WO2011028922A1 - Substituted xanthine derivatives - Google Patents

Substituted xanthine derivatives Download PDF

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Publication number
WO2011028922A1
WO2011028922A1 PCT/US2010/047708 US2010047708W WO2011028922A1 WO 2011028922 A1 WO2011028922 A1 WO 2011028922A1 US 2010047708 W US2010047708 W US 2010047708W WO 2011028922 A1 WO2011028922 A1 WO 2011028922A1
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WO
WIPO (PCT)
Prior art keywords
compound
deuterium
disease
patient
compounds
Prior art date
Application number
PCT/US2010/047708
Other languages
French (fr)
Inventor
Roger D. Tung
Julie F. Liu
Scott L. Harbeson
Original Assignee
Concert Pharmaceuticals, Inc.
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority to EA201270339A priority Critical patent/EA023809B1/en
Priority to BR112012008108A priority patent/BR112012008108B1/en
Application filed by Concert Pharmaceuticals, Inc. filed Critical Concert Pharmaceuticals, Inc.
Priority to MX2012002705A priority patent/MX2012002705A/en
Priority to ES10814504.6T priority patent/ES2542401T3/en
Priority to EP20100814504 priority patent/EP2473053B1/en
Priority to CA2771123A priority patent/CA2771123C/en
Priority to JP2012528054A priority patent/JP5292516B2/en
Priority to CN201080039090.5A priority patent/CN102480968B/en
Priority to DK10814504.6T priority patent/DK2473053T3/en
Priority to IN1641DEN2012 priority patent/IN2012DN01641A/en
Priority to MEP-2015-108A priority patent/ME02177B/en
Priority to PL10814504T priority patent/PL2473053T3/en
Priority to AU2010289465A priority patent/AU2010289465B2/en
Priority to SI201030985T priority patent/SI2473053T1/en
Publication of WO2011028922A1 publication Critical patent/WO2011028922A1/en
Priority to ZA2012/01295A priority patent/ZA201201295B/en
Priority to HK12111125.1A priority patent/HK1170377A1/en
Priority to HRP20150797TT priority patent/HRP20150797T1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D473/00—Heterocyclic compounds containing purine ring systems
    • C07D473/02—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6
    • C07D473/04—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms
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    • C07D473/06—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3
    • C07D473/10—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3 with methyl radicals in positions 3 and 7, e.g. theobromine
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    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • A61K31/52—Purines, e.g. adenine
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    • C07D473/02—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6
    • C07D473/04—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms
    • C07D473/06—Heterocyclic compounds containing purine ring systems with oxygen, sulphur, or nitrogen atoms directly attached in positions 2 and 6 two oxygen atoms with radicals containing only hydrogen and carbon atoms, attached in position 1 or 3

Definitions

  • ADME absorption, distribution, metabolism and/or excretion
  • a metabolic inhibitor will be co-administered with an important drug that is rapidly cleared.
  • an important drug that is rapidly cleared.
  • these drugs are typically co- dosed with ritonavir, an inhibitor of cytochrome P450 enzyme CYP3A4, the enzyme responsible for their metabolism.
  • Ritonavir itself has side effects and it adds to the pill burden for HIV patients who must already take a combination of different drugs.
  • dextromethorphan which undergoes rapid CYP2D6 metabolism is being tested in combination with the CYP2D6 inhibitor quinidine for the treatment of pseudobulbar disease.
  • combining drugs with cytochrome P450 inhibitors is not a satisfactory strategy for decreasing drug clearance.
  • the inhibition of a CYP enzyme activity can affect the metabolism and clearance of other drugs metabolized by that same enzyme. This can cause those other drugs to accumulate in the body to toxic levels.
  • a potentially attractive strategy, if it works, for improving a drug's metabolic properties is deuterium modification.
  • Deuterium is a safe, stable, non-radioactive isotope of hydrogen. Deuterium forms stronger bonds with carbon than hydrogen does. In select cases, the increased bond strength imparted by deuterium can positively impact the ADME properties of a drug, creating the potential for improved drug efficacy, safety, and tolerability.
  • the size and shape of deuterium are essentially identical to hydrogen, replacement of hydrogen by deuterium would not be expected to affect the biochemical potency and selectivity of the drug as compared to the original chemical entity that contains only hydrogen.
  • This invention relates to novel compounds that are substituted xanthine derivatives and pharmaceutically acceptable salts thereof.
  • this invention relates to novel substituted xanthine derivatives that are structurally related to pentoxifylline.
  • This invention also provides compositions comprising one or more compounds of this invention and a carrier and the use of the disclosed compounds and compositions in methods of treating diseases and conditions for which pentoxifylline and related compounds are beneficial.
  • FIGS. 1 A and IB depict the serum levels of a compound of this invention, pentoxifylline and certain of their respective metabolites in four individual dogs following oral administration of a combination of pentoxifylline and that compound of this invention.
  • FIG. 2 depicts the time course of the production of the specific metabolites measured in FIG. 3 following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-Ml with rat whole blood.
  • FIG. 3 depicts the relative amount of specific metabolites produced following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-Ml with rat whole blood.
  • FIG. 4 depicts the time course of the production of the specific metabolites measured in FIG. 5 following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-M1 with human liver microsomes.
  • FIG. 5 depicts the relative amount of specific metabolites produced following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-Ml with human liver microsomes DETAILED DESCRIPTION OF THE INVENTION
  • ameliorate and “treat” are used interchangeably and include both therapeutic and prophylactic treatment. Both terms mean decrease, suppress, attenuate, diminish, arrest, or stabilize the development or progression of a disease (e.g., a disease or disorder delineated herein), lessen the severity of the disease or improve the symptoms associated with the disease.
