefully quenched with ice water, adjusted to pH 7.0, evaporated, and purified by reverse-phase preparative HPLC (5 ⁇ 30% CH 3 CN/H 2 O, 0.1% TFA). The desired fractions were neutralized with NaHCO 3 , extracted with dichloromethane: isopropanol (85:15, 10mLx3), and evaporated to give the title compound as a white solid (8.0 mg, purity 99.6%, yield 20.0%).
  • Step 3 1 -(2-amino-4-chloro-7-((4-methoxy-3, 5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)ethanol (XXXI)
  • the resulting yellow suspension was stirred at 0 0 C for 5 minutes, and treated sequentially with 4-oxo-piperidine-1-carboxylic acid tert-butyl ester (20.0 g, 0.100 mol) in anhydrous tetrahydrofuran (50 mL) and ethyl malonate (15.2 mL, 0.100 mol).
  • the reaction mixture was stirred at 0 0 C for 30 minutes and then treated with a solution of dry pyridine (32.5 mL, 0.402 mol) in anhydrous tetrahydrofuran (60 mL). The mixture was stirred at 0 0 C for 1 hour and then at room temperature for 72 hours.
  • the resulting black solution was quenched with aqueous NH 4 CI (300 mL) and diluted with dichloromethane (500 mL).
  • the organic layer was separated and washed with aqueous NH 4 CI (200mLx6) containing 5% NH 4 OH until the organic layer turned to light yellow.
  • the organic layer was dried over Na 2 SO 4 , and evaporated.
  • the crude was purified by flash chromatography (methylene chloride: methanol 100:0 to 100:5). The desired fractions were evaporated to give the product as a light yellow oil (9.2Og, purity 90.0%, yield 85.0%).
  • Step 8 1-(2'-amino-4'-chloro-7'-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6', 7'- dihydrospiro[piperidine-4,5'-pyrrolo[2,3-d]pyrimidine]-1-yl)ethanone (XXXIII) [00273] A mixture of 1-(2'-amino-4'-chloro-6',7'-dihydrospiro[piperidine-4,5'-pyrrolo[2,3- d]pyrimidine]-1-yl)ethanone (4H, 16.4 mg, 0.0582 mmol), and 2-(chloromethyl)-4-methoxy-3,5- dimethylpyridine (10.8 mg, 0.0582 mol), NaH (2.93 mg, 0.122 mmol) in anhydrous dimethyl sulfoxide (1.0 mL) was stirred at 100 0 C for 1hr.
  • N.N-Dimethylformamide (0.9 mL) was treated with (2-Ethenyl)tri-n-butyltin (113 ⁇ L, 0.387 mmol) and Tetrakis(triphenylphosphine)palladium(0) (13 mg, 0.012 mmol) in microwave vial.
  • the mixture was microwaved at 100 0 C for 30 minutes, after which water was added.
  • the metal solids were filtered off and rinsed with ethyl acetate.
  • the resulting filtrate was extracted 3 times with ethyl ether, which was washed twice with aqueous potassium fluoride. Purification by silica gel chromatography using ethyl acetate/hexane gave the title compound (22 mg, yield 75%).
  • Step 4 4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-5-iodo-7H-pyrrolo[2,3-d]pyrimldin-2- amine
  • the title compound was obtained from 5-bromo-4-methoxy-2,3-dimethyl-pyridine 1 -oxide by following the 3 step sequence (1. m-CPBA, 2. Ac 2 O, 3. OH " ) described in Kasibhatla S. R. et al. J.
  • Step 4 (4-methoxy-3-methyl-5-(prop- 1 -en-2-yl)pyridin-2-yl)methanol
  • Step 8 Methyl 2-amino-4-chloro-7-((5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methyl)-7H- pyrrol ⁇ [2,3-d]pyrimidine-5-carboxylate
  • the resulting reaction mixture was stirred at RT for 16h.
  • the reaction mixture was diluted with cold water (1500 mL) and extracted with EtOAc (3 x 500 mL). The combined organic layers were washed with ice cold water and dried over Na 2 SO 4 .
  • the solvent was evaporated under reduced pressure and the crude material was purified by precipitation from 20% dichloromethane/Hexane (100 mL) to afford the title compound (25.2 g, 71%) as a solid.
  • Step 4 (2-amino-4-chloro-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol
  • a mixture of (2-amino-4-chloro-7-(4-methoxybenzyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (5 g, 15 mmol) and TFA (35 ml) was stirred at 80 C. After 1h, H 2 SO 4 (2 ml) was added to the reaction mixture and the heating was continued for another 8 hours. After consumption of the starting material (by TLC), the volatiles were evaporated under reduced pressure.
  • Tetrahydrofuran (30 mL) was treated at rt with 3.0 M of Methylmagnesium bromide in Ether (2.2 mL, 6.6 mmol). The reaction was instantaneous and clean. The reaction was quenched with MeOH, evaporated, and extracted into EtOAc to give the desired product, which was used in the next step without further purification.
  • Step 2 1 -Methyl-4, 5, 6, 7-tetrahydro- 1 H-indazole-3-carboxylic acid ethyl ester
  • the mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (1.0 mL,
  • reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water
  • reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water

Abstract

Provided are certain Hsp90 inhibitors, i.e., compounds of Formula I and pharmaceutically acceptable salts thereof, pharmaceutical compositions thereof, and methods for their use and preparation.

Description

CERTAIN SUBSTITUTED PYRIMIDINES, PHARMACEUTICAL COMPOSITIONS THEREOF, AND
METHODS FOR THEIR USE
[0001] This application claims priority to U.S. Application No. 61/165,453, filed March, 31 , 2009 and to U.S. Application No. 61/165,457, filed March 31 , 2009, each of which is incorporated herein by reference in its entirety.
[0002] Provided are certain Hsp90 inhibitors, pharmaceutical compositions thereof, and methods for their use and preparation.
[0003] Hsp90s are ubiquitous chaperone proteins that are involved in folding, activation and assembly of a wide range of proteins, including key proteins involved in signal transduction, cell cycle control and transcriptional regulation. Researchers have reported that Hsp90 chaperone proteins are associated with important signaling proteins, such as steroid hormone receptors and protein kinases, including, e.g., Raf-1 , AKT, KIT, EGFR, v-Src family kinases, Cdk4, and ErbB-2. Studies further indicate that certain co-chaperones, e.g., Hsp70, p60/Hop/Sti1 , Hip, Bag1 , Hsp40/Hdj2/Hsj1 , immunophilins, p23, and p50, may assist Hsp90 in its function.
[0004] Hsp90 possesses a binding pocket at its N-terminus. This pocket is highly conserved and has weak homology to the ATP-binding site of DNA gyrase. Further, ATP and ADP have both been shown to bind this pocket with low affinity and to have weak ATPase activity. In vitro and in vivo studies have demonstrated that occupancy of this N-terminal pocket by ansamycins and other Hsp90 inhibitors alters Hsp90 function and inhibits protein folding. At high concentrations, ansamycins and other Hsp90 inhibitors have been shown to prevent binding of protein substrates to Hsp90. Hsp90 inhibitors, e.g. ansamycins, have also been demonstrated to inhibit the ATP-dependent release of chaperone-associated protein substrates. In either event, the substrates are degraded by an ubiquitin-dependent process in the proteasome.
[0005] Hsp90 is required for the stability, folding, and activity of specific "client" proteins that are involved in tumor cell signaling such as HER-2, ER, EGFR, cKIT, BRaf. Inhibition of Hsp90 results in client protein degradation leading to tumor cell stasis or death.
[0006] The combination of genetic instability and a stressed environment ensures enhanced cancer cell dependence on Hsp90 function. Inhibition of Hsp90 results in degradation of proteins essential for the maintenance of multiple oncogenic pathways and results in antitumor activity. Hsp90 inhibitors described herein, either as monotherapy or as combination therapy with standard of care therapies, can be used to inhibit multiple tumor signaling pathways, enhance the activity of these agents and lead to greater antitumor activity.
[0007] Including use as anti-cancer agents, Hsp90 inhibitors have also been implicated in a wide variety of other utilities, such as, anti-inflammation agents, anti-infectious disease agents, agents for treating autoimmunity, agents for treating stroke, ischemia, multiple sclerosis, cardiac disorders, central nervous system related disorders and agents useful in promoting nerve regeneration. Overlapping somewhat with the above, there are reports in the literature that fibrogenic disorders also may be treatable with Hsp90 inhibitors. Fibrogenic disorders include, but are not limited to, connective tissue diseases, such as scleroderma (or systemic sclerosis), polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders, including liver cirrhosis, keloid formation, interstitial nephritis, and pulmonary fibrosis.
[0008] Medications for the treatment of systemic lupus erythematosis (SLE) are limited, and no new medication for SLE has been approved in the past 30 years. The mainstay of therapy continues to be corticosteroids for flares of moderate to severe disease. Nonsteroidal anti-inflammatory drugs (NSAIDs), antimalarials, corticosteroids, and other immunosuppressive agents such as cyclophosphamide, azathioprine or 6-mercaptopurine, methotrexate (MTX), and mycophenolate mofetil (MMF) are used alone or in combination for the many manifestations of this disease. Azathioprine, cyclophosphamide, MTX, and MMF are common immunosuppressives for subsets of SLE patients exhibiting chronic active or relapsing disease activity who are not responsive to antimalarials and prednisone alone. Recently, belimumab (anti-BAFF monoclonal antibody) has been shown in two large, randomized, controlled Phase 3 studies to be effective in SLE, and is awaiting an approval decision from the FDA.
[0009] Current rheumatoid arthritis (RA) therapies consist of non-steroidal anti-inflammatory drugs (NSAIDs), glucocorticoids, DMARDs (disease modifying anti-rheumatic drugs) which include methotrexate (MTX), and newer biologic agents that target specific pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α) and interleukin-1 (IL-1) or anti-CD20 and antagonists of CD80/CD86. NSAIDs treat inflammation and pain but do not modify the course of disease and are associated with well known complications (gastrointestinal ulceration, exacerbation of hypertension, headache and depression, and exacerbation of underlying renal disease). Glucocorticoids treat acute symptomatic joint flares, inflammation, and pain when NSAIDs alone are inadequate. However, while the short-term effects are powerful, the long-term toxicities are substantial. When RA becomes established and is not adequately responding to NSAIDs or low-dose glucocorticoids, DMARDs are the next line of treatment (sulfasalazine, hydroxychloroquine, chloroquine, gold salts, D-penicillamine, and MTX). MTX in combination with NSAIDs has become the mainstay of initial DMARD therapy because MTX has proven efficacy, treats synovitis, and slows progression of structural damage. However, many patients either have an inadequate response to initial MTX therapy or lose responsiveness over time. For those patients, the biologic agents have been shown to be beneficial. Examples include etanercept (soluble TNF receptor fusion protein), infliximab (chimeric anti-TNF-α), adalimumab (humanized anti-TNF-α), rituximab (chimeric anti-CD20), abatacept (human CTLA4-Fc) and tocilizumab (humanized anti-IL-6R). Each of these drugs has been shown to improve the signs and symptoms of RA, improve physical function, and inhibit or slow the progression of structural damage. Yet, there remains significant unmet medical need for patients with RA. [0010] Large Phase 3 studies with the TNF-blocking agents (etanercept, infliximab, and adalimumab) conducted in patients with an inadequate response to MTX highlight the unmet medical need. As many as 33% to 50% of patients with an inadequate response to MTX also had minimal detectable clinical response to the TNF-blocking agents, since the proportion of patients who achieved American College of Rheumatology (ACR) responses at 6 months ranged from 50% to 67%. Recently approved biologic treatments for RA have been shown to be effective in patients with an inadequate response to TNF inhibitors (rituximab, abatacept and tocilizumab). However, response rates to these agents remain relatively low and many patients respond suboptimally or not at all. [0011] Thus, a need remains for additional Hsp90 inhibitors.
[0016] Provided is a compound represented by Formula I
Figure imgf000004_0001
or a pharmaceutically acceptable salt thereof, wherein:
X is a bond or an alkylene chain of the formula (CRaRb)n where n is 1 to 3, for each occurrence, Ra and Rb are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which, except for hydrogen, is optionally substituted, and a (CRaRb) unit is optionally replaced by -O-, -N(Ra)-, or -S-; or when n is 2 or 3, Ra in a first (CRaRb) unit is optionally taken together with Ra in a different (CRaRb) unit to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or
Ra is optionally taken together with Rb in the same (CRaRb) unit to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; R1 is selected from hydrogen, halogen, -OR11, -SR12, amino, and optionally substituted lower alkyl;
R2 is selected from -NR21R22, -N=C-NR21 R22, -NR21-C(O)R23 and -NR21-SO2R24; R3 and R4 are independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alicyclic, optionally substituted alicyclicalkyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -C(=J)RX, -C(=J)N(Ry)(Rz), C(=J)N(Rx)S(O)2Rχ, -C(=J)N(RX)N(RX)S(O)2RX, -C(RxJ=N(ORx), -C(RJ=NN(Ry)(R1), -C≡N, -Cf=J)ORx, -Cf=J)SRx, -N(Ry)(R1), -N(Rx)Ct=J)Rx, -N(Rx)Ct=J)N(Ry)(R1), -N(RX)C(=J)ORXI -N(RX)C(=J)SRX> -N(RX)S(O)2RW> -N(Rx)S(O)2ORx, -S(O)1ORx (where t is 1 or 2), - S(O)NRyRz, -NH-C(-NH2)=C-NO2, -CO-NH-SO2-NH2, -N(Rx)S(O)2N(Ry)(R1), N3, NO2, -ORx, -OC(=J)RX, -OC(=J)ORX, -OCt=J)N(Ry)(Rx), -OC(=J)SRX, -OP(O)(Rv)2, -OS(O)2Rx, -OS(O)2N(Ry)(R2), -OSi(RJ3, -P(O)(Rv)2, -SRx, -S(O)tRx (where t is 1 or 2), -S(O)2N(Ry)(Rz), -Si(Rw)3, and halogen, wherein each J is independently O, NRU or S wherein each Ru is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rv is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -ORx or -N(Ry)(Rz); each Rw is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rx is independently hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; and Ry and Rz are each independently hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; or Ry and Rz, together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl;
R3 is optionally taken together with R4 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, n is 1 , and the (CRaRb) unit is replaced by -N(Ra)-, then R3 and R4 taken together optionally form oxo; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, an Ra unit is optionally taken together with R1 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, an Ra in a first (CRaRb) unit is optionally taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is a bond, R1 is optionally taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90;
R5 and R6 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; or
R5 is optionally taken together with R6, to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, n is 1 , and the (CRaRb) unit is replaced by -N(Ra)-, then R5 and R6 taken together optionally form oxo; or
R3 is optionally taken together with R5, to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90;
R7 is selected from a bond, optionally substituted alkylene, -C(O)-, -C(S)-, -S(O)-, and -SO2-;
R8 is selected from alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which is optionally substituted;
R11 and R12 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; and
R21 and R22 are independently selected from hydrogen, alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl, each of which, except for hydrogen, is optionally substituted, or
R21 is optionally taken together with R22 to form an optionally substituted ring of 3-8 ring atoms wherein said optionally substituted ring optionally includes one or two additional ring atoms that are heteroatoms independently selected from O, S, =N, and NR90;
R23 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
R24 is selected from alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and
R90 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and wherein from 1 to 5 hydrogen atoms in the compound of Formula I are optionally replaced with deuterium, and provided that when X is an optionally substituted alkylene chain of the formula (CRaRb)n , n is 1 , the (CRaRb) unit is replaced by -N(Ra)-, and R7 is -CH2-, then R8 is not 3,5-dimethyl-4-methoxy-pyridin-2- yi- [0017] Also provided is a pharmaceutical composition comprising one or more pharmaceutically acceptable excipients and at least one compound or pharmaceutically acceptable salt thereof described herein.
[0018] Also provided is a method of treating an individual having an Hsp90 mediated disorder comprising administering to said individual at least one compound or pharmaceutically acceptable salt described herein or a pharmaceutical composition described herein. BRIEF DESCRIPTION OF THE FIGURES
[0019] Figure 1 shows the effect of Compound A on client proteins in MCF7 cells.
[0020] Figure 2 shows the effect of Compound A on client proteins in N87 cells.
[0021] Figure 3 shows that Compound A down modulates CD4 expression on human blood T cells.
[0022] Figure 4 shows that in a mouse macrophage cell line Compound A decreased phosphorylation of MEK1/2, ERK1/2, and JNK1/2 in response to LPS. [0023] Figure 5 summarizes Compound A tumor efficacy studies.
[0024] Figure 6 shows the antitumor activity of Compound A in the high HER-2 N87 gastric carcinoma xenograft model.
[0025] Figure 7 shows the antitumor activity of Compound A in the high HER-2 BT474 breast carcinoma xenograft model.
[0026] Figure 8 shows the antitumor activity of Compound A in the HT29 colon carcinoma xenograft model.
[0027] Figure 9 shows that Compound A inhibits LPS-induced systemic TNF-α release in DBA/1 mice.
[0028] Figure 10 shows the efficacy of orally administered Compound A in a rat collagen- induced arthritis model.
[0029] As used in the present specification, the following words and phrases are generally intended to have the meanings as set forth below, except to the extent that the context in which they are used indicates otherwise.
[0030] A "pharmaceutically acceptable salt" may be prepared for any compound having a functionality capable of forming a salt, for example, an acid or base functionality. Pharmaceutically acceptable salts may be derived from organic or inorganic acids and bases. Compounds that contain one or more basic functional groups, e.g., amino or alkylamino, are capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable organic and inorganic acids. These salts can be prepared in situ during the final isolation and purification of the compounds, or by separately reacting a purified compound in its free base form with a suitable organic or inorganic acid, and isolating the salt thus formed. Examples of suitable acid salts include acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2- hydroxyethanesulfonate, lactate, maleate, malonate, methanesulfonate, 2-napthalenesulfonate, nicotinate, nitrate, oxalate, palmoate, pectinate, persulfate, 3-phenylpropionate, phosphate, picrate, pivalate, propionate, salicylate, succinate, sulfate, tartrate, thiocyanate, and tosylate. Other acids, such as oxalic, while not in themselves pharmaceutically acceptable, may be employed in the preparation of salts useful as intermediates in obtaining the compounds described herein and their pharmaceutically acceptable acid addition salts. See, e.g., Berge et al. "Pharmaceutical Salts", J. Pharm. Sci. 1977, 66:1-19.
[0031] Compounds that contain one or more acidic functional groups are capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable bases. The term "pharmaceutically acceptable salts" in these instances refers to the relatively non-toxic, inorganic and organic base addition salts of compounds described herein. These salts can likewise be prepared in situ during the final isolation and purification of the compounds, or by separately reacting the purified compound in its free acid form with a suitable base, such as the hydroxide, carbonate or bicarbonate of a pharmaceutically acceptable metal cation, with ammonia, or with a pharmaceutically acceptable organic primary, secondary or tertiary amine. Representative alkali or alkaline earth salts include the lithium, sodium, potassium, calcium, magnesium, and aluminum salts and the like. Illustrative examples of some of the bases that can be used include sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate, N+(C^ alkyl)4, and the like. Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine and the like. Also provided are salts wherein one or more basic nitrogen-containing groups are quarternized. Water or oil-soluble or dispersible products may be obtained by such quaternization. See, for example, Berge et al., supra.
[0032] The term "alkyl," alone or in combination, refers to an optionally substituted straight- chain, or optionally substituted branched-chain saturated hydrocarbon radical having from one to thirty carbons, such as from one to twelve carbons. Examples of alkyl radicals include methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, tert-butyl, tert-amyl, pentyl, hexyl, heptyl, octyl and the like. A "lower alkyl" is an alkyl having from one to six carbons.
[0033] The term "alkylene" refers to an optionally substituted straight-chain, or optionally substituted branched-chain saturated hydrocarbon radical having from one to thirty carbons, such as from one to twelve carbons, and having two points of attachment. Alkylene groups will usually have from 1 to 20 carbon atoms, for example 1 to 8 carbon atoms, such as from 1 to 6 carbon atoms. For example, C1 alkylene is a methylene group (-CH2-). A "lower alkylene" is an alkylene having from one to six carbons. [0034] The term "alkenyl," alone or in combination, refers to an optionally substituted straight- chain, or optionally substituted branched-chain hydrocarbon radical having one or more carbon-carbon double-bonds and having from two to thirty carbon atoms, such as two to eighteen carbons. Examples of alkenyl radicals include ethenyl, propenyl, butenyl, 1 ,3-butadienyl and the like. A "lower alkenyl" refers to an alkenyl having from two to six carbons.
[0035] The term "alkynyl," alone or in combination, refers to an optionally substituted straight- chain or optionally substituted branched-chain hydrocarbon radical having one or more carbon-carbon triple-bonds and having from two to thirty carbon atoms, such as from two to twelve carbon atoms, from two to six carbon atoms as well as those having from two to four carbon atoms. EΞxamples of alkynyl radicals include ethynyl, 2-propynyl, 2-butynyl, 1 ,3-butadiynyl and the like. A "lower alkynyl" refers to an alkynyl having from two to six carbons.
[0036] The term "carbon chain" embraces any alkyl, alkenyl, or alkynyl group, which is linear, cyclic, or any combination thereof. If the chain is part of a linker and that linker comprises one or more rings as part of the core backbone, for purposes of calculating chain length, the "chain" only includes those carbon atoms that compose the bottom or top of a given ring and not both, and where the top and bottom of the ring(s) are not equivalent in length, the shorter distance shall be used in determining the chain length.
[0037] The term "membered ring" can embrace any cyclic structure, including aryl, heteroaryl, alicyclic, heterocyclic and polycyclic fused ring systems as described below. The term "membered" is meant to denote the number of skeletal atoms that constitute the ring. Thus, for example, pyridine, pyran, and pyhmidine are six-membered rings and pyrrole, tetrahydrofuran, and thiophene are five-membered rings.
[0038] The term "optionally substituted ring" can embrace any any cyclic structure, including optionally substituted aryl, optionally substituted heteroaryl, optionally substituted alicyclic, optionally substituted heterocyclic and optionally substituted polycyclic fused ring systems as described below. [0039] The term "aryl," alone or in combination, refers to an optionally substituted aromatic hydrocarbon radical of six to twenty ring atoms, and includes mono-aromatic rings and fused aromatic rings. A fused aromatic ring radical contains from two to four fused rings where the ring of attachment is an aromatic ring, and the other individual rings within the fused ring may be aromatic, heteroaromatic, alicyclic or heterocyclic. Further, the term aryl includes mono-aromatic ring and fused aromatic rings containing from six to twelve carbon atoms, as well as those containing from six to ten carbon atoms. Examples of aryl groups include, without limitation, phenyl, naphthyl, anthryl, chrysenyl, and benzopyrenyl ring systems.
[0040] The term "heteroaryl" refers to optionally substituted aromatic radicals containing from five to twenty skeletal ring atoms and where one or more of the ring atoms is a heteroatom such as, for example, oxygen, nitrogen, sulfur, selenium and phosphorus. The term heteroaryl includes optionally substituted mono-heteroaryl radicals and fused heteroaryl radicals having at least one heteroatom (e.g., quinoline, benzothiazole). A fused heteroaryl radical may contain from two to four fused rings and where the ring of attachment is a heteroaromatic ring, the other individual rings within the fused ring system may be aromatic, heteroaromatic, alicyclic or heterocyclic. The term heteroaryl also includes mono- heteroaryls or fused heteroaryls having from five to twelve skeletal ring atoms, as well as those having from five to ten skeletal ring atoms. Examples of heteroaryls include, without limitation, furanyl, benzofuranyl, chromenyl, pyridyl, pyrrolyl, indolyl, quinolinyl, pyridyl-N-oxide, pyrimidyl, pyrazinyl, imidazolyl, pyrazolyl, oxazolyl, isoxazolyl, benzothiozolyl, benzimidazolyl, benzoxazolyl, benzothiadiazolyl, benzoxadiazolyl, benzotriazolyl, quinolinyl, isoquinolinyl, indolyl, purinyl, indolizinyl, thienyl and the like and their oxides.
[0041] The term "alicyclic" alone or in combination, refers to an optionally substituted saturated or unsaturated nonaromatic hydrocarbon ring system, including both monocyclic and multicyclic ring systems, including fused ring systems, containing from three to twenty ring atoms. Monocyclic alicylic groups typically have from 3 to about 8 carbon ring atoms or from 3 to about 7 carbon ring atoms. Multicyclic alicyclic groups may have 2 or 3 fused rings or contain bridged or caged groups. A fused alicyclic radical contains from two to four fused rings where the ring of attachment is an alicyclic ring, and the other individual rings within the fused ring may be aromatic, heteroaromatic, alicyclic or heterocyclic. Alicyclic substituents may be pendant from a substituted nitrogen or carbon atom, or a substituted carbon atom that may have two substituents may have an alicyclic group, which is attached as a spiro group. Examples of alicylic groups include cyclopropyl, cyclopropenyl, cyclobutyl, cyclopentyl, or cyclohexyl as well as bridged or caged saturated ring groups such as norbornyl or adamantyl. [0042] The term "heterocyclic" refers to optionally substituted saturated or unsaturated nonaromatic ring radicals containing from five to twenty ring atoms where one or more of the ring atoms are heteroatoms such as, for example, oxygen, nitrogen, sulfur, and phosphorus. The term heterocyclic includes mono-heterocyclic and fused heterocyclic ring radicals, as well as spiro-heterocyclic ring radicals. A fused heterocyclic radical may contain from two to four fused rings where the attaching ring is a heterocyclic, and the other individual rings within the fused heterocyclic radical may be aromatic, heteroaromatic, alicyclic or heterocyclic. The term heterocyclic also includes mono-heterocyclic and fused heterocyclic radicals having from five to twelve skeletal ring atoms, as well as those having from five to ten skeletal ring atoms. Example of heterocyclics include without limitation, tetrahydrofuranyl, benzodiazepinyl, tetrahydroindazolyl, dihydroquinolinyl, and the like.
[0043] The term "arylalkyl," alone or in combination, refers to an alkyl radical as defined above in which one H atom is replaced by an aryl radical as defined above, each of which may be optionally substituted. Examples of arylalkyl groups include benzyl, 2-phenylethyl and the like. [0044] The term "heteroarylalkyl" refers to an alkyl radical as defined above in which one H atom is replaced by a heteroaryl radical as defined above, each of which may be optionally substituted. [0045] The term "alicyclicalkyl" refers to an alkyl radical as defined above in which one H atom is replaced by an alicyclic radical as defined above, each of which may be optionally substituted. [0046] The term "heterocyclicalkyl" refers to an alkyl radical as defined above in which one H atom is replaced by a heterocyclic radical as defined above, each of which may be optionally substituted. [0047] The term "alkoxy," alone or in combination, refers to an alkyl ether radical, alkyl-O-, wherein the term alkyl is defined as above and wherein the alkyl group may be optionally substituted.
EΞxamples of alkoxy radicals include methoxy, ethoxy, n-propoxy, iso-propoxy, n-butoxy, iso-butoxy, sec- butoxy, tert-butoxy and the like. A "lower alkoxy" is an alkoxy having from one to six carbons.
[0048] The term "aryloxy," alone or in combination, refers to an aryl ether radical wherein the term aryl is defined as above and wherein the aryl group may be optionally substituted. Examples of aryloxy radicals include phenoxy and the like.
[0049] The term "alkylthio," alone or in combination, refers to an alkyl thio radical, alkyl-S-, wherein the term alkyl is as defined above.
[0050] The term "arylthio," alone or in combination, refers to an aryl thio radical, aryl-S-, wherein the term aryl is as defined above.
[0051] The term "heteroarylthio" refers to the group heteroaryl-S-, wherein the term heteroaryl is as defined above.
[0052] The term "acyl" refers to a radical -C(O)R where R includes alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl or heteroarylalkyl wherein the alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl or heteroarylalkyl groups may be optionally substituted.
[0053] The term "acyloxy" refers to the ester group -OC(O)R, where R is H, alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl, or heteroarylalkyl wherein the alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl or heteroarylalkyl may be optionally substituted.
[0054] The term "carboxyesters" refers to -C(O)OR where R is alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, heteroarylalkyl or arylalkyl, wherein each is optionally substituted.
[0055] The term "BOC" refers to -C(O)Otbutyl.
[0056] The term "carboxamido" refers to
where each of R and R' are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, heteroaryl, alicyclic, heterocyclic, arylalkyl and heteroarylalkyl, each of which, other than the hydrogen, being optionally substituted.
[0057] The term "oxo" refers to =0.
[0058] The term "halogen" refers to F, Cl, Br and I.
[0059] The terms "haloalkyl, haloalkenyl, haloalkynyl and haloalkyloxy" include alkyl, alkenyl, alkynyl and alkoxy structures, as described above, that are substituted with one or more fluorines, chlorines, bromines or iodines, or with combinations thereof.
[0060] The terms "perhaloalkyl, perhaloalkyloxy and perhaloacyl" refer to alkyl, alkyloxy and acyl radicals as described above, wherein all the H atoms are substituted with fluorines, chlorines, bromines or iodines, or combinations thereof. "Perhaloalkyl," "perhaloalkyloxy," and "perhaloacyl" are species, respectively, of "haloalkyl," "haloalkyloxy," and "haloacyl". [0061] The term "amino" refers to the group -NH2.
[0062] The term "alkylamino", refers to the group -NHR where R is independently selected from alkyl, arylalkyl, and heteroarylalkyl.
[0063] The term "dialkylamino", refers to the group -NRR' where R and R' are independent alkyl, arylalkyl, and heteroarylalkyl.
[0064] The term "aminocarbonyl" refers to the group -C(O)NRR' where R and R' are independently selected from hydrogen, alkyl, alkenyl, alkynyl, aryl, alicyclic, heteroaryl, heterocyclic, hydroxy, alkoxy, amino, alkylamino, dialkylamino, and carboxamido, and where R and R' are optionally joined together with the nitrogen bound thereto to form a heterocyclic or substituted heterocyclic group. [0065] The terms "optional" or "optionally" mean that the subsequently described event or circumstance may but need not occur, and that the description includes instances where the event or circumstance occurs and instances in which it does not. For example, "aryl optionally mono- or di- substituted with an alkyl" means that the alkyl may but need not be present, or either one alkyl or two may be present, and the description includes situations where the aryl is substituted with one or two alkyls and situations where the aryl is not substituted with an alkyl.
[0066] "Optionally substituted" groups may be substituted or unsubstituted. Unless stated otherwise, optionally substituted alkyl, alkylene, alkenyl and alkynyl refer to alkyl, alkylene, alkenyl or alkynyl radicals, as defined herein, that may be optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) independently selected from alkyl, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted by one, two, or three groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three alkyl groups, heteroarylalkyl, -C(=E)R,, -Ct=E)N(RO(Ri), -Cf=E)N(R1)S(O)2R1, -Ct=E)N(R1)N(R1)S(O)2R1, -C(R1J=N(OR1), -C(R1J=NN(RK)(R1), -C≡N, -C(=E)OR,, -Cf=E)SR1, -N(Rk)(R1), -N(R1)Cf=E)R1, -N(R1)Cf=E)N(RO(R1), -N(R1)Cf=E)OR1, -N(R1)Cf=E)SR1, -N(R1)S(O)2R1, -NR1S(O)2OR1, -S(O)1OR1 (where t is 1 or 2),
-S(O)NRkR|, -NH-C(-NH2)=C-NO2, -CO-NH-SO2-NH2, -N(R1)S(O)2N(RO(R,), -N3, -NO2, -OR1, -OC(=E)R,, -OCf=E)OR1, -OCf=E)N(RO(Ri), -Oq=E)SR1, -OP(O)(Rh)2, -OS(O)2R1, -OS(O)2N(RO(R1), -OSi(R1J3, -SR1, -S(O)1R, (where t is 1 or 2), -S(O)2N(RO(Ri), -P(O)(Rh)2, -Si(R1J3, and halogen, wherein each E is independently O, NR9 or S; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR1 or -N(RO(Ri); each R, is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R1 is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alkenyl, alkynyl, optionally substituted alicyclic, alicyclicalkyl, optionally substituted heterocyclic, heterocyclicalkyl, optionally substituted aryl, arylalkyl, optionally substituted heteroaryl, heteroarylalkyl, or hydroxy; or Rk and R|, together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclic or optionally substituted heteroaryl.
[0067] In some embodiments, optionally substituted alkyl, alkylene, alkenyl and alkynyl refer to alkyl, alkylene, alkenyl or alkynyl radicals, as defined herein, that may be optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) independently selected from alicyclic, heterocyclic, heteroaryl optionally substituted by one, two, or three alkyl groups, -C(=E)R,, -C(=E)N(Rk)(R|), -C(=E)N(R,)S(O)2R|, -CC=E)N(R1)N(R1)S(O)2R1, -C≡N, -Cf=E)OR1, -C(=E)SR,, -N(R11)(R1), -N(R1)Cf=E)R1, -N(R1)Cf=E)N(RO(R1), -N(R1)Ct=E)OR1, -N(R1)Ct=E)SR1, -N(R1)S(O)2R1, -N(R1)S(O)2N(RO(Ri), -OR1, -OCf=E)R1, -OCf=E)OR1, -OCf=E)N(ROfRi), -OCf=E)SR1, -SR1, -S(O)1R1 (where t is 1 or 2), S(O)2N(RO(Ri), and halogen, wherein each E is independently O, NR9 or S; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR1 or -NfRO(Ri); each R1 is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R1 is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alkenyl, alkynyl, optionally substituted alicyclic, alicyclicalkyl, optionally substituted heterocyclic, heterocyclicalkyl, optionally substituted aryl, arylalkyl, optionally substituted heteroaryl, heteroarylalkyl, or hydroxy; or Rk and R|, together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclic or optionally substituted heteroaryl.
[0068] In some embodiments, optionally substituted alkyl, alkylene, alkenyl and alkynyl refer to alkyl, alkylene, alkenyl or alkynyl radicals, as defined herein, that may be optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) independently selected from alicyclic, heterocyclic, heteroaryl optionally substituted by one, two, or three alkyl groups, -Cf=E)R1, -C(=E)N(R0(Rι), -C≡N, -Cf=E)OR,, -N(RO(R1), -NfR1)Cf=E)R1, -N(R1)Cf=E)N(RO(R1), N(R1)Cf=E)OR1, -OR1, -OCf=E)R1, -OCf=E)OR1, -OC(=E)N(Rk)(Rι), and halogen, wherein each E is independently O, NR9 or S; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -ORj or -N(Rk)(Ri); each R1 is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R1 is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alkenyl, alkynyl, optionally substituted alicyclic, alicyclicalkyl, optionally substituted heterocyclic, heterocyclicalkyl, optionally substituted aryl, arylalkyl, optionally substituted heteroaryl, heteroarylalkyl, or hydroxy; or Rk and R|, together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclic or optionally substituted heteroaryl. [0069] In some embodiments, optionally substituted alkyl, alkylene, alkenyl and alkynyl refer to alkyl, alkylene, alkenyl or alkynyl radicals, as defined herein, that may be optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) independently selected from alicyclic, heterocyclic, heteroaryl optionally substituted by one, two, or three alkyl groups, -Cf=E)R1, -C(=E)N(Rk)(R|), -C≡N, -C(=E)OR,, -N(Rk)(Ri), -N(R1)Cf=E)R1, -N(R1)C(=E)N(Rk)(R,), -N(R))C(=E)OR1, -OR1, -OC(=E)Rj, -OC(=E)N(Rk)(R,), -OC(=E)OR|, and halogen, wherein each E is independently O, NR9 or S; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR1 or -N(Rk)(Rι); each R1 is independently alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R, is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, optionally substituted alicyclic, alicyclicalkyl, optionally substituted heterocyclic, heterocyclicalkyl, optionally substituted aryl, arylalkyl, optionally substituted heteroaryl, heteroarylalkyl, or hydroxy; or Rk and Rh together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclic or optionally substituted heteroaryl.
[0070] In some embodiments, optionally substituted alkyl, alkylene, alkenyl and alkynyl refer to alkyl, alkylene, alkenyl or alkynyl radicals, as defined herein, that may be optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) independently selected from alicyclic, heterocyclic, heteroaryl optionally substituted by one, two, or three alkyl groups, -C(=E)R,, -C(=E)N(Rk)(R|), -C≡N, -C(=E)OR,, and halogen, wherein each E is independently O, NR9 or S; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR, or -N(Rk)(R|); each R, is independently alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rj is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, optionally substituted alicyclic, alicyclicalkyl, optionally substituted heterocyclic, heterocyclicalkyl, optionally substituted aryl, arylalkyl, optionally substituted heteroaryl, heteroarylalkyl, or hydroxy; or Rk and Rh together with the nitrogen atom to which they are attached, form an optionally substituted heterocyclic or optionally substituted heteroaryl. [0071] Unless stated otherwise, "optionally substituted aryl", including "optionally substituted phenyl", "optionally substituted alicyclic", "optionally substituted heteroaryl", and "optionally substituted heterocyclic" refer to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted by one, two, or three groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three alkyl groups, heteroarylalkyl, -RrCt=E)R,, -RrC(=E)N(Rk)(R,), -RrC(=E)N(R,)S(O)2R,, -RrCC=E)N(R1)N(R1)S(O)2R1, -RrC(R1J=N(OR1),
Figure imgf000015_0001
-R,-C(=E)OR1, -R,-C(=E)SR1, -R,-N(Rk)(R,), -RrN(R1)C(=E)R1, -RrN(R1)Cf=E)N(Rk)(R1), -RrN(R1)Ct=E)OR1, -RrN(R1)C(=E)SR1, -RrN(R1)S(O)2R1, -RrN(R1)S(O)2OR1, - RfS(O)1OR1 (where t is 1 or 2), -R,-SO-NRkR|, -RrNH-C(-NH2)=C-NO2, -RrCO-NH-SO2-NH2, -RrN(R1)S(O)2N(Rk)(R1), -RfN3, -RfN02, -RrOR1, -Rf-OC(=E)R1, -RrOCt=E)OR1, -Rf-0C(=E)N(Rk)(R|), -Rf-OC(=E)SR,, -RrOP(O)(Rh)2, -RrOS(O)2R1, -Rf-OS(O)2N(Rk)(R,), -RrOSi(R1)S, -RrP(O)(Rh)2, -R.-SR,, -Rf-S(O)|R| (where t is 1 or 2), -RrS(O)2N(Rk)(R|), -RrSi(R1J3, and halogen, wherein each E is independently O, NR9 or S; each R1 is independently alkylene or a direct bond; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR1 or -N(Rk)(R|); each R, is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rj is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, or hydroxy; or Rk and R|, together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl.
[0072] In some embodiments, "optionally substituted aryl", including "optionally substituted phenyl", "optionally substituted alicyclic", "optionally substituted heteroaryl", and "optionally substituted heterocyclic" refer to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three alkyl groups, heteroarylalkyl, -RrCf=E)R1, -RrCf=E)N(RK)(R,), -RrCt=E)N(R1)S(O)2R1, -Rf-C(=E)N(RJ)N(RJ)S(O)2RI, -R1CsN1 -RrCt=E)OR1, -RrCf=E)SR,, -Rf-N(Rk)(R,), -RrN(R,)C(=E)Rl, -RrN(RJ)C(=E)N(Rk)(Rl), -RrN(RJ)C(=E)ORJ, -RrN(R,)C(=E)SR,, -RrN(R1)S(O)2R1, -RrN(R1)S(O)2N(RO(R1), -R1NO2, -RrOR1, -RrOC(=E)RJ, -Rf-OC(=E)OR1, -R,-OC(=E)N(Rk)(Rι), -Rf-OC(=E)SR,, -RrSR,, -R,-S(O),R, (where t is 1 or 2), -Rf-S(O)2N(Rk)(R|), and halogen, wherein each E is independently O, NR9 or S; each Rf is independently alkylene or a direct bond; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR1 or -N(Rk)(R|); each R1 is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R, is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, or hydroxy; or Rk and R|, together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl. [0073] In some embodiments, "optionally substituted aryl", including "optionally substituted phenyl", "optionally substituted alicyclic", "optionally substituted heteroaryl", and "optionally substituted heterocyclic" refer to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three alkyl groups, heteroarylalkyl, -RrCf=E)R,, -RrC(=E)N(Rk)(Rι), -R1C=N, -RrC(=E)ORJ, -RrN(RO(Ri), -R,-N(RJ)C(=E)RJ, -RrN(R,)C(=E)N(RO(Rι), -R,-N(RI)C(=E)ORJ, -RfNO2, -RrOR1, -Rf-OC(=E)R,, -Rf-OC(=E)ORJ, -Rf-OC(=E)N(Rk)(R|), and halogen, wherein each E is independently O, NR9 or S; each Rf is independently alkylene or a direct bond; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -OR, or -N(Rk)(R|); each R1 is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R1 is independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy, hydroxy, and amino, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen , arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and R| are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, or hydroxy; or Rk and Rh together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl.
[0074] In some embodiments, "optionally substituted aryl", including "optionally substituted phenyl", "optionally substituted alicyclic", "optionally substituted heteroaryl", and "optionally substituted heterocyclic" refer to aryl, including phenyl, alicyclic, heterocyclic, and heteroaryl radicals, respectively, as defined herein, that are optionally substituted by one or more substituents (e.g., by one to six substituents, such as one to four substituents, for example, one or two substituents, such as one substituent) selected from alkyl optionally substituted by amino, haloalkyl, haloalkyloxy, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted by one, two, or three halogen atoms, arylalkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, heteroaryl optionally substituted by one, two, or three alkyl groups, heteroarylalkyl, -RrC(=E)R,, -RrC(=E)N(RO(Rι). -RfC≡N, -RrCt=E)OR1, -R,-N(Rk)(R,), -RrN(RJ)C(=E)RJ, -Rf-N(Rl)C(=E)N(Rk)(Rl), -Rf-N(R,)C(=E)OR,, -RfNO2, -Rf-OR1, -RrOC(=E)R,, -R,-OC(=E)N(Rk)(R|), -R,-OC(=E)OR,, and halogen, wherein each E is independently O, NR9 or S; each R1 is independently alkylene or a direct bond; each R9 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rh is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -0R| or -N(Rk)(R|); each R1 is independently alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each R, is independently hydrogen, alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl optionally substituted with one to five groups independently chosen from alkyl and halogen, arylalkyl, heteroaryl, or heteroarylalkyl; and Rk and Ri are each independently hydrogen, alkyl optionally substituted with one or two groups independently chosen from alkoxy and hydroxy, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, or hydroxy; or Rk and R|, together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl.
[0075] Unless stated otherwise specifically in the specification, it is understood that the substitution can occur on any atom of the alkyl, alkylene, alkenyl, alkynyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, alicyclic, alicyclicalkyl, heterocyclic, and heterocyclicalkyl groups. It will be understood by those skilled in the art, with respect to any group containing one or more substituents, that such groups are not intended to introduce any substitution or substitution patterns that are sterically impractical, synthetically non-feasible and/or inherently unstable.
[0076] Optionally substituted alicyclic and optionally substituted heterocyclic may additionally be substituted with oxo, thiono, imino, oxime or hydrazone, on a saturated carbon of their respective ring system.
[0077] Optionally substituted heteroaryl wherein the heteroatom(s) is/are nitrogen may additionally be substituted by oxo on the nitrogen atom of the ring system.
[0078] The term "substituted amino" refers to the group -NHRd or -NRdRe wherein Rd is selected from hydroxy, optionally substituted alkoxy, optionally substituted alkyl, optionally substituted alicyclic, optionally substituted acyl, optionally substituted carbamoyl, aminocarbonyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclic, optionally substituted alkoxycarbonyl, and sulfonyl, and Re is selected from optionally substituted alkyl, optionally substituted alicyclic, optionally substituted aryl, optionally substituted heteroaryl, and optionally substituted heterocyclic, and wherein substituted alkyl, substituted alicyclic, substituted aryl, substituted heteroaryl, and substituted heterocyclic are as described herein. The term "substituted amino" also refers to N-oxides of the groups -NHRd, and NRdRe each as described above. N-oxides can be prepared by treatment of the corresponding amino group with, for example, hydrogen peroxide or m-chloroperoxybenzoic acid. The person skilled in the art is familiar with reaction conditions for carrying out the N-oxidation. [0079] The term "pyridine-1 -oxy" also means "pyhdine-N-oxy."
[0080] The term "aminocarbonyl" refers to the group -CONRbRc, where Rb is selected from hydrogen, optionally substituted alkyl, optionally substituted alicyclic, optionally substituted heterocyclic, optionally substituted aryl, and optionally substituted heteroaryl; and Rc is independently selected from hydrogen and optionally substituted alkyl; or Rband Rc taken together with the nitrogen to which they are bound, form an optionally substituted 5- to 7-membered nitrogen-containing heterocyclic which optionally includes 1 or 2 additional heteroatoms selected from O, N, and S in the heterocyclic; wherein substituted alkyl, substituted alicyclic, substituted heterocyclic, substituted aryl, and substituted heteroaryl are as described herein. [0081] Some of the compounds described herein may contain one or more chiral centers and therefore may exist in enantiomeric and diastereomeric forms. Thus, the term "compound" is intended to cover all isomers per se, as well as mixtures of cis and trans isomers, mixtures of diastereomers and racemic mixtures of enantiomers (optical isomers) as well. Further, it is possible using well known techniques to separate the various forms, and some embodiments described herein may feature purified or enriched species of a given enantiomer or diastereomer.
[0082] A "pharmaceutical composition" refers to a mixture of one or more of the compounds or pharmaceutically acceptable salts thereof, described herein, with one or more pharmaceutically acceptable carriers. The purpose of a pharmaceutical composition is to facilitate administration of a compound to an organism.
[0083] The phrase "pharmaceutically acceptable carrier" as used herein means a pharmaceutically-acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting the subject agent from one organ, or portion of the body, to another organ, or portion of the body. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Some examples of materials which can serve as pharmaceutically-acceptable carriers include: (1) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethyl alcohol; (20) phosphate buffer solutions; and (21) other non-toxic compatible substances employed in pharmaceutical formulations. Pharmaceutically acceptable carriers include excipients and diluents.
[0084] The term "sulfonyl" refers to the groups: -S(O2)-OH, -S(O2)-(optionally substituted alkyl), -
S(O2)-(optionally substituted alicyclic), -S(O2)-(optionally substituted amino), -S(O2)-(optionally substituted aryl), -S(O2)-(optionally substituted heteroaryl), and -S(O2)-(optionally substituted heterocyclic). [0085] A "therapeutically effective amount" means an amount which is capable of providing a therapeutic effect. The specific dose of substance administered to obtain therapeutic effect will, of course, be determined by the particular circumstances surrounding the case, including, for example, the specific substance administered, the route of administration, the condition being treated, and the individual being treated. A typical daily dose (administered in single or divided doses) will contain a dosage level of from about 0.01 mg/kg to about 50-100 mg/kg of body weight of the active substance. In some embodiments, daily doses generally will be from about 0.05 mg/kg to about 20 mg/kg and ideally from about 0.1 mg/kg to about 10 mg/kg. Factors such as clearance rate, half-life and maximum tolerated dose (MTD) have yet to be determined but one of ordinary skill in the art can determine these using standard procedures.
[0086] In some embodiments, the therapeutic effect is the inhibition, to some extent, of the growth of cells characteristic of a proliferative disorder, e.g., a cancer. A therapeutic effect will also normally, but need not, relieve to some extent one or more of the symptoms other than cell growth or size of cell mass. A therapeutic effect may include, for example, one or more of 1) a reduction in the number of cells; 2) a reduction in cell size; 3) inhibition (i.e., slowing to some extent, or even stopping) of cell infiltration into peripheral organs, e.g., in the instance of cancer metastasis; 4) inhibition (i.e., slowing to some extent, or even stopping) of tumor metastasis; 5) inhibition, to some extent, of cell growth; and/or 6) relieving to some extent one or more of the symptoms associated with the disorder. In some embodiments, the therapeutic effect is the decrease, to some extent, in the metabolic rate of the tumor, as measured, e.g,. by 18F-glucose PET scan.
[0087] As used herein, the term IC50 refers to an amount, concentration or dosage of a particular test compound that achieves a 50% inhibition of a maximal response in an assay that measures such response. In some embodiments, the "IC50" value of a compound can be greater for normal cells than for cells exhibiting a proliferative disorder, e.g., cancer cells. The value depends on the assay used. [0088] By a "standard" is meant a positive or negative control. A negative control in the context of HER2 expression levels is, e.g., a sample possessing an amount of HER2 protein that correlates with a normal cell. A negative control may also include a sample that contains no HER2 protein. By contrast, a positive control does contain HER2 protein, for example, of an amount that correlates with overexpression as found in proliferative disorders, e.g., cancers. The controls may be from cell or tissue samples, or else contain purified ligand (or absent ligand), immobilized or otherwise. In some embodiments, one or more of the controls may be in the form of a diagnostic "dipstick." [0089] By "selectively targeting" is meant affecting one type of cell to a greater extent than another, e.g., in the case of cells with high as opposed to relatively low or normal HER2 levels. [0090] Provided is a compound represented by Formula I
Figure imgf000020_0001
or a pharmaceutically acceptable salt thereof, wherein: X is a bond or an alkylene chain of the formula (CRaRb)n where n is 1 to 3, for each occurrence, Ra and Rb are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which, except for hydrogen, is optionally substituted, and a (CRaRb) unit is optionally replaced by -O-, -N(Ra)-, or -S-; or when n is 2 or 3, Ra in a first (CRaRb) unit is optionally taken together with Ra in a different (CRaRb) unit to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or
Ra is optionally taken together with Rb in the same (CRaRb) unit to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; R1 is selected from hydrogen, halogen, -OR11, -SR12, amino, and optionally substituted lower alkyl; or
R2 is selected from -NR21R22, -N=C-NR21R22, -NR21-C(O)R23 and -NR21-SO2R24; R3 and R4 are independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alicyclic, optionally substituted alicyclicalkyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -C(=J)RX, -C(=J)N(Ry)(Rz), Cf=J)N(Rx)S(O)2Rx, -C(=J)N(RX)N(RX)S(O)2RX, -C(RX)=N(ORX), -C(Rx)=NN(Ry)(R1), -CSN, -C(=J)ORX, -C(=J)SRX, -N(Ry)(R2), -N(RX)C(=J)RX, -N(Rx)C(=J)N(Ry)(Rz), -N(RX)C(=J)ORX, -N(RX)C(=J)SRX, -N(Rx)S(O)2R*,, -N(Rx)S(O)2ORx, -S(O)1ORx (where t is 1 or 2), - S(O)NRyR2, -NH-C(-NH2)=C-NO2, -CO-NH-SO2-NH2, -N(Rx)S(0)2N(Ry)(Rz), N3, NO2, -ORx, -OC(=J)RX, -OC(=J)ORX, -OC(=J)N(Ry)(Rz), -OC(=J)SRX, -OP(O)(Rv)2, -OS(O)2Rx, -OS(O)2N(Ry)(Rz), - OSi(Rw)3, -P(O)(RV)2, -SRx, -S(O)1Rx (where t is 1 or 2), -S(O)2N(Ry)(Rz), -Si(RJ3, and halogen, wherein each J is independently O, NRU or S wherein each Ru is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rv is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -ORx or -N(Ry)(Rz); each Rw is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rx is independently hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; and Ry and Rz are each independently hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; or Ry and Rz, together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl; or
R3 is optionally taken together with R4 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, n is 1 , and the (CRaRb) unit is replaced by -N(Ra)-, then R3 and R4 taken together optionally form oxo; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, an Ra unit is optionally taken together with R1 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, an Ra in a first (CRaRb) unit is optionally taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is a bond, R1 is optionally taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90;
R5 and R6 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; or
R5 is optionally taken together with R6, to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n , n is 1 , and the (CRaRb) unit is replaced by -N(Ra)-, then R5 and R6 taken together optionally form oxo; or
R3 is optionally taken together with R5, to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90;
R7 is selected from a bond, optionally substituted alkylene, -C(O)-, -C(S)-, -S(O)-, and -SO2-;
R8 is selected from alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which is optionally substituted;
R11 and R12 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; and
R21 and R22 are independently selected from hydrogen, alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl, each of which, except for hydrogen, is optionally substituted, or R21 is optionally taken together with R22 to form an optionally substituted ring of 3-8 ring atoms wherein said optionally substituted ring optionally includes one or two additional ring atoms that are heteroatoms independently selected from O, S, =N, and NR90;
R23 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
R24 is selected from alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and
R90 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and wherein from 1 to 5 hydrogen atoms in the compound of Formula I are optionally replaced with deuterium, and provided that when X is an optionally substituted alkylene chain of the formula (CRaRb)n , n is 1 , the (CRaRb) unit is replaced by -N(R3)-, and R7 is -CH2- then R8 is not 3,5-dimethyl-4-methoxy-pyridin-2-yl. [0091] In some embodiments, X is selected from -O-, CH2, -CH2N(Ra)-, and -N(Ra)-, or X is a bond.
[0092] In some embodiments, R1 is halogen selected from fluoro, chloro, and bromo. In some embodiments, R1 is chloro.
[0093] In some embodiments, X is a bond and R1 and R3, taken together with the atoms to which they are attached, form an optionally substituted ring of 3-8 ring atoms wherein one of the 3-8 ring atoms optionally is O or S.
[0094] In some embodiments, R21 is hydrogen.
[0095] In some embodiments, R22 is hydrogen.
[0096] In some embodiments, R3 is selected from hydrogen, optionally substituted alkyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, and halogen.
[0097] In some embodiments, R3 is lower alkyl. In some embodiments, R3 is methyl. In some embodiments, R3 is f-butyl.
[0098] In some embodiments, R3 is alkyl substituted with one or two groups selected from
-OR31 wherein R31 is selected from hydrogen, alkyl, trialkylsilyl, and -C(O)R40 wherein R40 is selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl, and heteroaryl; -S(O)nR32 or -OS(O)2R32 wherein R32 is selected from alkyl, phenyl, and -NR51R52 wherein R51 and R52 are independently selected from hydrogen and alkyl, and n is 1 or 2; -NR33R34 wherein R33 and R34 are independently selected from hydrogen, lower alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, -C(O)R60 wherein R60 is selected from hydrogen, alkyl, aryl, and amino, and -S(O)2-alkyl or R33 and R ,34 taken together with the nitrogen to which they are bound form an optionally substituted heterocyclic or optionally substituted heteroaryl ring; -C(O)YR35 wherein Y is selected from -NR36 and O and wherein R35 and R36 are independently selected from hydrogen, alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, alicyclic, heterocyclic, aryl, and heteroaryl or R35 and R36 optionally join together to form an optionally substituted 5-6 membered heteroaryl or optionally substituted 5-6 membered heterocyclic ring; halogen; azido; and cyano.
[0099] In some embodiments, R3 is alkyl substituted with one or two groups selected from
-OR31 wherein R31 is selected from hydrogen, alkyl, and -C(O)R40 wherein R40 is selected from hydrogen, alkyl, and aryl;
-OS(O)2R32 wherein R32 is selected from alkyl and phenyl; -NR33R34 wherein R33 and R34 are independently selected from hydrogen, lower alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, -C(O)R60 wherein R60 is selected from hydrogen, alkyl, aryl, and amino, and -S(O)2-alkyl or R33 and R34 taken together with the nitrogen to which they are bound form an optionally substituted heterocyclic or optionally substituted heteroaryl ring; -C(O)YR35 wherein Y is selected from -NR36 and O and wherein R35 and R36 are independently selected from hydrogen, alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, alicyclic, heterocyclic, aryl, and heteroaryl; and azido.
[00100] In some embodiments, R3 is selected from -CH2-OH, -CH2OCH3, -CH2CH2-OH, -
CH(OH)CH3, -C(CHa)2CH2OH, -CH2-OC(O)CH3, -CH2-NH2, -CH2-NHCH3, -CH2N(CH3)2, -CH2CH2NH2, - CH2-NHCH2CH2OCH3, -CH2-N(CH2CH2OCH3)2, -CH2-N(CH2CH2OH)2, -CH2-NHC(O)CH3, -CH2- NHC(O)Ph, -CH2-NHC(O)NH2, -CH2-NHS(O)2CH3, -CH2-C(O)OH, -CH2-C(O)OCH3, -CH2C(O)NH2, and - CH2OS(O)2CH3.
[00101] In some embodiments, R3 is alkyl substituted with -NR33R34 wherein R33 and R34 optionally taken together with the nitrogen to which they are bound form an optionally substituted heterocyclic or optionally substituted heteroaryl ring selected from isoindolinyl, triazolyl, and piperidinyl. In some embodiments, the optional substitutents on the isoindolinyl or triazolyl rings are independently selected from oxo and -CH2N(CH3J2.
[00102] In some embodiments, R3 is alkyl substituted with -C(O)YR35 wherein Y is -NR36 and wherein R35 and R36 are independently selected from hydrogen and alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy. [00103] In some embodiments, R3 is selected from -CH2C(O)NH2, -CH2C(O)NHCH3,
CH2C(O)NHCH2CH3, -CH2C(O)N(CH3)2, -CH2C(O)NHCH2CH2OCH3, -CH2C(O)NHCH2CH(OH)CH2OH, -
CH2C(O)NHCH(CH3)CH2OH, and -CH2CH2C(O)NH2.
[00104] In some embodiments, R3 is alkyl substituted with -C(O)YR35 wherein Y is -NR36 and wherein R35 and R36 join together to form an optionally substituted heterocyclic ring. In some embodiments, the optionally substituted heterocyclic ring is selected from morpholinyl, azetidinyl, pyrrolidinyl, and piperidinyl, each of which is optionally substituted. In some embodiments, the substituents on the morpholinyl, azetidinyl, pyrrolidinyl, and piperidinyl rings are selected from -CH2OH, -
CH2CH2OH, -C(O)NH2, and -OH.
[00105] In some embodiments, R3 is alkyl substituted -C(O)YR35 wherein Y is -NR36 and wherein
,36
R and R are independently selected from hydrogen, optionally substituted alicyclic ring, and optionally substituted heterocyclic ring.
[00106] In some embodiments, R35 and R36 are independently selected from hydrogen, optionally substituted cyclopropanyl, and optionally substituted oxetanyl.
[00107] In some embodiments, R3 is selected from optionally substituted phenyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, andoptionally substituted heteroarylalkyl.
[00108] In some embodiments, R is selected from:
Figure imgf000025_0001
Figure imgf000026_0001
[00109] In some embodiments, R4 is selected from hydrogen and lower alkyl. In some embodiments, R4 is hydrogen.
[00110] In some embodiments, R3 and R4, taken together with the carbon to which they are attached, form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of those ring atoms are heteroatoms independently selected from O, S, =N, and NR90. In some embodiments, R3 and
R4, taken together with the carbon to which they are attached, form a piperidinyl ring optionally substituted with one or two groups selected from acyl and arylalkyl. In some embodiments, R3 and R4, taken together with the carbon to which they are attached, form a piperidinyl ring substituted with -C(O)CH3 or benzyl. In some embodiments, R3 and R4, taken together with the carbon to which they are attached, form a piperidinyl ring wherein the nitrogen of the piperidinyl ring is substituted with -C(O)CH3 or benzyl.
[00111] In some embodiments, R3 and R5 taken together with the atoms to which they are attached form an optionally substituted ring of 3-8 ring atoms, optionally with 0-2 hetero ring atoms independently selected from O, S, =N, and NR90.
[00112] In some embodiments, R5 and R6 are independently selected from hydrogen and optionally substituted lower alkyl. In some embodiments, R5 is hydrogen. In some embodiments, R5 is -
CH2OH.
[00113] In some embodiments, R6 is hydrogen.
[00114] In some embodiments, R7 is a bond or optionally substituted alkylene. In some embodiments, R7 is optionally substituted lower alkylene. In some embodiments, R7 is -CH2-. In some embodiments, R7 is -CH(CH3)-.
[00115] In some embodiments, R8 is selected from alicyclic, aryl, heterocyclic, and heteroaryl, each of which is optionally substituted.
[00116] In some embodiments, R8 is phenyl, pyridinyl, pyrimidinyl, pyridazinyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, benzothiazolyl, pyrazolyl, thiazolyl, tetrahydroisoquinolinyl, imidazolyl, or benzodioxolyl, each of which is optionally substituted.
[00117] In some embodiments, R8 is selected from:
Figure imgf000027_0001
Figure imgf000027_0002
Figure imgf000027_0003
Figure imgf000028_0001
] In some embodiments, R8 is selected from:
Figure imgf000029_0001
Figure imgf000029_0002
Figure imgf000029_0003
Figure imgf000029_0006
Figure imgf000029_0004
Figure imgf000029_0007
Figure imgf000030_0001
Figure imgf000031_0001
Figure imgf000032_0001
[00119] In some embodiments, R is
Figure imgf000032_0002
Figure imgf000032_0003
In some embodiments, R8 is optionally substituted lower alkyl.
In some embodiments, from 1 to 5 hydrogen atoms of R8 are optionally replaced with
In some embodiments, R1 is chloro; R2 is -NH2; R3 is optionally substituted alkyl; R4 is hydrogen; R5 and R6 are hydrogen; R7 is -CH2-; and R8 is selected from optionally substituted aryl and optionally substituted heteroaryl. [00123] In some embodiments, the stereocenter to which R3 and R4 are attached is of the S- configuration.
[00124] In some embodiments, the stereocenter to which R3 and R4 are attached is of the R- configuration.
[00125] In some embodiments, the stereocenter to which R5 and R6 are attached is of the S- configu ration.
[00126] In some embodiments, the stereocenter to which R5 and R6 are attached is of the R- configuration.
[00127] Also provided is a compound of Formula Il
Figure imgf000033_0001
or a pharmaceutically acceptable salt thereof, wherein R1, R2, R3, R4, R5, R6, R7, and R8 are as described for compounds of Formula I.
[00128] Also provided is a compound of Formula III
Figure imgf000033_0002
or a pharmaceutically acceptable salt thereof, wherein R1, R2, R3, R4, R5, R6, R7, and R8 are as described for compounds of Formula I. [00129] Also provided are the following compounds, and pharmaceutically acceptable salts thereof,
Figure imgf000034_0001
Figure imgf000035_0001
Figure imgf000035_0003
Figure imgf000035_0002
Figure imgf000036_0001
Figure imgf000037_0001
Figure imgf000038_0001
Figure imgf000039_0001
Figure imgf000040_0001
Figure imgf000041_0001
Figure imgf000042_0001
Figure imgf000043_0001
Figure imgf000044_0001
Figure imgf000044_0002
Figure imgf000044_0003
Figure imgf000045_0001
Figure imgf000046_0001
Figure imgf000047_0001
Figure imgf000048_0001
Figure imgf000049_0001
Figure imgf000050_0001
Figure imgf000050_0002
Figure imgf000050_0004
Figure imgf000050_0003
Figure imgf000051_0001
Figure imgf000052_0001
Figure imgf000053_0001
Figure imgf000054_0001
Figure imgf000055_0001
Figure imgf000057_0001
Figure imgf000058_0001
Figure imgf000059_0001
or a pharmaceutically acceptable salt thereof.
[00130] Also provided is a compound selected from:
Figure imgf000059_0002
Figure imgf000059_0003
Figure imgf000059_0004
Figure imgf000060_0001
Figure imgf000061_0001
or a pharmaceutically acceptable salt thereof.
[00131] Also provided is a compound selected from:
Figure imgf000062_0001
Figure imgf000063_0001
Figure imgf000063_0002
or a pharmaceutically acceptable salt thereof.
[00132] The compounds and pharmaceutically acceptable salts described herein may be synthesized by various methods known in the art. See, e.g., U.S. Patent No. 7,138,402 and WO 2008/093075, each of which is incorporated herein by reference to the extent that it refers to preparation of compounds described therein. The chemical reagents are available commercially, e.g., from Aldrich Chemical Co., Milwaukee, Wis., USA. Otherwise their preparation is facile and known to one of ordinary skill in the art, or it is referenced or described herein. [00133] As will be apparent to those skilled in the art, conventional protecting groups may be necessary to prevent certain functional groups from undergoing undesired reactions. Suitable protecting groups for various functional groups as well as suitable conditions for protecting and deprotecting particular functional groups are well known in the art. For example, numerous protecting groups are described in T. W. Greene and P. G. M. Wuts, Protecting Groups in Organic Synthesis, Third Edition, Wiley, New York, 1999, and references cited therein.
[00134] If the compounds described herein contain one or more chiral centers, such compounds can be prepared or isolated as pure stereoisomers, i.e., as individual enantiomers or diastereomers, or as stereoisomer-enriched mixtures. All such stereoisomers (and enriched mixtures) are included within the scope of this invention, unless otherwise indicated. Pure stereoisomers (or enriched mixtures) may be prepared using, for example, optically active starting materials or stereoselective reagents well-known in the art. Alternatively, racemic mixtures of such compounds can be separated using, for example, chiral column chromatography, chiral resolving agents and the like.
[00135] Compounds of Formula I also include crystalline and amorphous forms of those compounds, including, for example, polymorphs, pseudopolymorphs, solvates (including hydrates), unsolvated polymorphs (including anhydrates), conformational polymorphs, and amorphous forms of the compounds, as well as mixtures thereof. "Crystalline form," "polymorph," and "novel form" may be used interchangeably herein, and are meant to include all crystalline and amorphous forms of the compound, including, for example, polymorphs, pseudopolymorphs, solvates (including hydrates), unsolvated polymorphs (including anhydrates), conformational polymorphs, and amorphous forms, as well as mixtures thereof, unless a particular crystalline or amorphous form is referred to. Likewise, pharmaceutically acceptable salts of compounds of Formula I also include crystalline and amorphous forms of those salts, including, for example, polymorphs, pseudopolymorphs, solvates (including hydrates), unsolvated polymorphs (including anhydrates), conformational polymorphs, and amorphous forms of the salts, as well as mixtures thereof. These crystalline forms may be prepared using methods known to the skilled artisan. For example, solvates may be prepared using standard low molecular weight solvents using methods known to the skilled artisan. See, e.g., Polymorphism in Pharmaceutical Solids, ed. Harry G. Brittan, Vol. 95, Marcel Dekker, Inc., New York, 1999 and more particularly, pages 202-208 which describe how hydrates and solvates of both organic and inorganic compounds are routinely prepared.
[00136] Compounds of Formula I also include different enriched isotopic forms, e.g., compounds enriched in the content of 2H, 3H, 11C, 13C and/or 14C. In some embodiments, the compounds are deuterated. Such deuterated forms can be made by the procedure described in U.S. Patent Nos. 5,846,514 and 6,334,997. As described in U.S. Patent Nos. 5,846,514 and 6,334,997, deuteration may improve the efficacy and increase the duration of action of drugs.
[00137] Deuterium substituted compounds can be synthesized using various methods such as described in: Dean, Dennis C; Editor. Recent Advances in the Synthesis and Applications of Radiolabeled Compounds for Drug Discovery and Development. [In: Curr. Pharm. Des., 2000; 6(10)] 2000, 110 pp.; Kabalka, George W.; Varma, Rajender S. The Synthesis of Radiolabeled Compounds via Organometallic Intermediates, Tetrahedron, 1989, 45(21), 6601-21 ; and Evans, E. Anthony. Synthesis of Radiolabeled Compounds, J. Radioanal. Chem., 1981 , 64(1-2), 9-32.
Figure imgf000065_0001
[00138] A general strategy for the synthesis of compounds of Formula I is outlined in Scheme 1.
Scheme 1 includes three methods to prepare compounds of Formula I and also shows that compounds of Formula I can be further elaborated at, for example R3, to generate additional species. It should be understood that other methods can be used to prepare compounds of Formula I as well. Scheme 2
Figure imgf000066_0001
[00139] Compounds of Formula I can be synthesized from compounds of Formula 1.1 according to Scheme 2. The compound of Formula 1.1 is, for example, reacted with phosphoryl chloride and benzyltriethylammonium chloride to yield a compound of Formula 1.2. The compound of Formula 1.2 is then alkylated using electrophiles such as LG-R7-R8, where LG is a leaving group, in a solvent such as dimethylsulfoxide or dimethylformamide optionally in the presence of a base such as sodium hydride or potassium carbonate. Leaving groups include, but are not limited to, e.g., mesylate, halogen, triflate, tosylate, etc.
Figure imgf000066_0002
[00140] The compounds of Formula 1.1 can be synthesized from aldehydes of Formula 2.1 and ketones of Formula 3.1 according to Scheme 3. Accordingly, a compound of Formula 2.1 wherein R3 is alkyl, is brominated with, for example 5,5-dibromobarbituric acid and hydrogen bromide in a solvent such as dichloromethane, to yield a compound of Formula 2.2. The brominated aldehyde of Formula 2.2 is then condensed with an amino substituted pyrimidine to yield a compound of Formula 2.3 which upon reduction results in a compound of Formula 1.1. [00141] Compounds of Formula 1.1 can also be synthesized from ketones of Formula 3.1 according to Scheme 3. Accordingly, a compound of Formula 3.1 wherein R3 and R4 are each independently alkyl or taken together with the carbon to which they are attached form a heterocyclic, is reacted with a diester, for example ethyl malonate, to yield a compound of Formula 3.2. Michael addition of nitromethane to the compound of Formula 3.2, followed by reduction of the nitro group to an amine results in a spontaneous cyclization to an α-carboxy-γ-lactam of Formula 3.4. The compound of Formula 3.4 is activated with, for example, triethyloxonium tetrafluoroborate in a solvent such as dichloromethane or with P2S5 to produce a compound of Formula 3.5 which is then treated with guanidine to yield a pyrrolopyrimidine-4-one of Formula 1.1.
Figure imgf000067_0001
[00142] An example of the syntheses outlined in Schemes 2 and 3 that proceeds through the diester of Formula 3.2 and yields a 4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro- pyrrolo[2,3-d]pyrimidine-2-amine of Formula I is shown in Scheme 4. Accordingly, as discussed in Scheme 3 above, an α-carboxy-γ-lactam of Formula 3.4 is obtained by Michael addition of nitromethane to a diester of Formula 3.2, followed by reduction of the nitro group to an amine, and subsequent spontaneous cyclization. The lactam is then activitated with Et3O+BF4 " or P2Ss and treated with guanidine to yield the compound of Formula 1.1. Protection of the NH2 group with Ac2O, deoxychlorination with POCI3 and Bn(Et3)2NCI, and deprotection with, for example, HCI yield the 4-chloro-6,7-dihydro- pyrrolo[2,3-d]pyrimidine scaffold. Finally, alkylation with 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine hydrochloride in dimethysulfoxide with sodium hydride provides the compound of Formula I.
Scheme 5
Figure imgf000067_0002
[00143] Compounds of Formula I can also be synthesized from compounds of Formula 4.1 according to Scheme 5. The compound of Formula 4.1 is, for example, alkylated using electrophiles such as LG-R7-R8, where LG is a leaving group, in a solvent such as dimethylsulfoxide or dimethylformamide. Leaving groups include, but are not limited to, e.g., mesylate, halogen, triflate, tosylate, etc. LG-R7-R8 can be, for example, a halide substituted arylalkyl.
Scheme 6
Figure imgf000068_0001
[00144] The compounds of formula 4.1 can be synthesized from amino substituted pyrimidines of
Formula 5.1 according to Scheme 6. Accordingly, the amino group of the pyrimidine of Formula 5.1 is optionally protected with, for example tert-butoxycarbonyl, by reacting the compound with di-tert-butyl dicarbonate under basic conditions. The protected amino pyrimidine of Formula 5.2 is then alkylated with an allyl halide, for example methyl 4-bromocrotonate. Note that the protected amino is recommended with some allyl halides (e.g., methyl bromocrotonate), but is unnecessary with others (e.g., allyl bromide). Next the 5-position is brominated with, for example, NBS or Br2 to set up the compound for radical cyclization. The radical cyclization can be mediated by hydrogen donors such as Bu3SnH or TMS3SiH inc onjunction with a radical initiator such as AIBN, Vazo 88, or Et3B/O2. At this stage, the resulting enantiomers may optionally be separated by chiral HPLC or enzymatic resolution. The amine protecting group is then removed with for example, trifluoroacetic acid in dichloromethane, to yield the compound of Formula 4.1. Scheme 7
Figure imgf000069_0001
[00145] An example of producing a compound of Formula I by proceeding through the synthesis outlined in Scheme 6 is shown in Scheme 7. The synthesis begins with a 6-chloropyrimidine-2,4-diamine of Formula 5.1a. The amino group of the 6-chloropyrimidine-2,4-diamine is protected with tert- butoxycarbonyl under basic conditions using potassium tert-butoxide. The Boc-protected NH is then alkylated with methyl 4-bromocrotonate. The 5-position is brominated with NBS followed by a radical cyclization mediated by TMS3SiH and Vazo88. The two enantiomers of the resulting racemic bicyclic scaffold are separated by chiral HPLC followed by cleavage of the Boc group with trifluoroacetic acid in dichloromethane. The 7-position of the pyrrolopyrimidine is then alkylated in dimethylsulfoxide with a chloro substituted arylalkyl and potassium tert-butoxide to yield an ester of Formula I.
Scheme 8
Figure imgf000069_0002
[00146] An alternate method of synthesizing the compounds of formula 4.1 from amino substituted pyrimidines is shown in Scheme 8. Accordingly, the amino group of the pyrimidine of Formula 6.1 is reacted with an α-chloro or α-bromoaldehyde to give, regioselectively, a pyrrolo[2,3-d]pyrimidine of Formula 6.2. After protection of the NH group with, for example, an electron-withdrawing group such tosyl, the double bond in the 5-membered ring is hydrogenated using, for example, H2 and a Pd-based catalyst to give a 6,7-dihydro-pyrrolo[2,3-d]pyrimidine of Formula 6.4. The protecting group is then cleaved with, for example, an acid such as H2SO4, to yield a compound of Formula 4.1.
Scheme 9
Figure imgf000070_0001
[00147] An example of producing a compound of Formula I by proceeding through the synthesis outlined in Scheme 8 is shown in Scheme 9. Accordingly, 2,6-diaminopyrimidin-4-ol is reacted with an α- bromoaldehyde to give, regioselectively, a pyrrolo[2,3-d]pyrimidine of Formula 6.2a. After protection of the NH2 and NH with electron-withdrawing groups (e.g. pivaloyl and tosyl, respectively), the double bond in the 5-membered ring is hydrogenated, using H2 and a Pd-based catalyst to give a 6,7-dihydro- pyrrolo[2,3-d]pyrimidine of Formula 6.4a. The protecting groups are then cleaved with H2SO4, and the hydroxy group is converted to a chlorine atom with, for example, POCI3. Alkylation with a chloride substituted arylalkyl yields a compound of Formula I.
Scheme 10
Figure imgf000071_0001
[00148] Compounds of Formula I can also be synthesized from iodo-pyrrolo[2,3-c/)pyrimidines of
Formula 7.1 according to Scheme 10. One route begins with the formation of an α,β-unsaturated ester of Formula 8.1 from an iodo-pyrrolo[2,3-c(|pyrimidine of Formula 7.1 by, for example, carbonylation with carbon monoxide in the presence of an alcohol and a catalyst. The α,β-unsaturated ester of Formula 8.1 is then reduced with, for example, a hindered borohydride such as lithium tri-sec-butylborohydride, to yield a compound of Formula I. [00149] A second route begins by Sonagashira coupling of an iodo-pyrrolo[2,3-cdpyrimidine of
Formula 7.1 with an alkyne, followed by hydrolysis of the triple bond, to yield an α,β-unsaturated ketone of Formula 9.2. The α,β-unsaturated ketone of Formula 9.2 is then reduced with, for example, a hindered borohydride such as lithium tri-sec-butylborohydride, to yield a compound of Formula I.
Scheme 11
Figure imgf000072_0001
[00150] An example of the routes outlined in Scheme 10, beginning with a 2-amino, 4-chloro pyrrolopyrimidine, is shown in Scheme 11. In one method, an α,β-unsaturated ester is obtained from the 2-amino, 4-chloro pyrrolopyrimidine by carbonylation with carbon monoxide in the presence of methanol and a palladium catalyst. The α,β-unsatu rated ester is then reduced with lithium tri-sec-butylborohydride (LiBH(sec-Bu)3) to yield a compound of Formula I. In the second method, an α,β-unsaturated ketone of Formula 9.2a is obtained from the 2-amino, 4-chloro pyrrolopyrimidine by Sonagashira coupling with an alkyne, followed by hydrolysis of the triple bond. The α,β-unsaturated ketone of Formula 9.2a is then reduced with lithium tri-sec-butylborohydride (LiBH(sec-Bu)3) to yield a compound of Formula I.
Scheme 12
Figure imgf000073_0001
Formula I
[00151] A variation of the method shown in Scheme 11 that proceeds through an α,β-unsaturated ester is shown in Scheme 12. The synthesis begins with protecting the NH moiety of the 2-amino, 4- chloro pyrrolopyrimidine with, for example, para-methoxy benzyl chloride. After carbon monoxide insertion and 1 ,4-reduction of the α,β-unsaturated ester, the para-methoxybenzyl group is cleaved under acidic conditions (e.g. TFA/H2SO4, 80 0C), and the OH group is protected with a silyl group, for example tert- butyldimethylsilyl. This affords building block 10.3, which is ideally suited for alkylation of the NH with a group such as Ar-CH2-X. Desilyation can be achieved with a fluoride source such as tetra-n- butylammonium fluoride (TBAF), or with an acid such as trifluoroacetic acid (TFA).
Scheme 13
Figure imgf000073_0002
[00152] In addition to the methods outlined in Scheme 1 , the compounds of Formula I can be synthesized by other methods as well. One example of another method is shown in Scheme 13. In Scheme 13, the five membered ring of the compound of Formula I is constructed by starting with a 4- aminopyrimidine of Formula 11.1 and forcing the nucelophilic 4-amino group to displace an appropriately positioned leaving group (LG), such as mesylate or an epoxide. If the leaving group is an epoxide then, as shown in Scheme 14, the ring formation results in a primary alcohol.
Scheme 15
Figure imgf000074_0001
[00153] Scheme 15 shows the further elaboration of the R3 substituent of compounds of Formula
I by substitution reactions wherein LG is a leaving group and Nuc is a nucleophile. Leaving groups include, but are not limited to, e.g., mesylate, halogen, hydroxyl substituents derivatized by the Mitsunobu reaction, triflate, tosylate, etc. Nucleophiles include, but are not limited to, e.g., halides, cyanide, azide, heterocycles, hydrides, etc. The substitution reactions can be carried out in solvents such as acetone, tetrahydrofuran, dimethylsulfoxide, dimethylacetamide, etc. and in the presence of bases such as potassium carbonate. Following substitution, further reactions such as oxidation and coupling reactions can then be performed. In addition, the R3 substituent can be elaborated by a variety of other reactions known to those skilled in the art such as cyclizations, reductions, acyl transfer reactions, sulfonation, etc. Examples of reactions that may be used to elaborate the R3 substitutent are shown and described in Schemes 16-24 below.
Scheme 16
Figure imgf000074_0002
[00154] As shown in Scheme 16, an OH substituted R3 moiety can displace a group on the 4- position of the pyrimidine ring to form an additional ring. The displacement of the group on the 4-position can be accomplished by heating a compound of Formual 12.1 in the presence of sodium hydroxide as shown in Scheme 17.
Figure imgf000075_0001
[00155] Amides can be generated from an ester substituted R i3 substituent by conversion of the ester group into an amide group as shown in Scheme 18. A wide variety of conditions can be employed for this conversion. For example, to prepare a primary amide, the ester can be hydrolyzed, activated (e.g., with pentafluorophenyl ester), and treated with ammonia as shown in Scheme 19. Alternatively, the ester group can be directly converted to the primary amide by heating with ammonia or by heating with an ammonia equivalent such as HCONH2/tBuOK. To prepare secondary or tertiary amides, a synthesis analogous to that shown in Scheme 20 can be used. Specifically, the ester can be hydrolyzed, activated (e.g., with pentafluorophenyl ester), and then treated with an amine other than NH3. Scheme 21
Figure imgf000076_0001
[00156] Primary amides can be also generated from a cyano substituted R3 substitent. As shown in Scheme 21 , the cyano substituted R3 substitent can be generated from a compound of Formula 14.1 by nucleophilic substitution with a CN reagent, such as potassium cyanide. The cyano compound of Formula 14.2 can then be treated with, for example, H2CVNaOH, to yield a primary amide of Formula I. An example of the route outlined in Scheme 19 starting with a 2-amino-4-chloro-6,7- dihydropyrrolopyrimidine is shown in Scheme 22. The OH group of the R3 substituent is activated by reaction with methanesulfonyl chloride which is then displaced by potassium cyanide. The cyano compound is treated with H2O2/NaOH to yield a primary amide of Formula I.