  • a disease e.g., a disease or disorder delineated herein
  • Disease means any condition or disorder that damages or interferes with the normal function of a cell, tissue, or organ.
  • a position designated as having deuterium when a particular position is designated as having deuterium, it is understood that the abundance of deuterium at that position is substantially greater than the natural abundance of deuterium, which is 0.015%.
  • a position designated as having deuterium typically has a minimum isotopic enrichment factor of at least 3340 (50.1% deuterium incorporation) at each atom designated as deuterium in said compound.
  • isotopic enrichment factor means the ratio between the isotopic abundance and the natural abundance of a specified isotope.
  • a compound of this invention has an isotopic enrichment factor for each designated deuterium atom of at least 3500 (52.5% deuterium incorporation at each designated deuterium atom), at least 4000 (60% deuterium incorporation), at least 4500 (67.5% deuterium incorporation), at least 5000 (75%o deuterium), at least 5500 (82.5% deuterium incorporation), at least 6000 (90%) deuterium incorporation), at least 6333.3 (95%> deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5%> deuterium incorporation).
  • any atom not specifically designated as a particular isotope is meant to represent any stable isotope of that atom.
  • a position is designated specifically as “H” or “hydrogen”, the position is understood to have hydrogen at its natural abundance isotopic composition. Also unless otherwise stated, when a position is designated
  • the position is understood to have deuterium at an abundance that is at least 3340 times greater than the natural abundance of deuterium, which is 0.015% (i.e., at least 50.1% incorporation of deuterium).
  • isotopologue refers to a species that differs from a specific compound of this invention only in the isotopic composition thereof.
  • a compound represented by a particular chemical structure containing indicated deuterium atoms will also contain lesser amounts of isotopologues having hydrogen atoms at one or more of the designated deuterium positions in that structure.
  • the relative amount of such isotopologues in a compound of this invention will depend upon a number of factors including the isotopic purity of deuterated reagents used to make the compound and the efficiency of incorporation of deuterium in the various synthesis steps used to prepare the compound. However, as set forth above, the relative amount of such isotopologues in toto will be less than 49.9% of the compound.
  • the invention also provides salts of the compounds of the invention.
  • a salt of a compound of this invention is formed between an acid and a basic group of the compound, such as an amino functional group, or a base and an acidic group of the compound, such as a carboxyl functional group.
  • the compound is a pharmaceutically acceptable acid addition salt.
  • pharmaceutically acceptable refers to a component that is, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and other mammals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.
  • pharmaceutically acceptable salt means any non-toxic salt that, upon administration to a recipient, is capable of providing, either directly or indirectly, a compound of this invention.
  • pharmaceutically acceptable counterion is an ionic portion of a salt that is not toxic when released from the salt upon administration to a recipient.
  • Acids commonly employed to form pharmaceutically acceptable salts include inorganic acids such as hydrogen sulfide, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid and phosphoric acid, as well as organic acids such as para-toluenesulfonic acid, salicylic acid, tartaric acid, bitartaric acid, ascorbic acid, maleic acid, besylic acid, fumaric acid, gluconic acid, glucuronic acid, formic acid, glutamic acid, methanesulfonic acid, ethanesulfonic acid,
  • Such pharmaceutically acceptable salts thus include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate,
  • pharmaceutically acceptable acid addition salts include those formed with mineral acids such as hydrochloric acid and hydrobromic acid, and especially those formed with organic acids such as maleic acid.
  • the invention also includes solvates and hydrates of the compound of the invention.
  • hydrate means a compound which further includes a stoichiometric or non-stoichiometric amount of water bound by non- covalent intermolecular forces.
  • solvate means a compound which further includes a stoichiometric or non-stoichiometric amount of solvent such as water, acetone, ethanol, methanol, dichloromethane, 2-propanol, or the like, bound by non-covalent intermolecular forces.
  • Formulae A, Al, I and B can be chiral in some instances (when Y , Y and R are different from one another) and in other instances it can be achiral (when at least two
  • chiral compounds of this invention can exist as either individual enantiomers, or as racemic or scalemic mixtures of enantiomers. Accordingly, a compound of the present invention will include racemic and scalemic enantiomeric mixtures, as well as individual respective stereoisomers that are substantially free from another possible stereoisomer.
  • substantially free of other stereoisomers means less than 25% of other stereoisomers, preferably less than 10% of other stereoisomers, more preferably less than 5% of other stereoisomers and most preferably less than 2% of other stereoisomers, or less than "X"% of other stereoisomers (wherein X is a number between 0 and 100, inclusive) are present.
  • stable compounds refers to compounds which possess stability sufficient to allow for their manufacture and which maintain the integrity of the compound for a sufficient period of time to be useful for the purposes detailed herein (e.g., formulation into therapeutic products, intermediates for use in production of therapeutic compounds, isolatable or storable intermediate compounds, treating a disease or condition responsive to therapeutic agents).
  • D refers to deuterium.
  • Stereoisomer refers to both enantiomers and diastereomers.
  • Tet refers to tertiary.
  • US refers to the United States of America.
  • alkylene means a straight or branched chain divalent hydrocarbon radical, preferably having from one to six carbon atoms (Ci_6alkylene). In some embodiments, the alkylene group has from one to four carbon atoms (Ci_ 4 alkylene).