Scheme 23
Figure imgf000077_0001
[00157] Additional methods that may be used to elaborate the R3 substituent are shown in
Scheme 23. For example, the R3 substituent can be activated and then substituted. Further, the R3 substituent can by cyclized or reduced and then acylated. As a representative example, a hydroxy group can be converted into a leaving group such as a mesylate, then displaced with a nucleophile such as azide as Shown in Scheme 24. The azide can then be reacted with a terminal alkyne to give a [2+3] dipolar cycloaddition or reduced to an amine that can be further acylated. [00158] The compounds and pharmaceutically acceptable salts described herein exhibit acceptable stability towards acids, bases, and amino acids. For example, the compounds and pharmaceutically acceptable salts described herein exhibited stability towards acids. The stability of the compounds and pharmaceutically acceptable salts described herein in acidic medium can be tested by incubating the compound or pharmaceutically acceptable salt in a 5M solution of methanesulfonic acid in DMSO/Water 1 :1 at about 40 0C. Certain of the compounds described herein displayed a half-life greater than 100 h when tested under such conditions. [00159] Furthermore, compounds and pharmaceutically acceptable salts described herein exhibit stability with regard to reactions with bases such as lysine, proteins, and the like. The stability of the compounds and pharmaceutically acceptable salts described herein with bases such as lysine can be tested by incubating the compound or pharmaceutically acceptable salt in a 10 mM solution of lysine in DMSO/Water 1 :1 at about 40 0C. Certain of the compounds described herein displayed a half-life greater than 10O h when tested under such conditions.
[00160] Also provided are methods for the use of the compounds and pharmaceutically acceptable salts described herein, in treatment of disorders that are mediated by Hsp90. [Examples of such disorders include, but are not limited to, inflammatory diseases, infections, autoimmune disorders, stroke, ischemia, cardiac disorder, neurological disorders, fibrogenic disorders, proliferative disorders, and metabolic diseases.
[00161] Fibrogenic disorders include but are not limited to connective tissue diseases, such as scleroderma (or systemic sclerosis), polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders, including liver cirrhosis, keloid formation, interstitial nephritis, and pulmonary fibrosis.
[00162] Proliferative disorders include cancer, psoriasis and benign prostatic hyperplasia (BPH).
Cancer includes, for example, malignant tumors, e.g., non-solid tumors such as leukemia, multiple myeloma or lymphoma, and also solid tumours, for example, oesophageal cancer, myeloma, hepatocellular, pancreatic, cervical cancer, ewings tumour, neuroblastoma, kaposis sarcoma, ovarian cancer, endometrial cancer, colorectal cancer, prostate cancer, bladder cancer, melanoma, head and neck cancer, renal cancer, bile duct cancer, bone cancer, neuronal cancer, testicular cancer, ovarian cancer. Additional forms of cancer include, for example, breast, ovary, lung (including small cell lung cancer, non-small cell lung cancer and bronchioalveolar cancer), colon, rectum, prostate, bile duct, bone, bladder, head and neck, kidney, liver, gastrointestinal tissue, oesophagus, pancreas, skin, testes, thyroid, uterus, cervix, vulva or other tissues, as well as leukemias and lymphomas including CLL and CML, tumors of the central and peripheral nervous system, and other tumor types such as melanoma, multiple myeloma, fibrosarcoma and osteosarcoma, and malignant brain tumors.
[00163] In some embodiments the cancer to be treated is identified by the expression or lack of expression of at least one molecular marker known to correlate with either or both of 1) dependency on Hsp90 for cancer cell growth and/or survival, and 2) susceptibility to treatment by administration of an Hsp90 inhibitor disclosed herein. For example, a cancer biopsy may be screened and identified as HER- 2 positive and that patent may then be selected for treatment by administration of a chemical entity disclosed herein. In some embodiments the molecular marker is chosen from HER-2, estrogen receptor, progesterone receptor, androgen receptor, EGF receptor, B Raf, AKT, and NF-kB. [00164] Other Hsp90-mediated disorders include multiple sclerosis, lupus, rheumatoid arthritis and irritable bowel syndrome CJD, Huntington's disease and Alzheimer's disease. [00165] Also provided are pharmaceutical compositions comprising one or more pharmaceutically acceptable carriers and at least one compound or pharmaceutically acceptable salt thereof, described herein.
[00166] Those of ordinary skill in the art are familiar with formulation and administration techniques that can be employed with the compounds and pharmaceutically acceptable salts described herein, e.g., as discussed in Goodman and Gilman, The Pharmacological Basis of Therapeutics, current ed.; Pergamon; and Remington's, Pharmaceutical Sciences (current edition), Mack Publishing Co., Easton, PA.
[00167] The compounds or salts thereof may be administered either alone or in combination with pharmaceutically acceptable carriers in a pharmaceutical composition, according to standard pharmaceutical practice. The compounds and pharmaceutically acceptable salts can be administered orally or parenterally, including the intravenous, intramuscular, intraperitoneal, subcutaneous, rectal and topical routes of administration.
[00168] For example, the pharmaceutical compositions described herein can be administered locally to the area in need of treatment. This may be achieved by, for example, but not limited to, local infusion during surgery, topical application, e.g., cream, ointment, injection, catheter, or implant, said implant made, e.g., out of a porous, non-porous, or gelatinous material, including membranes, such as sialastic membranes, or fibers. The administration can also be by direct injection at the site (or former site) of a tumor or neoplastic or pre-neoplastic tissue.
[00169] Still further, the compounds or pharmaceutically acceptable salts described herein can be delivered in a vesicle, e.g., a liposome (see, for example, Langer, Science 1990, 249,1527 1533; Treat et al., Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Bernstein and Fidler, Ed., Liss, N.Y., pp. 353 365, 1989).
[00170] The pharmaceutical compositions described herein can also be delivered in a controlled release system. In some embodiments, a pump may be used (see, Sefton, 1987, CRC Crit. Ref. Biomed. Eng. 14:201 ; Buchwald et al. Surgery, 1980 88, 507; Saudek et al. N. Engl. J. Med. 1989, 321 , (574). Additionally, a controlled release system can be placed in proximity of the therapeutic target. (See, Goodson, Medical Applications of Controlled Release, 1984, Vol. 2, pp. 115 138). [00171] The pharmaceutical compositions described herein can also contain the active ingredient in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. Compositions intended for oral use may be prepared according to any method known in the art for the manufacture of pharmaceutical compositions, and such compositions may contain one or more agents selected from sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations. Tablets contain the active ingredient in admixture with one or more pharmaceutically acceptable carriers which are suitable for the manufacture of tablets. These pharmaceutically acceptable carriers may be, for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as microcrystalline cellulose, sodium crosscarmellose, corn starch, or alginic acid; binding agents, for example starch, gelatin, polyvinyl-pyrrolidone or acacia, and lubricating agents, for example, magnesium stearate, stearic acid or talc. The tablets may be un-coated or coated by known techniques to mask the taste of the drug or delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a water soluble taste masking material such as hydroxypropylmethyl-cellulose or hydroxypropylcellulose, or a time delay material such as ethyl cellulose, or cellulose acetate butyrate may be employed as appropriate. [00172] Formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water soluble carrier such as polyethyleneglycol or an oil medium, for example peanut oil, liquid paraffin, or olive oil. [00173] Aqueous suspensions contain the active material in admixture with one or more pharmaceutically acceptable carriers suitable for the manufacture of aqueous suspensions. Such carriers are suspending agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethyl-cellulose, sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be a naturally-occurring phosphatide, for example lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example heptadecaethylene-oxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and a hexitol such as polyoxyethylene sorbitol monooleate, or condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example polyethylene sorbitan monooleate. The aqueous suspensions may also contain one or more preservatives, for example ethyl, or n-propyl p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents, such as sucrose, saccharin or aspartame. [00174] Oily suspensions may be formulated by suspending the active ingredient in a vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in mineral oil such as liquid paraffin. The oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an anti-oxidant such as butylated hydroxyanisol or alpha-tocopherol.
[00175] Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already mentioned above. Additional substances, for example sweetening, flavoring and coloring agents, may also be present. These compositions may be preserved by the addition of an anti-oxidant such as ascorbic acid. [00176] The pharmaceutical compositions described herein may also be in the form of an oil-in- water emulsion. The oily phase may be a vegetable oil, for example olive oil or arachis oil, or a mineral oil, for example liquid paraffin, or mixtures of these. Suitable emulsifying agents may be naturally- occurring phosphatides, for example soybean lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monooleate. The emulsions may also contain sweetening agents, flavoring agents, preservatives and antioxidants. [00177] Syrups and elixirs may be formulated with sweetening agents, for example glycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative, flavoring and coloring agents and antioxidant.
[00178] The pharmaceutical compositions may be in the form of a sterile injectable aqueous solution. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution.
[00179] The sterile injectable preparation may also be a sterile injectable oil-in-water microemulsion where the active ingredient is dissolved in the oily phase. For example, the active ingredient may be first dissolved in a mixture of soybean oil and lecithin. The oil solution may then be introduced into a water and glycerol mixture and processed to form a microemulsion. [00180] The injectable solutions or microemulsions may be introduced into a patient's bloodstream by local bolus injection. Alternatively, it may be advantageous to administer the solution or microemulsion in such a way as to maintain a constant circulating concentration of the instant compound. In order to maintain such a constant concentration, a continuous intravenous delivery device may be utilized. An example of such a device is the Deltec CADD-PLUS™ model 5400 intravenous pump. [00181] The pharmaceutical compositions may be in the form of a sterile injectable aqueous or oleagenous suspension for intramuscular and subcutaneous administration. This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in 1 ,3-butanediol. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid may find use in the preparation of injectables. [00182] The pharmaceutical compositions described herein may also be administered in the form of suppositories for rectal administration of the drug. These compositions can be prepared by mixing the inhibitors with a suitable nonirritating pharmaceutically acceptable carrier which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug. Such materials include cocoa butter, glycerinated gelatin, hydrogenated vegetable oils, mixtures of polyethylene glycols of various molecular weights and fatty acid esters of polyethylene glycol. [00183] For topical use, creams, ointments, jellies, solutions or suspensions, etc., containing a compound pharmaceutically acceptable salts described herein can be used. As used herein, topical application can include mouth washes and gargles.
[00184] The compounds and pharmaceutically acceptable salts described herein can be administered in intranasal form via topical use of suitable intranasal vehicles and delivery devices, or via transdermal routes, using those forms of transdermal skin patches well known to those of ordinary skill in the art. To be administered in the form of a transdermal delivery system, the dosage administration will, of course, be continuous rather than intermittent throughout the dosage regimen. [00185] The compounds and pharmaceutically acceptable salts described herein may also be used in conjunction with other well known therapeutic agents that are selected for their particular usefulness against the condition that is being treated. For example, the compounds and pharmaceutically acceptable salts described herein may be useful in combination with at least one additional anti-cancer and/or cytotoxic agents. Further, the compounds and pharmaceutically acceptable salts described herein may also be useful in combination with other inhibitors of parts of the signaling pathway that links cell surface growth factor receptors to nuclear signals initiating cellular proliferation. [00186] Such known anti-cancer and/or cytotoxic agents that may be used in combination with the compounds and pharmaceutically acceptable salts described herein include:
(i) other antiproliferative/antineoplastic drugs and combinations thereof, as used in medical oncology, such as alkylating agents (for example cis-platin, oxaliplatin, carboplatin, cyclophosphamide, nitrogen mustard, melphalan, chlorambucil, busulphan, temozolamide and nitrosoureas); antimetabolites (for example gemcitabine and antifolates such as fluoropyrimidines like 5-fluorouracil and tegafur, raltitrexed, methotrexate, cytosine arabinoside, and hydroxyurea); antitumour antibiotics (for example anthracyclines like adriamycin, bleomycin, doxorubicin, daunomycin, epirubicin, idarubicin, mitomycinC, dactinomycin and mithramycin); antimitotic agents (for example vinca alkaloids like vincristine, vinblastine, vindesine and vinorelbine and taxoids like taxol and taxotere and polokinase inhibitors); and topoisomerase inhibitors (for example epipodophyllotoxins like etoposide and teniposide, amsacrine, topotecan and camptothecin);
(ii) cytostatic agents such as antioestrogens (for example tamoxifen, fulvestrant, toremifene, raloxifene, droloxifene and iodoxyfene), antiandrogens (for example bicalutamide, flutamide, nilutamide and cyproterone acetate), LHRH antagonists or LHRH agonists (for example goserelin, leuprorelin and buserelin), progestogens (for example megestrol acetate), aromatase inhibitors (for example as anastrozole, letrozole, vorazole and exemestane) and inhibitors of 5a-reductase such as finasteride;
(iii) anti-invasion agents [for example c-Src kinase family inhibitors like 4-(6-chloro- 2,3methylenedioxyanilino)-7-[2-(4-methylpiperazin-l-yl)ethoxy]-5-tetrahydropyran-4yloxyquinazoline (AZD0530; International Patent Application WO 01/94341), N-(2- chloro-6-methylphenyl)-2- {6-[4-(2- hydroxyethyl)piperazin-l-yl]-2-methylpyrimidin-4ylamino}thiazole-5-carboxamide (dasatinib, BMS-354825; J. Med. Chern., 2004, 47, 66586661 )and bosutinib (SKI-606), and metalloproteinase inhibitors like marimastat, inhibitors of urokinase plasminogen activator receptor function or antibodies to Heparanase];
(iv) inhibitors of growth factor function: for example such inhibitors include growth factor antibodies and growth factor receptor antibodies (for example the anti-erbB2 antibody trastuzumab [Herceptin™], the anti-EGFR antibody panitumumab, the anti-erbB 1 antibody cetuximab [Erbitux, C225] and any growth factor or growth factor receptor antibodies disclosed by Stem et al. Critical reviews in oncology/haematology, 2005, Vol. 54, pp 11-29); such inhibitors also include tyrosine kinase inhibitors, for example inhibitors of the epidermal growth factor family (for example EGFR family tyrosine kinase inhibitors such as W-fS-cft/oro^-fluorophenyO^-methoxy-e-fS-morpholinopropoxyJquinazolin^-amine (gefitinib, ZD1839), N-(3-ethynylphenyl)-6,7-bis(2-methoxyethoxy)quinazolin-4-amine (erlotinib, OSI-774) and 6-acrylamido-N-(3-chloro-4-fluorophenyl)-7-(3- morpholinopropoxy)-quinazolin-4-amine (Cl 1033), erbB2 tyrosine kinase inhibitors such as lapatinib); inhibitors of the hepatocyte growth factor family; inhibitors of the insulin growth factor family; inhibitors of the platelet-derived growth factor family such as imatinib and/or nilotinib (AMN107); inhibitors of serine/threonine kinases (for example Ras/Raf signalling inhibitors such as farnesyl transferase inhibitors, for example sorafenib (BAY 43-9006), tipifarnib (Rl 15777) and lonafamib (SCH66336)), inhibitors of cell signalling through MEK and/or AKT kinases, c-kit inhibitors, abl kinase inhibitors, P13 kinase inhibitors, Plt3 kinase inhibitors, CSF-IR kinase inhibitors, IGF receptor (insulin like growth factor) kinase inhibitors; aurora kinase inhibitors (for example AZD1152, PH739358, VX-680, MLN8054, R763, MP235, MP529, VX-528 and AX39459) and cyclin dependent kinase inhibitors such as CDK2 and/or CDK4 inhibitors;
(v) antiangiogenic agents such as those which inhibit the effects of vascular endothelial growth factor, [for example the anti-vascular endothelial cell growth factor antibody bevacizumab (Avastin™) and for example, a VEGF receptor tyrosine kinase inhibitor such as vandetanib(ZD6474), vatalanib (PTK787), sunitinib (SU1 I248), axitinib (AG-013736), pazopanib (GW 786034) and 4 {4-fluoro-2- methylindol-5-yloxy)-6-methoxy-7-(3pyrrolidin-l-ylpropoxy)quinazoline (AZD2I7I; Example 240 within WO 00/47212), compounds such as those disclosed in International Patent Applications W097/22596, WO 97/30035, WO 97/32856 and WO 98/13354 and compounds that work by other mechanisms (for example linomide, inhibitors of integrin av~3 function and angiostatin));
(vi) vascular damaging agents such as Combretastatin A4 and compounds disclosed in International Patent Applications WO 99/02166, WO 00/40529, WO 00/41669, WO 01/92224, WO 02/04434 and WO 02/08213;
(vii) an endothelin receptor antagonist, for example zibotentan (ZD4054) or atrasentan;
(viii) antisense therapies, for example those which are directed to the targets listed above, such as ISIS 2503, an anti-ras antisense;
(ix) gene therapy approaches, including for example approaches to replace aberrant genes such as aberrant p53 or aberrant BRCAI or BRCA2, GDEPT (gene-directed enzyme pro-drug therapy) approaches such as those using cytosine deaminase, thymidine kinase or a bacterial nitroreductase enzyme and approaches to increase patient tolerance to chemotherapy or radiotherapy such as multidrug resistance gene therapy; and
(x) immunotherapy approaches, including for example ex-vivo and in-vivo approaches to increase the immunogenicity of patient tumour cells, such as transfection with cytokines such as interleukin 2, interleukin 4 or granulocyte-macrophage colony stimulating factor, approaches to decrease T-cell anergy, approaches using transfected immune cells such as cytokine-transfected dendritic cells, approaches using cytokine-transfected tumour cell lines and approaches using anti-idiotypic antibodies. [00187] In certain embodiments, the compound or pharmaceutically acceptable salt is administered in combination with one or more agents chosen from pacliataxel, bortezomib, dacarbazine, gemcitabine, trastuzumab, bevacizumab, capecitabine, docetaxel, erlotinib, aromatase inhibitors, such as AROMASIN™ (exemestane), and estrogen receptor inhibitors, such as FASLODEX™ (fulvestrant). [00188] In addition, the compounds and pharmaceutically acceptable salts described herein may be useful in combination with at least one additional anti-inflammatory and/or immuno-modulatory agent. [00189] Such known additional anti-inflammatory and/or immuno-modulatory agents that may be used in combination with the compounds and pharmaceutically acceptable salts described herein include:
(i) TNF-blocking agents such as etanercept, infliximab, and adalimumab;
(ii) anti-B cell therapeutics such as rituximab;
(iii) nonsteroidal anti-inflammatory drugs such as diclofenac, etodolac, fenoprofen, flurbiprofen, ibuprofen, indomethacin, ketoprofen, meclofenamate, nabumetone, naproxen, oxaprozin, piroxicam, sulindac and tolmetin;
(iv) COX-2 inhibitors such as celecoxib and meloxicam;
(v) disease modifying anti-rheumatic drugs (DMARDs) such as sulfasalazine, hydroxychloroquine, chloroquine, gold salts, D-penicillamine, and methotrexate;
(vi) anti-malarials such as hydroxychloroquine, chloroquine, and quinacrine; and
(vii) corticosteroids such as prednisone, betamethasone, dexamethasone, hydrocortisone, methylprednisolone, and triamcinolone;
[00190] In certain embodiments, the compound or pharmaceutically acceptable salt is administered in combination with one or more agents chosen from hydroxychloroquine and methotrexate. [00191] When a compound or pharmaceutically acceptable salt described herein is administered into a human subject, the daily dosage will normally be determined by the prescribing physician with the dosage generally varying according to the age, weight, and response of the individual patient, as well as the severity of the patient's symptoms.
[00192] In one exemplary application, a suitable amount of compound is administered to a mammal undergoing treatment for cancer, for example, breast cancer. Administration typically occurs in an amount of between about 0.01 mg/kg of body weight to about 100 mg/kg of body weight per day (administered in single or divided doses), such as at least about 0.1 mg/kg of body weight per day. A particular therapeutic dosage can include, e.g., from about 0.01 mg to about 1000 mg of compound, such as including, e g , from about 1 mg to about 1000 mg The quantity of active compound in a unit dose of preparation may be varied or adjusted from about 0 1 mg to 1000 mg, such as from about 1 mg to 300 mg, for example 10 mg to 200 mg, according to the particular application The amount administered will vary depending on the particular IC50 value of the compound used and the judgment of the attending clinician taking into consideration factors such as health, weight, and age In combinational applications in which the compound is not the sole active ingredient, it may be possible to administer lesser amounts of compound and still have therapeutic or prophylactic effect
[00193] In some embodiments, the pharmaceutical preparation is in unit dosage form In such form, the preparation is subdivided into unit doses containing appropriate quantities of the active component, e g , an effective amount to achieve the desired purpose
[00194] The actual dosage employed may be varied depending upon the requirements of the patient and the severity of the condition being treated Determination of the proper dosage for a particular situation is within the skill of the art Generally, treatment is initiated with smaller dosages which are less than the optimum dose of the compound Thereafter, the dosage is increased by small amounts until the optimum effect under the circumstances is reached For convenience, the total daily dosage may be divided and administered in portions during the day if desired
[00195] The amount and frequency of administration of the compounds and pharmaceutically acceptable salts described herein, and if applicable other chemotherapeutic agents and/or radiation therapy, will be regulated according to the judgment of the attending clinician (physician) considering such factors as age, condition and size of the patient as well as severity of the disease being treated [00196] The chemotherapeutic agent and/or radiation therapy can be administered according to therapeutic protocols well known in the art It will be apparent to those skilled in the art that the administration of the chemotherapeutic agent and/or radiation therapy can be varied depending on the disease being treated and the known effects of the chemotherapeutic agent and/or radiation therapy on that disease Also, in accordance with the knowledge of the skilled clinician, the therapeutic protocols (e g , dosage amounts and times of administration) can be varied in view of the observed effects of the administered therapeutic agents (ι e , antineoplastic agent or radiation) on the patient, and in view of the observed responses of the disease to the administered therapeutic agents
[00197] Also, in general, the compounds and pharmaceutically acceptable salts described herein need not be administered in the same pharmaceutical composition as a chemotherapeutic agent, and may, because of different physical and chemical characteristics, be administered by a different route For example, the compounds/compositions may be administered orally to generate and maintain good blood levels thereof, while the chemotherapeutic agent may be administered intravenously The determination of the mode of administration and the advisability of administration, where possible, in the same pharmaceutical composition, is well within the knowledge of the skilled clinician The initial administration can be made according to established protocols known in the art, and then, based upon the observed effects, the dosage, modes of administration and times of administration can be modified by the skilled clinician.
[00198] The particular choice of compound (and where appropriate, chemotherapeutic agent and/or radiation) will depend upon the diagnosis of the attending physicians and their judgment of the condition of the patient and the appropriate treatment protocol.
[00199] The compounds and pharmaceutically acceptable salts described herein (and where appropriate chemotherapeutic agent and/or radiation) may be administered concurrently (e.g., simultaneously, essentially simultaneously or within the same treatment protocol) or sequentially, depending upon the nature of the proliferative disease, the condition of the patient, and the actual choice of chemotherapeutic agent and/or radiation to be administered in conjunction (i.e., within a single treatment protocol) with the compound/composition.
[00200] In combinational applications and uses, the compound/composition and the chemotherapeutic agent and/or radiation need not be administered simultaneously or essentially simultaneously, and the initial order of administration of the compound/composition, and the chemotherapeutic agent and/or radiation, may not be important. Thus, the compounds and pharmaceutically acceptable salts described herein may be administered first followed by the administration of the chemotherapeutic agent and/or radiation; or the chemotherapeutic agent and/or radiation may be administered first followed by the administration of the compounds and pharmaceutically acceptable salts described herein. This alternate administration may be repeated during a single treatment protocol. The determination of the order of administration, and the number of repetitions of administration of each therapeutic agent during a treatment protocol, is well within the knowledge of the skilled physician after evaluation of the disease being treated and the condition of the patient. For example, the chemotherapeutic agent and/or radiation may be administered first, and then the treatment continued with the administration of the compounds and pharmaceutically acceptable salts described herein followed, where determined advantageous, by the administration of the chemotherapeutic agent and/or radiation, and so on until the treatment protocol is complete.
[00201] Thus, in accordance with experience and knowledge, the practicing physician can modify each protocol for the administration of a compound/composition for treatment according to the individual patient's needs, as the treatment proceeds.
[00202] The attending clinician, in judging whether treatment is effective at the dosage administered, will consider the general well-being of the patient as well as more definite signs such as relief of disease-related symptoms, inhibition of tumor growth, actual shrinkage of the tumor, or inhibition of metastasis. Size of the tumor can be measured by standard methods such as radiological studies, e.g., CAT or MRI scan, and successive measurements can be used to judge whether or not growth of the tumor has been retarded or even reversed. Relief of disease-related symptoms such as pain, and improvement in overall condition can also be used to help judge effectiveness of treatment. [00203] A variety of in vitro and in vivo assays are available to test the effect of the compounds and pharmaceutically acceptable salts described herein on Hsp90. Hsp90 competitive binding assays and functional assays can be performed as known in the art by substituting in the compounds and pharmaceutically acceptable salts described herein. Chiosis et al. Chemistry & Biology 2001 , 8, 289 299, describe some of the known ways in which this can be done. For example, competition binding assays using, e.g., geldanamycin or 17-AAG as a competitive binding inhibitor of Hsp90 can be used to determine relative Hsp90 affinity of the compounds and pharmaceutically acceptable salts described herein by immobilizing the compound or salt of interest or other competitive inhibitor on a gel or solid matrix, preincubating Hsp90 with the other inhibitor, passing the preincubated mix over the gel or matrix, and then measuring the amount of Hsp90 that retains or does not retain on the gel or matrix. Alternatively, competition binding assays using a geldanamycin fluorescent probe, e.g., reduced FITC- geldanamycin, as a competitive binding inhibitor of Hsp90 or an Hsp90 paralog can be be performed by pre-incubating Hsp90 or the Hsp90 paralog with the geldanamycin fluorescent probe, adding the compound or salt of interest, incubating the reaction with gentle shaking, and then measuring the fluorescence. Maximum signal and background controls contain no compound or no Hsp90 (or Hsp90 paralog), respectively.
[00204] Downstream effects can also be evaluated based on the known effect of Hsp90 inhibition on function and stability of various steroid receptors and signaling proteins including, e.g., Raf1 and HER2. Compounds and pharmaceutically acceptable salts described herein induce dose-dependent degradation of these molecules, which can be measured using standard techniques. For example, Hsp90 inhibition can be determined with a cell based assay that quantifies the expression level of HER2 in tumor cells treated with the compound or salt of interest. Inhibition of Hsp90 also results in up- regulation of Hsp90 and related chaperone proteins that can similarly be measured. Antiproliferative activity on various cancer cell lines can also be measured, as can morphological and functional differentiation related to Hsp90 inhibition. For example, the tumor cell growth inhibitory activity of the compound or salt of interest may be determined by treating tumor cells with various concentrations of the compound or salt of interest followed by measurement of cell viability. [00205] Many different types of methods are known in the art for determining protein concentrations and measuring or predicting the level of proteins within cells and in fluid samples. Indirect techniques include nucleic acid hybridization and amplification using, e.g., polymerase chain reaction (PCR). These techniques are known to the person of skill and are discussed, e.g., in Sambrook, Fritsch & Maniatis Molecular Cloning: A Laboratory Manual, 2nd ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y., 1989; Ausubel, et al. Current Protocols in Molecular Biology, John Wiley & Sons, NY, 1994, and, as specifically applied to the quantification, detection, and relative activity of HER2/Neu in patient samples, e.g., in U.S. Pat. Nos. 4,699,877, 4,918,162, 4,968,603, and 5,846,749. A brief discussion of two generic techniques that can be used follows. [00206] The determination of whether cells overexpress or contain elevated levels of HER2 can be determined using well known antibody techniques such as immunoblotting, radioimmunoassays, western blotting, immunoprecipitation, enzyme-linked immunosorbant assays (ELISA), and derivative techniques that make use of antibodies directed against HER2. As an example, HER2 expression in breast cancer cells can be determined with the use of an immunohistochemical assay, such as the Dako Hercep™ test (Dako Corp., Carpinteria, Calif.)- The Hercep™ test is an antibody staining assay designed to detect HER2 overexpression in tumor tissue specimens. This particular assay grades HER2 expression into four levels: 0, 1 , 2, and 3, with level 3 representing the highest level of HER2 expression. Accurate quantitation can be enhanced by employing an Automated Cellular Imaging System (ACIS) as described, e.g., by Press, M. et al. Modern Pathology 2000, 13, 225A.
[00207] Antibodies, polyclonal or monoclonal, can be purchased from a variety of commercial suppliers, or may be manufactured using well-known methods, e.g., as described in Harlow et al. Antibodies: A Laboratory Manual, 2nd ed; Cold Spring Harbor Laboratory, Cold Spring Harbor, N. Y., 1988.
[00208] HER2 overexpression can also be determined at the nucleic acid level since there is a reported high correlation between overexpression of the HER2 protein and amplification of the gene that codes for it. One way to test this is by using RT-PCR. The genomic and cDNA sequences for HER2 are known. Specific DNA primers can be generated using standard, well-known techniques, and can then be used to amplify template already present in the cell. An example of this is described in Kurokawa, H. et al. Cancer Res. 2000, 60, 5887 5894. PCR can be standardized such that quantitative differences are observed as between normal and abnormal cells, e.g, cancerous and noncancerous cells. Well known methods employing, e.g., densitometry, can be used to quantitate and/or compare nucleic acid levels amplified using PCR.
[00209] Similarly, fluorescent in situ hybridization (FISH) assays and other assays can be used, e.g., Northern and/or Southern blotting. These rely on nucleic acid hybridization between the HER2 gene or mRNA and a corresponding nucleic acid probe that can be designed in the same or a similar way as for PCR primers, above. See, e.g., Mitchell M S, and Press M. F. Oncol., Suppl. 1999, 12, 108 116. For FISH, this nucleic acid probe can be conjugated to a fluorescent molecule, e.g., fluorescein and/or rhodamine, that does not interfere with hybridization, and which fluorescence can later be measured following hybridization. See, e.g., Kurokawa, H et al, Cancer Res. 2000, 60, 5887-5894 (describing a specific nucleic acid probe having sequence δ'-FAM-NucleicAcid-TAMRA-p-S' sequence). ACIS-based approaches as described above can be employed to make the assay more quantitative (de Ia Torre- Bueno, J., et al. Modern Pathology 2000, 13, 221A).
[00210] lmmuno and nucleic acid detection can also be directed against proteins other than
Hsp90 and HER2, which proteins are nevertheless affected in response to Hsp90 inhibition. [00211] In vitro and in vivo assays are also available to determine the anti-inflammatory and immunomodulatory activities of the compounds and pharmaceutically acceptable salts described herein. For example, the stability and function of newly synthesized and activated/phosphorylated forms of a proximal signaling molecule in the T cell receptor activation pathway, the Src family kinase p56fck, is dependent upon Hsp90, with Hsp90 inhibition resulting in p56"* degradation and subsequent internalization of CD4. Thus, the ability of the compound or salt of interest to inhibit human CD4+ T cell proliferation in response to an allogeneic non-T cell stimulus and the ability of the compound or salt of interest to down modulate T cell surface CD4 can be measured. In addition, the ability of the compound or salt of interest to inhibit lipopolysaccharide (LPS)-induced activation of mitogen-activated protein kinase (MAPK) pathways can be determined.
[00212] The following examples are offered by way of illustration only and are not intended to be limiting of the full scope and spirit of the invention.
EXAMPLES
[00213] The chemical reagents used to create the novel products of the invention are all available commercially, e.g., from Aldrich Chemical Co., Milwaukee, Wis., USA. Otherwise their preparation is facile and known to one of ordinary skill in the art, or it is referenced or described herein.
General Procedure for the Suzuki Reaction and TBS/BOC Deprotection
[00214] A mixture of the aryl iodide (typically 100 mg, 0.188 mmol), the boronic acid or ester (1.2 equivalents), 1 ,1 '[bis(diphenylphosphino)ferrocene] dichloropalladium(ll) (25 mg), DMSO (4 ml_), and 2.0
M K2CO3 in water (0.5 mL, 1 mmol) was degassed by bubbling nitrogen through it. The mixture was heated at 90 0C for 1 h. Water was added to precipitate the crude product which was then collected by filtration. The precipitate was redissolved in MeOH (6 mL, 100 mmol) and TFA (2 mL, 20 mmol) for deprotection of the TBS and/or BOC group. The reaction was monitored via LC/MS. Upon completion, the solvent was removed and the product was purified by preparative, reverse phase HPLC.
General Alkylation Procedure
[00215] A mixture of the 6,7-dihydropyrrrolo[2,3-d]pyrimidine (1 equiv.), the alkylating agent of formula Aryl-CH2CI (1-2 equiv.), and base (NaH or tBuOK, 1-2 equiv.) is stirred in DMF or DMSO at 0 0C or room temperature for 10-20 minutes. Work-up (EtOAc/water), drying (brine, Na2SO4) and evaporation gives the crude alkylated material.
General Amide Formation Procedure
[00216] The carboxylic acid (either in its acid form or as its lithium salt, 1 equiv.) is dissolved/suspended in THF. Pentafluorophenyl trifluoroacetate (1-3 equiv.) and the mixture is stirred ar
RT for 5-60 minutes. The desired amine is added (2-10 equivalents) and the mixture is stirred for another
5-60 minutes. Work-up (EtO Ac/water), drying (brine, Na2SO4) and evaporation gives the crude amide.
General Analytical Methods
[00217] The analytical HLPC and LC-MS chromatograms were obtained using a C18 column
(Agilent Zorbax SB-C18, 5 microns; 4.6 mm x 150 mm). A gradient was applied between solvent A (0.1%
TFA in H2O) and solvent B (0.05% TFA in CH3CN) increasing the proportion of B linearly from 5% to 100% over 5 minutes (5-100-5 method) or over 7 minutes (5-100-7 method) with a constant flow rate of 1 mL/min. The column was not heated, and the purity was determined at 254 nM. Example 1
(2-Amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (III)
Step 1. Methyl 2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7H-pyrrolo[2,3- d]pyrimidine-5-carboxylate (H)
[00218] Into an autoclave vessel was added 4-chloro-5-iodo-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine (I1 30.0 g, 67.6 mmol), [(R)-(+)-2,2'-bis(diphenylphosphino)- 1 ,1'-binaphthyl]palladium(ll) chloride (541 mg, 0.676 mmol, Aldrich# 342335), triethylamine (18.8 ml_, 135 mmol) and anhydrous methanol (54.8 ml_, 1350 mmol). The autoclave was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 800C while stirring for 3 hours. The reaction was cooled to room temperature and diluted with water (20OmL), and extracted with a mixture of dichloromethane: isopropanol (95:5, 300mLx3). The organic layer was filtered over a celite pad, concentrated, and the solid was collected, washed with cold methanol (5mLx3), and dried over high vacuum pump to give the title compound as a white solid (16.0 g, purity 85.0%, yield 63.0%). 1HNMR (DMSO): 8.05 (s, 1 H), 7.87 (s, 1 H), 6.79 (s, 2H), 5.32 (s, 2H), 3.75 (s, 3H), 3.73 (s, 3H), 2.28 (s, 3H), 2.16 (s, 3H). LS-MS: 375.6 (M, 100%), 377.6 (M+2, 32.7%), 376.6 (M+1 , 19.2 %), 378.6 (M+3, 5.7%). Step 2. (2-Amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (III)
[00219] To a suspension of methyl 2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate (II, 10.0 g, 26.6 mmol) in anhydrous tetrahydrofuran (150 mL, 1.8 mol) was added 1.0 M of L-selectride solution (160 mL, 160mmol) dropwise via syringe at 0 0C. The reaction mixture was warmed to room temperature, allowed to stir for 1 hr, carefully quenched with ice water, adjusted to pH 7.0 with 2N HCI solution, and concentrated under reduced pressure. The solid was collected, and the solid was crystallized with a mixture of dichloromethane: methanol (10:1 ) to give the title compound (4.5Og) as a white powder. The mother liquor was purified by flash chromatography (dichloromethane: methanol =100:1 to 100:5), and evaporated to give the title compound (0.50 g) as a white power. (Total 5.00 g, purity 98.0%, yield 53.0%). 1HNMR (DMSO-d6): 8.16 (s, 1 H), 6.41 (s, 2H), 4.86 (t, 1 H), 4.65 (d, 1 H), 4.48 (d, 1 H), 3.72 (s, 3H), 3.64 (m, 1 H), 3.54 (t, 1 H), 3.31-3.27 (m, 2H), 3.24 (m, 1 H), 2.19 (s, 3H), 2.15 (s, 3H). LC-MS: 349.7 (M, 100%), 351.6 (M+2, 32.7%), 350.6 (M+1 , 19.2 %), 352.7 (M+3, 5.7%). Example 2
(R)^2-ammo-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (IV) and (S)-(2-am\no-4-chloro-7-((4-methoxy-3, S-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (V) [00221 ] Racemic (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihyclro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (III, 500 mg) was separated by chiral HPLC (Chiralpak AS-H, 25% methanol (0.1%DEA)/CO2, 100bar, 50mL/min, 200 nM) to provide the R-enantiomer IV (218 mg, purity > 99%, ee > 99%) and S-enantiomer V (213 mg, purity > 99%, ee > 99%). Example 3
Methanesulfoπic acid 2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (Vl)
[00222] A mixture of [2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (III, 6.00 g, 17.2 mmol), and triethylamine (12.0 ml_, 85.8 mmol) in anhydrous methylene chloride (30.0 ml_) was cooled to 0 0C and then was treated with methanesulfonyl chloride (1.39 ml_, 18.0 mol) dropwise via syringe. After addition, the reaction mixture was warmed to room temperature, allowed to stir for 30 min, poured into ice-water (10OmL), washed with sat. NH4CI solution, dried over sodium sulfate, and evaporated. The crude was purified by flash chromatography (dichloromethane: methanol =100:1 to 100:5), and evaporated to yield the title compound as a white solid (700 mg, purity 99.5%, yield 60.0%). 1H NMR (CD3CI-C/3 ): δ ppm 8.10 (s, 1H), 5.46 (s, 2H), 4.69 (d, 1 H), 4.45 (d, 1 H), 4.37 (m, 1 H), 4.17 (m, 1 H), 3.66 (s, 3H), 3.61 -3.56 (m, 2H), 3.39 (m, 1 H), 2.91 (s, 3H), 2.18 (s, 3H), 2.15 (s, 3H). LC-MS: 427.5 (M, 100%), 429.5 (M+2, 32.7%), 428.5 (M+1 , 19.2 %), 430.5 (M+3, 5.7%). Example 4
^Chloro-S-iodomethyl-T^^methoxy-SyS-dimethyl-pyridin-Σ-ylmethyty-βjT-dihydro-SH-pyrrolofcS- d]pyrimidin-2-ylamine (VII)
[00223] A mixture of methanesulfonic acid 2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (Vl, 200 mg, 0.5 mmol), and sodium iodide (150 mg, 5.0 mmol) in acetone (6.0 mL) was heated to reflux for overnight. The reaction mixture was evaporated and the crude was purified by flash chromatography (dichloromethane :methanol =100:1 to 100:5), and evaporated to give the title compound as a white solid (82.0 mg, purity 99.5%, yield 40.0%). 1H NMR (Acetone-d6 ): δ ppm 8.16 (s, 1 H), 5.85 (s, 2H), 4.73 (d, 1 H), 4.58 (d, 1 H), 3.80 (s, 3H), 3.75 (t, 1 H), 3.60-3.46 (m, 2H), 3.33 (m, 1 H), 2.28 (s, 3H), 2.27 (s, 3H). LC-MS: 459.5 (M, 100%), 461.5 (M+2, 32.7%), 460.5 (M+1 , 19.2 %), 462.4 (M+3, 5.7%). Example 5
^chloro-TW-methoxy-S.S-dimethylpyridin-Σ-ytymethylJ-S-methyl-βjT-dihydro-SH-pyrrolofcS- d]pyrimidin-2-amine (VIII)
[00224] A solution of 4-chloro-5-iodomethyl-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (VII1 25.0 mg, 0.0544 mmol) in anhydrous tetrahydrofuran (500 μL) was treated with 1.0 M of L-Selectride in tetrahydrofuran (326 μL, 0.326 mmol) at 0 0C. The reaction mixture was stirred for 20 minutes at room temperature, quenched with ice-water, adjusted to pH 2.0, purified by flash chromatography (dichloromethane: methanol =100:1 to 100:5), and evaporated to give the title compound as a white solid (VIII1 5.0 mg, purity 99.5%, yield 27.0%). 1H NMR (Acetone-c/6 ): δ ppm 8.17 (s, 1 H), 5.72 (s, 2H), 4.69 (d, 1 H), 4.57 (d, 1 H), 3.79 (s, 3H), 3.75 (t, 1H), 3.26(m, 1 H)1 3.11 (m, 1 H), 2.25 (s, 3H), 2.24 (s, 3H), 1.24 (d, 3H). LC-MS: 333.6 (M, 100%), 335.6 (M+2, 32.7%), 334.7 (M+1 , 19.2 %), 336.6 (M+3, 5.7%). Example 6
2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetonitrile (IX)
[00225] A mixture of methanesulfonic acid 2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (Vl, 200mg, 0.000467 mol) and potassium cyanide (91.3 mg, 0.00140 mol) in dimethyl sulfoxide (5.00 mL) was heated to 65 0C for overnight. After being cooled, the reaction mixture was poured into sat. sodium bicarbonate solution, extracted with dichloromethane: isopropanol (85:15, 20mLx3), evaporated, and the residue was purified by flash chromatography (dichloromethane: methanol =100:1 to 100:5) to give the title compound as a white solid (50.0 mg, purity 99.5%, yield 30.0%). 1H NMR (DMSO-crø ): δ ppm 8.14 (s, 1 H), 6.56 (s, 2H), 4.65 (d, 1H), 4.48 (d, 1H), 3.76 (s, 3H), 3.53(m, 1 H), 3.18 (m, 1 H), 2.83 (m, 2H), 2.17 (s, 3H), 2.15 (s, 3H). LC-MS: 358.6 (M, 100%), 360.6 (M+2, 32.7%), 359.6 (M+1 , 19.2 %), 361.6 (M+3, 5.7%). Example 7
4-Chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-5-piperidin-1-ylmethyl-6,7-dihydro-5H- pyrrol o[2, 3-d]pyrimidin-2-ylamine (X)
[00226] A mixture of methanesulfonic acid 2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (Vl, 100 mg, 0.200 mmol), piperidine (231 μL, 2.34 mmol), potassium carbonate (161 mg, 1.17 mmol), and N,N-dimethylacetamide (1.00 mL) was heated to 60 0C for 4 hours, diluted with water, adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white power (20.0 mg, purity 99.5%, yield 20.0%). 1H NMR (MeOH-d4 ): δ ppm 8.45 (s, 1H), 4.98 (d, 1H), 4.85 (d, 1 H), 4.13 (s, 3H), 3.96 (m, 2H), 3.79 (m, 1H), 3.53-3.45 (m, 2H), 3.33 (t, 2H), 2.47 (s, 3H), 2.40 (s, 3H), 1.90 (m, 6H). LC-MS: 416.6 (M, 100%), 418.6 (M+2, 32.7%), 417.6 (M+1 , 19.2 %), 419.6 (M+3, 5.7%). Example 8
4-CMoro-7-(4-methoxy-3)5-dimethyl-pyridin-2-ylmethyl)-5-morpholin-4-ylmethyl-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-2-ylamine (Xl)
[00227] A mixture of methanesulfonic acid 2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (Vl, 100 mg, 0.200 mmol), morpholine (204 μL, 2.34 mol), potassium carbonate (161 mg, 1.17 mmol) and N,N-dimethylacetamide (3.00 mL) was heated to 60 0C for 4 hours, diluted with water, adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5-→30% CH3CN/H2O, 0.1% TFA). The desired fraction were lyophilized to give the title compound as a white power (9.0 mg, purity 99.5%, yield 9.0%, TFA salt). 1H NMR (DMSO-ofβ ): δ ppm 8.42 (S, 1H), 4.92 (d, 1 H), 4.80 (d, 1 H), 4.12 (s, 3H), 3.95-3.88 (m, 6H), 3.79 (m, 1H), 3.55 (m, 4H)1 3.33 (m, 2H), 2.47 (s, 3H), 2.40 (s, 3H). LC-MS: 418.6 (M, 100%), 420.6 (M+2, 32.7%), 419.6 (M+1 , 19.2 %), 421.6 (M+3, 5.7%). Example 9
5-((1H-pyrazol-1-yl)methyl)-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-2-amine (XII)
[00228] A mixture of methanesulfonic acid 2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (Vl, 50.0 mg, 0.117 mmol), 1 H-pyrazole (8.35 mg, 0.123 mmol) and potassium carbonate (48.4 mg, 0.350 mol) in anhydrous dimethyl sulfoxide (2.0 mL) was heated to 60 0C for 4 hours, diluted with water, adjusted to pH 2.0, purified by reverse- phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fraction was neutralized and extracted with dichloromethane: isopropanol (85:15, 10mLx3) and evaporated to give the title compound as a white solid (5.0 mg, purity 99.5%, yield 10.0%). 1H NMR (DMSO-ctø ): δ ppm 8.46 (dd, 1 H), 8.16 (s, 1 H), 7.86 (d, 1 H), 6.52 (dd, 1 H), 6.38 (s, 2H), 4.77 (d, 1 H), 4.53 (d, 1 H), 3.97 (m, 1 H), 3.89 (m, 1 H), 3.71 (s, 3H), 3.63 (m, 2H), 3.37 (m, 1 H), 2.19 (s, 6H). LC-MS: 399.6 (M, 100%), 401.6 (M+2, 32.7%), 400.6 (M+1 , 19.2 %), 402.6 (M+3, 5.7%). Example 10
5-A2idomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (XIII)
[00229] A mixture of [2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (III, 1.00 g, 2.86 mmol), diisopropyl azodicarboxylate (1.69 mL, 8.58 mmol), and triphenylphosphine (2.25 g, 8.58 mmol) in anhydrous tetrahydrofuran (50.0 mL) was treated with diphenylphosphonic azide (1.85 mL, 8.58 mmol) at room temperature. The mixture was stirred overnight, evaporated, diluted with water (20OmL), adjusted to pH 4.0 with 2N HCI solution, and washed with dichloromethane (30ml_x3); the organic layer was discarded. The aqueous layer was neutralized with 2N NaOH solution, extracted with dichloromethane: isopropanol (85:15, 30mLx3), evaporated, and purified by flash chromatography (dichloromethane: methanol =100:1 to 100:8) to give the title compound as a white solid (1.00 g, purity 93.5%, yield 87.0%). 1H NMR (DMSO-c/6 ): δ ppm 8.13 (s, 1 H), 6.48 (s, 2H), 4.63 (d, 1 H), 4.48 (d, 1 H), 3.69 (s, 3H), 3.61 (t, 1 H), 3.57 (m, 2H), 3.40 (m, 1 H), 3.20 (m, 1 H), 2.16 (s, 3H), 2.14 (s, 3H). LC-MS: 374.6 (M, 100%), 376.6 (M+2, 32.7%), 375.6 (M+1 , 19.2 %), 377.6 (M+3, 5.7%). Example 11
5-Aminomethyl^-chloro-7^4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (XIV)
[00230] A mixture of 5-azidomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIII, 1.0 g, 2.7 mmol), triphenylphosphine (2.10 g, 8.00 mmol), water (0.50 mL, 28.0 mmol) in tetrahydrofuran (25.0 mL) was stirred at room temperature for 12 hrs. The solvent was evaporated. The resulting crude was treated with 1N HCI solution (10OmL) and impurities were extracted with dichloromethane (50mLx2). The aqueous layer was purified by reverse- phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with sat. NaHCO3 solution, extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (400 mg, purity 99.6%, yield 40.0%). 1H NMR (DMSO-d6 ): δ ppm 8.13 (s, 1 H), 6.35 (S1 2H), 4.59 (d, 1 H), 4.50 (d, 1 H), 3.69 (s, 3H), 3.51 (t, 1 H), 3.35 (m, 1 H), 3.12 (m, 1 H), 2.85 (dd, 1 H), 2.53 (dd, 1H), 2.16 (s, 3H), 2.14 (s, 3H). LC-MS: 348.6 (M, 100%), 350.6 (M+2, 32.7%), 349.6 (M+1 , 19.2 %), 351.6 (M+3, 5.7%). Example 12
(1-((2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methyl)-1H-1,2,3-triazol-4-yl)methanol (XV)
[00231] A mixture of 5-azidomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIII, 70.0 mg, 0.187 mmol), copper(l) iodide (7.11 mg, 0.0374 mmol), dimethyl sulfoxide (1.50 mL), water (0.50 mL) and 2-propyn-1 -ol (54.4 μL, 0.934 mmol) was stirred at room temperature for 6 hours. The reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white powder (60.0 mg, purity 99.5%, yield 70.0%). 1H NMR (DMSO-d6 ): δ ppm 8.42 (s, 1H), 7.90 (s, 1 H), 6.74 (s, 2H), 4.69- 4.54 (m, 4H), 4.47 (s, 2H), 3.94 (s, 3H), 3.80 (m, 1H), 3.58 (t, 1 H), 3.38 (dd 1H), 2.23 (s, 3H), 2.18 (s, 3H). LC-MS: 430.6 (M, 100%), 432.5 (M+2, 32.7%), 431.6 (M+1 , 19.2 %), 433.6 (M+3, 5.7%). Example 13
2-(1^(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydrθ'5H-pyrrolo[2,3- d]pyrimidin-5-yl)methyl)-1H-1,2,3-triazol-4-yl)ethanol (XVI)
[00232] A mixture of 5-Azidomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIII, 70.0 mg, 0.187 mmol), copper(l) iodide (7.11 mg, 0.0374 mmol), dimethyl sulfoxide (1.50 mL), water (0.50 mL) and 3-butyn-1 -ol (70.8 μL, 0.934 mmol) was stirred at room temperature for 6 hours. The reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white powder (60.0 mg, purity 99.5%, yield 70.0%). 1H NMR (DMSO-d6): δ ppm 8.45 (s, 1H), 7.80 (s, 1 H), 6.77 (s, 2H), 4.73 (dd, 1 H), 4.63 (m, 2H), 4.50 (m, 2H), 3.94 (s, 3H), 3.78 (m, 1 H), 3.58 (m, 3H), 3.37 (dd 1 H), 2.74 (t, 2H), 2.34 (s, 3H), 2.19 (s, 3H). LC-MS: 444.6 (M, 100%), 446.5 (M+2, 32.7%), 445.6 (M+1 , 19.2 %), 447.5 (M+3, 5.7%). Example 14
4-Chloro-5-((4-((dimethylamino)methyl)-1H-1,2,3-triazol-1-yl)methyl)-7-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (XVII) [00233] A mixture of 5-Azidomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIII, 100 mg, 0.267 mmol), copper(l) iodide (25.4 mg, 0.133 mmol), dimethyl sulfoxide (1.50 ml_), water (0.50 mL) and propargyl(dimethylamine) (0.100 ml_, 0.934 mol) was stirred at room temperature for 6 hours. The reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with sat. NaHCO3, extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (20.0 mg, purity 99.6%, yield 20.0%). 1H NMR (DMSO-d6 ): δ ppm 8.14 (s, 1 H), 7.87 (s, 1 H), 6.51 (s, 2H), 4.61-4.43 (m, 4H), 4.08 (m, 1 H), 3.72 (s, 3H), 3.54 (d, 2H), 3.16 (d, 2H), 2.17 (s 9H), 2.10 (s, 3H). LC-MS: 457.6 (M, 100%), 459.6 (M+2, 32.7%), 458.6 (M+1 , 19.2 %), 460.6 (M+3, 5.7%). Example 15 δ-ff^S-aminopentan-S-ylf-IH-I^S-triazol-i-ylfmethyO^-chloro-T-^-methoxy-S^-dimethylpyridin- 2-yl)methyl)-6J-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (XVIII)
[00234] A mixture of 5-azidomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIII, 100 mg, 0.267 mmol), copper(l) iodide (25.4 mg, 0.133 mmol), dimethyl sulfoxide (1.50 mL), water (0.50 mL) and 1 ,1-diethyl-prop-2-ynylamine (0.125 mL, 0.934 mol) was stirred at room temperature for 6 hours. The reaction mixture was diluted with water (2OmL), adjusted to pH 2.0 with 2N HCI solution, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with sat. NaHCO3 solution, extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (20.0 mg, purity 99.6%, yield 20.0%). (DMSO-d6 ): δ ppm 8.14 (s, 1 H), 7.74 (s, 1H), 6.50 (s, 2H), 4.59-4.41 (m, 4H), 4.09 (m, 1 H), 3.72 (s, 3H), 3.54 (t, 1H), 3.27 (m, 1 H), 2.18 (s, 3H), 2.11 (s, 3H), 0.90 (q, 4H), 0.65 (t, 6H). LC-MS: 485.6 (M, 100%), 487.6 (M+2, 32.7%), 486.6 (M+1 , 19.2 %), 488.6 (M+3, 5.7%). Example 16
N-((2-amino-4-chloro-7^(4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methyl)acetamide (XIX)
[00235] A mixture of 5-aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIV, 50.0 mg, 0.143 mmol), 4-DMAP (17.5 mg, 0.143 mmol) and acetic anhydride (13.5 μL, 0.143 mmol) in dry N,N-dimethylformamide (0.50 mL) was stirred at room temperature for 2 hours. The reaction mixture was diluted with water (2OmL), adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the product (TFA salt) as a white powder (20.0 mg, purity 99.5%, yield 40.0%). 1H NMR (DMSO-ctø): δ ppm 8.43 (s, 1 H), 7.98 (t, 1 H), 4.80 (dd, 1 H)1 4.63 (dd, 1 H), 3.93 (s, 3H), 3.60 (t, 1 H), 3.47-3.35 (m, 2H), 3.28 (dd, 1H), 3.15 (m, 1 H), 2.33 (s, 3H), 2.24 (s, 3H), 1.75 (s, 3H). LC-MS: 390.6 (M, 100%), 392.6 (M+2, 32.7%), 391.6 (M+1 , 19.2 %), 393.6 (M+3, 5.7%). Example 17 N^Σ-amino-A-chloro-T^-methoxy^S-dimethylpyridin-Σ-y^methyiyβJ-dihydro-SH-pyrrolol∑^- d]pyrimidin-5-yl)methyl)benzamide (XX)
[00236] A mixture of 5-aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIV, 50.0 mg, 0.143 mmol), 4-DMAP (1.75 mg, 0.0143 mmol), and benzoic anhydride (32.4 mg, 0.143 mmol) in dry N.N-dimethylformamide (0.50 ml_) was stirred at room temperature for 2 hours. The reaction mixture was diluted with water (20 mL), adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the product (TFA salt) as a white powder (30.0 mg, purity 99.5%, yield 50.0%). 1H NMR (DMSO-d6 ): δ ppm 8.63 (t, 1 H)1 8.34 (s, 1 H), 7.75 (t, 2H), 7.53 (t, 1 H), 7.44 (t, 2H), 6.77 (s, 2H), 4.79 (dd, 1 H), 4.60 (dd, 1 H), 3.90 (s, 3H), 3.60-3.50 (m, 3H), 3.42 (m, 2H), 2.30 (s, 3H), 2.22 (s, 3H). LC-MS: 452.5 (M, 100%), 454.5 (M+2, 32.7%), 453.5 (M+1 , 19.2 %), 455.5 (M+3, 5.7%). Example 18
2-((2-amino^-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methyl)isoindoline-1 ,3-dione (XXI)
[00237] A mixture of [2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (III, 100.0 mg, 0.286 mmol), diisopropyl azodicarboxylate (0.169 mL, 0.858 mmol) and triphenylphosphine (225 mg, 0.858 mmol) in anhydrous tetrahydrofuran (5.00 mL) was treated with phthalimide (44.2 mg, 0.300 mmol). The reaction mixture was stirred at room temperature overnight, evaporated, diluted with water (10OmL), adjusted to pH 4.0 with 2N HCI. The impurities were washed with dichloromethane (30mLx3). The aqueous layer was purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1 % TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (70.0 mg, purity 99.5%, yield 50.0%). 1H NMR (DMSO-d6 ): δ ppm 8.05 (S, 1 H), 7.84 (s, 4H), 6.47 (s, 2H), 4.69 (dd, 1H), 4.41 (dd, 1 H), 3.80 (m, 1H), 3.72 (s, 3H), 3.68- 356 (m, 3H), 3.42-3.26 (m, 2H), 2.17 (s, 3H), 2.14 (s, 3H). LC-MS: 478.5 (M, 100%), 480.5 (M+2, 32.7%), 479.5 (M+1 , 19.2 %), 481.5 (M+3, 5.7%). Example 19 i-ffΣ-amino-^chloro-T-W-methoxy-SjS-dimethylpyridin-Σ-yOmethyty-βS-dihydro-SH-pyrrolofcS- d]pyrimidin-5-yl)methyl)piperidine-2, 6-dione (XXII)
[00238] A mixture of [2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (III, 100.0 mg, 0.285 mmol), diisopropyl azodicarboxylate (0.169 mL, 0.858 mmol) and triphenylphosphine (225 mg, 0.858 mmol) in anhydrous tetrahydrofuran (5.00 mL) was treated with glutarimide (34.0 mg, 0.300 mmol) at room temperature. The reaction mixture was stirred at room temperature overnight, evaporated, diluted with water (10OmL), and adjusted to pH 4.0 with 2N HCI. The impurities were washed with dichloromethane (30mLx3). The aqueous layer was purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with aqueous NaHCO3, extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (55.0 mg, purity 99.5%, yield 43.0%). 1H NMR (DMSO-d6 ): δ ppm 8.11 (s, 1 H), 6.42 (s, 2H), 4.63 (dd, 1H), 4.44 (dd, 1H), 3.85 (dd, 1H), 3.74 (m, 1 H), 3.72 (s, 3H), 3.65 (m, 1H), 3.52-3.40 (m, 2H), 3.06 (m, 1 H), 2.53 (t, 4H), 2.15 (s, 6H), 1.80 (t, 2H). LC-MS: 444.6 (M, 100%), 446.5 (M+2, 32.7%), 445.5 (M+1 , 19.2 %), 447.5 (M+3, 5.7%). Example 20
N-ii∑-amino-A-chloro-T-iiA-methoxy-S^-dimethylpyridin-Σ-y^methyiyβJ-dihydro-SH-pyrrolol∑^- d]pyrimidin-5-yl)methyl)methanesulfonamide (XXIII)
[00239] A mixture of 5-(aminomethyl)-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (XIV, 50mg, 0.143 mmol) and triethylamine (0.1 ml_, 0.717 mmol) in anhydrous methylene chloride (1.0 mL) was cooled to 0 0C and treated with methanesulfonyl chloride (22 μL, 0.287 mmol) dropwise via syringe. After addition, the reaction mixture was warmed to room temperature, allowed to stir for 30 minutes, and evaporated. The crude was diluted with water (3OmL), adjusted to pH 4.0 with 2N HCI and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (15.0 mg, purity 99.6%, yield 24.0%). 1H NMR (DMSO-d6 ): δ ppm 8.16 (s, 1 H), 7.28 (t, 1 H), 6.47 (s, 2H), 4.67 (d, 1 H), 4.48 (d, 1 H), 3.72 (s, 3H), 3.57 (t, 1 H), 3.30-3.25 (m, 3H), 2.99 (m,1 H), 2.82 (s, 3H), 2.19 (S, 3H), 2.16 (s, 3H). LC-MS: 426.5 (M, 100%), 428.5 (M+2, 32.7%), 427.5 (M+1 , 19.2 %), 429.5 (M+3, 5.7%). Example 21
4-chloro-5^(dimethylamino)methyl)-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6J-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-amine (XXIV)
[00240] To a suspension of 5-(aminomethyl)-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (XIV, 60mg, 0.171 mmol) in acetonitrile (1.4mL) was added 37% formaldehyde (36.7 μL, 0.492 mmol), followed by 30% aqueous acetic acid (28.0 μL, 0.492 mmol). The mixture was stirred for 5 minutes and then cooled to O0C, and 1.0 M of sodium cyanoborohydride in tetrahydrofuran (0.492 mL, 0.492 mmol) was added to the reaction mixture. The reaction mixture was warmed to room temperature, allowed to stir for 30 minutes, quenched with aqueous sodium bicarbonate solution, adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (12.0 mg, purity 99.6%, yield 20.0%). 1H NMR (DMSO-c/6 ): δ ppm 8.16 (s, 1 H), 5.73 (s, 2H), 4.71 (d, 1 H), 4.52 (d, 1H), 3.78 (s, 3H), 3.60-3.49 (m, 2H), 3.36 (m, 1 H), 2.50 (dd,1 H), 2.34 (t, 3H), 2.23 (s, 3H), 2.18 (s, 3H). LC-MS: 376.6 (M, 100%), 378.6 (M+2, 32.7%), 377.7 (M+1 , 19.2 %), 379.6 (M+3, 5.7%). Example 22
(2Ry2-amino-N^(2-amino^-chloro-7^(4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methyl)-3-methylbutanamide (XXV) [00241] A mixture of 5-Aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (XIV, 50.0 mg, 0.143 mol), (R)-2-(tert- butoxycarbonylamino)-3-methylbutanoic acid (31.1 mg, 0.143 mol), 1 -hydroxybenzotriazole (19.4 mg, 0.143 mol), N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (27.5 mg, 0.143 mol) and triethylamine (99.9 μl_, 0.717 mol) in N,N-dimethylformamide (2.0 mL) was stirred at room temperature overnight. The reaction mixture was diluted with water (60ml), and extracted with dichloromethane: isopropanol (85:15, 20ml_x3). The organic layer was evaporated and treated with trifluoroacetic acid (0.110 mL, 1.43 mol) overnight. The reaction mixture was evaporated, diluted with water (20 mL), adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 15mLx3), and evaporated to give the title compound as a white solid (18.0 mg, purity 99.6%, yield 28.0%). 1H NMR (Acetone-αtø ): δ ppm 8.16 (s, 1 H), 7.20-7.10 (s, 3H), 5.79 (d, 2H), 4.66-4.49 (m, 2H), 3.78 (m, 1 H), 3.78 (s, 3H), 3.76-3.32 (m, 5H), 2.22 (s, 3H), 2.21 (s, 3H), 2.04 (m, 1 H), 0.78 (m, 6H). LC- MS: 447.6 (M, 100%), 449.6 (M+2, 32.7%), 448.6 (M+1 , 19.2 %), 450.6 (M+3, 5.7%). Example 23
S-fΣ-aminoethylW-chloro-V^ft-methoxy-Sβ-dimethylpyridin-Σ-yOmethylJ-βJ-dihydro-SH- pyrrolo[2,3-d]pyrimidin-2-amine (XXVI)
[00242] A solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetonitrile (IX, 50mg, 0.0139 mmol) in anhydrous tetrahydrofuran (0.50 mL) was treated dropwise with 1.0 M of L-selectride in tetrahydrofuran (0.836 mL, 0.836 mmol) at 0 0C. The reaction mixture was warmed to room temperature, allowed to stir for 1hr, carefully quenched with ice water, adjusted to pH 7.0, evaporated, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 10mLx3), and evaporated to give the title compound as a white solid (8.0 mg, purity 99.6%, yield 20.0%). 1H NMR (Acetone-crø): δ ppm 8.15 (s, 1 H), 5.69 (s, 2H), 4.70 (d, 1 H), 4.52 (d, 1H), 3.78 (s, 3H), 3.66 (t, 1 H), 3.40-3.31 (m, 2H), 3.20 (q, 2H), 2.24 (s,3H), 2.22 (s, 3H), 1.76(m, 2H). LC-MS: 362.6 (M, 100%), 364.6 (M+2, 32.7%), 363.6 (M+1 , 19.2 %), 365.6 (M+3, 5.7%). Example 24
[Σ-Amino^-chloro-T^-methoxy-SyS-dimethyl-pyridin-Σ-ylmethyty-βS-dihydro-SH-pyrrolofcS- d]pyrimidin-5-yl]-acetic acid methyl ester (XXVII) and2-(2-amino-4-chloro-7-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyhmidin-5-yl)acetic acid (XXVIII) [00243] A mixture of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetonitrile (IX, 100 mg, 0.278 mol) in anhydrous methanol (1.00 mL) was cooled to -10 0C and treated with HCI (4.0 M in 1 ,4-dioxane, 0.418 mL, 1.70 mmol) for 1 hour at -10 0C. The mixture was placed in a refrigerator (0 °C to 5 0C) for 6 days, diluted with ice-water (5mL), neutralized with sodium bicarbonate solution, extracted with a mixture of dichloromethane: isopropanol (90:10, 10mLx3), and evaporated. The residue was purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 10ml_x3), and evaporated to give [2-Amino-4-chloro-7-(4-methoxy- S.S-dimethyl-pyridin^-ylmethyO-θ.y-dihydro-SH-pyrrolo^.S-dlpyrimidin-S-yll-acetic acid methyl ester as a white solid (37.0 mg, purity 99.6%, yield 30.0%). 1H NMR (CDCI3-d3 ): δ ppm 8.17 (s, 1 H), 5.11 (s, 2H), 4.62 (s, 2H)1 3.75 (s, 3H), 3.65 (s, 3H)1 3.57 (t, 1H)1 3.21 (m, 1 H)1 3.01 (m, 1 H)1 2.44 (m, 1 H)1 2.32 (m, 1 H), 2.24 (s, 3H), 2.21 (s, 3H). LC-MS: 391.6 (M1 100%), 393.6 (M+2, 32.7%), 392.6 (M+1 , 19.2 %), 394.6 (M+3, 5.7%).
[00244] The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 10ml_x3), and evaporated to give 2-(2-amino-4-chloro-7-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid as a white solid (52mg, purity 99.6%, yield 52.0%). 1H NMR (Acetone- d6 ): δ ppm 8.49 (s, 1H), 6.44 (s, 2H), 5.05 (d, 1 H), 5.97 (d, 1H), 4.16 (t, 1 H), 4.04 (s, 3H), 3.72 (m, 2H)1 2.97 (m, 1 H), 2.76 (m, 1 H), 2.46 (s,3H), 2.41 (s, 3H). LC-MS: 377.6 (M, 100%), 379.6 (M+2, 32.7%), 378.6 (M+1 , 19.2 %), 380.6 (M+3, 5.7%). Example 25
2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)ethanol (XXIX)
[00245] A solution of [2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid methyl ester (XXVII, 20.0 mg, 0.0510 mol) in anhydrous tetrahydrofuran (0.50 mL) was treated with 1.0 M of L-Selectride in tetrahydrofuran (0.204 mL) at 0 0C. The mixture was warmed to room temperature, stirred for 20 minutes, quenched with ice water, adjusted to pH 7.0, concentrated under reduced pressure, extracted with a mixture of dichloromethane: 2- propanol (85:15, 10mLx3), dried over Na2SO4, and evaporated to give the title compound as a white power (6.0 mg, purity 96.1%, yield 31.0%). 1H NMR (Acetone-c/6 ): δ ppm 8.15 (s, 1 H), 5.69 (s, 2H), 4.72 (d, 1 H), 4.52 (d, 1 H), 3.79 (s, 3H), 3.67 (t, 1 H)1 3.59 (m, 1 H), 3.43 (m, 1 H), 3.36 (m, 1 H), 2.24 (s, 3H), 2.23 (s, 3H). LC-MS: 363.6 (M, 100%), 365.6 (M+2, 32.7%), 364.6 (M+1 , 19.2 %), 366.6 (M+3, 5.7%). Example 26
5ΛBn-buXy\^-ch\oro-7^(4-methoxy-3,5-d\methy\pyr\d\n-2-y\)methy\)-6,7-d\hydro-5H-pyrro\o[2,3- d]pyrimidin-2-amine (XXX)
[00246] A procedure similar to that described in steps 1 and 2 below is reported in
WO2005/028434. A procedure similar to that described in step 3 below is reported in J. Org. Chem., 1996, 6/(4), 1261-1266.
Step 1. 2-Bromo-3,3-dimethyl-butyraldehyde (1B)
[00247] A mixture of 3,3-dimethyl-butyraldehyde (1 A, 10.0 mL, 0.0797 mol), 5,5-dibromobarbituric acid (14 g, 0.048 mol), 45% Hydrogen bromide (2.0 mL, 0.016 mol), acetic acid (1.0 mL, 0.018 mol) and methylene chloride (150 mL) was stirred at room temperature for 5 hours. The reaction was filtered over a celite pad, the filtrate was washed with aq. Na2SO3 (2x50mL) and then brine (2x50mL). The organic layer was evaporated to give the title compound as a yellow oil (8.5Og1 purity 99.0%, yield 59.0%).
1HNMR (DMSO-d6): 9.46 (d, 1 H), 4.35 (d, 1 H), 1.07 (s, 9H).
Step 2. 2-amino-5-tert-butyl-3H-pyrrolo[2,3-d]pyrimidin-4(7H)-one (1C)
[00248] To a suspension of sodium acetate (3.90 g, 0.0475 mol), water (6.84 mL, 0.380 mol),
N,N-dimethylacetamide (44.2 mL, 0.475 mol) and 2-bromo-3,3-dimethyl-butyraldehyde (1 B, 8.50 g,
0.0475 mol) was added 2,4-diamino-6-hydroxypyrimidine (5.99 g, 0.0475 mol). The reaction mixture was stirred at room temperature overnight whereupon the starting materials gradually dissolved and the product precipitated out. The solid was collected by filtration, washed with water (50mLx2), and dried over high vacuum to give the title compound as a white solid (6.1Og, purity 99.0%, yield 62.0%). 1HNMR
(DMSO-d6): 10.60 (s, 1 H), 10.10 (s, 1 H), 6.26 (d, 1 H), 5.98 (s, 2H), 1.31 (s, 9H). LC-MS: 207.0 (M,
100%), 208.0 (M+1 , 15.0%)
Step 3. N-(5-tert-butyl-4-oxo-4,7-dihydro-3H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
[00249] A mixtute of 2-amino-5-tert-butyl-3H-pyrrolo[2,3-d]pyrimidin-4(7H)-one (1C, 710 mg,
0.0034 mol) and 2,2-dimethylpropanoyl chloride (1.48 mL, 0.0120 mol) in pyridine (7.0 mL, 0.086 mol) was stirred for 2 hours at 90 0C. The reaction mixture was evaporated, the residue was taken up in 37%
NH4OH (3mL) and MeOH (1OmL). The mixture was stirred at room temperature for 30 minutes. The solid was collected, washed with water (5mlχ5), dried with high vacuum pump to give the title compound as a white solid (1.00 g, purity 80.0%, yield 80.0%). LC-MS: 290.7 (M, 100%).
Step 4. N-(5-tert-butyl-4'θxo-7-tosyl-4, 7-dihydro-3H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide (W)
[00250] The solid (800 mg, 0.003 mol) obtained in the previous step was dissolved in N1N- dimethylformamide (8.00 mL). The resulting solution was flushed with nitrogen at 0 0C, and then treated with sodium hydride (218 mg, 0.00909 mol) in N.N-dimethylformamide (3.0 mL, 0.039 mol) over 15min, followed by p-toluenesulfonyl chloride (0.398 mL, 0.00276 mol). The mixture was warmed to room temperature and stirred overnight, quenched with water (10OmL), adjusted to pH 7.0 with 2N HCI solution.
The solid was collected, washed with water (2OmL), dried over high vacuum pump to give the title compound as a white solid (1.22 g, purity 95.0%, yield 94.6%). LC-MS: 444.6 (M1 100%).
Step 5. N-(5-tert-butyl-4-oxo-4,5,6, 7-tetrahydro-3H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
[00251] A mixture of N-(5-tert-butyl-4-oxo-7-tosyl-4,7-dihydro-3H-pyrrolo[2,3-d]pyrimidin-2- yl)pivalamide (1 D, 1.00 g, 2.25 mmol), 20% Pd(OH)2 on carbon (Pearlman's catalyst, 1.00 g, 0.00712 mol) and ethanol (40 mL, 0.5 mol) was stirred under hydrogen (50 psi) for 10 days. The reaction was filtered over a celite pad, and evaporated to yield the product N-[5-tert-Butyl-4-oxo-7-(toluene-4-sulfonyl)-
4,5,6,7-tetrahydro-3H-pyrrolo[2,3-d]pyrimidin-2-yl]-2,2-dimethyl-propionamide as a white solid (1.00 g, purity 89.4%, yield 71.0%). LC-MS: 446.6 (M, 100%).
Step 6. 2-Amino-5-tert-butyl-3,5,6,7-tetrahydro-pyrrolo[2,3-d]pyrimidin-4-one (1E)
[00252] A mixture of N-[5-tert-Butyl-4-oxo-7-(toluene-4-sulfonyl)-4,5,6,7-tetrahydro-3H- pyrrolo[2,3-d]pyrimidin-2-yl]-2,2-dimethyl-propionamide (0.80 g, 0.0016 mol) and sulfuric acid (5.00 mL,
0.0938 mol) was heated to 100 0C for 3 hours. The mixture was cooled, diluted with ice-water (120 mL), neutralized to pH 7 with 1 M NaOH solution, extracted with a mixture of dichloromethane: isopropanol
(85:15, 100ml_x2), and evaporated to give the title compound as a white solid (330.0 mg, purity 97.0%, yield 96.0%). 1HNMR (DMSO-d6): 9.48 (s, 1 H), 6.20-6.19 (m, 3H), 3.31-3.16 (m, 2H), 2.78 (q, 1 H), 0.87
(s, 9H). LC-MS: 208.9 (M, 100%), 209.9 (M+1 , 15.0).
Step 7. 5-tert-butyl-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (1F)
[00253] A mixture of 2-amino-5-tert-butyl-3,5,6,7-tetrahydro-pyrrolo[2,3-d]pyrimidin-4-one (1E,
0.200 g, 0.000960 mol) and acetic anhydride (0.906 ml_, 0.00960 mol) was heated to 100 0C for 20 minutes. The reaction mixture was evaporated to dryness. To the residue were added benzyltriethylammonium chloride (0.437 g, 0.00192 mol) and phosphoryl chloride (0.537 ml_, 0.00576 mol). The resulting mixture was heated to 100 °C for 30 minutes, and evaporated. The residue was taken up in 6N HCI solution (2OmL). The mixture was stirred at 60 0C for 1.0 hour. After being cooled, the mixture was adjusted to pH 4.0, purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O,
0.1% TFA). The desired fractions were neutralized with NaHCθ3, extracted with dichloromethane: isopropanol (85:15, 30mLx3), and evaporated to give the title compound as a white solid (105 mg, purity
97.0%, yield 96.0%). 1HNMR (DMSO-d6): 9.48 (s, 1 H), 6.20-6.19 (m, 3H), 3.31 -3.16 (m, 2H), 2.78 (q,
1 H), 0.87 (s, 9H). LC-MS: 208.9 (M, 100%), 209.9 (M+1 , 15.0).
Step 8. 5-tert-Butyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-2-ylamine (XXX)
[00254] A mixture of 5-tert-Butyl-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (1F,
100 mg, 0.441 mol), 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine (86.0 mg, 0.000463 mol) and sodium hydride (22.2 mg, 0.000926 mol) in N,N-dimethylformamide (1.50 mL, 0.0194 mol) was flushed with an atmosphere of nitrogen. The mixture was heated to 100 °C for 1 hour. After cooling, the mixture was diluted with ice-water (3OmL), adjusted to pH 4.0 with 6N HCI, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with
NaHCO3, extracted with dichloromethane: isopropanol (85:15, 20mLx3), and evaporated to give the title compound as a white solid (130 mg, purity 97.4%, yield 76.0%). 1HNMR (DMSO-d6): 8.14 (s, 1 H), 6.39
(s, 2H), 4.75 (d, 1H), 4.31 (d, 1H), 3.74 (s, 3H), 3.47 (t, 1 H), 3.23 (m, 1 H), 2.77 (m, 1 H), 2.16 (s, 3H)1 2.14
(s, 3H), 0.78 (s, 9H). LC-MS: 376.1 (M, 100%), 378.1 (M+2, 32.7%), 377.1 (M+1 , 19.2 %), 379.1 (M+3,
5.7%).
Example 27
1-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)ethanol (XXXI)
Step 1. 4-Chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-5-trimethylsilanylethynyl-7H- pyrrolo[2, 3-d]pyrimidin-2-ylamine (2A)
[00255] A round bottom flask was charged with 4-chloro-5-iodo-7-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine (I, 5.0O g, 11.3 mmol) and Dimethyl sulfoxide (100 mL, 1410 mmol). The mixture was degassed. (Trimethylsilyl)acetylene (1.67 mL, 11.8 mmol), triethylamine (8 ml_, 60 mmol), tetrakis(triphenylphosphine)palladium(0) (500 mg, 0.4 mmol), and copper(l) iodide (200 mg, 1 mmol) were added. The reaction was stirred under an atmosphere of
Nitrogen at 60 °C for 22h. Water (250 ml_) was added, and the mixture was cooled in an ice bath. The precipitate was collected by filtration, re-dissolved in 125 ml. EtOAc and 25 ml_ THF. Hexane (125 ml_) was added to precipitate the desired product. The mixture was let stand overnight and cooled. Filtration gave the title compound.
Step 2. 1^2-ammo^-chloro-7-((A-methoxy-3,5-dimethy\pyndin-2-yl)methyl)-7H-pyTrolo[2,3- d]pyrimidin-5-yl)ethanone (2B)
[00256] 4-Chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-5-trimethylsilanylethynyl-7H- pyrrolo[2,3-d]pyrimidin-2-ylamine (100 mg, 0.3 mmol) was treated with 2 ml of 20% sulfuric acid. After the reaction was complete, the pH was adjusted to 8 with 10M NaOH, and purified by reverse-phase HPLC
(10→35% CH3CN/H2O, 0.1 %TFA). The desired fractions were neutralized with NH4OH, extracted with dichloromethane, and evaporated to yield the title compound as a white solid.
Step 3. 1 -(2-amino-4-chloro-7-((4-methoxy-3, 5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)ethanol (XXXI)
[00257] A solution of 1 -(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7H- pyrrolo[2,3-d]pyrimidin-5-yl)ethanone (2B, 33mg, 0.0917mmol) in anhydrous tetrahydrofuran (0.50 mL) was treated with 1.0 M of L-selectride in tetrahydrofuran (0.550 mL, 0.550 mmol ) dropwise at 0 0C. The mixture was warmed to room temperature, allowed to stir for 20 min, quenched with ice water, adjusted to pH 7.0, concentrated under reduced pressure, and purified by reverse-phase preparative HPLC (5→30%
CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 10mLx3), and evaporated to give the title compound as a white solid (7.0 mg, purity 99.6%, yield 21.0%). 1HNMR (Acetone-d6): 8.17 (s, 1 H), 5.72 (s, 2H), 4.88 (d, 1 H),
4.37 (d, 1H), 4.28 (m, 1 H), 4.09 (s, 1 H), 3.78 (s, 3H), 3.62 (t, 1 H), 3.53 (t, 1 H), 3.45 (m, 1 H), 2.08 (s,6H),
0.93 (d, 3H). LC-MS: 363.6 (M, 100%), 365.6 (M+2, 32.7%), 364.6 (M+1 , 19.2 %), 366.6 (M+3, 5.7%).
Example 28
A-chioro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyiy5,5-dimethyi-6J-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-amine (XXXII)
[00258] A procedure similar to that described in step 1 below is reported in Tetrahedron:
Asymmetry, 2004, 15, 3323-3327. A procedure similar to that described in step 2 below is reported in
WO2006/110783. A procedure similar to that described in steps 3 and 4 below is reported in
Heterocycles, 1993, 35(2), 925-936.
Step 1. Diethyl 2-(2-methyl-1-nitropropan-2-yl)malonate (3B)
[00259] A mixture of 2-isopropylidene-malonic acid diethyl ester (20.0 mL, 0.102 mol), nitromethane (55.4 mL, 1.02 mol) and N,N,N',N'-tetramethylguanidine (6.4 mL, 0.051 mol) was stirred at room temperature for 16 hours. The reaction mixture was diluted with 2N HCI (200 mL). The organic layer was separated, washed with a saturated sodium chloride solution, dried over sodium sulfate, and evaporated to yield the title compound as yellow oil (26.7g, purity 94.0%, yield 70.6%). 1HNMR (DMSO- d6): 4.75 (s, 2H), 4.16 (q, 2H), 4.12 (q, 2H)1 3.67 (s, 1H), 1.19 (t, 6H), 1.15 (s, 6H).