  • alkylene as used herein include, but are not limited to, methylene (-CH 2 -), ethylene (-CH 2 CH 2 -), propylene (-CH 2 CH 2 CH 2 -), and branched versions thereof such as (-CH(CH 3 )-), -CH 2 CH(CH 3 )- and the like.
  • Halo means chloro, bromo, fluoro, or iodo.
  • Alkyl means an aliphatic hydrocarbon group which may be straight or branched having 1 to 15 carbon atoms in the chain. Preferred alkyl groups have 1 to 12 carbon atoms in the chain, and more preferably 1 to 6 carbon atoms. Branched means that one or more lower alkyl groups such as methyl, ethyl or propyl are attached to a linear alkyl chain. “Lower alkyl” means about 1 to about 4 carbon atoms in the chain which may be straight or branched.
  • Exemplary alkyl groups include methyl, fluoromethyl, difluoromethyl, trifluoromethyl, cyclopropylmethyl, cyclopentylmethyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, n-pentyl, 3-pentyl, heptyl, octyl, nonyl, decyl and dodecyl; preferred are methyl, difluoromethyl and i- propyl.
  • Alkyl groups may be optionally substituted with one or more groups selected from halo, cyano, hydroxyl, carboxy, alkoxy, alkoxycarbonyl, oxo, amino, alkylamino, dialkylamino, cycloheteroalkyl, alkylcycloheteroalkyl, aryl, alkylaryl, heteroaryl, and alkylheteroaryl.
  • any alkyl or alkoxy moiety of the alkyl substituent group has 1 to 6 carbon atoms.
  • Aryl means an aromatic carbocyclic radical containing 6 to 10 carbon atoms.
  • exemplary aryl groups include phenyl or naphthyl.
  • Aryl groups may be optionally substituted with one or more groups which may be the same or different, and which are selected from alkyl, aryl, aralkyl, alkoxy, aryloxy, aralkyloxy, halo, and nitro.
  • any alkyl or alkoxy moiety of the aryl substituent group has 1 to 6 carbon atoms.
  • Heteroaryl means a 5- to a 10-membered aromatic monocyclic or multicyclic hydrocarbon ring system in which one or more of the carbon atoms in the ring system is or are element(s) other than carbon, for example nitrogen, oxygen or sulfur. Heteroaryl groups may be optionally substituted with one or more groups which may be the same or different, and which are selected from alkyl, aryl, aralkyl, alkoxy, aryloxy, aralkyloxy, halo, and nitro.
  • heteroaryl groups include pyrazinyl, furanyl, thienyl, pyridyl, pyrimidinyl, isoxazolyl, isothiazolyl, pyridazinyl, 1,2,4-triazinyl, quinolinyl, and isoquinolinyl.
  • Alkyl means an aryl-alkyl group in which the aryl and alkyl components are as previously described. Preferred aralkyls contain a lower alkyl moiety.
  • Exemplary aralkyl groups include benzyl and 2-phenethyl.
  • Heteroaralkyl means a heteroaryl-alkyl group in which the heteroaryl and alkyl components are as previously described.
  • Cycloalkyl means a non-aromatic mono-, multicyclic, or bridged ring system of 3 to 10 carbon atoms.
  • the cycloalkyl group is optionally substituted by one or more halo, or alkyl.
  • Exemplary monocyclic cycloalkyl rings include cyclopentyl, fluorocyclopentyl, cyclohexyl and cycloheptyl.
  • Heterocycloalkyl means a non-aromatic mono-, bi- or tricyclic, or bridged hydrocarbon ring system in which one or more of the atoms in the ring system is or are element(s) other than carbon, for example nitrogen, oxygen or sulfur.
  • Preferred heterocycloalkyl groups contain rings with a ring size of 3-6 ring atoms.
  • Exemplary heterocycloalkyl groups pyrrolidine, piperidine, tetrahydropyran, tetrahydrofuran, tetrahydrothiopyran, and tetrahydrothiofuran.
  • Cycloalkylalkyl means a group in which the cycloalkyl and alkyl components are as previously described.
  • Heteroycloalkylalkyl means a group in which the cycloalkyl and alkyl components are as previously described.
  • variable may be referred to generally
  • each R may be referred to specifically (e.g., R , R , R , etc.). Unless otherwise indicated, when a variable is referred to generally, it is meant to include all specific embodiments of that particular variable.
  • the resent invention provides a compound of Formula A:
  • R 1 and R 2" are each independently selected from hydrogen, -(Ci-C 4 )alkyl, or - (Ci-C 4 )alkylene-0-(Ci-C 2 )alkyl, wherein the alkyl and alkylene groups at each instance are independently and optionally substituted with deuterium;
  • R 3 is selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R 4 is n-butylene optionally substituted with deuterium
  • R 5 is selected from hydrogen, deuterium, alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, heterocycloalkylalkyl, aryl, and heteroaryl, wherein each of the alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, heterocycloalkylalkyl, aryl, and heteroaryl is optionally substituted and wherein one or more hydrogen atoms in the alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, heterocycloalkylalkyl, aryl, or heteroaryl or optional substituent thereof is optionally replaced with a corresponding number of deuterium atoms; and
  • the compound of Formula A is other than the following:
  • R 1 and R 2 are each methyl optionally substituted with deuterium and R 5 is hydrogen or deuterium, then either: (i) Y 1 is fiuoro ; or (ii) Y 1 is OH, and Y 2 is selected from -CH 3 , -CH 2 D, -CHD 2 and -CD . In one aspect of this embodiment, the compound is not
  • At least one of Y ⁇ R , R ⁇ R and R 4 bears at least one deuterium atom.
  • R 1 and R 2 are each methyl optionally substituted with deuterium;
  • R 5 is hydrogen or deuterium; and either: (a)
  • Y 1 and Y 2" are taken together with the carbon atom to which they are bound to form
  • Y 1 is -OH and Y 2 is selected from hydrogen and deuterium, with the provisos that:
  • R 5 is D, the compound having
  • Y and Y" are as defined for Formula A.
  • R 1 and R 2 are each independently selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R 3 is selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R 4 is selected from -(CH 2 ) 4 -, -(CD 2 ) 4 -, ⁇ - (CD 2 ) 3 CH 2 , and ⁇ -CD 2 (CH 2 ) 3 -, wherein
  • R 1 and R 2 are each independently selected from -CH 3 and -CD 3 ;
  • R 3 is selected from -CH 3 and -CD 3 ;
  • R and R are each independently selected from -CH 3 and -CD 3 ;
  • R 3 is selected from -CH 3 and -CD 3 ;
  • R 4 is selected from
  • the present invention provides a compound of Formula A, wherein R 5 is hydrogen, the compound having Formula I: (I), or a salt thereof, wherein:
  • R 1 and R 2" are each independently selected from hydrogen, -(Ci-C4)alkyl, or - (Ci-C4)alkylene-0-(Ci-C2)alkyl, wherein the alkyl and alkylene groups at each instance are independently and optionally substituted with deuterium;
  • R 3 is selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R 4 is n-butylene optionally substituted with deuterium
  • R 1 and R 2 are each
  • R is selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R 4 is selected from -(CH 2 ) 4 -, -(CD 2 ) 4 -, ⁇ - (CD 2 ) CH 2 , and ⁇ -CD 2 (CH 2 ) -, wherein " ⁇ " represents the portion of the R 4 moiety
  • R and R are each independently selected from -CH and -CD ;
  • R 3 is selected from -CH and -CD ;
  • R 4 is selected from
  • R and R are each independently selected from -CH and -CD ;
  • R 3 is selected from -CH and -CD ;
  • R 4 is selected from
  • the compound of Formula I is other than the following:
  • the compound of Formula I is other than the following:
  • the compound of Formula I is other than the following:
  • R 1 and R 2" are each independently selected from hydrogen, -(Ci-C4)alkyl, or - (Ci-C4)alkylene-0-(Ci-C2)alkyl, wherein the alkyl and alkylene groups at each instance are independently and optionally substituted with deuterium;
  • R 3 is selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R 4 is n-butylene optionally substituted with deuterium
  • R , R and R bears at least one deuterium atom.
  • One embodiment relates to a compound of Formula A, A 1 , 1, or II, wherein
  • R" and R J are each independently selected from -CH 3 , -CH 2 D, -CHD 2 and -CD 3 .
  • Another embodiment relates to a compound of Formula A, A 1 , 1, or II,
  • R and R are each independently selected from -CH 3 , and -CD 3 .
  • Another embodiment relates to a compound of Formula A, A 1 , 1, or II, wherein R 1 is selected from hydrogen, (Ci-C 3 )alkyl, and (Ci-C 2 )alkylene-0(Ci- C 2 )alkyl.
  • Another embodiment relates to a compound of Formula A, A 1 , 1, or II, wherein R 1 is hydrogen, -CH 3 , -CD 3 , -CH 2 CH 2 CH 3 , -CD 2 CH 2 CH 3 , -CD 2 CD 2 CH 3 , - CD 2 CD 2 CD 3 , -CH 2 OCH 2 CH 3 , -CH 2 OCD 2 CH 3 , -CH 2 OCD 2 CD 3 , -CD 2 OCH 2 CH 3 , - CD 2 OCD 2 CH 3 , or -CD 2 OCD 2 CD 3 .
  • R 5 is selected from hydrogen, deuterium, alkyl, cycloalkyl, heterocycloalkyl,
  • cycloalkylalkyl, and heterocycloalkylalkyl wherein each of alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, and heterocycloalkylalkyl may be optionally substituted.
  • each methylene unit in R 4 is selected from -CH 2 - and -CD 2 -; more
  • R is selected from -(CH 2 )4-, -(CD 2 ) 4 -, '-CD 2 (CH 2 )3- and
  • Y 1 is F
  • Y 2 is selected from hydrogen, -CH 3 , -CH 2 D, -CHD 2 and -CD 3 ;
  • R is -CD 3 ; R and R are
  • R is selected from -(CH 2 ) 4 -, -(CD 2 ) 4 -, ⁇ -CD 2 (CH 2 ) 3 - and ⁇ -(CD 2 ) 3 CH 2 -.
  • R is -CD 3 ; R and R are
  • R is selected from -(CH 2 ) 4 -, and -(CD 2 ) 4 -.
  • R is -CD 3 ; R and R are -
  • R is -(CH 2 ) 4 -; Y is fluoro; and Y is selected from deuterium, -CH 2 D, -CHD 2 and -CD 3 .
  • R is -CD 3 ; R and R are - CH 3 ; R 4 is -(CH 2 ) 4 -; Y 1 is fluoro; and Y 2 is fluorine.
  • R is -CD 3 ; R and R are -CH 3 ; R 4 is -(CH 2 ) 4 -; R 5 is deuterium; Y 1 is fluoro; and Y 2 is selected from deuterium, -CH 2 D, -CHD 2 and -CD 3 .
  • R is -CD 3 ; R and R are -CH 3 ; R 4 is -(CH 2 ) 4 -; R 5 is deuterium; Y 1 is fluoro; and Y 2 is fluorine.