Step 2. ethyl 4,4-dimethyl-2-oxopyrrolidine-3-carboxylate (3C)
[00260] A mixture of 2-(1 ,1-Dimethyl-2-nitro-ethyl)-malonic acid diethyl ester (3B, 21.0 g, 0.0804 mol) and nickel chloride hexahydrate (8.2 g, 0.034 mol) in methanol (200 mL) was treated with sodium tetrahydroborate (5.47 g, 0.145 mol) in portions at 0 0C for 6 hours. The resulting black solution was quenched with aqueous NH4CI (2000 mL) and diluted with dichloromethane (1000 mL). The organic layer was separated and washed with aqueous NH4CI (200mLx6) containing 5% NH4OH until the organic layer turned to light yellow. The organic layer was dried over Na2SO4, and evaporated. The crude was purified by flash chromatography (methylene chloride: methanol 100:0 to 100:5). The desired fractions were evaporated to give the title product as a light green oil (10.5g, purity 95.0%, yield 67.0%). 1HNMR
(DMSO-d6): 7.89 (s, 1H), 4.11 (q, 2H), 3.07 (d, 1H), 2.95 (d, 1H), 2.83 (s, 1H), 1.21 (t, 3H), 1.16 (s, 3H),
1.02 (s, 3H).
Step 3. Ethyl 5-ethoxy-3,3-dimethyl-3,4-dihydro-2H-pyrrole-4-carboxylate (3D)
[00261] A solution of 4,4-dimethyl-2-oxo-pyrrolidine-3-carboxylic acid ethyl ester (3C, 10.0 g,
0.0540 mol) in anhydrous methylene chloride (40.0 mL) under N2 was treated with a solution of triethyloxonium tetrafluoroborate (11.9 g, 0.0626 mol) in anhydrous methylene chloride (40.0 mL), stirred at room temperature overnight, poured into water (8OmL), and allowed to stand for 30 min. The organic layer washed with sat. NaHCO3, water, dried, and evaporated to give the title product as a light green oil
(11.5g, purity 95.0%, yield 59.0%). 1HNMR (DMSO-d6): 4.15 (q, 2H), 4.07 (q, 2H), 3.30 (s, 1 H), 3.28 (s,
1 H), 3.17 (s, 1 H), 1.20 (t, 3H), 1.18 (s, 3H), 1.13 (s, 3H), 0.98 (s, 3H). LC-MS: 213.9 (M, 100%), 214.9
(M+1 , 22.5%).
Step 4. 2-amino-5,5-dimethyl-6,7-dihydro-3H-pyrrolo[2,3-d]pyrimidin-4(5H)-one (3E)
[00262] A mixture of sodium ethoxide (4.94 g, 0.0727 mol) in anhydrous ethanol (70 mL) was treated successively with guanidine hydrochloride (2.78 g, 0.0291 mol) and 2-ethoxy-4,4-dimethyl-4,5- dihydro-3H-pyrrole-3-carboxylic acid ethyl ester (3D, 6.20 g, 0.0291 mol). The mixture was heated to reflux overnight and evaporated. The residue was diluted with water (50 mL), adjusted to pH 5.0 with 1 N
HCI solution, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white solid (0.8Og, purity
99.0%, yield 15.2%). 1HNMR (DMSO-d6): 10.6 (s, 1 H), 7.16 (s, 2H), 3.19 (s, 2H), 1.23 (s, 6H). LC-MS:
180.9 (M, 100%), 181.9 (M+1 , 10.0%).
Step 5. 4-chloro-5,5-dimethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (3F)
[00263] A mixture of 2-amino-5,5-dimethyl-3,5,6,7-tetrahydro-pyrrolo[2,3-d]pyrimidin-4-one (3E,
0.80 g, 4.4 mmol) and acetic anhydride (4.19 mL, 44.4 mmol) was heated to reflux for 1 hour. The mixture was evaporated to dryness, and the residue was treated with benzyltriethylammonium chloride
(2.02 g, 8.88 mmol) and phosphoryl chloride (2.48 mL, 26.6 mmol). The mixture was heated to reflux for
40 minutes, and evaporated. The residue was poured into ice-water (5 mL), diluted with 6N HCI (2OmL), and stirred at 50 °C for 1.5 hours. After cooling, the mixture was adjusted to pH 4.0, purified by reverse- phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white solid (0.35g, purity 99.0%, yield 39.0%). 1HNMR (DMSO- d6): 8.12 (s, 1 H), 6.85 (s, 1 H), 3.32 (s, 2H), 1.30 (s, 6H). LC-MS: 198.8 (M, 100%), 200.8 (M+2, 32.7%), 199.9 (M+1 , 19.2 %), 201.8 (M+3, 5.7%).
Step 6. 4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5,5-dimethyl-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-2-amine (XXXII)
[00264] A mixture of 4-chloro-5,5-dimethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (3F,
100.0 mg, 0.503 mmol) and 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine (98.1 mg, 0.528 mmol), NaH (25.4 mg, 1.06 mmol) in anhydrous dimethyl sulfoxide (1.00 mL) was stirred at 100 0C for 1 hr. After cooling, the mixture was diluted with ice-water (3OmL), adjusted to pH 4.0 with 6N HCI, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were neutralized with NaHCO3, extracted with dichloromethane: isopropanol (85:15, 10mLx3), and evaporated to give the title compound as a white solid (50 mg, purity 99.4%, yield 30.0%). 1HNMR (DMSO-d6): 8.16 (s, 1 H), 6.38 (s, 2H), 4.59 (s, 2H), 3.72 (s, 3H), 3.18 (s, 1 H), 2.19 (s, 3H), 2.17 (s, 3H), 1.28 (s, 6H). LC- MS: 347.7 (M, 100%), 349.7 (M+2, 32.7%), 348.7 (M+1 , 19.2 %), 350.7 (M+3, 5.7%). Example 29
1-(2'-amino^'-chloro-7'-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6',7'-dihydrospiro[piperidine- 4,5'-pyrrolo[2,3-d]pyrimidine]-1-yl)ethanone (XXXIII)
[00265] A procedure similar to that described in step 1 below is reported in J. Med. Chem.,
2004, 47, 2587-2598. A procedure similar to that described in step 2 below is reported in Tetrahedron: Asymmetry, 2004, 15, 3323-3327. A procedure similar to that described in step 3 below is reported in WO2006/110783. A procedure similar to that described in steps 3, 4, and 5 below is described in Heterocycles, 1993, 35(2), 925-936.
Step 1. 2-{1-tert-Butoxycarbonyl-piperidin-4-ylidene)-malonic acid diethyl ester (4B) [00266] A mixture of anhydrous tetrahydrofuran (350 mL) and anhydrous carbon tetrachloride (50 mL) was cooled to 0 0C and treated with titanium tetrachloride (22.1 mL, 0.201 mol). The resulting yellow suspension was stirred at 0 0C for 5 minutes, and treated sequentially with 4-oxo-piperidine-1-carboxylic acid tert-butyl ester (20.0 g, 0.100 mol) in anhydrous tetrahydrofuran (50 mL) and ethyl malonate (15.2 mL, 0.100 mol). The reaction mixture was stirred at 0 0C for 30 minutes and then treated with a solution of dry pyridine (32.5 mL, 0.402 mol) in anhydrous tetrahydrofuran (60 mL). The mixture was stirred at 0 0C for 1 hour and then at room temperature for 72 hours. The reaction mixture was quenched with water (3OmL) and extracted with dichloromethane (200mLx2). The combined organic layers were washed with brine (5OmL), sat. NaHCO3 solution (5OmL), dried over Na2SO4, and evaporated to give the title compound as yellow oil (15.0 g, purity 90.0%, yield 39.0%). 1HNMR (DMSO-d6): 4.17 (q, 4H), 3.41 (t, 4H), 2.55 (t, 4H), 1.41 (s, 9H), 1.20 (t, 3H), 1.18 (t, 3H). Step 2. 2-(1-tert-Butoxycarbonyl-4-nitromethyl-piperidin-4-yl)-malonic acid diethyl ester (4C) [00267] A mixture of 2-(1-tert-butoxycarbonyl-piperidin-4-ylidene)-malonic acid diethyl ester (4B,
15 g, 0.044 mol), nitromethane (23.8 mL, 0.439 mol) and N,N,N',N'-tetramethylguanidine (2.8 ml_, 0.022 mol) was stirred at room temperature for 16 hours. The mixture was diluted with 2N HCI solution (20OmL) and shaken in a separatory funnel. The organic layer was washed with saturated NaCI solution, dried with sodium sulfate and evaporated to yield the title compound as a yellow oil (12.6g, purity 90.0%, yield 64.0%). 1HNMR (DMSO-d6): 5.01 (s, 2H), 4.15 (t, 4H), 3.48 (m, 2H), 3.17 (m, 2H)1 1.72-1.60 (m, 4H), 1.40 (s, 9H), 1.20 (t, 3H), 1.18 (t, 3H).
Step 3. 3-Oxo-2,8-diaza-spiro[4.5]decane-4,8-dicarboxylic acid 8-tert-butyl ester 4-ethyl ester (4D) [00268] A mixture of 2-(1-tert-Butoxycarbonyl-4-nitromethyl-piperidin-4-yl)-malonic acid diethyl ester (4C, 12.0 g, 0.0298 mol) and nickel chloride hexahydrate (3.5 g, 0.015 mol) in methanol (100 mL) was treated with sodium tetrahydroborate (6.77 g, 0.179 mol) in portions at 0 0C for 6 hours. The resulting black solution was quenched with aqueous NH4CI (300 mL) and diluted with dichloromethane (500 mL). The organic layer was separated and washed with aqueous NH4CI (200mLx6) containing 5% NH4OH until the organic layer turned to light yellow. The organic layer was dried over Na2SO4, and evaporated. The crude was purified by flash chromatography (methylene chloride: methanol 100:0 to 100:5). The desired fractions were evaporated to give the product as a light yellow oil (9.2Og, purity 90.0%, yield 85.0%). 1HNMR (DMSO-d6): 7.95 (s, 1H), 4.15 (t, 2H), 3.45-3.30 (m, 4H), 3.14 (s, 2H), 3.00 (s, 1H), 1.55-1.35 (m, 4H), 1.38 (s, 9H), 1.20 (t, 3H), 1.18 (t, 3H).
Step 4. 3-Ethoxy-2,8-diaza-spiro[4.5]dec-2-ene-4,8-dicarboxylic acid 8-tert-butyl ester 4-ethyl ester (4E)
[00269] To a mixture of triethyloxonium tetrafluoroborate (6.21 g, 0.0327 mol) in anhydrous methylene chloride (40 mL) was added dropwise a solution of 3-oxo-2,8-diaza-spiro[4.5]decane-4,8- dicarboxylic acid 8-tert-butyl ester 4-ethyl ester (4D, 9.20 g, 0.0282 mol) in anhydrous methylene chloride (40 mL) under nitrogen. The mixture was stirred at room temperature overnight, and poured into water (8OmL). The organic layer was washed with sat. NaHCO3 and water, dried over Na2SO4, and evaporated to give the title product as a light yellow oil (6.4Og, purity 90.0%, yield 58.0%). LC-MS: 354.8 (M, 100%), 355.7 (M+1 , 22.5%).
Step 5. tert-Butyl 2'-amino-4'-oxo-3',4',6', 7'-tetrahydrospiro[piperidine-4,5'-pyrrolo[2,3- djpyrimidine]- 1 -car boxy late (4F)
[00270] A mixture of sodium ethoxide (3.69 g, 0.0542 mol) in anhydrous ethanol (50 mL, 0.8 mol) was treated successively with guanidine hydrochloride (1.72 g, 0.0180 mol) and 3-Ethoxy-2,8-diaza- spiro[4.5]dec-2-ene-4,8-dicarboxylic acid 8-tert-butyl ester 4-ethyl ester (4E, 6.40 g, 0.0180 mol). The mixture was heated to reflux overnight and evaporated. The residue was diluted with water (5OmL), adjusted to pH 5.0 with 1 N HCI solution, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white solid (0.37g, purity 90.0%, yield 5.7%). 1HNMR (DMSO-d6): 9.56 (s, 1 H), 7.28 (s, 1 H)1 6.68(s, 2H), 4.10 (m, 2H), 3.86 (m, 2H), 3.18 (s, 1 H), 2.00 (m, 2H), 1.41 (s, 9H), 1.35 (m, 2H). LC-MS: 321.7 (M, 100%), 322.7 (M+1 , 20.0%).
Step 6. 4'-chloro-6', 7'-dihydrospiro[piperidine-4,5'-pyrrolo[2,3-d]pyrimidin]-2'-amine (4G) [00271] A mixture of tert-butyl 21-amino-4'-oxo-3',4',6',7'-tetrahydrospiro[piperidine-4,5'- pyrrolo[2,3-d]pyrimidine]-1-carboxylate (4F, 0.37 g, 1.2 mmol) and acetic anhydride (1.09 ml_, 11.5 mmol) was heated to reflux for 1 hour, and evaporated. To the residue were added benzyltriethylammonium chloride (0.524 g, 2.30 mmol) and phosphoryl chloride (0.644 ml_, 6.91 mmol). The mixture was heated to reflux for 40 min, and evaporated. The residue was poured into ice-water (5mL), diluted with 6N HCI solution (2OmL), and stirred at 50 0C for 1.5 hours. After cooling, the mixture was adjusted to pH 4.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white solid (120mg, purity 95.0%, yield 41.0%). LC-MS: 239.7 (M, 100%), 241.7 (M+2, 32.7%), 240.7 (M+1 , 19.2 %), 242.7 (M+3, 5.7%). Step 7. 1-(2'-amino-4'-chloro-6',7'-dihydrospiro[piperidine-4,5'-pyrrolo[2,3-dJpyrimidine]-1- yl)ethanone (4H)
[00272] A mixture of 4'-chloro-6',7'-dihydrospiro[piperidine-4,5'-pyrrolo[2,3-d]pyrimidin]-21-amine
(4G, 50.0 mg, 0.208 mmol), acetic anhydride (19.7 μL, 0.208mmol), and 4-dimethylaminopyridine (2.5mg, 0.021mmol) in anhydrous N,N-dimethylformamide (1.0 mL) was stirred at room temperature for 2 hours. The reaction was evaporated, diluted with water (20ml), adjusted to pH 4.0, and purified by reverse-phase preparative HPLC (5→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white solid (23 mg, purity 95.0%, yield 39.0%). 1HNMR (DMSO-d6): 8.53 (s, 1 H), 7.10 (s, 1 H), 3.77 (t, 2H), 3.57(s, 2H), 3.08 (t, 2H), 2.00 (s, 3H), 1.88 (t, 2H), 1.60 (t, 2H). LC-MS: 281.7 (M, 100%), 283.7 (M+2, 32.7%), 282.7 (M+1 , 19.2 %), 284.7 (M+3, 5.7%).
Step 8. 1-(2'-amino-4'-chloro-7'-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6', 7'- dihydrospiro[piperidine-4,5'-pyrrolo[2,3-d]pyrimidine]-1-yl)ethanone (XXXIII) [00273] A mixture of 1-(2'-amino-4'-chloro-6',7'-dihydrospiro[piperidine-4,5'-pyrrolo[2,3- d]pyrimidine]-1-yl)ethanone (4H, 16.4 mg, 0.0582 mmol), and 2-(chloromethyl)-4-methoxy-3,5- dimethylpyridine (10.8 mg, 0.0582 mol), NaH (2.93 mg, 0.122 mmol) in anhydrous dimethyl sulfoxide (1.0 mL) was stirred at 100 0C for 1hr. After cooling, the mixture was diluted with water (15 mL), adjusted to pH 4.0 with 6N HCI, and purified by reverse-phase preparative HPLC (5-→30% CH3CN/H2O, 0.1% TFA). The desired fractions were lyophilized to give the title compound (TFA salt) as a white solid (2.1 mg, purity 90.0%, yield 7.5%). 1HNMR (Acetone-d6): 8.43 (s, 1 H), 4.98 (s, 2H), 4.56 (t, 1 H), 3.97 (s, 3H), 3.92 (t, 1H), 3.89 (s, 2H), 3.15 (t, 1 H), 2.54 (t, 1H), 2.38 (s, 6H), 2.23-2.14 (m, 2H), 2.15 (s, 3H), 90-1.81 (m, 2H). LC-MS: 430.6 (M, 100%), 432.6 (M+2, 32.7%), 431.6 (M+1 , 19.2 %), 433.6 (M+3, 5.7%). Example 30
(2-Amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-6-yl)methanol (XXXIV) Step 1. diethyl 2-allylmaloπate
[00274] To a stirred solution of diethyl malonate (1Og, 62.43 mmol) in DMF (150 ml) was added
K2CO3 (17.3g, 124.86 mmol) and allylbromide (5.9 ml, 68.67 mmol). The resulting reaction mixture was stirred at RT for 7h. The progress of the reaction was monitored (by TLC), the reaction mixture was diluted with water (200 ml) and extracted with EtOAc (3x 100 ml). The combined organic layers was dried over Na2SO4 and concentrated under reduced pressure, the resulting crude was purified by column chromatography (10% EtOAc/ Hexane) to give diethyl 2-allylmalonate (8.1g, 64.8%). 1H-NMR [HSP-004- A8051-062, B270109A65] (CDCI3, 500 MHz): δ 5.83- 5.75 (m, 1H), 5.15 (d, J = 1.5 Hz, 1 H), 5.13 (d, J = 10 Hz, 1 H), 4.24- 4.18 (m, 4H), 3.43 (t, J = 7.5 Hz, 1H), 2.67- 2.64 (m, 2H), 1.29- 1.24 (m, 6H); Mass [HSP-004-A8051-062 (270109A)]: 201.2[MVl]; TLC system: 10% EtOAc/Hexane, Rf: 0.6. Step 2. 5-allyl-2-aminopyrimidine-4,6-diol
[00275] To a stirred solution of diethyl 2-allylmalonate (4g, 20 mmol) in MeOH (50 ml) was added
NaOMe (2.48g, 46 mmol), guanidine HCI (2.1g, 22 mmol) at 0 0C under inert condition. The reaction mixture was refluxed for 16h. After completion of the starting material (by TLC), the volatiles was evaporated under reduced pressure. The crude material was diluted with water and acidifies with 4N HCI pH ~ 2- 4. The crude solid was filtered off and washed with water. The solid was allowed to high vacuum to afford 5-allyl-2-aminopyrimidine-4,6-diol (2.1 g, 62.8%) as white solid. 1H-NMR [HSP-004-A8051-063, 29-01-2009 (AA3)] (DMSO-D6,200 MHz): δ 10.4 (bs, 1 H), 6.44 (bs, 1 H), 5.84- 5.62 (m, 1 H), 4.91- 4.74 (m, 2H), 2.85 (d, J = 6.2 Hz, 2H); LC-Mass: [HSP-004-A8031-063 (290109M07)]: 167 [MVl] at 2.42 RT (82.5% HPLC Purity); TLC system: 10% EtOAc/Hexane, Rf: 0.2. Step 3. 5-allyl-4,6-dichloropyrimidin-2-amine
[00276] To a stirred solution of 5-allyl-2-aminopyrimidine-4,6-diol (3g, 17.96 mmol) in POCI3 (30 ml) was stirred at 100 0C for 7h. After completion of the starting material (By TLC), POCI3 was removed under reduced pressure and crude material was diluted with ice cold water and basified with saturated K2CO3 solution till PH - 8 and extract with EtOAc (2x 10 ml). The combined the organic extracts were dried over Na2SO4, and evaporated. The resulting crude was purified by column chromatography (10% EtOAc/ Hexane) to afford 5-allyl-4,6-dichloropyrimidin-2-amine (2.4g, 66%) as white solid. 1H-NMR [HSP- 004- A8051 -079, 13-11-2008 (AA89)] (CDCI3, 200 MHz): δ 5.94- 5.79 (m, 1 H), 5.34 (bs, 2H), 5.14- 5.04 (m, 2H), 3.45 (d, J = 6.4 Hz, 2H); LC-Mass: [HSP-004-A8035-079 (141108M04)]: 203 [MVl], at 4.91 RT (92.11% HPLC Purity); TLC system: 10% MeOH/DCM, Rf: 0.7.
Step 4. 5-allyl-6-chloro-N4-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)pyrimidine-2,4-diamine [00277] To a stirred solution of 5-allyl-4,6-dichloropyrimidin-2-amine (1.85g, 0.0091 mol) in DMF
(10 ml) was added (4-methoxy-3,5-dimethylpyridin-2-yl)methylamine (1.66g, 0.010 mol) and K2CO3 (3.1g, 0.022 mol) at 00C. The resulting reaction mixture was heated at 70 0C for 2h and stirred at RT for 1 h. The progress of reaction monitored (By TLC). The reaction mass was diluted with water (25 ml) and extracted with EtOAc (2x 20 ml). The combined organic layers was dried over Na2SO4 and concentrated under reduced pressure, the resulting crude was purified by column chromatography (50% EtOAc/Hexane) to afford 5-allyl-6-chloro-N4-((4-methoxy-315-dimethylpyridin-2-yl)methyl)pyrimidine-2,4-diarnine (0.6g). 1H- NMR [HSP-004-A8053-010, 08-01-2009 (AA54)] (DMSO-D6 200 MHz): δ 8.21 (s,1 H), 7.06 (bs, 1 H), 6.33 (s, 2H), 5.87- 5.72 (m, 1 H), 5.16- 5.01 (m, 2H), 4.50 (d, J = 4.4 Hz, 2H), 3.72 (s, 3H), 3.28- 3.25 (m, 2H), 2.17 (s, 3H), 2.16 (s, 3H); LC-Mass: [HSP-004-A8053-010 (080109M10)]: 334 [M++1], at 3.69 RT (80.51% HPLC Purity); TLC system: 50% EtOAc/Hexane R,: 0.4.
Step 5. 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)pyrimidin-5- yl)propane-1,2-diol
[00278] To a stirred solution of 5-allyl-6-chloro-N4-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)pyrimidine-2,4-diamine (0.6g, 0.0018 mol) in DMF (3 ml) was added TFA (10 ml) was 30 % H2O2 (15 ml) at 0 0C. The resulting reaction mixture was stirred at RT for 12h, (TLC shows 50% completion), added 30 % H2O2 (10 ml) and stirred at RT for 6h (TLC shows 70% completion), added 30 % H2O2 (10 ml) and stirred at RT for 6h (TLC shows 80% completion). The reaction mass was diluted with water (25 ml) and basified with NaHCO3 solution and extracted with EtOAc (2x 15 ml). The combined organic layers was dried over Na2SO4 and concentrated under reduced pressure, the resulting crude was purified by column chromatography (10% MeOH/DCM) to afford 3-(2-amino-4-chloro-6-((4-methoxy-3,5- dimethylpyridin-2-yl)methylamino)pyrimidin-5-yl)propane-1 ,2-diol (0.4g). 1H-NMR [HSP-004-A8053-012, 15-01 -2009 (AA37)] (DMSO-D6 200 MHz): δ 8.18 (s,1 H), 7.20 (bs, 1 H), 6.27 (s, 2H), 4.91 (d, J= 4.8 Hz, 1H), 4.74- 4.69 (m, 1H), 4.51 (bs, 2H), 3.72 (s, 3H), 3.69- 3.60 (m, 2H), 2.72- 2. 54 (m, 1 H), 2.46- 2. 40 (m, 1 H) 2.20 (s, 3H), 2.18 (s, 3H); LC-Mass: [HSP-004-A8053-012 (090109M18)]: 368 [M++1], at 2.35 RT (85.70% HPLC Purity); TLC system: 10% MeOH/DCM, R,: 0.5.
Step 6. 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)pyrimidin-5-yl)-2- hydroxypropyl methanesulfonate
[00279] To a stirred solution of 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2- yl)methylamino)pyrimidin-5-yl)propane-1 ,2-diol (0.35g, 0.00095 mol) in DMF (10 ml) was added DIPEA (0.24 ml, 0.0014 mol) and Mesyl chloride (0.08 ml, 0.0010 mol) at 0 0C. The resulting reaction mixture was stirred at RT for 2h. After Completion of the starting material (By TLC), the reaction mixture was diluted with water (15 ml) and extracted with EtOAc (2x 15 ml). The combined organic layers was dried over Na2SO4 and concentrated to afford 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2- yl)methylamino)pyrimidin-5-yl)-2-hydroxypropyl methanesulfonate (0.36g). LC-Mass: [HSP-004-A8053- 014 (120109M09)]: 446 [M++1], at 2.83 RT (38.59% HPLC Purity);TLC system: 10% MeOH/DCM, Rf: 0.6. Step 7. 6-chloro-l^-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5^oxiran-2-ylmethyl)pyrimidine- 2,4-diamine
[00280] To a stirred solution of 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2- yl)methylamino)pyrimidin-5-yl)-2-hydroxypropyl methanesulfonate (0.36g, 0.0008 mol) in MeOH (3 ml) was added NaOMe (0.058g, 0.00097 mol) at 00C. The resulting reaction mixture was stirred at RT for 2h. The resulting reaction mixture was stirred at RT for 2h. After Completion of the starting material (By TLC), the reaction mass concentrated under reduced pressure, the resulting crude was purified by column chromatography (50% EtOAc/ Hexane) to afford 6-chloro-N*-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-5-(oxiran-2-ylmethyl)pyrimidine-2,4-diamine (0.08g). 1H-NMR [HSP-004-A8053-015, 15-01- 2009 (AA39)] (DMSO-D6.200 MHz): δ 8.20 (s, 1 H), 7.18- 7.16 (m, 1 H), 6.35 (bs, 2H), 4.51 (d, 4.4 Hz1 1 H), 3.72 (S1 3H), 3.13- 2.98 (m, 1 H), 2.83- 2.81 (m, 1H)1 2.78- 2. 71 (m, 2H)1 2.61- 2.50 (m 1 H), 2.19 (s, 3H), 2.18 (S1 3H); LC-Mass: [HSP-004-A8053-015 (150109M06)]: 350 [M++1] at 3.13 RT (98.3% HPLC Purity); TLC system: 10% MeOH/DCM, Rf: 0.5.
Step 8. (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-SH- pyrrolo[2,3-d]pyrimidin-6-yl)methanol (XXXIV)
[00281] To a stirred solution of 6-chloro-bf-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5-
(oxiran-2-ylmethyl)pyrimidine-2,4-diamine (20 mg) in DMF (3 ml) at 100 0C for 4h. After completion of the starting material (By TLC)1 the reaction mixture was cooled RT and diluted with water (5 ml) and extracted with EtOAc (2x 5 ml). The combined organic layers was dried over Na2SO4 and concentrated under reduced pressure, the resulting crude was purified by preparative TLC (50% EtOAc/Hexane) to afford (2- amino-4-chloro-7-((4-methoxy-315-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-6- yl)methanol (3 mg). 1 H-NMR [HSP-004-A8053-021 , B290109A11] (CDCI3, 500 MHz): δ 8.05 (s,1 H), 5.76 (d, J = 16.5 Hz, 1 H), 4.53 (bs, 2H), 4.30 (s, 1 H), 3.93 (d, J = 16.5 Hz1 1 H)1 3.78 (s, 3H), 3.67 (d, J= 12 Hz, 1H), 3.53-3.50 (m, 1 H)1 3.12-3.00 (m, 1H)1 2.67- 2.63 (m, 1 H), 2.23 (s, 3H)1 2.21 (s, 3H); Mass: [HSP- 004-A8053-021 (01272009)]: 350.4 [M++1]; HPLC Purity: [HSP-004-A8053-015 (01272009)]: 3.27 RT (81.65%); TLC system: 10% MeOH/DCM, R1: 0.5. Example 31
(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyπdin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (XXXV)
Step 1. (4-bromo-3,5-dimethylpyridin-2-yl)methyl acetate
[00282] A solution of (3,5-dimethyl-4-nitropyridin-2-yl)methanol (5.00 g, 27.4 mmol) in Acetyl
Bromide (20.3 mL, 274 mmol) was stirred at reflux for 2 hours. After cooling to ambient temperature the mixture was poured onto crushed ice and stirred vigorously. The solution was brought to pH 10 with careful addition of saturated sodium carbonate. The resulting mixture was extracted 3 times with ethyl acetate and the combined organic layers washed with brine. Purification was performed by silica gel chromatography, eluting with ethyl acetate/hexane to give the title product as a clear oil (5.822 g, yield
Step 2. (3,5-dimethyl-4-vinylpyridin-2-yl)methyl acetate
[00283] A solution of (4-bromo-3,5-dimethylpyridin-2-yl)methyl acetate (50.0 mg, 0.194 mmol) in
N.N-Dimethylformamide (0.9 mL) was treated with (2-Ethenyl)tri-n-butyltin (113 μL, 0.387 mmol) and Tetrakis(triphenylphosphine)palladium(0) (13 mg, 0.012 mmol) in microwave vial. The mixture was microwaved at 1000C for 30 minutes, after which water was added. The metal solids were filtered off and rinsed with ethyl acetate. The resulting filtrate was extracted 3 times with ethyl ether, which was washed twice with aqueous potassium fluoride. Purification by silica gel chromatography using ethyl acetate/hexane gave the title compound (22 mg, yield 75%).
Step 3. (3,5-dimethyl-4-vinylpyridin-2-yl)methanol
[00284] A solution of (3,5-dimethyl-4-vinylpyridin-2-yl)methyl acetate (1.347 g, 6.562 mmol) in
Methanol (20.0 mL), Tetrahydrofuran (20.0 ml_) was treated with 2.00 M Potassium carbonate in
Water(9.842 mL, 19.68 mmol). After 1 hour the reaction was diluted with water and extracted 3 times with ethyl acetate. The combined organic layers were washed with brine and saturated sodium bicarbonate, then dried over magnesium sulfate. The crude material was carried on without further purification.
Step 4. (4-ethyl-3,5-dimethylpyridin-2-yl)methanol
[00285] A solution of (3,5-dimethyl-4-vinylpyridin-2-yl)methanol (1.378 g, 8.443 mmol) in
Methanol (50 mL) in a bomb was treated with Platinum dioxide (958.6 mg, 4.221 mmol) and hydrogenated at 2 atm for 30 minutes. The bomb was then recharged from 0.75 to 2 atm and hydrogenated for another 45 minutes. The catalyst was carefully filtered off using glass filter paper and rinsed with ethanol. The filtrate was then concentrated to an oil which crystallized on standing to give the title product (1 g, yield 72%).
Step 5. 2-(chloromethyl)~4-ethyl-3,5-dimethylpyridine
[00286] A solution of (4-ethyl-3,5-dimethylpyridin-2-yl)methanol (476.0 mg, 2.881 mmol) in
Methylene chloride (35 mL) was treated with Thionyl chloride (1.051 mL, 14.40 mmol) and stirred for 2 hours. The mixture was then condensed to a solid and taken up in ethyl acetate and saturated sodium bicarbonate. Following 3 extractions with ethyl acetate, the crude mixture was purified by silica gel chromatography using ethyl acetate/hexane to give the title compound.
Step 6. 4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2- amine
[00287] A suspension of 2-(chloromethyl)-4-ethyl-3,5-dimethylpyridine (130 mg, 0.71 mmol), 4- chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (210 mg, 0.7 mmol) and K2CO3 in anhydrous DMF
(5mL) was stirred for 2 h at it Work-up (EtOAc/water) and silica gel column chromatography (20%
EtOAc/Hexane) gave the title compound (290 mg, purity >90%, yield 92%). 1H NMR (DMSOd6): δ ppm
8.20 (s, 1H), 7.65 (S, 1 H), 5.39 (s, 2H), 5.00 (s, 2H), 3.84 (s, 3H), 2.68 (q, 2H), 2.31 (s, 3H), 2.26 (s, 3H),
1.11 (t, 3H), LC/MS: rt (5-100-7 method) = 5.713min.; 441.3 (M+1 , 100 %), 443.4 (M+3, 35%).
Step 7. Ethyl 2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-7H-pyrrolo[2,3- d]pyrimidine-5-carboxylate
[00288] In an autoclave vessel was added (60mg, 0.1 mmol), [(R)-(+)-2,2'- bis(diphenylphosphino)-1 ,1'-binaphthyl]palladium(ll) chloride (10 mg, 0.012 mmol, Aldrich# 342335), and anhydrous methanol (10 mL,200 mmol). The reaction mixture was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 80°C while stirring for 3 hours. The reaction was cooled to room temperature, diluted with water, and extracted with a mixture of dichloromethane: isopropanol
(95:5). The solid product was purified by reverse phase HPLC (19 mg, purity >90%, yield 38%). 1H NMR (CDCI3): δ ppm 8.05 (s, 1 H) 7.87 (s, 1 H), 6.79 (s, 2H), 5.32 (s, 2H), 3.75 (s, 3H), 3.73 (s, 3H), 2.28 (s, 3H), 2.16 (s, 3H), 1.11 (t, 3H), LS-MS: rt (5-100-7 method) = 5.086 min.; 373.6 (M+1 , 100 %), 375.6(M+3,
Step 8. (2-amino-4-chloro-7-((4-ethyl-3,5-dimethytpyridin-2-yl)methyl)-6,7-dihydro-SH-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (XXXV)
[00289] To a suspension of methyl 2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate (15mg, 0.40 mmol) in anhydrous THF (10 ml.) was added 1.0 M of L-selectride solution (1 ml_, 1mmol) dropwise via syringe at 0 0C. The reaction mixture was warmed to room temperature, allowed to stir for 1 h, then carefully quenched with ice water before being adjusted to pH 7.0 using 2N HCI solution. The organic layer was concentrated under reduced pressure. The solid product was purified by reverse phase HPLC (6.95 mg, purity 95%, yield 49.6%). 1H NMR (CDCI3): δ ppm 8.11 (s, 1H), 5.07 (d, 1 H), 4.8 (s, 2H), 4.34 (d, 1 H), 3.91 (m, 1H), 3.80 (dd, 2H), 3.60 (m, 1 H), 3.4Q (m, 1 H), 2.26 (d, 6H) 1.12 (t, 3H). LC/MS: rt (5-100-7 method) = 3.949 min.; 348.6 (M+1 , 100 %), 349.6(M+3,
Example 32
(2-amino-4-chloro-7-((5-methoxy-4,6-dimethylpyridin-3-yl)niethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidiπ-5-yl)methanol (XXXVI)
Step 1. 4-Chloro-5-iodo-7-((5-methoxy-4, 6-dimethylpyridin-3-yl)methyl)-7H-pyrrolo[2,3- d]pyrimidin-2-amine
[00290] A suspension of 5-(chloromethyl)-3-methoxy-2,4-dimethylpyridine (100 mg, 0.54 mmol),
4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (110 mg, 0.37 mmol) and K2CO3 (IOI mg, 0.73 mmol) in anhydrous DMF (3 mL) was stirred for for 2 h at 700C. Work-up (EtOAc/water) gave a beige solid (110 mg, purity >90%, yield 60%). 1H NMR (CDCI3): δ ppm 8.13 (s, 1 H), 6.84 (s, 1 H), 5.19 (s, 2H), 5.01 (s,
2H), 3.74 (s, 3H), 2.54 (s, 3H), 2.17 (s, 3H), LC/MS: rt (5-100-7 method) = 5.532min.; 443.4(M+1 , 100 %),
445.4 (M+3, 35%).
Step 2. Methyl 2-amino-4-chlorth7-((5-methoxy-4, 6-dimethylpyridin-3-yl)methyl)-7H-pyrrolo[2,3- d]pyrimidine-5-carboxylate
[00291] In an autoclave vessel was added 4-chloro-5-iodo-7-((5-methoxy-4,6-dimethylpyridin-3- yl)methyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine (100mg, 0.2 mmol), [(R)-(+)-2,2'-bis(diphenylphosphino)-
1 ,1'-binaphthyl]palladium(ll) chloride (10 mg, 0.012 mmol, Aldrich# 342335), and anhydrous methanol (10 mL,200 mmol). The autoclave vessel was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 500C while stirring for 16 hours. The reaction was cooled to room temperature and diluted with water, before being extracted with a mixture of dichloromethane: isopropanol (95:5). The solid product was purified by silica gel column chromatography (52 mg, purity
80.0%, yield 50%). 1H NMR (CDCI3): δ ppm 8.13 (s, 1 H) 7.38 (s, 1 H), 5.19 (s, 2H), 5.00 (s, 2H), 3.79 (s,
3H), 3.68 (s, 3H), 2.50 (s, 3H), 2.10(s, 3H)1 LS-MS: rt (5-100-7 method) = 4.818min.; 375.6 (M+1 , 100 %),
377.6(M+3, 35%). Step 3. (2-amino-4-chloro-7-((5-methoxy-4, 6-dimethylpyridin-3-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XXXVI)
[00292] A suspension of methyl 2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-7H- pyrrolo[2,3-d]pyrimidine-5-carboxylate (40 mg, 0.1 mmol) in anhydrous tetrahydrofuran (10 mL) was cooled at 0C and treated dropwise with 1.0 M of L-selectride solution (0.7 mL, 0.7 mmol). The reaction mixture was warmed to room temperature, then allowed to stir for 1 h before being carefully quenched with ice water and adjusted to pH 7.0 with 2N HCI solution. The solid product was purified by silica gel column chromatography (5%MeOH/EtOAc),(14 mg, purity 90%, yield 30%). 1H NMR (CDCI3): δ ppm 8.08
(s, 1 H), 4.98 (s, 2H), 4.63 (d, 1 H), 4.45 (d, 1 H), 3.80 (d, 2H), 3.73 (s, 3H), 3.50 (d, 3H), 3.40 (m, 1 H), 2.51
(s, 3H), 2.23 (s, 3H)). LC/MS: rt (5-100-7 method) = 3.725 min.; 349.6 (M+1 , 100 %), 351.6 (M+3, 35%).
Example 33
(4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine
(XXXVII)
Step 1. 4-bromo-2,3-dimethylpyridine
[00293] 2,3-dimethylpyridin-4-ol, ( 2.0 g, 20 mmol) and phosphorus tribromide oxide (14.0 g, 49 mmol) were left stirring at 1300C for 4 hours under nitrogen. The reaction mixture was poured onto ice and made basic by adding aqueous NaOH. The mixture was then extracted with diethyl ether and water.
The brown needle solid was obtained after ether was gently removed (2.03g, purity 90%, yield 60%). 1H
NMR (CDCI3): δ ppm 8.10 (d, 1 H), 7.34 (d, 1 H), 2.58 (s, 3H), 2.39 (s, 3H). LC/MS: rt (5-100-7 method) =
3.555 min.; 185.7 (M+1 , 100 %) 187.7 (M+3, 97%).
Step 2. 4-ethyl-2,3-dimethylpyridine
[00294] 4-bromo-2,3-dimethylpyridine (2.03 g, 10.9 mmol) and tetrakis(triphenylphosphine)palladium (0) (0.4 g, 0.3 mmol) were suspended in THF (20 mL). The reaction mixture was degassed and placed on an ice bath before slowly adding diethyl zinc (2.05g, 16.6mmol).
After the diethyl zinc addition was completed, the reaction was brought to 50 °C for 1 hour. The reaction mixture was poured onto ice and made basic by adding aq. NaOH. The mixture was then extracted with diethyl ether and water. Evaporation of the organic layer gave the desired product as a viscous yellow oil
(0.98g, purity 80%, yield 66%). 1H NMR (CDCI3): δ ppm 8.21 (d, 1 H), 6.91 (d, 1 H), 2.64 (q, 2H), 2.51 (s,
3H), 2.21 (s, 3H), 1.21 ( t, 3H). LC/MS: rt (5-100-7 method) = 3.897 min.; 135.9 (M+1 , 100 %).
Step 3. 2-(bromomethyl)-4-ethyl-3-methylpyridine
[00295] The title compound was obtained from 4-ethyl-2,3-dimethylpyridine by following the 4 step sequence (1. m-CPBA, 2. Ac2O, 3. OH", 4. PPh3ZCBr4) described in Kasibhatla S. R. et al. J. Med.
Chem. 2007, 50(12), 2767-2778. 1H NMR (CDCI3): δ ppm 8.32 (d, 1 H), 7.08 (d, 1 H), 4.64 (s, 2H), 2.64 (q,
2H), 2.36 (s, 3H)1 1.24 (t, 3H). LC/MS: rt (5-100-7 method) = 4.385 min.; 213.7 (M+1 , 100 %) 215.7 (M+3,
Step 4. 4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-5-iodo-7H-pyrrolo[2,3-d]pyrimldin-2- amine [00296] A suspension of 2-(bromomethyl)-4-ethyl-3-methylpyridine (0.71 g, 3.3 mmol), 4-chloro-5- iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (770 mg, 0.2.6 mmol) and K2CO3 (605 mg, 4.38 mmol) in anhydrous DMF (3 ml_) was stirred for 2 h at 70 °C. Work-up (EtOAc/water) gave the title compound (0.82 g, purity 85.0%, yield 62%). 1H NMR (DMSOd6): δ ppm 8.11 (d, 1 H), 7.24 (s, 1 H), 7.09 (d, 1 H), 6.70 (s, 2H), 5.33(s, 2H), 2.63 (q, 2H), 2.27 (s, 3H), 1.13 (t, 3H), LC/MS: rt (5-100-7 method) = 5.782min.; 427.5(M+1 , 100 %), 429.5 (M+3, 35%).
Step 5. Methyl 2-amino-4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-7H-pyrrolo[2,3- d]pyrimidine-5-carboxylate
[00297] In an autoclave vessel was added 4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-5- iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (200mg, 0.5 mmol), [(R)-(+)-2,2'-bis(diphenylphosphino)-1 ,1 '- binaphthyl]palladium(ll) chloride (10 mg, 0.012 mmol, Aldrich# 342335), and anhydrous methanol (10 ml_,200 mmol). The autoclave vessel was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 500C while stirring for 16 hours. The solvent was evaporated. The white solid product was purified by silica gel column chromatography (75% EtOAc/Hexane) (112 mg, purity >90%, yield 60%). 1H NMR (CDCI3): δ ppm 8.35 (d, 1 H), 7.68 (s, 1 H), 7.11 (d, 1 H), 5.43 (s, 2H), 5.01 (s, 2H), 3.85(s, 3H), 2.65 (t, 2H), 2.27 (s, 3H), 1.23 (t, 3H), LS-MS: rt (5-100-7 method) = 4.998min.; 359.6 (M+1 , 100 %), 361.6(M+3, 35%).
Step 6. (2-amino-4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (XXXVII)
[00298] To a suspension of methyl 2-amino-4-chloro-7-((4-ethyl-3-methylpyridin-2-yl)methyl)-7H- pyrrolo[2,3-d]pyrimidine-5-carboxylate (90 mg, 0.2 mmol) in anhydrous tetrahydrofuran (10 mL) was added 1.0 M of L-selectride solution (1.9 mL, 8.9 mmol) dropwise via syringe at 0 0C. The reaction mixture was warmed to room temperature, then allowed to stir for 1 hr before being carefully quenched with ice water and adjusted to pH 7.0 with 2N HCI solution. The solid product was purified by silica gel chromatography (5%MeOH/EtOAc),(52 mg, purity 90%, yield 60%). 1H NMR (CDCI3): δ ppm 8.27 (d, 1 H), 7.07 (d, 1H), 5.13 (d, 1 H), 4.76 (s, 2H), 4.38 (d, 1 H), 4.14 (m, 1 H), 3.96 (m, 1H), 3.82 (q, 2H), 3.63 (m, 1H), 3.43 (m, 1 H),2.70 (q, 2H), 2.27 (s, 3H), 1.24 (t, 3H). LC/MS: rt ((5-100-7 method) = 3.794 min., 333.7(M+1 , 100 %), 335.7 (M+3, 35%). Example 34
(2-Amino-4-chloro-7-((5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XXXVIII) Step 1. 4-Methoxy-2,3-dimethyl-pyridine 1 -oxide
[00299] To a mixture of 2,3-Dimethyl-4-nitro-pyridine 1 -oxide (50.0 g, 0.297 mol) and methanol
(500 mL) was added in portions Potassium carbonate (50.0 g, 0.362 mol) at 0 °C. After the addition, the mixture was heated to reflux for 2 hours, evaporated, taken up in CH3CN (1.5 L), refluxed for 30 minutes and filtered. The solid was washed with acetonitrile (10OmL X 3) and the combined filtrates were evaporated to yield the desired product as a yellow solid (24.0 g, Yield: 47%) 1H NMR (DMSO-d6): δ ppm
8.11 (s, 1H), 6.95 (s, 1 H), 3.84 (s, 3H), 2.35 (s, 3H), 2.12(s, 3H).
Step 2. 5-bromo-4-Methoxy-2,3-dimethyl-pyridine 1 -oxide
[00300] A mixture of 4-methoxy-2,3-dimethyl-pyridine 1 -oxide (306 mg, 0.00200 mol), trifluoroacetic Acid (0.42 ml_, 0.0054 mol) and Sulfuric acid (0.56 ml_, 0.010 mol) was treated with N- bromosuccinimide (712 mg, 0.00400 mol) portionwise, and the resulting reaction mixture was stirred for
16 hours at room temperature. The reaction was quenched with ice-water (1OmL), neutralized with 6N
NaOH to pH 7.0, and extracted with dichloromethane (100 mL X 2). The combined organic layers were then evaporated to give the title compound. Yield : 48% 1 H NMR (DMSOd6): 8.48 (s, 1 H), 3.75 (s, 3H),
2.31 (s, 3H), 2.22(s, 3H).
Step 3. (5-bromo-4-methoxy-3-methylpyridin-2-yl)methanol
[00301] The title compound was obtained from 5-bromo-4-methoxy-2,3-dimethyl-pyridine 1 -oxide by following the 3 step sequence (1. m-CPBA, 2. Ac2O, 3. OH") described in Kasibhatla S. R. et al. J.
Med. Chem. 2007, 50(12), 2767-2778. 1H NMR (CDCI3): δ ppm 8.51 (s, 1 H), 4.64 (d, 2H), 4.45 (t, 1 H),
3.90 (s, 3H), 2.19 ( S, 3H).
Step 4. (4-methoxy-3-methyl-5-(prop- 1 -en-2-yl)pyridin-2-yl)methanol
[00302] (5-bromo-4-methoxy-3-methylpyridin-2-yl)methanol (1g, 4 mmol), isopropenylboronic acid pinacol ester (1.2 g, 7.1 mmol), tetrakis(triphenylphosphine) palladium(O) (300 mg, 0.2 mmol), and potassium carbonate (2M, 8 mL, 20 mmol) were suspended in DMF (30 mL). The mixture was degassed and left stirring under nitrogen gas in a screw top glass vessel at 80 0C for 2 hours. The reaction mixture was extracted with EtOAc and water. The organic solvent was removed to yield the product as a viscous oil (400 mg, purity 90%, yield 50%). 1H NMR (CDCI3): δ ppm 8.24 (s, 1 H),5.26 (d, 2H), 4.77 (b, s 1 H),
4.67 (s, 2H), 3.79 (s, 3H), 2.15 ( d, 3H), 2.13 (s, 3H), LS-MS: rt (5-100-7 method) = 4.227min.; 193.9
(M+1 , 100 %).
Step 5. (5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methanol
[00303] To (4-methoxy-3-methyl-5-(prop-1-en-2-yl)pyridin-2-yl)methanol (400mg, 2 mmol) in ethanol (40 mL) was added 10% Pd/C (40 mg). The mixture was degassed before being placed under hydrogen gas (atmospheric pressure) via a balloon for 2 hours. The Pd/C particulates were removed by filtration, and the filtrate was evaporated to give a white solid residue. (402 mg, purity 90%, yield 100%).
1H NMR (CDCI3): δ ppm 8.33 (s, 1 H), 4.77 (b, s 1H), 4.66 (s, 2H), 3.80 (s, 3H), 3.25 (m, 1 H), 2.15 (s, 3H),
1.31 (d, 6H).
Step 6. 2-{bromomethyl)-5-isopropyl-4-methoxy-3-methylpyridine
[00304] (5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methanol (402 mg, 2.06 mmol) was dissolved in dichloromethane (2OmL) before adding triphenylphosphine (725 mg, 2.76 mmol) and carbon tetrabromide (910 mg, 2.7 mmol). The reaction mixture was stirred at room temperature for 1 hour. Silica gel chromatography gave a white-purple solid (295 mg, purity 90%, yield 50%). 1H NMR (CDCI3): δ ppm
8.33 (s, 1H), 4.60 (s 2H), 3.80 (s, 3H), 3.28 (m, 1H), 2.36 (s, 3H), 1.29 (d, 6H). LS-MS: rt (5-100-7 method) = 5.135 min.; 257.7 (M+1 , 100 %), 259.7 (M+1 , 99%).
Step 7. 4-chloro-5-iodo-7-((5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methyl)-7H-pyrrolo[2,3- d]pyrimidin-2-amine
[00305] 2-(bromomethyl)-5-isopropyl-4-methoxy-3-methylpyridine (210 mg, 0.82 mmol), 4-chloro-
5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (220 mg, 0.75 mmol) and K2CO3 (605 mg, 4.38 mmol) were suspended in anhydrous DMF (1OmL) for 2 hours at 70 0C. Water was added to precipitate the crude product. (224 mg, purity >90%, yield 57%). 1H NMR (DMSOd6): δ ppm 8.19 (s, 1 H), 7.31 (s, 1H), 7.09 (s,
1H), 6.73 (s, 2H), 5.30 (s, 2H), 3.73 (s, 3H), 3.15 (m, 1H), 2.27 (s, 3H), 1.19 (d, 6H), LC/MS: rt (5-100-7 method) = 6.206 min.; 471.4 (M+1 , 100 %), 473.4 (M+3, 35%).
Step 8. Methyl 2-amino-4-chloro-7-((5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methyl)-7H- pyrrolσ[2,3-d]pyrimidine-5-carboxylate
[00306] In an autoclave vessel was added 4-chloro-5-iodo-7-((5-isopropyl-4-methoxy-3- methylpyridin-2-yl)methyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine (100 mg, 0.2 mmol), [(R)-(+)-2,2'- bis(diphenylphosphino)-1 ,1'-binaphthyl]palladium(ll) chloride (10 mg, 0.012 mmol, Aldrich# 342335), and anhydrous methanol (5 mL,100 mmol). The autoclave vessel was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 500C while stirring for 16 hours. The solvent was evaporated. The white solid product was purified by preparative TLC (32 mg, purity >90%, yield 30%). 1H
NMR (CDCI3): δ ppm 8.30 (s, 1 H), 7.71 (s, 1 H), 5.34(s, 2H), 5.05(s, 2H), 3.86 (s, 3H), 3.74(s, 3H), 3.22
(m, 1H), 2.25 (s, 3H), 1.27 (d, 6H), LC/MS: rt (5-100-7 method) = 5.856 min.; 403.6(M+1 , 100 %),
405.6(M+3, 100 %).
Step 9. (2-Amino-4-chloro-7-((5-isopropyl-4-methoxy-3-methylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XXXVIII)
[00307] To a suspension of methyl 2-amino-4-chloro-7-((5-isopropyl-4-methoxy-3-methylpyridin-
2-yl)methyl)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate (30 mg, 0.2 mmol) in anhydrous tetrahydrofuran
(10 mL) was added 1.0 M of L-selectride solution (0.5mL, 0.5 mmol) dropwise via syringe at 0 0C. The reaction mixture was warmed to room temperature, then allowed to stir for 1 hour before being carefully quenched with ice water and adjusted to pH 7.0 with a 2N HCI solution. Preparative TLC (100% EtOAc) gave the title compound as a solid (17 mg, purity 90%, yield 50%). 1H NMR (CDCI3): δ ppm 8.24(s, 1 H),
5.05 (d, 1H), 4.90 (s, 2H), 4.33(d, 1 H), 3.98 (dd, 1 H), 3.83 (m, 2H), 3.77 (s, 3H), 3.41 (m, 1 H), 3.21 (m,
1H), 2.24 (s, 3H)), 1.27 (d, 6H), LC/MS: rt (5-100-7 method) = 4.347 min.; 377.6 (M+1 , 100 %), 379.6
(M+3, 100 %).
Example 35
2-amino-4-chloro-7-((5-ethyl-4-methoxy-3-methylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (XXXIX)
Step 1. (4-methoxy-3-methyl-5-vinylpyridin-2-yl)methanol
[00308] (5-bromo-4-methoxy-3-methylpyridin-2-yl)methanol (0.5g, 2 mmol), dibutyl vinylboronate
(0.70 g, 4 mmol), tetrakis(triphenylphosphine) palladium(O) (150 mg, 0.1 mmol), potassium carbonate (2M, 3ml, 6 mmol) were suspended in DMF (10 ml_). The mixture was degassed and left stirring under nitrogen gas in a screw top glass vessel at 8OC for 2 hours. The reaction mixture was extracted with EtOAc and water. The organic solvent was removed to yield the product as an off white solid (354 mg, purity 85%, yield 80%). 1H NMR (CDCI3): δ ppm 8.51 (s, 1H),6.90 (q, 1 H), 5.89 (d, 1 H), 5.44 (d, 1 H), 4.67 (s, 2H), 3.79 (s, 3H), 3.68 (b, s, 1 H)1 2.13 (s, 3H), LS-MS: rt (5-100-7 method) = 3.995min.; 179.9 (M+1 , 100 %). Step 2. (5-ethyl-4-methoxy-3-methylpyridin-2-yl)methanol
[00309] To (4-methoxy-3-methyl-5-(prop-1-en-2-yl)pyridin-2-yl)methanol (400 mg, 2 mmol) in ethanol (40 mL) was added 10% Pd/C (40 mg). The mixture was degassed before being placed under hydrogen gas (atmospheric pressure) via a balloon for 2 hours. The Pd/C particulates were removed by filtration, and the solvent was removed to give a white solid residue. (400 mg, purity 90%, yield 100%). 1H NMR (CDCI3): δ ppm 8.25(s, 1 H), 4.82 (b, s 1 H), 4.65 (s, 2H), 3.80 (s, 3H), 2.71 (q, 1H), 2.14 (s, 3H), 1.28 (t, 3H), LS-MS: rt (5-100-7 method) = 3.888min.; 181.9 (M+1, 100 %). Step 3. 2-(bromomethyl)-5-ethyl-4-methoxy-3-methylpyridine
[00310] (5-ethyl-4-methoxy-3-methylpyridin-2-yl)methanol (400 mg, 2.0 mmol) was dissolved in dichloromethane (2OmL) before adding triphenylphosphine (800 mg, 3.0 mmol) and carbon tetrabromide (1400 mg, 4.2 mmol). The reaction mixture was left stirring at room temperature for 1 hour. The solvent was evaporated, and silica gel chromatography (15% EtOAc/Hexane) gave a white purple solid (290 mg, purity 90%, yield 50%). 1H NMR (CDCI3): δ ppm 8.27(s, 1 H), 4.60 (s, 2H), 3.80 (s, 3H), 2.71 (q, 1 H), 2.35 (s, 3H), 1.28 (t, 3H), LS-MS: rt (5-100-7 method) = 4.897min.; 245.7 (M+1 , 100 %), 247.7 (M+1 , 100 %). Step 4. 4-chloro-7-((5-ethyl-4-methoxy-3-methylpyridin-2-yl)methyl)-5-iodo-7H-pyrrolo[2,3- d]pyrimidin-2-amine
[00311] 2-(bromomethyl)-5-ethyl-4-methoxy-3-methylpyridine (200 g, 0.82 mmol), 4-chloro-5- iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (220 mg, 0.75 mmol) and K2CO3(200 mg, 1 mmol) were suspended in anhydrous DMF (10 mL) for 2 hours at 70 0C. Water was added to precipitate the crude product. (224 mg, purity >90%, yield 57%). 1H NMR (DMSO-d6): δ ppm 8.19 (s, 1 H), 7.28 (s, 1 H), 6.73 (s, 2H), 5.30(s, 2H), 3.73 (s, 3H), 2.57(m, 2H), 2.27 (s, 3H), 1.15 (t, 3H), LC/MS: rt (5-100-7 method) = 6.073 min.; 457.4 (M+1 , 100 %), 459.4 (M+3, 35%).
Step 5. Methyl 2-amino-4-chloro-7-((5-ethyl-4-methoxy-3-methylpyridin-2-yl)methyl)-7H- pyrrolo[2,3-d]pyrimidine-5-carboxylate
[00312] In an autoclave vessel was added 4-chloro-7-((5-ethyl-4-methoxy-3-methylpyridin-2- yl)methyl)-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (100 mg, 0.2 mmol), [(R)-(+)-2,2'- bis(diphenylphosphino)-1 ,1'-binaphthyl]palladium(ll) chloride (10 mg, 0.012 mmol, Aldrich# 342335), and anhydrous methanol (5 mL,100 mmol). The autoclave vessel was degassed and was pressurized to 100 psi with carbon monoxide. It was then heated to 500C while stirring for 16 hours. The solvent was evaporated, and the white solid product was purified by preparative TLC (50 mg, purity >90%, yield 63%). 1H NMR (CDCI3): δ ppm 8.26 (s, 1 H), 7.72 (s, 1 H), 5.37(s, 2H), 5.02(s, 2H), 3.86 (s, 3H)1 3.77(s, 3H), 2.68 (q, 2H), 2.27 (s, 3H), 1.27 (s, 3H), LC/MS: rt (5-100-7 method) = 5.300 min.; 389.6 (M+1 , 100 %), 391.6
(M+3, 100 %).
Step 6. 2-amino-4-chloro-7-((5-ethyl-4-methoxy-3-methylpyridin-2'yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XXXIX)
[00313] To a suspension of methyl 2-amino-4-chloro-7-((5-ethyl-4-methoxy-3-methylpyridin-2- yl)methyl)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate (50 mg, 0.1 mmol) in anhydrous tetrahydrofuran (10 ml_) was added 1.0 M of L-selectride solution (1 mL, 1 mmol) dropwise via syringe at 0 °C. The reaction mixture was warmed to room temperature, then allowed to stir for 1hr before being carefully quenched with ice water and adjusted to pH 7.0 with 2N HCI solution. Preparative TLC (100% EtOAc) gave the title compound as a solid product (25 mg, purity 90%, yield 50%). 1H NMR (CDCI3): δ ppm 8.16(s, 1 H), 4.98
(d, 1 H), 4.94 (s, 2H), 4.34(d, 1 H), 3.92 (dd, 1 H), 3.81 (m, 2H), 3.77 (s, 3H), 3.63 (m, 1 H), 3.40 (m, 1 H),
2.65 (q, 2H), 2.22 (s, 3H)), 1.23 (t, 3H), LC/MS: rt (5-100-7 method) = 4.251 min.; 363.6 (M+1 , 100 %),
365.6 (M+3, 100 %).
Example 36
(2-amino-4-chloro-7-ethyl-6, 7-dihydro-5H-pyrrolo[2, 3-d]pyrimidin-5-yl)methanol (XL)
Step 1. 4-chloro-7-ethyl-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine
[00314] A solution of 4-Chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-ylamine (1.00 g, 3.40 mmol) and N,N-Dimethylformamide (50 mL) was stirred in an ice bath under nitrogen, lodoethane (271.6 μL,
3.396 mmol) was added and the reaction stirred overnight at room temperature. The crude product was crashed out from solution upon addition of water. The crude solid was filtered, rinsed with water, and dried overnight to provide the title compound which was carried on without further purification (850 mg, yield 78%).
Step 2. Methyl 2-amino-4-chloro-7-ethyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate
[00315] A solution of 4-chloro-7-ethyl-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (850.0 mg, 2.635 mmol) in N,N-Dimethylformamide (20 mL) and methanol (53.4 mL) was treated with triethylamine (0.742 mL, 5.32 mmol). The reaction was degassed with nitrogen then treated with [(R)-(+)-2,2'-
Bis(diphenylphosphino)-1 ,1 '-binaphthyl]palladium(ll) chloride (105.4 mg, 0.1318 mmol). The solution was then refluxed overnight under 1 atm of carbon monoxide at 45° C. The reaction was cooled to room temperature before adding water to crash out a purple solid. The solid was filtered off and the filtrate extracted 4 times with dichloromethane. Purification was performed by silica gel chromatography in dichloromethane/methanol to provide the title compound as a white solid (455 mg, yield 68%).
Step 3. (2-amino-4-chloro-7-ethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XL)
[00316] A solution of methyl 2-amino-4-chloro-7-ethyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate
(200.0 mg, 0.7853 mmol) in tetrahydrofuran (13 mL) was stirred in a flame-dried flask under nitrogen at
00C. A 4.0 M solution of L-Selectride in THF (1.174 mL, 5.497 mmol) was added via syringe. The reaction was removed from the ice bath after five minutes of stirring. Upon completion of the reaction Rochelle's salt was added slowly to the vigorously stirred solution. Saturated sodium bicarbonate was added and the mixture was extracted 3 times with dichloromethane. The combined organic phases were dried over MgSO4. Purification was performed by silica gel chromatography in dichloromethane/methanol to provide the title compound as a white solid (50 mg, yield 27.6%). LC-MS (5-100-5 method) : Rt = 1.527 min; M+1 = 229.0; 1H NMR (400 MHz, DMSO-cfe) δ ppm 6.37 (s, 2 H)1 4.77 - 4.95 (m, 1 H), 3.61 - 3.71 (m, 1 H), 3.52 - 3.60 (m, 1 H), 3.45 - 3.52 (m, 1 H), 3.33 - 3.39 (m, 1 H), 3.15 - 3.31 (m, 3 H), 1.06 (t, J= 7.2 Hz, 3 H).
Example 37
(2-amino-4-chloro-7-(4-methoxybenzyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XU) Step 1. 4-chloro-5-iodo-7-(4-methoxybenzyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine [00317] To a stirred solution of 4-chloro-5-iodo-7H-pyrrolo[2,3-d]pyrimidin-2-amine (25 g, 0.085 mol) in DMF (500 mL) was added K2CO3 (23.5 g, 0.17 mol) and PMB-CI (13.8 mL, 0.10 mol) at 0 0C. The resulting reaction mixture was stirred at RT for 16h. The reaction mixture was diluted with cold water (1500 mL) and extracted with EtOAc (3 x 500 mL). The combined organic layers were washed with ice cold water and dried over Na2SO4. The solvent was evaporated under reduced pressure and the crude material was purified by precipitation from 20% dichloromethane/Hexane (100 mL) to afford the title compound (25.2 g, 71%) as a solid. 1HNMR (500 MHz, DMSO-cfe)] δ 7.40 (s, 1 H), 7.22 (d, J = 9.0 Hz, 2H), 6.88 (d, J = 8.5 Hz, 2H), 6.82 (br s, 2H), 5.13 (s, 2H), 3.71 (s, 3H). HPLC (10-90[3]-10 method): Rt = 2.69 min. (89%).
Step 2. methyl 2-amino-4-chloro-7-(4-methoxybenzyl)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxylate [00318] The mixture of 4-chloro-5-iodo-7-(4-methoxybenzyl)-7H-pyrrolo[2,3-d]pyrimidin-2-amine
(15 g, 0.036 mol), Pd(PPh3J4 (418.6 mg, 0.0003 mol), racemic BINAP (225 mg, 0.0003 mol) and Et3N (10.1 mL, 0.07 mol) in MeOH (300 mL) was taken in an autoclave. The resulting reaction mixture was stirred at 800C for 2h under CO atmosphere at 100 psi. After the consumption of the starting material (By TLC), the reaction mixture was cooled to RT and CO gas was removed carefully. The reaction mixture was diluted with water (300 mL) and extracted with CH2CI2 (3 x 200 mL). The combined organic extracts were dried over Na2SO4 and evaporated under reduced pressure. The crude material was purified by precipitation from 70% EtOAc/Hexane (50 mL) to afford the title compound (8.9 g, 71%) as a solid. 1HNMR: (500 MHz, DMSO-cfe)] δ 7.97 (s, 1 H), 7.25 (d, J = 8.5 Hz, 2H), 6.90-6.88 (m, 4H), 5.21 (s, 2H), 3.74 (s, 3H), 3.71 (s, 3H). HPLC (10-90[3]-10 method): Rt = 2.29 min. (93%). Step 3. (2-amino-4-chloro-7-(4-methoxybenzyl)-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)methanol (XLI)
[00319] To a stirred solution of methyl 2-amino-4-chloro-7-(4-methoxybenzyl)-7H-pyrrolo[2,3- d]pyrimidine-5-carboxylate (10 g, 0.028 mol) in THF (200 mL) was added L-Selectride (21.9 g, 0.115 mol) at 0 0C. The resulting reaction mixture was stirred at RT for 2h. The reaction mixture was allowed to cool to 0 0C and diluted with MeOH (200 mL). The resulting mixture was warmed to RT and stirred for 2h. The reaction mixture was concentrated under reduced pressure and the crude material was purified by column chromatography [silica gel; 4% MeOH/dichloromethane] to afford the title compound (5.9 g, 63.85%) as a solid. 1H NMR (500 MHz, DMSO-Cf6)] δ 7.19 (d, J = 8.0 Hz, 2H), 6.90 (d, J = 8.0 Hz, 2H), 6.44 (s, 2H), 4.83- 4.81 (m 1 H), 4.50 (d, J = 15 Hz, 1 H), 4.36 (d, J = 15 Hz, 1 H), 3.73 (s, 3H), 3.66- 3.62 (m, 1 H), 3.45- 3.37 (m, 2H), 3.29-3.27 (m, 1 H), 3.24-3.19 (m, 1 H). HPLC (10-90[3]-10 method): Rt = 1.54 min. (93%).
Step 4. (2-amino-4-chloro-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol [00320] A mixture of (2-amino-4-chloro-7-(4-methoxybenzyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (5 g, 15 mmol) and TFA (35 ml) was stirred at 80 C. After 1h, H2SO4 (2 ml) was added to the reaction mixture and the heating was continued for another 8 hours. After consumption of the starting material (by TLC), the volatiles were evaporated under reduced pressure. To the residue was added crushed ice, and the pH was adusted to - 10 with aqueous NaOH. The precipitated solid was filtered, washed with water and dried under reduced pressure to afford (2-amino-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (5 g, 30%) as a brown solid. The obtained crude material was directly used for next step without further purification. 1H NMR (DMSO-ofe, 500 MHz): δ ppm 7.19 (bs, 1 H), 6.26 (bs, 2H), 4.83 (bs, 1 H), 3.64- 3.59 (m, 1 H), 3.54- 3.50 (m 1 H), 3.45- 3.42 (m, 1H), 3.39- 3.30 (m 1 H), 3.25- 3.21 (m, 1H); HPLC (10-90-15 method, Atlantis C-18 (250x4.6 mm, 5 μM), 5 mM NH4OAc) rt = 9.12 min; TLC system: 10% MeOH/DCM R,: 0.2.
Step 5. 5^(tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- amine (XLI)
[00321] To a stirred solution of (2-amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)methanol (6.4 g, 0.03 mol) in DMF (192 ml) were added imidazole (6.53 g, 0.096 mol) and TBDMS-CI (9.64 g, 0.064 mol) at 0 0C. The resulting reaction mixture was stirred at RT for 2h. After consumption of the starting material (By TLC), the reaction mass was diluted with water (600 ml) and extracted with EtOAc (3x 200 ml). The combined organic layers were evaporated and reduced pressure. The crude material was purified by column chromatography [60-120 silica gel; 3% MeOH/DCM] to afford the title compound (6.5 g, 64.6%) as an off-white solid. 1H-NMR (DMSO-Of6, 500 MHz): δ 7.20 (br s, 1H), 6.28 (br s, 2H), 3.77- 3.74 (m, 1H), 3.58- 3.53 (m 2H), 3.40- 3.37 (m, 1 H), 3.30-3.28 (m, 1H), 0.89 (s, 9H), 0.02 (s, 6H); LC-MS (0-90-5 method, Ascentis(R) Express C-18 75x2.7mm, 3.0 μM) rt = 2.25 min. m/z = 315.0 (M+1). TLC system: 10% MeOH /DCM, Rf: 0.6.
Example 38 [2-Amino-4-chloro-7-(4-fluoro-benzyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyhmidin-5-yl]- methanol (XLII)
[00322] A mixture of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (73.1 mg, 0.232 mmol) and Potassium tert-Butoxide (27 mg, 0.24 mmol) in DMSO was treated with 1 -(chloromethyl)-4-fluoro-benzene (28 μL, 0.24 mmol)] for 10 min at rt. Work-up (EtOAc/water) and concentration gave crude 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-(4-fluoro- benzyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (86.3 mg; Yield = 87.9%). This crude material was dissolved in MeOH, and treated with 4 drops of cone. aq. HCI at 4 0C overnight. Concentration and work-up (EtOAc vs. ammonia) gave the desired alochol, which was crystallized from EtOAc/hexane to give the title compound (32 mg; Yield = 45%). 1H NMR (400 MHz, DMSO-Of6) δ ppm 7.30 (dd, J=8.3, 6.1 Hz, 2 H), 7.17 (t, J=8.6 Hz, 2 H), 6.48 (s, 2 H), 4.85 (t, J=5.2 Hz, 1 H), 4.57 (d, J=15.7 Hz, 1 H), 4.43 (d, J=15.2 Hz, 1 H), 3.65 (dt, J=9.6, 4.5 Hz, 1 H), 3.48 (t, J=9.6 Hz, 1 H), 3.41 (dd, J=9.9, 3.5 Hz, 1 H), 3.21 - 3.28 (m, 1 H). LC-MS (5-100-5 method): Rt = 2.12 min; m/z = 309.1 (M+1). Example 39
[2-Amino-4-chloro-7-(1,4,5-trimethyl-1H-imidazol-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-methanol (XLIII)
[00323] A 4-mL vial was charged with Sodium-tert-butoxide (27 mg, 0.28 mmol) and 5-(tert-Butyl- dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (28.7 mg, 0.091 mmol). Dimethyl sulfoxide (1.0 ml_) was added, and the vial was shaken for 1 minute to dissolve most of the solids. Commercial 2-(chloromethyl)-1 ,4,5-trimethyl-1 H-imidazole hydrochloride (25 mg, 0.13 mmol) was added portionwise over 1 min. The alkylation was complete within 2 min. 1.0 M of hydrogen chloride in Water (1.5 ml_, 1.5 mmol) was added to induce desilylation. The reaction was complete after 30 min, and was purified by reverse-phase HPLC. The desired fraction was neutralized with Dowex 66 resin (polyamine) until pH = 5. The resin was filtered off, and evaporation gave the title compound as a solid residue (6.33 mg; Yield = 21.5%). 1H NMR (400 MHz, CDCI3) δ ppm 4.97 (d, J=16.7Hz, 1 H), 4.72 (d, Λ=16.4Hz, 1H), 3.80 - 3.85 (m, 1H), 3.72 - 3.78 (m, 4H), 3.61 - 3.70 (m, 1H), 3.42-3.50 (d, m, 1H), 2.28 (s, 3H), 2.26 (s, 3H). LC-MS (5-100-5 method): Rt = 1.58 min.; m/z = 323.0 (M+1). Example 40
(2-amino-4-chloro-7-((5-ethyl-1,4-dimethyl-1H-pyrazol-3-yl)methyl)-6,7-dihydro-SH-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (XLIV) Step 1. Ethyl 3-methyl-2,4-dioxohexanoate
[00324] Sodium (5.13 g, 223 mmol) was dissolved in Ethanol (300 mL) at rt over 1h. 3-Pentanone
(21.28 mL, 200.8 mmol) was added, and the mixture was stirred for 45 min at rt. Diethyl oxalate (44.02 g, 301.2 mmol) was added, and the mixture was stirred at rt for for 30 min. The solvent was evaporated on a rotary evaporator (Tbath = 45 0C). The residue was taken in brine (200 mL), and washed with EtOAc (3x 150 mL). The organic layers were discarded. The pH was adjusted to 5 with cone. HCI, which brought the pH to 1 , followed by K3PO4- H2O (5.6 g), which neutralized the mixture to pH= 5. The aqueous layer was extracted with EtOAc (3x150 mL). Evaporation gave 17.2 g of crude that was quite pure, except for residual diethyl oxalate (about 33 mol%). Flash chromatography (240 g SiO2, EtOAc/Hexane 10:90 to 30:70) gave 9.3 g of Ethyl 3-methyl-2,4-dioxohexanoate as a colorless oil (diketo tautomer in DMSO-d6). Step 2. Ethyl 5-ethyl-1 ,4-dimethyl-1 H-pyrazole-3-carboxylate
[00325] A solution of Ethyl 3-methyl-2,4-dioxohexanoate (2.05 g, 7.16 mmol) in ethanol (18 mL) was treated with N-Methylhydrazine (381 uL, 7.16 mmol) at rt for 10 min. The mixture was evaporated. The crude NMR spectrum indicated two isomeric products in 1 :0.7 ratio, and TLC showed very different Rf values (0.9 and 0.5 in EtOAc/hexane 1 :1 ). Chromatography (EtOAc/hexane 0:100 to 100:0 gradient) gave the undesired ethyl 3-ethyl-1 ,4-dimethyl-1H-pyrazole-5-carboxylate (0.83 g, Rf = 0.9 in EtOAc/hexane 1 :1 ) and the desired, more polar ethyl 5-ethyl-1 ,4-dimethyl-1 H-pyrazole-3-carboxylate
(0.84 g, Rf = 0.5 in EtOAc/hexane 1 :1 ).
Step 3. (5-ethyl-1,4-dimethyl-1H-pyrazol-3-yl)methanol
[00326] A solution of ethyl 5-ethyl-1 ,4-dimethyl-1 H-pyrazole-3-carboxylate (630 mg, 3.21 mmol) in tetrahydrofuran (15 ml_) was treated with super-hydride (0.79 g, 7.50 mmol) at rt for 5 min. The reduction was very clean. The reaction was quenched with 5 mL MeOH and evaporated. Brine (50 mL) was added, followed by H2O2 30% (5 mL) (caution! very vigorous and exothermic fizz). Extraction with /-
PrOH/CH2CI2 5:100 (3x50 mL) afforded crude (5-ethyl-1 ,4-dimethyl-1 H-pyrazol-3-yl)methanol, which was very clean by NMR and used without without further purification.
Step 4. 3-(chloromethyl)-5-ethyl-1,4-dimethyl-1H-pyrazole
[00327] A solution of (5-ethyl-1 ,4-dimethyl-1 H-pyrazol-3-yl)methanol (148 mg, 0.960 mmol) in
Methylene chloride (1 mL) was treated wuth Thionyl chloride (110 μL, 1.5 mmol) (caution! thionyl chloride is extremely reactive-add slowly). After 10 min, the mixture was concentrated on a rotary evaporator, and dried on high vacuum overnight to give clean 3-(chloromethyl)-5-ethyl-1 ,4-dimethyl-1 H- pyrazole hydrochloride, which was used without further purification.
Step 5. 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((5-ethyl-1,4-dimethyl-1H-pyrazol-3- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
[00328] A mixture of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (77 mg, 0.24 mmol) and 3-(chloromethyl)-5-ethyl-1 ,4-dimethyl-1 H-pyrazole (57 mg,
0.33 mmol) in Dimethyl sulfoxide (0.5 mL) was treated with 95% Sodium hydride (25 mg, 1.0 mmol) at rt.
No reaction occurred after 10 min. Upon addition of another portion of Sodium hydride (12 mg, 0.50 mmol), the reaction went to completion within 10 min. Work-up (EtOAc/water) and evaporation gave the crude title compound (124 mg) which was used without further purification.
Step 6. (2-amino-4-chloro-7-((5-ethyl-1,4-dimethyl-1H-pyrazol-3-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XLIV)
[00329] A solution of crude 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((5-ethyl-1 ,4-dimethyl-
1 H-pyrazol-3-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (134 mg, 0.297 mmol) in
Methanol (2.0 mL) and Water (0.30 mL, 17 mmol) was treated wth Trifluoroacetic Acid (0.30 mL, 3.9 mmol) at rt for 45 min. The mixture was injected on a preparative HPLC (20-70% MeOH/water, 0.1%
TFA). The desired fractions were combined, the MeOH was evaporated, and the ammonia was added.
Extraction with EtOAc (4 times) gives 45 mg of the title compound. 1H NMR (400 MHz, CD3OD:CDCI3
1 :2) δ 4.45 (s, 2H), 3.82 (dd, J= 3.54, 10.86 Hz, 1 H), 3.72 (s, 3H), 3.51 - 3.58 (m, 2H), 3.49 (dd, J = 3.79,
10.36 Hz, 1H), 3.33 - 3.39 (m, 1 H), 2.58 (q, J= 7.58 Hz, 2H), 1.90 (s, 3H), 1.12 (t, J= 7.58 Hz, 3H). LC-
MS (5-100-5 method): Rt = 2.06 min.; m/z = 337.1 (M+1 ).
Example 41
{2-Amino-4-chloro-7-[1-(5-ethyl-1,4-dimethyl-1H-pyrazol-3-yl)-ethyl]-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl}-methanol (XL V) Step 1. S-EthyMJ-dimethyl-IH-pyrazole-S-carbaldehyde
[00330] A solution of(5-ethyl-1 ,4-dimethyl-1 H-pyrazol-3-yl)methanol (1.4 g, 9.1 mmol) in
Methylene chloride (28 mL) was treated with Dess-Martin periodinane (4.7 g, 11 mmol) at it The reaction was very rapid but incomplete. Addition of a second portion of Dess-Martin periodinane (1.1 g, 2.6 mmol) drove the reaction to completion. NaOH 1 M (60 mL) was added, and the aldehyde was extracted. The crude was chromatographed (24g SiO2, EtOAc/Hexane 0:100 to 100:0) to afford the title aldehyde as a white solid wich has a tendency to sublime.
Step 2. 1-f5-Ethyl-1,4-dimethyl-1H-pyrazol-3-yl)^thanol
[00331] A solution of 5-Ethyl-1 ,4-dimethyl-1 H-pyrazole-3-carbaldehyde (1.00 g, 6.57 mmol) in
Tetrahydrofuran (30 mL) was treated at rt with 3.0 M of Methylmagnesium bromide in Ether (2.2 mL, 6.6 mmol). The reaction was instantaneous and clean. The reaction was quenched with MeOH, evaporated, and extracted into EtOAc to give the desired product, which was used in the next step without further purification.
Step 3. 3-(1-Chloro-ethyl)-5-ethyl-1,4-dimethyl-1 H-pyrazole
[00332] A solution of [1 -(5-Ethyl-1 ,4-dimethyl-1 H-pyrazol-3-yl)-ethanol (0.45 g, 2.7 mmol) in methylene chloride (9 mL, 100 mmol) was treated at 0 °C with Thionyl chloride (450.0 μL, 6.169 mmol).
The reaction was complete within 10 min. Aqueous NaHCO3 was added, and the desired product was extracted in methylene chloride. Chromatography (EtOAc/hexane 0:100 to 100:0) gave 100 mg of the desired product (impure). Loading on the column an additional 1 mL of JPr2NEt, and eluting with 100%
EtOAc 100% gave another 79 mg of material, very clean by NMR.
Step 4. 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-[1-(5-ethyl-1,4-dimethyl-1H-pyrazol-3- yl)-ethyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine
[00333] A solution of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (75.0 mg, 0.238 mmol) and Sodium hydride (6.29 mg, 0.262 mmol) in DMSO (0.7 mL) was treated with 3-(1 -Chloro-ethyl)-5-ethyl-1 ,4-dimethyl-1 H-pyrazole (48.9 mg, 0.262 mmol) at rt for 5 min. Another 7 mg of Sodium hydride and 20 mg of 3-(1-Chloro-ethyl)-5-ethyl-1 ,4-dimethyl-1 H-pyrazole were added to bring the reaction to completion. Work-up (EtOAc and Water) gave a 1 :1 mixture of isomers, that were used in the next step without further separation/purification.