  • Y is F; Y is selected from hydrogen; R 3 is -CH 3 ; and R 4 is -(CH 2 ) 4 -.
  • Y is F; Y is fluorine; R is - CH 3 ; and R 4 is -(CH 2 ) 4 -.
  • One embodiment provides a compound of Formula B:
  • One embodiment provides a compound of Formula B, wherein each Z 3 , Z 4 5 1 2 5 and Z is hydrogen.
  • R and R are each -CD 3 .
  • R is deuterium.
  • Y 1 and Y 2 are taken together with the carbon to which
  • each Z , Z 4 and Z 5 is deuterium.
  • R 1 and R 2 are each -CD 3 .
  • R 5 is deuterium.
  • Y 1 and is OH, and Y 2 is hydrogen or deuterium.
  • R 1 and R 2 are each -CD 3 .
  • R 5 is deuterium.
  • each Z , Z and Z 5 is hydrogen and R 5 is deuterium.
  • each Z 3 , Z 4 and Z 5 is deuterium and R 5 is deuterium.
  • R 5 is deuterium.
  • each Z 3 , Z 4 and Z 5 is hydrogen and R 5 is deuterium.
  • each Z 3 , Z 4 and Z 5 is deuterium and R 5 is deuterium.
  • R 1 and R2 are each 1
  • R and R2 are each R 5 is deuterium. In another aspect, R 1
  • -CD 3 and and R2 are each -CD 3 , and each Z 3 , Z 4 and Z 5 is deuterium.
  • R 1 and R 2 are each -CD 3 , each Z 3 , Z 4 and Z 5 is deuterium and R 5 is deuterium.
  • R 1 and R 2 are each -CD 3 , and each Z 3 , Z 4 and Z 5 is hydrogen.
  • R 1 and R 2 are each -CD 3 , each Z 3 , Z 4 and Z 5 is hydrogen and R 5 is deuterium
  • a still further embodiment provides a compound of Formula B, Y 1 and is
  • OH, and Y 2 is hydrogen or deuterium.
  • R 5 is deuterium.
  • each Z 3 , Z 4 and Z 5 is hydrogen and R 5 is deuterium.
  • each Z 3 , Z 4 and Z 5 is deuterium and R 5 is deuterium.
  • R 1 and R 2 are each R 1 and R2 are each 5
  • -CD 3 In another aspect, -CD 3 and R is deuterium. In another aspect, R 1 and R 2 are each -CD 3 , and each Z 3 , Z 4 and Z 5 is deuterium. In another aspect, R 1 and R 2 are each -CD 3 , each Z 3 , Z 4 and Z 5 is deuterium and R 5 is deuterium. In another aspect, R 1 and R 2 are each -CD 3 , and each Z 3 , Z 4 and Z 5 is hydrogen. In another aspect, R 1 and R 2 are each -CD 3 , each Z 3 , Z 4 and Z 5 is hydrogen and R 5 is deuterium
  • Another embodiment provides a compound of Formula B, wherein R 5 is deuterium.
  • Another embodiment provides a compound of Formula B, wherein R 5 is deuterium, Z 3 , Z 4 and Z 5 is hydrogen and R 1 is -CD 3 .
  • containing a chiral carbon atom bound to Y and Y are intended to represent a racemic mixture of enantiomers.
  • Table 1 above shows examples of specific compounds of Formula I. These examples are deuterated and/or fluorinated analogs of pentoxifylline and its metabolites.
  • Table 2 above shows examples of specific compounds of Formula I where R is H and Y is CH 3 or CD 3 . These compounds include deuterated and fluorinated analogs of Albifylline (HWA-138). Albifylline has been studied for uses that are associated with pentoxifylline.
  • CD 261 CD 2 OCD 2 CD 3 CD 3 CD 3 T CD 2 (CH 2 ) 3 OH CD 3
  • Table 3 above shows examples of specific compounds of Formula I where R is -CH 2 -0-CH 2 CH 3 , optionally substituted with deuterium.
  • Y 1 is OH or F and Y is CH 3 or CD 3 .
  • These compounds include deuterated and fluorinated analogs of torbafylline (HWA-448). Torbafylline has been studied for the treatment of depression, urinary incontinence, irritable bowel syndrome and multiple sclerosis.
  • CD2CD2CD3 CD 3 CD 3 (CD 2 ) 4 OH CD 3
  • CD2CD2CD3 CD 3 CD 3 (CD 2 ) 4 F CD 3
  • Table 4 above shows examples of specific compounds of Formula I where R is -CH 2 CH 2 CH 3 optionally substituted with deuterium.
  • Y 1 is OH or F and Y is CH 3 or CD 3 .
  • These compounds include deuterated and fluorinated analogs of A-802715.
  • A-802715 has been studied for the treatment of septic shock and inhibition of effects of allograft reaction.
  • Table 5 above shows examples of specific compounds of Formula I where R is -CH 2 CH 2 CH 3 optionally substituted with deuterium.
  • Y 1 and Y are taken together with their intervening carbon to form a carbonyl.
  • These compounds include deuterated analogs of propentofylline.
  • Propentofylline has been studied for the treatment of Alzheimer's disease, neuropathic pain, traumatic brain injury, dysuria, retinal or optic nerve head damage, and peptic ulcers. It has also been studied for controlling intraocular pressure, stabilization of auto-regulation of cerebral blood flow and inhibition of effects of allograft reaction.
  • Table 6 above shows examples of specific compounds of Formula A. These examples are deuterated and/or fluorinated analogs of pentoxifylline and its metabolites where R 5 is deuterium.
  • the compound is not any one of Compounds 100, 116, or 149.
  • Examples of specific compounds of this invention include the following:
  • the present invention provides a compound of Formula C
  • R is selected from
  • R is selected from -CH 3 and -CD 3 ; one of Y and Y is -OH; and the other of Y 1 and Y 2 is deuterium or hydrogen ⁇
  • One embodiment provides a compound of Formula C, wherein R 1 is -CH 3 .
  • One embodiment provides a compound of Formula C, wherein R 1 is -CD 3 .
  • One embodiment provides a compound of Formula C, wherein R is -CH 3 .
  • One embodiment provides a compound of Formula C, wherein R is -CD 3 .
  • One embodiment provides a compound of Formula C, wherein R 1 is -CH 3 and R 2 is -CH 3 .
  • Y 1 is -OH.
  • Y2 is OH.
  • One embodiment provides a compound of Formula C, or a pharmaceutically acceptable salt thereof, wherein the compound has the structure:
  • Y is deuterium. In another aspect, Y is hydrogen.
  • One embodiment provides a compound of Formula C, or a pharmaceutically acce table salt thereof, wherein the compound has the structure:
  • Y 2 is deuterium. In another aspect, Y 2 is hydrogen.
  • Examples of the compounds of the formula C include the following compounds and pharmaceutically acceptable salts thereof:
  • the present invention also provides a compound of Formula D:
  • R is selected from
  • R is selected from -CH 3 and -CD 3 ; one of Y and Y is -OH; and the other of Y 1 and Y 2 is deuterium or hydrogen;
  • One embodiment provides a compound of Formula D, wherein R 1 is -CH 3 .
  • One embodiment provides a compound of Formula D, wherein R 1 is -CD 3 .
  • One embodiment provides a compound of Formula D, wherein R is -CH 3 .
  • One embodiment provides a compound of Formula D, wherein R is -CD 3 .
  • One embodiment provides a compound of Formula D, wherein R 1 is -CH 3 and R 2 is -CH 3 .
  • Y 1 is -OH.
  • Y2 is OH.
  • One embodiment provides a compound of Formula D, or a pharmaceutically acce table salt thereof, wherein the compound has the structure:
  • Y is deuterium. In another aspect, Y is hydrogen.
  • One embodiment provides a compound of Formula D, or a pharmaceutically acce table salt thereof, wherein the compound has the structure:
  • Y 2 is deuterium. In another aspect, Y 2 is hydrogen.
  • Examples of the compounds of the formula D include the following compounds and pharmaceutically acceptable salts thereof:
  • any atom not designated as deuterium in any of the embodiments set forth above is present at its natural isotopic abundance.
  • Such methods can be carried out utilizing corresponding deuterated and optionally, other isotope-containing reagents and/or intermediates to synthesize the compounds delineated herein, or invoking standard synthetic protocols known in the art for introducing isotopic atoms to a chemical structure.
  • deuterated compound 10 is alkylated with deuterated intermediate 11 (wherein X is chloride, bromide or iodide) in the presence of potassium carbonate to afford compounds of Formula I.
  • deuterated intermediate 11 wherein X is chloride, bromide or iodide
  • sodium hydroxide in aqueous methanol may be employed to afford compounds of Formula I according to the methods of US Patent 4289776.
  • compounds of Formula II can be used to make compounds where Y 1 is OH.
  • compounds of Formula II are reduced with either sodium borohydride or sodium borodeuteride (commercially available at 99 atom %D) according to the general method of European Patent publication 0330031 to form compounds wherein Y 1 is OH and Y 2 is hydrogen or deuterium.
  • the enantiomeric alcohol products may be separated, for example through the method of Nicklasson, M et al, Chirality, 2002, 14(8): 643-652.
  • enzymatic reduction affords an enantiomerically-enriched alcohol product using the methods disclosed in Pekala, E et al., Acta Bote Pharmaceutica, 2007, 64(2): 109-113, or in Pekala, E et al, Biotech J, 2007, 2(4): 492-496. 1 2
  • ketoreductase or carbonyl reductase at an appropriate pH with an enantiomeric excess of at least 80% may be, for example, any one of ALMAC Carbonyl Reductases CRED A131 , CRED A801 , CRED A901 , CRED A251 , or CRED A271 (each
  • the enzyme is selected from CRED A131 , CRED A251 , KRED-NADH 101 , KRED-NADH 102, KRED-NADH 1 12, KRED-NADH 126, ES-KRED-121 , ES-KRED-128, ES- KRED-130, ES-KRED-142, ES-KRED-169, or ES-KRED-171.
  • the enzyme is selected from CRED A131 , CRED A251 , and KRED-NADH 101.
  • the ketoreductase or carbonyl reductase that favors formation of a compound wherein the stereochemistry at the C(H)OH or C(D)OH group is (R) may be, for example, any one of KRED-NADP-1 18, CRED A601-NADP, CRED A291-NADP, CRED A31 1-NADP. KRED- AD-1 10, ES-KRED-120, ES CRED-I 31 , CRED A 1G1-NADP.
  • the amount of ketoreductase or carbonyl reductase used in the reaction ranges from 0.05 wt% to 10 wt% as a percentage of the weight of the substrate, such as 0.5 wt% to 5 wt%.
  • the amount of enzyme is between 1.0 wt% and 2.0 wt%. In a more specific aspect, the amount of enzyme is about 1.0 wt%.
  • the hydride source is a compound or mixture that is capable of providing a hydride anion or a synthon of a hydride anion.