Step 5. {2-Amino-4-chloro-7-[1-(5-ethyl-1,4-dimethyl-1H-pyrazol-3-yl)-ethyl]-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (XL V)
[00334] The crude mixture from the previous step (110 mg) was dissolved in 4 mL MeOH and treated with cone. Aq. HCI (-200 μL) at rt. Desylation occured within minutes to give two isomers in a 1 :1 ratio. Preparative reverse-phase HPLC gave (20 to 70% CH3CN, 0.1% TFA) two isomers in a 1 :1 ratio. 1H
NMR (400 MHz, Acetone-d6) δ ppm 5.95 (br. s., 1 H), 5.40 (q, J=7.1 Hz, 1 H), 3.82 (dd, J=10.6, 3.5 Hz, 1
H), 3.74 (s, 3 H), 3.61 (dd, J=10.1 , 3.5 Hz, 1 H), 3.56 (dd, J=WS, 7.8 Hz, 1 H), 3.30 - 3.37 (m, 2 H), 3.20
- 3.29 (m, 1 H), 2.60 (q, J=7.6 Hz, 2 H), 1.86 (s, 3 H), 1.51 (d, Λ=7.1 Hz, 3 H), 1.10 (t, J=7.6 Hz, 3 H). LC-
MS (5-100-5 method): Rt = 2.16 min.; m/z = 351.1 (M+1). Example 42
(2-amino-4-chloro-7-((2-methyl-2,4,5,6-tetrahydrocyclopenta[c]pyrazoh3-yl)methyl)-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl)methanol (XL Vl)
Step 1. Hydroxy-[2-oxo-cyclopent-(Z)-ylidene]-acetic acid ethyl ester
[00335] Anhydrous ethanol (80.0 ml_) was treated with sodium hydride (3.39 g, 141 mmol) at 0 0C and then stirred for 20 minutes at room temperature until hydrogen gas evolution ceased. Diethyl oxalate
(9.60 mL, 70.7 mmol) was added, followed by cyclopentanone (6.26 ml_, 70.7 mmol) in anhydrous ethanol (20.0 mL). The reaction mixture was stirred at room temperature for 4 hours and evaporated. The residue was diluted with EtOAc (20OmL). The organic layer was washed with brine (30.0 mLx3), dried over Na2SO4, and evaporated to give the title compound as a colorless oil (9.0 g, purity ≥ 90%, yield
62.0%). 1H NMR (CDCI3): δ ppm 12.95 (s, 1 H), 4.36 (q, 2H), 2.97 (t, 2H), 2.49 (t, 2H), 2.00 (m, 2H), 1.39
(t, 3H).
Step 2. 2-Methyl-2,4,5,6-tetrahydro-cyclopentapyrazole-3-carboxylic acid ethyl ester (A) and 1-
Methyl-1,4,5,6-tetrahydro-cyclopentapyrazole-3-carboxylic acid ethyl ester (B)
[00336] A solution of hydroxy-[2-oxo-cyclopent-(Z)-ylidene]-acetic acid ethyl ester (7.20 g, 39.1 mmol) in ethanol (100 mL, 2000 mmol) was treated with N-methylhydrazine (2.08 mL, 39.1 mmol) at room temperature for 10 minutes and then heated to reflux for 30 minutes. After cooling, the reaction mixture was evaporated to dryness, and the crude was purified by flash chromatography (hexane/EtOAc, 4:1 to
1 :4) to give 2-methyl-2,4,5,6-tetrahydro-cyclopentapyrazole-3-carboxylic acid ethyl ester as a white solid
(2.87g, purity > 90%, yield 34.0%). 1H NMR (DMSO-d6): δ ppm 4.19 (q, 2H), 3.95 (s, 3H), 2.65 (t, 2H),
2.56 (t, 2H), 2.28 (m, 2H), 1.23 (t, 3H). LC-MS (5-100-7 method):Rt=6.533 min; M+1 =194.8.
[00337] The chromatography also gave i-methyl-i ^.δ.θ-tetrahydro-cyclopentapyrazole-S- carboxylic acid ethyl ester as a yellow solid (0.26 g, purity ≥ 90%, yield 3.40%). 1H NMR (DMSO-d6): δ ppm 4.17 (q, 2H), 3.73 (s, 3H), 2.66 (m, 4H), 2.50 (m, 2H), 1.23 (t, 3H). LC-MS (5-100-7 method):Rt=5.655 min; M+1 =194.8.
Step 3. 3-(hydroxymethyl)-2-methylene-2,4,5, 6-tetrahydrocyclopenta[c]pyrazol-2-ium
[00338] A solution of 2-methyl-2,4,5,6-tetrahydro-cyclopentapyrazole-3-carboxylic acid ethyl ester
(1.50 g, 7.72 mmol) in tetrahydrofuran (10.0 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (17.8 mL, 17.8 mmol) at room temperature for 20 min. The reaction mixture was quenched with MeOH (5.00 mL) and evaporated. The residue was diluted with brine (50.0 mL) and then
30% H2O2 (5.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a white solid. (1.18 g, purity 99.0%, yield 77.0%). 1H NMR (DMSO-d6): δ ppm 5.08 (t, 1 H),
4.38 (d, 2H), 3.67 (s, 3H), 2.48 (m, 4H), 2.25 (m, 2H). LC-MS (5-100-7 method) :Rt=3.440 min;
M+1 =152.9.
Step 4. 3-{chloromethyl)-2-methyl-2,4,5,6-tetrahydrocyclopenta[c]pyrazole
[00339] A solution of 3-(hydroxymethyl)-2-methylene-2,4,5,6-tetrahydrocyclopenta[c]pyrazol-2- ium (500.0 mg, 3.285 mmol) in methylene chloride (5.00 mL) was treated with thionyl chloride (718.9 μL,
9.856 mmol).The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water (5OmL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane: isopropanol (100/5, 20.0 ml_x3). The organic layer was dried over Na2SO4, and evaporated to give the title compound as a white solid (460 mg, purity > 90%, yield 74.0%). LC-MS (5-100-7 method):Rt=5.621 min; M+1 =170.8.
Step 5. (2-amino-4-chloro-7-((2-methyl-2,4,5, 6-tetrahydrocyclopenta[c]pyrazol-3-yl)methyl)-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol
[00340] A mixture of 3-(chloromethyl)-2-methyl-2,4,5,6-tetrahydrocyclopenta[c]pyrazole (37.6 mg,
0.221 mol) in dimethyl sulfoxide (1.00 mL) was treated with sodium hydride (5.56 mg, 0.232 mmol) at 0
0C for 10 minutes and then allowed to reach room temperature. 5-[2-(terf-butyl-dimethyl-silanyloxy)-1 ,1- dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (69.5 mg, 0.221 mmol) in dimethyl sulfoxide (1.00 mL) was added, and stirring was continued at room temperature for 20 minutes.
The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (1.00 mL, 0.0060 mol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (36.0 mg, purity 99.0%, yield 48.7%). 1H NMR (DMSO-d6): δ ppm 6.61 (br. s, 2H), 4.91 (t, 1 H), 4.89 (d, 1 H), 4.53 (d,
1H), 3.74 (s, 3H), 3.65 (m,1H), 3.47-3.27 (m, 4H), 2.50 (m, 2H), 2.38 (t, 2H), 2.26 (m, 2H). LC-MS (5-100-
7 method) :Rt=4.097 min; M+1 =334.6.
Example 43 p-Amino^-chloro-T-fi-methyl-IΛS.β-tetrahydro-cyclopentapyrazol-S-ylmethyO-βJ-dihydro-SH- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (XL VII)
Step 1. (1-Methyl-1,4,5,6-tetrahydro-cyclopentapyrazol-3-yl)-methanol
[00341] A solution of 1 -methyl-1 ,4,5,6-tetrahydro-cyclopentapyrazole-3-carboxylic acid ethyl ester
(250 mg, 1.3 mmol) in tetrahydrofuran (10.0 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (3.86 mL, 3.86 mmol) at room temperature for 20 min. The reaction mixture was quenched with MeOH (5.00 mL) and evaporated. The residue was diluted with brine (50.0 mL) and then
30% H2O2 (5.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a white solid (250 mg, purity 99.0%, yield 77.0%). 1H NMR (DMSO-d6): δ ppm 4.79 (t, 1 H),
4.27 (d, 2H), 3.61 (s, 3H), 2.63 (t, 2H), 2.50 (t, 2H), 2.44 (m, 2H). LC-MS (5-100-7 method): Rt=3.352 min;
M+1 =152.9.
Step 2. 3-{chloromethyl)-1-methyl-1,4,5,6-tetrahydrocyclopenta[c]pyrazole
[00342] A solution of (1 -methyl-1 ,4,5,6-tetrahydro-cyclopentapyrazol-3-yl)-methanol (200 mg, 1 mmol) in methylene chloride (1.68 mL) was treated with thionyl chloride (288 μL, 3.94 mmol). The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 mLx3). The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/EtOAc, 1 :3) to give the title compound as a colorless oil (89.0 mg, purity > 90%, yield 40.0%). 1H NMR (DMSOd6): δ ppm 4.57 (s, 2H), 3.65 (s, 3H), 2.63 (t, 2H), 2.53 (t, 2H), 2.46 (m,
2H). LC-MS (5-100-7 method):Rt=5.573 min; M+1=170.8
Step 3. [2-Amino-4-chloro-7-(1-methyl-1,4,5,6-tetrahydro-cyclopentapyrazol-3-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (XL VII)
[00343] A mixture of S-chloromethyl-i-methyl-i ^.δ.e-tetrahydro-cyclopentapyrazole (42.3 mg,
0.248 mmol) in dimethyl sulfoxide (0.50 mL) was treated with sodium hydride (5.56 mg, 0.232 mmol) at 0
0C for 10 minutes and then allowed to reach room temperature. A solution of 5-[2-(tert-butyl-dimethyl- silanyloxy)-1 , 1 -dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (78.0 mg, 0.248 mmol) in dimethyl sulfoxide (1.00 mL) was added, and stirred was continued at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water
(1.00 mL, 6.00 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 40% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (50.0 mg, purity 99.0%, yield 60%). 1H NMR (DMSOd6): δ ppm 6.40 (br. s, 2H), 4.82 (t, 1 H)1 4.27 (dd, 2H), 3.59 (m,
1 H), 3.57 (s, 3H), 3.39-3.11 (m, 4H), 2.56 (t, 2H), 2.33 (m, 2H), 2.26 (t, 2H). LC-MS (5-100-7 method):Rt=4.200 min; M+1 =334.6.
Example 44
(2-amino-4-chloro-7-((2, 6, 6-trimethyl-4,5, 6, 7-tetrahydro-2H-indazol-3-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XL VIII)
Step 1. [4,4-Dimethyl-2-oxo-cyclohex-(Z)-ylidene]-hydroxy-acetic acid ethyl ester
[00344] Anhydrous ethanol (80.0 mL) was treated with sodium hydride (4.30 g, 179 mmol) at 0 0C and then stirred for 20 minutes at room temperature until hydrogen gas evolution ceased. Diethyl oxalate
(12.2 mL, 89.6 mmol) was added, followed by 3,3-dimethylcyclohexanone (12.4 mL, 89.6 mmol) in anhydrous ethanol (20.0 mL). The reaction mixture was stirred at room temperature for 6 hours and evaporated. The residue was diluted with EtOAc (20OmL). The organic layer was washed with brine (30.0 mLx3), dried over Na2SO4, and evaporated to give the title compound as colorless oil (15.0 g, purity ≥
90%, yield 66.6%). 1H NMR (CDCI3): δ ppm 12.20 (s, 1 H), 4.34 (q, 2H), 2.50 (t, 2H), 2.22 (s, 2H), 1.48 (t,
2H), 1.38 (t, 3H), 0.98 (s, 6H).
Step 2. Ethyl 2, 6, 6-trimethyl-4,5, 6, 7-tetrahydro-2H-indazole-3-carboxylate and ethyl 1, 6, 6-trimethyl-
4,5,6,7-tetrahydro-1H-indazole-3-carboxylate
[00345] A solution of [4,4-Dimethyl-2-oxo-cyclohex-(Z)-ylidene]-hydroxy-acetic acid ethyl ester
(11.1 g, 49.0 mmol) in ethanol (120 mL) was treated with N-methylhydrazine (2.61 mL, 49.0 mmol) at room temperature for 10 minutes and then heated to reflux for 30 minutes. After cooling, the reaction mixture was evaporated. The crude was purified by flash chromatography (hexane/EtOAc, 4:1 to 1 :4) to give ethyl 2,6,6-trimethyl-4,5,6,7-tetrahydro-2H-indazole-3-carboxylate as a white solid (2.87g, purity ≥
90%, yield 34.0%). 1H NMR (DMSO-d6): δ ppm 4.23 (q, 2H), 3.96 (s, 3H), 2.61 (t, 2H), 2.29 (s, 2H), 1.44 (t, 2H), 1.27 (t, 3H), 0.90 (s, 6H). LC-MS (5-100-7 method): Rt=7.634 min; M+1 =236.8.
[00346] The chromatography also gave ethyl 1 , 6, 6-trimethyl-4,5,6,7-tetrahydro-1H-indazole-3- carboxylate as colorless oil (7.23 g, purity 99.0%, yield 61.7%). LC-MS (5-100-7 method):Rt=6.859 min;
M+1 =236.8.
Step 3. (2, 6, 6-Trimethyl-4, 5, 6, 7-tetrahydro-2H-indazol-3-yl)-methanol
[00347] A solution of ethyl 2,6,6-trimethyl-4,5,6,7-tetrahydro-2H-indazole-3-carboxylate (2.14g,
9.06mmol) in tetrahydrofuran (10.0 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (22.6 mL, 22.6 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (5.00 mL) and evaporated. The residue was diluted with brine (50.0 mL) and then
30% H2O2 (5.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a yellow solid (1.76 g, purity 99.0%, yield 84.4%). 1H NMR (DMSO-d6): δ ppm 5.04 (t, 1 H),
4.38 (d, 2H), 3.70 (s, 3H), 2.42 (t, 2H), 2.25 (s, 2H), 1.43 (t, 2H), 0.93 (s, 6H). LC-MS (5-100-7 method):Rt=4.489 min; M+1 =194.9.
Sf ep 4. 3-Chloromethyl-2, 6, 6-trimethyl-4, 5, 6, 7-tetrahydro-2H-indazole
[00348] A solution of (2, 6, 6-trimethyl-4,5,6,7-tetrahydro-2H-indazol-3-yl)-methanol (1.50 g, 7.72 mmol) in methylene chloride (10.0 mL) was treated with thionyl chloride (2.25 mL, 30.9 mmol). The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3).
The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/EtOAc, 1 :3) to give the title compound as a white solid (1.34 g, purity ≥ 90%, yield 81.6%). 1H NMR (DMSO-d6): δ ppm 4.83 (s, 2H), 3.75 (s, 3H), 2.44 (t, 2H), 2.27 (s, 2H), 1.45 (t, 2H),
0.93 (s, 6H). LC-MS (5-100-7 method):Rt=6.584 min; M+1=212.8.
Step 5. (2-Amino^t-chloro-7-((2,6,6-trimethyl-4,S,6J-tetrahydro-2H-indazol-3-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XL VIII)
[00349] A mixture of 3-chloromethyl-2,6,6-trimethyl-4,5,6,7-tetrahydro-2H-indazole (54.8 mg,
0.258 mmol) in dimethyl sulfoxide (0.50 mL) was treated with sodium hydride (6.50 mg, 0.271 mol) at 0
0C for 10 minutes and then brought to room temperature. 5-[2-(fe/t-butyl-dimethyl-silanyloxy)-1 ,1- dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (81.2 mg, 0.258 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes.
The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (1.00 mL, 6.00 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 40% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (70.0mg, purity 99.0%, yield 70.0%). 1H NMR (DMSO-d6): δ ppm 6.53 (br. s, 2H), 4.87 (t, 1 H), 4.55 (d, 1 H), 4.36 (d,
1H), 3.67 (s, 3H), 3.62 (m, 1 H), 3.41-3.23 (m, 4H), 2.39 (t, 2H), 2.27 (s, 2H), 1.44 (t, 2H), 0.94 (s, 3H),
0.93 (S, 3H). LC-MS (5-100-7 method): Rt=4.824 min; M+1 =376.6.
Example 45 [2-Amino-4-chloro-7-(1, 6, 6-trimethyl-4, 5, 6, 7-tetrahydro-1H-indazol-3-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (XLIX)
Step 1. (1,6,6-Trimethyl-4,5,6,7-tetrahydro-1H-indazol-3-yl)-methanol
[00350] A solution of ethyl 1 ,6,6-trimethyl-4,5,6,7-tetrahydro-1 H-indazole-3-carboxylate (3.0Og,
12.7mmol) in tetrahydrofuran (20.0 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (38.1 mL, 38.1 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (5.00 mL) and evaporated. The residue was diluted with brine (50.0 mL) and then
30% H2O2 (5.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a colorless oil (2.30 g, purity ≥ 90%, yield 83.9%). 1H NMR (DMSO-d6): δ ppm 4.67 (t, 1 H),
4.25 (d, 2H), 3.52 (s, 3H), 2.37 (t, 2H), 2.26 (s, 2H), 1.37 (t, 2H), 0.92 (s, 6H). LC-MS (5-100-7 method):Rt=4.445 min; M+1 =194.9.
Step 2. 3-Chloromethyl-1,6,6-trimethyl-4,5,6, 7-tetrahydro-1 H-indazole
[00351] A solution of (1 ,6,6-trimethyl-4,5,6,7-tetrahydro-1 H-indazol-3-yl)-methanol (1.50 g, 7.72 mmol) in methylene chloride (10.0 mL) was treated with thionyl chloride (1.69 mL, 23.2 mmol). The reaction mixture was stirred at room temperature for 20 minutes, and evaporated. The residue was diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3). The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/EtOAc, 1 :3) to give the title compound as a pale yellow oil (1.54 g, purity 95.0%, yield 89.1%). 1H NMR (DMSO-d6): δ ppm 4.62(s,
2H), 3.61 (s, 3H), 2.43 (t, 2H), 2.34 (s, 2H), 1.45 (t, 2H), 0.97 (s, 6H). LC-MS (5-100-7 method):Rt=6.891 min; M+1=212.8
Step 3. [2-Amino-4-chloro-7-(1, 6, 6-trimethyl-4,5, 6, 7-tetrahydro- 1 H-indazol-3-ylmethyl)-6, 7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (XLIX)
[00352] A solution of 3-chloromethyl-1 ,6,6-thmethyl-4,5,6,7-tetrahydro-1 H-indazole (47.0 mg,
0.221 mmol) in dimethyl sulfoxide (0.50 mL) was treated with sodium hydride (5.56 mg, 0.232 mol) at 0
0C for 10 minutes and then brought to room temperature. A solution of 5-[2-(fert-butyl-dimethyl- silanyloxy)-1 ,1-dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (69.5 mg, 0.221 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water
(1.00 mL, 6.00 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 45% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (56.0mg, purity 99.0%, yield 67.0%). 1H NMR (DMSO-d6): δ ppm 6.46 (br. s, 2H), 4.83 (t, 1 H), 4.45 (d, 1 H), 4.28 (d,
1H), 3.64 (m, 1 H), 3.64 (s, 3H), 3.44-3.19 (m, 4H), 2.32 (s, 2H), 2.26 (t, 2H)1 1.38 (t, 2H), 0.96 (s, 3H),
0.94 (s, 3H). LC-MS (5-100-7 method): Rt=5.078 min; M+1 =376.6.
Example 46
[2-Amino-4-chloro-7-(1-methyl-4,5)6,7-tetrahydro-1H-indazol-3-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (L)
Step 1. Hydroxy-[2-oxo-cyclohex-(Z)-ylidene]-acetic acid ethyl ester
[00353] Anhydrous ethanol (80.0 mL) was treated with sodium hydride (4.30 g, 179 mmol) at 0 0C and then stirred for 20 minutes at room temperature until hydrogen gas evolution ceased. Diethyl oxalate
(12.2 mL, 89.6 mmol) was added to the mixture, followed by cyclohexanone (9.28 mL, 89.6 mmol) in anhydrous ethanol (40.0 mL). The reaction mixture was stirred at room temperature for 6 hours and evaporated. The residue was diluted with EtOAc (20OmL) and the organic layer was washed with brine
(30.0 mLx3), dried over Na2SO4, and evaporated to give the title compound as a yellow oil (9.50 g, purity
95.0%, yield 51.0%). 1H NMR (DMSO-d6): δ ppm 11.92 (br. s, 1 H), 4.20 (q, 2H), 2.34 (t, 2H), 2.14 (t, 2H),
1.64 (m, 2H), 1.54 (m, 2H), 1.25 (t, 3H).
Step 2. 1 -Methyl-4, 5, 6, 7-tetrahydro- 1 H-indazole-3-carboxylic acid ethyl ester
[00354] Hydroxy-[2-oxo-cyclohex-(Z)-ylidene]-acetic acid ethyl ester (4.50 g, 22.7 mmol) in ethanol (60.0 mL, 1030 mmol) was treated with N-methylhydrazine (1.21 mL, 22.7 mmol) at room temperature overnight and evaporated. The crude was purified by flash chromatography (hexane/EtOAc,
4:1 to 1:4) to give the title compound as a yellow oil (2.00 g, purity 95.0%, yield 42.3%). 1H NMR (DMSO- d6): δ ppm 4.23 (q, 2H), 3.96 (s, 3H), 2.61 (t, 2H), 2.49 (s, 2H), 1.66 (m, 4H), 1.27 (t, 3H). LC-MS (5-100-7 method):Rt=6.851 min; M+1 =208.8
Step 3. (1-Methyl-4,5,6,7-tetrahydro-1H-indazol-3-yl)-methanol
[00355] A solution of 1 -methyl-4,5,6,7-tetrahydro-1 H-indazole-3-carboxylic acid ethyl ester (1.50 g, 7.20 mmol) in tetrahydrofuran (10.0 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (18.0 mL, 18.0 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (5.00 mL) and evaporated. The residue was diluted with brine (50.0 mL) and then
30% H2O2 (5.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a white solid (1.10 g, purity 95.0%, yield 92.0%). 1H NMR (DMSO-d6): δ ppm 4.69 (t, 1 H),
4.27 (d, 2H), 3.56 (s, 3H), 2.49 (t, 2H), 2.39 (t, 2H), 1.71 (m, 2H), 1.58 (m, 2H). LC-MS (5-100-7 method):Rt=3.836 min; M+1=166.9
Step 4. 3-Chloromethyl-1-methyl-4,5,6,7-tetrahydro-1H-indazole
[00356] A solution of (1-methyl-4,5,6,7-tetrahydro-1 H-indazol-3-yl)-methanol (0.500 g, 3.01 mmol) in methylene chloride (10.0 mL) was treated with thionyl chloride (0.658 mL, 9.02 mmol). The reaction mixture was stirred at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3). The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography
(hexane: EtOAc=I :3) to give the title compound as a yellow solid (400 mg, purity ≥ 90%, yield 60.0%). 1H
NMR (DMSO-d6): δ ppm 4.61 (s, 2H), 3.62 (s, 3H), 2.54 (t, 2H), 2.42 (t, 3H), 1.74 (m, 2H), 1.63 (m, 2H).
LC-MS (5-100-7 method): Rt=6.076 min; M+1=184.8
Step 5. [Σ-Amino-J-chloro-T-fi-methyM&βJ-tetrahydro-IH-indazol-S-ylmethyiyβJ-dihydro-SH- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (L)
[00357] A mixture of 3-chloromethyl-1 -methyl-4,5,6,7-tetrahydro-1 H-indazole (17.6 mg, 0.0953 mmol) in dimethyl sulfoxide (0.50 mL) was treated with sodium hydride (2.40 mg, 0.100 mmol) at 0 0C for
10 minutes and then brought to room temperature. A solution of 5-[2-(fe/?-butyl-dimethyl-silanyloxy)-1 ,1- dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (30.0 mg, 0.0953 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes.
The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (1.0 mL,
6.00 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (21.5 mg, purity 99.0%, yield 64.7%). 1H NMR (DMSOd6): δ ppm 6.42 (br. s, 2H), 4.83 (t, 1 H), 4.41 (d, 1 H), 4.29 (d,
1 H), 3.65 (m, 1 H), 3.60 (s, 3H), 3.41 (t, 1 H), 3.27-3.17 (m, 4H), 2.53 (m, 2H), 2.25 (t, 2H), 1.71 (m, 2H),
1.59 (m, 2H). LC-MS (5-100-7 method): Rt=4.540 min; M+1 =348.6.
Example 47
[2-Amino-7-(3-bromo-5,6-dihydro-4H-pyrrolo[1,2-b]pyrazol-2-ylmethyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl]-methanol (Ll)
Step 1. i-Nitroso-pyrrolidine-2-carboxylic acid
[00358] A mixture of DL-proline (10.0 g, 86.8 mmol) and sodium nitrite (8.39 g, 122 mmol) in water (20.0 mL) was treated with 12 M of hydrogen chloride in water (10.1 mL, 122 mmol) at 0 0C for 2 hours and then at room temperature for another 2 hours. The reaction mixture was extracted with tert- butyl methyl ether (50.0 mLx5). The organic layer was washed with brine (15.0 mLx2), dried over
Na2SO4, and evaporated to give the title compound as a white solid (9.50 g, purity 95.0%, yield 72.1%).
LC-MS (5-100-7 method):Rt=2.948 and 3.068 min; M+1=144.8.
Step 2. 3-Oxo-3,4,5,6-tetrahydropyrrolo[1,2-c][1,2,3]oxadiazol-7-ium-3a-ide
[00359] A mixture of i-nitroso-pyrrolidine-2-carboxylic acid (9.50 g, 65.9 mmol) in tetrahydrofuran
(20.0 mL) was treated with trifluoroacetic anhydride (14.0 mL, 98.9 mmol) under nitrogen at 0 0C and then at room temperature for 2 hours. The reaction mixture was evaporated, the residue was purified by flash chromatography (dichloromethane/MeOH, 100:5 to 100:10) to give the title compound as a brown oil
(3.50 g, purity > 90%, yield 42.0%). %). LC-MS (5-100-7 method): Rt=2.623 min; M+1 =126.9
Step 3. 5,6-Dihydro-4H-pyrrolo[1,2-b]pyrazole-2-carboxylic acid ethyl ester
[00360] A mixture of 3-oxo-3,4,5, 6-tetrahydropyrrolo[1 ,2-c][1 ,2,3]oxadiazol-7-ium-3a-ide (1.00 g,
7.93 mmol) in 1 ,2-diethoxyethane (5.00 mL) was purged with nitrogen three times and heated to 125 °C.
Ethyl propiolate (1.04 mL, 10.3 mmol) was added dropwise over 10 minutes. The reaction was stirred at
125 °C for 3 hours and evaporated. The residue was purified by purified by reverse-phase preparative
HPLC (30 to 70% CH3CN/H2O, 0.1% TFA) to give the title compound as a brown oil (80.0 mg, purity >
90%, yield 6.0%). LC-MS (5-100-7 method):Rt=5.116 min; M+1=180.8
Step 4. 3-Bromo-5,6-dihydro-4H-pyrrolo[1,2-b]pyrazole-2-carboxylic acid ethyl ester
[00361] A solution of 5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazole-2-carboxylic acid ethyl ester (80.0 mg, 0.444 mmol) in tetrahydrofuran (1.00 ml_) was treated with N-bromosuccinimide (86.9 mg, 0.488 mmol) at room temperature for 20 min. The reaction mixture was evaporated, diluted with dichloromethane (30.0 mL), washed with sat. Na2SO3 solution (5.00 ml_) and brine (5.00 mL), dried over Na2SO4, and evaporated to give the title compound as a yellow oil (100.0 mg, purity 95.0%, yield 80.0%). LC-MS (5-
100-7 method): Rt=5.871 ; M+1 =258.6
Step 5. (3-Bromo-5, 6-dihydro-4H-pyrrolo[1,2-b]pyrazol-2-yl)-methanol
[00362] A solution of 3-bromo-5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazole-2-carboxylic acid ethyl ester
(80.0 mg, 0.309 mmol) in tetrahydrofuran (1.00 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran(1.54 mL, 1.54 mmol) at room temperature for 20 min. The reaction mixture was quenched with MeOH (2.00 mL) and evaporated. The residue was diluted with brine (30.0 mL) and then
30% H2O2 (3.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a brown solid (52.0 mg, purity 95.0%, yield 78.0%). LC-MS (5-100-7 method): Rt=4.514min;
M+1 =216.7
Step 6. (3-bromo-5, 6-dihydro-4H-pyrrolo[1,2-b]pyrazol-2-yl)methanol
[00363] A solution of (3-bromo-5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazol-2-yl)-methanol (50.0 mg,
0.230 mmol) in methylene chloride (0.500 mL) was treated with thionyl chloride (0.0605 mL, 0.829 mmol).
The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water
(50.0 mL), adjusted to pH 7.0, extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 mLx3). The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/ EtOAc, 1 :3) to give the title compound as a brown solid (40.0 mg, purity 95.0%, yield 70.0%). LC-MS (5-100-7 method):Rt=6.249 min; M+1 =236.6
Step 7. [2-Amino-7-(3-bromo-5, 6-dihydro-4H-pyrrolo[1,2-b]pyrazol-2-ylmethyl)-4-chloro-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (Ll)
[00364] A mixture of (3-bromo-5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazol-2-yl)methanol (40.0 mg,
0.170 mmol) in dimethyl sulfoxide (0.50 mL) was treated with sodium hydride (4.28 mg, 0.178 mmol) at 0
°C for 10 minutes and then brought to room temperature. A solution of 5-[2-(fert-butyl-dimethyl- silanyloxy)-1 ,1 -dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (53.5 mg, 0.170 mmol) in dimethyl sulfoxide (1.50 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water
(1.00 mL, 6.00 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (41.5 mg, purity 99.0%, yield 61.1%). 1H NMR (DMSO-d6): δ ppm 6.29 (br. s, 2H), 4.73 (t, 1 H), 4.44 (d, 1 H), 4.35 (d,
1H), 4.07 (t, 2H), 3.66 (m, 1H), 3.45 (m, 2H), 3.21 (m, 2H), 2.78 (t, 1 H), 2.54 (m, 2H). LC-MS (5-100-7 method):Rt=4.621 min; M+1 =400.5
Example 48
[2-Amino-4-chloro-7-(4)5, 6, 7-tetrahydro-pyrazolo[1,5-a]pyridin-2-ylmethyl)-6) 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LII)
Step 1. i-Nitroso-piperidine-2-carboxylic acid
[00365] A mixture of piperidine-2-carboxylic acid (11.2 g, 86.8 mmol) and sodium nitrite (8.39 g,
122 mmol) in water (20.0 ml_) was treated with 12 M of hydrogen chloride in water (10.1 ml_, 122 mmol) at
0 0C for 2 hours and then at room temperature for another for 2 hours. The reaction mixture was extracted with fert-butyl methyl ether (50.0 ml_x5). The organic layer was washed with brine (15.0 ml_x2), dried over Na2SO4, and evaporated to give the title compound as a white solid (10.4 g, purity 95.0%, yield
75.7%). LC-MS (5-100-7 method): Rt=4.219 min; M+1=158.8
Step 2. 3-Oxo-4,5, 6,7-tetrahydro-3H-[1,2,3]oxadiazolo[3,4-a]pyridin-8-ium-3a-ide
[00366] A mixture of i-nitroso-piperidine-2-carboxylic acid (10.4 g, 65.9 mmol) in tetrahydrofuran
(20.0 mL) was treated with trifluoroacetic anhydride (14.0 mL, 98.9 mmol) under nitrogen at 0 0C and then at room temperature for 2 hours. The reaction mixture was evaporated. The residue was purified by flash chromatography (dichlororπethane/MeOH, 100:5 to 100:10) to give the title compound as a brown oil
(4.50 g, purity 95.0%, yield 49.0%). LC-MS (5-100-7 method):Rt=4.336 min; M+1=140.9
Step 3. 4,5,6, 7-Tetrahydro-pyrazolo[1,5-a]pyridine-2-carboxylic acid ethyl ester
[00367] A mixture of 3-oxo-4,5,6,7-tetrahydro-3H-[1 ,2,3]oxadiazolo[3,4-a]pyridin-8-ium-3a-ide
(1.00 g, 7.14 mmol) in 1 ,2-diethoxyethane (5.00 mL) was purged with nitrogen three times and heated to
125 0C. Ethyl propiolate (1.04 mL, 10.3 mmol) was added dropwise over 10 minutes. The reaction was stirred at 125 0C for 3 hours and evaporated. The residue was purified by purified by reverse-phase preparative HPLC (30 to 70% CH3CN/H2O, 0.1% TFA) to give the title compound as a brown oil (280.0 mg, purity 99.0%, yield 20.0%). LC-MS (5-100-7 method):Rt=5.474 min; M+1 =194.8
Step 4. (4, 5, 6, 7-Tetrahydro-pyrazolo[1 , 5-a]pyridin-2-yl)-methanol
[00368] A solution of 4,5,6,7-tetrahydro-pyrazolo[1,5-a]pyridine-2-carboxylic acid ethyl ester
(140.0 mg, 0.721 mmol) in tetrahydrofuran (2.00 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (2.52 mL, 2.52 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (2.00 mL) and evaporated. The residue was diluted with brine (30.0 mL) and then
30% H2O2 (3.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/isopropanol (100:5, 50.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a colorless oil (110.0 mg, purity 99.0%, yield 100%).
Step 5. 2-Chloromethyl-4,5,6, 7-tetrahydro-pyrazolo[1,5-a]pyridine
[00369] A solution of (4,5,6,7-tetrahydro-pyrazolo[1 ,5-a]pyridin-2-yl)-methanol (110.0 mg, 0.723 mmol) in methylene chloride (0.500 mL) was treated with thionyl chloride (0.211 mL, 2.89 mmol). The reaction mixture was stirred at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 mLx3).
The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/ EtOAc, 1 :3) to give the title compound as a pale yellow oil (95.0 mg, purity
99.0%, yield 77.0%). LC-MS (5-100-7 method): Rt=5.548 min; M+1=170.8. Step 6. [2-Amino-4-chloro-7-(4,5,6, 7-tetrahydro-pyrazolo[1,5-a]pyridin-2-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl]-methanol (LII)
[00370] A solution of 2-chloromethyl-4,5,6,7-tetrahydro-pyrazolo[1 ,5-a]pyridine (32.5 mg, 0.190 mmol) in dimethyl sulfoxide (1.00 ml_) was treated with sodium hydride (4.80 mg, 0.200 mmol) at 0 0C for
10 minutes and then brought to room temperature. 5-[2-(tert-butyl-dimethyl-silanyloxy)-1 ,1-dimethyl-ethyl]-
4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (60.0 mg, 0.190 mmol) in dimethyl sulfoxide
(1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (0.476 mL, 2.86 mmol) for
15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to
50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (32.3 mg, purity 99.0%, yield
50.6%). 1H NMR (CDCI3): δ ppm 5.83 (s, 2H), 5.05 (br. s, 2H), 4.53 (d, 1 H), 4.36 (d, 1 H), 4.05 (t, 2H),
3.80 (m, 2H), 3.58 (m, 2H), 3.35 (m, 1 H), 2.72 (t, 1H), 1.97 (m, 2H), 1.80 (m, 2H). LC-MS (5-100-7 method):Rt=4.310 min; M+1=334.6.
Example 49
[2-Ammo-7-(3-bromo-4,S,6J^etrahydro-pyrazolo[1,S-a]pyridm-2-ylmethyl)-4-chloro-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LIII)
Step 1. 3-Bromo-4,5,6, 7-tetrahydro-pyrazolo[1,5-a]pyridine-2-carboxylic acid ethyl ester
[00371] A solution of 4,5,6,7-tetrahydro-pyrazolo[1 ,5-a]pyridine-2-carboxylic acid ethyl ester
(140.0 mg, 0.721 mmol) in tetrahydrofuran (1.00 mL) was treated with N-bromosuccinimide (86.9 mg,
0.488 mmol) at room temperature for 20 minutes. The reaction mixture was evaporated, diluted with dichloromethane (30.0 mL), washed with sat. Na2SO3 solution (5.00 mL) and brine (5.00 mL), dried over
Na2SO4, and evaporated to give the title compound as a yellow oil (140.0 mg, purity 95.0%, yield 68.0%).
LC-MS (5-100-7 method):Rt=6.281 min; M+1 =272.6
Step 2. (3-Bromo-4, 5, 6, 7-tetrahydro-pyrazolo[1 , 5-a]pyridin-2-yl)-methanol
[00372] A solution of 3-bromo-4,5,6,7-tetrahydro-pyrazolo[1 ,5-a]pyridin-2-carboxylic acid ethyl ester (140.0 mg, 0.721 mmol) in tetrahydrofuran (2.00 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (1.28 mL, 1.28 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (2.00 mL) and evaporated. The residue was diluted with brine
(30.0 mL) and then 30% H2O2 (3.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/ isopropanol (100:5, 30.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a colorless oil (107.0 mg, purity 95.0%, yield 90.3%). LC-MS (5-100-7 method):Rt=4.806 min; M+1 =230.7
Step 3. 3-Bromo-2-chloromethyl-4,5,6,7-tetrahydro-pyrazolo[1,5-a]pyridine
[00373] A solution of (3-bromo-4,5,6,7-tetrahydro-pyrazolo[1 ,5-a]pyridin-2-yl)-methanol (107 mg,
0.463 mmol) in methylene chloride (0.500 mL) was treated with thionyl chloride (0.118 mL, 1.62 mmol).
The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water
(50.0 mL), adjusted to pH 7.0, and extracted with dichloromethane/isopropanol (100:5, 20.0 ml_x3). The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/ EtOAc=I :3) to give the title compound as a brown solid (98.0 mg, purity 95.0%, yield 84.8%). LC-MS (5-100-7 method):Rt=6.619 min; M+1 =250.6
Step 4. [2-Amino-7-(3-bromo-4,5,6,7-tetrahydro-pyrazolo[1,5-a]pyridin-2-ylmethyl)-4-chloro-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LIII)
[00374] A mixture of 3-bromo-2-chloromethyl-4,5,6,7-tetrahydro-pyrazolo[1 ,5-a]pyridine (47.5 mg,
0.190 mmol) in dimethyl sulfoxide (1.00 mL) was treated with sodium hydride (4.80 mg, 0.200 mmol at 0 0C for 10 minutes and then brought to room temperature. A solution of 5-[2-(ferf-butyl-dimethyl- silanyloxy)-1 , 1 -dimethyl-ethylH-chloro-ej-dihydro-δH-pyrrolo^.S-dlpyrimidin^-ylamine (60.0 mg, 0.190 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (0.476 mL, 2.86 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse- phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (46.2 mg, purity 99.0%, yield 58.6%). 1H NMR (CDCI3): δ ppm 5.06 (br. s, 2H), 4.66 (d, 1 H), 4.36 (d, 1 H), 4.05 (t, 2H), 3.85 (m, 1 H), 3.76 (m, 1 H), 3.66-3.55 (m, 2H), 3.36 (m, 1 H), 2.65 (t, 1 H), 1.98 (m, 2H), 1.87 (m, 2H). LC-MS (5-100-7 method):Rt=4.886 min; M+1=414.5 Example 50
[Σ-Amino^-chloro-T^S-chloro-Sjδ-dihydro^H-pyrrolop^-bJpyrazol-Σ-ylmethylf-β^-dihydro-SH- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LIV)
Step 1. 3-Chloro-5,6-dihydro-4H-pyrrolo[1,2-b]pyrazole-2-carboxylic acid ethyl ester [00375] A solution of 5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazole-2-carboxylic acid ethyl ester (100.0 mg, 0.555 mmol) in tetrahydrofuran (1.00 mL) was treated with N-chlorosuccinimide (81.5 mg, 0.610 mmol) at room temperature for 60 minutes. The reaction mixture was evaporated, diluted with dichloromethane (3OmL), washed with sat. Na2SO3 solution (5.00 mL) and brine (5.00 mL), dried over Na2SO4, and evaporated. The residue was purified by reverse-phase preparative HPLC (5 to 60% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (64.0 mg, purity 95.0%, yield 53.7%). LC-MS (5-100-7 method):Rt=5.766 min; M+1 =214.7 Step 2. (3-Chloro-5, 6-dihydro-4H-pyrrolo[1,2-b]pyrazol-2-yl)-methanol
[00376] A solution of 3-chloro-5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazole-2-carboxylic acid ethyl ester
(64.0 mg, 0.298 mmol) in tetrahydrofuran (1.00 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (1.04 mL, 1.04 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (2.00 mL) and evaporated. The residue was diluted with brine (30.0 mL) and then 30% H2O2 (3.00 mL) was added dropwise. The mixture was extracted with dichloromethane/isopropanol (100:5, 30.0 mLx3), dried over Na2SO4, and evaporated to afford the title compound as a white solid (43.0 mg, purity 95.0%, yield 83.5%). LC-MS (5-100-7 method) :Rt=4.364 min; M+1 =182.8 Step 3. 3-Chloro-2-chloromethyl-5, 6-dihydro-4H-pyrrolo[1,2-b]pyrazole [00377] A solution of (3-chloro-5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazol-2-yl)-methanol (43.0 mg, 0.249 mmol) in methylene chloride (0.500 mL) was treated with thionyl chloride (0.0454 ml_, 0.623 mmol). The reaction mixture was stirred at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 mLx3). The organic layer was dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane: EtOAc=I :3) to give the title compound as a pale yellow oil (19.0 mg, purity 95.0%, yield 39.9%). LC-MS (5-100-7 method) :Rt=6.127 min; M+1=190.7 Step 4: [2-Amino-4-chloro-7-(3-chloro-5, 6-dihydro-4H-pyrrolo[1,2-b]pyrazol-2-ylmethyl)-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LIV)
[00378] A mixture of 3-chloro-2-chloromethyl-5,6-dihydro-4H-pyrrolo[1 ,2-b]pyrazole (19.0 mg,
0.0994 mmol) in dimethyl sulfoxide (0.500 mL) was treated with sodium hydride (2.50 mg, 0.104 mmol) at 0 0C for 10 minutes and then brought to room temperature. A solution of 5-[2-(fert-butyl-dimethyl- silanyloxy)-1 ,1-dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (31.3 mg, 0.0994 mmol) in dimethyl sulfoxide (0.500 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (0.249 mL, 1.49 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse- phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (10.8 mg, purity 99.0%, yield 30.6%). 1H NMR (CDCI3): δ ppm 4.53 (dd, 2H), 4.14 (t, 2H), 3.86 (m, 1 H), 3.66-3.51 (m, 3H), 3.34 (m, 1 H), 2.90 (t, 2H), 2.65 (m, 2H). LC-MS (5-100-7 method):Rt=4.554 min; M+1 =354.6 Example 51
{2-Amino-4-chloro-7-[1-(4-fluoro-phenyl)-ethyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}- methanol (LV)
[00379] A mixture of 1 -(1-bromo-ethyl)-4-fluoro-benzene (38.7 mg, 0.190 mmol) in dimethyl sulfoxide (0.500 mL) was treated with sodium hydride (4.80 mg, 0.200 mmol) at 0 0C for 10 minutes and then brought to room temperature. A solution of 5-[2-(fe/t-butyl-dimethyl-silanyloxy)-1 ,1-dimethyl-ethyl]-4- chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (60.0 mg, 0.190 mmol) in dimethyl sulfoxide (0.500 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (0.500 mL, 3.00 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 60% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (26.0 mg, purity 99.0%, yield 42.3%). 1H NMR (CDCI3): δ ppm 7.32 (m, 1 H), 7.09 (m, 1 H)1 6.99 (m, 2H), 5.49 (m, 1 H), 4.94 (br. s, 2H), 3.85 (m, 2H), 3.60 (m, 1 H), 3.34 (m, 2H), 1.60 (d, 2H). LC-MS (5-100-7 method): Rt=4.966 min; M+1 =322.6 Example 52
{2-Amino-4-chloro-7-[5-ethyl- 1 -(4-methoxy-benzyl)-4-methyl- 1 H-pyrazol-3-ylmethyl]-6, 7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (L Vl) Step 1. Ethyl S-ethyl^-methyl-IH-pyrazoleS-carboxylate [00380] (Z)-ethyl 2-hydroxy-3-methyl-4-oxohex-2-enoate (3.46 g, 18.6 mmol) was treated with hydrazine (0.583 mL, 18.6 mmol) at O 0C and then was stirred at room temperature overnight. The resulting solid was collected and dissolved in dichloromethane (50.0 mL), washed with water (20.0 ml_x2), and evaporated to give the title compound as a yellow solid (2.90 g, purity ≥ 90%, yield 85.6%).
LC-MS (5-100-7 method):Rt=5.625min; M+1=182.8
Step 2. 5-Ethyl-1-(4-methoxy-benzyl)-4-methyl-1H-pyrazole-3-carboxylic acid ethyl ester and 5-
Ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazole-3-carboxylic acid ethyl ester
[00381] A mixture of ethyl δ-ethyM-methyl-I H-pyrazole-S-carboxylate (2.80 g, 15.4 mmol), and potassium carbonate (4.25 g, 30.7 mmol) in N,N-dimethylformamide (12.0 mL) was treated with p- methoxybenzyl chloride (2.50 mL, 18.4 mmol) at 0 0C and then the mixture was stirred overnight at room temperature. The mixture was diluted with sat. NH4CI solution (300 mL), extracted with dichloromethane
(50.0 mLx3), dried over Na2SO4, and concentrated. The residue was purified by flash chromatography
(hexane/ EtOAc, 8:1 to 2:1 ) to yield 5-ethyl-1-(4-methoxy-benzyl)-4-methyl-1 H-pyrazole-3-carboxylic acid ethyl ester as a colorless oil (2.30 g, purity > 90%, yield 50.0%). %). LC-MS (5-100-7 method): Rt=7.710 min; M+1 =302.7
[00382] The chromatography also gave 5-ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazole-3- carboxylic acid ethyl ester as a colorless oil (1.40 g, purity > 90%, yield 30.0%). LC-MS (5-100-7 method):Rt=7.127 min; M+1=302.7
Step 3. [5-Ethyl-1-(4-methoxy-benzyl)-4-methyl-1H-pyrazol-3-yl]-methanol
[00383] A solution of 5-ethyl-1-(4-methoxy-benzyl)-4-methyl-1 H-pyrazole-3-carboxylic acid ethyl ester (1.00 g, 3.31 mmol) in tetrahydrofuran (2.00 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (8.27 mL, 8.27 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (2.00 mL) and evaporated. The residue was diluted with brine
(30.0 mL) and then 30% H2O2 (3.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/ isopropanol (100:5, 30.0 mLx3), dried over Na2SO4, and evaporated. The residue was purified by reverse-phase preparative HPLC (5 to 60% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (0.43 g, purity 99.0%, yield 50.0%). LC-MS (5-100-7 method): Rt=5.207 min;
M+1 =260.8
Step 4. 3-Chloromethyl-5-ethyl-1-(4-methoxy-benzyl)-4-methyl-1H-pyrazole
[00384] A solution of [5-ethyl-1-(4-methoxy-benzyl)-4-methyl-1 H-pyrazol-3-yl]-methanol (0.400 g,
1.54 mmol) in methylene chloride (4.00 mL) was treated with thionyl chloride (0.280 mL, 3.84 mmol. The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water
(5OmL), adjusted to pH 7.0, extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 mLx3).
The organic layer was dried over Na2SO4, and evaporated to give the title compound as a pale yellow oil
(0.42 g, purity 99.0%, yield 98.0%). LC-MS (5-100-7 method):Rt=7.091 min; M+1 =278.7
Step 5. {2-Amino-4-chloro-7-[5-ethyl-1-(4-methoxy-benzyl)-4-methyl-1H-pyrazol-3-ylιnethyl]-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (L Vl) [00385] A mixture of 3-chloromethyl-5-ethyl-1-(4-methoxy-benzyl)-4-methyl-1 H-pyrazole (88.5 mg, 0.318 mmol) in dimethyl sulfoxide (0.50 ml.) was treated with sodium hydride (8.00 mg, 0.333 mmol) at 0 °C for 10 minutes and then brought to room temperature. 5-[2-(fert-butyl-dimethyl-silanyloxy)-1 ,1 - dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (100.0 mg, 0.318 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes.
The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (0.794 mL, 4.76 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (94.0 mg, purity 99.0%, yield 67.0%). 1H NMR (DMSO-d6): δ ppm 6.90 (d, 2H), 6.77 (d, 2H), 6.50 (br. s, 2H), 5.18
(d, 1 H), 5.09 (d, 1H), 4.77 (t, 1 H), 4.45 (s, 2H), 3.70 (s, 3H), 3.56 (m, 1 H), 3.18 -2.96 (m, 4H), 2.50 (q,
2H), 1.96 (s, 3H), 1.15 (t, 3H). LC-MS (5-100-7 method):Rt=5.051 min; M+1=442.6
Example 53
{2-Amino-4-chloro-7-[5-ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazol-3-ylmethyl]-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (L VII)
Step 1. [5-Ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazol-3-yl]-methanol
[00386] A solution of 5-ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazole-3-carboxylic acid ethyl ester (1.00 g, 3.31 mmol) in tetrahydrofuran (2.00 mL) was treated with 1.0 M of lithium triethylborohydride in tetrahydrofuran (8.27 mL, 8.27 mmol) at room temperature for 20 minutes. The reaction mixture was quenched with MeOH (2.00 mL) and evaporated. The residue was diluted with brine
(30.0 mL) and then 30% H2O2 (3.00 mL) was added dropwise. The mixture was extracted with a mixture of dichloromethane/ isopropanol (100:5, 30.0 mLx3), dried over Na2SO4, and evaporated. The residue was purified by reverse-phase preparative HPLC (5 to 60% CH3CN/H2O, 0.1 % TFA) to give the title compound as a white solid (0.46 g, purity ≥ 90%, yield 53.0%). LC-MS (5-100-7 method): Rt=5.461 min;
M+1 =260.8
Step 2. 5-Chloromethyl-3-ethyl-1-(4-methoxy-benzyl)-4-methyl-1 H-pyrazole
[00387] A solution of [5-ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazol-3-yl]-methanol (0.400 g,
1.54 mmol) in methylene chloride (4.00 mL) was treated with thionyl chloride (0.280 mL, 3.84 mmol) at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 ml_x3). The organic layer was dried over Na2SO4, and evaporated to give the title compound as a pale yellow oil (0.42 g, purity ≥ 90%, yield
98.0%). LC-MS (5-100-7 method): Rt=7.187 min; M+1=278.7
Step 3. {2-Amino-4-chloro-7-[5-ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazol-3-ylmethyl]-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (L VII)
[00388] A solution of 5-chloromethyl-3-ethyl-1 -(4-methoxy-benzyl)-4-methyl-1 H-pyrazole (88.5 mg, 0.318 mmol) in dimethyl sulfoxide (0.500 mL) was treated with sodium hydride (8.00 mg, 0.333 mmol) at 0 0C for 10 minutes and then brought to room temperature. A solution of 5-[2-(tert-butyl-dimethyl- silanyloxy)-1,1-dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (100.0 mg, 0.318 mmol) in dimethyl sulfoxide (1.00 ml_) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (0.794 mL, 4.76 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse- phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (96.0 mg, purity 99.0%, yield 68.0%). 1H NMR (DMSOd6): δ ppm 7.04 (d, 2H), 6.87 (d, 2H), 6.44 (br. s, 2H), 5.17 (d, 1 H), 5.09 (d, 1 H), 4.83 (t, 1 H), 4.44 (s, 2H), 4.35 (d, 1H), 3.71 (s, 3H), 3.66 (m, 1 H), 3.46 - 3.18 (m, 4H), 2.50 (q, 2H), 1.87 (s, 3H), 0.92 (t, 3H). LC-MS (5-100-7 method):Rt=5.569 min; M+1=442.6 Example 54
[Σ-Amino-A-chloro-T-iδ-ethyl-A-methyl-ΣH-pyrazol-S-ylmethyiyβJ-dihydro-SH-pyrrolofcS- d]pyrimidin-5-yl]-methanol (L VIII)
[00389] A mixture of {2-amino-4-chloro-7-[5-ethyl-2-(4-methoxy-benzyl)-4-methyl-2H-pyrazol-3- ylmethyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (50.0 mg, 0.113 mmol), trifluoroacetic acid (1.00 mL, 13.0 mmol) and 12 M of hydrogen chloride in water (1.00 mL, 12.0 mmol) was heated to 130 0C for 3 hours. The mixture was diluted with ice -water (2OmL), adjusted to pH 7.0, filtered, and purified by reverse-phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (10.0 mg, purity 99.0%, yield 27.4%). 1H NMR (acetone-d6): δ ppm 5.81 (br. s, 2H), 4.54 (d, 1 H), 4,43 (d, 1H), 3.82 (m, 1 H), 3.62-3.50 (m, 3H), 3.30 (m, 1 H), 2.60 (q, 2H), 1.94 (s, 3H), 1.20 (t, 3H). LC-MS (5-100-7 method):Rt=4.006 min; M+1 =322.7 Example 55
[2-Amino-7^2-bromo^,5,6-trimethyl-pyridin-3-ylmethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-methanol (LIX) Step 1. (Z)-4-Amino-3-methyl-pent-3-en-2-one
[00390] A mixture of, 3-methyl-2,4-pentanedione (25.5 mL, 219 mmol), silica gel (1.20 g, 15.3 mmol) and 17.0 M of ammonia in water (14.2 mL, 241 mmol) was stirred at room temperature overnight, whereupon the mixture solidified to give a mass of pale yellow needles. The reaction mixture was triturated in hexanes (50.0 mL) and filtered. The solid was taken up in MeOH (500 mL) and heated until dissolved, filtered, and evaporated to give the title compound as a white solid (15.0 g, purity ≥ 90%, yield 54.5%). 1H NMR (DMSOd6): δ ppm 10.21 (br. s, 1 H), 7.15 (br. s, 1 H), 2.02 (s, 3H), 1.94 (s, 3H), 1.77 (s, 1 H).
Step 2. 4,5,6-Trimethyl-2-oxo-1,2-dihydro-pyridine-3-carbonitrile
[00391] A solution of malononitrile (3.85 g, 58.3 mmol) in anhydrous tetrahydrofuran (5.00 mL) was slowly added dropwise to (Z)-4-Amino-3-methyl-pent-3-en-2-one (6.60 g, 58.3 mmol) in anhydrous tetrahydrofuran (10.0 mL) at 0 0C. The mixture was stirred at room temperature until a precipitate appeared. The solid was separated by filtration and crystallized from ethanol to give the title compound as a white solid (7.00 g, purity > 90%, yield 66.0%). LC-MS (5-100-7 method):Rt=4.271 min; M+1=162.8 Step 3. 2-Bromo-4,5,6-trimethyl-nicotinonitrile [00392] A mixture 4,5,6-trimethyl-2-oxo-1,2-dihydro-pyridine-3-carbonitrile (4.50 g, 27.7 mmol), phosphorus pentoxide (8.27 g, 29.1 mmol) and tetra-N-butylammonium bromide (10.7 g, 33.3 mmol) in anhydrous toluene (70.0 mt_) was heated to reflux for 4 hours. The mixture was cooled to room temperature and ice-cold water (10.0 ml_) was added carefully with stirring and cooling. The mixture was stirred for 2 hours and then filtered through a pad of celite, and the layers were separated. The toluene layer was washed with brine (30.0 ml. x2) and evaporated. The crude was crystallized with a mixture of dichloromethane/ MeOH (9:1 , 200 mL) to give the title compound as a white solid (4.20 g, purity 99.0%, yield 66.0%). LC-MS (5-100-7 method) :Rt=6.395 min; M+1 =224.7 Step 4. 2-Bromo-4,5,6-trimethyl-pyridine-3-carbaldehyde
[00393] A solution of 2-bromo-4,5,6-trimethyl-nicotinonitrile (3.00 g, 13.3 mmol) in toluene (20.0 mL) was treated with 1.0 M of diisobutylaluminum hydride in toluene (15.0 mL, 15.0 mmol) at -70 °C for 2 hours. The reaction mixture was brought to room temperature and was carefully quenched with 2M sulfuric acid (35.0 mL). The resulting mixture was stirred overnight at room temperature and diluted with water (30.0 mL). The organic layer was washed with water (20.0 mL) and evaporated to give the title compound as a pale yellow solid (2.60 g, purity 95.0%, yield 81.0%). LC-MS (5-100-7 method): Rt=6.336 min; M+1 =227.7
Step 5. (2-Bromo-4,5,6-trimethyl-pyridin-3-yl)-methanol
[00394] A solution of 2-bromo-4,5,6-trimethyl-pyridine-3-carbaldehyde (2.10 g, 9.21 mmol) in ethanol (50.0 mL) was treated with sodium tetrahydroborate (1.74 g, 46.0 mmol) potionwise at 10 °C. After addition, the mixture was stirred at room temperature for 1 hour, quenched with water (5.0 mL). The reaction mixture was evaporated. The residue was dissolved in dichloromethane (15OmL). The organic layer was washed with water (5OmL) and evaporated. The crude was purified by flash chromatography (dichloromethane/MeOH, 10:1 ) to give the title compound as a white solid (1.4O g, purity 95.0%, yield 62.8%). LC-MS (5-100-7 method):Rt=4.245 min; M+1 =229.7 Step 6. 2-Bromo-3-chloromethyl-4,5,6-trimethyl-pyridine
[00395] A solution of (2-bromo-4,5,6-trimethyl-pyridin-3-yl)-methanol (200.0 mg, 0.869 mmol) in anhydrous methylene chloride (2.00 mL) was treated with thionyl chloride (0.158 mL, 2.17 mmol). The reaction mixture was stirred at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (95:5, 20.0 mLx3). The organic layer was dried over Na2SO4 and evaporated to give the title compound as a white solid (170.0 mg, purity 95.0%, yield 75.0%). LC-MS (5-100-7 method):Rt=6.857 min; M+1=249.6 Step 7. [2-Amino-7-(2-bromo-4,5, 6-trimethyl-pyridin-3-ylmethyl)-4-chloro-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LIX)
[00396] A mixture of 2-bromo-3-chloromethyl-4,5,6-trimethyl-pyridine (170.0 mg, 0.684 mmol) in dimethyl sulfoxide (1.00 mL) was treated with sodium hydride (17.2 mg, 0.718 mmol) at 0 0C for 10 minutes and then brought to room temperature. 5-[2-(fe/t-butyl-dimethyl-silanyloxy)-1 ,1-dimethyl-ethyl]-4- chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (215 mg, 0.684 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (2.28 mL, 13.7 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a pale yellow solid (126.0 mg, purity 95.0%, yield 44.6%). 1H NMR (DMSO-d6): δ ppm 6.52 (br. s, 2H), 4.80 (m, 2H), 4.51 (d, 1 H), 3.57 (m, 1 H), 3.28 (m, 1 H), 3.18 (m, 3H)1 2.43 (s, 3H), 2.25 (s, 3H), 2.16 (s, 3H). LC-MS (5-100-7 method): Rt=4.702 min; M+1=413.5. Example 56
3-(2-Amino-4-chloro-5-hydroxymethyl-5, 6-dihydro-pyrrolo[2, 3-d]pyrimidin-7-ylmethyl)-4, 5, 6- trimethyl-pyridine-2-carbonitrile (LX)
[00397] A mixture of [2-amino-7-(2-bromo-4,5,6-trimethyl-pyridin-3-ylmethyl)-4-chloro-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (60.0 mg, 0.145 mmol) and copper cyanide (13.0 mg, 0.145 mmol) in dimethyl sulfoxide (3.00 mL, 42.3 mmol) was subjected to microwave irradiation at 100 0C for 60 minutes. After cooling, the resulting reaction mixture was diluted with water (10.0 mL), purified by reverse- phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a brown solid (10.0 mg, purity 99.0%, yield 20.0%). 1H NMR (DMSO-d6): δ ppm 6.56 (br. s, 2H), 4.84 (m, 3H), 4.48 (d, 2H), 3.57 (m, 2H), 3.25-3.18 (m, 3H), 2.54 (s, 3H), 2.17 (s, 3H), 2.08 (s, 3H). LC-MS (5-100-7 method):Rt=4.674 min; M+1 =358.6 Example 57
3-(2-Amino-4-chloro-5-hydroxymethyl-5,6-dihydrθ'pyrrolo[2,3-d]pyrimidin-7'ylmethyl)-4,5,6- trimethyl-pyridine-2-carboxylic acid amide (LXI)
[00398] A mixture of 3-(2-amino-4-chloro-5-hydroxymethyl-5,6-dihydro-pyrrolo[2,3-d]pyrimidin-7- ylmethyl)-4,5,6-trimethyl-pyridine-2-carbonitrile (7.00 mg, 0.0200 mmol), dimethyl sulfoxide (0.500 mL, 7.00 mmol), 2 M of sodium hydroxide in water (19.5 μL, 0.0390 mmol), 30% hydrogen peroxide (9.96 μL, 0.0975 mmol) and ethanol (0.50 mL, 8.60 mmol) was stirred at room temperature for 20 minutes. The reaction mixture was quenched with sat. Na2S2O3 solution (2.00 mL), adjusted to pH 3.0, and filtered. The filtrate was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (3.0 mg, purity 99.0%, yield 41.0%). 1H NMR (DMSO-d6): δ ppm 8.07 (br. s, 1 H), 7.67 (br. s, 1 H), 4.89 (dd, 2H), 3.65-3.18 (m, 5H), (d, 2H), 2.54 (s, 3H), 2.22 (s, 3H), 2.18 (s, 3H). LC-MS (5-100-7 method): Rt=3.725 min; M+1=376.6 Example 58
/2-4/n/no-4-c/i/oro-7-<4,5,6-fr//nef/iy/-pyr/d/n-3-y//nef/iy/>6,7-d//jydro-5H- pyrrolo[2,3-d]pyrimidin-5- yl]-methanol (LXII)
Step 1. (4,5,6-Trimethyl-pyridin-3-yl)-methanol
[00399] A mixture of (2-bromo-4,5,6-trimethyl-pyridin-3-yl)-methanol (160.0 mg, 0.6953 mmol),
5% Palladium on CaCO3 (70.0 mg, 0.0329 mmol) in ethanol (10.0 mL) was stirred overnight under hydrogen (balloon). The catalyst was removed by filtration through a pad of celite, and the filtrate was evaporated to give the title compound as a white solid (100.0 mg, purity 95.0%, yield 90.0%). LC-MS (5- 100-7 method): Rt=3.258 min; M+1=151.9
Step 2. 5-Chloromethyl-2,3,4-trimethyl-pyridine
[00400] A solution of (4,5,6-trimethyl-pyridin-3-yl)-methanol (100.0 mg, 0.661 mmol) in anhydrous methylene chloride (2.00 mL) was treated with thionyl chloride (0.241 ml_, 3.31 mmol). The reaction mixture was stirred at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (95:5, 20.0 ml_x3). The organic layer was dried over Na2SO4 and evaporated to give the title compound as a yellow oil (100.0 mg, purity 95.0%, yield 80.0%). LC-MS (5-100-7 method):Rt=3.966 min; M+1 =169.8
Step 3. β-Amino^-chloro-T-ft&β-trimethyl-pyridin-S-ylmethylJ-βJ-dihydro-SH-pyrrolofcS- d]pyrimidin-5-yl]-methanol (LXII)
[00401] A mixture of 5-chloromethyl-2,3,4-trimethyl-pyridine (102 mg, 0.603 mmol) in dimethyl sulfoxide (1.00 mL) was treated with sodium hydride (17.2 mg, 0.718 mmol) at 0 0C for 10 minutes and then brought to room temperature. 5-[2-(fert-butyl-dimethyl-silanyloxy)-1 ,1-dimethyl-ethyl]-4-chloro-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (190.0 mg, 0.603 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes. The mixture was diluted with water (2.00 mL) and treated with 6 M of hydrogen chloride in water (2.28 mL, 13.7 mmol) for 15 minutes, diluted with water (30.0 mL), filtered, and purified by reverse-phase preparative HPLC (5 to 50%
CH3CN/H2O, 0.1% TFA) to give the title compound as a pale brown solid (103.0 mg, purity 98.0%, yield
51.1%). 1H NMR (DMSOd6): δ ppm 8.08 (s, 1 H), 6.48 (br. s, 2H), 4.85 (t, 1 H), 4.65 (d, 2H), 4.33 (d, 1 H),
3.60 (m, 1 H), 3.28-3.07 (m, 4H), 3.18 (m, 3H), 2.43 (s, 3H), 2.18 (s, 3H), 2.17 (s, 3H). LC-MS (5-100-7 method): Rt=3.531 min; M+1 =333.6
Example 59
3-[3-(2-Amino-4-chloro-5-hydroxymethyl-5,6-dihydro-pyrrolo[2,3-d]pyrimidin-7-ylmethyl)-4,5,6- trimethyl-pyridin-2-yl]-propan-1 -ol (LXIII)
Step 1. 2-[3-(4-Methoxy-benzyloxy)-prop-1-ynyl]-4,5,6-trimethyl-nicotinonitrile
[00402] To a mixture of 2-bromo-4,5,6-trimethyl-nicotinonitrile (400.0 mg, 1.777 mmol), potassium carbonate (614.0 mg, 4.443 mmol), copper(l) iodide (13.54 mg, 71.08 μmol), triphenylphosphine (37.2 mg, 0.142 mmol) in N,N-dimethylformamide (3.44 mL) and water (3.40 mL) was added 20% Pd(OH)2 on carbon (20.0 mg, 0.0285 mmol). The mixture was degassed by bubbling nitrogen at room temperature for
30 minutes. 1-methoxy-4-((prop-2-ynyloxy)methyl)benzene (0.7829 g, 4.443 mmol) was added to the mixture and heated to 80 0C overnight under nitrogen. The reaction mixture was filtered through a pad of celite, washed with dichloromethane (15.0 mL) and water (5.00 mL). The combined filtrates were washed with brine (5.00 mLx3). The organic layer was dried over Na2SO4, filtered, and evaporated. The residue was purified by flash chromatography (hexane/EtOAc, 2:1) to give the title compound as a white solid
(450.0 mg, purity 99.0%, yield 78.0%). LC-MS (5-100-7 method) :Rt=7.198 min; M+1=320.7
Step 2. 2-[3-(4-Methoxy-benzyloxy)-propyl]-4, 5, 6-trimethyl-nicotinonitrile
[00403] Hydrogen gas (60 psi) was conducted to a mixture of 2-[3-(4-methoxy-benzyloxy)-prop-1- ynyl]-4,5,6-trimethyl-nicotinonitrile (450 mg, 1.4 mmol) and 10% palladium on carbon (60.0 mg, 0.0564 mmol) in methanol (13.9 ml.) for 7 hours. After removal of the catalyst by filtration through a pad of celite, the filtrate was evaporated to give the title compound as a white solid (410.0 mg, purity 95.0%, yield
85.0%). LC-MS (5-100-7 method):Rt=5.780 min; M+1=324.7
Step 3. 2-[3-(4-Methoxy-benzyloxy)-propyl]-4, 5, β-trimethyl-pyridine-3-carbaldehyde
[00404] A solution of 2-[3-(4-methoxy-benzyloxy)-propyl]-4,5,6-trimethyl-nicotinonitrile (410 mg,
1.3 mmol) in toluene (5.00 mL) was treated with 1.0 M of diisobutylaluminum hydride in toluene (1.42 ml_,
1.42 mmol) at -70 °C for 2 hours. The reaction mixture was brought to room temperature and was carefully quenched with 2M sulfuric acid (35.0 mL). The resulting mixture was stirred overnight at room temperature and diluted with toluene (50.0 mL). The organic layer was washed with water (20.0 mL) and evaporated to give the title compound as a pale yellow solid (215.0 mg, purity ≥ 90%, yield 47.0%). LC-
MS (5-100-7 method):Rt=5.403 min; M+1 =327.7
Step 4. {2-[3-(4-Methoxy-benzyloxy)-propyl]-4, 5, 6-trimethyl-pyridin-3-yl}-methanol
[00405] A solution of 2-[3-(4-methoxy-benzyloxy)-propyl]-4,5,6-trimethyl-pyridine-3-carbaldehyde
(150 mg, 0.46 mmol) in ethanol (2.14 mL) was treated with sodium tetrahydroborate (86.7 mg, 2.29 mmol) portionwise at 10 0C. After addition, the mixture was stirred at room temperature for 1 hour, quenched with water (5.00 mL), and evaporated. The residue was dissolved in dichloromethane (15.0 mL). The organic layer was washed with water (5.00 mL) and evaporated. The crude was purified by flash chromatography (dichloromethane/MeOH, 10:1) to give the title compound as a white solid 84.0 mg, purity > 90%, yield 56.0%). LC-MS (5-100-7 method): Rt=5.120 min; M+1=329.7
Step 5. 3-Chloromethyl-2-[3-(4-methoxy-benzyloxy)-propyl]-4,5, 6-trimethyl-pyridine
[00406] A solution of {2-[3-(4-methoxy-benzyloxy)-propyl]-4,5,6-trimethyl-pyridin-3-yl}-methanol
(80.0 mg, 0.243 mmol) in anhydrous methylene chloride (2.00 mL) was treated with thionyl chloride
(0.241 mL, 3.31 mmol). The reaction mixture was stirred at room temperature for 20 minutes, evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, and extracted with a mixture of dichloromethane/isopropanol (95:5, 20.0 mLx3). The organic layer was dried over Na2SO4 and evaporated to give the title compound as a yellow oil (55.0 mg, purity ≥ 90%, yield 65.0%). LC-MS (5-100-
7 method):Rt=6.068 min; M+1 =347.7
Step 6. 3-[3-(2-Amino-4-chloro-5-hydroxymethyl-5, 6-dihydro-pyrrolo[2, 3-d]pyrimidin-7-ylmethyl)-
4,5,6-trimethyl-pyridin-2-yl]-propan-1 -ol (LXIII)
[00407] A mixture of 3-chloromethyl-2-[3-(4-methoxy-benzyloxy)-propyl]-4,5,6-trimethyl-pyridine
(55.0 mg, 0.158 mmol) in dimethyl sulfoxide (1.00 mL) was treated with sodium hydride (4.17 mg, 0.174 mmol) at 0 0C for 10 minutes and then brought to room temperature. 5-[2-(fert-butyl-dimethyl-silanyloxy)-
1 ,1-dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (49.8 mg, 0.158 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 20 minutes.
The mixture was diluted with water (2.00 mL) and then treated with 6 M of hydrogen chloride in water
(0.264 mL, 1.58 mmol) and trifluoroacetic acid (1.00 mL, 13.0 mmol) at 80 0C for 1 hour. After cooling, the mixture was diluted with water (30.0 ml_), filtered, and purified by reverse-phase preparative HPLC (5 to 50% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (27.9 mg, purity 99.0%, yield 45.0%). 1H NMR (CDCI3): δ ppm 5.71 (br. s, 2H), 4.92 (d, 2H), 4.60 (d, 1 H), 3.74 (m, 1 H), 3.64 (t, 2H), 3.58 (t, 2H), 3.55-3.31 (m, 4H), 2.83 (s, 3H), 2.52 (s, 3H), 2.37 (s, 3H), 1.95 (m, 2H). LC-MS (5-100-7 method):Rt=3.627 min; M+1=391.6 Example 60
7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3-yl)methyl)-5-((tert-bυtyldimethylsilyloxy)methyl)-4- chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (LXIV) Step 1. 2-bromo-3-(chloromethyl)-5-ethyl-4,6-dimethylpyridine
[00408] A solution of (2-bromo-5-ethyl-4,6-dimethyl-pyridin-3-yl)-methanol (1.0 g, 6 mmol) in anhydrous dichloromethane (8 mL, 100 mmol) was cautiously treated with thionyl chloride (0.6 mL, 8.5 mmol).The reaction mixture was stirred at room temperature for 20 minutes before being evaporated to dryness. The residue was diluted with water (5 mL), then adjusted to pH 7.0 with aq. NaHCO3 before extracting with dichloromethane (3X10 mL). The combined organic layers were dried over anhydrous Na2SO4, then evaporated to give the title compound as a viscous cloudy colorless oil. 1H NMR (CDCI3): δ ppm 4.80 (s, 1 H), 2.66 (q, 2H), 2.55 (s, 3H), 2.46 (s, 3H), 1.15 (t, 3H); LC/MS : rt (5-100-5 method) = 3.286 min.; 261.6 (M+1 , 77.4 %), 263.6 (M+3, 100%), 265.6 (M+5, 24.1%).
Step 2. 7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3-yl)methyl)-5-((tert-butyldimethylsilyloxy)methyl)- 4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (LXIV)
[00409] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-
2-ylamine (1.0 g, 3.182 mmol) was dissolved in anhydrous DMF (10 mL, 100 mmol), cooled to 0 0C1 and treated with NaH (125 mg, 5.21 mmol). To the reaction mixture, 2-bromo-3-(chloromethyl)-5-ethyl-4,6- dimethylpyridine (0.98 g, 3.7 mmol) in 1 mL DMF was added. The reaction was left stirring at rt for 20 additional minutes. It was then extracted with EtOAc and water. The organic layer was dried and the solvent was removed to give a white solid (0.98 g, purity >90%, yield 51.4%). 1H NMR (DMSO-cfe) δ ppm 6.56 (br. s., 2H), 4.83 (d, 1H), 4.46 (d, 1 H), 3.60 (m, 1 H), 3.41 (m, 2H), 3.25 (m., 1 H), 3.15 (m, 1 H), 2.61 (m, 2H), 2.44 (s., 3H), 2.28 (s., 3H), 1.06 (t, 3H), 0.82 (s, 9H), -0.08 (d, 3H), -0.16 (d, 3H). LC/MS: rt (5- 100-7 method) = 7.749 min.; 539.5 (M+1 , 77.4 %), 541.5 (M+3, 100%), 543.5 (M+5, 32.4%) Example 61
(2-amino-4-chloro-7-((5-ethyl-4,6-dimethyl-2-(1H-pyrazol-4-yl)pyridin-3-yl)methyl)-6,7-dihydro-SH- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXV)
[00410] The compound was obtained by reacting 7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3- yl)methyl)-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (102 mg, 0.19 mmol) with tert-butyl 4-(4,4,5,5-tetramethyl-1 ,3,2-dioxaborolan-2-yl)-1H-pyrazole-1- carboxylate (105 mg, 0.35 mmol), using the general procedure for Suzuki reaction and TBS deprotection. A white solid was collected (16 mg, purity >90%, yield 18%). 1H NMR (CDCI3) δ ppm 7.69 (s., 2H), 5.22 (s, 2H), 4.64 (m, 2H), 3.68 (d, 2H), 3.22 (m, 3H), 2.71 (m, 2H), 2.61 (s, 3H), 2.30 (s, 3H), 1.16 (t, 3H). LC/MS : rt (5-100-7 method) = 3.805 min.; 413.6 (M+1, 100 %), 415.6 (M+3, 35%).
Example 62
(2-amlno-4-chloro-7^(S^thyl^,6-dimethyl-2^3,3,3-trifluoroprop-1-en-2-yl)pyridin-3-yl)methyl)-6,7- dihydro-5H'pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXVI)
[00411] The title compound was obtained by reacting 7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3- yl)methyl)-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
(102 mg, 0.19 mmol) with dibutyl 3,3,3-trifluoroprop-1-en-2-ylboronate (90 mg, 0.36 mmol), using the general procedure for Suzuki reaction and TBS deprotection. A white solid was collected (2.7 mg, purity
>90%, yield 3%). 1H NMR (CDCI3) δ ppm 6.24 (d, 1 H), 5.61 (s., 1H), 4.88 (s., 2H), 4.81 (d., 2H), 4.50 (d,
1 H), 3.75 (m, 2H), 3.34 (m, 2H), 3.21 (d, 1 H), 2.71 (m, 2H), 2.60 (s, 3H), 2.23 (s, 3H), 1.17 (t, 3H). LC/MS
: rt (5-100-5 method) = 2.057 min.; 442.0 (M+1 , 100 %), 263.6 (M+3, 32.4 %).
Example 63
(2-amino-4-chloro-7-((5-ethyl-2-(1-ethyl-1H-pyrazol-4-yl)-4,6-dimethylpyridin-3-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXVII)
[00412] The title compound was obtained by reacting 7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3- yl)methyl)-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
(102 mg, 0.19 mmol) with 1-ethyl-4-(4,4,5,5-tetramethyl-1 ,3,2-dioxaborolan-2-yl)-1 H-pyrazole (79 mg,
0.35 mmol), using the general procedure for Suzuki reaction and TBS deprotection. A white solid was collected (40 mg, purity >90%, yield 40%). 1H NMR (CDCI3) δ ppm 7.70 (s, 2H), 4.89 (s, 2H), 4.82 (d, 1 H),
4.68 (d, 1 H), 4.22 (m, 2H), 3.71 (m, 2H),3.29 (m, 2H), 3.18 (m, 1 H), 2.71 (m, 2H), 2.60 (s, 3H), 2.27 (s,
3H), 1.52 (t, 3H), 1.16 (t, 3H). LC/MS : rt (5-100-5method) = 1.872 min.; 442.1 (M+1 , 100 %), 414.1 (M+3,
Example 64
2-amino-4-chloro-7-((5-ethyl-2-(isoxazol-4-yl)-4,6-dimethylpyridin-3-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXVIII)
[00413] The title compound was obtained by reacting 7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3- yl)methyl)-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
(50 mg, 0.09 mmol) with 4-(4,4,5,5-tetramethyl-1 ,3,2-dioxaborolan-2-yl)isoxazole (28 mg, 0.14 mmol), using the general procedure for Suzuki reaction and TBS deprotection. A white solid was collected (1.13 mg, purity >90%, yield 3%). 1H NMR (CDCI3) δ ppm 9.07 (s, 1 H), 5.32 (s, 1 H), 5.14 (q, 2H), 4.84 (s, 2H),
3.80 (m, 1 H), 3.6 (q, 2H), 3.48 (dd, 1 H), 3.40 (m, 1 H), 2.67 (q, 2H), 2.60 (s, 3H), 2.04 (s, 3H), 1.13 (t, 3H).
LC/MS : rt (5-100-5 method) = 2.133 min.; 415.1 (M+1 , 100 %), 417.1 (M+3, 35%)
Example 65
(2-amino-4-chloro-7-((5-ethyl-4,6-dimethyl-2-(1H-pyrazol-5-yl)pyridin-3-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXIX)
[00414] The title compound was obtained by reacting 7-((2-bromo-5-ethyl-4,6-dimethylpyridin-3- yl)methyl)-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (50 mg, 0.09 mmol) with 1 H-pyrazol-5-ylboronic acid (16 mg, 0.14 mmol), using the general procedure for
Suzuki reaction and TBS deprotection. A white solid was collected (3.07 mg, purity >90%, yield 7%). 1H
NMR (CDCI3) δ ppm 7.60 (d, 1 H), 6.60 (d, 1 H), 5.06 (s, 2H), 4.85(dd, 2H), 3.71 (d, 2H), 3.6 (q, 2H), 3.48
(dd, 1 H), 3.40 (m, 1 H), 2.67 (q, 2H), 2.60 (s, 3H), 2.04 (s, 3H), 1.13 (t, 3H). LC/MS : rt (5-100-5method) =
2.133 min.; 415.1 (M+1 , 100 %), 417.1 (M+3, 35%).
Example 66
5-((2-amino-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-5H-pyrrolo[2,3-d]pyrimidin-7(6H)- yl)methyl)-3-methoxy-2,4-dimethylpyridine 1 -oxide (LXX)
Step 1. 5-(chloromethyl)-3-methoxy-2,4-dimethylpyridine 1 -oxide
[00415] 5-Chloromethyl-3-methoxy-2,4-dimethyl-pyridine (500 mg, 3 mmol) and m-CPBA (852 mg, 4.94 mmol) (77% pure) were dissolved in dichloromethane (10 mL, 200 mmol). The reaction was monitored reaction via LC/MS. Excess m-CPBA was added as needed. The reaction mixture was extracted with aq. NaOH (1 N) and dichloromethane. After the organic solvent was removed, the crude product was purified by silica gel column chromatography (100% EtOAc to 10%MeOH/EtOAc) to give an off-white solid. 1H NMR (CDCI3) δ ppm 8.18 (s, 1 H), 4.49 (s, 2H), 3.78 (s, 3H), 2.53 (s, 3H), 2.35 (s, 3H).
LC/MS : rt (5-100-5 method) = 2.174 min.; 202.0 (M+1 , 100 %), 204.0 (M+3, 35%).
Step 2. 5^(2-amino-5-((tert-butyldimethy\si\y\oxy)methy\)-4-ch\oro-5H-pyrro\o[2,3-d]pyr\m\dm-
7(6H)-yl)methyl)-3-methoxy-2,4-dimethylpyridine 1 -oxide (LXX)
[00416] The title compound was obtained by alkylating 5-((tert-butyldimethylsilyloxy)methyl)-4- chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine with 5-(chloromethyl)-3-methoxy-2,4- dimethylpyridine 1 -oxide using the general alkylation procedure. The product precipitated from the reaction mixture (165 mg, purity >90%, yield 48.2%). 1H NMR (CDCI3) δ ppm 8.07 (s, 1H), 4.95 (s, 2H),
4.83 (d, 1H), 4.46 (d, 1 H), 3.60 (m, 1 H), 3.41 (m, 2H), 3.25 (m., 1 H), 3.15 (m, 1 H), 2.61 (m, 2H), 2.44 (s.,
3H), 2.28 (s., 3H), 1.06 (t, 3H), 0.82 (s, 9H), -0.08 (d, 3H), -0.16 (d, 3H). LC/MS: rt (5-100-7 method) =
7.749 min.; 539.5 (M+1 , 77.4 %), 541.5 (M+3, 100%), 543.5 (M+5, 32.4%)
Example 67
S-ifΣ-amino-A-chloro-S^hydroxymethyiySH-pyrrolol∑^-dJpyrimidin^βHyy^methyiys-methoxy-
2,4-dimethylpyridine 1 -oxide (LXXI)
[00417] 5-((2-amino-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-5H-pyrrolo[2,3-d]pyrimidin-
7(6H)-yl)methyl)-3-methoxy-2,4-dimethylpyridine 1 -oxide (80 mg, 0.2 mmol) was dissolved in TFA (2 mL,
10 mmol), dichloromethane (8 mL, 100 mmol) and water (20 mg, 1 mmol). The reaction was left stirring at
60 0C for 30 minutes. The solvent was removed before being extracted with dichloromethane and aqueous saturated bicarbonate. The product was obtained as a glassy solid after a reverse phase HPLC purification. (48 mg, purity >90%, yield 80%). 1H NMR (acetone-d6) δ ppm 8.62 (s, 1 H), 5.02 (m, 2H), 4.03
(t, 1 H), 3.89 (m, 1 H), 3.84 (s, 3H), 3.81 (dd, 1 H), 3.73 (dd, 1 H), 3.57 (m, 1H), 2.61 (s, 3H), 2.47 (s, 3H),
LC/MS: rt (5-100-5 method) = 1.825 min.; 366.0 (M+1 , 100 %), 368.0 (M+3, 35%).
Example 68 Methyl β-ffΣ-amino-S-fftert-butyldimethylsilyloxyfmethyl^-chloroSH-pyrrolofΣ^-dJpyrimidin-
7(6H)-yl)methyl)-4-methoxy-5-methylnicotinate (LXXII)
Step 1. methyl 6-(chloromethyl)-4-methoxy-5-methylnicotinate
[00418] A solution of (methyl 6-(hydroxymethyl)-4-methoxy-5-methylnicotinate (3.45g, 16.3 mmol) in anhydrous dichloromethane (40 ml_, 100 mmol) was cautiously treated with thionyl chloride (1.26 ml_,
17.3 mmol).The reaction mixture was stirred at room temperature for 20 minutes, and evaporated to dryness. The reaction was quenched with aq. NaOH and extracted with dichloromethane. The solid product was purified by silica gel chromatography (1 :1 EtOAc/Hexane) (3.34 g, purity 90%, yield 80%). 1H
NMR (CDCI3) δ ppm 8.76 (s, 1 H), 4.4.70 (s, 2H), 3.95 (s, 3H), 3.91 (s, 3H), 2.40 (s, 3H); LC/MS : rt (5-
100-7 method) = 5.245 min.; 229.7 (M+1 , 100 %), 231.7 (M+3, 35%).
Step 2. Methyl 6-((2-amino-5-((tert-bυtyldimethylsilyloxy)methyl)-4-chloro-5H-pyrrolo[2,3- d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-5-methylnicotinate (LXXII)
[00419] 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- amine (98 mg, 0.31 mmol) was dissolved in anhydrous DMF (3 ml_). The reaction was cooled to 0 0C and treated with NaH (12 mg, 0.5 mmol). To that reaction mixture, methyl 6-(chloromethyl)-4-methoxy-5- methylnicotinate (85 mg, 0.37 mmol) in 1 ml_ DMF was added, and the reaction was stirred at rt for 20 min. The reaction mixture was quenched with water and extracted with EtOAc. The organic solvent was removed to yield a white solid which was then washed with diethyl ether (85 mg, purity 95%, yield 51%).
1H NMR (CDCI3): δ ppm 8.76 (s, 1 H), 4.79 (s, 2H), 4.75 (d, 1 H), 4.64 (d, 1 H), 3.97 (s, 3H), 3.93 (s, 1 H),
3.90 (t, 1 H), 3.66 (m, 1 H), 3.40 (m, 1 H), 3.55 (m, 1 H),3.41 (m, 1 H), 2.28 (s, 3H), LC/MS: rt (5-100-7 method) = 6.973 min.; 507.5 (M+1 , 100 %), 509.5 (M+3, 45%).
Example 69 methyl 6-((2-amino-4-chloro-5-(hydroxymethyl)-5H-pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4- methoxy-5-methylnicotinate (LXXUI)
[00420] A solution of methyl 6-((2-amino-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-5H- pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-5-methylnicotinate (80 mg, 0.2 mmol) in TFA (2 ml_,
10 mmol), dichloromethane (8 mL, 100 mmol), and water (20 mg, 1 mmol) was stirred at 60 0C for 30 minutes. The reaction was quenched with aq. NaOH and extracted with dichloromethane. The organic solvent was removed to yield a white solid which was washed with diethyl ether (56 mg, purity 85%, yield
80%). 1H NMR (CDCI3): δ ppm 8.75 (s, 1 H), 5.05(d, 1 H), 4.92 (s, 2H), 4.39 (d, 1 H), 3.94 (s, 3H), 3.92 (s,
3H), 3.91 (t, 1 H), 3.83 (m, 2H), 3.62 (m, 1 H), 3.45 (m, 1H), 2.28 (s, 3H), LC/MS: rt (5-100-7 method) =
4.282 min.; 393.5 (M+1 , 100 %), 395.5 (M+3, 35%).
Example 70
(β-fβ-amino^-chloro-S^hydroxymethyl^SH-pyrrolofcS-dJpyrimidin^βHΪ-yOmethyl^methoxyS- methylpyridin-3-yl)methanol (LXXIV)
[00421] A solution of ethyl 6-((2-amino-4-chloro-5-(hydroxymethyl)-5H-pyrrolo[2,3-d]pyrimidin-
7(6H)-yl)methyl)-4-methoxy-5-methylnicotinate (15 mg, 0.038 mmol) in anhydrous THF (10 mL) was cooled at O CC and treated with lithium triethylborohydride (40 mg, 0.4 mmol, 1 M in THF). The reaction was quenched with water and extracted with EtOAc. The white solid product was purified by reverse phase HPLC (1.4 mg, purity 90%, yield 9%). 1H NMR (CDCI3): δ ppm 8.34 (s, 1 H), 5.07(d, 1 H), 4.78 (d,
2H), 3.87 (s, 3H), 3.80 (m, 2H), 4.35 (d, 1 H), 3.68 (m, 1H), 3.62 (m, 1 H), 3.45 (m, 1H), 2.24 (s, 3H),
LC/MS: rt (5-100-7 method) = 3.538 min.; 365.6 (M+1 , 100 %), 367.6 (M+3, 35%).
Example 71
[2-Amino-7-(5-bronio-4-methoxy-3-methyl-pyndin-2-ylmethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]ρyrimidin-5-yl]-methanol (LXXV)
Step 1. (5-Bromo-4-methoxy-3-methyl-pyridin-2-yl)-methanol
[00422] Acetic acid 5-bromo-4-methoxy-3-methyl-pyridin-2-ylmethyl ester (100 mg, 0.365 mmol) in tetrahydrofuran (1.5 ml_), methanol (1.0 ml_), and water (0.5 ml.) was let stir with lithium hydroxide, monohydrate (76.5 mg, 1.82 mmol). After 2 hours the reaction was diluted with ethyl acetate and water.
The organic layer was separated, dried over Na2SO4, filtered and condensed to yield 67 mg (79% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.50 (s, 1 H), 4.63 (d, J= 4.3 Hz,
2H), 4.45 (t, J= 4.5 Hz, 1 H), 3.89 (s, 3H), 2.17 (s, 3H). LC-MS (5-100-5 method): Rt = 1.13 min.; m/z =
231.9 (M+1 ).
Step 2. 5-Bromo-2-chloromethyl-4-methoxy-3-methyl-pyridine
[00423] (5-Bromo-4-methoxy-3-methyl-pyridin-2-yl)-methanol (67 mg, 0.29 mmol) in methylene chloride (3.0 mL) was let stir with thionyl chloride (100 μM, 1.4 mmol). After 40 minutes the reaction was condensed in vacuo. The residue was then partitioned between ethyl acetate and sat. aq. NaHCO3. The organic layer was separated, washed with brine, dried over Na2SO4, filtered and condensed to yield 72 mg (100% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.49 (s, 1 H),
4.66 (s, 2H), 3.90 (s, 3H), 2.40 (d, J= 0.5 Hz, 3H). LC-MS (5-100-5 method): Rt = 2.85 min.; m/z = 249.9
(M+1 ).
Step 3. 7-(5-Bromo-4-methoxy-3-methyl-pyridin-2-ylmethyl)-5-(tert-butyl-dimethyl- silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine
[00424] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- ylamine (47.8 mg, 0.152 mmol) in N,N-dimethylformamide (1.1 mL) was treated with sodium hydride (3.9 mg, 0.15 mmol) at 0 0C. 5-Bromo-2-chloromethyl-4-methoxy-3-methyl-pyridine (38 mg, 0.15 mmol) in
N,N-dimethylformamide (1.1 mL) was then added. After 25 minutes the reaction was diluted with water, extracted with ethyl acetate, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography with a gradient of 50-100% ethyl acetate/hexanes.
The title compound was obtained in 69 mg (86 % yield) as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.46 (s, 1 H), 4.86 (s, 2H), 4.71 (d, J= 14.9 Hz, 1 H), 4.53 (d, J= 14.9 Hz, 1 H), 3.84 - 3.90 (m, 4H),
3.46 - 3.64 (m, 3H), 3.32 - 3.43 (m, 1 H), 2.29 (s, 3H), 0.81 (s, 9H), 0.00 (s, 3H), -0.04 (s, 3H). LC-MS (5-
100-5 method): Rt = 3.37 min.; m/z = 528.0 (M+1).
Step 4. [2-Amino-7-(5-bromo-4-methoxy-3-methyl-pyridin-2-ylmethyl)-4-chloro-6) 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LXXV)
[00425] 7-(5-Bromo-4-methoxy-3-methyl-pyridin-2-ylmethyl)-5-(tert-butyl-dimethyl- silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (69 mg, 0.13 mmol) in methanol (100 ml_) was stirred with 1.00 M of hydrogen chloride in water (0.50 mL, 0.50 mmol). After 6 hours the reaction was condensed and the residue partitioned between ethyl acetate and saturated aq. NaHCθ3. The organic layer was separated, washed with brine, dried over Na2SO4, filtered and condensed to yield 57 mg (86% yield, 98.7% HPLC purity) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.44 (s, 1 H), 4.92 (d, J= 16.4 Hz, 1 H), 4.77 (s, 2H), 4.36 (d, J= 16.2 Hz, 1 H), 3.88 (s, 4H), 3.72 - 3.85 (m, 2H), 3.58 (dd, J= 9.9, 3.3 Hz, 1 H), 3.38 - 3.49 (m, 1 H), 3.04 (br. s., 1 H), 2.29 (s, 3H). LC-MS (5-100-5 method): Rt = 2.22 min. (98.7% HPLC purity); m/z = 413.9 (M+1). Example 72
[2-Amino-4-chloro-7-(2,3-dihydro-turo[3,2-c]pyridm' -4-ylmethyl)-6,7-d\hydro-5H-pyrroloi2,3- d]pyrimidin-5-yl]-methanol (LXXVI) Step 1: ethyl fυro[3,2-c]pyridine-4-carboxylate
[00426] 4-chlorofuro[3,2-c]pyridine (800 mg, 5.21 mmol) in ethanol (200 mL) was treated with sodium acetate (856 mg, 10.4 mmol) and nitrogen was bubbled through the solution. The [1 ,1'- bis(diphenylphosphino)ferrocene]dichloropalladium(ll), complex with dichloromethane (1 :1 ) (340 mg, 0.42 mmol) was then added and carbon monoxide was bubbled through the solution. The reaction was then heated at 75 0C under a carbon monoxide balloon. After 6 hours the reaction was filtered through celite and condensed. The residue was then partitioned between ethyl acetate and water, separated, and the organic layer was washed with brine, dried over MgSO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, 50-100% gradient) to yield 908 mg (91% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.67 (d, J= 5.6 Hz, 1 H), 7.79 (d, J= 2.0 Hz, 1 H), 7.64 (d, J= 5.6 Hz, 1 H), 7.45 (d, J= 1.3 Hz, 1 H), 4.56 (q, J= 7.1 Hz, 2H), 1.51 (t, J= 7.1 Hz, 3H). LC-MS (5-100-5 method): Rt = 1.95 min.; m/z = 192.0 (M+1). Step 2. 2,3-Dihydro-furo[3,2-c]pyridine-4-carboxylic acid ethyl ester
[00427] Ethyl f uro[3,2-c]pyridine-4-carboxylate (2.40E2 mg, 1.26 mmo) in ethanol (8.0 mL) was treated with 10% palladium on carbon (1:9, Palladiunrcarbon black, 49 mg, 0.046 mmol) and shaken on the Parr hydrogenator at 75 psi. After 48 hours LC-MS showed some starting material remained. Another 50 mg of Pd/C was added and the reaction was shaken on the Parr hydrogenator at 75 psi. After another 24 hours LC-MS showed the reaction had gone to completion. The reaction was then filtered through celite and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient of 30-80%) to yield 177 mg (73% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.44 (d, J= 5.6 Hz, 1 H), 6.88 (d, J= 5.3 Hz, 1 H), 4.71 (t, J= 8.8 Hz, 2H), 4.47 (q, J= 7.1 Hz, 2H), 3.61 (t, J= 8.8 Hz, 2H), 1.45 (t, J= 7.2 Hz, 3H). LC-MS (5-100-5 method): Rt = 1.49 min. m/z = 194.0 (M+1). Step 3. (2,3-Dihydro-furo[3,2-c]pyridin-4-yl)-methanol [00428] 2,3-Dihydro-furo[3,2-c]pyridine-4-carboxylic acid ethyl ester (79 mg, 0.41 mmol) in tetrahydrofuran (1.8 ml_) at -78 0C was treated with 1.00 M of lithium tetrahydroaluminate in THF (0.409 mL, 0.409 mmol). After 35 minutes the reaction was treated with 0.020 mL of water (while still at -78 °C ). 0.020 mL of 15% NaOH was then added followed by 0.060 mL of water and the reaction was let stir for 15 minutes at room temperature. MgSO4 was then added and the reaction was stirred for an additional 15 minutes. The reaction was then filtered through celite and condensed to yield 69 mg (56% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.27 (d, J= 5.6 Hz, 1 H), 6.71 (d, J= 5.6 Hz, 1H), 4.70 (t, J= 8.8 Hz, 2H), 4.65 (s, 2H), 3.16 (t, J= 8.8 Hz, 2H). LC-MS (5-100-5 method): Rt = 0.26 min.; m/z = 152.0 (M+1).
Step 4: 4-Chloromethyl-2,3-dihydro-furo[3,2-c]pyridine
[00429] (2,3-Dihydro-furo[3,2-c]pyridin-4-yl)-methanol (69 mg, 0.46 mmol) was stirred in methylene chloride (3.3 mL) and thionyl chloride (170 μM, 2.3 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 1 hour the reaction was condensed. The residue was then partitioned between ethyl acetate and sat. aq. NaHCO3. The layers were separated and the organics were dried over Na2SO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient of 30-100%) to provide 21 mg (27% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.27 (d, J= 5.6 Hz, 1 H), 6.72 (d, J= 5.6 Hz, 1 H), 4.70 (t, J= 8.8 Hz, 2H), 4.61 (s, 2H), 3.33 (t, J= 8.8 Hz, 2H). LC-MS (5-100-5 method): Rt = 0.27 min.; m/z = 170.0 (M+1 ).
Step 5. S-ftert-Butyl-dimethyl-silanyloxymethyl^-chloro-T-fΣ^-dihydro-furo^Σ-cJpyridin^- ylmethyl)-6, 7-dihydro-5H-pyrrolo[2, 3-d]pyrimidin-2-ylamine
[00430] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-
2-ylamine (44.6 mg, 0.142 mmol) in dimethyl sulfoxide (0.95 mL) was treated with sodium hydride (3.6 mg, 0.14 mmol). 4-chloromethyl-2,3-dihydro-furo[3,2-c]pyridine (24 mg, 0.14 mmol) in dimethyl sulfoxide (0.95 mL) was then added. After 1 hour the reaction still contained starting materials by LC-MS. An additional 1 mg of NaH was added. After 1.5 hours, LC-MS indicated all of the 4-chloromethyl-2,3- dihydro-furo[3,2-c]pyridine had been consumed. After 2 hours the reaction was diluted with water and extracted with ethyl acetate. The organic layer was then washed with brine, dried over Na2SO4, filtered, and condensed. The crude material was then purified on by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient of 30-100%) to yield 40.0 mg (60% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.25 (d, J= 5.6 Hz, 1 H), 6.69 (d, J= 5.6 Hz, 1H), 4.90 (br. s., 2H), 4.52 - 4.71 (m, 4H), 3.88 (dd, J= 10.0, 3.7 Hz, 1 H), 3.51 - 3.69 (m, 3H), 3.31 - 3.45 (m, 1H), 3.16 (t, J= 8.8 Hz, 2H), 0.81 (s, 9H), -0.03 (s, 3H). LC-MS (5-100-5 method): Rt = 2.71 min.; m/z = 448.1 (M+1 ). Step 6. [2-Amino-4-chloro-7-(2, 3-dihydro-furo[3,2-c]pyridin-4-ylmethyl)-6, 7-dihydro-5H-pyrrolo[2, 3- d]pyrimidin-5-yl]-methanol (LXXVI)
[00431] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-(2,3-dihydro-furo[3,2-c]pyridin-4- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (40.0 mg, 0.0893 mmol) in methanol (6.8 mL) was stirred with 1.00 M of Hydrogen chloride in water (0.34 ml_, 0.34 mmol). After 6 hours the reaction was condensed and the residue partitioned between ethyl acetate and sat. aq. NaHCO3. The organic layer was separated and the aqueous layer was extracted 2 more times with ethyl acetate. The combined organics were then washed with brine, dried over Na2SO4, filtered, and condensed to yield 31 mg (100% yield) of the title compound as a white solid. 1H NMR (400 MHz, MeOD) δ ppm 8.16 (d, J= 5.6 Hz, 1 H), 6.77 (d, J= 5.6 Hz, 2H), 4.75 (d, J= 15.4 Hz, 1 H), 4.60 - 4.69 (m, 2H), 4.52 (d, J= 15.2 Hz, 1 H), 3.78 (dd, J= 10.9, 3.5 Hz, 1 H), 3.57 - 3.70 (m, 2H), 3.50 (dd, J= 10.0, 3.9 Hz, 1 H), 3.39 (dt, J= 9.5, 3.6 Hz, 1 H), 3.22 (m, 2H). LC-MS (5-100-5 method): Rt = 0.72 min. (96.9% HPLC purity); m/z = 334.0 (M+1 ). Example 73
(2-Amino-4-chloro-7-fυro[3,2-c]pyridin-4-ylmethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)- methanol (LXXVII)
Step 1. 4-Chloromethyl-furo[3,2-c]pyridine
[00432] 2,3-Dihydro-furo[3,2-c]pyridine-4-carboxylic acid ethyl ester (1.00E2 mg, 0.523 mmol) in tetrahydrofuran (2.4 mL) at -78 0C was treated with 1.00 M of lithium tetrahydroaluminate in THF (0.523 mL, 0.523 mmol). After 1 hour the reaction was treated with 0.020 mL of water (while still at -78 0C ). 0.020 mL of 15% NaOH was then added followed by 0.060 mL of water and the reaction was let stir for 15 minutes at room temperature. MgSO4 was then added and the reaction was stirred for an additional 15 minutes. The reaction was then filtered through celite and condensed. The crude alcohol (78 mg, 0.52 mmol) was then stirred in methylene chloride (3.8 mL) and thionyl chloride (190 μM, 2.6 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 30 minutes the reaction was condensed. The residue was then partitioned between ethyl acetate and sat. aq. NaHCO3. The organic layer was separated and washed with brine, dried over Na2SO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient of 50-100%) to yield 54 mg (62% yield over 2 steps) of an oil. 1H NMR (400 MHz, CDCI3) δ ppm 8.45 (d, J= 5.8 Hz, 1 H), 7.72 (d, J= 2.3 Hz, 1 H), 7.44 (dd, J= 5.6, 1.0 Hz, 1 H), 7.02 (dd, J= 2.3, 1.0 Hz, 1 H), 4.92 (s, 2H). LC-MS (5-100-5 method): Rt = 0.98-1.2 min. (broad); m/z = 168.0 (M+1 ). Step 2. (2-Amino-4-chloro-7-furo[3,2-c]pyridin-4-ylmethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin- 5-yl)-methanol (LXXVII)
[00433] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- ylamine (50.2 mg, 0.159 mmol) in dimethyl sulfoxide (1.1 mL) was treated with sodium hydride (4.0 mg, 0.16 mmol). 4-Chloromethyl-furo[3,2-c]pyridine (26.7 mg, 0.159 mmol) in dimethyl sulfoxide (1.1 mL) was then added. After 50 minutes an additional 2 mg of NaH was added. After 2 hours an additional 1 mg of NaH was added. After 2.5 hours the reaction was diluted with water and extracted with ethyl acetate. The organics were then washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by reverse phase-preparative HPLC (mobile phase MeCN/water, TFA buffer) to yield 26.2 mg (48% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3 -fCD3OD) δ ppm 8.55 (d, J= 6.3 Hz, 1 H), 7.92 (d, J= 2.3 Hz, 1 H), 7.77 (d, J= 6.1 Hz, 1 H), 7.23 (d, J= 1.5 Hz, 1 H), 5.11 - 5.23 (m, 2H), 3.81 - 3.90 (m, 1 H), 3.71 - 3.79 (m, 2H), 3.61 - 3.69 (m, 1H), 3.39 (dd, J= 8.5, 4.4 Hz, 1 H). LC-MS (5-100-5 method): Rt = 1.08 min. (97.6 % HPLC purity); m/z = 332.0 (M+1). Example 74
(2-Amino-4-chloro-7-pyrrolo[1,2-a]pyrazin-1-ylmethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)- methanol (LXXVUI)
Step 1. Pyrrolo[1,2-a]pyrazine-1-carboxylic acid ethyl ester.
[00434] 1-Chloro-pyrrolo[1 ,2-a]pyrazine (200 mg, 1.31 mmol) in ethanol (5 mL) was treated with sodium acetate (215 mg, 2.62 mmol) and nitrogen was bubbled through the solution. The [1 ,1'- bis(diphenylphosphino)ferrocene]dichloropalladium(ll),complex with dichloromethane (1 :1) (86 mg, 0.10 mmol) was then added and CO was bubbled through the solution. The reaction was then heated at 75 0C and let stir under a CO balloon. After 2 hours and 25 minutes the reaction was filtered through celite, rinsing the filter cake with ethanol and methanol, and condensed. The residue was then partitioned between ethyl acetate and water, separated, and the organic layer was washed with water and brine, dried over Na2SO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient from 50-100%) to provide 158 mg (63% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 7.97 (dd, J= 4.5, 1.0 Hz, 1 H), 7.68 (d, J= 4.5 Hz, 1 H), 7.55 (dd, J= 2.5, 1.3 Hz, 1 H), 7.45 (dt, J= 4.2, 1.0 Hz, 1 H), 7.03 (dd, J= 4.3, 2.5 Hz, 1 H), 4.54 (q, J= 7.2 Hz, 2H), 1.49 (t, J= 7.2 Hz, 3H). LC-MS (5-100-5 method): Rt = 1.90 min.; m/z = 191.1 (M+1). Step 2. 1-Chloromethyl-pyrrolo[1,2-a]pyrazine
[00435] Pyrrolo[1 ,2-a]pyrazine-1-carboxylic acid ethyl ester (116 mg, 0.610 mmol) in tetrahydrofuran (2.8 mL) at -78 0C was treated with 1.00 M of lithium tetrahydroaluminate in THF (0.610 mL, 0.610 mmol). After 35 minutes the reaction was treated with 0.020 mL of water (while still at -78 0C ). 0.020 mL of 15% NaOH was then added followed by 0.060 mL of water and the reaction was let stir for 15 minutes at room temperature. MgSO4 was then added and the reaction was stirred for an additional 15 minutes. The reaction was then filtered through celite and condensed. The crude pyrrolo[1 ,2-a]pyrazin-1- yl-methanol (90 mg, 0.3 mmol was stirred in methylene chloride (4.4 mL) and thionyl chloride (220 μM, 3.0 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 1 hour the reaction was condensed. The residue was then partitioned between ethyl acetate and sat. Aq. NaHCO3). The layers were separated and the organics were dried over Na2SO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: methanol/dichloromethane, gradient of 0-10%) to provide 66 mg (66% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 7.79 (d, J= 4.8 Hz1 1 H), 7.50 (d, J= 4.8 Hz, 1 H), 7.47 (dd, J= 2.1 , 1.4 Hz, 1 H), 6.88 - 6.97 (m, 2H), 4.82 (s, 2H). LC-MS (5-100-5 method): Rt = 0.27 min.; m/z = 170.0 (M+1).
Step 3. 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-pyrrolo[1,2-a]pyrazin-1-ylmethyl-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine
[00436] δ-Oert-Butyl-dimethyl-silanyloxymethyO^-chloro-ej-dihydro-δH-pyrrolo^.S-djpyrimidin^- ylamine (39.7 mg, 0.126 mmol) in dimethyl sulfoxide (1.0 mL) was treated with sodium hydride (4.14 mg, 0.164 mmol). 1-chloromethyl-pyrrolo[1 ,2-a]pyrazine (21 mg, 0.13 mmol) in dimethyl sulfoxide (1.0 mL) was then added. After 16 hours an additional 2 mg of NaH was added. After 20 hours the reaction was diluted with water and extracted with ethyl acetate. The organic layer was washed with brine, dried over Na2SO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient of 30-100%) to yield 31 mg (60% purity, 53% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 7.76 (dd, J= 4.8, 0.8 Hz, 1H), 7.45 (d, J= 4.8 Hz, 1 H), 7.42 (dd, J= 2.5, 1.3 Hz, 1 H), 6.88 (dt, J= 4.1 , 1.1 Hz, 1 H), 6.83 (dd, J= 4.0, 2.5 Hz, 1 H), 4.87 - 5.04 (m, 4H), 4.71 - 4.81 (m, 1H), 3.86 (dd, J= 9.7, 3.7 Hz, 1 H), 3.58 - 3.63 (m, 1 H), 3.51 - 3.58 (m, 1 H), 3.41 - 3.45 (m, 1 H), 3.31 - 3.39 (m, 1 H), 0.77 (s, 9H), -0.06 (s, 3H), -0.11 (s, 3H). LC-MS (5-100-5 method): Rt = 2.86 min.; m/z = 445.1 (M+1).
Step 4. (2-Amino-4-chloro-7-pyrrolo[1,2-a]pyrazin-1-ylmethyl-6, 7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-methanol (LXXVIII)
[00437] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-pyrrolo[1 ,2-a]pyrazin-1-ylmethyl-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (31 mg, 0.069 mmol) in methanol (5.3 mL) was stirred with 1.00 M of hydrogen chloride in water (0.554 mL, 0.554 mmol). After 16 hours the reaction was condensed and the residue purified by reverse phase-preparative HPLC (mobile phase: MeCN/water, TFA buffer) to provide 9 mg (40% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 7.77 (d, J= 5.1 Hz, 1 H), 7.45 (s, 1 H), 7.42 (d, J= 4.8 Hz, 1 H), 6.86 (s, 1 H), 5.16 (d, J= 16.4 Hz, 1 H), 4.96 (br. s., 2H), 4.65 (d, J= 16.4 Hz, 1 H), 4.07 (t, J= 6.7 Hz, 1H), 3.77 - 3.88 (m, 2H), 3.66 (dd, J= 9.9, 3.3 Hz, 1 H), 3.38 - 3.48 (m, 1 H). LC-MS (5-100-5 method): Rt = 1.46 min. (98.55% HPLC purity); m/z = 331.0 (M+1 ). Example 75
(2-Amino-4-chloro-7-furo[3,2-b]pyridin-6-ylmethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)- methanol (LXXIX)
Step 1. Furo[3,2-b]pyridin-6-yl-methanol
[00438] Furo[3,-b]pyridin-6-carboxaldehyde (2.00E2 mg, 1.36 mmol) in tetrahydrofuran (2.0 mL) at 0 °C was treated with Sodium tetrahydroborate (51.4 mg, 1.36 mmol) and let stir under a nitrogen balloon. After 15 minutes the reaction was diluted with ethyl acetate, washed with water and brine, dried over Na2SO4, filtered, and condensed to provide 204 mg (100% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.53 (s, 1H), 7.76 - 7.90 (m, 2H), 6.97 (d, J= 2.0 Hz, 1 H), 4.86 (s, 2H), 2.20 (br. s., 1 H). LC-MS (5-100-5 method): Rt = 0.26 min.; m/z = 150.0 (M+1). Step 2. 6-Chloromethyl-furo[3,2-b]pyridine
[00439] Furo[3,2-b]pyridin-6-yl-methanol (204 mg, 1.37 mmol) was stirred in methylene chloride
(4.9 mL) and thionyl chloride (250 μM, 3.4 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 1 hour the reaction was condensed. The residue was then partitioned between ethyl acetate and sat.aq. NaHCO3. The layers were separated and the organics were dried over Na2SO4, filtered, and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, gradient of 30-100%) to yield 174 mg (76% yield) of the title compound. 1H
NMR (400 MHz, CDCI3), δ ppm 8.58 (s, 1 H), 7.89 (d, J= 2.3 Hz, 1H), 7.84 (s, 1H), 7.00 (d, J= 1.3 Hz, 1 H),
4.75 (s, 2H). LC-MS (5-100-5 method): Rt = 1.79 min.; m/z = 168.0 (M+1).
Step 3. 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-furo[3,2-b]pyridin-6-ylmethyl-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine
[00440] δ-Oert-Butyl-dimethyl-silanyloxymethylJ^-chloro-ΘJ-dihydro-SH-pyrrolo^.S-dlpyrimidin^- ylamine (54.8 mg, 0.174 mmol) in dimethyl sulfoxide (1.4 ml.) was treated with the sodium hydride (4.84 mg, 0.191 mmol). 6-Chloromethyl-furo[3,2-b]pyridine (35 mg, 0.21 mmol) in dimethyl sulfoxide (1.4 ml_) was then added. After 45 minutes LC-MS indicated still some starting materials left, therefore, an additional 2 mg of NaH was added. After 1.5 hours the reaction was diluted with water, extracted with ethyl acetate, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (mobile phase: ethyl acetate/hexanes, 30-100% gradient) to provide 77 mg (99% yield) of the title compound as a white solid. LC-MS (5-100-5 method): Rt = 2.81 min.; m/z = 446.1 (M+1 ).
Step 4. (2-Arnino-4-chloro-7-furo[3,2-b]pyridin-6-ylmethyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-
5-yl)-methanol (LXXIX)
[00441] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-furo[3,2-b]pyridin-6-ylmethyl-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (77 mg, 0.17 mmol) in methanol (100 mL) was stirred with
1.00 M of Hydrogen chloride in water (1.0 mL, 1.0 mmol) and the reaction was heated at 55 0C. After 1 hour the reaction was diluted with sat. aq. NaHCO3 and extracted twice with ethyl acetate. The combined organics were then dried over Na2SO4, filtered and condensed to yield 34 mg (53% yield) of the title compound. 1H NMR (400 MHz, DMSO-d6) δ ppm 8.50 (d, J= 1.8 Hz, 1 H), 8.30 (d, J= 2.3 Hz, 1 H), 7.94
(s, 1 H), 7.13 (dd, J= 2.3, 1.0 Hz, 1 H), 6.51 (s, 2H), 4.87 (t, J= 5.3 Hz, 1 H), 4.71 - 4.79 (m, 1 H), 4.55 - 4.62
(m, 1 H), 3.62 (dt, J= 10.5, 4.4 Hz, 1 H), 3.50 - 3.58 (m, 1 H), 3.44 - 3.49 (m, 1 H), 3.36 - 3.41 (m, 1 H), 3.20 -
3.28 (m, 1H). LC-MS (5-100-5 method): Rt = 1.57 min. (90.2% HPLC purity); m/z = 332.0 (M+1).
Example 76
[2-Amino-4-chloro-7-(2,3-dihydro-furo[3,2-b]pyridin-6-ylmethyl)-6,7-dihydro-5H-pyrroto[2,3- d]pyrimidin-5-yl]-methanol (LXXX)
Step 1: (2,3-Dihydro-furo[3,2-b]pyridin-6-yl)-methanol
[00442] Furo[3,-b]pyridin-6-carboxaldehyde (400 mg, 2.72 mmol) in ethanol (10 mL) was treated with 10% Palladium on Carbon (1 :9, Palladiumxarbon black, 145 mg, 0.136 mmol) and shaken on the
Parr hydrogenator at 75 psi. After 24 hours the compound was filtered through celite and condensed to provide 332 mg of the title compound (70% purity, 56% yield). 1H NMR (400 MHz, CDCI3), δ ppm 7.97 (s,
1 H), 7.06 (d, J= 1.3 Hz, 7H), 4.62 - 4.71 (m, 4H), 3.30 (t, J= 9.0 Hz, 2H).
Step 2. 6-Chloromethyl-2,3-dihydro-furo[3,2-b]pyridine
[00443] (2,3-Dihydro-furo[3,2-b]pyridin-6-yl)-methanol (209 mg, 1.38 mmol) was stirred in methylene chloride (5.0 mL) and thionyl chloride (151 μM, 2.07 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 1 hour the reaction was condensed. The residue was then partitioned between ethyl acetate and sat. aq. NaHCO3. The layers were separated and the organics were dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (30-100% gradient of ethyl acetate/hexanes) to provide 89 mg (38% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.05 (d, J= 1.5 Hz, 1 H), 7.08 (d, J= 1.8 Hz, 1 H), 4.71 (t, J=
8.8 Hz, 2H), 4.56 (s, 2H), 3.35 (t, J= 8.8 Hz, 2H). LC-MS (5-100-5 method): Rt = 1.68 min. (not uv active); m/z = 170.0 (M+1).
Step 3. 5-{tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-(2,3-dihydro-furo[3,2-b]pyridin-6- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine
[00444] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- ylamine (74.3 mg, 0.236 mmol) in dimethyl sulfoxide (1.9 mL) was treated with sodium hydride (6.55 mg,
0.259 mmol). 6-Chloromethyl-2,3-dihydro-furo[3,2-b]pyridine (52 mg, 0.31 mmol) in dimethyl sulfoxide
(1.9 mL) was then added. After 2 hours LC-MS indicated still some starting materials left. Therefore an additional 3.4 mg of NaH was added. After 24 hours the reaction was diluted with water, extracted with ethyl acetate, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (30-100% ethyl acetate/hexanes gradient) to yield 88 mg (83% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 7.97 (d, J= 1.5 Hz, 1 H),
6.94 (d, J= 1.5 Hz, 1 H), 4.93 (br. s., 2H), 4.67 (t, J= 8.8 Hz, 2H), 4.41 - 4.57 (m, 2H), 3.88 (dd, J= 9.9, 3.8
Hz, 1 H), 3.58 (dd, J= 10.0, 8.0 Hz, 1 H), 3.45 - 3.53 (m, 2H), 3.26 - 3.42 (m, 3H), 0.79 - 0.88 (m, 9H), 0.03
(s, 3H), 0.00 (s, 3H). LC-MS (5-100-5 method): Rt = 2.86 min.; m/z = 448.1 (M+1).
Step 4. [2-Amino-4-chloro-7-(2, 3-dihydro-furo[3,2-b]pyridin-6-ylmethyl)-6, 7-dihydro-5H-pyrrolo[2, 3- d]pyrimidin-5-yl]-methanol (LXXX)
[00445] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-(2,3-dihydro-furo[3,2-b]pyridin-6- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (88 mg, 0.20 mmol) in methanol (10 mL) was stirred with 1.00 M of Hydrogen chloride in water (1 mL). After 16 hours the reaction was condensed and the residue partitioned between ethyl acetate and sat. aq. NaHCO3. The organic layer was separated, washed with brine, dried over Na2SO4, filtered and condensed to provide 59.6 mg (90% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3 + CD3OD) δ ppm 7.90 (d, J= 1.8 Hz, 1 H), 6.97 (d,
J= 1.5 Hz, 1 H), 4.69 (t, J= 9.0 Hz, 2H), 4.42 - 4.59 (m, 2H), 3.83 (dd, J= 10.9, 3.8 Hz, 1 H), 3.58 - 3.68 (m,
1H), 3.49 - 3.58 (m, 2H), 3.39 - 3.44 (m, 1 H), 3.29 - 3.34 (m, 2H). LC-MS (5-100-5 method): Rt = 1.49 min. (99.6% HPLC purity); m/z = 334.0 (M+1).
Example 77
[2-Amino-4-chloro-7-(4-chloro-1-methyl-1H-pyrrolo[2,3-b]pyridin-5-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LXXXI)
Step 1. 4-Chloro-1H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester
[00446] 4-Chloro-5-iodo-1H-pyrrolo[2,3-b]pyridine (400 mg, 1.44 mmol) in ethanol (4.0 mL) was treated with Sodium acetate (236 mg, 2.88 mmol) and nitrogen was bubbled through the solution. The 1 ,1'-bis(di-t-butylphosphino)ferrocene palladium dichloride (75 mg, 0.12 mmol) was then added and CO was bubbled through the solution. The reaction was then heated at 75 0C under a CO balloon. After 36 hours the reaction was filtered through celite and condensed. The residue was then partitioned between ethyl acetate and water, separated, and the aqueous layer was extracted 3 more times with ethyl acetate. The combined organic layers were then washed with water, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (0-50% gradient of ethyl acetate/hexanes) to provide 210 mg (65% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 11.35 (br. s., 1 H), 8.90 (s, 1 H), 7.46 (dd, J= 3.4, 2.1 Hz, 1 H), 6.75 (dd, J= 3.5, 1.8 Hz, 1 H), 4.47 (q, J= 7.2 Hz, 2H), 1.46 (t, J= 7.1 Hz, 3H). LC-MS (5-100-5 method): Rt = 2.70 min.; m/z = 278.8 (M+1). Step 2. 4-Chloro-1 -methyl-1 H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester [00447] 4-Chloro-1 H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester (50 mg, 0.22 mmol) in
N,N-dimethylformamide (1.6 mL) at 0 °C was treated with Sodium hydride (6.18 mg, 0.245 mmol) and methyl iodide (15.2 μM, 0.245 mmol). After 45 minutes an additional 3 mg of NaH was added. After 3 hours the reaction was diluted with ethyl acetate, washed 2x with water, washed with brine, dried over Na2SO4, filtered and condensed to provide 52 mg (98% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.86 (s, 1 H), 7.24 (d, J= 3.5 Hz, 1H), 6.67 (d, J= 3.5 Hz, 1 H), 4.37 - 4.50 (m, 2H), 3.91 (s, 3H), 1.43 (t, J= 7.1 Hz, 3H). LC-MS (5-100-5 method): Rt = 2.98 min.; m/z = 239.0 (M+1). Step 3. (4-Chloro-1 -methyl-1 H-pyrrolo[2,3-b]pyridin-5-yl)-methanol
[00448] 4-Chloro-1 -methyl-1 H-pyrrolo^.S-blpyridine-δ-carboxylic acid ethyl ester (52 mg, 0.22 mmol) in tetrahydrofuran (1.4 mL) at 0 0C was treated with 1.00 M of Lithium tetrahydroaluminate in THF(0.220 mL, 0.220 mmol). After 10 minutes the ice bath was removed. After 1 hour an additional 0.220 mL of LiAIH4 was added. After 1 hour and 20 minutes, the reaction was treated with 0.020 mL of water, 0.020 mL of 15% NaOH, followed by 0.060 mL of water and the reaction was let stir for 15 minutes at room temperature. MgSO4 was then added and the reaction was stirred for an additional 15 minutes. The reaction was then filtered through celite and condensed to provide 53 mg (-80% purity by NMR, -90 % yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.31 (s, 1 H), 7.21 (d, J= 3.5 Hz, 1 H), 6.54 (d, J= 3.5 Hz, 1 H), 4.90 (s, 2H), 3.88 (s, 3H). LC-MS: Rt = 2.18 min.; m/z = 197.0 (M+1). Step 4. 4-Chloro-5-chloromethyl-1-methyl-1H-pyrrolo[2,3-b]pyridine
[00449] (4-Chloro-1 -methyl-1 H-pyrrolo[2,3-b]pyridin-5-yl)-methanol (43 mg, 0.22 mmol) was stirred in methylene chloride (2.0 mL) and thionyl chloride (24 μM, 0.33 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 1 hour the reaction was condensed. The residue was then partitioned between ethyl acetate and sat. aq. NaHCO3. The layers were separated and the organics were dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (20-100% ethyl acetate/hexanes gradient) to provide 25 mg of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.33 (s, 1 H), 7.23 (d, J= 3.5 Hz, 1 H), 6.57 (d, J= 3.5 Hz, 1 H), 4.87 (s, 2H), 3.88 (s, 3H). LC-MS (5-100-5 method): Rt = 2.99 min.; m/z = 215.0 (M+1). Step 5. 5-(tert-Butyl-dimetbyl-silany\oxymethyl)-4-chloro-7-(4-chloro-1-methyl-1H-pyrrolo[2,3- b]pyridin-5-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine
[00450] The title compound was made from 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (30.5 mg, 0.0969 mmol) and 4-Chloro-5-chloromethyl-1- methyl-1 H-pyrrolo[2,3-b]pyridine (25 mg, 0.12 mmol) according to the general alkylation procedure. The crude material thus obtained was purified by flash chromatography (20-100% ethyl acetate/hexanes gradient) to provide 42 mg (88% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm -0.05 (3 H, s), -0.01 (3 H, s), 0.79 (9 H, s), 3.35 (1 H, td, J= 6.1, 3.5 Hz), 3.49 - 3.59 (3 H, m), 3.83 - 3.94 (4 H, m), 4.68 - 4.93 (4 H, m), 6.57 (1 H, d, J= 3.5 Hz), 7.24 (1 H, d, J= 3.5 Hz), 8.35 (1 H, s). LC- MS (5-100-5 method): Rt = 3.34 min.; m/z = 493.1 (M+1).
Step 6. [2-Amino-4-chloro-7-(4-chloro-1-methyl-1H-pyrrolo[2,3-b]pyridin-5-ylmethyl)-6,7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-5-yl]-methanol (LXXXI)
[00451] A solution of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-7-(4-chloro-1-methyl-1 H- pyrrolo[2,3-b]pyridin-5-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (42 mg, 0.085 mmol) in methanol (4.3 ml_) was stirred with 1.00 M of Hydrogen chloride in water (0.4 ml_, 0.4 mmol) and the reaction was heated at 55 0C. After 16 hours the reaction was condensed and the residue partitioned between ethyl acetate and sat. aq. NaHCO3. The organic layer was separated, and the aqueous layer was back extracted with ethyl acetate. The combined organics were washed with brine, dried over Na2SO4, filtered and condensed to provide 26.6 mg (79% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3 + CD3OD) δ ppm 3.36 - 3.47 (3 H, m), 3.66 - 3.77 (5 H, m), 4.56 - 4.77 (2 H, m), 6.42 (1 H, d, J= 3.5 Hz), 7.14 (1 H, d, J= 3.5 Hz), 8.12 (1 H, s). LC-MS (5-100-5 method): Rt = 2.23 min.; m/z = 379.0 (M+1). Example 78
{2-Amino-4-chlσro-7-[4-methyl-1-(toluene-4-sυlfonyl)-1H-pyrrolo[2,3-b]pyridin-5-ylmethyl]-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (LXXXII)
Step 1. 4-Chloro-1-(toluene-4-sulfonyl)-1H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester [00452] A solution of 4-Chloro-1H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester (100 mg,
0.445 mmol) in N,N-dimethylformamide (1.7 mL) at 0 0C was treated with Sodium hydride (16.9 mg, 0.668 mmol) and p-Toluenesulfonyl chloride (93.4 mg, 0.490 mmol). After 3 days the reaction was diluted with ethyl acetate, washed 2x with water, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (50-100% ethyl acetate/hexanes gradient) to provide the title compound (174 mg, 100% yield) as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 1.41 (3 H, t, J= 7.1 Hz), 2.39 (3 H, s), 4.42 (2 H, q, J= 7.2 Hz), 6.79 (1 H, d, J= 4.0 Hz), 7.29 (2 H, d, J= 8.6 Hz), 7.81 (1 H, d, J= 4.0 Hz), 8.07 (2 H, d, J= 8.3 Hz), 8.90 (1 H, s). LC-MS (5-100-5 method): Rt = 3.51 min.; m/z = 378.9 (M+1).
Step 2: 4-Methyl-1-(toluene-4-sulfonyl)-1H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester [00453] 4-Chloro-1 -(toluene-4-sulfonyl)-1 H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester
(170 mg, 0.449 mmol) in 1 ,4-dioxane (2 mL) was treated with tetrakis(triphenylphosphine)palladium(0) (25.9 mg, 0.0224 mmol) and 2.00 M of trimethylaluminum in toluene (247 μM, 0.494 mmol). The reaction was then heated at 100 0C. After 17 hours the reaction was cooled, diluted with water, and extracted with ethyl acetate. The organics were then washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (0-100% ethyl acetate/hexanes gradient) to provide the title compound (150 mg, 93% yield). 1H NMR (400 MHz, CDCI3) δ ppm 1.40 (3 H, t, J= 7.2 Hz), 2.37 (3 H, s), 2.75 (3 H, s), 4.38 (2 H, q, J= 7.1 Hz), 6.71 (1 H, d, J= 4.0 Hz), 7.22 - 7.31 (2 H, m), 7.74 (1 H, d, J= 4.0 Hz), 8.07 (2 H, d, J= 8.3 Hz), 8.96 (1 H, s). LC-MS (5-100-5 method): Rt = 3.40 m/z = 359.0 (M+1).
Step 3. [4-Methyl-1-(toluene-4-sulfonyl)- 1 H-pyrrolo[2, 3-b]pyridin-5-yl]-methanol [00454] 4-Methyl-1 -(toluene-4-sulfonyl)-1 H-pyrrolo[2,3-b]pyridine-5-carboxylic acid ethyl ester (79 mg, 0.22 mmol) in tetrahydrofuran (5.0 mL, 62 mmol) at 0 0C was treated with 1.00 M of lithium tetrahydroaluminate in THF (441 μM, 0.441 mmol). After 1 hour and 20 minutes the reaction was treated with 0.020 mL of water. 0.020 mL of 15% NaOH was then added followed by 0.060 mL of water and the reaction was let stir for 15 minutes at room temperature. MgSO4 was then added and the reaction was stirred for an additional 15 minutes. The reaction was then filtered through celite and condensed to provide 62 mg (89% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 2.35 (3 H, s), 2.53 (3 H, s), 4.77 (2 H, s), 6.63 (1 H, d, J= 4.0 Hz), 7.25 (2 H, d, J= 8.1 Hz), 7.69 (1 H, d, J= 4.0 Hz), 8.04 (2 H, d, J= 8.3 Hz), 8.32 (1 H, s). LC-MS (5-100-5 method): Rt = 2.72 min.; m/z = 317.0 (M+1). Step 4. 5-Chloromethyl-4-methyl-1-(toluene-4-sυlfonyl)-1H-pyrrolo[2,3-b]pyridine [00455] [4-Methyl-1-(toluene-4-sulfonyl)-1 H-pyrrolo[2,3-b]pyridin-5-yl]-methanol (62 mg, 0.20 mmol) was stirred in methylene chloride (1.8 mL, 28 mmol) and thionyl chloride (28 μM, 0.39 mmol) was added. The reaction was let stir under a nitrogen balloon at room temperature. After 1 hour the reaction was diluted with ethyl acetate, washed with sat. aq. NaHCO3, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (50-100% ethyl acetate/hexanes gradient) to yield 45 mg (68% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 2.36 (3 H, s), 2.55 (3 H, s), 4.70 (2 H, s), 6.63 (1 H, d, J= 4.0 Hz), 7.26 (2 H, d, J= 8.1 Hz), 7.71 (1 H, d, J= 4.0 Hz), 8.05 (2 H, d, J= 8.3 Hz), 8.32 (1 H, s). LC-MS (5-100-5 method): Rt = 3.27 min.; m/z = 355.0 (M+1).
Step 5. S^tert-Butyl-dimethyl-sHanyloxymethylW-chloro-T-μ-methyl-i-ζtoluene^-sulfonyO-IH- pyrrolo[2,3-b]pyridin-5-ylmethyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine [00456] 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- ylamine (38.5 mg, 0.122 mmol) was alkylated with 5-Chloromethyl-4-methyl-1-(toluene-4-sulfonyl)-1 H- pyrrolo[2,3-b]pyridine (45 mg, 0.13 mmol) according to the general alkylation procedure. The crude material thus obtained was then purified by flash chromatography (20-100% ethyl acetate/hexanes gradient) to provide 58 mg (77% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm -0.12 (3 H, s), -0.07 (3 H, s), 0.72 (9 H, s), 2.36 (3 H, s), 2.43 (3 H, s), 3.27 - 3.42 (3 H, m), 3.49 (1 H, dd, J= 10.0, 7.7 Hz), 3.81 (1 H, dd, J= 10.0, 3.7 Hz), 4.48 (1 H, d, J= 14.9 Hz), 4.72 (1 H, d, J= 14.7 Hz), 4.93 (2 H, s), 6.61 (1 H, d, J= 4.0 Hz), 7.27 (2 H, d, J= 8.1 Hz), 7.71 (1 H, d, J= 4.0 Hz), 8.07 (2 H1 d, J= 8.3 Hz), 8.26 (1 H, s). LC-MS (5-100-5 method): Rt = 3.55 min.; m/z = 613.1 (M+1). Step 6. {2-Amino-4-chloro-7-[4-methyl-1-(toluene-4-sulfonyl)-1H-pyrrolo[2,3-b]pyridin-5-ylmethyl]- 6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (LXXXII)
[00457] S-ftert-Butyl-dimethyl-silanyloxymethylH-chloro^-^-methyl-i-Ooluene^-sulfonylJ-I H- pyrrolo[2,3-b]pyridin-5-ylmethyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (58 mg, 0.094 mmol) in methanol (4.8 ml_) was stirred with 1.00 M of Hydrogen chloride in water (0.5 ml_, 0.5 mmol) and the reaction was heated at 55 °C. After 2 hours the reaction was condensed and the residue partitioned between ethyl acetate and sat. aq. NaHCO3. The organic layer was separated, and the aqeous layer was back extracted with ethyl acetate. The combined organics were washed with brine, dried over Na2SO4, filtered and condensed to provide 43 mg (91% yield) of the title compound. 1H NMR (400 MHz, CDCI3 + MeOD) δ ppm 2.38 (4 H, s), 2.45 (3 H, s), 3.29 - 3.53 (3 H, m), 3.59 (1 H, m, J= 10.9, 7.3 Hz), 3.79 (1 H, dd, J= 10.9, 3.5 Hz), 4.53 (1 H, d, J= 14.9 Hz), 4.71 (1 H, d, J= 14.7 Hz), 6.65 (1 H, d, J= 4.3 Hz), 7.29 (2 H, d, J= 8.1 Hz), 7.70 (1 H, d, J= 4.0 Hz), 8.02 (2 H, d, J= 8.3 Hz), 8.22 (1 H, s). LC-MS (5-100-5 method): Rt = 2.64 min.; m/z = 499.1 (M+1). Example 79
Acetic acid 2-amino-4-chloro-7-(4-chloro-1-methyl-1 H-pyrrolo[2,3-b]pyridin-5-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-ylmethyl ester (LXXXIII)
[00458] The title compound was made as a by-product of the alkylation of 5-(tert-Butyl-dimethyl- silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (110 mg, 0.349 mmol) with 4- Chloro-5-chloromethyl-1-methyl-1 H-pyrrolo[2,3-b]pyridine (90 mg, 0.42 mmol) via the general alkylation procedure. The crude material thus generated was then purified by flash chromatography (20-100% ethyl acetate/hexanes gradient) to yield 129 mg (75% yield) of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro- 7-(4-chloro-1-methyl-1 H-pyrrolo[2,3-b]pyridin-5-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2- ylamine and 27 mg (18% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.32 (s, 1 H), 7.23 (d, J= 3.5 Hz, 1 H), 6.55 (d, J= 3.5 Hz, 1 H), 5.02 (s, 2H), 4.83 - 4.91 (m, 1 H), 4.67 - 4.78 (m, 1 H), 4.32 (dd, J= 11.0, 4.2 Hz, 1 H), 4.00 - 4.07 (m, 1 H), 3.89 (s, 3H), 3.45 - 3.64 (m, 2H), 3.35 (dd, J= 9.5, 3.2 Hz, 1 H), 1.99 (s, 3H). LC-MS (5-100-5 method): Rt = 2.53 (95.4%) m/z = 421.0 (M+1 ). Example 80
[2-Amint>-4-chloro-7-(4-methyl-1H-pyrrolo[2,3-b]pyridin-5-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-methanol (LXXXIV)
[00459] {2-Amino-4-chloro-7-[4-methyl-1 -(toluene-4-sulfonyl)-1 H-pyrrolo[2,3-b]pyridin-5-ylmethyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl}-methanol (19.1 mg, 0.0383 mmol) in methanol (1.9 mL) was stirred with 5.0 M of potassium hydroxide in water (76 μM, 0.38 mmol) and heated at 60 0C. After 18 hours the reaction was diluted with ethyl acetate and the reaction was washed with water, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by preperative HPLC (gradient of 5-100% acetonitrile/water 0.1% TFA buffer) to provide 3 mg (20% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3 + MeOD) δ ppm 8.07 (br. s., 1 H), 7.32 (d, J= 3.3
Hz, 1 H), 6.53 (d, J= 3.5 Hz, 1H), 5.34 (s, 1 H), 4.77 (d, J= 14.4 Hz, 1 H), 4.60 (d, J= 14.7 Hz, 1 H), 3.81 (d,
J= 11.1 Hz, 1 H), 3.69 - 3.75 (m, 1 H), 3.49 - 3.57 (m, 1 H), 3.39 - 3.49 (m, 2H), 2.54 (s, 3H). LC-MS (5-100-
5 method): Rt = 1.79 (98.8% HPLC purity) m/z = 345.1 (M+1).
Example 81
(2-Amino^-chloro-7^(4-methoxy-3-rnethyl-5-propylpyridin-2-yl)rnethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (LXXXV)
Step 1. (5-bromo-4-methoxy-3-methylpyridin-2-yl)methyl acetate
[00460] A mixture of 5-Bromo-4-methoxy-2,3-dimethyl-pyridine 1 -oxide (10.0 g, 43.1 mmol) and
Acetic anhydride (100 rπL, 1.06 mol) was refluxed at 1200C for 30 minutes. The solution was removed from heat, cooled to ambient temperature, and condensed under vacuum to a crude solid. The solid was purified by silica gel chromatography in ethyl acetate/hexane to give the title compound as a yellow solid
(7.49 g, yield 63.4%).
Step 2. (5-bromo-4-methoxy-3-methylpyridin-2-yl)methanol
[00461] A solution of (5-bromo-4-methoxy-3-methylpyridin-2-yl)methyl acetate (3.453 g, 12.60 mmol) in Methanol (20.0 mL), Tetrahydrofuran (40.0 mL), and Water (10.0 mL) was treated with
Potassium carbonate (5.22 g, 37.8 mmol). When complete, the reaction was diluted with water and brine, extracted 3 times with ethyl acetate, and dried over magnesium sulfate. Evaporation of the solvent under vacuum gave the title product as a viscous oil (2.8 g, yield 95.8%).
Step 3. 5-bromo-4-methoxy-2-((methoxymethoxy)methyl)-3-methylpyridine
[00462] A solution of (5-Bromo-4-methoxy-3-methyl-pyridin-2-yl)-methanol (2.8 g, 12.1 mmol) in methylene chloride (116.0 mL) and N,N-Diisopropylethylamine (6.30 mL, 36.2 mmol) was cooled to 00C.
Chloromethyl Methyl Ether (1.37 mL, 18.1 mmol) was added in a dropwise manner. Upon completion, the solution was condensed under vacuum to give a solid which was purified by silica gel chromatography eluting with ethyl acetate/hexane to yield the title product as an oil (1.488 g, yield 44.7%).
Step 4. 5-allyl-4-methoxy-2-((methoxymethoxy)methyl)-3-methylpyridine
[00463] A solution of 5-bromo-4-methoxy-2-((methoxymethoxy)methyl)-3-methylpyridine (400.0 mg, 1.449 mmol) in N,N-Dimethylformamide (2.41 mL) was treated with allyltributyltin (0.576 g, 1.74 mmol) and purged with nitrogen for 15 minutes. Tetrakis(triphenylphosphine)palladium(0) (170 mg, 0.15 mmol) was added and the mixture was microwaved for 10 minutes at 1200C. The catalyst was then filtered off and rinsed with ethyl acetate. The crude filtrate was purified by silica gel chromatography eluting with ethyl acetate/hexane to give the title product as a yellow liquid (320 mg, yield 93.1%).
Step S. 4-methoxy-2-((methoxymethoxy)methyl)-3-methyl-5-propylpyridine
[00464] A solution of 5-allyl-4-methoxy-2-((methoxymethoxy)methyl)-3-methylpyridine (75.0 mg,
0.316 mmol) in Ethanol (5.00 mL) was treated with 10% Palladium on carbon (10.0 mg, 0.0940 mmol).
The mixture was hydrogenated under 1 atm of hydrogen. Upon completion the catalyst was removed by filtration and rinsed with ethanol. Purification by silica gel chromatography eluting with ethyl acetate/hexane gave the title compound as a clear oil (43 mg, yield 56.9%).
Step 6. (4-methoxy-3-methyl-5-propylpyridin-2-yl)methanol
[00465] A solution of 4-methoxy-2-((methoxymethoxy)methyl)-3-methyl-5-propylpyridine (43.0 mg, 0.180 mmol) in 2.9 M HCI in Ethanol (1.50 ml_, 4.35 mmol) was stirred at room temperature. Upon removal of the methoxymethyl protecting group, the solution was treated with 7.5M NaOH in Water (750 μL, 5.6 mmol) and stirred for 5 minutes. The solution was the diluted with water, extracted 3 times with dichloromethane, and dried over magnesium sulfate. Purification by silica gel chromatography eluting with ethyl acetate/hexane gave the title compound (28 mg, yield 64%).
Step 7. 2-(chloromethyl)-4-methoxy-3-methyl-5-propylpyridine
[00466] A solution of (4-methoxy-3-methyl-5-propylpyridin-2-yl)methanol (24.0 mg, 0.123 mmol) in Methylene chloride (1.6 ml_) was treated with thionyl chloride (89.66 μL, 1.229 mmol). After 30 minutes the reaction was condensed under vacuum to a dry solid, then taken up in ethyl acetate and washed twice with saturated aqueous sodium bicarbonate. The organic phase was dried over magnesium sulfate and purified by silica gel chromatography eluting with ethyl acetate/hexane to give the title compound (19 mg, yield 72%).
Step 8. 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((4-methoxy-3-methyl-5-propylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
[00467] A solution of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (28.0 mg, 0.0889 mmol) in Dimethyl sulfoxide (0.60 ml_) was treated with Sodium hydride (3.3 mg, 0.090 mmol) and stirred for 5 minutes. To this was added a solution of 2-(chloromethyl)-
4-methoxy-3-methyl-5-propylpyridine (19.0 mg, 0.0889 mmol) in dimethyl sulfoxide (0.60 ml_). After 2 hours the reaction was diluted with water, extracted 3 times with ethyl acetate, washed with brine, and dried over magnesium sulfate. Purification by silica gel chromatography eluting with ethyl acetate/hexane gave the title compound (15 mg, yield 34.3%)
Step 9. (2-amino-4-chloro-7-((4-methoxy-3-methyl-5-propylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXV)
[00468] A solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((4-methoxy-3-methyl-5- propylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (15.0 mg, 0.0305 mmol) in
Methanol (2.3 mL) was treated with 1 M HCI in water (0.12 ml_, 0.12 mmol) at room temperature. After 6 hours, the reaction was condensed under vacuum and the residue partitioned between ethyl acetate and saturated NaHCO3. The organic layer was separated, washed with brine, dried over magnesium sulfate and condensed under vacuum. The crude product was purified by reverse-phase HPLC (CH3CN/H2O,
0.1% TFA). The title product was isolated as a white solid (2.48 mg, yield 21.3%); LC-MS (5-100-5 method) : Rt = 2.059 min; M+1 = 378.1 ; 1H NMR (400 MHz, CDCI3) δ ppm 8.16 (s, 1 H), 5.03 (d, J= 16.7
Hz, 1 H), 4.90 (s, 2 H), 4.30 (d, J= 16.4 Hz, 1 H), 3.94 (dd, J= 11.0, 4.2 Hz, 1 H), 3.79 - 3.86 (m, 2 H), 3.77 (S, 3 H)1 3.64 (dd, J= 10.1 , 3.0 Hz, 1 H), 3.37 - 3.46 (m, 1 H), 2.51 - 2.63 (m, 2 H), 2.23 (s, 3 H), 1.55
- 1.70 (m, 2 H), 0.97 (t, J= 7.3 Hz, 3 H).
Example 82
(2-Amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridln-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXVI)
Step 1. 4-methoxy-2-((methoxymethoxy)methyl)- 3-methyl-5-trideυteromethylpyridine
[00469] A solution of 0.50 M of ZnCI2 in tetrahydrofuran (59.0 ml_) in a flame-dried sealed tube under nitrogen was treated with 1.0 M methyl-d3-magnesium iodide in diethyl ether (27.6 ml_, 27.6 mmol).
After stirring for 20 minutes N-Methylpyrrolidinone (22.1 ml_) was added. After 5 minutes, Bis(tri-tert-butyl- phosphine)palladium(O) (0.19 g, 0.37 mmol) was added, followed by a solution of 5-bromo-4-methoxy-2-
((methoxymethoxy)methyl)-3-methylpyridine (5.09 g, 18.4 mmol) in N-Methylpyrrolidinone (18.4 ml_). The tube was sealed and heated to 8O0C for 3 days. The reaction was then cooled, treated with water, and filtered to remove any solids. The filtrate was extracted 5 times with ethyl acetate then condensed under vacuum. Purification was performed by silica gel chromatography in ethyl acetate/hexane to give the title compound as a yellow oil (2.13 g, yield 54%).
Step 2. (4-methoxy-3-methyl-5-trideuteromethylpyridin-2-yl)methanol
[00470] 4-Methoxy-2-((methoxymethoxy)methyl)- 3-methyl-5-trideuteromethylpyridine (5.33 g,
24.9 mmol) was added to a 2.9 M HCI in ethanol (88.8 mL, 258 mmol) and the mixture was stirred at room temperature. Upon removal of the methoxymethyl protecting group, the solution was treated with7.5M NaOH in Water (50 mL, 373 mmol) and stirred for 5 minutes. The solution was diluted with water, extracted 3 times with dichloromethane, and dried over magnesium sulfate. Purification by silica gel chromatography eluting with ethyl acetate/hexane gave the title compound as a colorless oil (4.07 g, yield 96%).
Step 3. 2-(chloromethyl)-4-methoxy-3-methyl-5-trideuteromethylpyridine
[00471] A solution of (4-methoxy-3-methyl-5-trideuteromethylpyridin-2-yl)methanol (100 mg,
0.588 mmol) in Methylene chloride (7.5 mL) was treated with thionyl chloride (214 μL, 2.94 mmol). After
30 minutes the reaction was evaporated under vacuum to a dry solid, taken up in ethyl acetate, and rinsed twice with saturated aqueous sodium bicarbonate. The organic phase was dried over magnesium sulfate and purified by silica gel chromatography eluting with ethyl acetate/hexane to give the title compound (98 mg, yield 88%).
Step 4. (2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2-yl)methyl)-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXVI)
[00472] A solution of S-^ert-Butyl-dimethyl-silanyloxymethyO^-chloro-ΘJ-dihydro-δH-pyrrolo^.S- d]pyrimidin-2-ylamine (159 mg, 0.506 mmol) in Dimethyl sulfoxide (7.2 mL) was stirred in a flame-dried flask under nitrogen. Potassium tert-Butoxide (56.8 mg, 0.506 mmol) was added, followed by 10 minutes of stirring. 2-(chloromethyl)-4-methoxy-3-methyl-5-trideuteromethylpyridine (98.0 mg, 0.519 mmol) was then added at room temperature. Upon consumption of the starting material, the reaction diluted with water and extracted 3 times with ethyl acetate. The combined organic phases were condensed under vacuum and stirred overnight in Methanol (21 mL) and 1.0 M HCI in Water (2.025 ml_, 2.025 mmol). The mixture was treated with aqueous sodium bicarbonate and extracted 3 times with ethyl acetate.
Purification by silica gel chromatography column using dichloromethane/methanol gave the title compound as a white powder (126 mg, yield 70%). LC-MS (5-100-5 method) : Rt = 1.777 min; M+1 =
353.0; 1H NMR (400 MHz, CDCI3) δ ppm 8.15 (s, 1 H), 5.02 (d, J= 16.4 Hz, 1 H), 4.76 (s, 2 H), 4.30 (d, J=
16.4 Hz, 1 H), 3.89 - 4.08 (m, 2 H), 3.83 (d, J= 2.5 Hz, 1 H), 3.77 (s, 3 H), 3.74 - 3.82 (m, 1 H), 3.61 (dd,
J= 10.1 , 3.0 Hz, 1 H), 3.37 - 3.46 (m, 1 H), 2.23 (s, 3 H)
Example 83
(2-Amino-7'((5-bromo-2,3-dihydrofuro[3,2-b]pyridin-6-yl)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXVII)
Step 1. (2,3-dihydrofuro[3,2-b]pyridin-6-yl)methanol
[00473] A solution of Furo[3,-b]pyridin-6-carboxaldehyde (0.500 g, 3.40 mmol) in Ethanol (11.9 mL) was stirred in a bomb. 10% Palladium on Carbon (1 :9, Palladiumxarbon black, 181 mg, 0.170 mmol) was added and the mixture hydrogenated at 80 psi for 18 hours. The catalyst was removed by filtration and rinsed with ethanol. Concentration of the filtrate gave a crude mixture of reduction products which were carried on without further purification (384 mg, crude product).
Step 2. (5-bromo-2,3-dihydrofuro[3,2-b]pyridin-6-yl)methanol
[00474] A solution of (2,3-Dihydro-furo[3,2-b]pyridin-6-yl)-methanol (100.0 mg, 0.4631 mmol) in
Acetonitrile (4.8 mL) was treated with N-Bromosuccinimide (123.6 mg, 0.6946 mmol) and stirred overnight. The reaction was then condensed under vacuum and partitioned between dichloromethane, methanol, and brine. Extraction with dichloromethane was performed 4 times and the combined organic layers were dried over magnesium sulfate. Purification was performed by silica gel chromatography, eluting with ethyl acetate/hexane to afford the title product as a white solid (64 mg, yield 60%).
Step 3. 5-bromo-6-(chloromethyl)-2,3-dihydrofuro[3,2-b]pyridine
[00475] A solution of (5-bromo-2,3-dihydrofuro[3,2-b]pyridin-6-yl)methanol (100.0 mg, 0.278 mmol) in Methylene chloride (3.6 mL) was treated with thionyl chloride (203 μL, 2.785 mmol). After 30 minutes the reaction was condensed under vacuum to a dry solid then taken up in ethyl acetate and rinsed twice with saturated aqueous sodium bicarbonate. The organic phase was dried over magnesium sulfate and purified by silica gel chromatography eluting with ethyl acetate/hexane to give the title compound (55 mg, yield 79%).
Step 4. 7-((5-bromo-2,3-dihydrofuro[3,2-b]pyridin-6-yl)methyl)-5-((tert- butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
[00476] A solution of 5-(tert-Butyl-dimethyl-silanyloxymethyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (35 mg, 0.11 mmol) in N,N-Dimethylformamide (0.81 mL) was treated with the
Sodium hydride (2.8 mg, 0.11 mmol). To that was added a solution of 5-bromo-6-(chloromethyl)-2,3- dihydrofuro[3,2-b]pyridine (27.6 mg, 0.111 mmol) in N,N-Dimethylformamide (0.81 mL) followed by stirring for 2 hours. Water was then added, followed by three ethyl acetate extractions and drying over magnesium sulfate. Purification by silica gel chromatography using ethyl acetate/hexane gave the title compound as a yellow solid (26 mg, yield 40%).
Step 5. (2-amino-7-((5-bromo-2,3-dihydrofuro[3,2-b]pyridin-6-yl)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXVII)
[00477] A solution of 7-((5-bromo-2,3-dihydrofuro[3,2-b]pyridin-6-yl)methyl)-5-((tert- butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (26.0 mg, 0.0493 mmol) in Methanol (3.8 ml.) was treated with "IM HCI in water (0.19 ml_, 0.19 mmol). Upon completion the reaction was condensed under vacuum and the residue partitioned between ethyl acetate and saturated sodium bicarbonate. The organic layer was separated, washed with brine, dried over MgSO4 and condensed. The crude material was purified by silica gel chromatography, eluting with dichloromethane/methanol to give the title compound as a white powder (10.0 mg, yield 48%). LC-MS (5-
100-5 method) : Rt = 2.056 min; M+1 = 413.9; NMR not taken due to solubility.
Example 84
(2-Amino-4-chloro-7-((2,2-difluoro-4-methylbenzo[d][1,3]dioxol-5-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXVIII)
Step 1. 2,2-difluoro-4-methylbenzo[d][1,3]dioxole-5-carboxylic acid
[00478] To a stirred solution of 2,2-difluorobenzo[d][1 ,3]dioxole-5-carboxylic acid (1g, 4.95 mmol) in dry THF (20 mL) was added n-Buϋ (6.8 ml_, 10.8 mmol of 1.6M) drop wise at - 78 0C. After being stirred for 6h, the resulting reaction mixture was then treated with MeI (0.61 mL, 9.90 mmol) at RT. The resultant reaction mixture was stirred at RT for 16h. The progress of the reaction was monitored by TLC.
The reaction mixture was quenched with a saturated solution of NH4CI, extracted with EtOAc (3x 30 mL) and dried over Na2SO4. The resulting material was concentrated under reduced pressure to afford the title compound (1 g, as crude) as a brown solid. TLC system: 10% MeOH/dichloromethane, Rf = 0.5.
Step 2. methyl 2,2-difluoro-4-methylbenzo[d][1,3]dioxole-5-carboxylate
[00479] To a stirred solution of 2,2-difluoro-4-methylbenzo[d][1 ,3]dioxole-5-carboxylic acid (1g,
4.62 mmol) in MeOH (20 mL) was added thionyl chloride (1 mL, 13.8 mmol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 5h. The volatiles were evaporated under reduced pressure and the crude material was diluted with water and extracted with dichloromethane. The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure. The crude material was purified by column chromatography to afford the title compound (250 mg, 23.5%). 1H NMR (200 MHz, CDCI3): δ 7.80 (d, J = 8.2 Hz, 1 H), 6.95 (d, J = 8.2 Hz, 1H), 3.89 (s, 3H),
2.55 (s, 3H). TLC system: 30% EtOAc/Hexane, Rf = 0.7.
Step 3. (2,2-difluoro-4-methylbenzo[d][1, 3]dioxol-5-yl)methanol
[00480] To a stirred solution of methyl 2,2-difluoro-4-methylbenzo[d][1 ,3]dioxole-5-carboxylate
(250 mg, 0.001 mol) in THF (10 mL) was added DIBAL-H (1.4 mL, 2.17 mmol of 1.6M in Toluene) at 0 0C.
The resulting reaction mixture was stirred at RT for 4h. The reaction mixture was quenched with a saturated solution of sodium potassium tartarate. The volatiles were evaporated under reduced pressure and the crude material was diluted with water and extracted with EtOAc (3x 10 ml_). The combined organic layers were dried over Na2SO4. The solvent was evaporated under reduced pressure. The crude material was purified by column chromatography to afford the title compound (180 mg, 82%). 1H NMR (200 MHz, CDCI3): δ 7.08 (d, J = 8 Hz, 1 H), 6.88 (d, J = 8 Hz, 1 H), 4.68 (bs, 2H), 2.32 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.83 min. (70%); m/z = 205.1 (M+2). Step 4. 5-(chloromethyl)-2,2-difluoro-4-methylbenzo[d][1, 3]dioxole
[00481] To a stirred solution of (2,2-difluoro-4-methylbenzo[d][1 ,3]dioxol-5-yl)methanol (80mg,
0.396 mmol) in dry dichloromethane (5 ml_) was added thionyl chloride (0.08 ml_, 1.189 mmol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 4h. The crude material was diluted with dichloromethane, washed with a saturated NaHCO3 solution and extracted with EtOAc (2x 15 ml_). The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure to afford the title compound (70 mg, 80% as crude). This material was used for next step with out any purification. 1H NMR (200 MHz, CDCI3): δ 7.08 (d, J = 8.4 Hz, 1H), 6.92 (d, J= 8.4 Hz, 1 H), 4.62 (s, 2H)1 2.41 (s, 3H). TLC system: 10% EtOAc/Hexane, Rf = 0.7. Step 5. 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((2,2-difluoro-4- methylbenzo[d][1,3]dioxol-5-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine [00482] To a stirred solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-amine (40 mg, 0.12 mmol) in dry DMF (4 mL) were added 60% NaH (5 mg, 0.12 mmol) and 5-(chloromethyl)-2,2-difluoro-4-methylbenzo[d][1 ,3]dioxole (42 mg, 0.19 mmol) at 0° C. The resulting reaction mixture was stirred at 0° C for 10 min. The reaction mass was diluted with ice water and extracted with EtOAc (2x 10 mL). The combined organic layers were washed with water and brine, dried over Na2SO4 and evaporated under reduced pressure. The resulting crude was purified by column chromatography (60-120 silica gel; 2% MeOH/dichloromethane) to afford the title compound (50 mg, 79%) as an off-white solid. 1H NMR 0(500 MHz, DMSO-d6): δ 7.18 (d, J = 8 Hz, 1 H), 7.09 (d, J = 8 Hz, 1H), 6.48 (s, 2H), 4.78 (d, J = 15 Hz, 1 H), 4.23 (d, J = 15 Hz, 1 H), 3.70- 3.68 (m, 1 H), 3.54- 3.52 (m, 1 H), 3.45- 3.43 (m, 1 H), 3.22- 3.20 (m, 2H), 2.21 (s, 3H), 0.74 (s, 9H), -0.004 to -0.113 (m, 6H). LC-MS (10- 90[4]-10 method): Rt = 3.90 min. (91%); m/z = 499 (M+1).
Step 6. (2-amino-4-chloro-7-((2,2-difluoro-4-methylbenzo[d][1,3]dioxol-5-yl)methyl)-6,7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXVIII)
[00483] To a stirred solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((2,2-difluoro-4- methylbenzo[d][1 ,3]dioxol-5-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (50 mg, 0.10 mmol) in dry THF (5 mL) was added TBAF (0.55 mL, 0.50 mmol) at 0° C. The resulting reaction mixture was stirred at RT for 1h. The reaction mass was diluted with ice water and extracted with EtOAc (2x 20 mL).The combined organic extracts were washed with water, brine and dried over Na2SO4 and evaporated under reduced pressure. The resulting crude was purified by preparative HPLC to afford the title compound (8 mg, crude) as a white solid. 1H NMR (500 MHz, DMSO-d6): δ 7.19 (d, J = 8.5 Hz, 1 H), 7.07 (d, J = 8.5 Hz, 1 H)1 6.45 (s, 2H), 4.84 (t, J = 5Hz, 1 H), 4.65 (d, J = 15.5 Hz, 1 H), 4.36 (d, J = 15.5 Hz,
1 H), 3.63- 3.59 (m, 1 H), 3.50- 3.41 (m, 1 H), 3.36- 3.34 (m, 1 H), 3.24- 3.201 (m, 2H) 2.22 (s, 3H). LC-MS
(10-90[4]-10 method): Rt = 3.33 min. (98%); m/z = 385.3 (M+1 ).
Example 85
(2-Amino-7-((6-bromo-4-methylbenzo[d][1,3]dioxol-5-yl)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXIX)
Step 1. 4-methylbenzo[d][1,3]dioxole-5-carboxylic acid
[00484] To a stirred solution of benzo[d][1 ,3]dioxole-5-carboxylic acid (1g, 0.006 mol) in dry THF
(70 mL) was added n-BuLi (8.3 ml_, 0.013 mol of 1.6M) drop wise at - 70 °C. The resulting reaction mixture was stirred at -20 0C for 2h and then MeI (0.75 mL, 0.012 mol) was added at -70 0C. The resultant reaction mixture was stirred at 0 0C for 4h (-70% product formation was observed by TLC), and quenched with 1 N HCI at 0 C. The THF was evaporated under reduced pressure. The resulting solid was filtered off to provide the title compound (600 mg, as crude). LC-MS (10-90[4]-10 method): Rt = 2.74 min.
(70%); m/z = 181 (M+1 ).
Step 2. methyl 4-methylbenzo[d][1,3]dioxole-5-carboxylate
[00485] To a stirred solution of 4-methylbenzo[d][1 ,3]dioxole-5-carboxylic acid (300 mg, 0.00166 mol) in MeOH (10 mL) was added thionyl chloride (0.12 mL, 0.0033 mol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 16h. The volatiles were evaporated under reduced pressure and the crude material was diluted with saturated NaHCO3 solution extracted with EtOAc (2 x 10 mL). The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure. The crude material was purified by column chromatography to afford the title compound (150 mg, 46.2%). 1H NMR (200 MHz, CDCI3): δ 7.60 (d, J = 8.2 Hz, 1 H), 6.70 (d, J = 8.2 Hz,
1 H), 6.01 (s, 2H), 3.85 (s, 3H), 2.46 (s, 3H). TLC system: 10% EtOAc/Hexane, R, = 0.8.
Step 3. methyl 6-bromo-4-methylbenzo[d][1,3]dioxole-5-carboxylate
[00486] To a stirred solution of methyl 4-methylbenzo[d][1 ,3]dioxole-5-carboxylate (100 mg,
0.0005 mol) in AcOH (1 mL) was added Br2 (82 mg, 0.0005 mol) at -20 0C. The reaction mixture was stirred at -20 0C for 2h. The reaction mass was diluted with water and extracted with EtOAc (2 x 10 mL).
The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure. The crude material was purified by column chromatography to afford the title compound (60 mg, 42%). 1H NMR (200 MHz, CDCI3): δ 6.87 (s, 1 H), 5.99 (s, 2H), 3.92 (s, 3H), 2.18 (s,
3H). TLC system: 30% dichloromethane/Hexane, Rf = 0.4.
Step 4. (6-bromo-4-methylbenzo[d][1,3]dioxol-5-yl)methanol
[00487] To a stirred solution of methyl 6-bromo-4-methylbenzo[d][1 ,3]dioxole-5-carboxylate (200 mg, 0.0007 mol) in THF (2 mL) was added DIBAL-H (179 mg, 0.001 mol of 1.6M in Toluene) at 0 0C. The resulting reaction mixture was stirred at 0 0C for 2h. The reaction mixture was quenched with 2N HCI and extracted with EtOAc (3 x 10 mL). The combined organic layers were dried over Na2SO4. The solvent was evaporated under reduced pressure and the crude was washed with pentane to afford the title compound (150 mg, 79%). 1H NMR (200 MHz, CDCI3): δ 6.905 (s, 1 H), 5.96 (s, 2H), 4.76 (d, J = 3.2 Hz, 2H), 2.33 (s, 3H), 1.78 (bs, 1H). TLC system: 20% EtOAc/Hexane, R, = 0.3. Step 5. 6-bromo-5-(chloromethyl)-4-methylbenzo[d][1,3]dioxole
[00488] To a stirred solution of (6-bromo-4-methylbenzo[d][1 ,3]dioxol-5-yl)methanol (120 mg,
0.0004 mol) in CHCI3 (1.2 mL) was added thionyl chloride (172 mg, 0.001 mol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 2h. The reaction mass was quenched with saturated NaHCO3 solution and extracted with EtOAc (2 x 15 mL). The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure and the crude was washed with hexane to afford the title compound (100 mg, 78%) as a white solid. 1H NMR (200 MHz, CDCI3): δ 6.926 (s, 1 H), 5.982 (s, 2H), 4.755 (s, 2H), 2.33 (s, 3H); TLC system: 30% EtOAc/Hexane, R, = 0.7.