  • the deuteride source is a compound or mixture that is capable of providing a deuteride anion or a synthon of a deuteride anion.
  • a hydride or deuteride source comprises a catalytic co-factor and optionally, a co-factor regeneration system.
  • a catalytic co-factor used with the ketone reductase or carbonyl reductase in the process of this invention is selected from NAD, NADP, NADH, NADPH, NAD 2 H and NADP 2 H.
  • the choice of co-factor may be based upon (a) the presence or absence of a co-factor regeneration system; (b) the requirement for a hydride versus a deuteride source; and (c) an appropriate pH to perform the method according to the present invention means buffer conditions that maintain the pH at between 6.0 and 7.5 throughout the reaction.
  • the pH of the reaction was maintained at between 6.5 and 7.3.
  • the pH of the reaction was maintained between 6.0 and 7.0.
  • dropwise addition of KOH is used to maintain the desired pH because the enzymatic reaction generates acid.
  • the pH of the reaction is maintained between 6.90 and 7.05.
  • the process may be performed at a temperature of about 20° C to 37° C.
  • the temperature is about 29° C to 32° C.
  • the process may be performed over a time period of about 12 hours to about 24 hours. In one embodiment, the time period is about 24 hours to about 40 hours. In one embodiment, the time period is about 40 hours to about 72 hours. In one embodiment, the time period is a time period sufficient for less than about 5% of
  • CH 3 and R" is -CD 3 ; and (c) R and R" are -CD 3 are all known. See Benchekroun, Y et al, J Chromatogr B, 1977, 688: 245; Ribon, B et al, Coll INSERM, 1988, 164: 268; and Horning, MG et al, Proc Int Conf Stable Isot 2 nd , 1976, 41-54. 3-Methyl-
  • N-alkylurea 13 A synthesis of compound 10 is depicted in Scheme 2 starting with commercially-available N-nitroso-N-methylurea.
  • Urea 13 may be treated with 2-cyanoacetic acid and acetic anhydride to provide cyanoacetamide derivative 14, which is treated first with aqueous NaOH and then with aqueous HCl to provide cyclized pyrimidinedione 15 according to the methods of Dubey, PK et al., Indian Journal of Heterocyclic Chemistry, 2005, 14(4): 301-306.
  • cyanoacetamide 14 may be treated with trimethylsilylchloride and
  • useful deuterated amine reagents 12 include, but are not limited to, commercially-available compounds such as n-propyl-d7-amine, or known compounds such as l-propan-l,l-d2-amine (Moritz, F et al, Organic Mass
  • Useful deuterated urea reagents 13 may include, but are not limited to, commercially-available compounds such as N-
  • Useful deuterated electrophiles 18 may include, but are not limited to,
  • the alcohol moiety in 21 is converted to the chloride upon treatment with triphenylphosphine and carbon tetrachloride to yield chloride lla-i/ 5 following the general procedures of Clement, J-L, Org Biomol Chem, 2003, 1 : 1591-1597.
  • Scheme 4 depicts a synthesis of compound lla-i 9 and compound lla-i 77.
  • commercially-available 4-phenylbutyric acid 22 may be heated in D 2 0 (99 atom %D) in the presence of Pd/C and hydrogen gas to afford deuterated acid 23 according to the general methods of Esaki, et al, Chem Eur J, 2007, 13: 4052-4063.
  • Addition of deuterated methyllithium in the presence of trimethylsilyl chloride provides ketone 24, according to the general method of Porta, A et al., J Org Chem, 2005, 70(12): 4876-4878.
  • Ketone 24 is converted to acetal 25 by treatment with D2SO4 (99 atom %D) and commercially-available ethyleneglycol- ⁇ (99 atom %D).
  • Treatment of 25 with NaI0 4 and RuCl 3 according to the general method of Gamier, J-M et al., Tetrahedron: Asymmetry, 2007, 18(12): 1434-1442 provides carboxylic acid 26.
  • Y is fluorine; Y is selected from hydrogen and deuterium;
  • a deuterated (or nondeuterated) benzyl-protected alcohol 27 such as known [[[(5R)-5-fluorohexyl]oxy]methyl]-benzene (PCT publication WO2000031003) is deprotected by hydrogenation in the presence of Pd/C to provide alcohol 28.
  • the alcohol is chlorinated with thionyl chloride according to the general procedure of Lacan, G et al., J Label Compd Radiopharm, 2005, 48(9): 635-643 to afford chloride llb-(R).
  • a deuterated (or nondeuterated) alcohol 29, such as known (S)-(+)-5-fluorohexanol (Riswoko, A et al, Enantiomer, 2002, 7(1): 33-39) is chlorinated to afford chloride llb-(S).
  • Scheme 5 depicts a synthesis of other intermediates 11c and lie.
  • DAST diethylaminosulfur trifluoride
  • halides can be used to make compounds 11 as disclosed in Scheme 5.
  • commercially-available 5-chlorovaleryl chloride, or commercially-available 5-bromovaleryl chloride, or commercially- available ethyl 5-bromovalerate may be useful as reagents 30 or 31.
  • Grignard reagent 32 affords electrophile 11 wherein R and Y are simultaneously CD 3 .
  • commercially-available 4-hydroxy-butanoic acid ethyl ester 37 is treated with DHP and CSA, or with DHP, TsOH, and pyridine to provide ester 38.