Step 6. 7-((6-bromo-4-methylbenzo[d][1,3]dioxol-5-yl)methyl)-5-((tert- butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine [00489] To a stirred solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-amine (30 mg, 0.00008 mol) in DMF (1.5 mL) was added NaH (5 mg, 0.0001 mol) and 6-bromo-5-(chloromethyl)-4-methylbenzo[d][1 ,3]dioxole (34 mg, 0.0001 mol) at 0 0C. The resulting reaction mixture was stirred at 0 0C for 2h. The reaction mass was quenched with water and extracted with EtOAc (2 x 5 mL).The combined organic layers was washed with brine, dried over Na2SO4 and evaporated under reduced pressure. The resulting crude was purified by column chromatography (60-120 silica gel; 40% EtOAc/Hexane) to afford the title compound (20 mg, 42%) as a white solid. 1H NMR (500 MHz, DMSO-d6): δ 7.11 (s, 1 H), 6.49 (s, 2H), 6.05 (d, J = 11.5 Hz, 2H), 4.76 (d, J = 14 Hz1 1 H), 4.43 (d, J= 14 Hz, 1 H), 3.66- 3.64 (m, 1 H), 3.43- 3.39 (m, 2H), 3.22 (bs, 1 H), 3.15- 3.13 (m, 1 H), 2.13 (s, 3H), 0.74 (s, 9H), -0.004 (s, 3H), -0.113 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 5.95 min. (85%); m/z = 540 (M+1).
Step 7. (2-amino-7-((6-bromo-4-methylbenzo[d][1,3]dioxol-5-yl)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (LXXXIX)
[00490] To a stirred solution of 7-((6-bromo-4-methylbenzo[d][1 ,3]dioxol-5-yl)methyl)-5-((tert- butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (20 mg, 0.03 mmol) in dry THF (1 mL) was added TBAF (0.5 mL, 015 mmol) at 0° C. The resulting reaction mixture was stirred at RT for 30 min. The reaction mass was quenched with water and extracted with EtOAc (2 x 20 mL).The combined organic extracts were washed with water and brine, dried over Na2SO4, evaporated under reduced pressure, and the crude was washed with Hexane to afford the title compound (10 mg, 66% yield) as a white solid. 1H NMR (500 MHz, DMSO-c/6): δ 7.12 (s, 1 H), 6.48 (s, 2H), 6.08 (bs, 2H), 4.79 (bs, 1 H), 4.73 (d, J = 14 Hz, 1 H), 4.47 (d, J = 14 Hz, 1 H), 3.57 (bs, 1 H), 3.34 (s, 1 H), 3.30-3.16 (m, 4H), 2.14 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.75 min. (90%); m/z = 427 (M+1). Example 86 (2-Amino-4-chloro-7-((6-methoxy-5-methyl-1H-benzo[d][1,2,3]triazol-4-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XC)
Step 1: methyl 3-methoxy-2-methyl-6-nitrobenzoate
[00491] To a stirred solution of methyl 3-methoxy-2-methylbenzoate (250 mg, 1.38 mmol) in glacial acetic acid (0.75 ml.) was added cone. HNO3 (0.25 mL) dropwise at RT. The resulting mixture was stirred at 70 0C for 2 h. After completion of the reaction (by TLC), the reaction mixture was diluted with cold water and the aqueous layer was extracted with EtOAc. The organic layer was washed with water,
0.5M NaOH, and brine, and was dried over Na2SO4. The solvent was evaporated under reduced pressure and the crude material was purified by column chromatography to afford the title compound (115 mg,
36.8%) as an off-white solid. 1H NMR (200 MHz, CDCI3,): δ 8.14 (d, J = 9.0 Hz, 1 H), 6.94 (d, J = 9.4 Hz,
1 H)1 3.97 (s, 6H), 2.19 (s, 3H). TLC system: 15% EtOAc/Hexane, Rf = 0.3.
Step 2. methyl 6-amino-3-methoxy-2-methylbenzoate
[00492] A solution of methyl 3-methoxy-2-methyl-6-nitrobenzoate (150 mg, 0.66 mmol) and 10%
Pd/C (60 mg) in EtOAc (10 mL) was taken in an autoclave reactor and stirred under hydrogen pressure
(25 psi) at RT for 2h. The reaction mixture was filtered through a celite bed, rinsing with EtOAc. The filtrate was washed with water, dried over Na2SO4 and evaporated under reduced pressure. The crude material was purified by column chromatography to afford the title compound (75 mg, 58 % yield) as a brown syrup. 1H NMR (200 MHz, CDCI3): δ 6.84 (d, J = 8.8 Hz, 1 H), 6.56 (d, J = 8.8 Hz, 1 H), 4.37 (bs,
2H), 3.92 (s, 3H), 3.78 (s, 3H), 2.25 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 2.89 min. (98%); m/z =
196 (M+1 ).
Step 3. methyl 6-acetamido-3-methoxy-2-methylbenzoate
[00493] To a stirred solution of methyl 6-amino-3-methoxy-2-methylbenzoate (200 mg, 1.02 mmol) in chloroform (10 mL) were added TEA (0.15 mL, 1.12 mmol) and acetyl chloride (0.07 mL, 1.02 mmol) at RT under anhydrous conditions. The resulting mixture was stirred at RT for 3 hrs. After completion of the reaction (by TLC), the volatiles were evaporated under reduced pressure. The crude material was then taken up in EtOAc, washed with water, washed with brine and then concentrated under reduced pressure to afford the title compound (200 mg, 80.2% yield) as an off-white solid. 1H NMR (200
MHz, DMSO-cfe): δ 9.41 (bs, 1 H), 7.18 (d, J = 8.8 Hz, 1 H), 7.02 (d, J = 8.8 Hz, 1 H), 3.80 (s, 3H), 3.78 (s,
3H), 2.08 (S, 3H), 1.94 (s, 3H). TLC system: 50% EtOAc/Hexane: R,= 0.2.
Step 4. methyl 2-acetamido-5-methoxy-6-methyl-3-nitrobenzoate
[00494] To a stirred solution of methyl 6-acetamido-3-methoxy-2-methylbenzoate (1.2 g, 5.06 mmol) in TFA (20 mL, 15 vol) was added KNO3 (0.51 g, 5.06 mmol) at RT and the resulting mixture was stirred at 55 0C for 30 min. The volatiles were evaporated under reduced pressure. The reaction mass was diluted with water (15 mL), the pH was adjuste to ~8 with a saturated NaHCO3 solution and extracted with dichloromethane (2x 30 mL). The combined organic layer was washed with water, brine, dried over anhydrous Na2SO4 and concentrated under reduced pressure. The crude material was purified by column chromatography to afford the title compound (1.1 g, 78.5% yield) as a brown solid. 1H NMR (200 MHz,
CDCI3): δ 8.60 (bs, 1 H), 7.53 (s, 1 H), 3.92 (s, 6H), 2.30 (s, 3H), 2.16 (s, 3H). Mass = 283 (M +1). Step 5. methyl 2-amino-5-methoxy-6-methyl-3-nitrobenzoate
[00495] A solution of methyl 2-acetamido-5-methoxy-6-methyl-3-nitrobenzoate (1.1 g, 3.90 mmol) in cone. HCI (5 mL) was stirred at 90 0C for 4h. The reaction mass was adjusted to pH~8 with a saturated
NaHCO3 solution and extracted with EtOAc (2x 30 mL). The combined organic layer was washed with water, brine, dried over anhydrous Na2SO4 and concentrated under reduced pressure to afford the title compound (750 mg, 80.1%) as a brick red solid. 1H NMR (200 MHz, CDCI3): δ 7.68 (s, 1H), 6.97 (bs, 2H),
3.96 (s, 3H), 3.83 (s, 3H), 2.29 (s, 3H).
Step 6. methyl 2,3-diamino-5-methoxy-6-methylbenzoate
[00496] A solution of methyl 2-amino-5-methoxy-6-methyl-3-nitrobenzoate (750 mg, 3.125 mmol),
10% Pd/C (375 mg) in EtOAc (10 mL) was placed in an autoclave reactor and was stirred under hydrogen pressure (40 psi) at RT for 2h. After the consumption of the starting material (by TLC), the reaction mixture was filtered through a celite bed, rinsing with EtOAc. The filtrate was washed with water, dried over Na2SO4 and evaporated under reduced pressure to afford the title compound (500 mg, 76.2 % yield) as a brown solid. 1H NMR (200 MHz, CDCI3): δ 6.45 (s, 1 H), 3.91 (s, 5H), 3.73 (s, 5H), 2.18 (s, 3H). LC-
MS (10-90[4]-10 method): Rt = 2.99 min. (91%); m/z = 211.3 (M+1).
Step 7. methyl 6-methoxy-5-methyl-1 H-benzo[d][1 ,2,3]triazole-4-carboxylate
[00497] To a stirred solution of methyl 2,3-diamino-5-methoxy-6-methylbenzoate (350 mg, 1.66 mmol) in AcOH (5 mL) was added sodium nitrite (6M, 126 mg, 1.83 mmol) at 0 0C. The reaction mixture was stirred for 1 h at same temperature. The reaction mass was diluted with water and basified with a saturated NaHCO3 solution (pH~ 8). The aqueous layer was extracted with EtOAc, and the combined organic extracts were washed with water, brine, dried over anhydrous Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography to afford the title compound (6 mg, 50%). 1H NMR (200 MHz, CDCI3): δ 12.79 (bs, 1 H), 7.60 (s, 1 H), 4.08 (s, 3H), 3.96 (s,
3H), 2.70 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.23 min. (97%); m/z = 222.4 (M+1).
Step 8. (6-methoxy-5-methyl-1H-benzo[d][1,2,3]triazol-4-yl)methanol
[00498] To a stirred solution of methyl 6-methoxy-5-methyl-1 H-benzo[d][1 ,2,3]triazole-4- carboxylate (200 mg, 0.904 mmol) in THF (5 mL) was added DIBAL-H (1.8 mL, 2.71 mmol of 1.5M in
Toluene) at 0 °C. The resulting reaction mixture was stirred at RT for 3h. The reaction mixture was quenched with MeOH (-3 mL) and diluted with sodium tartarate (10 mL) solution. The volatiles were evaporated under reduced pressure and the residue was diluted with water and extracted with EtOAc (3x
25 mL). The combined organic layers were dried over Na2SO4. The solvent was evaporated under reduced pressure to afford the title compound (140 mg, 80%, crude) as a brown solid. 1H NMR (200 MHz,
DMSO-cfe): δ 7.18 (s, 1 H), 5.20 (bs, 1 H), 4.92 (s, 2H), 3.87 (s, 3H), 2.26 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 1.97 min. (98%); m/z = 194 (M+1).
Step 9. 4-(chloromethyl)-6-methoxy-5-methyl-1H-benzo[d][1,2,3]triazole
[00499] To a stirred solution of (6-methoxy-5-methyl-1 H-benzo[d][1 ,2,3]triazol-4-yl)methanol (150 mg, 0.77 mmol) in CHCI3 (5 mL) was added thionyl chloride (0.24 mL, 1.88 mmol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 3h. The volatiles were evaporated under reduced pressure and the residue was diluted with saturated NaHCO3 solution (pH~ 8). The aqueous layer was extracted with EtOAc (2x 50 ml_). The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure. The crude material was purified by silica gel column chromatography to afford the title compound (120 mg, 73% yield) as an off-white solid. 1H NMR (500 MHz, DMSO-d6): δ 14.60 (bs, 1 H), 7.13 (s, 1H), 5.25 (s, 1H), 3.90 (s, 3H), 2.30 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.42 min. (96%); m/z = 212.2 (M+1).
Step 10. 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((6-methoxy-5-methyl-1H- benzo[d][1,2,3]triazol-4-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine [00500] To a stirred solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2 -amine (35 mg, 0.111 mmol) in DMF (2 mL) were added 4-(chloromethyl)-6- methoxy-5-methyl-1 H-benzo[d][1 ,2,3]triazole (28 mg, 0.0.133 mmol) and NaH (6.5 mg, 0.16 mmol) at 0 0C under anhydrous conditions. The reaction mass was quenched with ice water and extracted with EtOAc (2x 20 mL). The organic layer was washed with water, brine and dried over Na2SO4. The solvent was evaporated under reduced pressure to afford the title compound (50 mg, 92.5%, crude) as an off- white solid. 1H NMR (500 MHz, DMSO-cfe): δ 15.40 (bs, 1 H), 7.14 (d, J = 6.8 Hz, 1 H), 6.54 (bs, 2H), 5.12 (bs, 2H), 4.98 (bs, 1 H), 3.98 (s, 3H), 3.62- 3.60 (m, 1 H), 3.30 (s, 3H), 2.18 (s, 3H), 0.65 (s, 9H), -0.90 (s, 3H), -0.88 (s, 3H). Mass: 490 (M+1 ).
Step 11. (2-amino-4-chloro-7-((6-methoxy-5-methyl-1H-benzo[d][1,2,3]triazol-4-yl)methyl)-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XC)
[00501] To a stirred solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((6-methoxy-5- methyl-1 H-benzo[d][1 ,2,3]triazol-4-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (45 mg, 0.092 mmol) in THF (1 mL) was added TBAF (0.2 mL, 0.184 mmol, 1 M in THF) at 0 °C under anhydrous conditions. The reaction mixture was stirred at 10 °C for 30 min. After completion of reaction (by TLC), the volatiles were evaporated under reduced pressure and the residue was diluted with water (50 mL). The aqueous layer was extracted with EtOAc (2x 50 mL). The organic layer was washed with water, brine and dried over Na2SO4. The solvent was evaporated under reduced pressure and the crude material was purified by preparative HPLC to afford the title compound (18 mg, 53% yield) as an off-white solid. 1H NMR (500 MHz, DMSO-Of6): δ 7.18 (bs, 1 H), 6.97 (bs, 2H), 5.09 (d, J = 8.0 Hz, 1 H), 4.89 (d, J = 8.0 Hz, 1 H), 4.60 (bs, 2H), 3.91 (s, 3H), 3.78- 3.74 (m, 1 H), 3.38- 3.35 (m, 1 H), 3.25- 3.23 (m, 1 H), 2.97 (bs, 2H), 2.21 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.78 min. (97%); m/z = 376.5 (M+1). Example 87
(2-amino-4-chloro-7-(5,6,7,8-tetrahydroquinolin-5-yl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)methaπol (XCI)
Step 1. 5,6,7,8-tetrahydroquinolin-5-ol
[00502] To a stirred solution of 7,8-dihydroquinolin-5(6H)-one (300 mg, 2.038 mmol) in MeOH (5 mL) was added portionwise NaBH4 (155 mg, 4.076 mmol) at 0 0C. The reaction mixture was stirred at RT for 5h. The reaction mass was quenched with cold water, and the volatiles were evaporated under reduced pressure. The crude was extracted with EtOAc (3x 25 ml_). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (250 mg, 82% yield). 1H NMR: (200 MHz, CDCI3): δ 8.44 (d, J = 3.4 Hz, 1 H), 7.79 (d, J = 7.8 Hz, 1 H), 7.17 (t, J = 7.8 Hz,
1H), 4.82 (bs, 1H), 2.97 (t, J = 4.8 Hz, 2H), 2.10- 1.99 (m, 4H), 1.93- 1.86 (m, J= 4 Hz, 1H). Mass: 150
(M+1).
Step 2. 5-chloro-5, 6, 7, 8-tetrahydroquinoline
[00503] To a stirred solution of 5,6,7, 8-tetrahydroquinolin-5-ol (200 mg, 1.34 mmol) in CHCI3 (10 ml.) was added thionyl chloride (292 ml_, 4.026 mmol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 5h. The volatiles were evaporated under reduced pressure and the crude material was diluted with saturated NaHCO3 solution. The aqueous layer was extracted with CHCI3 (2 x
15 mL), and the combined organic layers were washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure to afford the title compound (150 mg, 67% yield) as a colorless liquid. 1H NMR: (200 MHz, CDCI3): δ 8.48 (d, J = 1.6 Hz, 1 H), 7.71 (d, J = 8.0 Hz, 1 H), 7.17- 7.14 (m, 1 H),
5.24- 5.21 (m, 1 H), 3.06- 2.90 (m, 2H), 2.30- 2.21 (m, 3H), 2.02- 1.94 (m, 1 H).
Step 3. (2-amino-4-chloro-7-(5, 6, 7,8-tetrahydroquinolin-5-yl)-6, 7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)methanol (XCI)
[00504] To a stirred solution of NaH (5.5 mg, 0.114 mmol) in DMF (1 mL) was added 5-((tert- butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (30 mg, 0.095) in
DMF (1 mL) at 0 0C. After being stirred at RT for 5 min, 5-chloro-5,6,7,8-tetrahydroquinoline (19.1 mg,
0.114 mmol) was added to the reaction mixture at RT and stirring was continued for an additional 4 h.
After completion of reaction (by TLC), the reaction mixture was quenched with water and extracted with
EtOAc. The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting crude was purified by preparative TLC eluting with 6% MeOH/dichloromethane to afford the title compound as a 1 :1 mixture of diastereomers (10 mg, 32% yield). 1H NMR (500 MHz,
DMSO-cfe): δ 8.39 (s, 1 H), 7.46 (d, J= 8.0 Hz, 1 H), 7.35 (d, J= 8.0 Hz, 1 H), 7.18 (t, J = 8.0 Hz, 1 H), 6.45
(s, 2H), 5.33 (bs, 1H), 4.83- 4.80 (m, 1H), 3.63- 3.60 (m, 2H), 3.40- 3.20 (m, 2H), 3. 19- 3.16 (m, 1H),
2.85- 2.81 (m, 2H), 2.02- 1.85 (m, 1 H), 1.99- 1.96 (m, 3H). LC-MS (10-90[4]-10 method): Rt = 2.69 min.
(52%); m/z = 331 (M+1) and Rt = 2.76 (44%); m/z = 331 (M+1 ) (-1 :1 mix of diastereomers).
Example 88
(2-amino-4-chloro-7-((6-methoxy-5-methyl-1H-benzo[d]imidazol-4-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XCII)
Step 1. methyl 6-methoxy-5-methyl-1H-benzo[d]imidazole-4-carboxylate
[00505] To a stirred solution of methyl 2,3-diamino-5-methoxy-6-methylbenzoate (250 mg, 1.19 mmol) in 4N HCI (3 mL) was added HCO2H (0.22 mL, 5.95 mmol) at RT. The reaction mixture was then refluxed for 2h. After consumption of the starting material (monitored by TLC), the reaction mixture was brought to RT and was diluted with water and basified with a saturated NaHCO3 solution (to pH ~8). The aqueous layer was extracted with EtOAc (2 x 15ml) and the combined organic extracts were washed with water, brine and dried over anhydrous Na2SO4. The solvent was removed under reduced pressure and the crude material was purified by silica gel column chromatography to afford the title compound (180 mg,
69% yield) as a brown solid. 1H NMR (200 MHz, CDCI3): δ 10.45 (bs, 1 H; Exc), 8.01 (s, 1H), 7.50 (s, 1 H),
4.02 (s, 3H), 3.91 (s, 3H), 2.63 (s, 3H).
Step 2. (6-methoxy-5-methyl-1H-benzo[d]imidazol-4-yl)methanol
[00506] To a stirred solution of methyl 6-methoxy-5-methyl-1H-benzo[d]imidazole-4-carboxylate
(150 mg, 0.68 mmol) in dry THF (5 mL) was added DIBAL-H (1.8 mL, 2.72 mmol of 1.5M in Toluene) at 0
0C. The reaction mixture was brought to RT and stirred for 3 h. After consumption of the starting material
(monitored by TLC), the reaction mixture was quenched with MeOH (~2 mL) and diluted with sodium potassium tartarate solution (5 mL). The volatiles were removed under reduced pressure and the residue was diluted with water and extracted with EtOAc (3 x 20 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (90 mg, 70% yield, crude) as a brown solid. 1H NMR (200 MHz, DMSO-cfe): δ 12.06 (bs, 1 H; Exc), 7.99 (s, 1 H), 7.09 (s, 1 H),
5.10-5.07 (m, 1 H; Exc), 4.92-4.90 (m, 1 H), 4.79- 4.77 (m, 1 H), 3.80 (s, 3H), 2.23 (s, 3H).
Step 3. 4-(chloromethyl)-6-methoxy-5-methyl- 1 H-benzo[d]imidazole
[00507] To a stirred solution of (6-methoxy-5-methyl-1 H-benzo[d]imidazol-4-yl)methanol (85 mg,
0.44 mmol) in CHCI3 (3 mL) was added SOCI2 (0.14 mL, 1.77 mmol) at 00C under anhydrous conditions and the reaction was slowly brought to RT and stirred for 3h. After consumption of the starting material
(monitored by TLC), the volatiles were removed under reduced pressure and the residue was diluted with
H2O. The aqueous layer was extracted with EtOAc (2 x 50 mL). The combined organic extracts were washed with brine, dried over anhydrous Na2SO4 and concentrated in vacuo. The crude material was purified by silica gel column chromatography to afford the title compound (60 mg, 64.5%) as a brown solid. 1H NMR (200 MHz, DMSO-cfe): δ 12.41 (bs, 1 H; Exc), 8.09 (s, 1 H), 7.09 (s, 1 H), 5.13 (s, 2H), 3.82
(s, 3H), 2.27 (s, 3H).
Step 4. 1-(4-(chloromethyl)-6-methoxy-5-methyl-1H-benzo[d]imidazol-1-yl)ethanone
[00508] To a stirred solution of 4-(chloromethyl)-6-methoxy-5-methyl-1 H-benzo[d]imidazole (100 mg, 0.47 mmol) in CHCI3 (5 mL) was added AcCI (0.03 mL, 0.52 mmol) at O 0C under inert atmosphere.
The reaction mixture was brought to RT and stirred for 2h. After consumption of the starting material
(monitored by TLC), the reaction mixture was diluted with dichloromethane. The organic layer was washed with NaHCO3 solution, brine (20 mL), dried over anhydrous Na2SO4, and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography to afford the title compound (95 mg, 80%) as an off-white solid. 1H NMR (200 MHz, DMSO-cfe): δ 8.74 (s, 1H), 7.70 (s, 1 H),
5.13 (s, 2H), 3.85 (s, 3H), 2.75 (s, 3H), 2.30 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 4.05 min. (84%); m/z = 253.4 (M+1).
Step 5. i^-ffΣ-aminchS-tftert-butyldimethylsilyloxyJmethyl^-chloro-SH-pyrrolofcS-dJpyrimidin-
7(6H)-yl)methyl)-6-methoxy-5-methyl-1H-benzo[d]imidazol-1-yl)ethanone [00509] To a stirred solution of 1 -(4-(chloromethyl)-6-methoxy-5-methyl-1 H-benzo[d]imidazol-1 - yl)ethanone (10 mg, 0.03 mmol) in DMF (1 mL) was added NaH (1.5 mg, 0.03 mmol) at 0 0C under anhydrous conditions. After 10 min, a solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (8.8 mg, 0.03 mmol) in DMF (0.3 mL) was added, and the reaction mixture and further stirred for 3 min. The reaction mixture was quenched with ice cold water and extracted with EtOAc (2 x 5 mL). The combined organic extracts were washed with water, brine and dried over Na2SO4. The crude material was purified by silica gel column chromatography to yield the title compound (3 mg, 17.8%). Mass: 531 (M+ 1).
Step 6. (2-amino-4-chloro-7-((6-methoxy-5-methyl-1H-benzo[d]imidazol-4-yl)methyl)-6, 7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XCII)
[00510] To a stirred solution of 1-(4-((2-amino-5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-5H- pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-6-methoxy-5-methyl-1 H-benzo[d]imidazol-1 -yl)ethanone (15 mg, 0.02 mmol) in THF (1 mL) was added TBAF (0.05 mL, 0.05 mmol, 1M in THF) at 0 0C under inert atmosphere. The reaction mixture was stirred at 0 °C for 30 min. After consumption of the starting material (monitored by TLC), the volatiles were evaporated under reduced pressure. The residue was diluted with water (3 mL) and extracted with EtOAc (2 x 5 mL). The combined organic extracts were washed with water and brine, and dried over Na2SO4. The solvent was evaporated under reduced pressure and the crude material was purified by preparative TLC to afford the title compound (4 mg, 40%) as an off-white solid. 1H NMR (500 MHz, CD3OD): δ 8.06 (s, 1H), 7.12 (s, 1 H), 5.01 (bs, 2H), 3.89 (s, 3H), 3.76-3.73 (m, 1 H), 3.48-3.46 (m, 4H), 2.32-2.30 (m, 3H). LC-MS (10-90[4]-10 method): Rt = 1.32 min. (81%); m/z = 375.5 (M+1). Example 89
(2-amino-4-chloro-7-((4-methyl-6,7-dihydro-5H-cyclopenta[c]pyridin-3-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XCIII)
Step 1. W-methyl-βJ-dihydroSH-cyclopentalcJpyridin-S-ytymethyl acetate [00511] To a stirred solution of 3,4-dimethyl-6,7-dihydro-5H-cyclopenta[c]pyridine (Tius, M. A.;
Thurkauf, A.; Truesdell, J. W. Aromatic annulation. I. Synthesis of pyridines. Tet. Lett. 1983, 23, 2819- 2822) (200 mg, 1.36 mmol) in CHCI3 (20 mL) was added m-CPBA (352 mg, 2.04 mmol) portionwise over a period of 5 min at 0 0C under N2 atmosphere and stirred for 30 min. The reaction mixture was allowed to warm to RT and stirred for 12 h. The volatiles were evaporated under vacuum, diluted with water (20 mL) and basified to pH -8 using saturated NaHCO3 solution. The aqueous layer was extracted with CH2CI2 (2 x 35 mL). The combined organic extracts were dried over anhydrous Na2SO4 to obtain 3,4-dimethyl-6,7- dihydro-5H-cyclopenta[c]pyridine 2-oxide (200 mg). A mixture of 3,4-dimethyl-6,7-dihydro-5H- cyclopenta[c]pyridine 2-oxide (200 mg, 1.22 mmol) in Ac2O (15 mL) was refluxed for 12 h. Upon completion of the reaction (by TLC), the reaction mass was allowed to cool to RT and concentrated under reduced pressure to afford the title compound (200 mg, crude) as brown color syrup. Mass: 206 (M+1). Step 2. (4-methyl-6, 7-dihydro-5H-cyclopenta[c]pyridin-3-yl)methanol [00512] To a stirred solution of (4-methyl-6,7-dihydro-5H-cyclopenta[c]pyridin-3-yl)methyl acetate
(200 mg, 0.97 mmol) in MeOH: H2O (15 mL, 2: 1) was added K2CO3 (269 mg, 1.95 mmol) at 0 0C and stirred for 12 h at RT. After completion of the reaction (by TLC), the precipitated solid was filtered off and the filtrate was concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 3-4% MeOH/CH2CI2 to afford the title compound (100 mg, 62 %) as a light green thick syrup. 1H NMR (200 MHz, CDCI3): δ 8.24 (s, 1 H), 4.65 (s, 2H), 2.99-2.82 (m, 4H), 2.12 (s, 3H), 2.08- 2.04 (m, 3H). Mass: 164 (M+1 ).
Step 3. 3-(chloromethyl)-4-methyl-6, 7-dihydro-5H-cyclopenta[c]pyridine
[00513] To a stirred solution of (4-methyl-6,7-dihydro-5H-cyclopenta[c]pyridin-3-yl)methanol (100 mg, 0.61 mmol) in dry CH2CI2 (20 mL) was added thionyl chloride (0.13 mL, 1.84 mmol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 4h. Upon completion of the reaction (By TLC), the reaction mass was diluted with CH2CI2 (50 mL) and washed with saturated NaHCO3 solution (35 mL). The organic layer was dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (70 mg, 63%) as a brown, thick syrup. This material was used for the next step without further purification. 1H NMR (200 MHz, CDCI3): δ 8.28 (s, 1 H), 4.78 (s, 2H), 3.02- 2.87 (m, 4H), 2.37 (s, 3H), 2.19- 2.11 (m, 2H). Mass: 182 (M+1 ).
Step 4. 5-{(tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((4-methyl-6, 7-dihydro-5H- cyc\openta[c]pyr\d\n-3-y\)methy\}-6,7-d\hydro-5H-pyrrolo[2,3-d]pyrim\dm-2-amm' e [00514] To a stirred suspension of NaH (6.0 mg, 0.15 mmol) in dry DMF (50 mL) was added a solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (50 mg, 0.15 mmol) in dry DMF (10 mL) followed by a solution of 3-(chloromethyl)-4-methyl-6,7-dihydro- 5H-cyclopenta[c]pyridine (28 mg, 0.15 mmol) in dry DMF slowly at 0 0C. After stirring at RT for 12h, the reaction mixture was diluted with water and extracted with EtOAc (2 x 30 mL). The combined organic extracts were washed with water, dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 3-4% MeOH/dichloromethane to afford the title compound (50 mg, 68%) as an off-white solid. 1H NMR (500 MHz, DMSO-d6): δ 8.17 (s, 1H), 6.41 (s, 2H), 4.77 (d, J = 8.0 Hz, 1 H), 4.41 (d, J = 8.0 Hz, 1 H), 3.74- 3.71 (m, 1 H), 3.55- 3.46 (m, 2H), 3.30- 2.89 (m, 2H), 2.88- 2.73 (m, 4H), 2.18 (s, 3H), 2.05- 2.01 (m, 2H), 1.33 (s, 9H), 0.86 (s, 6H). LC-MS (10-90[4]-10 method): Rt = 4.31 min. (89%); m/z = 460.9 (M+1). Step 5. (2-amino-4-chloro-7-((4-methyl-6, 7-dihydro-5H-cyclopenta[c]pyridin-3-yl)methyl)-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (XCIII)
[00515] To a stirred solution of 5-((tert-butyldimethylsilyloxy)methyl)-4-chloro-7-((4-methyl-6,7- dihydro-SH-cyclopentatclpyridin-S-yOmethylJ-ej-dihydro-SH-pyrrolo^.S-dJpyrimidin^-amine (50 mg, 0.108 mmol) in dry THF (5 mL) was added TBAF (0.48 mL, 0.43 mmol) at 0 °C under anhydrous conditions. The reaction mixture was stirred at RT for 4h. Upon completion of the reaction (By TLC), the volatiles were evaporated under reduced pressure and the residue was diluted with water (10 mL) and extracted with EtOAc (2 x 20 mL). The combined organic extracts were washed with water, dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 4-5% MeOH/CH2CI2 to afford the title compound (28 mg, 75%) as an off- white solid. 1H NMR (500 MHz, DMSO-cfe): δ 8.17 (s, 1 H), 6.38 (s, 2H), 4.82- 4.80 (m, 1 H)1 4.68 (s, 1 H), 4.51 (s, 1H), 3.66- 3.53 (m, 1 H), 3.53- 3.49 (m, 1 H), 3.34- 3.28 (m, 2H), 3.23- 3.20 (m, 1H), 3.19- 2.82 (m, 4H), 2.18 (s, 3H)1 2.06- 1.98 (m, 2H). LC-MS (10-90[4]-10 method): Rt = 2.98 min. (97%); m/z = 346.7 (M+1 ). Example 90 methyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl)acetate (XCIV) Step 1. tert-butyl 2-amino-6-chloropyrimidin-4-ylcarbamate
[00516] To a stirred solution of 6-chloropyrimidine-2,4-diamine (10 g, 69.4 mmol) in DMF (100 ml.) was added potassium tertiary butoxide (7.7 g, 69.4 mmol) followed by (BOc)2O (15.1 g, 69.4 mol) at rt under inert atmosphere. The mixture was stirred for 16 h. The reaction mixture was diluted with water and extracted with EtOAc (4 x 50 ml_). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 50% EtOAc/Hexane to afford the title compound (5.0 g, 30%). 1H NMR (500 MHz, DMSO-cfe): δ 10.0 (s, 1H), 7.01 (s, 1 H), 6.81 (s, 2H), 1.45 (s, 9H). LC-MS (20-90[10]-20 method): Rt = 9.13 min. (97%); m/z = 245 (M+1).
Step 2. (E)-methyl 4-((2-amino-6-chloropyrimidin-4-yl)(tert-butoxycarbonyl)amino)but-2-enoate [00517] To a stirred solution of tert-butyl 2-amino-6-chloropyrimidin-4-ylcarbamate (2.0 g, 8.23 mmol) in acetone (10 mL) was added K2CO3 (0.56 g, 4.05 mmol) followed by methyl-4-bromo crotonate (2.2 g, 12.2 mmol) slowly at RT under N2 atmosphere. The resulting reaction mixture was heated to 80 0C for 8 h. The volatiles were evaporated under reduced pressure and the crude material was purified by neutral alumina column chromatography eluting with ether/Hexane followed by CH2CI2 to afford the title compound (2.0 g, 71%) as a white solid. 1H NMR (500 MHz, CDCI3): δ 7.46 (s, 1 H), 6.98- 6.92 (m, 1 H), 5.85 (d, J = 15 Hz, 1 H), 4.99 (s, 2H), 4.72 (d, J = 5 Hz, 2H), 3.73 (s, 3H), 1.52 (s, 9H). Step 3. (E)-methyl 4-((2-amino-5-bromo-6-chloropyrimidin-4-yl)(tert-bυtoxycarbonyl)amino)but-2- enoate
[00518] To a stirred solution of (E)-methyl 4-((2-amino-6-chloropyrimidin-4-yl)(tert- butoxycarbonyl)amino)but-2-enoate (1 g, 2.91 mmol) in EtOH (15 mL) was added NaOAc (0.23 g, 2.92 mmol) at rt under nitrogen atmosphere. The resulting reaction mixture was allowed to cool to 0 0C; bromine (0.15 mL, 2.92 mmol) was added to the reaction mixture and stirring was continued for another 5 min. After completion of the reaction (by TLC), the volatiles were evaporated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 30% EtOAc/ Hexane to afford the title compound (1.0 g, 80%) as a colorless semi solid. 1H NMR (500 MHz, CDCI3): δ 6.98- 6.92 (m, 1 H), 6.01 (d, J= 16 Hz, 1 H), 5.13 (s, 2H), 4.45 (q, 2H), 3.73 (s, 3H), 1.51 (s, 9H). Step 4. (R)- and (S)- tert-butyl 2-amino-4-chloro-5-(2-methoxy-2-oxoethyl)-5H-pyrrolo[2,3- d]pyrimidine-7(6H)-carboxylate
[0098] A mixture of (E)-methyl 4-((2-amino-5-bromo-6-chloropyrimidin-4-yl)(tert- butoxycarbonyl)amino)but-2-enoate (1.0 g, 3.36 mmol) in dry benzene (15 mL) was degassed with argon gas for 30 min. A solution of (Me3Si)3SiH (1.4 mL, 4.73 mmol) and AIBN (0.77 g, 4.73 mmol) in dry benzene was degassed with argon for 15 min and the mixture was added to the reaction mixture at 0 0C. The resulting reaction mixture was stirred at 90 0C for 16-30 h. The volatiles were evaporated under vacuum, diluted with water and extracted with EtOAc (2 x 50 mL). The organic layer was dried over anhydrous Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 30% EtOAc/Hexane to obtain the title compound as a racemate (500 mg, 71.4%). 1H NMR (500 MHz, DMSO-d6): δ 6.85 (s, 2H), 4.02 (t, J = 10 Hz, 1 H), 3.65 (d, J = 4.5 Hz, 1 H), 3.60 (S1 3H), 3.48- 3.44 (m, 1 H), 2.80 (d, J = 3.5 Hz, 1 H), 2.66- 2.60 (m, 1 H), 1.47 (s, 9H). LC- MS (20-90[10]-20 method): Rt = 9.30 min. (98); m/z = 343 (M+1). The two enantiomers could be separated by supercritical fluid HPLC using a Chiralpak IC (3x15 cm) column and eluting with 50% MeOH (0.1% diethylamine)/CO2 100 bar, 95 mUmin to give the (R) enantiomer (fast-running) and the (S)- enantiomer (slow-running).
Step 5. (R)- and (S)- methyl 2-(2-amino-4-chloro-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)acetate
[00519] Racemic tert-butyl 2-amino-4-chloro-5-(2-methoxy-2-oxoethyl)-5H-pyrrolo[2,3- d]pyrimidine-7(6H)-carboxylate (8.0 g, 23.3 mmol) was added to TFA:dichloromethane (50 mL, 1 : 4) at 0 0C and stirred at RT for 4 h. The volatiles were evaporated under reduced pressure. The residue was quenched with ice and saturated NaHCO3 solution at 0 0C, and the aqueous layer was extracted with EtOAc (2 x 100 mL). The organic layer was dried over Na2SO4 and concentrated under reduced pressure. The crude material was washed with n-hexane and dried under vacuum to afford the title compound (5.5 g, 89.2%) as an off-white solid. 1H NMR (500 MHz, DMSO-cfe): δ 7.26 (s, 1 H)1 6.33 (s, 2H), 3.66 (X1 J = IO Hz1 1 H), 3.60 (s, 3H), 3.52- 3.47 (m, 1 H), 3.20 (d, J = 4.5 Hz, 1 H)1 2.76 (d, J = 3.5 Hz1 1 H)1 2.40 (d, J = 10.5 Hz, 1H). LC-MS (10-90[4]-10 method): Rt = 2.79 min. (98%); m/z = 243.4 (M+1). The two enantiomers could be separated by supercritical fluid HPLC using a Chiralpak AD-H (2x15 cm) column and eluting with 25% iPrOH (0.1% diethylamine)/CO2 100 bar, 65 mL/min to give the (R) enantiomer (fast- running) and the (S)-enantiomer (slow-running).
Step 6. (R)- and (S)- methyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)- 6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (XCIV)
[0099] To a stirred suspension of NaH (2.0 g, 43.21 mmol) in DMF (50 mL) was added methyl 2-
(2-amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (5.0 g, 20.57 mmol) at 00C. After being stirred for 5 min, 2-chloromethyl-3,5-dimethyl-4-methoxy-pyridine hydrochloride (4.56 g, 20.57 mmol) was added at 0 0C and stirred at RT for 2 h. The reaction mixture was quenched with water (200 mL) and extracted with EtOAc (3 x 50 mL). The organic extract was washed with water (50 mL), brine, dried over anhydrous Na2SO4 and concentrated under vacuo. The crude material was purified by recrystallization using EtOAc/Hexane to afford the title compound (4.7 g, 58%); 1H NMR (DMSO-cfe, 500 MHz): δ 8.14 (s, 1 H), 6.44 (s, 2H), 4.56 (s, 2H), 3.73- 3.66 (m, 1 H), 3.71 (s, 3H), 3.56 (s, 3H), 3.50- 3.46 (m, 1 H), 3.19- 3.16 (m, 1H), 2.84- 2.80 (m, 1 H), 2.52- 2.50 (m, 1 H), 2.18 (s, 3H), 2.16 (s, 3H); LC-MS (10- 90-3 method, Acquity BEH-C18 50 x 2.1 mm, 1.7 μM), purity: 97.7%, rt = 1.47 min, m/z = 392 (M+1). TLC system: 80% EtOAc/Hexane, R(: 0.3. The two enantiomers could be separated by supercritical fluid HPLC using a Chiralpak AS-H (2x15 cm) column and eluting with 35% MeOH (0.1% diethylamine)/CO2 100 bar, 60 mL/min to give the (R) enantiomer (fast-running) and the (S)-enantiomer (slow-running). Example 91
4-Chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-5-piperidin-4-yl-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-ylamine (XCV)
Step 1. 5-Bromo-6-chloro-N(4)-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-pyrimidine-2,4- diamine
[00520] 5-Bromo-6-chloro-pyrimidine-2,4-diamine (Kosogof, C; Liu, B.; Liu, G.; Nelson, L.T.J. ;
Serby, M. D.; Sham, H. L.; Szczepankiewicz, B. G.; Xin, Z.; Zhao, H., Preparation of diaminopyrimidines as growth hormone secretagogue receptor (GHS-R) antagonists. U.S. patent 2005171131 , Aug 4, 2005) (2.420 g, 10.83 mmol) and 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine, hydrochloride salt (2.40 g, 10.8 mmol) in dimethyl sulfoxide (200 mL) was treated with sodium hydride (624 mg, 26.0 mmol) and let stir at room temperature under a nitrogen balloon. After 12 hours the reaction was diluted with water and the resulting solid was collected by vacuum filtration. The solid was then washed with ether and dried in vacuoto provide 1.764 g (38% yield) of the title compound. 1H NMR (400 MHz, DMSO-cfe) δ ppm 8.25 (s, 1 H), 7.51 (t, J= 4.2 Hz, 1 H), 6.73 (s, 2H), 4.51 (d, J= 4.0 Hz, 2H), 3.74 (s, 3H), 2.22 (s, 3H), 2.17 (s, 3H). LC-MS (5-100-5 method): Rt = 2.32 (86% HPLC purity) m/z = 371.9 (M+1 ). Step 2. 4-{2-[(2-Amino-5-bromo-6-chloro-pyrimidin-4-yl)-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-amino]-ethylidene}-piperidine-1-carboxylic acid tert-butyl ester [00521] 5-Bromo-6-chloro-N(4)-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-pyrimidine-2,4- diamine (467 mg, 1.25 mmo) in dimethyl sulfoxide (2.0 mL) was treated with sodium hydride (50.6 mg, 2.00 mmol). After stirring for 10 minutes a solution of 4-(2-Chloro-ethylidene)-piperidine-1-carboxylic acid tert-butyl ester (Barba, O.; Bradley, S. E.; Fyfe, M.C.T.; Bertram, LS.; Gatrell W.; Procter, M.J.; Rasamison, CM.; Swain, S.A., N-arylpiperazine derivatives as G-protein coupled receptor agonists and their preparation, pharmaceutical compositions, and use in the treatment of obesity and diabetes. PCT Int. App. WO 2007003964, Jan 11 , 2007) (308 mg, 1.25 mmol) in dimethyl sulfoxide (1.6 mL) was added via syringe. After 1 hour an additional 30 mg of Sodium hydride was added. After 18 hours the reaction was diluted with ethyl acetate, washed with water, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (gradient of 20-100% ethyl acetate/hexanes) to provide 148 mg (20% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm
8.19 (s, 1H), 5.38 (t, J= 6.9 Hz, 1 H), 4.80 (br. s., 2H), 4.75 (s, 2H), 4.25 (d, J= 6.8 Hz, 2H), 3.76 (s, 3H),
3.20 - 3.44 (m, 4H), 2.25 (s, 3H), 2.15 (s, 3H), 2.10 (dt, J= 19.8, 5.6 Hz, 4H), 1.45 (s, 9H). LC-MS (5-100- 5 method): Rt = 3.21 m/z = 581.1 (M+1).
Step 3. 4-Chloro-7-(4-methoxy-3, 5-dimethyl-pyridin-2-ylmethyl)-5-piperidin-4-yl-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-ylamine (XCV)
[00522] 4-{2-[(2-Amino-5-bromo-6-chloro-pyrimidin-4-yl)-(4-πnethoxy-3,5-dinnethyl-pyridin-2- ylmethylj-aminoj-ethylidenej-piperidine-i-carboxylic acid tert-butyl ester (49 mg, 0.067 mmol) in benzene (5.1 ml_) was treated with tri-n-butyltin hydride (19.0 μM, 0.0707 mmol) followed by 1.00 M of triethylborane in hexane (70.7 μM, 0.0707 mmol) and let stir open to the atmosphere. After 2.5 hours the reaction was condensed. The crude material was then purified by flash chromatography (50-100% ethyl acetate/hexanes gradient) to provide 12 mg (35% yield) of 4-[2-Amino-4-chloro-7-(4-methoxy-3,5- dimethyl-pyridin^-ylmethylJ-θ^-dihydro-SH-pyrrolo^.S-dJpyrimidin-S-yll-piperidine-i-carboxylic acid tert- butyl ester. 4-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-piperidine-1-carboxylic acid tert-butyl ester (12 mg, 0.024 mmol) in 1 ,2- dichloroethane (1.0 mL) was let stir with trifluoroacetic Acid (18.4 μM, 0.238 mmol) under a nitrogen balloon. After 1 hour the reaction was heated at 40 0C. After 18 hours the reaction was heated at 50 0C. After 24 hours an additional 0.0184 mL of trifluoroacetic acid was added. After 2 days the reaction was condensed. The crude material was then purified by preparative HPLC (mobile phase: acetonitrile/water with 0.1% TFA buffer, gradient of 5-95%). The HPLC fractions containing the desired compound were condensed. The resulting residue was then taken up in dichloromethane, washed with sat. aq. NaHCO3, dried over Na2SO4, filtered and condensed to yield 3.9 mg (39% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.18 (s, 1 H), 4.71 - 4.87 (m, 3H), 4.46 (d, J= 14.7 Hz, 1H), 3.75 (S, 3H), 3.39 - 3.53 (m, 1 H), 3.29 - 3.37 (m, 1 H), 3.18 - 3.27 (m, 1 H), 2.95 - 3.13 (m, 2H), 2.45 - 2.69 (m, 2H), 2.26 (s, 3H), 2.21 (s, 3H), 2.00 - 2.15 (m, 1 H), 1.45 - 1.57 (m, 1H), 1.14 - 1.39 (m, 2H), 0.98 - 1.13 (m, 1 H). LC-MS (5-100-5 method): Rt = 2.32 min. (96% HPLC purity); m/z = 403.1 (M+1). Example 92
4-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)piperidine-1 -sulfonamide (XCVI)
[00523] In a flame-dried flask under nitrogen was stirred 4-Chloro-7-(4-methoxy-3,5-dimethyl- pyridin-2-ylmethyl)-5-piperidin-4-yl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (10.0 mg, 0.0248 mmol) in 1 ,4-Dioxane (0.29 mL). Triethylamine (8.65 μL, 0.0620 mmol) was added, followed by Sulfamide (4.77 mg, 0.0496 mmol). The mixture was microwaved for 2 hours at 120° C. Water was added and the mixture was extracted with ethyl acetate. Purification on a silica gel prep plate using EtOAc/MeOH gave the product as a white solid. (4.1 mg, yield 34%). LC-MS (5-100-5 method): Rt = 1.981 min; M+1 = 482.1. Example 93
2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-2-methylpropan- 1 -ol (XC VII) and 2-(4-chloro-7-((4-methoxy-3, 5-dimethylpyridin-2- yl)methyl)-2-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-2-methylpropan-1-ol (XCVIII) Step 1. 3-(4-Methoxy-benzyloxy)-2,2-dimethyl-propan-1-ol
[00524] A solution of neopentyl glycol (20.0 g, 192 mmol) in N,N-dimethylformamide (100 mL) was treated with sodium hydride (4.84 g, 202 mmol) in N,N-dimethylformamide (100 mL) at 0 0C for 30 minutes. p-Methoxybenzyl chloride (26.0 mL, 192 mmol) was added to the mixture at 0 0C, followed by tetra-n-butylammonium iodide (50.0 mg, 0.1 mmol). The mixture was stirred at 60 0C for 18 hours, diluted with sat. NH4CI solution (30OmL), extracted with fert-butyl methyl ether (150 mLx3), dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (heptane/EtOAc, 3:1 to 2:1) to afford the title compound as a colorless oil (26.2g, purity 95.0%, yield 57.8%). 1H NMR (CDCI3): δ ppm δ ppm 7.23 (d, 2H), 6.88 (d, 2H), 4.44 (s, 2H), 3.81 (s, 3H), 3.44 (d, 2H), 3.29 (s, 2H), 2.60 (t, 1H), 0.92 (s, 6H). Step 2. 3-(4-Methoxy-benzyloxy)-2,2-dimethyl-propionaldehyde
[00525] A solution of 3-(4-methoxy-benzyloxy)-2,2-dimethyl-propan-1-ol (26.0 g, 116 mmol) in anhydrous methylene chloride (200 mL) was treated with a suspension of pyridinium chlorochromate (42.5 g, 197 mmol) in anhydrous methylene chloride (200 mL) at room temperature for 2 hours. The reaction was diluted with anhydrous diethyl ether (60OmL) and filtered. The diethyl ether was dried over Na2SO4, and concentrated. The residue was purified by flash chromatography (hexane/EtOAc, 10:1 to 10:3) to yield the title compound as a colorless oil (15.8g, purity 95.0%, yield 58.2%). 1H NMR (CDCI3): δ ppm 9.56 (s, 1 H), 7.24 (d, 2H), 6.89 (d, 2H), 4.45 (s, 2H), 3.83 (s, 3H), 3.44 (s, 2H), 1.10 (s, 6H). Step 3. Diethyl 2-(3-(4-methoxybenzyloxy)-2,2-dimethylpropylidene)malonate [00526] A mixture of anhydrous tetrahydrofuran (250 mL) and anhydrous carbon tetrachloride
(35.0 mL) was treated with titanium tetrachloride (15.5 mL, 0.141 mol) at 0 °C for 5 minutes. The resulting yellow suspension was then treated sequentially with 3-(4-methoxy-benzyloxy)-2,2-dimethyl- propionaldehyde (15.7 g, 0.0706 mol) in anhydrous tetrahydrofuran (35.0 mL, 0.432 mol) and ethyl malonate (10.7 mL, 0.0706 mol) at 0 0C for 30 minutes. A solution of anhydrous pyridine (22.8 mL, 0.282 mol) in anhydrous tetrahydrofuran (40.0 mL) was added to the mixture at 0 0C for 1 hour and then allowed to reach room temperature overnight. The reaction mixture was quenched with water (20OmL) and extracted with methylene chloride (200mLx2). The combined organic layers were washed with brine and sat. NaHCO3, dried over Na2SO4, and evaporated to give the title compound as a yellow oil (24.7g, purity > 90%, yield 86.4%). 1H NMR (CDCI3): δ ppm 7.27 (d, 2H), 7.00 (d, 1 H), 6.89 (d, 2H), 4.47 (s, 2H), 4.25 (m, 4H), 3.82 (s, 3H), 3.27 (s, 2H), 1.32 (m, 6H), 1.15 (s, 6H).
Step 4. Diethyl 2-(4-(4-methoxybenzyloxy)-3,3-dimethyl-1-nitrobutan-2-yl)malonate [00527] A mixture of diethyl 2-(3-(4-methoxybenzyloxy)-2,2-dimethylpropylidene)malonate (24.0 g, 0.0658 mol), nitromethane (35.7 mL, 0.658 mol) and N,N,N',N'-tetramethylguanidine (4.13 mL, 0.0329 mol) was stirred at room temperature for 16 hours. To the mixture was added a 2N HCI solution. The organic layer was separated and washed with sat. NaCI solution (50.0 ml_x2), dried over Na2SO4, and evaporated. The residue was purified by flash chromatography (hexane/EtOAc, 10:1 to 10:3) to yield the title compound as a yellow oil (26.Og, purity > 90%, yield 84.0%). 1H NMR (CDCI3): δ ppm 7.21 (d, 2H), 6.86 (d, 2H), 4.98 (dd, 1 H), 4.77 (dd, 1 H), 4.32 (d, 2H), 4.20 (m, 4H), 3.80 (s, 3H), 3.35 (s, 1 H), 3.15 (dd, 2H), 1.26 (m, 6H), 0.93 (s, 3H) 0.92 (s, 3H).
Step 5. 4-[2-(4-Methoxy'benzyloxy)-1, 1-dimethyl-ethyl]-2-oxo-pyrrolidine-3-carboxylic acid ethyl ester
[00528] Sodium tetrahydroborate (13.9 g, 0.367 mol) was added to the mixture of diethyl 2-(4-(4- methoxybenzyloxy)-3,3-dimethyl-1-nitrobutan-2-yl)malonate (26.0 g, 0.0611 mol) and nickel chloride hexahydrate (7.26 g, 0.0306 mol) in methanol (150 mL) at 0 0C. After addition, the resulting reaction mixture was stirred at room temperature for 24 hours. The reaction mixture was quenched with sat. NH4CI solution (30OmL), diluted with methylene chloride (500 mL), The organic layer was separated, washed with sat. NH4CI solution (200 mLx6), dried over Na2SO4, and evaporated to provide the title compound as a pale yellow oil (16.4 g, purity > 90%, yield 69.1%). 1H NMR (CDCI3): δ ppm 7.22 (d, 2H), 6.88 (d, 2H),
6.10 (s, 1 H), 4.37 (s, 2H), 4.20 (q, 2H), 3.77 (s, 3H), 3.41 (m, 2H), 3.19 (m, 1 H), 3.14 (s, 2H), 3.08 (m,
1H), 1.27 (t, 3H), 0.91 (s, 6H).
Step 6. 2-Ethoxy-4-[2-(4-methoxy-benzyloxy)-1, 1-dimethyl-ethyl]-4,5-dihydro~3H-pyrrole-3- carboxylic acid ethyl ester
[00529] A solution of 4-[2-(4-methoxy-benzyloxy)-1 , 1 -dimethyl-ethyl]-2-oxo-pyrrolidine-3- carboxylic acid ethyl ester (8.00 g, 22.9 mmol) in anhydrous methylene chloride (20.0 mL) was treated with 1.0 M of triethyloxonium tetrafluoroborate in anhydrous methylene chloride (26.6 mL, 26.6 mmol) under nitrogen at room temperature overnight. The reaction mixture was diluted with water (80.0 mL) and allowed to stand for 30 minutes. The organic layer was washed with sat. NaHCO3 (10.0 mLx2), water
(10.0 mL), dried over Na2SO4, and evaporated to give the title compound as a pale green oil (6.8Og, purity
> 90%, yield 71.0%). 1H NMR (DMSO-d6): δ ppm 9.48 (s, 1 H), 7.23 (d, 1H), 6.89 (d, 1 H), 6.23 (s, 2H),
4.40 (d, 1H), 4.32 (d, 1 H), 3.75 (s, 3H), 3.27 (m, 4H), 3.00 (m, 1 H), 0.89 (s, 3H), 0.84 (s, 3H).
Step 7. 2-Amino-5-(1-hydroxy-2-methylpropan-2-yl)-6,7-dihydro-3H-pyrrolo[2,3-d]pyrimidin-4(5H)- one
[00530] Sodium hydride (2.16 g, 90.1 mmol) was added to anhydrous ethanol (80.0 mL) at 0 0C and stirred for 20 minutes until hydrogen gas evolution ceased. Guanidine hydrochloride (8.60 g, 90.1 mmol) was added. The mixture was stirred at 50 0C for 20 minutes and cooled to 100C. The NaCI was filtered off, and to the filtrate was added 2-ethoxy-4-[2-(4-methoxy-benzyloxy)-1 ,1-dimethyl-ethyl]-4,5- dihydro-3H-pyrrole-3-carboxylic acid ethyl ester (6.8 g, 18.0 mmol). The mixture was stirred until clear and then concentrated in vacuo to remove most of the alcohol. The residue was heated to 90 0C overnight while maintaining a vacuum of 10 mmHg. The mixture was evaporated and the residue was taken up in water (50.0 mL) and 6N HCI solution (10.0 mL). The mixture was heated to 60 0C for 3 hours. After filtration, the aqueous layer was purified by reverse-phase preparative HPLC (5 to 30%, CH3CN/H2O,
0.1% TFA) to give the title compound as a white solid (360mg, purity > 90%, yield 8.0%). 1H NMR
(DMSO-d6): δ ppm 10.50 (s, 1 H), 7.25 (s, 1 H), 6.84 (s, 1 H), 3.48 (t, 1 H), 3.37 (t, 1 H), 3.18 (d, 1 H), 3.07
(m, 1 H), 2.97 (m, 1 H), 0.87 (s, 3H), 0.61 (s, 3H).
Step 8. 2-(2-Amino-4-chloro-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-2-methyl-propan-1-ol [00531] A mixture of 2-Amino-5-(1-hydroxy-2-methylpropan-2-yl)-6,7-dihydro-3H-pyrrolo[2,3- d]pyrimidin-4(5H)-one (350 mg, 1.60 mmol) and acetic anhydride (1.47 ml_, 15.6 mmol) was heated to reflux for 1 hour and evaporated. The residue was treated with benzyltriethylammonium chloride (0.711 g, 3.12 mmol) and phosphoryl chloride (0.873 ml_, 9.36 mmol at 90 0C for 30 min and evaporated. The residue was poured into ice-water (10.0 ml_) and 11 M of hydrogen chloride in water (1.80 mL, 20.0 mmol). The mixture was stirred at 45 °C overnight. After cooling, the mixture was adjusted to pH 4.0 with 4N NaOH solution, filtered and purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (242mg, purity 99.0%, yield 63.0%). 1H NMR (DMSOd6): δ ppm 7.10 (s, 1 H), 6.29 (s, 2H), 3.40 (m, 2H), 3.28 (m, 2H), 3.13 (m, 1 H), 3.03 (m, 1H), 0.85 (s, 3H), 0.73(s, 3H).
Step 9. 5-[2-(tert-Butyl-dimethyl-silanyloxy)-1, 1-dimethyl-ethyl]-4-chloro-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-ylamine
[00532] A mixture of 2-(2-amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-2-methyl- propan-1-ol (242 mg, 0.997 mmol), 1 H-imidazole (170 mg, 2.49 mmol) in N,N-dimethylformamide (3.00 mL) was treated with fert-butyldimethylsilyl chloride (316 mg, 2.09 mmol) at 60 °C for 2 hours. The mixture was diluted with water (20.0 mL), extracted with a mixture of dichloromethane/isopropanol (100:5, 20.0 mLx2), washed with brine (10.0 mL), and evaporated. The residue was purified by preparative TLC (hexane/EtOAc, 1 :1) to give the title compound as a white solid (106 mg, purity 99.0%, yield 29.5%). LC- MS (5-100-7 method):Rt=7.020 min; M+1 =356.7
Step 10. 2-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl]-2-methyl-propan- 1 -ol (XCVII) and 2-(4-chloro-7-((4-methoxy-3, 5- dimethylpyridin-2-yl)methyl)-2-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)-2-methylpropan-1 -ol (XCVIII)
[00533] A mixture of 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine (41.0 mg, 0.221 mmol) in dimethyl sulfoxide (1.00 mL) was treated with sodium hydride (7.56 mg, 0.315 mmol) at 0 0C for 10 minutes and then brought to room temperature. A solution of 5-[2-(fert-butyl-dimethyl-silanyloxy)-1 ,1- dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (75.0 mg, 0.210 mmol) in dimethyl sulfoxide (1.00 mL) was added and the mixture was stirred at room temperature for 1 h. The mixture was diluted with sat. NH4CI solution, extracted with a mixture of dichloromethane/isopropanol (85:15, 20mLx3), dried over Na2SO4, and evaporated. The crude was dissolved with DMSO (5.00 mL) and 6.0 M hydrogen chloride in water (3.50 mL). The mixture was stirred at room temperature for 20 minutes and evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% TFA) to give 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-2-methylpropan-1 -ol as a white solid (40.0mg, purity 99.0%, yield 48.6%). 1H NMR (Acetone-d6): δ ppm 8.16 (s, 1 H), 5.79 (s, 2H), 4.94 (d, 2H), 4.20 (d, 2H), 3.79 (s, 3H), 3.64 (m, 1 H), 3.48 (m, 1 H), 3.34 (d, 1 H), 3.23 (d, 1 H), 3.12 (m, 1 H), 2.26 (s, 6H), 0.94 (s, 3H), 0.78(s, 3H). LC-MS (5-100-7 method): Rt=4.265 min; M+1=391.6 [00534] In addition, the chromatography also gave 2-(4-chloro-7-((4-methoxy-3,5-dimethylpyridin-
2-yl)methyl)-2-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-
5-yl)-2-methylpropan-1-ol as a white solid (8.0mg, purity 99.0%, yield 7.0%). 1H NMR (DMSOd6): δ ppm
8.22 (s, 1H), 8.20 (s, 1 H)1 6.63 (s, 1 H), 5.19 (d, 1 H)1 4.58 (s, 2H)1 4.02 (m, 1H)1 3.98 (d, 1H), 3.80 (s, 6H)1
3.45 (d, 1 H)1 3.36 (t, 1 H)1 3.19 (d, 1 H)1 3.09 (d, 1 H)1 2.29 (s, 6H), 2.17 (s, 6H), 1.05 (s, 3H)1 0.95 (s, 3H).
LC-MS (5-100-7 method): Rt=4.876 min; M+1 =540.5
Example 94
4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H'pyrrolo[2,3-d]pyrimidin-2- amine (XCIX)
Step 1. N-(4-hydroxy-7H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
[00535] To a stirred solution of 2-amino-7H-pyrrolo[2,3-d]pyrimidin-4-ol (100 mg, 0.00066 mol) in pyridine (1 mL) was added pivaloyl chloride (255 mg, 0.00213 mol) at RT. The resulting reaction mixture was stirred at 90 0C for 2h. The volatiles were evaporated under reduced pressure and the crude material was diluted with aqueous NH3 solution (1 mL) and stirred at RT for 30 min. The excess of aqueous NH3 solution was evaporated and the crude material was washed with 10% EtOAc/Hexane to afford the title compound (100 mg, 64.5%) as a brown solid. 1H NMR (200 MHz1 DMSO-cfe): δ 11.7 (s, 1 H)1 7.04 (bs,
1 H)1 6.96 (d, J = 3.4 Hz1 1 H), 6.71 (bs, 1 H)1 6.40 (d, J = 3.4 Hz, 1 H)1 1.06 (s, 9H). LC-MS (10-90[4]-10 method): Rt = 2.36 min. (98.8% HPLC purity); m/z = 235 (M+1 ).
Step 2. N-(4-hydroxy-7-tosyl-7H-pyrrolo[2, 3-d]pyrimidin-2-yl)pivalamide
[00536] To a stirred solution of N-(4-hydroxy-7H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide (200 mg,
0.00085 mol) in DMF (2 mL) was added NaH (58 mg, 0.0025 mol) and tosyl chloride (162 mg, 0.00085 mol) at 0 0C. The resulting reaction mixture was stirred at RT for 12h. The reaction mass was cooled to
RT and diluted with water and extracted with EtOAc (2x 15 mL). The combined organic layers was evaporated under reduced pressure to afford the title compound (150 mg, 45%) as an off-white solid. 1H
NMR (200 MHz, DMSO-c/6l) δ 12.2 (bs, 1 H)1 10.92 (bs, 1 H), 8.13 (d, J = 8.6 Hz1 2H), 7.49-7.36 (m, 3H)1
6.64 (d, J = 3.8 Hz, 1 H), 2.34 (s, 3H)1 1.25 (S1 9H). LC-MS (10-90[4]-10 method): Rt = 4.12 min. (76%
HPLC purity); m/z = 388.4 (M+1 ).
Step 3. N-(4-hydroxy-7-tosyl-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
[00537] To a stirred solution of N-(4-hydroxy-7-tosyl-7H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
(150 mg, 0.00038 mol) in EtOH (1.5 mL) was added 5% Pd(OH)2/C (15 mg) at RT under anhydrous conditions. The reaction mixture was stirred at RT for 5h under Hydrogen balloon pressure. The reaction mixture was filtered off and washed with EtOH. The filtrate was evaporated under reduced pressure to afford the title comopund (100 mg, 60%) as an off-white solid. 1H NMR (200 MHz, DMSO-cfe) δ 11.95 (bs,
1H)1 10.97 (bs, 1 H), 8.05 (d, J = 8.4 Hz, 2H)1 7.39 (d, J= 8.4 Hz, 2H), 3.89 (t, J = 9.2 Hz, 2H), 2.65 (t, J =
8.3 Hz1 2H), 2.36 (s, 3H), 1.27 (s, 9H). LC-MS (10-90[4]-10 method): Rt = 4.34 min. (78% HPLC purity); m/z = 391 (M+1).
Step 4. N-(4-chloro-7-tosyl-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide [00538] A solution of N-(4-hydroxy-7-tosyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
(125 mg, 0.776 mol) in POCI3 (1.5 ml_) was stirred at 70 0C for 2h. The reaction mixture was cooled to RT and quenched with saturated NaHCO3 solution and extracted with EtOAc (2x 15 mL). The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure to afford the title compound (20 mg, 38.7%) as an off-white solid. Mass: 408.9 (M++1). TLC system: 5% MeOH/dichloromethane, Rf = 0.7.
Step 5. 4-chloro-6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine
[00539] A solution of N-(4-chloro-7-tosyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-yl)pivalamide
(200 mg, 0.77 mmol) in H2SO4 (5 mL) was stirred at 100 0C for 8h. The reaction mixture was cooled to RT and quenched with saturated K2CO3 solution and extracted with EtOAc (3x 15 mL). The organic layer was washed with brine and dried over Na2SO4. The solvent was evaporated under reduced pressure; the crude material was washed with hexane to afford the title compound (30mg, 36%) as an off-white solid. 1H NMR (500 MHz, DMSO-Of6): δ 7.20 (bs, 1H), 6.25 (bs, 2H), 3.48 (t, J= 9 Hz, 1 H), 2.82 (t, J = 8.5 Hz, 1 H). LC-MS (10-90[4]-10 method): Rt = 3.28 min. (99% HPLC purity); m/z = 171 (M+1). Step 6. 4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-2-amine (XCIX)
[00540] To a stirred solution of 4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (30 mg,
0.00017 mol) in DMF (0.3 mL) was added NaH (60% oil suspension) (165 mg, 1.2 mmol) and 2- (chloromethyl)-4-methoxy-3,5-dimethylpyridine (32 mg, 0.00017 mol) at 0 0C. The resulting reaction mixture was stirred at 10 0C for 20 min. The reaction mass was quenched with water and extracted with EtOAc (2x 5 mL).The combined organic layers were washed with brine and dried over Na2SO4 and evaporated under reduced pressure. The resulting crude material was washed with n-pentane to afford the title compound (6 mg, 10%) as a white solid. 1H NMR (500 MHz, DMSO-d6): δ 8.19 (s, 1 H), 6.38 (bs, 2H), 4.58 (s, 2 H), 3.72 (s, 3H), 3.52 (t, J = 8.5 Hz, 2H), 2.82 (t, J = 8.0 Hz, 2H), 2.19 (s, 6H). HPLC (10- 90[3]-6 method): Rt = 1.27 min. (99% purity). Mass: m/z = 319.9 (M+1). Example 95
(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-6-yl)methanol (C)
Step 1. 5-allyl-6-chloro-N4-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)pyrimidine-2,4-diamine [00541] To a stirred solution of 5-allyl-4,6-dichloropyrimidin-2-amine (Barbosa, J.; Dong, L.; Fink,
C. A.; Lanthorn, T. H.; Schmidt, J. M.; Wang, J.; Zipp, G. G. Preparation of biphenyl heteroaryl methanones for improving cognition. PCT Int. Appl. WO 2006124897) (1.85g, 0.0091 mol) in DMF (10 mL) was added (4-methoxy-3,5-dimethylpyridin-2-yl)methylamine (1.66g, 0.010 mol) and K2CO3 (3.1g, 0.022 mol) at 0 0C. The resulting reaction mixture was heated at 70 0C for 2h and stirred at RT for 1h. The reaction mass was diluted with water (25 mL) and extracted with EtOAc (2x 20 mL). The combined organic layers was dried over Na2SO4 and concentrated under reduced pressure, the resulting crude was purified by column chromatography (50% EtOAc/Hexane) to afford the title compound (0.6 g). 1H NMR (200 MHz, DMSO-CZ6): δ 8.21 (s,1 H), 7.06 (bs, 1 H), 6.33 (s, 2H), 5.87- 5.72 (m, 1 H), 5.16- 5.01 (m, 2H),
4.50 (d, J = 4.4 Hz, 2H), 3.72 (s, 3H), 3.28- 3.25 (m, 2H), 2.17 (s, 3H), 2.16 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.69 min. (81%); m/z = 334 (M+1).
Step 2. 3^2-amino^-chloro-6-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)pyrimidin-5- yl)propane-1,2-diol
[00542] To a stirred solution of 5-allyl-6-chloro-N4-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)pyrimidine-2,4-diamine (0.6 g, 0.0018 mol) in DMF (3 mL) was added TFA (10 mL) and 30 %
H2O2 (15 mL) at 0 0C. The resulting reaction mixture was stirred at RT for 12h (TLC shows 50% completion). More 30 % H2O2 (IO mL) was added, and the mixture was stirred at RT for another 6h (TLC shows 70% completion). More 30 % H2O2 (10 mL) was added, and the mixture was stirred at RT for another 6 (TLC shows 80% completion). The reaction mass was diluted with water (25 mL) and basified with NaHCO3 solution and extracted with EtOAc (2x 15 mL). The combined organic layers was dried over
Na2SO4 and concentrated under reduced pressure; the resulting crude was purified by column chromatography (10% MeOH/dichloromethane) to afford the title compound (0.4 g). 1H NMR (200 MHz,
DMSO-Cf6): δ 8.18 (s,1 H), 7.20 (bs, 1 H), 6.27 (s, 2H), 4.91 (d, J= 4.8 Hz, 1 H), 4.74- 4.69 (m, 1 H), 4.51 (bs,
2H), 3.72 (s, 3H), 3.69- 3.60 (m, 2H), 2.72- 2. 54 (m, 1H), 2.46- 2. 40 (m, 1 H) 2.20 (s, 3H), 2.18 (s, 3H).
LC-MS (10-90[4]-10 method): Rt = 2.35 min. (86%); m/z = 368 (M+1 ).
Step 3. 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)pyrimidin-5-yl)-2- hydroxypropyl methanesulfonate
[00543] To a stirred solution of 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2- yl)methylamino)pyrimidin-5-yl)propane-1 ,2-diol (0.35g, 0.00095 mol) in DMF (10 mL) was added DIPEA
(0.24 mL, 0.0014 mol) and Mesyl chloride (0.08 mL, 0.0010 mol) at 0 °C. The resulting reaction mixture was stirred at RT for 2h. The reaction mixture was diluted with water (15 mL) and extracted with EtOAc
(2x 15 mL). The combined organic layers were dried over Na2SO4 and concentrated to afford the title compound (0.36 g). LC-MS (10-90[4]-10 method): Rt = 2.83 min. (40%); m/z = 446 (M+1).
Step 4. 6-chloro-N4-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5-(oxiran-2-ylmethyl)pyrimidine-
2,4-diamine
[00544] To a stirred solution of 3-(2-amino-4-chloro-6-((4-methoxy-3,5-dimethylpyridin-2- yl)methylamino)pyrimidin-5-yl)-2-hydroxypropyl methanesulfonate (0.36g, 0.0008 mol) in MeOH (3 mL) was added NaOMe (0.058g, 0.00097 mol) at 0 CC. The resulting reaction mixture was stirred at RT for 2h.
The resulting reaction mixture was stirred at RT for 2h. The reaction mass concentrated under reduced pressure, the resulting crude was purified by column chromatography (50% EtOAc/ Hexane) to afford the title compound (0.08g). 1H NMR (200 MHz, DMSO-cfe): δ 8.20 (s, 1 H), 7.18- 7.16 (m, 1 H), 6.35 (bs, 2H),
4.51 (d, 4.4 Hz, 1 H), 3.72 (s, 3H), 3.13- 2.98 (m, 1 H), 2.83- 2.81 (m, 1 H), 2.78- 2. 71 (m, 2H), 2.61- 2.50 (m 1 H), 2.19 (s, 3H), 2.18 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.13 min. (98%); m/z = 350 (M+1 ). Step 5. (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-6-yl)methanol (C) [00545] 6-chloro-N4-((4-methoxy-3,5-dirnethylpyridin-2-yl)methyl)-5-(oxiran-2-ylmethyl)pyrimidine-
2,4-diamine (20 mg) was stirred in DMF (3 ml_) at 100 0C for 4h. The reaction mixture was cooled RT and diluted with water (5 mL) and extracted with EtOAc (2x 5 ml_). The combined organic layers was dried over Na2SO4 and concentrated under reduced pressure, and the resulting crude was purified by preparative TLC (50% EtOAc/Hexane) to afford the title compound (3 mg). 1H NMR (500 MHz, CDCI3): δ 8.05 (s,1 H), 5.76 (d, J = 16.5 Hz, 1 H), 4.53 (bs, 2H), 4.30 (s, 1 H)1 3.93 (d, J = 16.5 Hz, 1 H), 3.78 (s, 3H), 3.67 (d, J = 12 Hz, 1H), 3.53-3.50 (m, 1 H), 3.12-3.00 (m, 1 H), 2.67- 2.63 (m, 1 H), 2.23 (s, 3H), 2.21(s, 3H). Mass: 350.4 (M+1 ). HPLC: (10-90[4]-10 method): Rt = 3.27 min. (82%). Example 96
1-(4-Methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-3,3-dimethyl-2,2a,3,4-tetrahydro-1H-5-oxa-1,6,8- triaza-acenaphthylen-7-ylamine (Cl)
[00546] A mixture of 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine hydrochloride (18.9 mg,
0.0853 mmol) in dimethyl sulfoxide (1.0 mL) was treated with sodium hydride (4.09 mg, 0.171 mmol) at 0 0C for 10 minutes and then allowed to reach room temperature. A solution of 5-[2-(terf-butyl-dimethyl- silanyloxy)-1 ,1-dimethyl-ethyl]-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (29.0mg, 0.0812 mmol) in dimethyl sulfoxide (0.50 mL) was added. The mixture was heated to 100 °C for 1h and then was cooled to room temperature. The pH was adjusted to 4.0 with 6N HCI, and the solution was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (1.80 mg, purity 97.0%, yield 6.0%). 1H NMR (Acetone-d6): δ ppm 8.12 (s, 1 H), 5.49 (s, 2H), 4.85 (d, 1 H), 4.03 (d, 1 H), 3.83 (s, 2H), 3.76 (s, 3H), 3.52 (t, 1H), 3.25 (t, 1 H), 3.08 (t, 1 H), 2.24 (s, 3H), 2.21 (s, 3H), 0.96 (s, 3H), 0.75 (s, 3H). LC-MS (5-100-7 method): Rt=4.559 min; M+1=355.7. Example 97
(2-amino-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)methanol (CII)
[00547] A mixture of (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (98 mg, 0.28 mmol), TFA (50 μL, 0.6 mmol), 10% Pd/C (20 mg) and EtOH (30 mL) was placed on a Parr hydrogenator for 1 h at 40 psi. The Pd/C was removed by filtration, and the solvents were evaporated to yield a beige solid. (60 mg, purity 90%, yield 60%). 1H NMR (CDCI3): δ ppm 8.10 (s, 1 H), 7.50 (s, 1 H), 4.88 (s, 2H), 4.71 (d, 1H), 4.51 (d, 1 H), 3.70 (s, 3H), 3.59 (m, 3H), 3.27 (m, 2H), 2.19 (s, 3H), 2.17 (s, 3H)1.12 (t, 3H). LC/MS: rt (5-100-7 method) = 3.471 min.; 315.7(M+1 , 100 %), 316.7(M+3, 20%) Example 98
(R)-2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetamide (CV) and (S)- 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (CVI) Step 1. 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide
[00548] A solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetonitrile (1.0 g, 2.7 mmol) in DMSO (10 ml), was cooled to 0
°C, and treated with 1 N NaOH (7.5 mL) followed by 30% H2O2 (5 mL). The resulting reaction mixture was brought to RT and stirred for 24h. Work-up (EtO Ac/water), drying (Na2SO4), evaporation, purification by preparative HPLC gave the title compound (210 mg, 20%) as a white solid. 1H NMR (DMSO-d6t200
MHz): δ 8.15 (s,1 H), 7.35 (s, 1 H; Exc), 6.83 (s, 1 H; Exc), 6.39 (s, 2H; Exc), 4.59-4.52 (m, 2H), 3.71 (s,
3H), 3.62-3.58 (m, 1 H), 3.45-3.41 (m, 1 H), 3.13-3.10 (m, 1H), 2.69-2.66 (m, 1 H), 2.20-2.15 (m, 7H); LC-
MS (5-100-5 method) rt = 1.73 min, m/z = 377.1 (M+1 ). TLC system: 10% MeOH/DCM R,: 0.5.
Step 2. (R)-2-(2-amino-4-chloro-7-((4-methoxy-3, 5-dimethylpyridι'n-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (CV) and (S)- 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (CVI)
Step 1. 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide
[00549] Racemic 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetamide was separated on a Chiralpak AS-H (2x15 cm) eluting with
35% MeOH (0.1%Et2NH)/CO2 100 psi, at a flow rate of 60 mLΛnin with detection at 220 nM to give the
(R)-enantiomer (fast running) and the (S) enantiomer (slow running).
Example 99
(R)-2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetamide (CVII)
[00550] Alternative procedure. A flame-dried flask was charged with formamide (615.5 uL, 15.44 mmol) and anhydrous Dimethyl sulfoxide (4.00 mL). Potassium tert-Butoxide (890.0 mg, 7.931 mmol) was added in portions, and the resulting suspension was stirred for 5 minutes. A solution of (R)-methyl 2-(2- amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)acetate (1.939 g, 4.948 mmol) in anhydrous Dimethyl sulfoxide (5.50 mL, 77.5 mmol) was added. After
15 minutes, the reaction was quenched with water. Work-up (EtOAc/water), drying (MgSO4) and evaporation gave the title material as a solid (1.467 g, 77.5% yield). See previous example for spectral data.
Example 100
2-[(R)-2-Amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-acetamide (CVIII)
Step 1. 5-Ethyl-4,6-dimethyl-2-oxo-1,2-dihydro-pyridine-3-carbonitrile
[00551] 2-Cyanoacetamide (7.35 g, 87.4 mmol) was dissolved in boiling ethanol (50.0 mL, 814 mmol). 3-Ethyl-2,4-pentanedione (10.5 mL, 78.0 mmol) was added to the mixture and the system gently refluxed with stirring while N-ethylethanamine (4.04 mL, 39.0 mmol) was added dropwised over 20 minutes. After refluxing for an additional hour, the mixture was cooled to 0 0C. The white crystals that formed were collected and dried to give the title compound as white crystals (9.00 g, purity 96.0%, yield
65.5%). LC-MS (5-100-7 method):Rt=4.722 min; M+1 =176.8
Step 2. 2-Bromo-5-ethyl-4,6-dimethyl-nicotinonitrile
[00552] A mixture 5-ethyl-4,6-dimethyl-2-oxo-1 ,2-dihydro-pyridine-3-carbonitrile (9.00 g, 51.1 mmol), phosphorus pentoxide (15.2 g, 53.6 mmol) and tefra-N-butylammonium bromide (19.8 g, 61.3 mmol) in anhydrous toluene (120 ml_) was heated to reflux for 4 hours. The mixture was cooled to room temperature and ice-water (18.4 mL) was added carefully with stirring and cooling. The mixture was stirred for 2 hours and then filtered through a pad of celite, and the layer was separated. The toluene layer was washed with brine (30.0 mL x2) and evaporated. The crude was purified by flash chromatography (hexane/ EtOAc, 100:0 to 0:100) to give the title compound as a white solid (10.0 g, purity 96.0%, yield 81.9%). LC-MS (5-100-7 method):Rt=6.862 min; M+1=238.7
Step 3. 2-Bromo-5-ethyl-4,6-dimethyl-pyridine-3-carbaldehyde
[00553] A solution of 2-bromo-5-ethyl-4,6-dimethyl-nicotinonitrile (10.0 g, 41.8 mmol) in toluene
(45.0 mL) was treated with 1.0 M of diisobutylaluminum hydride in toluene (46.8 mL, 46.8 mmol) at -70
0C for 2 hours. The reaction mixture was brought to room temperature and was carefully quenched with
2M sulfuric acid (188 mL). The resulting mixture was stirred overnight at room temperature and diluted with toluene (100.0 mL). The organic layer was washed with water (30.0 mL) and evaporated to give the title compound as a colorless oil (9.72 g, purity 96.0%, yield 96.0%). LC-MS (5-100-7 method):Rt=6.842 min; M+1 =241.7
Step 4. (2-Bromo-5-ethyl-4,6-dimethyl-pyridin-3-yl)-methanol
[00554] A solution of 2-bromo-5-ethyl-4,6-dimethyl-pyridine-3-carbaldehyde (9.00 g, 37.2 mmol) in ethanol (100 mL) was treated with sodium tetrahydroborate (3.52 g, 92.9 mmol) portionwise at 10 0C.
After addition, the mixture was stirred at room temperature for 1 hour, quenched with water (6.70 mL, 372 mmol) and 5.0 M of ammonium chloride in water (100.0 mL). The reaction mixture was evaporated. The residue was dissolved in dichloromethane (150 mL). The organic layer was washed with water (50.0 mL) and evaporated. The crude was purified by flash chromatography (dichloromethane/ MeOH, 100:1 to
100:10) to give the title compound as a white solid (6.20 g, purity 95.0%, yield 65.0%). LC-MS (5-100-7 method) :Rt=4.607 min; M+1 =243.7
Step 5. (5-Ethyl-4,6-dimethyl-pyridin-3-yl)-methanol
[00555] A mixture of (2-bromo-5-ethyl-4,6-dimethyl-pyridin-3-yl)-methanol (6.00 g, 24.6 mmol) and 5% palladium on CaCO3 (2.62 g, 1.23 mmol) in ethanol (100.0 mL) was stirred under hydrogen
(balloon) overnight. After removal of the catalyst by filtration through a pad of celite, the filtrate was evaporated. The crude was purified by flash chromatography (dichloromethane/ MeOH, 100:1 to 100:10) to give the title compound as a white solid (3.30 g, purity 95.0%, yield 77.2%). LC-MS (5-100-7 method):Rt=3.702 min; M+1 =165.9
Step 6. 5-Chloromethyl-3-ethyl-2,4-dimethyl-pyridine [00556] A solution of (5-ethyl-4,6-dimethyl-pyridin-3-yl)-methanol (3.15 g, 19.1 mmol) in anhydrous methylene chloride (20.0 mL) was treated with thionyl chloride (3.48 mL, 47.7 mmol). The reaction mixture was stirred at room temperature for 20 minutes, and evaporated, diluted with water (50.0 mL), adjusted to pH 7.0, extracted with a mixture of dichloromethane/isopropanol (95:5, 50.0 ml_x3). The organic layer was dried over Na2SO4 and evaporated to give the title compound as reddish oil (3.10 g, purity 95.0%, yield 88.0%). LC-MS (5-100-7 method):Rt=4.350 min; M+1 =183.8
Step 7. [(R)-2-Amino-4-chloro-7-(5-ethyl-4, 6-dimethyl-pyridin-3-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid
[00557] A mixture of (f?)-methyl 2-(2-amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)acetate (400.0 mg, 1.65 mmol) and potassium ferf-butoxide (203 mg, 1.81 mmol) in anhydrous dimethyl sulfoxide (2.50 mL) was treated with 5-chloromethyl-3-ethyl-2,4-dimethyl-pyridine (318 mg, 1.73 mmol) in dimethyl sulfoxide (4.00 mL) at room temperature for 1 hour. 6 M of hydrogen chloride in water (2.75 mL,
16.5 mmol) was added and the reaction mixture was heated at 70 °C for 1.5 hours. The mixture was diluted with water (70.0 mL) and purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O,
0.1% HCOOH) to give the title compound as a white solid (520.0 mg, purity 95.0%, yield 80.0%). LC-MS
(5-100-5 method) :Rt=1.837 min; M+1=376.1
Step 8. 2-[(R)-2-Amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetamide (CVIII)
[00558] A solution of [(fl)-2-amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (200.0 mg, 0.532 mmol) in tetrahydrofuran (10.0 mL) and triethylamine (371 μL, 2.66 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (183 μL, 1.06 mmol) at room temperature for 20 minutes. 0.5 M of ammonia in dioxane (5.32 mL, 2.66 mmol) was added to the mixture and was stirred for 20 minutes at room temperature. The reaction mixture was evaporated. The residue was purified by flash chromatography (dichloromethane/ methanol, 100:1 to
100:10) to give the title compound as a white solid (90.0 mg, purity 99.0%, yield 40.0%). 1H NMR (DMSO- d6): δ ppm 8.07 (s, 1 H), 7.37 (br. s, 1 H), 6.85 (br. s, 1 H), 6.50 br. s, 1 H), 4.53 (d, 1 H), 4.41 (d, 1 H), 3.46
(m, 2H), 3.03 (m, 1 H), 2.66 (m, 3H), 2.46 (s, 3H), 2.20 (s, 3H), 2.16 (m, 1 H), 1.20 (t, 3H). LC-MS (5-100-7 method):Rt=3.847 min; M+1 =374.6
Example 101
(S)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetamide (CIX)
Step 1. (S)-methyl 2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetate
[00559] In a flame-dried flask under nitrogen was stirred (S)-methyl 2-(2-amino-4-chloro-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (100.0 mg, 0.4121 mmol) and 2-(chloromethyl)-4-ethyl-
3,5-dimethylpyridine (79.5 mg, 0.433 mmol) in Dimethyl sulfoxide (4.2 mL). The mixture was treated with
Potassium tert-Butoxide (55.5 mg, 0.494 mmol) and stirred for 30 minutes before being diluted with water. The solution was then extracted 3 times with ethyl acetate and dried over magnesium sulfate. Purification by silica gel chromatography using ethyl acetate/hexane gave an impure mixture that was carried on crude.
Step 2. Lithium (S)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate
[00560] A solution of (S)-methyl 2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (37.0 mg, 0.0949 mmol) in Methanol (0.500 mL) and
Water (0.250 mL) was treated with Lithium hydroxide (2.27 mg, 0.0949 mmol) for 5 hours at 50°C. The reaction was then condensed to dryness to yield the title product as a white solid.
Step 3. (S)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (CIX)
[00561] A solution of Lithium (S)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (18.7 mg, 0.0498 mmol) in Tetrahydrofuran (2 mL) and N,N-Diisopropylethylamine (63 μL, 0.36 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (21.44 μL, 0.1244 mmol) at room temperature for 5 minutes, followed by addition of 0.5 M NH3 in dioxane (0.02615 g, 0.2488 mmol). After 10 minutes the reaction was diluted with water, extracted 3 times with dichloromethane, and dried over magnesium sulfate. Purification by silica gel chromatography using dichloromethane/methanol gave the final product as a white solid. (11 mg, yield
59%). LC-MS (5-100-5 method) : Rt = 1.748 min; M+1 = 380.1 ; 1H NMR (400 MHz, CDCI3) δ ppm 8.16 (s,
1 H), 5.34 - 5.64 (m, 2 H), 4.86 (s, 2 H), 4.64 (s, 2 H), 3.77 (s, 3 H), 3.73 (d, J= 10.1 Hz, 1 H), 3.62 - 3.71
(m, 1 H), 3.35 (dd, J= 10.0, 3.9 Hz, 1 H), 2.94 (dd, J= 15.5, 3.2 Hz, 1 H), 2.38 (dd, J= 15.5, 10.2 Hz, 1 H),
2.22 (s, 3 H).
Example 102
2-(2-amino-4-chloro-7-((5-(hydroxymethyl)-4-methoxy-3-methylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl)acetamide (CX)
Part A. Methyl 6-(chloromethyl)-4-methoxy-5-methylnicotinate
Step 1. 4-Methoxy-2,3-dimethyl-pyridine 1 -oxide (3)
[00562] A solution of 4-nitro-2,3-lutidine N-oxide (1 , 50.0 g, 0.297 mol, Aldrich) in Methanol (500 mL, 10 mol) was cooled by means of an ice-bath, and treated portionwise with potassium carbonate
(50.0 g, 0.362 mol) (Caution: without the ice bath, the reaction can be violently exothermic). The reaction mixture was heated to reflux for 2 hours, after which time the reaction was cooled to room temperature and the solvent evaporated. The crude solid was taken up in CH3CN (1.5 L), heated to reflux for 30 minutes, filtered, and the cake was washed with CH3CN (100mLX3). The CH3CN solutions were combined, and filtered again to remove solids. The solid was tentatively assigned as a dimer (2,2'-
(ethane-1 ,2-diyl)bis(4-methoxy-3-methylpyridine 1 -oxide, 2) (8.2 g, 6.3% yield, 70% purity). The filtrate was evaporated to yield the product 3 as yellow solid (24.0 g, 47% yield, 90% purity). Step 2. 5-Bromo-4-methoxy-2,3-dimethyl-pyridine 1 -oxide (4)
[00563] Refer to Duan, J. et al., Synlett, 1999, 1245-1246. A solution of 4-Methoxy-2,3-dimethyl- pyridine 1 -oxide (3, 20.0 g, 0.130 mol) in Trifluoroacetic Acid (27.2 ml_, 0.352 mol) and Sulfuric acid (36.2 mL, 0.679 mol) was treated with N-Bromosuccinimide (46.5 g, 0.261 mol) portionwise. The reaction mixture was stirred overnight at room temperature, diluted with ice-cold water (50OmL), quenched with saturated aqueous Na2S2O3 (50OmL), neutralized with 6N NaoH to pH 7.0, extracted with DCM (
800mLX2), washed with saturated aqueous NH4CI, and evaporated to give product the desired product as a yellow solid (18.0 g, 42% yield, 70% purity).
Step 3. 4-methoxy-5-(methoxycarbonyl)-2,3-dimethylpyridine 1 -oxide (5)
[00564] A steel autoclave (Parr "bomb") was charged with 5-Bromo-4-methoxy-2,3-dimethyl- pyridine 1 -oxide (4, 14.0 g, 60.3 mmol), [(R)-(+)-2,22-Bis(diphenylphosphino)-1 ,12- binaphthyl]palladium(ll) chloride (483 mg, 0.603 mmol), Triethylamine (18.0 mL, 129 mmol) and Methanol
(70.0 mL) and a stir bar. The autoclave was degassed and pressurized to 100 PSI with carbon monoxide gas, then heated at 80 0C while stirring for 8 hrs. The reaction vessel was cooled to room temperature, and the reaction mixture was evaporated to yield the crude material 5 (16.9 g, 85% yield, 64% purity).
Refer to Albaneze-Walker J. et. al, Organic Letters, 2004 , 6, 2097-2100.
Step 4. Methyl β-facetoxymethyl^-methoxyS-methylnicotinate (6)
[00565] A solution of 4-methoxy-5-(methoxycarbonyl)-2,3-dimethylpyridine 1 -oxide (5, 17.6 g,
53.3 mmol) in acetic anhydride (130 mL) was heated to 100 C for 2 hours under nitrogen, and evaporated. The crude material was purified by flash chromatography (Hexane: acetone=4:1) to yield the product 6 as a yellow oil (4.6 g, 21% yield, 61% purity).
Step 5. Methyl 6-(acetoxymethyl)-4-methoxy-5-methylnicotinate (7)
[00566] A heavy-wall, pressure-resistant glass vessel was charged with methyl 6-
(acetoxymethyl)-4-methoxy-5-methyl nicotinic acid (6, 6.00 g, 14.4 mmol), triethylamine (60.OmL,
0.430mol) and MeOH (60.OmL). The sealed vessel was heated to 90 0C for 12 hours, and evaporated to give 7 as a crude material which was used in the next step without further purification. (5.0 g, 82% yield,
50% purity).
Step 6. Methyl 6-(chloromethyl)-4-methoxy-5-methylnicotinate (8)
[00567] A solution of methyl 6-(hydroxymethyl)-4-methoxy-5-methylnicotinate (7, 5.0 g, 12 mmol) in dry methylene chloride (30 mL) was treated with thionyl chloride (3.1 mL, 43 mmol) and stirred at room temperature for 30 min (HPLC Rt=5.102 min). The mixture was evaporated, diliuted with water, and purified by preparative HPLC (50X150mm C18 column, 50mL/min; Solvent A: 0.1%TFA water; solvent B:
0.05% TFA Acetonitrile; Gradient: 5% B to 20%B over 5 min then 20% to 60% over 20min). The desired fractions were neutralized and extracted with DCM (300 mL x 2) to yield the desired product. (1.1 g, 39% yield, 95% purity).
Part B. 2-(2-amino-4-chloro-7-((5-(hydroxymethyl)-4-methoxy-3-methylpyridin-2-yl)methyl)-6, 7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (CX) Step 1. (6-(chloromethyl)-4-methoxy-5-methylpyridin-3-yl)methanol
[00568] To a stirred solution of methyl 6-(chloromethyl)-4-methoxy-5-methylnicotinate (0.4 g, 1.74 mmol) in dry ether (30 ml_) was added LiAIH4 (79 mg, 2.09 mmol) portionwise at 0 0C, and the reaction mixture was stirred for 30 min. After consumption of the starting material (by TLC), the reaction was quenched with water (35 mL) and extracted with EtOAc (3 x 35 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (0.28 g, 81%), which was used in the next step without any further purification. 1H NMR (200 MHz, CDCI3) δ 8.36 (s, 1 H),
4.74 (s, 2H), 4.69 (s, 2H), 3.85 (s, 3H), 2.36 (s, 3H), 2.21 (br, 1 H). Mass: 202.0 (M+1).
Step 2. 2-(chloromethyl)-4-methoxy-3-methyl-5-((tetrahydro-2H-pyran-2-yloxy)methyl)pyridine
[00569] To a stirred solution of (6-(chloromethyl)-4-methoxy-5-methylpyridin-3-yl)methanol (0.28 g, 1.39 mmol) in dry CH2CI2 (30 mL) was added 3, 4-dihydroxy 2-H pyran (0.14 g, 1.667 mmol) followed by a catalytic amount of PTSA (10 mg) and stirred at 0 0C for 30 minutes. After consumption of the starting material (by TLC), the reaction mixture was diluted with water (20 mL) and extracted with CH2CI2
(2 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (0.20 g, 50%). 1H NMR (200 MHz, CDCI3) δ 8.44 (s, 1 H), 4.85-4.77
(m, 1 H), 4.73 (s, 3H), 4.54 (d, J = 12Hz, 1 H), 3.86 (s, 3H), 3.65-3.58 (m, 2H), 2.34 (s, 2H), 1.86-1.53 (m,
7H). Mass: 285.9 M+1).
Step 3. methyl 2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-((tetrahydro-2H-pyran-2- yloxy)methyl)pyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate
[00570] To a stirred suspension of NaH (0.03 g, 0.77 mmol, 60% dispersion in mineral oil) in DMF
(20 mL) was added a solution of methyl 2-(2-amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)acetate (0.17 g, 0.70 mmol) in DMF (5 mL) followed by dropwise addition of a solution of 2-
(chloromethyl)-4-methoxy-3-methyl-5-((tetrahydro-2H-pyran-2-yloxy)methyl)pyridine (0.2 g, 0.70 mmol) in
DMF (5 mL) and the resulting solution was stirred a RT for 1 h. After consumption of the starting material
(by TLC), the reaction mixture was quenched with water (5 mL) and extracted with EtOAc (2 X 25 mL).
The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 3% MeOH/ CH2CI2 to afford the title compound (0.22 g, 63.8%). 1H NMR (200 MHz, DMSO-c/6) δ 8.28 (br s, 1 H), 6.46 (br s, 2H), 4.71-
4.60 (m, 4H), 4.43 (d, J = 12 Hz, 1 H), 3.77 (s, 3H) 3.75-3.68 (m, 2H), 3.51 (s, 3H), 3.53-3.44 (m, 2H),
3.26-3.18 (m, 1 H), 2.88-2.79 (m, 1 H), 2.29-2.25 (m, 1H), 2.18 (s, 3H), 1.69-1.39 (m, 6H). Mass: 492.2
(M+1).
Step 4. 2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-((tetrahydro-2H-pyran-2- yloxy)methyl)pyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetamide
[00571] A mixture of methyl 2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-((tetrahydro-2H-pyran-
2-yloxy)methyl)pyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (0.02 g, 0.041 mmol) and NH4OH (100 mL) was stirred in a sealed tube at RT for 12 h. After consumption of the starting material (by TLC), the reaction mixture was extracted with EtOAc (3 x 100 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 10% MeOH/ CH2CI2, to afford the title compound ( 0.1 g, 51%). 1H NMR (500 MHz, DMSO-Of6) δ 8.29 (s, 1 H), 7.36 (s, 1H, Exc), 6.84 (s, 1 H, EΞxc ), 6.39 (s, 2H, Exc), 4.71-4.67 (m, 3H), 4.59 (s, 2H), 4.46-4.42 (m, 1H), 3.79-3.76 (m, 1 H), 3.77 (s, 3H), 3.65 (t, J = 10 Hz, 1 H), 3.57-3.47 (m, 2H), 3.17-3.15 (m, 1 H), 2.70-2.63 (m, 1 H), 2.37-2.35 (m, 1 H), 2.18 (s, 3H) 1.71 -1.64 (m, 2H), 1.49-1.47 (m, 4H). Mass: 477.2 (M+1).
Step 5. 2-(2-amino-4-chloro-7-((5-(hydroxymethyl)-4-methoxy-3-methylpyridin-2-yl)methyl)-6, 7- dihydro-5H-pyrrolo[2, 3-d]pyrimidin-5-yl)acetamide (CX)
[00572] To a stirred solution of 2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-((tetrahydro-2H- pyran-2-yloxy)methyl)pyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (0.1 g, 0.21 mmol) in methanol (10 ml.) was added a catalytic amount of p-tolyl sulfonic acid (0.01 g) at 0 0C, and stirring was continued at RT for 1 h. After consumption of the starting material (by TLC), the reaction mixture was adjusted to pH ~ 8 with sat aq. NaHCO3 solution and extracted with EtOAc (13 X 50 ml_). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by silica gel column chromatography eluting with 10-12% MeOH/CH2CI2 to afford the title compound (30 mg, 36.5%). 1H NMR (500 MHz, DMSO-J6) δ 8.30 (s, 1 H), 7.36 (s, 1 H, Exc), 6.83 (s, 1H, Exc), 6.39 (s, 2H, Exc), 5.16 (t, J = 5.5 Hz, 1 H, Exc), 4.62-4.54 (m, 2H), 4.51 (d, J= 5Hz, 2H), 3.76 (s, 3H), 3.63-3.59 (m, 1 H), 3.46-3.40 (m, 1 H), 3.17-3.11 (m, 1 H), 2.70-2.66 (m, 1 H), 2.21 -2.16 (m, 4H). LC-MS (10-90[12]-25 method): Rt = 10.06 min. (92%); m/z = 393 (M+1). Example 103
2-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-morpholin-4-yl-ethanone (CXI)
[00573] A solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (18.0 mg, 0.0476 mmol) in N,N-dimethylformamide (1.00 mL) was treated with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (12.2 mg, 0.0635 mmol), triethylamine (36.9 μL, 0.265 mmol) and 1 -hydroxybenzotriazole (1.43 mg, 0.0106 mmol) at 50 0C overnight. The mixture was diluted with water (30.0 mL), the pH was adjusted to 4.0 with 2N HCI, and the resulting solution was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH). The desired fractions were lyophilized to give the title compound as a white power (3.2 mg, purity ≥ 90%, yield 14.0%). 1H NMR (Acetone-d6): δ ppm 8.13 (s, 1 H), 5.78 (s, 2H), 4.62(dd, 2H), 3.78 (s, 3H), 3.76 (m, 1 H), 3.57 (m, 4H), 3.50 (m, 4H), 3.20 (m, 1 H), 2.92 (m, 1 H), 2.55 (m, 1 H), 2.24 (s, 6H). LC- MS (5-100-7 method) :Rt=4.105 min; M+1 =446.6. Example 104
2-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N,N-dimethyl-acetamide (CXII)
Step 1. [2-Amino-4-chloro-7-(4-methoxy-3,5-diιnethyl-pyridin-2-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid pentafluorophenyl ester [00574] A solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (150 mg, 0.400 mmol) in tetrahydrofuran (5.00 mL) and triethylamine (276.7 μL, 1.98 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (102.6 μl_, 0.595 mmol) at room temperature for 30 minutes. The reaction mixture was evaporated, diluted with sat. NH4CI solution (20.0 mL), extracted with dichloromethane (20.0 ml_x3), dried over Na2SO4, and evaporated to give the title compound as a white solid (210 mg, purity 30.0%, yield 29.0%). LC-MS (5- 100-7 method):Rt=6.051 min; M+1=543.4.
Step 2. 2-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimldin-5-yl]-N,N-dimethyl-acetamide (CXII)
[00575] [2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid pentafluorophenyl ester (70.0 mg, 0.129 mmol) was treated with 1.0 M of dimethylamine in tetrahydrofuran (0.460 mL, 0.460 mmol) at room temperature for 30 minutes and evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (8.80 mg, purity 99.0%, 17.0%). 1H NMR (DMSO-d6): δ ppm 8.12 (s, 1H), 5.72 (br. s, 2H), 4.62 (d, 1H), 4.54 (d, 1 H), 3.75 (s, 3H), 3.71 (m, 1 H), 3.54 (m, 1 H), 3.15 (m, 1 H), 2.95 (m, 1 H), 2.92 (s, 3H), 2.90 (s, 3H), 2.52 (m, 1 H), 2.20 (s, 6H). LC-MS (5- 100-7 method): Rt=4.135 min; M+1 =404.6. Example 105
2-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-cyclopropyl-acetamide (CXIII)
[00576] A solution of [2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid pentafluorophenyl ester (70.0 mg, 0.129 mmol) in tetrahydrofuran (1.50 mL) was treated with cyclopropylamine (300 μL, 4.00 mmol) at room temperature for 30 minutes and evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (10.0 mg, purity 99.0%, 20.0%). 1H NMR (acetone-d6): δ ppm 8.10 (s, 1 H), 7.77 (br. s, 1 H), 4.62 (d, 1 H), 4.55 (d, 1 H), 3.75 (s, 3H), 3.58- 3.54 (m, 2H), 3.23 (m, 1 H), 2.73 (m, 1 H), 2.58 (m, 1 H), 2.20 (m, 1 H), 2.19 (s, 6H), 0.60 (m, 2H), 0.35 (m, 2H). LC-MS (5-100-7 method): Rt=4.200 min; M+1=416.6. Example 106
2-[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-azetidin-1-yl-ethanone (CXIV)
[00577] A solution of [2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid pentafluorophenyl ester (70.0 mg, 0.129 mmol) was treated with azetidine (100 mg, 2.00 mmol) at room temperature for 30 minutes and evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (9.70 mg, purity 99.0%, yield 18.0%). 1H NMR (acetone-d6): δ ppm 8.14 (s, 1H), 5.74 (br. s, 2H), 4.65 (d, 1H), 4.51 (d, 1H), 4.10 (m, 2H), 3.80 (m, 2H), 3.75 (s, 3H), 3.47 (t, 1H), 3.41 (m, 1 H), 3,21 (m, 1 H), 2.85 (m, 1 H)1 2.62 (m, 1 H), 2.23 (s, 6H), 2.18 (m, 2H). LC-MS (5-100-7 method) :Rt=4.167 min; M+1=416.6.
Example 107
(R)-methyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2, 3-d]pyrimidin-5-yl)acetate (CXV)
[00578] A mixture of (fl)-methyl 2-(2-amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5- yl)acetate (200.0 mg, 0.824 mmol) and potassium fert-butoxide (102 mg, 0.907 mmol) in anhydrous dimethyl sulfoxide (5.00 mL) was treated with a solution of 2-(chloromethyl)-4-methoxy-3,5- dimethylpyridine (161 mg, 0.865 mmol) in dimethyl sulfoxide (1.00 mL) at room temperature for 1 hour.
The reaction mixture was diluted with ice-water (20.0 mL) and extracted with a mixture of dichloromethane/isopropanol (95:5, 20.0 mLx3). The organic layer was dried over Na2SO4 and evaporated. The residue was purified by flash chromatography (dichloromethane/MeOH, 100:0 to 100:5) to give the title compound as a white solid (220.0 mg, purity 96.0%, yield 65.0%). 1H NMR (DMSOd6): δ ppm 8.16 (s, 1 H), 6.46 (br. s, 2H), 4.57 (s, 2H), 3.74 (s, 3H), 3.72 (m, 2H), 3.57 (s, 3H), 3.50 (m, 1 H), 3.20
(m, 1 H), 2.82 (m, 1 H), 2.19 (s, 3H), 2.17 (s, 3H).
Example 108
2-[(R)-2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-methyl-acetamide (CXVI)
Step 1. (R)-perfluorophenyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
6, 7-dihydro-5H-pyrrolo[2, 3-d]pyrimidin-5-yl)acetate
[00579] A solution of lithium (fl)-2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (160.0 mg, 0.325 mmol) in tetrahydrofuran
(5.00 mL) and triethylamine (226.7 μL) was treated with trifluoroacetic acid pentafluorophenyl ester (84.10 μL, 0.488 mmol) at room temperature for 30 minutes. The reaction mixture was evaporated, diluted with sat. NH4CI solution, extracted with dichloromethane (20.0 ml_x3), dried over Na2SO4, and evaporated to give the title compound (140.0 mg, purity 95.0%, yield 79.0%). LC-MS (5-100-7 method):Rt=6.311 min;
M+1 =543.4.
Step 2. 2-[(R)-2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-N-methyl-acetamide
[00580] A mixture of (f?)-perfluorophenyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-
2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (80.0 mg, 0.147 mmol) in anhydrous tetrahydrofuran (1.00 mL) was treated with 2.0 M of methylamine in tetrahydrofuran (0.322 mL, 0.644 mmol) at room temperature for 30 minutes and evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (29.0 mg, purity 99.0%, yield 50.4%). 1H NMR (CDCI3): δ ppm 8.30 (br. s, 1 H), 5.01 (br. s, 2H), 4.73 (d, 2H),
4.62 (d, 1H), 3.86 (s, 3H)1 3.72 (m, 2H), 3.52 (m, 2H), 2.86 (m, 1 H), 2.80 (d, 3H), 2.48 (m, 1 H), 2.33 (s,
3H), 2.31 (s, 3H). LC-MS (5-100-7 method): Rt=3.972 min; M+1=390.6 Example 109
(R)-2^2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-N-(2-methoxyethyl)acetamide (CXVII)
[00581] A mixture of (fi)-perfluorophenyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-
2-yl)methyl)-6,7-dihydro-5H-pyrro!o[2,3-cl]pyrimidin-5-yl)acetate (60.0 mg, 0.110 mmol) in anhydrous tetrahydrofuran (1.00 ml_) was treated with 2-methoxyethylamine (55.9 μl_, 0.644 mmol) at room temperature for 30 minutes and evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (23.0 mg, purity 99.0%, yield 47.9%). 1H NMR (CDCI3): δ ppm 8.28 (br. s, 1H), 5.12 (br. s, 2H), 4.70 (s, 2H), 3.85 (s, 3H), 3.74 (m, 2H), 3.46 (m, 5H), 3.37 (s, 3H), 2.89 (m, 1 H), 2.44 (m, 1 H), 2.31 (s, 3H), 2.27 (s, 3H). LC-MS (5-100-7 method):Rt=4.077 min; M+1 =434.6 Example 110
[(R)-2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-acetic acid (CXVIII)
[00582] A mixture of (fl)-methyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (120.0 mg, 0.306 mmol) was alkylated with 5-(chloromethyl)-2,3,4-trimethylpyridine using the general alkylation procedure. The resulting crude material was dissolved in dimethyl sulfoxide (1.00 mL) and was treated with 6 M of hydrogen chloride in water (0.425 mL, 2.55 mmol) at 80 0C for 2.0 hours. After cooling, the mixture was diluted with water (25.0 mL) and purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (100.0 mg, purity 99.0%, yield 90.0%). 1H NMR (DMSO-d6): δ ppm 12.34 (br. s, 1 H), 8.11 (s, 1 H), 6.55 (br. s, 2H), 4.50 (dd, 2H), 3.51 (m, 1H), 3.43 (m, 1 H), 3.07 (m, 1 H), 2.77 (m, 1 H), 2.45 (s, 3H), 2.35 (m, 1 H), 2.18 (s, 6H). LC-MS (5-100-7 method):Rt=3.737 min; M+1=361.6 Example 111
2-[(R)-2-Amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-acetamide (CXIX)
[00583] A solution of [(fl)-2-amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (210.0 mg, 0.580 mmol) in tetrahydrofuran (10.0 mL, 0.123 mol) and triethylamine (385 μL, 2.76 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (143 μL, 0.829 mmol) at room temperature for 20 minutes. 0.5 M of ammonia in dioxane (11.0 mL, 5.53 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The residue was purified by flash chromatography (dichloromethane/ methanol, 100:1 to 100:15) to give the title compound as a white powder (200.0 mg, purity 99.0%, yield 100.0%). 1H NMR (DMSO-d6): δ ppm 8.26 (s, 1H), 7.39 (br. s, 1 H), 6.85 (br. s, 1H), 6.53 (br. s, 1 H), 4.60 (d, 1H), 4.49 (d, 1 H), 3.52 (m, 1 H), 3.46 (m, 1 H), 3.09 (m, 1 H), 2.69 (m, 1 H), 2.53 (s, 3H), 2.29 (s, 3H), 2.24 (s, 3H), 2.19 (m, 1 H). LC-MS (5- 100-7 method): Rt=3.520 min; M+1 =360.6 Example 112 2-[(R)-2-Amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2)3- d]pyrimidin-5-yl]-N-methyl-acetamide (CXX)
[00584] A solution of [(fl)-2-Amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (50.0 mg, 0.138 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (96.3 μL) was treated with trifluoroacetic acid pentafluorophenyl ester (47.6 μl_, 0.276 mmol) at room temperature for 20 minutes. 2.0 M of methylamine in tetrahydrofuran (0.345 mL, 0.691 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (32.0 mg, purity 99.0%, yield 62.0%). 1H NMR (DMSO-d6): δ ppm 8.07 (s, 1 H), 7.82 (m, 1 H), 6.51 (br. s, 1 H), 4.53 (d, 1 H), 4.42 (d, 1 H), 3.43 (m, 2H), 3.03 (m, 1 H), 2.66 (m, 1 H), 2.50 (s, 3H), 2.43 (s, 3H), 2.18 (s, 3H), 2.17 (m, 1 H). LC-MS (5-100-7 method) :Rt=3.670 min; M+1=374.6 Example 113
2-[(R)-2-Amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-ethyl-acetamide (CXXI)
[00585] A solution of [(fl)-2-amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (50.0 mg, 0.138 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (96.3 μL) was treated with trifluoroacetic acid pentafluorophenyl ester (47.6 μL, 0.276 mmol) at room temperature for 20 minutes. 2.0 M of ethylamine in tetrahydrofuran (0.345 mL, 0.691 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (25.6 mg, purity 99.0%, yield 47.6%). 1H NMR (DMSO-d6): δ ppm 8.07 (s, 1 H), 7.87 (t, 1 H), 6.51 (br. s, 1 H), 4.54 (d, 1 H), 4.40 (d, 1 H), 3.43 (m, 2H), 3.02 (m, 3H), 2.66 (m, 1 H), 2.43 (s, 3H), 2.17 (s, 6H), 2.15 (m, 1 H), 0.95 (t, 3H). LC-MS (5-100-7 method):Rt=3.849 min; M+1 =388.6 Example 114
2-[(R)-2-Amino-4-chloro-7-(4, 5, 6-trimethyl-pyridin-3-ylmethyl)-6, 7-dihydro-5H-pyrrolo[2, 3- d]pyrimidin-5-yl]-N-(2-methoxy-ethyl)-acetamide (CXXII)
[00586] A solution of [(fl)-2-amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (50.0 mg, 0.138 mmol) in tetrahydrofuran (2.00 mL, 0.0246 mol) and triethylamine (96.3 μL, 0.691 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (47.6 μL, 0.276 mmol) at room temperature for 20 minutes. 2-methoxyethylamine (51.9 mg, 0.691 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (45.0 mg, purity 99.0%, yield 78.0%). 1H NMR (DMSO-d6): δ ppm 8.06 (s, 1 H), 8.00 (t, 1 H), 6.52 (br. s, 1 H), 4.54 (d, 1 H), 4.40 (d, 1 H), 3.44 (m, 2H), 3.26 (m, 2H), 3.19 (s, 3H), 3.15 (m, 2H), 3.01 (m, 1 H), 2.65 (m, 1 H), 2.42 (s, 3H)1 2.17 (s, 6H), 2.15 (m, 1 H). LC-MS (5-100-7 method): Rt=3.822 min; M+1=418.6 Example 115 2-[(R)-2-Ammo-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-SH-pyrrolo[2,3- d]pyrimidin-5-yl]-N-ethyl-acetamide (CXXIII)
[00587] A solution of [(/:?)-2-amino-4-chloro-7-(4-nnethoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (50.0 mg, 0.132 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.2 μL, 0.662 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (68.4 μL, 0.397 mmol) at room temperature for 20 minutes. 2.0 M of Ethylamine in tetrahydrofuran (0.331 mL, 0.662 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (23.2 mg, purity 99.0%, yield 43.3%). 1H NMR (DMSO-d6): δ ppm 8.15 (s, 1 H), 7.87 (t, 1 H), 6.41 (br. s, 1 H), 4.56 (s, 2H), 3.72 (s, 3H), 3.60 (t, 3H), 3.57 (m, 1 H), 3.13 (m, 1 H), 3.03 (m, 2H), 2.65 (m, 1 H), 2.18 (s, 3H), 2.16 (s, 3H), 2.15 (m, 1 H), 0.97 (t, 1 H). LC-MS (5-100-7 method):Rt=4.200 min; M+1=404.6 Example 116
2-[(R)-2-Amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-oxetan-3-yl-acetamide (CXXIV)
[00588] A solution of [(/?)-2-amino-4-chloro-7-(4,5,6-trimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H- pyrrolo[2,3-dJpyrimidin-5-yl]-acetic acid (50.0 mg, 0.138 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (96.3 μL) was treated with trifluoroacetic acid pentafluorophenyl ester (47.6 μL, 0.276 mmol) at room temperature for 20 minutes. Oxetan-3-ylamine (50.5 mg, 0.691 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (45.0 mg, purity 99.0%, yield 78.0%). 1H NMR (DMSO-d6): δ ppm 8.67 (d, 1 H), 8.06 (s, 1 H), 6.52 (br. s, 1 H), 4.73 (m, 1 H), 4.66 (m, 2H), 4.53 (d, 1 H), 4.42-4.35 (m, 2H), 4.31 (m, 1 H), 3.44 (m, 2H), 3.03 (m, 1 H), 2.71 (m, 1 H), 2.42 (s, 3H), 2.18 (s, 3H), 2.17 (s, 3H), 2.15 (m, 1 H). LC-MS (5-100-7 method):Rt=3.719 min; M+1=416.6 Example 117
(R)-2-(2-am\no-A-chloro-7-((4-methoxy-3,5-dimethyipyndin-2-y\)methyl)-6J-dibydTO-5H-pyrro\o[2,3- d]pyrimidin-5-yl)-N-(oxetan-3-yl)acetamide (CXXV)
[00589] A solution of [(fl)-2-amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (50.0 mg, 0.132 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.2 μL, 0.662 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (68.4 μL, 0.397 mmol) at room temperature for 20 minutes. Oxetan-3-ylamine (48.4 mg, 0.662 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (19.2 mg, purity 99.0%, yield 33.5%). 1H NMR (DMSO-d6): δ ppm 8.67 (d, 1 H), 8.15 (s, 1 H), 6.42 (br. s, 1 H), 4.74 (m, 1 H), 4.67 (m, 2H), 4.56 (s, 2H), 4.39-4.32 (m, 4H), 3.72 (s, 3H), 3.61 (t, 1 H), 3.45 (m, 1 H), 3.13 (m, 1 H), 2.71 (m, 1 H), 2.42 (s, 3H), 2,25 (m, 1 H), 2.18 (s, 3H), 2.17 (s, 3H). LC-MS (5-100-7 method): Rt=4.011 min; M+1=432.5 Example 118
2-[(R)-2-Amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-methyl-acetamide (CXXVI)
[00590] A solution of [(/:?)-2-amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (120.0 mg, 0.319 mmol) in tetrahydrofuran (4.00 ml_) and triethylamine (222 μl_, 1.60 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (110 μL, 0.638 mmol) at room temperature for 20 minutes. 2.0 M of methylamine in tetrahydrofuran (0.798 mL, 1.60 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (53.0 mg, purity 99.0%, yield 43.0%). 1H NMR (DMSO-d6): δ ppm 8.05 (s, 1 H), 7.83 (d, 1H), 6.51 (br. s, 1 H), 4.53 (d, 1H), 4.40 (d, 1 H), 3.45 (m, 2H), 3.05 (m, 1 H), 2.66 (m, 3H), 2.51 (d, 3H), 2.46 (s, 3H), 2.19 (s, 3H), 2.16 (m, 1 H), 1.09 (t, 3H). LC-MS (5- 100-7 method): Rt=3.948 min; M+1 =388.6 Example 119
2-[(R)-2-Amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-ethyl-acetamide (CXXVII)
[00591] A solution of [(R)-2-amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyhdin-3-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (120.0 mg, 0.319 mmol) in tetrahydrofuran (4.00 mL) and triethylamine (222 μL, 1.60 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (110.0 μL, 0.638 mmol) at room temperature for 20 minutes. 2.0 M of ethylamine in tetrahydrofuran (0.798 mL, 1.60 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (55.0 mg, purity 99.0%, yield 43.0%). 1H NMR (DMSO-d6): δ ppm 8.06 (s, 1 H), 7.88 (t, 1 H), 6.51 (br. s, 1 H), 4.54 (d, 1 H), 4.39 (d, 1 H), 3.45 (m, 2H), 3.03 (m, 3H), 2.63 (m, 3H), 2.46 (s, 3H), 2.46 (s, 3H), 2.19 (s, 3H), 2.13 (m, 1 H), 1.09(t, 3H), 0.95 (t, 3H). LC-MS (5-100-7 method): Rt= 4.160 min; M+1 =402.7 Example 120
2-[(R)-2-Amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7-dihydrt)-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-((S)-2-hydroxymethyl-morpholin-4-yl)-ethanone (CXXVIII) [00592] A solution of [(f?)-2-amino-4-chloro-7-(5-ethyl-4,6-dimethyl-pyridin-3-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl]-acetic acid (80.0 mg, 0.213 mmol) in tetrahydrofuran (5.00 mL) and triethylamine (148 μL, 1.06 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (73.4 μL, 0.426 mmol) at room temperature for 20 minutes. (S)-1-Morpholin-2-yl-methanol (125 mg, 1.06 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (45.0 mg, purity 99.0%, yield 44.0%). 1H NMR (DMSO-d6): δ ppm 8.05 (s, 1 H)1 6.52 (br. s, 1 H), 4.52 (m, 1 H), 4.42 (d, 1 H), 4.27-3.75 (m, 4H), 3.68-2.38 (m, 12H), 2.46 (s, 3H), 2.19 (s, 3H), 1.06 (t, 3H). LC-MS (5-100-7 method): Rt= 3.997 min; M+1 =474.6 Example 121
2-[(R)-2-Amino-4-chloro-7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-(2-hydroxymethyl-morpholin-4-yl)-ethanone (CXXIX)
[00593] A solution of lithium (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μl_, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. Morpholin-2-yl-methanol (77.9 mg, 0.665 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (32.5 mg, purity 99.0%, yield 51.4%). 1H NMR (DMSO-d6): δ ppm 8.13 (s, 1H), 6.52 (br. s, 1 H), 4.93 (m, 1 H), 4.77 (m, 2H), 4.29-2.76 (m, 15H), 2.34 (m, 1 H), 2.26 (s, 3H), 2.21 (s, 3H), 1.05 (t, 3H). LC-MS (5-100-7 method):Rt= 4.183 min; M+1=474.6 Example 122
2-[(R)-2-Amino-4-chlorθ'7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-((S)-2-hydroxymethyl-morpholin-4-yl)-ethanone (CXXX)
[00594] A solution of lithium (fl)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μL, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. (S)-1 -Morpholin-2-yl-methanol (77.9 mg, 0.665 mol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (34.5 mg, purity 99.0%, yield 54.6%). 1H NMR (DMSO-d6): δ ppm 8.15 (s, 1H), 6.47 (br. s, 1H), 4.77 (m, 1 H), 4.62 (m, 2H), 4.29-2.76 (m, 15H), 2.36 (m, 1H), 2.27 (s, 3H), 2.22 (s, 3H), 1.06 (t, 3H). LC-MS (5-100-7 method):Rt= 4.182 min; M+1 =474.6 Example 123
2-[(R)-2-Amino-4-chloro-7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-((3S,4S)-3,4-dihydroxy-pyrrolidin-1-yl)-ethanone (CXXXI) [00595] A solution of lithium (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μL, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. (3S,4S)-Pyrrolidine-3,4-diol (68.6 mg, 0.665 mmol) was added to the mixture and was stirred for 20 minutes at room temperature. The reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (8.10 mg, purity 99.0%, yield 13.0%). 1H NMR (DMSO-de): δ ppm 8.09 (s, 1H)1 6.41 (br. s, 1H), 5.34 (d, 1H), 5.16 (d, 1H), 4.58 (s, 2H), 4.05-2.60 (m, 12H), 2.43 (m, 1H), 2.23 (s, 3H), 2.21 (s, 3H), 1.06 (t, 3H). LC-MS (5-100-7 method):Rt= 3.952 min; M+1 =460.6 Example 124
2-[(R)-2-Amino-4-chloro-7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-((S)-2-hydroxymethyl-pyrrolidin-1-yl)-ethanone (CXXXII) [00596] A solution of lithium (f7)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 ml_) and triethylamine (92.7 μl_, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μl_, 0.266 mmol) at room temperature for 20 minutes. L-Prolinol (67.3 mg, 0.665 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (30.0 mg, purity 99.0%, yield 50.0%). 1H NMR (DMSO-d6): δ ppm 8.11 (s, 1 H), 6.44 (br. s, 1 H), 4.69-4.48 (m, 3H), 3.90-2.61 (m, 15H), 2.38 (m, 1 H), 2.25 (s, 3H), 2.22 (s, 3H), 1.05 (t, 3H). LC-MS (5-100-7 method):Rt= 4.463 min; M+1=458.6 Example 125
2-[(R)-2-Amino-4-chloro-7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-((R)-3-hydroxy-pyrrolidin-1-yl)-ethanone (CXXXIII)
[00597] A solution of lithium (fl)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μL, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. (S)-Pyrrolidin-3-ol (57.9 mg, 0.665 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (33.6 mg, purity 99.0%, yield 56.8%). 1H NMR (DMSO-d6): δ ppm 8.14 (s, 1 H), 6.46 (br. s, 1 H), 4.95 (m, 1 H), 4.62 (m, 2H), 4.25 (m, 1 H), 3.68-2.23 (m, 11 H), 2.27 (s, 3H), 2.24 (s, 3H), 1.86-1.71 (m, 2H), 1.05 (t, 3H). LC-MS (5-100-7 method):Rt= 4.165 min; M+1 =444.6 Example 126
2-[(R)-2-Amino-4-chloro-7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-N-((R)-2,3-dihydroxy-propyl)-acetamide (CXXXIV)
[00598] A solution of lithium (/7)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μL, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. (S)-3-Amino-propane-1 ,2-diol (60.6 mg, 0.665 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (19.7 mg, purity 99.0%, yield 33.0%). 1H NMR (DMSO-d6): δ ppm 8.14 (s, 1H), 7.92 (m, 1 H), 6.44 (br. s, 1 H), 4.70-4.50 (m, 4H), 3.60 (t, 1 H), 3.44-3.12 (m, 7H), 2.98 (m, 1H), 2.74- 2.63 (m, 3H), 2.25 (s, 3H), 2.22 (s, 3H), 1.05 (t, 3H). LC-MS (5-100-7 method): Rt= 3.960 min; M+1 =448.6 Example 127
2-((R)-2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-N-((S)-1-hydroxypropan-2-yl)acetamide (CXXXV)
[00599] A solution of lithium (fl)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μl_, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. (_?S)-2-Aminopropan-1-ol (50.0 mg, 0.665 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% formic acid) to give the title compound as a white solid (34.7 mg, purity 99.0%, yield 60.2%). 1H NMR (DMSOd6): δ ppm 8.09 (s, 1H), 7.68 (d, 1 H), 6.41 (br. s, 1 H), 4.62 (m, 1 H), 4.56 (dd, 2H), 3.86 (m, 1 H), 3.71 (m, 1 H), 3.58 (m, 1H), 3.44 (m, 1H), 3.12 (m, 2H), 2.64 (m, 3H), 2.23 (s, 3H), 2.20 (s, 3H), 2.13 (m, 1H), 1.07 (t, 3H), 0.996 (t, 3H). LC-MS (5-100-7 method):Rt= 4.218 min; M+1=432.6 Example 128
2-[(R)-2-Amino-4-chloro-7-(4-ethyl-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl]-1-[2-(2-hydroxy-ethyl)-morpholin-4-yl]-ethanone (CXXXVI)
[00600] A solution of lithium (fl)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (50.8 mg, 0.133 mmol) in tetrahydrofuran (2.00 mL) and triethylamine (92.7 μL, 0.665 mmol) was treated with trifluoroacetic acid pentafluorophenyl ester (45.8 μL, 0.266 mmol) at room temperature for 20 minutes. 2-Morpholin-2-yl-ethanol (87.2 mg, 0.665 mmol) was added. After stirring for 20 minutes at room temperature, the reaction mixture was evaporated. The crude was purified by reverse-phase preparative HPLC (5 to 30% CH3CN/H2O, 0.1% HCOOH) to give the title compound as a white solid (31.6 mg, purity 99.0%, yield 48.6%). 1H NMR (DMSO-d6): δ ppm 8.14 (s, 1H), 6.46 (br. s, 1 H), 4.62 (s, 2H), 4.20 (m, 1 H), 4.11-2.60 (m, 14H), 2.35 (m, 1H), 2.27 (s, 3H), 2.23 (s, 3H)1 1.54 (m, 2H), 1.07 (t, 3H), 0.996 (t, 3H). LC-MS (5-100-7 method):Rt= 4.297 min; M+1 =488.6 Example 129
(R)-methyl 2-(2-amino-4-chloro-7-((5-methoxy-4,6-dimethylpyridin-3-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetate (CXXXVII)
[00601] The title compound was obtained by reacting (fl)-methyl 2-(2-amino-4-chloro-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (501 mg, 2.06 mmol) with 5-(chloromethyl)-3-methoxy-2,4- dimethylpyridine (502 mg, 1.31 mmol), using the general alkylation procedure. A white solid was collected. (221 mg, purity >90%, yield 27%). 1H NMR (CDCI3) δ ppm 8.09 (s, 1 H), 4.89 (s, 2H),4.54 (q, 2H), 3.75 (s, 3H), 3.67 (s, 3H), 3.69 (m, 2H), 3.16 (q, 1 H), 3.05(dd, 1 H), 2.53 (s, 3H), 2.42 (m, 1 H), 2.24 (s, 3H), LC/MS: rt (5-100-5 method) = 1.962 min.; 392.1 (M+1 , 100 %), 394.1 (M+3, 35%). Example 130
(Rj-Σ^Σ-amino^-chloro-T-tfS-methoxy^β-dimethylpyridin-S-ylJmethyO-βJ-dihydro-SH-pyrrolofcS- d]pyrimidiπ-5-yl)acetic acid (CXVIII)
[00602] The title compound was obtained by hydrolysis of (/?)-methyl 2-(2-amino-4-chloro-7-((5- methoxy-4,6-dimethylpyridin-3-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (220 mg,
0.56 mmol) with LiOH (1 eq) in THF/water at 25-50 °C for 1 h, and evaporation. A white solid was collected. (180 mg, purity >90%, yield 76%). 1H NMR (DMSO-d6) δ ppm 8.04 (s, 1H), 6.52 (s, 2H), 4.55
(q, 2H), 3.67 (s, 3H), 3.59(t, 1 H), 3.44 (m, 1 H), 3.28 (dd, 1 H), 3.09 (m, 1H), 2.77 (dd, 1 H), 2.40 (s, 3H),
2.31 (m, 1 H), 2.17 (s, 3H), LC/MS: rt (5-100-5 method) = 1.751 min.; 378.1 (M+1 , 100 %), 380.1 (M+3,
35%).
Example 131
(R)-2^2-amino^-chloro-7-((5-methoxy^,6-dimethylpyridin-3-yl)methyl)-6J-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetamide (CXXXIX)
[00603] The title compound was obtained by reacting (fl)-2-(2-amino-4-chloro-7-((5-methoxy-4,6- dimethylpyridin-S-yOmethyO-ey-dihydro-δH-pyrrolo^.S-dJpyrimidin-S-yOacetic acid (180 mg, 0. 48 mmol) with trifluoroacetic acid pentafluorophenyl ester (600 μl_, 3 mmol), followed by ammonia in dioxane (2ml,
0.5M, 20 mmol), using the general amide formation procedure. A white solid was collected (160 mg, purity >90%, yield 89%). 1H NMR (CDCI3) δ ppm 9.70 (b, s, 1 H), 8.15 (s, 1 H), 6.38 (d, 1 H), 6.100 (b, s,
1 H), 4.88 (dd, 2H), 3.83 (t, 1 H), 3.79 (s, 3H), 3.65 (m, 2H), 2.78 (dd, 3H), 2.70 (m, 1 H), 2.43 (s, 3H), 2.30
(m, 1 H) 2.27 (s, 3H), LC/MS: rt (5-100-7 method) = 3.991 min.; 406.6 (M, 100 %), 408.6 (M, 35%).
Example 132
(fy-Σ^Σ-amino^-chloro-T-tfS-methoxy^β-dimethylpyrMin-S-yOmethyty-βJ-dihydro-SH-pyrroloRS- d]pyrimidin-5-yl)-N-methylacetamide (CXL)
[00604] The title compound was obtained by reacting (/?)-2-(2-amino-4-chloro-7-((5-methoxy-4,6- dimethylpyridin-3-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (45 mg, 0. 12 mmol) with trifluoroacetic acid pentafluorophenyl ester (150 μL, 0.75 mmol), followed by methylamine (2 mL, 2M,
4 mmol) using the general amide formation procedure. A white solid was collected (34.6 mg, purity >90%, yield 75%). 1H NMR (CDCI3) δ ppm 8.10 (s, 1 H), 5.63 (s, 1H), 4.95 (s, 2H), 4.56 (q, 2H), 3.74(s, 3H), 3.62
(m, 2H), 3.29 (m, 1 H), 2.89 (m, 1 H), 2.78 (d, 3H), 2.51 (s, 3H), 2.27 (m, 1 H) 2.23 (s, 3H), LC/MS: rt (5-
100-5 method) = 1.738 min.; 391 (M+1 , 100 %), 393 (M+3, 35%).
Example 133
(R) -2-(2-amino-4-chloro-7-((5-methoxy-4, 6-dimethylpyridin-3-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)-N-(2-methoxyethyl)acetamide (CXLI)
[00605] The title compound was obtained by reacting (f?)-2-(2-amino-4-chloro-7-((5-methoxy-4,6- dimethylpyridin-3-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (30 mg, 0.08 mmol) with trifluoroacetic acid pentafluorophenyl ester (150 μL, 0.75 mmol), followed by 2-methoxyethanamine
(0.1g, 1 mmol) using the general amide formation procedure. A white solid was collected (24 mg, purity >90%, yield 69%). 1H NMR (CDCI3) δ ppm 8.08 (s, 1 H), 6.32 (s, 1 H), 5.01 (m, 1 H), 4.93 (s, 4H), 4.56 (m,
4H), 3.75 (s, 3H), 3.68 (m, 2H), 3.27 (d, 1 H), 2.92 (d, 1 H), 2.52 (s, 3H), 2.27 (m, 1H) 2.24 (s, 3H), LC/MS: rt (5-100-5 method) = 1.756 min.; 433.1 (M+1 , 100 %), 435.2 (M+3, 35%).
Example 134
(flJ-Σ^-amino-α-chloro-T-ttS-methoxy^.e-dimethylpyridin-S-yOmethylJ-β.T-dihydro-SH-pyrrolo^.a- d]pyrimidin-5-yl)-N-(oxetan-3-yl)acetamide (CXLH)
[00606] The title compound was obtained by reacting (f?)-2-(2-amino-4-chloro-7-((5-methoxy-4,6- dimethylpyridin-3-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (30 mg, 0.08 mmol) with trifluoroacetic acid pentafluorophenyl ester (150 ml_, 0.75 mmol), followed by oxetan-3-amine (0.1 g,
1 mmol) using the general amide formation procedure. A white solid was collected (18 mg, purity >90%, yield 50%). 1H NMR (CDCI3) δ ppm 8.09 (s, 1 H), 5.99 (s, 1 H), 4.93 (s, 2H), 4.52 (s, 2H), 3.75 (s, 3H), 3.68
(m, 2H), 3.42 (s, 3H), 3.35 (s, 3H), 3.29 (m, 1H), 2.89 (m, 1 H), 2.52 (s, 3H), 2.27 (m, 1H) 2.23 (s, 3H),
LC/MS: rt (5-100-5 method) = 1.831 min.; 433.1 (M+1 , 100 %), 435.2 (M+3, 35%).
Example 135
(Symethyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetate (CXLIII)
[00607] The title compound was obtained by reacting (S)-methyl 2-(2-amino-4-chloro-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (195 mg, O.βOmmol) with 2-(chloromethyl)-4-methoxy-3,5- dimethylpyridine (182 mg, 0.98 mmol), using the general alkylation procedure. A white solid was collected (190 mg, purity >90%, yield 54%). 1H NMR (CDCI3) δ ppm 8.19 (s, 1 H), 4.86 (s, 2H),4.64 (s,
2H), 3.78 (S1 3H), 3.76 (t, 1 H), 3.67 (s, 3H), 3.64 (m, 1H), 3.24 (dd, 1H), 3.03(dd, 1 H), 2.49 (m, 1H), 2.26
(s, 3H),2.23 (s, 3H). LC/MS: rt (5-100-7 method) = 4.520 min.; 391.6 (M+1 , 100 %), 393.6 (M+3, 35%).
Example 136
(S)-2-(2-amino^-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetic acid (CXLIV)
[00608] The title compound was obtained by hydrolysis of (S)-methyl 2-(2-amino-4-chloro-7-((4- methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (95 mg, 0.24 mmol), with LiOH (1 eq) in THF/water at 25-50 0C for 1 h, and evaporation. A white solid was collected.
(65 mg, purity >90%, yield 64%). 1H NMR (DMSO-d6) δ ppm 12.33 (b, s, 1 H), 8.23 (s, 1 H), 6.52 (s, 2H),
4.61 (s, 2H), 3.78 (s, 3H), 3.73(m, 2H), 3.20 (dd, 1H)1 2.80 (dd, 1 H), 2.43 (dd, 1H), 2.23 (s, 3H), 2.18 (s,
3H), LC/MS: rt (5-100-7method) = 4.055min.; 377.6 (M+1 , 100 %), 379.6 (M+3, 35%).
Example 137
(S)-2^2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-N-(oxetan-3-yl)acetamide (CXL V)
[00609] The title compound was obtained by reacting (S)-2-(2-amino-4-chloro-7-((5-methoxy-4,6- dimethylpyridin-S-ylJmethylJ-ej-dihydro-SH-pyrrolo^.S-dlpyrimidin-δ-yOacetic acid (45 mg, 0.12 mmol) with trifluoroacetic acid pentafluorophenyl ester (102.6 μL, 0.60 mmol), followed by oxetan-3-amine (68 mg, 0.93mmol) using the general amide formation procedure. A white solid was collected (27 mg, purity
>90%, yield 47%). 1H NMR (CDCI3) δ ppm 8.17 (s, 1 H), 6.53 (d, 1 H), 5.02 (m, 1 H)1 4.98 (s, 2H), 4.93 (q,
2H), 4.65 (d, 2H), 4.52 (t, 1H), 4.41 (t, 1H), 3.77 (s, 3H), 3.65 (m, 2H), 3.33 (dd,1H), 2.93 (dd, 1 H), 2.33
(m, 1H), 2.26 (s, 3H), 2.23 (s, 3H), LC/MS: rt (5-100-5 method) = 1.913 min.; 433.1 (M+1 , 100 %), 435.2
(M+3, 35%).
Example 138
2-((2-amino-5-(2-amino-2-oxoethyl)-4-chloro-5H-pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4- methoxy-3,5-dimethylpyridine 1 -oxide (CXLVI)
Step 1. 2-(chloromethyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide
[00610] 2-(chloromethyl)-A solution of 4-methoxy-3,5-dimethylpyridine (500 mg, 3 mmol) in dichloromethane (15ml) was treated with m-CPBA (1g, 5.8 mmol). The reaction was monitored via
LC/MS. Excess m-CPBA was added as needed. The reaction mixture was extracted with aqueous NaOH
(1 N) and dichloromethane. The organic layer was dried and removed to yield a white crude solid (0.45 g, purity 85%, yield 70%). 1H NMR (CDCI3) δ ppm 8.06 (s, 1 H), 4.93 (s, 2H), 3.77 (s, 3H), 2.36 (s, 3H), 2.23
(s, 3H)1. LC/MS : rt (5-100-5 method) = 1.955 min.; 202.0 (M+1 , 100 %), 204.0 (M+3, 35%)
Step 2. 2-((2-amino-5-(carboxymethyl)-4-chloro-5H-pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4- methoxy-3,5-dimethylpyridine 1 -oxide
[00611] 2-Amino-4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-acetic acid methyl ester
(420 mg, 1.7 mmol) was dissolved in N,N-Dimethylformamide (5 mL, 150 mmol). The reaction was cooled to 00C and treated with NaH (95 mg, 4.0 mmol). To that reaction mixture, 2-(chloromethyl)-4-methoxy-
3,5-dimethylpyridine 1 -oxide (425 mg, 2.11 mmol) in 1 mL DMF was added. The reaction was stirred at rt for 20 minutes, by which time alkylation had taken place and most of the CO2Me group was hydrolyzed.
Water was used to quench the reaction and TFA was added. The beige solid product was purified by reverse phase HPLC. (450 mg, purity >90%, yield 59%). 1H NMR (DMSO-d6) δ ppm 12.31 (s, 1 H), 8.13
(s, 1 H), 6.48 (s, 2H), 4.7 (s, 2H), 3.70 (s, 3H), 3.68 (t, 1H), 3.40 (m, 1 H), 3.28 (dd, 1 H), 2.75 (dd, 1 H), 2.35
(m, 1 H), 2.28 (s, 3H), 2.17 (s, 3H), LC/MS: rt (5-100-7 method) = 4.104 min.; 393.6 (M+1 , 100 %), 395.6
(M+3, 35%).
Step 3. 2-((2-amino-5-(2-amino-2-oxoethyl)-4-chloro-5H-pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4- methoxy-3,5-dimethylpyridine 1 -oxide (CXLVI)
[00612] To a solution of 2-((2-amino-5-(carboxymethyl)-4-chloro-5H-pyrrolo[2,3-d]pyrimidin-7(6H)- yl)methyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide (30 mg, 0.08 mmol) in THF (5 mL) and DIEA (200 μL,
1 mmol) was treated with thfluoroacetic acid pentafluorophenyl ester (60 μL, 0. 3 mmol) at room temperature for 5 minutes. The reaction was monitored by LC/MS which showed the disappearance of the acid and the formation of the perfluorophenyl ester. Excess ammonia (0.02 g, 1 mmol) in dioxane (0.5
M from Aldrich) was added to the reaction mixture. The reaction was completed within 10 minutes. After removing the solvent, the solid was purified by reverse phase HPLC. (13 mg, purity >90%, yield 40%). 1H
NMR (CDCI3) δ ppm 8.10 (s, 1 H)1 6.42 (b, s, 1H), 6.20 (b, s, 2H), 5.97 (b, S, 1 H), 4.88 (dd, 2H)1 3.88 (t, 1 H), 3.79 (s, 3H), 3.65 (m, 2H), 2.78 (dd, 1 H), 2.42 (s, 3H), 2.35 (m, 1H) 2.26 (s, 3H), LC/MS: rt (5-100-5 method) = 1.807 min.; 393.0 (M+1 , 100 %), 395.0 (M+3, 35%).
Example 139
2-((2-amino-4-chloro-5-(2^methylamino)-2-oxoethyl)-5H-pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4- methoxy-3,5-dimethylpyridine 1 -oxide (CXLVII)
[00613] The title compound was obtained by reacting 2-((2-amino-5-(carboxymethyl)-4-chloro-5H- pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide (30 mg, 0. 08 mmol) with trifluoro-acetic acid pentafluorophenyl ester (60 μL, 0.3 mmol), followed by methyl amine (40 mg, 1 mmol) in THF (2M solution from Aldrich), using the general amide formation procedure. (19 mg, purity >90%, yield 59%). 1H NMR (CDCI3) δ ppm 9.70 (b, s, 1 H), 8.15 (s, 1 H), 6.38 (d, 1 H), 6.100 (b, s, 1 H), 4.88 (dd,
2H), 3.83 (t, 1 H), 3.79 (s, 3H), 3.65 (m, 2H), 2.78 (dd, 3H), 2.70 (m, 1 H), 2.43 (s, 3H), 2.30 (m, 1 H) 2.27
(s, 3H), LC/MS: rt (5-100-7 method) = 3.991 min.; 406.6 (M, 100 %), 408.6 (M, 35%).
Example 140
2-((2-amino-4-chloro-5-(2^dimethylamino)-2-oxoethyl)-5H'pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-
4-methoxy-3,5-dimethylpyridine 1 -oxide (CXLVIII)
[00614] The title compound was obtained by reacting 2-((2-amino-5-(carboxymethyl)-4-chloro-5H- pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide (30 mg, 0. 08 mmol) with trifluoroacetic acid pentafluorophenyl ester (60 μL, 0.3 mmol), followed by dimethyl amine (60 mg, 1 mmol) in THF (2M solution from Aldrich), using the general amide formation procedure. (20 mg, purity
>90%, yield 50%). 1H NMR (CDCI3) δ ppm 8.10 (s, 1 H), 5.9 (br. s, 2H), 4.88 (q, 2H), 3.87 (t, 1 H), 3.76 (s,
3H), 3.65 (m, 1 H), 3.41 (m, 1 H), 3.0 (m, 1 H), 2.96 (s, 3H) 2.90 (s, 3H), 2.45 (m, 1H), 2.33 (s, 3H), 2.23 (s,
3H), LC/MS: rt (5-100-7 method) = 4.205 min.; 420.6 (M, 100 %), 422.6 (M+2, 35%)
Example 141
2-((2-amino-4-chloro-5-(2-(methylamino)-2-oxoethyl)-5H-pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4- methoxy-3,5-dimethylpyridine 1 -oxide (CXLIX)
[00615] The title compound was obtained by reacting 2-((2-amino-5-(carboxymethyl)-4-chloro-5H- pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide (30 mg, 0. 08 mmol) with trifluoroacetic acid pentafluorophenyl ester (60 μL, 0.0003 mol), followed by diethyl amine (100 mg, 1 mmol) using the general amide formation procedure (12 mg, purity >90%, yield 30%). 1H NMR (CDCI3) δ ppm 8.06 (s, 1 H), 5.20 (s, 2H), 4.88 (q, 2H), 3.83 (t, 1 H), 3.75 (s, 3H), 3.65 (m, 1 H), 3.40 (m, 2H), 3.25
(m, 4H), 3.02 ( m, 1H), 2.40 (m, 1H), 2.32 (s, 3H), 2.23 (s, 3H), 1.15 (s, 3H), 1.09 (s, 3H). LC/MS: rt (5-
100-7 method) = 4.701 min.; 448.6 (M, 100 %), 450.6 (M+2, 35%).
Example 142
2-((2-amino-4-chloro-5-(2-(methyl(1-methylpiperidin-4-yl)amino)-2-oxoethyl)-5H-pyrrolo[2,3- d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide (CL)
[00616] The title compound was obtained by reacting 2-((2-amino-5-(carboxymethyl)-4-chloro-5H- pyrrolo[2,3-d]pyrimidin-7(6H)-yl)methyl)-4-methoxy-3,5-dimethylpyridine 1 -oxide (30 mg, 0. 08 mmol) with trifluoro-acetic acid pentafluorophenyl ester (60 μL, 0.3 mmol), followed by N,1-dimethylpiperidin-4-amine (200 mg, 1 mmol), using the general amide formation procedure. (13 mg, purity >90%, yield 30%). 1H NMR (CDCI3) δ ppm 8.11 (s, 1 H), 5.40 (s, 2H), 4.88 (q, 2H), 4.74 (m, 1 H), 3.90 (t, 1H), 3.78 (s, 3H), 3.65 (m, 1 H), 3.60 (b, d, 2H), 3.45 (dd, 1 H), 3.0 (m, 2H), 2.85 (s, 3H), (2.80 m, 2H), 2.75 (s, 3H), 2.71 (t, 1 H), 2.47 9m, 1 H), 2.39 (s, 3H), 2.23 (s, 3H), 2.21 (m, 2H), 1.80(m, 2H). LC/MS: rt (5-100-7 method) = 3.889 min.; 503.6 (M, 100 %), 505.6 (M+2, 35%). Example 143
(R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetamide (CLI)
[00617] (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide was prepared in a manner similar to its (S) enantiomer but starting from (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetate. Example 144
(R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetamide (CLII)
[00618] A solution of (R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (131.0 mg, 0.3440 mmol) in Tetrahydrofuran (14.6 mL) and N,N-Diisopropylethylamine (440 μL, 2.5 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (300.4 μL, 1.743 mmol) at room temperature. After 5 minutes, 0.5 M ammonia in dioxane (0.3664 g, 3.485 mmol) was added. Upon completion, water was added and the mixture extracted with DCM and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (99 mg, yield 75%). LC-MS (5-100-5 method) : Rt = 1.748 min; M+1 = 380.1 ; 1H NMR (400 MHz, CDCI3) δ ppm 8.16 (s, 1 H), 5.34 - 5.64 (m, 2 H), 4.86 (s, 2 H), 4.64 (s, 2 H), 3.77 (s, 3 H), 3.73 (d, J= 10.1 Hz, 1 H), 3.62 - 3.71 (m, 1 H), 3.35 (dd, J= 10.0, 3.9 Hz, 1 H), 2.94 (dd, J= 15.5, 3.2 Hz, 1 H), 2.38 (dd, J= 15.5, 10.2 Hz, 1 H), 2.22 (s, 3 H). Example 145
(R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-methylacetamide (CLIII)
[00619] A solution of (R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (45.0 mg, 0.118 mmol) in Tetrahydrofuran (5 mL) and N,N-Diisopropylethylamine (150 μL, 0.86 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (103.2 μL, 0.5986 mmol) at room temperature for 5 minutes. To this was added 2.0 M Methylamine in Tetrahydrofuran (0.5986 mL, 1.197 mmol). Upon completion, water was added and the mixture extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (31.3 mg, yield 66.6%). LC-MS (5-100-5 method) : Rt = 1.843 min; M+1 = 394.1 ; 1H NMR (400 MHz, CDCI3) δ ppm 8.16 (s, 1 H), 5.62 (br. s., 1 H), 4.89 (s, 2 H), 4.52 - 4.72 (m, 2 H), 3.76 (s, 3 H), 3.55 - 3.71 (m, 2 H), 3.33 (d, J= 6.1 Hz1 1 H), 2.80 - 2.92 (m, 1 H), 2.77 (d, J= 4.8 Hz, 3 H), 2.28 (dd, J= 15.0, 10.0 Hz, 1 H), 2.20 (s, 3 H). Example 146
(R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideυteromethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-ethylacetamide (CUV)
[00620] A solution of (R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (45.0 mg, 0.118 mmol) in Tetrahydrofuran (5 ml_) and N,N-Diisopropylethylamine (150 μl_, 0.86 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (103.2 μl_, 0.5986 mmol) at room temperature for 5 minutes. To this was added 2.0 M Ethylamine in Tetrahydrofuran (0.5986 ml_, 1.197 mmol). Upon completion, water was added and the mixture was extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid (35 mg, yield 72%). LC-MS (5-100-5 method) : Rt = 1.959 min; M+1 = 408.2; 1H NMR (400 MHz, CDCI3) δ ppm 8.17 (s, 1 H), 5.47 (br. s., 1 H), 4.80 (s, 2 H), 4.64 (dd, J= 27.0, 12.1 Hz, 2 H), 3.73 - 3.85 (m, 3 H), 3.57 - 3.73 (m, 2 H), 3.31 - 3.42 (m, 1 H), 3.17 - 3.30 (m, 2 H), 2.86 (dd, J= 15.3, 2.9 Hz, 1 H), 2.28 (dd, J= 14.9, 9.9 Hz, 1 H), 2.21 (s, 3 H), 1.10 (t, J= 7.2 Hz, 3 H) Example 147
(R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-(oxetan-3-yl)acetamide (CLV)
[00621] A solution of (R)-2-(2-amino-4-chloro-7-((4-methoxy-3-methyl-5-trideuteromethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (36.0 mg, 0.0945 mmol) in Tetrahydrofuran (4 mL) and N,N-Diisopropylethylamine (120 μL, 0.69 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (82.54 μL, 0.4789 mmol) at room temperature for 5 minutes. To this was added Oxetan-3-ylamine (70.01 mg, 0.9578 mmol). Upon completion, water was added and the mixture was extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (35 mg, yield 82%). LC-MS (5-100-5 method) : Rt = 1.869 min; M+1 = 436.1 ; 1H NMR (400 MHz, CDCI3) δ ppm 8.16 (s, 1 H), 6.19 (d, J= 7.3 Hz, 1 H), 4.95 - 5.08 (m, 1 H), 4.90 (q, J= 7.2 Hz, 2 H), 4.84 (br. s., 2 H), 4.65 (s, 2 H), 4.50 (t, J= 6.4 Hz, 1 H), 4.42 (t, J= 6.4 Hz, 1 H), 3.79 (s, 3 H), 3.61 - 3.71 (m, 2 H), 3.32 (d, J= 6.1 Hz, 1 H), 2.90 (dd, J= 15.2, 3.0 Hz, 1 H), 2.38 (dd, J= 15.3, 9.7 Hz, 1 H), 2.23 (s, 3 H). Example 148
(R)-2^2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-N-methylacetamide (CL Vl)
[00622] A solution of (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (120.0 mg, 0.3143 mmol) in Tetrahydrofuran (13 mL) and N,N-Diisopropylethylamine (400 μl_, 2.3 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (274.4 μl_, 1.592 mmol) at room temperature for 5 minutes. To this was added 2.0 M Methylamine in Tetrahydrofuran (1.592 ml_, 3.185 mmol). Upon completion, water was added and the mixture was extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (85 mg, yield 69%). LC-MS (5-100-5 method) : Rt = 1.771 min; M+1 = 389.1 ; 1H NMR (400 MHz, CDCI3) δ ppm 8.11 (s, 1 H), 5.55 - 5.76 (m, 1 H), 4.90 (s, 2 H), 4.54 - 4.76 (m, 2 H), 3.66 (dd, J= 7.8, 2.3 Hz, 2 H), 3.30 (d, J= 6.1 Hz, 1 H), 2.82 - 2.95 (m, 1 H), 2.76 (d, J= 4.8 Hz, 3 H), 2.66 (q, J= 7.6 Hz, 2 H), 2.27 (s, 3 H), 2.23 (s, 3 H), 2.30 (s, 1 H), 1.10 (t, J= 7.6 Hz, 3 H). Example 149
(R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-N-ethylacetamide (CL VII)
[00623] A solution of (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-N-ethylacetamide (120.0 mg, 0.3143 mmol) in Tetrahydrofuran (25 mL) and N,N-Diisopropylethylamine (0.40 ml_, 2.3 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (274.4 μl_, 1.592 mmol) at room temperature for 5 minutes. To this was added 2.0 M Ethylamine in Tetrahydrofuran (1.592 mL, 3.185 mmol). Upon completion, water was added and the mixture was extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (105 mg, yield 80%). LC-MS (5-100-5 method) : Rt = 2.075 min; M+1 = 403.2; 1H NMR (400 MHz, CDCI3) δ ppm 8.10 (s, 1 H), 5.57 (br. s., 1 H), 4.89 (br. s., 2 H), 4.56 - 4.77 (m, 2 H), 3.59 - 3.72 (m, 2 H), 3.31 (d, J= 5.8 Hz, 1 H), 3.19 - 3.28 (m, 2 H), 2.86 (dd, J= 15.0, 2.9 Hz, 1 H), 2.68 (q, J= 7.6 Hz, 2 H), 2.29 (s, 3 H), 2.25 (s, 3 H), 2.23 - 2.32 (m, 1 H), 1.12 (t, J= 7.6 Hz, 3 H), 1.09 (t, J= 7.3 Hz, 3 H). Example 150
(Rj-Σ-fΣ-amino^-chloro-T-tftethyl-S^-dimethylpyridin-Σ-yOmethylJ-βJ-dihydro-SH-pyrrolofcS- d]pyrimidin-5-yl)-1-(3-(hydroxymethyl)azetidin-1-yl)ethanone (CLVIII)
[00624] In a flame-dried flask under nitrogen was stirred (R)-2-(2-amino-4-chloro-7-((4-ethyl-3,5- dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-1-(3-(hydroxymethyl)azetidin-1- yl)ethanone (50.0 mg, 0.131 mmol) and N,N-Diisopropylethylamine (0.17 mL, 0.96 mmol) in Tetrahydrofuran (10 mL).
Trifluoroacetic acid pentafluorophenyl ester (80.0 μL, 0.464 mmol) was added and the mixture was stirred for 5 minutes followed by addition of 3-(hydroxymethyl)azetidine (115.6 mg, 1.327 mmol). Upon completion, water was added and the mixture was extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (22 mg, yield 37.4%). LC-MS (5-100-5 method) : Rt = 1.937 min; M+1 = 445.2; 1H NMR (400 MHz, CDCI3) δ ppm 8.12 (s, 1 H), 4.79 (s, 2 H), 4.53 - 4.76 (m, 2 H), 4.08 - 4.21 (m, 1 H), 4.03 (dt, J= 13.3, 9.2 Hz, 1 H), 3.89 (dt, J= 9.0, 4.7 Hz, 1 H), 3.71 - 3.84 (m, 3 H), 3.56 - 3.70 (m, 2 H), 3.19 - 3.31 (m, 1 H), 2.71 - 2.86 (m, 2 H), 2.61 - 2.71 (m, 2 H), 2.28 (s, 3 H)1 2.24 (d, J= 2.0 Hz, 3 H), 2.12 - 2.22 (m, 1 H), 1.11 (t, J= 7.6 Hz, 3 H). Example 151
2-((R)-2-amino-4-chloro-7-((4^thyl-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)-1-((S)-3-(hydroxymethyl)pyrrolidin-1-yl)ethanone (CLIX) [00625] A solution of 2-((R)-2-amino-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)-1-((S)-3-(hydroxymethyl)pyrrolidin-1-yl)ethanone (50.0 mg, 0.131 mmol) in Tetrahydrofuran (10 mL) and N,N-Diisopropylethylamine (0.17 ml_, 0.96 mmol) was treated with Trifluoroacetic acid pentafluorophenyl ester (28.21 μl_, 0.1637 mmol) at room temperature for 5 minutes. To this was added (S)-pyrrolidin-3-ylmethanol (134.2 mg, 1.327 mmol). Upon completion, water was added and the mixture was extracted 3 times with dichloromethane and dried over MgSO4. The residue was purified by silica gel chromatography using dichloromethane/methanol to give the title product as a white solid. (38 mg, yield 62%). LC-MS (5-100-5 method) : Rt = 1.945 min; M+1 = 459.2; 1H NMR (400 MHz, CDCI3) δ ppm 8.11 (d, J= 5.3 Hz, 1 H), 4.71 - 4.99 (m, 3 H), 4.40 - 4.63 (m, 1 H), 3.66 - 3.74 (m, 2 H), 3.57 - 3.66 (m, 1 H), 3.47 - 3.56 (m, 2 H), 3.33 - 3.47 (m, 2 H), 3.10 - 3.29 (m, 1 H), 2.91 - 3.07 (m, 1 H), 2.66 (qd, J= 7.5, 1.9 Hz, 2 H), 2.47 - 2.57 (m, 1 H), 2.30 - 2.46 (m, 1 H), 2.28 (d, J= 3.3 Hz, 3 H), 2.19 - 2.25 (m, 3 H), 2.09 (dd, J= 12.1 , 5.6 Hz, 1 H), 2.00 (dd, J= 14.4, 6.3 Hz, 1 H), 1.76 - 1.88 (m, 1 H), 1.67 (dd, J= 12.9, 8.1 Hz, 1 H), 1.05 - 1.16 (m, 3 H). Example 152
1-(2-((S)-2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetyl)piperidine-3-carboxamide (CLX)
[00626] The title compound (40 mg, 31%, 1 :1 mixture of 2 diastereomers) was made from (S)-2-
(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin- 5-yl)acetic acid and piperidine-3-carboxamide using the general amide formation procedure. 1H NMR (500 MHz, DMSO-d6): 5 8.14 (s, 1 H), 7.35-7.31 (m, 1 H), 6.89-6.80 (m, 1 H), 6.44 (br s, 2H), 4.65-4.52 (m, 2H), 4.32 (d, J = 13Hz, 1 H), 4.09-4.03 (m, 1 H), 3.75 (s, 3H), 3.68-3.63 (m, 1 H), 3.50-3.40 (m, 2H), 3.17-3.05 (m, 2H), 2.95-2.89 (m, 1 H), 2.82-2.79 (m, 1H), 2.70-2.63 (m, 1 H), 2.57-2.54 (m, 1 H), 2.17 (s, 3H), 2.13 (s, 3H), 1.80-1.69 (m, 1 H), 1.65-1.52 (m, 2H). LC-MS (10-90[4]-10 method): Rt = 2.63 min. (96%); m/z = 488 (M+1).
Example 153
1-(2-((Ry2-amino^-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetyl)piperidine-3-carboxamide (CLXI)
To a stirred solution of (R)-2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (0.225 g, 0.66 mmol) in DMF (5 mL) at RT was added DCC (0.165 g, 0.801 mmol), followed by 2,3,4,5,6-pentafluorophenol (0.147 mg, 0.801 mmol) and continued stirring for 3 h at RT. After the completion of starting material (by TLC), piperidine-3- carboxamide (85 mg, 0.66 mmol) was added and the stirring was continued for another 2 h at RT. After consumption of the starting material (by TLC), the reaction mixture was quenched with water (20 ml_) and extracted with EtOAc (3 X 10 ml_). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure to obtain crude material, this was purified by silica gel column chromatography eluting with 7% MeOH/dichloromethane, to afford the title compound (0.15 g, 52%). 1H NMR (500 MHz, DMSO-cfe): δ 8.14 (s, 1 H), 7.35-7.31 (m, 1 H), 6.89-6.80 (m, 1H), 6.39 (br s, 2H)1 4.59- 4.52 (m, 2H), 4.32 (d, J = 13Hz, 1 H), 4.08-4.04 (m, 1 H), 3.76-3.70 (m, 1 H), 3.71 (s, 3H), 3.65 (t, J = 10 Hz, 1 H), 3.48-3.44 (m, 1 H), 3.09-3.03 (m, 2H), 2.94-2.89 (m, 1 H), 2.81 -2.78 (m, 1 H), 2.67-2.64 (m, 1 H), 2.53-2.49 (m, 1 H), 2.18 (s, 3H), 2.16 (s, 3H), 1.84-1.80 (m, 1 H)1 1.65-1.52 (m, 2H). LC-MS (10-90[4]-10 method): Rt = 1.30 min. (96%); m/z = 488.2 (M+1). Example 154
5-{[Bis^2-methoxy-ethyl)-amino]-methyl}-4-chloro-7-(4-methoxy-3)5-dimethyl-pyridin-2-ylmethyl)- 6, 7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (CLXII)
[00627] A mixture of 5-aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (20.0 mg, 0.0573 mmol) and potassium carbonate (23.8 mg, 0.172 mmol) in N.N-dimethylformamide (1.00 mL) was treated with 1-bromo-2-methoxyethane (16.2 μL, 0.172 mmol) at room temperature overnight. The mixture was diluted with water (10.0 mL), adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5 to 60% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (3.10 mg, purity 80.0%, yield 9.30%). 1H NMR (CDCI3): δ ppm 8.20 (s, 1 H), 4.88 (br. s, 2H), 4.78 (d, 1 H), 4,53 (d, 1 H), 3.81 (m, 1 H), 3.80 (s, 3H), 3.65-3.43 (m, 7H), 3.39 (s, 6H), 2.99 (m, 1 H), 2.85 (m, 2H), 2.76 (m, 2H), 2.26 (s, 3H), 2.24 (s, 3H). LC-MS (5-100-7 method): Rt=4.092 min; M+1 =464.6 Example 155
2-[[2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-ylmethyl]-(2-hydroxy-ethyl)-amino]-ethanol (CLXIII)
[00628] A mixture of 5-aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (20.0 mg, 0.0573 mmol) and potassium carbonate (23.8 mg, 0.172 mmol) in N,N-dimethylformamide (1.00 mL) was treated with 2-bromoethanol (12.2 μL, 0.172 mmol) at room temperature overnight. The mixture was diluted with water (10.0 mL), adjusted to pH 2.0, and purified by reverse-phase preparative HPLC (5 to 60% CH3CN/H2O, 0.1% TFA) to give the title compound as a white solid (2.80 mg, purity > 90%, yield 10.0%). 1H NMR (CDCI3): δ ppm 8.19 (s, 1 H), 4.90 (br. s, 2H), 4.85 (m, 3H), 4.38 (d, 1 H), 3.81 (m, 1 H), 3.80 (s, 3H), 3.59 (m, 4H), 3.44-3.20 (m, 3H), 2.90-2.62 (m, 5H), 2.28 (s, 3H), 2.25 (s, 3H). LC-MS (5-100-7 method):Rt=3.633 min; M+1 =436.6 Example 156
(R)-5-(aminomethyl)-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-amine (CLXIV) and (S)-5-(aminomethyl)-4-chloro-7-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H-pyrrolo[2, 3-d]pyrimidin-2-amine (CLXV) [00629] A racemic mixture of 5-aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2- ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (1.65g) was separated by chiral HPLC (Chiralpak iC, 40% IPA (0.1%DEA)/CO2> 100bar, 80mL/min, 220 nM) to provide the fl-enantiomer (632 mg, purity>99%, ee>99%) and S-enantiomer (654mg, purity>99%, ee>99%). Example 157 i-tfΣ-amino^-chloro-T-W-methoxy-Sβ-dimethylpyridin-Σ-yOmethyO-βJ-dihydro-SH-pyrrolofcS- d]pyrimidin-5-yl)methyl)-2-cyanoguanidine (CLXVI)
[00630] A solution of 5-Aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (30 mg, 0.09 mmol) and diphenyl cyanocarbonimidate (60 mg, 0.25 mmol) in anhydrous THF (5 ml_) was stirred at rt for 30 minutes. The reaction was monitored via LC/MS. Ammonia (400 mg, 20 mmol, 7N in MeOH) was added and the mixture was stirred at 60 C for 3 hours. The white solid product was purified by preparative TLC. (15 mg, purity 90%, yield 40%). 1H NMR (CDCl3/CD3OD): δ ppm 8.11 (s, 1 H), 6.8 (s, 1 H), 5.9 (s, 2H), 5.19 (s, 2H), 4.87(d, 1H), 4.34 (d, 1 H), 3.78 (s, 3H), 2.25 (s, 3H), 2.22 (s, 3H), LC/MS:. rt (5-100-7 method) = 4.111 min.; 415.6(M+1 , 100 %), 417.6(M+3, 35%). Example 158
4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5-((methylamino)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-amine (CLXVII)
[00631] A mixture of 5-Aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (50 mg, 0.1 mmol), methyl iodide (25 mg, 0.18 mmol) and K2CO3 (101 mg, 0.73 mmo) in anhydrous DMF (5mL) was stirred for 2 h at rt. The white solid product was purified by reverse phase HPLC and preparative TLC (5 mg, purity 90%, yield 9 %). 1H NMR (CDCI3ZCD3OD): δ ppm 8.19 (s, 1 H), 4.82 (s, 2H), 4.58 (d, 1 H), 3.78 (s, 3H), 3.58 (t, 1 H), 3.38 (m, 2H), 2.93 (m, 1 H), 2.72 (dd, 1H), 2.43 (s, 3H), 2.26 (s, 3H), 2.23 (s, 3H), LC/MS:. rt (5-100-7 method) = 3.976 min.; 362.6(M+1 , 100 %), 364.6(M+3, 35%). Example 159
[(R)-2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-ylmethyl]-υrea (CLXVIII)
[00632] (f?)-5-Aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (30.0 mg, 0.0860 mmol) in tetrahydrofuran (2.0 mL) was stirred under a nitrogen balloon. The trimethylsilyl isocyanate (14.0 μM, 0.103 mmol) was then added. After 1 hour the reaction was diluted with ethyl acetate, washed with sat. NaHCO3, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by preparative HPLC (mobile phase: acetonitrile/water with 0.1% TFA buffer, gradient of 5-95%). The HPLC fractions containing the desired compound were combined, made basic with sat. aq. NaHCO3, and extracted 2x with ethyl acetate. The organics were then washed with brine, dried Na2SO4, filtered and condensed to yield 11 mg (32% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.22 (s, 1 H), 6.56 (br. s., 1H), 5.02 (br. s., 2H), 4.93 (d, J= 16.2 Hz, 1H), 4.64 (s, 2H), 4.29 (d, J= 16.2 Hz, 1H), 3.77 (s, 3H), 3.69 (dt, J= 13.3, 4.3 Hz, 1H), 3.56 - 3.64 (m, 1 H), 3.48 - 3.56 (m, 2H), 3.39 - 3.47 (m, 1 H), 2.24 (s, 3H), 2.22 (s, 3H). LC-MS (5-100-5 method): Rt = 1.78 (99.1% HPLC purity) m/z = 392.0 (M+1). Example 160
[(S)-2-Amino-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-ylmethyl]-urea (CLXIX)
[00633] (S)-5-Aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (70.0 mg, 0.201 mmol) in tetrahydrofuran (4.7 mL) was stirred under a nitrogen balloon. The trimethylsilyl isocyanate (32.6 μM, 0.241 mmol) was then added. After 2 hours the reaction was diluted with ethyl acetate, washed with sat. aq. NaHCO3, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by preparative HPLC (mobile phase: acetonitrile/water with 0.1% TFA buffer, gradient of 5-95%). The HPLC fractions containing the desired compound were combined, made basic with sat. aq. NaHCO3), and extracted 1x with ethyl acetate and 2x with 10% MeOH/dichloromethane. The combined organics were then washed with brine, dried over Na2SO4, filtered and condensed to yield 36 mg (45% yield) of the title compound as a white solid. 1H NMR (400 MHz, CDCI3) δ ppm 8.22 (s, 1 H), 6.48 (br. s., 1 H), 4.85 - 5.06 (m, 3H), 4.52 (br. s., 2H), 4.27 (d, J= 16.4 Hz, 1 H), 3.67 - 3.82 (m, 4H), 3.57 - 3.67 (m, 1 H), 3.40 - 3.55 (m, 3H), 2.24 (s, 3H), 2.22 (s, 3H). LC-MS (5-100-5 method): Rt = 1.78 (99.9% HPLC purity) m/z = 392.1 (M+1). Example 161
N-[(S)-2-Amino^-chloro-7^4-methoxy-3,5-dimethylψyridin-2-ylmethyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-ylmethyl]-acetamide (CLXX)
[00634] (S)-5-Aminomethyl-4-chloro-7-(4-methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-2-ylamine (58 mg, 0.17 mmol) in tetrahydrofuran (3.9 mL) was stirred under a nitrogen balloon and the triethylamine (46.3 μM, 0.332 mmol) was added. The acetyl chloride (14.2 μM, 0.200 mmol) was then added. After 20 hours the reaction was diluted with ethyl acetate, washed with sat. aq. NaHCO3, washed with brine, dried over Na2SO4, filtered and condensed. The crude material was then purified by flash chromatography (0-10% MeOH/ dichloromethane gradient) to provide 37 mg (54% yield) of the title compound. 1H NMR (400 MHz, CDCI3) δ ppm 8.17 (s, 1 H), 6.93 (br. s., 1 H), 4.89 - 5.08 (m, 3H), 4.31 (d, J= 15.9 Hz, 1 H), 3.71 - 3.88 (m, 4H), 3.59 - 3.70 (m, 1 H), 3.37 - 3.52 (m, 3H), 2.26 (s, 3H), 2.24 (s, 3H), 1.93 (s, 3H). LC-MS (5-100-7 method): Rt = 3.97 min. (94.2% HPLC purity); m/z = 390.6 (M+1).
Example 162
(S)-5-((1,3,4-oxadiazol-2-yl)methyl)-4-chloro-7-((4-ethyl-3,S-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2, 3-d]pyrimidin-2-amine (CLXXI)
[00635] In a flame-dried flask under nitrogen (S)-2-(2-amino-4-chloro-7-((4-ethyl-3,5- dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetohydrazide (20.0 mg, 0.0513 mmol) was stirred in Ethyl orthoformate (2.50 mL, 15.0 mmol). Concentrated Sulfuric acid (1.00 μL, 0.0188 mmol) was added and the mixture was heated in a microwave at 80 0C for 15 minutes. The crude mixture was purified by reverse-phase HPLC (2% to 30% CH3CN/H2O, 0.1% TFA) to give the product as a white solid (1.7 mg, yield 8.2%); LC-MS (5-100-5 method) : Rt = 2.010 min; M+1 = 400.0; 1H NMR (400 MHz, CDCI3) δ ppm 8.32 (s, 1 H), 8.12 (s, 1 H), 4.94 (s, 2 H), 4.51 - 4.76 (m, 2 H), 3.63 - 3.85 (m, 2 H),
3.30 - 3.52 (m, 2 H), 3.10 (dd, J= 15.7, 9.1 Hz, 1 H), 2.67 (q, J= 7.6 Hz, 2 H)1 2.27 (s, 3 H), 2.23 (s, 3 H), 1.12 (t, J= 7.6 Hz, 3 H)
Example 163
(R)-5-((1,3,4-oxadiazol-2-yl)methyl)-4-chloro-7-((4-ethyl-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (CLXXII)
[00636] In a flame-dried flask under nitrogen (R)-2-(2-amino-4-chloro-7-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetohydrazide (47.0 mg, 0.120 mmol) was stirred in Ethyl orthoformate (4.50 mL, 27.0 mmol). Concentrated Sulfuric acid (2.34 μL, 0.0439 mmol) was added and the mixture was heated in a microwave at 80 0C for 15 minutes. The crude mixture was purified by HPLC (2% to 30% CH3CN/H2O, 0.1% TFA) to give the product as a white solid. (1.11 mg, yield 2.3%). LC-MS (5-100-5 method) : Rt = 1.900 min; M+1 = 402.0; 1H NMR (400 MHz, CDCI3) δ ppm 8.32 (s, 1 H), 8.16 (s, 1 H), 4.88 (s, 2 H), 4.50 - 4.74 (m, 2 H), 3.78 (s, 3 H), 3.72 - 3.83 (m, 2 H), 3.35 - 3.51 (m, 2 H), 3.05 - 3.24 (m, 1 H), 2.25 (s, 3 H), 2.21 (s, 3 H). Example 164
4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5-(methoxymethyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-2-amine (CLXXIII)
[00637] To a stirred solution of (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methanol (20 mg, 0.057 mmol) in THF (1 mL) was added NaH (3 mg, 0. 11 mmol) and MeI (12 mg, 0. 11 mmol) at 0 0C under anhydrous conditions. The reaction mixture was stirred at RT for 12h. The volatiles were evaporated under reduced pressure and the crude material was purified by column chromatography [60-120 silica gel; 10g, 1 -2% MeOH/dichloromethane] to afford the title compound (10 mg, 50%) as an off-white solid. 1H NMR (500 MHz, DMSO-cfe): δ 8.15 (s, 1H), 6.41 (s, 2H), 4.63 (d, J = 15 Hz, 1 H), 4.52 (d, J = 15 Hz, 1 H), 3.68 (s, 3H), 3.53- 3.47 (m, 2H), 3.33-
3.31 (m, 2H), 3.19- 3.17 (m, 1H), 3.14 (s, 3H), 2.15 (s, 3H), 2.11 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 3.05 min. (91%); m/z = 364 (M+1).
Example 165
4-chloro-7^(4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5-((2-methoxyethylamino)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amine (CLXXIV)
[00638] To a stirred solution of (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methyl methanesulfonate (300 mg, 0.7 mmol) in DMF (30 mL) was added K2CO3 (242 mg, 1.75 mmol) followed by 2-methoxy-ethylamine (57 mg, 0.77 mmol) at 0 0C under an inert atmosphere. The reaction mixture was heated at 60 °C for 12 h. The reaction mixture was cooled to RT, diluted with water (50 mL) and extracted with EtOAc (2 x 50 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude material was purified by preparative TLC to afford the title compound (56 mg, 19.7%). 1HNMR: (500 MHz, CDCI3): δ 8.20 (S, 1H), 4.81 (s, 2H), 4.72 (d, J = 14.5 Hz, 1 H), 4.58 (d, J = 10.5 Hz, 1 H), 3.75 (s, 3H), 3.61 (t, J = 9.5 Hz, 1H), 3.45-3.43 (m, 3H), 3.38-3.33 (m, 1 H), 3.26 (s, 3H), 3.02- 2.99 (m, 1H), 2.81 - 2.71 (m, 3H), 2.26 (s, 3H), 2.21 (S, 3H). LC-MS (10-90[4]-10 method): Rt = 2.72 min. (90%); m/z = 407 (M+1). Example 166
4-chloro-7^(4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5-((oxetan-3-ylamino)methyl)-6,7-dihydro- 5H-pyrrolo[2,3-d]pyrimidin-2-amine (CLXXV)
[00639] To a stirred solution of (2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)methyl methanesulfonate (0.3 g, 0.70 mmol) in dry DMF (15 mL) was added K2CO3 (0.241 g, 1.75 mmol) followed by oxetan-3-amine (56 mg, 0.77 mmol). The reaction was stirred at 60° C for 12 h. After consumption of the starting material (by TLC), the reaction was cooled to RT, quenched with water (15 mL) and the aqueous layer was extracted with EtOAc (2 x 30 mL). The combined organic extracts was washed with water (25 mL), dried over Na2SO4 and concentrated under reduced pressure to obtain the crude material, which was purified by silica gel column chromatography eluting with 10% MeOH/dichloromethane, to afford 45% HPLC pure material. This was subjected to preparative HPLC purification to afford the title compound (4 mg, 2%). 1H NMR (500 MHz, CDCI3): δ 8.19 (s, 1H), 4.81 -4.72 (m, 4H), 4.54 (d, J = 15 Hz, 1 H), 4.36-4.34 (m, 2H), 3.94-3.89 (m, 1 H), 3.76 (s, 3H), 3.61 (t, J = 10 Hz, 1 H), 3.46-3.45 (m, 1 H), 3.34-3.30 (m, 1 H), 2.84-2.76 (m, 2H), 2.25 (s, 3H), 2.22 (s, 3H). LC-MS (10-90[4]-10 method): Rt = 7.63 min. (94%); m/z = 405.7 (M+1 ). Example 167
3-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)propanamide (CLXXVI)
Step 1. 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)ethanol
[00640] To a stirred solution of methyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrirnidin-5-yl)acetate (0.9 g, 2.30 mmol) in dry THF (10 mL) was added LAH (0.4 g, 11.47 mmol) portion wise at 0 °C and stirred under reflux for 7 h. After consumption of the starting material (by TLC), the reaction mixture was cooled to 0 0C, quenched with EtOAc (50 mL) and filtered through silica gel bed, the bed was washed several times with EtOAc (2 x 35 mL). The combined filtrate was concentrated under reduced pressure to afford the title compound (0.58 g, 69%). This was used in the next step without any further purification. 1H-NMR (500 MHz, DMSO-O6): δ 8.14 (s, 1H), 6.35 (br s, 2H, Exc), 4.61 -4.47 (m, 3H)1 3.71 (s, 3H), 3.57 (t, J= 9.5 Hz, 1 H), 3.41-3.38 (m, 2H), 3.25-3.19 (m, 2H), 2.18 (s, 3H), 2.16 (s, 3H), 1.98-1.94 (m, 1 H), 1.54-1.50 (m, 1 H). Mass: m/z = 364 (M+1 ). Step 2. 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)ethyl methanesulfonate
[00641] To a stirred solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)ethanol (0.58 g, 1.59 mmol) in CH2CI2 (10 mL) was added Et3N (0.46 ml_, 3.19 mmol) followed by Ms-Cl (0.19 ml_, 2.34 mmol) drop wise at 0 0C under inert atmosphere and stirred at RT for 2 h. After the consumption of starting material (by TLC), the reaction mixture was quenched with water (5 mL) and extracted with CH2CI2 (2 x 25 mL). The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure. The obtained crude material was purified by silica gel column chromatography eluting with 3% MeOH/CH2CI2 to afford the title compound (0.5 g, 70%). 1H-NMR (500 MHz, DMSO-d6): δ 8.15 (s, 1 H), 6.42 (br s, 2H, Exc), 4.62- 4.53 (m, 2H), 4.26-4.22 (m, 2H), 3.71 (s, 3H), 3.64 (t, J = 9 Hz, 1 H), 3.30-3.27 (m, 2H), 3.17(s, 1 H), 3.13 (s, 3H), 2.18 (s, 3H), 2.17 (s, 3H), 1.90-1.84 (m, 1 H). Mass: m/z = 442.0 (M+1). Step 3. 3-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)propanenitrile
[00642] To a stirred solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl methanesulfonate (0.58 g, 1.59 mmol) in DMSO (4 mL) under inert atmosphere was added KCN (0.4 g, 0.68 mmol) at RT and stirred at 70 0C for 5 h. After the consumption of starting material (by TLC), the reaction mixture was quenched with saturated NaHCO3 solution (5 mL) and extracted with EtOAc (2 x 25 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The obtained crude material was purified by silica gel column chromatography eluting with 3% MeOH/CH2CI2 to afford the title compound (0.1 g, 59%). 1H-NMR (500 MHz, DMSO-O6): δ 8.14 (s, 1 H), 6.43 (br s, 2H, Exc), 4.62-4.51 (m, 2H), 3.71 (s, 3H), 3.63 (t, J = 9.5 Hz, 1 H), 3.30-3.23 (m, 2H), 2.57-2.44 (m, 2H), 2.18 (s, 3H), 2.16 (s, 3H), 2.04-1.94 (m, 1 H), 1.82-1.78 (m, 1 H). Mass: m/z = 373.1 (M+1).
Step 4. 3-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)propanamide
[00643] To a stirred solution of 3-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)propanenitrile (0.1 g, 0.30 mmol) in DMSO (4 mL) was added 1 N NaOH (0.5 mL, 405 mmol) followed by H2O2 (0.7 mL, 6.3 mmol) and stirred at RT for 6 h. After the consumption of starting material (by TLC), the reaction mixture was diluted with water (5 mL) and extracted with EtOAc (2 x 25 mL). The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure. The obtained crude material was purified by Preparative TLC by eluting with 7% MeOH/CH2CI2 to afford the title compound (28 mg, 28% yield). 1H-NMR (500 MHz, DMSO-O6): δ 8.15 (s, 1 H), 7.24 (br s, 1 H, Exc), 6.68 (br s, 1 H, Exc), 6.37 (br s, 2H, Exc), 4.61-4.51 (m, 2H), 3.71 (s, 3H), 3.58 (t, J= 9.5 Hz, 1H), 3.19-3.12 (m, 2H), 2.18 (s, 3H), 2.16 (s, 3H), 2.01-1.96 (m, 3H), 1.66-1.63 (m, 1 H). Mass: m/z = 391.1 (M++1 ). HPLC (10-90[4]-10 method): Rt = 1.19 min. (99%). Example 168
2-(4-Methoxy-3,5-dimethyl-pyridin-2-ylmethyl)-1,2,7,8,9,9a-hexahydro-6-thia-2,3,5-triaza- benz[cd]azulen-4-ylamine (CLXXVII)
Step 1. S-2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)ethyl ethanethioate [00644] To a stirred solution of 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl methanesulfonate (0.2 g, 0.45 mmol) in DMF (5 mL) was added KSAc (0.1 g, 0.90 mmol) and stirred at RT for 3 h. After consumption of the starting material (by TLC), the reaction mixture was quenched with water (5 mL) and extracted with EtOAc (2 x 50 mL). The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure. The obtained crude material was purified by preparative TLC eluting with 70% EtOAc/Hexane to afford the title compound (0.1 g, 34% yield). 1H-NMR (500 MHz, DMSO-de) δ 8.14 (s, 1 H), 6.40 (br s, 2H, Exc), 4.63 (d, J = 16 Hz, 1 H), 4.50 (d, J = 15.5 Hz, 1 H), 3.71 (s, 3H) 3.62 (t, J = 10 Hz, 1 H), 3.30-3.21 (m, 2H), 2.84-2.78 (m, 1 H), 2.77-2.73 (m, 1 H), 2.29 (s, 3H) 2.18 (s, 3H), 2.16 (s, 3H), 1.91-1.89 (m, 1H), 1.74-1.71 (m, 1 H). Mass: m/z = 422.1 (M+1 ).
Step 2. 2-(4-Methoxy-3, 5-dimethyl-pyridin-2-ylmethyl)- 1,2, 7, 8, 9, 9a-hexahydro-6-thia-2,3, 5-triaza- benz[cd]azulen-4-ylamine
[00645] To a stirred solution of S-2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl ethanethioate (70 mg, 0.16 mmol) in EtOH (15 mL) was added NaBH4 (94 mg, 0.25 mmol) portion wise at 0 0C and stirred at 100 0C for 2 h. After consumption of the starting material (by TLC), the volatiles were evaporated under reduced pressure and the residue was extracted with EtOAc (2 x 50 mL). The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure. The obtained crude material was purified by silica gel column chromatography eluting with 4% MeOH/CH2CI2 to afford the title compound (24 mg, 40.6% yield). 1H-NMR (500 MHz, DMSO-d6) δ 8.15 (s, 1 H), 6.05 (br s, 2H, Exc), 4.80 (d, J = 15 Hz, 1 H), 4.07 (d, J = 15 Hz, 1 H), 3.72 (s, 3H), 3.53-3.55 (m, 1H), 3.14-3.10 (m, 2H), 3.08-3.04 (m, 3H), 2.30-2.28 (m, 2H), 2.19 (s, 6H), 1.35-1.33 (m, 1 H). LCMS (10-90[4]-10 method): Rt = 2.73 min. (99%), m/z = 344 (M+1).
Example 169
2-(2-amino-4-bromo-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3- d]pyrimidin-5-yl)acetamide (CLXXVIII)
Step 1: 2-(2-amino-4-bromo-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3'd]pyrimidin-5-yl)acetic acid
[00646] A stirred solution of methyl 2-(2-amino-4-chloro-7-((4-methoxy-3,5-dimethylpyridin-2- yl)methyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetate (30 mg, 0.076 mmol) in 36% HBr/acetic acid (0.3 mL) was stirred at 60 0C for 16 h. After consumption of the starting material (by TLC), the reaction mixture was basified to pH ~ 10 using aqueous NaOH solution at 0 CC and stirred for 5 minutes. Then the reaction mixture was neutralized to pH - 7 using aqueous citric acid solutions and extracted with EtOAc (2 x 25 mL). The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure to afford the title compound (18 mg, 56%) as an off white solid. 1H-NMR (500 MHz, DMSO-de): δ 12.18 (br s, 1 H, Exc), 8.15 (s, 1 H), 6.43 (s, 2H, Exc), 4.56-4.54 (m, 2H), 3.72 (s, 3H), 3.17-3.16 (m, 2H), 2.83-2.79 (m, 1 H), 2.61-2.55 (m, 1 H), 2.40-2.35 (m, 1 H), 2.18 (s, 3H), 2.16 (s, 3H). Step 2. 2-(2-amino-4-bromo-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6, 7-dihydro-5H- pyrrolo[2,3-d]pyrimidin-5-yl)acetamide
[00647] A mixture of 2-(2-amino-4-bromo-7-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-6,7- dihydro-5H-pyrrolo[2,3-d]pyrimidin-5-yl)acetic acid (15 mg, 0.035 mmol), EDCI (10.2 mg, 0.053 mmol),
HOBt (7.0 mg, 0.053 mmol), DIPEA (13.7 mg, 0.106 mmol) and NH4CI (3.8 mg, 0.071 mmol) in dry DMF
(0.15 mL) was stirred at RT for 4 h. After the consumption of starting material (by TLC), the reaction mixture was diluted with water and extracted with EtOAc (2 x 20 mL). The combined organic extracts were dried over anhydrous Na2SO4 and concentrated under reduced pressure. The obtained crude material was purified by preparative TLC eluting with 10% MeOH/CH2CI2 to afford the title compound (8 mg, 57%). 1H-NMR (500 MHz, DMSO-d6) δ 8.15 (s, 1H), 7.35 (s, 1 H, Exc), 6.84 (s, 1 H, Exc), 6.41 (s, 2H,
Exc), 4.59-4.50 (m, 2H), 3.72 (s, 3H), 3.60 (t, J = 10 Hz, 1 H), 3.11-3.09 (m, 2H), 2.66 (s, 2H), 2.18 (s, 3H),
2.15 (s, 3H). LCMS (10-90[4]-10 method): Rt = 3.72 min. (94%), m/z = 421 (M+1 ).
Example 171
2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8-dihydropyrido[2,3- d]pyrimidin-6-yl)methanol and 2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
5,6,7,8-tetrahydropyrido[2,3-d]pyrimidin-6-yl)methanol (CLXXIX)
Step 1. Ethyl 3-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)propanoate
[00648] The reaction was performed as described in WO2009/139834. 4-methoxy-3,5-dimethyl- pyridine-2-carbaldehyde (2.2 g, 13 mmol), 3-amino-propionic acid ethyl ester HCI (2.5 g, 16 mmol) and
N,N-diisopropylethylamine (2.8 mL, 16 mmol) were suspended in 1 ,2-dichloroethane (50 mL). The reaction mixture was stirred at room temperature for 10 minutes before adding sodium triacetoxyborohydride (4.2 g, 20 mmol) was added. The reaction mixture was poured on ice-water and extracted with DCM (2 X 30 ml). The combined organic layers was washed with water (2 x 30 ml) and dried with MgSO4. Evaporation gave the crude product as an oil (2.1 g, purity 90.0%, yield 53%). 1H
NMR (CDCI3): 8.20 (s, 1H), 4.16 (q, 2H), 3.87 (s, 2H), 3.76 (s, 3H), 3.00 (t, 2H), 2.58 (t, 2H), 2.45 (b, m,
1 H), 2.24 (d, 6H), 1.27 (t, 3H). LC/MS: rt (5-100-5 method) = 2.12 min.; 267.1 (M+1 , 100 %), 268.1 (M+2,
16%).
Step 2. Ethyl 3-((2-amino-6-chloro-5-formylpyrimidin-4-yl)((4-methoxy-3, 5-dimethylpyridin-2- yl)methyl)amino)propanoate
[00649] Ethyl 3-((4-methoxy-3,5-dimethylpyridin-2-yl)methylamino)propanoate [(2.1 g, 7.9 mmol) and 2-amino-4,6-dichloropyrimidine-5-carbaldehyde (1.7 g, 8.8 mmol) were dissolved in a mixture of N1N-
Dimethylformamide (20 mL) and N,N-Diisopropylethylamine (1.6 mL, 9.2 mmol). The reaction was left stirring at RT for 1 hour and was monitored by LC/MS. The reaction was extracted with EtOAc (40 ml) and water (40 ml). The aqueous layer was again extracted with EtOAc (2 x 40 ml). The combined EtOAc layers were dried with MgSO4. The solvent was removed under reduced pressure to give a white solid residue. 1H NMR (CDCI3)) 10.06 (s, 1 H), 8.17 (s, 1 H), 5.30 (s, 2H), 4.77 (s, 2H), 4.10 (t, 2H), 3.76 (s, 3H),
3.71 (t, 2H), 2.71 (t, 2H), 2.25 (s, 3H), 2.10 (s, 3H), 1.22 (t, 3H). LC/MS : rt (5-100-5 method) = 2.382 min.; 422.0 (M+1 , 100 %), 424 (M+3, 36 %)
Step 3. Synthesis of Ethyl 2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8- dihydropyrido[2,3-d]pyrimidine-6-carboxylate, Methyl 2-amino-4-chloro-8-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-7,8-dihydropyrido[2,3-d]pyrimidine-6-carboxylate, and 2-amino-4- chloro-5-hydroxy-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8-dihydropyrido[2,3- d]pyrimidine-6-carboxylic acid
[00650] A suspension of ethyl 3-((2-amino-6-chloro-5-formylpyrimidin-4-yl)((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)amino)propanoate (1.00 g, 2.37 mmol) and cesium carbonate (1.00 g, 3.07 mmol) in N,N-Dimethylformamide (10 mL) was stirred for 72 hours at room temperature. A LC/MS sample indicated that the reaction progressed with less than 10% conversion and with many side products. Addition of sodium methoxide (0.25 g, 4.6 mmol) and heating at 50 °C for 2 hours brought the reaction to completion, as added to the reaction mixture. The reaction was extracted with EtOAc (40 ml) and water (40 ml). The aqueous layer was again extracted with EtOAc (2 x 40 ml). The 2-amino-4- chloro-5-hydroxy-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8-dihydropyrido[2,3-d]pyrimidine-6- carboxylic acid went into the aqueous layer, while the other two products went into the organic layer The solvents were removed and the products were purified via prep HPLC.
[00651 ] Ethyl 2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8- dihydropyrido[2,3-d]pyrimidine-6-carboxylate was a yellow solid (0.065 g; Yield = 6.1%; Purity = 90%). 1H
NMR (CDCI3) 8.22 (s, 1 H), 7.54 (s, 1 H), 4.99 (s, 2H), 4.84 (s, 2H), 4.41 (s, 2H), 4.23 (t, 2H), 3.78(s, 3H),
2.26 (s, 3H), 2.24 (s, 3H), 1.32 (t, 3H). LC/MS : rt (5-100-5 method) = 2.532min.; 404.0 (M+1 , 100 %),
406(M+3, 36 %).
[00652] Methyl 2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8- dihydropyrido[2,3-d]pyrimidine-6-carboxylate was a yellow solid (0.052 g; Yield = 5.1%; Purity = 90%;).
1H NMR (CDCI3) 8.21 (s, 1 H), 7.54 (s, 1 H), 4.93 (s, 2H), 4.84 (s, 2H), 4.42 (s, 2H), 3.78(s, 3H), 3.77 (s,
3H), 2.26 (s, 3H), 2.24 (s, 3H). LC/MS : rt (5-100-5 method) = 2.383min.; 390.0 (M+1 , 100 %), 392(M+3,
36 %)
[00653] 2-amino-4-chloro-5-hydroxy-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8- dihydropyrido[2,3-d]pyrimidine-6-carboxylic acid seemed to slowly decompose in water.
Step 4: Synthesis of 2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8- dihydropyrido[2,3-d]pyrimidin-6-yl)methanol and 2-amino-4-chloro-8-((4-methoxy-3,5- dimethylpyridin-2-yl)methyl)-5,6,7,8-tetrahydropyrido[2,3-d]pyrimidin-6-yl)methanol
[00654] A suspension of methyl 2-amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-
7,8-dihydropyrido[2,3-d]pyrimidine-6-carboxylate (30 mg, 0.08 mmol) in tetrahydrofuran (5 mL) was cooled at 0 0C and treated with L-Selectride (0.5 mL, 2 mmol) dropwise. The reaction was complete within 20 minutes. The reaction was quenched with water and extracted with EtOAc (2 x 20 mL). The combined organic layers were evaporated, and the products were purified via preparative HPLC.
[00655] 2-Amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-7,8-dihydropyrido[2,3- d]pyrimidin-6-yl)methanol was obtained as a white solid (10 mg, purity 90.0%, yield 30%). 1H NMR (CDCI3) 8.20 (s, 1 H), 6.48 (s, 1H), 4.8 (s, 2H), 4.77 (s, 2H), 4.3 (s, 2H), 4.06 (s, 2H), 3.78 (s, 3H), 2.26 (s, 3H), 2.25 (s, 3H). LC/MS : rt (5-100-5 method) = 1.955 min.; 362.0 (M+1 , 100 %), 364 (M+3, 37 %). [00656] 2-Amino-4-chloro-8-((4-methoxy-3,5-dimethylpyridin-2-yl)methyl)-5,6,7,8- tetrahydropyrido[2,3-d]pyrimidin-6-yl)methanol was obtained as a white solid (1.26 mg, purity 90.0%, yield 4%). 1H NMR (CDCI3) 8.15 (s, 1H), 5.78 (d, 1 H)1 4.51 (s, 2H), 3.98 (d, 1 H), 3.82 (m, 1H), 3.80 (s, 3H), 3.60 (m, 2H), 3.44 (m, 1 H), 2.80 (dd, 1 H), 2.55 (m, 1 H), 2.39 (m, 1 H), 2.36 (s, 3H), 2.25 (s, 3H). LC/MS : rt (5-100-5 method) = 1.911 min.; 364.0 (M+1 , 100 %), 366 (M+3, 37 %)
ACID STABILITY EXAMPLES Example A
Stability Towards Acids
[00657] The test article was dissolved in a 5M solution of methanesulfonic acid in DMSO/water
1 :1 (pH = -0.5), heated at 40 0C, and monitored by LC-MS.
[00658] Under these conditions, the chlorine atom of 6-chloroguanines of type A (see scheme below) was hydrolyzed with a half-life T1/2 = 1 h.
[00659] The chlorine atom of 4-chloro-7H-pyrrolo[2,3-d]pyrimidin-2-amines of type B had a half- life ranging from 5 to 130 hours, depending on the nature of the R group. In each case, the chlorine group was hydrolyzed.
[00660] In contrast, the chlorine atom of 4-chloro-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-2-amines of type C was substantially more stable, and the chlorine atom did not undergo hydrolysis, and remained stable with T1/2 > 1000 h.
Rate of chlorine hydrolysis in 5M MsOH solution (DMSO/water 1:1 ) at 40 0C
Figure imgf000217_0001
BIOLOGY EXAMPLES
Example A
Hsp90α, Hsp90β, Grp94, and TRAP-1 Competitive Binding Assays
Recombinant human Hsp90α (Stressgen Bioreagents, 0.8 nM) [or in-house expressed human Hsp90β (1.5 nM), TRAP1 (20 nM) (Hsp90 mitochondrial paralog) or Grp94 (3.1 nM) (Hsp90 endoplasmic reticulum paralog)] and reduced FITC-geldanamycin (Biomol International, 2 nM; reduced with TCEP for 3 h. at room temperature; Onuahoa, S. C. et al. J. MoI. Biol., 2007, 372:287-297) were incubated in a 96- well microplate at room temperature for 3 h. in assay buffer containing 20 mM Hepes, (pH 7.4), 50 mM KCI1 5 mM MgCI2, 20 mM Na2MoO4, 2 mM DTT, 0.1 mg/mL bovine gamma globulin, 0.1% (v/v) CHAPS. Following the pre-incubation, Compound A, a compound that falls within the scope of Formula I and exemplified in the working examples above, in 100% DMSO was added to final concentrations between 0.04 nM and 10 μM (final volume 100 μl, 2% DMSO). The reaction was incubated for 16 h. at room temperature with gentle shaking. Fluorescence was measured in an EnVision plate reader (Perkin Elmer) using 485 nm excitation and 535 nm emission wavelengths. Maximum signal and background controls contained no compound or no Hsp90 protein (or Hsp90 paralog), respectively. The data were fit to a 4- parameter curve using GraphPad Prism, and IC50 values were generated. Example B
Cell Lysate Binding Assay
[00661] MCF7 breast carcinoma cell lysates were prepared by douncing in lysing buffer (20 mM
HEPES, pH 7.3, 1 mM EDTA, 5 mM MgCI2, 100 mM KCI), and then incubated with or without test compound for 30 mins at 4° C, followed by incubation with biotin-GM linked to BioMag™ streptavidin magnetic beads (Qiagen) for 1 hr at 4° C. The tubes were placed on a magnetic rack, and the unbound supernatant removed. The magnetic beads were washed three times in lysis buffer and boiled for 5 mins at 95° C in SDS-PAGE sample buffer. Samples were analyzed on SDS protein gels, and Western blots were done for rHsp90. Bands in the Western Blots were quantitated using the Bio-rad Fluor-S Multilmager, and the % inhibition of binding of rHsp90 to the biotin-GM was calculated. Example C
HER2 Degradation Assay
[00662] MCF7 breast carcinoma cells (ATCC) were grown in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS). 10 mM HEPES, pH 7.3, and Antimycotic Antibiotic (100 units/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B; Invitrogen), and plated in 24 well plates (0.8 to 1.0 x 105 cells in 1.0 mL medium), and incubated at 37°C. Twenty-four hours later (cells were 40-50% confluent), test compounds that were serially diluted (1 :3) in 96 well plates in DMSO were added (2 μL per well). An additional 1.0 mL warm medium was added to each well and plates incubated for 16-18 hours) at 37°C. All compounds were tested at 10 concentrations ranging between 10 μM and 0.5 nM. The medium was aspirated from each well, wells were washed with 1 mL phosphate buffered saline (PBS), and 200 μL 0.25% trypsin was added to each well. After trypsinization was complete, 50 μL of FBS was added to each well. Cells (200 μL from each well) were transferred to 96 well plates and 100 μL cold PBS containing 0.2% BSA, 0.2% sodium azide (BA buffer) was added to each well. The plates were centrifuged at 1 ,200 rpm for 5 min at 4°C using a Beckman Coulter Allegra® 6R tabletop centrifuge (Beckman Coulter, Brea, CA). Wells were aspirated andcells were washed once in 300 μL BA buffer. Phycoerythrin (PE) conjugated anti-HER2/Neu antibody (BD Biosciences, #340552) was added at a dilution of 1 :20 (final concentration was 1 μg/mL) and the cells were pipeted up and down to form a single cell suspension, and incubated at 4°C for 30 mins. The plates were centrifuged and washed twice with 250 μl_ BA buffer, and each well resuspended in 100 μl_ BA buffer containing Propidium Iodide (Pl1 final concentration 10 μg/mL; Invitrogen #P3566). Cells were mixed well and samples analyzed in the 96 well plate using a BD FACSArray™ Bioanalyzer (BD Biosciences, San Jose, Calif.)- Pl positive dead cells were excluded, and 5,000 total events were collected per sample. Compensation was adjusted using unstained cells, Pl only treated cells, and PE-anti-HER2/Neu only treated cells. A fluorescence histogram was generated from the live cell gate and the mean fluorescence intensity (MFI) of each sample was determined using BD FACSArray software. MFI data were exported to Microsoft Excel and analyzed in GraphPad Prism® (GraphPad Software, Inc.) for calculation of EC50 values.
[00663] EC50 may be defined as the concentration resulting in a 50% decrease in HER2 MFI relative to 0.1% DMSO treated cells.
TABLE 2. Biological activities of selected compounds in the Hsp90α competitive binding assay
(Example A) and the HER2 degradation assay (Example C).
Figure imgf000219_0001
Figure imgf000220_0001
Figure imgf000221_0001
Figure imgf000222_0001
Figure imgf000223_0001
Figure imgf000224_0001
Figure imgf000225_0001
Figure imgf000226_0001
Figure imgf000227_0001
Figure imgf000228_0001
Figure imgf000229_0001
Figure imgf000230_0001
Figure imgf000231_0001
Figure imgf000232_0001
Figure imgf000233_0001
Figure imgf000234_0001
Figure imgf000235_0001
Figure imgf000236_0001
Figure imgf000237_0001
Figure imgf000238_0001
Figure imgf000239_0001
Figure imgf000240_0001
Figure imgf000241_0001
Figure imgf000242_0001
Figure imgf000243_0001
Figure imgf000244_0001
Figure imgf000245_0001
Figure imgf000246_0001
TABLE 3. Biological activity of Compound A in the Hsp90α, Hsp90β, Grp94, and TRAP-1 competitive binding assays (Example A).
Figure imgf000246_0002
TABLE 4. Biological activity of Compound A in the HER2 degradation assay (Example C).
Figure imgf000247_0001
Example D
Tumor Cell Growth Inhibition Assay
[00664] The cell growth inhibitory activity of Compound A was determined by treating tumor
(BT474, MCF7 and N87) cells with various concentrations of Compound A for 5 days followed by colorimetric measurement of cell viability (MTS assay).
[00665] Compound A was diluted and added at a concentration range to the plated cells. DMSO
(0.03 to 0.003%) was included as a vehicle control. Cells were incubated in the presence of Compound A for 5 days. Phenazine methosulfate (PMS, stock concentration 1 mg/mL) and MTS (3-(4,5- dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium inner salt, stock concentration 2 mg/mL from Promega, Wl) were mixed at a concentration of 1 :20 and added to each well of a 96 well plate. Reduction of MTS gave rise to a soluble formazan product that was secreted into the culture medium. After 4 hours incubation, the formazan product was quantitated spectrophotometrically at a wavelength of 490 nm. Data was acquired using SOFTmaxΘPRO software. 100% viability was defined as the OD 490 of DMSO treated cells stained with MTS (the mean OD490 of cells treated with DMSO at a range of 0.03 to 0.003%). Percent viability of Compound A was calculated from the OD490 values as follows: % viability = (OD490 nm sample/OD490 nm DMSO treated cells x 100). The IC50 was defined as the concentration that gave rise to 50% inhibition of cell viability. The IC5O values for Compound A observed in the three tumor cell lines are shown in Table 5.
TABLE 5. Biological activity of Compound A in the tumor cell growth inhibition assay (Example D).
Figure imgf000247_0002
Example E
Tumor Client Protein Inhibition Assay
[00666] The effects of Compound A on Hsp90 client and heat shock proteins was tested over a range of concentrations in MCF7 and N87 tumor cells. The cells were exposed to 8-1000 nM of
Compound A for 24 hours. Treated cells were collected and levels of expression and/or phosphorylation of various client and heat shock proteins was determined by Western blotting. Figure 1 shows the results in MCF7 cells. The phosphorylation and protein levels of Akt, Raf, and Rb were affected at a concentration of 125-250 nM Compound A. No change in actin or p85PI3K levels were observed. Figure
2 shows the results in N87 cells. The phosphorylation and protein levels of HER2 were affected at a concentration of 63-125 nM of Compound A. Effects on several other client proteins, including EGFR,
Akt, ERK and Rb, were observed at 63-250 nM Compound A. No change in the actin protein levels was observed.
Example F
Human Mixed Lymphocyte Reaction Assay
[00667] A mixed lymphocyte reaction represents the T cell receptor dependent proliferative response of CD4+ T cells to histocompatibility antigens on non-T cells from a non-identical individual. The stability and function of newly synthesized and phosphorylated forms of a critical proximal signaling molecule in the T cell receptor activation pathway, the Src family kinase p56/c/<, is dependent upon Hsp90. The mixed lymphocyte reaction was used to measure the ability of Compound A to inhibit human CD4+ T cell activation as quantified by the uptake of tritium-labeled thymidine by dividing cells (Table 6).
[00668] Blood from two in-house donors (150 mL per donor) was collected into heparinized vacuum tubes (Becton Dickenson). Peripheral blood mononuclear cells (PBMCs) were isolated on Histopaque (Sigma) and CD4+ cells separated from both donors using MACS Human CD4+ T cell Isolation Kit Il (Miltenyi Biotec). Non-CD4+ cells were irradiated (2000 rads). CD4+ cells from one donor (7.5 x 104) were combined with non-CD4+ cells of the other donor at a ratio of 1 :3 (responders:stimulators) in round bottom 96 well plates (Costar). Solutions of Compound A were prepared in 100% DMSO and serially diluted 3-fold for a final concentration range of 0.5 nM to 10 μM. Compound A was added to cells for a final DMSO concentration of 0.1%. Culture was for 5 days at 370C in X-VIVO™ medium (Lonza), supplemented with 10% FBS (Hyclone), 4 mM L-Glutamine, 1 mM Sodium Pyruvate, 1x MEM Non-essential amino acids, 10 mM HEPES, pH7.3, and Antimycotic Antibiotic (100 units/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B; Invitrogen). Eighteen hours prior to harvest [3H]thymidine (American Radiolabeled Chemicals) was added (1 μCi/well). Cells were harvested using a Packard Filtermate 196 cell harvester (PerkinElmer) onto UniFilter GF/C plates (PerkinElmer). Scintillation fluid (50 μl_) was added to the filters and [3H]thymidine incorporated into DNA was quantified using a TopCount NXT (PerkinElmer). The EC50 was defined as the concentration resulting in a 50% decrease in CD4+ T cell proliferation relative to 0.1% DMSO treated cells.
TABLE 6. Biological activity of Compound A in the human mixed lymphocyte reaction assay.
Figure imgf000248_0001
Example G
CD4 Modulation Assay
[00669] The Src family kinase p56/cΛ is associated with surface CD4 in resting T cells. This association between CD4 and pSδ'1* inhibits CD4 endocytosis by preventing its entry into coated pits. The stability of newly synthesized and activated/phosphorylated forms of p56fc;t is dependent upon Hsp90, with Hsp90 inhibition resulting in p56fc* degradation and subsequent internalization of CD4. A cell-based assay designed to quantify CD4 expression level on human peripheral blood T cells was utilized to measure Hsp90 inhibition with an intention to translate the readout as an in vivo pharmacodynamic marker.
[00670] The CD4 modulation assay was used to quantify the ability of Compound A to down modulate T cell surface CD4 in vitro over a dose response (0.5 nM to 10 μM) and time course up to 3 days (Figure 3). Maximal CD4 down modulation of approximately 70% was obtained after 72 h continuous exposure to Compound A at or above 370 nM. Initial time course experiments established that the earliest detectable decrease in CD4 levels occurred after 6 to 8 h of in vitro compound exposure (not shown). Over the 72 h time course the EC50 value of Compound A in this assay remained constant at approximately 120 nM (Figure 3).
[00671] Blood from in-house donors (150 mL/per donor) was collected into heparinized vacuum tubes (Becton Dickenson). Peripheral blood mononuclear cells were isolated on Histopaque (Sigma) and CD4+ cells purified by negative selection using magnetic microbeads (MACS®) (Miltenyi Biotec, MACS CD4+ T Cell Isolation Kit II, Human). Purified CD4+ cells were cultured in 96-well plates between 4 and 40 x 105 cells/mL in medium [X-Vl VO™ Medium (Lonza) supplemented with 10% FBS (Hyclone), 4 mM L- glutamine, 1 mM sodium pyruvate, 10 mM HEPES, pH 7.3, 0.1 mM MEM Non-essential amino acids, 100 units/mL penicillin, 100 μg/mL streptomycin, and 0.25 μg/mL amphotericin B (Antimycotic Antibiotic, Invitrogen)]. Dilutions of Compound A were made in DMSO, diluted 40-fold with medium and added to cells for a final DMSO concentration of 0.1% and final culture volume of 250 μl_. Cells and Compound A were incubated at 37°C for various times, harvested, washed, and stained with the following antibodies at 40C for 30 min: Phycoerythrin (PE)-Cy5 conjugated anti-human CD4, PE-anti-human CD45RA, PE-Cy7- anti-human CD2, and Allophycocyanin (APC)-Cy7-anti-human CD25 (all from BD Biosciences). Cells were washed, resuspended in PBS, 3% FCS, and multicolor fluorescence analyzed on an LSRII flow cytometer (BD Biosciences). Machine settings were maintained constant between runs. For each sample the geometric mean fluorescence intensities (MFI) of CD4 and CD2 staining were measured on live CD4+ cells. CD4 MFI was normalized to CD2 MFI and expressed as a percentage of control cells incubated in medium only.
[00672] Maximal CD4 down modulation of approximately 70% was obtained after 72 h continuous exposure to Compound A at or above 370 nM. Initial time course experiments established that the earliest detectable decrease in CD4 levels occurred after 6 to 8 h of in vitro compound exposure (not shown). Over the 72 h time course the EC50 value of Compound A in this assay remained constant at approximately 120 nM (Figure 3). Example H Macrophage TLR4 Signaling Assay [00673] LPS signaling in macrophages is mediated through Toll-like Receptor 4 (TLR4) and the
LPS coreceptor, CD14 (Beutler, B. Curr. Top. Microbiol. Immunol., 2002, 270:109-120). The antiinflammatory effects of geldanamycin on LPS-stimulated TLR4 signaling have been previously noted in the mouse macrophage-like cell line RAW 264.7 as an inhibition of TNF-D and IL-6 production (Byrd, CA. et al. Proc. Natl. Acad. Sci. USA, 1999, 96:5645-5650; Wax, S. et al. Arthritis Rheum., 2003, 48:541 -550; Zhu, F.G. et al. Infect. Immun., 2001 , 69:5546-5552). A dose-dependent and Hsp90-dependent inhibition of LPS-stimulated TNF-α and IL-6 production in RAW 264.7 cells was confirmed with an Hsp90 inhibitor using an optimized pretreatment time with compound (4 h) and an optimized time of LPS stimulation (results not shown).
[00674] In macrophages, the MAPKs, ERK1/2, JNK1/2 and p38 MAPK are all activated by LPS, and inhibition of any one of these MAPKs results in attenuation of TNF-α induction (Dumitru, CD. et al. Cell, 2000, 103:1071 -1083).
[00675] To determine effects of Compound A on LPS-induced MAPK activation, RAW 264.7 cells were obtained from the ATCC and maintained in DMEM supplemented with 10% FCS (Hyclone). Cells were plated in 6-well dishes and grown for approximately 18 h in fresh medium. Medium was replaced with fresh, warm medium and Compound A in DMSO or DMSO alone was added with the final concentration of DMSO not exceeding 0.5%. After 4 h at 37°C, LPS (100 ng/mL, Escherichia coli 0111 :B4, Sigma) was added for 30 minutes. Cells were rinsed with ice-cold PBS and lysed in RIPA buffer (Upstate) containing Halt Protease Inhibitor Cocktail (Thermo Scientific) and Halt EDTA (Thermo Scientific). Protein concentrations of cell lysates were determined by Bradford assay (BioRad). Samples were adjusted to 1 mg/mL in lysis buffer. NuPage SDS Sample Buffer (Invitrogen) and NuPage Reducing Sample Buffer (Invitrogen) were added, samples heated to 1000C for 10 min, then stored frozen at -80°C For immunoblots samples were thawed, heated to 1000C, and electrophoresed on a NuPAGE 4-12% SDS polyacrylamide gel (Invitrogen). Proteins were transferred to Invitrolon PVDF membranes (Invitrogen) for immunoblotting. Antibodies used in immunoblotting were obtained from Cell Signaling Technologies and were specific to the following proteins: phospho-JNK (Thr183/Thr185); phospho- ERK1/2 (Thr202/Tyr204); phospho-MEK1/2 (Ser217/221); phospho-p38MAPK (Thr180/Tyr182, 3D7); ERK1/2 (137F5); p38MAPK; JNK; MEK1/2. Anti-GAPDH was a rabbit polyclonal IgG (Santa Cruz Biotechnology). The secondary antibody was a goat polyclonal anti-rabbit IgG conjugated to HRP (BioRad). Optimal dilutions of primary antibodies were determined for individual antibody lots as indicated in the manufacturer's instructions. Binding of the secondary antibody (HRP-conjugated goat anti-rabbit IgG, 1 :2000 in 5% BSA) was revealed by either SuperSignal West Pico or Femto Chemiluminescent Substrate System (Pierce). To strip the membranes of bound antibody, membranes were incubated in Restore Western Blot Stripping Buffer (Thermo Scientific), rinsed, and blocked with 5% BSA (Sigma Aldrich). Stripped membranes were immunoblotted with antibody to GAPDH as above.
[00676] Treatment of RAW 264.7 cells with Compound A resulted in a dose dependent decrease of phospho-ERK1/2 and phospho-JNK1/2 (Figure 4). Levels of phospho-p38 MAPK were not significantly changed by Compound A treatment (Figure 4). Total levels of each signaling protein did not differ between Compound A and DMSO treated cells (Figure 4, right panels). The phosphorylation of MEK1/2, the MAP2K upstream of ERK1/2, was also decreased by Compound A in a dose dependent manner, without altering levels of total MEK1/2 (Figure 4, right panel). These results demonstrate that in the mouse macrophage cell line RAW 264.7 Hsp90 inhibition with Compound A blocks LPS-induced activation of the MAPK pathways that are required for LPS-induced TNF-α production.
Example I
In Vivo Oncology Studies
[00677] Preclinical antitumor activity of Compound A administered as a single agent was evaluated in human tumor bearing athymic nude or SCID mice. Tumor lines that were used as xenografts included the high HER-2 expressing tumor models, NCI-N87 (gastric cancer) and BT474
(breast cancer), and a model that did not express high levels of the HER-2 protein, HT-29 (colon).
[00678] The primary xenograft model for testing Hsp90 inhibitors is the NCI-N87 HER-2+ gastric carcinoma model. This model is dependent upon HER-2 (a sensitive Hsp90 client protein) for growth.
This model is more reproducible and more sensitive than other HER-2+ models. The BT474 model was run after Compound A showed activity in the N87 tumor model. This model is more difficult to run since the mice require estradiol supplementation for BT474 tumor growth. In some cases, estradiol supplementation leads to added toxicity and body weight loss in the animals bearing the BT474 tumors.
A third human tumor model, the HT29 colon carcinoma xenograft, was also used to test Compound A activity.
[00679] Compound A demonstrated single-agent antitumor activity in three different xenograft models at doses ranging from 10 to 40 mg/kg, administered orally, on a daily QDx5 schedule. Efficacy results from the models evaluated are summarized in Figure 5.
Example J
Primary Pharmacology Model Data
[00680] The NCI-N87 HER-2+ gastric carcinoma model was the primary testing model for
Compound A. This model is dependent upon HER-2 (a sensitive Hsp90 client protein) for growth. Five studies were completed in this human tumor xenograft model.
[00681] Compound A showed potent and significant antitumor activity in the NCI-N87 human gastric carcinoma xenograft model. Data from a representative study is shown in Figure 6. Compound A was administered at 10, 20, 30, 35, and 40 mg/kg po qdx5. The percentage tumor growth inhibition (%
TGI) value was calculated based on the difference in mean tumor volume on the first dosing day (14 days post tumor inoculation) and the last measurement day after 4 weeks of dosing (day 44). The % TGI for
10, 20, 30, 35, and 40 mg/kg given orally qdx5 was 45%, 85%, 102%, 107%, and 103%, respectively.
The mean tumor volume in the four highest dose groups was significantly different from the mean tumor volume of the control group (p = 0.07, 0.0005, 0.0001 , 0.0001 and 0.0001 , respectively).
Example K Secondary Pharmacaology Model Data
[00682] In the BT474 breast carcinoma xenograft model, Compound A was administered orally
QDx5 at doses of 10, 20, 35, and 40 mg/kg. Treatment with Compound A gave a dose dependent inhibition of BT474 tumor growth (Figure 7). The %TGI values for 10, 20, 35, and 40 mg/kg administered orally QDx5 were 72%, 140%, 149%, and 149% respectively. The mean tumor volumes in all 4 groups were significantly different from the mean tumor volume of the control group (p = 0.006 in the 10 mg/kg group and p < 0.0001 in the remaining groups.
[00683] In the HT-29 colon carcinoma model, Compound A was administered orally QDx5 at 10,
20 and 40 mg/kg. Compound A showed antitumor activity in this tumor model (Figure 8). The % TGI values were calculated after 4 weeks of dosing (39 days post tumor inoculation). For the 10, 20 and 40 mg/kg groups the % TGI values were 43%, 61% and 76%, respectively. The p-values for the treated groups were 0.26, 0.08, and 0.04, respectively.
Example L
LPS-lnduced TNF-α Pharmacology Model
[00684] Preclinical anti-inflammatory activity of Compound A was evaluated in the mouse LPS- induced TNF-α model. Compound A was administered orally to female DBA/1 mice as a split dose, three and six hours prior to a sublethal LPS challenge. TNF-α was quantified in serum collected one hour after
LPS challenge.
[00685] Formulation of Compound A was as a suspension in 0.5% carboxymethylcellulose, 0.1%
Tween-80 (Sigma-Aldrich). Female DBA/1 mice (5 animals/group) were dosed according to body mass by oral gavage in two gavages given 3 h apart. The dose listed is the total of the two gavages (for example, a 50 mg/kg dose was administered in two, 25 mg/kg gavages). Three hours after the second gavage, mice were injected intraperitoneal^ with LPS (200 μg/kg). One hour later, blood was collected by facial vein puncture in BD Microtainer Serum Separation Tubes (BD Biosciences). Sera were frozen at -8O0C and assayed for TNF-α by ELISA according to the manufacturer's instructions (R&D Systems). Sera samples were serially diluted and values in the linear range of the titration were used to calculate the original concentration of the cytokine in the undiluted samples. Limit of detection in the TNF-α ELISA was
23.6 pg/ml. Horizontal bars represent for each group the mean serum TNF-α values ± SEM.
[00686] Compound A demonstrated anti-inflammatory activity in the LPS-induced TNF-α model at doses ranging from 12.5 to 100 mg/kg (Figure 9). Complete inhibition of LPS-induced TNF-α was found at the 100 mg/kg dose, and the inhibitory response was dose dependent.
Example M
Rat Collagen-Induced Arthritis Model
[00687] Compound A was administered semi-therapeutically in a rat CIA model, starting oral dosing on day 12, two days after the type Il collagen boost and dosing once every 3 days (q3d).
Compound A demonstrated a dose responsive inhibition in arthritis index, with statistically significant disease inhibition in the 12.5 mg/kg treatment group (*, p < 0.05; Figure 10) and in the 25, 35, and 40 mg/kg treatment groups (Figure 10). Disease development as measured by arthritis index in the 5 mg/kg treatment group was indistinguishable from that in the vehicle control group.
[00688] Female DA rats were immunized intradermal^ with rat collagen type Il (CII; Chondrex;
250 μl 1 :1 emulsion of 2 mg/ml rat CII in 0.05 N acetic acid: Incomplete Freund's Adjuvant) in both pinnae and at one site in the abdominal skin. Ten days post immunization rats received a second injection of rat CII (500 μg CII in PBS, intraperitoneal^). Compound A suspension or vehicle (0.5% carboxymethylcellulose, 0.1% Tween-80) administration was every 3 days (q3d) by oral gavage beginning on d 12 (5 animals/group). Clinical severity of CIA was assessed by monitoring inflammation on all four paws, applying a scale ranging from 0 to 4. Each paw was graded as follows: 0, normal; 1 , mild but definite redness and swelling of the ankle or wrist, or redness and swelling of any severity for 1 or 2 digits; 2, moderate to severe redness and swelling of the ankle or wrist, or more than two digits; 3, redness and swelling (pronounced edema) of the entire paw; and 4, maximally inflamed limb with involvement of multiple joints. The sum of the four individual scores was the arthritis index, with a maximal possible score of 16 for each animal. Each point represents the mean arthritis index ± SEM for animals in a group.
[00689] The foregoing examples are not limiting and are merely illustrative of various aspects and embodiments of the present invention. All documents cited herein are indicative of the levels of skill in the art to which the invention pertains. None, however, is admitted to be prior art. [00690] One skilled in the art will readily appreciate that the present invention is well adapted to carry out the objects and obtain the ends and advantages mentioned, as well as those inherent therein. The methods and compositions described illustrate preferred embodiments, are exemplary, and are not intended as limitations on the scope of the invention. Certain modifications and other uses will occur to those skilled in the art, and are encompassed within the spirit of the invention, as defined by the scope of the claims.
[00691] The invention illustratively described herein suitably may be practiced in the absence of any element or elements, limitation or limitations which is not specifically disclosed herein. The terms and expressions which have been employed are used as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding any equivalents of the features shown and described, or portions thereof. It is recognized that various modifications are possible within the scope of the invention claimed. Thus, it should be understood that although the present invention has been specifically disclosed by preferred embodiments, optional features, modifications and variations of the concepts herein disclosed may be resorted to by those skilled in the art, and that such modifications and variations are considered to be within the scope of this invention as defined by the description and the appended claims.
[00692] In addition, where features or aspects of the invention are described in terms of Markush groups or other grouping of alternatives, e.g., genuses, those skilled in the art will recognize that the invention is also thereby described in terms of any individual member or subgroup of members of the Markush group or subgenus, and exclusions of individual members as appropriate, e.g., by proviso. [00693] Other embodiments are within the following claims.

Claims

WHAT IS CLAIMED IS:
1. A compound represented by Formula I
Figure imgf000255_0001
or a pharmaceutically acceptable salt thereof, wherein:
X is a bond or an alkylene chain of the formula (CR 3aarR>bD \)n where n is 1 to 3, for each occurrence, Ra and Rb are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which, except for hydrogen, is optionally substituted, and a (CRaRb) unit is optionally replaced by -O-, -N(Ra)-, or -S-; or when n is 2 or 3, Ra in a first (CRaRb) unit is optionally taken together with Ra in a different (CRaRb) unit to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or
Ra is optionally taken together with Rb in the same (CRaRb) unit to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; R1 is selected from hydrogen, halogen, -OR11, -SR12, amino, and optionally substituted lower alkyl;
R2 is selected from -NR21R22, -N=C-NR21R22, -NR21 -C(O)R23 and -NR21 -SO2R24; R3 and R4 are independently selected from hydrogen, optionally substituted alkyl, optionally substituted alkenyl, optionally substituted alkynyl, optionally substituted alicyclic, optionally substituted alicyclicalkyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, -C(=J)RX, -C(=J)N(Ry)(R2), C(=J)N(RX)S(O)2RX, -C(=J)N(RX)N(RX)S(O)2RX, -C(RX)=N(ORX), -C(Rx)=NN(Ry)(R1), -C≡N, -C(=J)ORX, -Cf=J)SRx, -N(Ry)(R2), -N(RX)C(=J)RX, -N(Rx)C(=J)N(Ry)(Rl), -N(RX)C(=J)ORX, -N(RX)C(=J)SRX, -N(RX)S(O)2RW, -N(Rx)S(O)2ORx, -S(O)1ORx (where t is 1 or 2), - S(O)NRxR2, -NH-C(-NH2)=C-NO2, -CO-NH-SO2-NH2, -N(Rx)S(O)2N(Ry)(Rz), N3, NO2, -ORx, -0C(=J)Rx, -0C(=J)0Rx, -0C(=J)N(Ry)(Rz), -0C(=J)SRx, -0P(0)(Rv)2, -OS(O)2Rx, -OS(O)2N(Ry)(Rz), -0Si(Rw)3, -P(O)(Rv)2, -SRx, -S(OJtRx (where t is 1 or 2), -S(0)2N(Ry)(Rz), -Si(RJ3, and halogen, wherein each J is independently O, NRU or S wherein each R11 is independently hydrogen, alkyl, alkenyl, alkynyl, cyano, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rv is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, heteroarylalkyl, hydroxy, -ORx or -N(Ry)(R2); each Rw is independently alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; each Rx is independently hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; and Ry and Rz are each independently hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, or heteroarylalkyl; or Ry and Rz, together with the nitrogen atom to which they are attached, form a heterocyclic or heteroaryl; or
R3 is optionally taken together with R4 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, n is 1 , and the (CRaRb) unit is replaced by -N(Ra)-, then R3 and R4 taken together optionally form oxo; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, an Ra unit is optionally taken together with R1 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n, an Ra in a first (CRaRb) unit is optionally taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is a bond, R1 is optionally taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90;
R5 and R6 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; or
R5 is optionally taken together with R6, to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90; or when X is an optionally substituted alkylene chain of the formula (CRaRb)n , n is 1 , and the (CRaRb) unit is replaced by -N(Ra)-, then R5 and R6 taken together optionally form oxo; or
R3 is optionally taken together with R5, to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S1 =N, and NR90;
R7 is selected from a bond, optionally substituted alkylene, -C(O)-, -C(S)-, -S(O)-, and -SO2-;
R8 is selected from alkyl, alicyclic, heterocyclic, aryl, and heteroaryl, each of which is optionally substituted;
R11 and R12 are independently selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl and heteroaryl, each of which, except for hydrogen, is optionally substituted; and
R21 and R22 are independently selected from hydrogen, alkyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl, each of which, except for hydrogen, is optionally substituted, or
R21 is optionally taken together with R22 to form an optionally substituted ring of 3-8 ring atoms wherein said optionally substituted ring optionally includes one or two additional ring atoms that are heteroatoms independently selected from O, S, =N, and NR90;
R23 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl;
R24 is selected from alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and
R90 is selected from hydrogen, alkyl, alkenyl, alkynyl, alicyclic, alicyclicalkyl, heterocyclic, heterocyclicalkyl, aryl, arylalkyl, heteroaryl, and heteroarylalkyl; and wherein from 1 to 5 hydrogen atoms in the compound of Formula I are optionally replaced with deuterium, and provided that when X is an optionally substituted alkylene chain of the formula (CRaRb)n , n is 1 , the (CRaRb) unit is replaced by -N(Ra)-, and R7 is -CH2- then R8 is not 3,5-dimethyl-4-methoxy-pyridin-2-yl.
2. The compound or pharmaceutically acceptable salt of claim 1 , wherein X is selected from -0-, CH2, -CH2N(R3)-, and -N(Ra)-, or X is a bond.
3. The compound or pharmaceutically acceptable salt of claim 1 , wherein the compound of Formula I is represented by Formula Il
Figure imgf000258_0001
4. The compound or pharmaceutically acceptable salt of claim 1 , wherein the compound of
Formula I is represented by Formula III
Figure imgf000258_0002
5. The compound or pharmaceutically acceptable salt of any one of claims 1 to 4 wherein R1 is halogen selected from fluoro, chloro, and bromo.
6. The compound or pharmaceutically acceptable salt of claim 5 wherein R1 is chloro.
7. The compound or pharmaceutically acceptable salt of any one of claims 1 to 3 wherein X is a bond and R1 is taken together with R3 to form an optionally substituted ring of 3-8 ring atoms wherein one of the 3-8 ring atoms optionally is O or S.
8. The compound or pharmaceutically acceptable salt of any one of claims 1 to 7 wherein i21
R is hydrogen.
9. The compound or pharmaceutically acceptable salt of any one of claims 1 to 8 wherein R22 is hydrogen.
10. The compound or pharmaceutically acceptable salt of any one of claims 1 to 9 wherein R5 and R6 are independently selected from hydrogen and optionally substituted lower alkyl.
11. The compound or pharmaceutically acceptable salt of claim 10 wherein R5 is hydrogen.
12. The compound or pharmaceutically acceptable salt of claim 10 wherein R5 is -CH2OH.
13. The compound or pharmaceutically acceptable salt of any one of claims 1 to 12 wherein R6 is hydrogen.
14. The compound or pharmaceutically acceptable salt of any one of claims 1 to 13 wherein R7 is a bond or optionally substituted alkylene.
15. The compound or pharmaceutically acceptable salt of claim 14 wherein R7 is optionally substituted lower alkylene.
16. The compound or pharmaceutically acceptable salt of claim 15 wherein R7 is -CH2-.
17. The compound or pharmaceutically acceptable salt of claim 15 wherein R7 is -CH(CH3)-.
18. The compound or pharmaceutically acceptable salt of any one of claims 1 to 17, wherein R8 is selected from alicyclic, aryl, heterocyclic, and heteroaryl, each of which is optionally substituted.
19. The compound or pharmaceutically acceptable salt of claim 18 wherein R8 is phenyl, pyridinyl, pyrimidinyl, pyridazinyl, pyrazinyl, quinolinyl, isoquinolinyl, tetrahydroquinolinyl, benzothiazolyl, pyrazolyl, thiazolyl, tetrahydroisoquinolinyl, imidazolyl, or benzodioxolyl, each of which is optionally substituted.
20. The compound or pharmaceutically acceptable salt of claim 18 wherein R8 is selected from:
Figure imgf000260_0001
Figure imgf000260_0002
Figure imgf000260_0003
Figure imgf000261_0001
21. The compound or pharmaceutically acceptable salt of claim 18 wherein R8 is selected from:
Figure imgf000262_0001
Figure imgf000262_0002
Figure imgf000262_0003
Figure imgf000262_0005
Figure imgf000262_0006
Figure imgf000262_0004
Figure imgf000262_0007
Figure imgf000263_0001
Figure imgf000264_0001
Figure imgf000265_0001
22. The compound or pharmaceutically acceptable salt of claim 18 wherein R8 is
Figure imgf000265_0002
23. The compound or pharmaceutically acceptable salt of claim 19 wherein from 1 to 5 hydrogen atoms of R8 are optionally replaced with deuterium.
24. The compound or pharmaceutically acceptable salt of any one of claims 1 to 17 wherein R8 is optionally substituted lower alkyl.
25. The compound or pharmaceutically acceptable salt of any one of claims 1 to 6 or 8 to 24 wherein R3 is selected from hydrogen, optionally substituted alkyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, optionally substituted aryl, optionally substituted arylalkyl, optionally substituted heteroaryl, optionally substituted heteroarylalkyl, and halogen.
26. The compound or pharmaceutically acceptable salt of claim 25 wherein R3 is lower alkyl.
27. The compound or pharmaceutically acceptable salt of claim 26 wherein R3 is methyl.
28. The compound or pharmaceutically acceptable salt of claim 26 wherein R3 is f-butyl.
29. The compound or pharmaceutically acceptable salt of claim 25 wherein R3 is alkyl substituted with one or two groups selected from
-OR31 wherein R31 is selected from hydrogen, alkyl, trialkylsilyl, and -C(O)R40 wherein R40 is selected from hydrogen, alkyl, alicyclic, heterocyclic, aryl, and heteroaryl;
-S(O)nR32 or -OS(O)2R32 wherein R32 is selected from alkyl, phenyl, and -NR51R52 wherein R51 and R52 are independently selected from hydrogen and alkyl, and n is 1 or 2;
-NR33R34 wherein R33 and R34 are independently selected from hydrogen, lower alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, -C(O)R60 wherein R60 is selected from hydrogen, alkyl, aryl, and amino, and -S(O)2-alkyl or R33 is optionally taken together with R34 to form an optionally substituted heterocyclic or optionally substituted heteroaryl ring;
-C(O)YR35 wherein Y is selected from -NR36 and O and wherein R35 and R36 are independently selected from hydrogen, alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, optionally substituted alicyclic, optionally substituted heterocyclic, optionally substituted aryl, and optionally substituted heteroaryl or R35 is optionally taken together with R36 to form an optionally substituted 5-6 membered heteroaryl or optionally substituted 5-6 membered heterocyclic ring; halogen; azido; and cyano.
30. The compound or pharmaceutically acceptable salt of claim 29 wherein R3 is alkyl substituted with one or two groups selected from
-OR31 wherein R31 is selected from hydrogen, alkyl, and -C(O)R40 wherein R40 is selected from hydrogen, alkyl, and aryl;
-OS(O)2R32 wherein R32 is selected from alkyl and phenyl;
-NR33R34 wherein R33 and R34 are independently selected from hydrogen, lower alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, -C(O)R60 wherein R60 is selected from hydrogen, alkyl, aryl, and amino, and -S(O)2-alkyl or R33 is optionally taken together with R34 to form an optionally substituted heterocyclic or optionally substituted heteroaryl ring; -C(O)YR35 wherein Y is selected from -NR36 and O and wherein R35 and R36 are independently selected from hydrogen, alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy, alicyclic, heterocyclic, aryl, and heteroaryl; and azido.
31. The compound or pharmaceutically acceptable salt of claim 30 wherein R3 is selected from -CH2-OH, -CH2OCH3, -CH2CH2-OH, -CH(OH)CH3, -C(CH3J2CH2OH, -CH2-OC(O)CH3, -CH2-NH2, -CH2-NHCH3, -CH2N(CH3J2, -CH2CH2NH2, -CH2-NHCH2CH2OCH3, -CH2-N(CH2CH2OCH3)Z, -CH2-N(CH2CH2OH)2, -CH2-NHC(O)CH3, -CH2-NHC(O)Ph, -CH2-NHC(O)NH2, -CH2-NHS(O)2CH3, -CH2-C(O)OH, -CH2-C(O)OCH3, -CH2C(O)NH2, and -CH2OS(O)2CH3.
32. The compound or pharmaceutically acceptable salt of claim 29 wherein R3 is alkyl substituted with -NR33R34 wherein R33 is taken together with R34 to form an optionally substituted heterocyclic or optionally substituted heteroaryl ring selected from isoindolinyl, triazolyl, and piperidinyl.
33. The compound or pharmaceutically acceptable salt of claim 32 wherein the optional substitutents on the isoindolinyl or triazolyl rings are independently selected from oxo and -CH2N(CH3)2.
34. The compound or pharmaceutically acceptable salt of claim 29 wherein R3 is alkyl substituted with -C(O)YR35 wherein Y is -NR36 and wherein R35 and R36 are independently selected from hydrogen and alkyl optionally substituted with one or two groups independently selected from alkoxy and hydroxy.
35. The compound or pharmaceutically acceptable salt of claim 34 wherein R3 is selected from -CH2C(O)NH2, -CH2C(O)NHCH3, CH2C(O)NHCH2CH3, -CH2C(O)N(CH3)2, -CH2C(O)NHCH2CH2OCH3, -CH2C(O)NHCH2CH(OH)CH2OH. -CH2C(O)NHCH(CH3)CH2OH, and -CH2CH2C(O)NH2.
36. The compound or pharmaceutically acceptable salt of claim 29 wherein R3 is alkyl substituted with -C(O)YR35 wherein Y is -NR36 and wherein R35 is taken together with R36 to form an optionally substituted heterocyclic ring.
37. The compound or pharmaceutically acceptable salt of claim 36 wherein the optionally substituted heterocyclic ring is selected from morpholinyl, azetidinyl, pyrrolidinyl, and piperidinyl, each of which is optionally substituted.
38. The compound or pharmaceutically acceptable salt of claim 37 wherein the substituents on the morpholinyl, azetidinyl, pyrrolidinyl, and piperidinyl rings are selected from -CH2OH, -CH2CH2OH, ■ C(O)NH2, and -OH.
39. The compound or pharmaceutically acceptable salt of claim 29 wherein R3 is alkyl substituted -C(O)YR35 wherein Y is -NR36 and wherein R35 and R36 are independently selected from hydrogen, optionally substituted alicyclic ring, and optionally substituted heterocyclic ring.
40. The compound or pharmaceutically acceptable salt of claim 39 wherein R35 and R36 are independently selected from hydrogen, optionally substituted cyclopropanyl, and optionally substituted oxetanyl.
41. The compound or pharmaceutically acceptable salt of claim 25 wherein R3 is selected from optionally substituted phenyl, optionally substituted heterocyclic, optionally substituted heterocyclicalkyl, and optionally substituted heteroarylalkyl.
42. The compound or pharmaceutically acceptable salt of claim 41 wherein R3 is selected from:
Figure imgf000268_0001
Figure imgf000268_0003
Figure imgf000268_0002
Figure imgf000269_0001
43. The compound or pharmaceutically acceptable salt of any one of claims 1 to 42 wherein R4 is selected from hydrogen and lower alkyl.
44. The compound or pharmaceutically acceptable salt of claim 43 wherein R4 is hydrogen.
45. The compound or pharmaceutically acceptable salt of any one of claims 1 to 6 or 8 to 24 wherein R3 is taken together with R4 to form an optionally substituted ring of 3-8 ring atoms wherein optionally one or two of those ring atoms are heteroatoms independently selected from O, S, =N, and NR90.
46. The compound or pharmaceutically acceptable salt of claim 45 wherein R3 is taken together with R4 to form a piperidinyl ring optionally substituted with one or two groups selected from acyl and arylalkyl.
47. The compound or pharmaceutically acceptable salt of claim 46 wherein R3 is taken together with R4 to form a piperidinyl ring substituted with -C(O)CH3 or benzyl.
48. The compound or pharmaceutically acceptable salt of claim 47 wherein R3 is taken together with R4 to form a piperidinyl ring wherein the nitrogen of the piperidinyl ring is substituted with - C(O)CH3 or benzyl.
49. The compound or pharmaceutically acceptable salt of any one of claims 1 to 6, 8, 9, or 14 to 24 wherein R3 is taken together with R5 to form an optionally substituted ring of 3-8 ring atoms, wherein optionally one or two of the 3-8 ring atoms are heteroatoms independently selected from O, S, =N, and NR90.
50. The compound or pharmaceutically acceptable salt of claim 3 wherein R1 is chloro; FT is -NH2;
R3 is optionally substituted alkyl;
R4 is hydrogen;
R5 and R6 are hydrogen;
R7 is -CH2-; and
R8 is selected from optionally substituted aryl and optionally substituted heteroaryl.
51. The compound or pharmaceutically acceptable salt of claim 4 wherein R1 is chloro;
R2 is -NH2;
R3 is optionally substituted alkyl;
R4 is hydrogen;
R5 and R6 are hydrogen;
R7 is -CH2-; and
R8 is selected from optionally substituted aryl and optionally substituted heteroaryl.
52. The compound or pharmaceutically acceptable salt of claim 1 wherein the stereocenter to which R3 and R4 are attached is of the S-configuration.
53. The compound or pharmaceutically acceptable salt of claim 1 wherein the stereocenter to which R3 and R4 are attached is of the f?-configu ration.
54. The compound or pharmaceutically acceptable salt of claim 1 wherein the stereocenter to which R5 and R6 are attached is of the S-configuration.
55. The compound or pharmaceutically acceptable salt of claim 1 wherein the stereocenter to which R5 and R6 are attached is of the fl-configuration.
56. A compound selected from:
Figure imgf000270_0001
Figure imgf000271_0001
Figure imgf000272_0001
Figure imgf000273_0001
Figure imgf000274_0001
Figure imgf000275_0001
Figure imgf000275_0003
Figure imgf000275_0002
Figure imgf000276_0001
Figure imgf000277_0001
Figure imgf000278_0001
Figure imgf000279_0001
Figure imgf000280_0001
Figure imgf000281_0001
Figure imgf000282_0001
Figure imgf000283_0001
Figure imgf000284_0001
Figure imgf000285_0001
Figure imgf000286_0001
Figure imgf000287_0001
Figure imgf000288_0001
Figure imgf000289_0001
Figure imgf000290_0001
Figure imgf000290_0002
Figure imgf000290_0003
Figure imgf000290_0004
Figure imgf000291_0001
Figure imgf000292_0001
Figure imgf000293_0001
Figure imgf000294_0001
Figure imgf000295_0001
Figure imgf000295_0003
Figure imgf000295_0004
Figure imgf000295_0002
Figure imgf000296_0001
or a pharmaceutically acceptable salt thereof.
57. A compound selected from:
Figure imgf000296_0002
Figure imgf000297_0001
Figure imgf000298_0001
Figure imgf000298_0003
Figure imgf000298_0002
Figure imgf000299_0001
or a pharmaceutically acceptable salt thereof.
58. A compound of the following formula:
Figure imgf000299_0002
or a pharmaceutically acceptable salt thereof.
59. A compound of the following formula:
Figure imgf000299_0003
or a pharmaceutically acceptable salt thereof.
60. A compound of the following formula:
Figure imgf000300_0001
or a pharmaceutically acceptable salt thereof.
61. A compound of the following formula:
Figure imgf000300_0002
or a pharmaceutically acceptable salt thereof.
62. A compound of the following formula:
Figure imgf000300_0003
or a pharmaceutically acceptable salt thereof.
63. A compound of the following formula:
Figure imgf000300_0004
or a pharmaceutically acceptable salt thereof.
64. A compound of the following formula:
Figure imgf000301_0001
or a pharmaceutically acceptable salt thereof.
65. A compound of the following formula:
Figure imgf000301_0002
or a pharmaceutically acceptable salt thereof.
66. A compound of the following formula:
Figure imgf000301_0003
or a pharmaceutically acceptable salt thereof.
67. A compound of the following formula:
Figure imgf000302_0001
or a pharmaceutically acceptable salt thereof.
68. A compound of the following formula:
Figure imgf000302_0002
or a pharmaceutically acceptable salt thereof.
69. A compound of the following formula:
Figure imgf000302_0003
or a pharmaceutically acceptable salt thereof.
70. A compound of the following formula:
Figure imgf000302_0004
or a pharmaceutically acceptable salt thereof.
71. A compound of the following formula:
Figure imgf000303_0001
or a pharmaceutically acceptable salt thereof.
72. A compound of the following formula:
Figure imgf000303_0002
or a pharmaceutically acceptable salt thereof.
73. A compound of the following formula:
Figure imgf000303_0003
or a pharmaceutically acceptable salt thereof.
74. A pharmaceutical composition comprising one or more pharmaceutically acceptable excipients and at least one compound or pharmaceutically acceptable salt thereof of any one of claims 1 to 73.
75. A method of treating an individual having an Hsp90 mediated disorder comprising administering to said individual at least one compound or pharmaceutically acceptable salt of any one of claims 1 to 73 or a pharmaceutical composition of claim 74.
76. The method of claim 75 wherein the Hsp90-mediated disorder is selected from inflammatory diseases, infections, autoimmune disorders, stroke, ischemia, cardiac disorder, neurological disorders, fibrogenic disorders, proliferative disorders, and metabolic diseases.
77. The method of claim 76 wherein the proliferative disorder is selected from cancers.
78. The method of claim 77 further comprising administering to said individual at least one anti-cancer and/or cytotoxic agent other than a compound of Formula I, or a pharmaceutically acceptable salt thereof.
79. The method of claim 76 wherein the fibrogenic disorders is selected from connective tissue diseases, polymyositis, systemic lupus erythematosis, rheumatoid arthritis, and other fibrotic disorders.
80. The method of claim 79 wherein the fibrogenic disorder is rheumatoid arthritis.
81. The method of claim 79 wherein the fibrogenic disorder is systemic lupus erythematosis.
82. The method of claim 75 or 79 further comprising administering to said individual at least one anti-inflammatory and/or immuno-modulatory agent other than a compound of Formula I, or a pharmaceutically acceptable salt thereof.
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