  • Scheme 8 depicts the synthesis of intermediate lle-i n R 3 and Y 2
  • THF- ⁇ 3 ⁇ 4 41 may be treated with DCl and ZnCl 2 according to the general methods of Yang, A et al., Huagong Shikan, 2002, 16(3): 37-39 to afford known chloride 42 (Aiken, Rudolf-Giesbert, WO 2003080598A1). Following the same methods as in Scheme 6, chloride 42 may be converted to l l -dg.
  • Scheme 10 depicts a preparation of llc-i/ 2 , wherein R and Y are the same.
  • deuterated ester 46 (Feldman, KS et al, Journal of Organic Chemistry, 2000, 65(25): 8659-8668) is treated with carbon tetrabromide and
  • Scheme 11 depicts the synthesis of a compound of Formula Al .
  • a compound of Formula I is treated with potassium carbonate in D 2 0 to effect a hydrogen-to-deuterium exchange reaction, providing a compound of Formula Al .
  • additional hydrogen-to-deuterium exchange reactions may also occur elsewhere in the molecule.
  • Y is selected from fluorine and OH; and Y is selected from hydrogen, deuterium, -CH 3 , -CH 2 D, -CHD 2 and -CD 3 .
  • a number of novel intermediates can be used to prepare compounds of Formula A.
  • the invention also provides such a compound which is selected from the following:
  • R 1 and R 2 are independently selected from hydrogen, deuterium, Ci_ 3 alkyl optionally substituted with deuterium, and Ci_ 3 alkoxyalkyl optionally substituted with deuterium.
  • R 1 and R 2 Ci_ 3 alkyl include -CH 3 , -CD 3 , -CH2CH 2 CH 3 , and -CD 2 CD 2 CD 3 .
  • Ci_ 3 alkoxyalkyl include -CH 2 OCH 2 CH 3 , - CD 2 OCH 2 CH 3 , -CD 2 OCD 2 CH 3 , and -CD 2 OCD 2 CD 3 .
  • formula III includes the following: CH,
  • III-a Ill-b, III-c,
  • W is hydrogen.
  • W is deuterium.
  • Salts of compounds of Formula III are also useful, including salts that are known to be useful with respect to known xanthines.
  • useful salts include, but are not limited to, the lithium salt, sodium salt, potassium salt, and cesium salt.
  • An example of a particularly useful salt is the potassium salt.
  • the invention also provides pyrogen-free compositions comprising an effective amount of a compound of this invention or pharmaceutically acceptable salts thereof; and an acceptable carrier.
  • a composition of this invention is formulated for pharmaceutical use ("a pharmaceutical composition"), wherein the carrier is a pharmaceutically acceptable carrier.
  • the carrier(s) are "acceptable” in the sense of being compatible with the other ingredients of the formulation and, in the case of a pharmaceutically acceptable carrier, not deleterious to the recipient thereof in an amount used in the medicament.
  • Pharmaceutically acceptable carriers, adjuvants and vehicles that may be used in the pharmaceutical compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, microcrystalline cellulose, cellulose- based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene -polyoxypropylene-block polymers, polyethylene glycol and wool fat.
  • ion exchangers alumina, aluminum stearate, lecithin
  • serum proteins such as human serum albumin
  • solubility and bioavailability of the compounds of the present invention in pharmaceutical compositions may be enhanced by methods well-known in the art.
  • One method includes the use of lipid excipients in the formulation. See “Oral Lipid-Based Formulations: Enhancing the Bioavailability of Poorly Water- Soluble Drugs (Drugs and the Pharmaceutical Sciences),” David J. Hauss, ed.
  • Another known method of enhancing bioavailability is the use of an amorphous form of a compound of this invention optionally formulated with a poloxamer, such as LUTROLTM and PLURONICTM (BASF Corporation), or block copolymers of ethylene oxide and propylene oxide. See United States patent 7,014,866; and United States patent publications 20060094744 and 20060079502.
  • compositions of the invention include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal)
  • the compound of the formulae herein is administered transdermally (e.g., using a transdermal patch or iontophoretic techniques).
  • Other formulations may conveniently be presented in unit dosage form, e.g., tablets, sustained release capsules, and in liposomes, and may be prepared by any methods well known in the art of pharmacy. See, for example, Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, PA (17th ed. 1985).
  • Such preparative methods include the step of bringing into association with the molecule to be administered ingredients such as the carrier that constitutes one or more accessory ingredients.
  • ingredients such as the carrier that constitutes one or more accessory ingredients.
  • the compositions are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers, liposomes or finely divided solid carriers, or both, and then, if necessary, shaping the product.
  • compositions of the present invention suitable for oral administration may be presented as discrete units such as capsules, sachets, or tablets each containing a predetermined amount of the active ingredient; a powder or granules; a solution or a suspension in an aqueous liquid or a non-aqueous liquid; an oil-in-water liquid emulsion; a water-in-oil liquid emulsion; packed in liposomes; or as a bolus, etc.
  • Soft gelatin capsules can be useful for containing such suspensions, which may beneficially increase the rate of compound absorption.
  • carriers that are commonly used include lactose and corn starch.
  • Lubricating agents such as magnesium stearate, are also typically added.
  • useful diluents include lactose and dried cornstarch.
  • aqueous suspensions are administered orally, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening and/or flavoring and/or coloring agents may be added.
  • compositions suitable for oral administration include lozenges comprising the ingredients in a flavored basis, usually sucrose and acacia or tragacanth; and pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia.
  • compositions suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.
  • the formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampules and vials, and may be stored in a freeze dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use.
  • Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets.
  • Such injection solutions may be in the form, for example, of a sterile injectable aqueous or oleaginous suspension.
  • This suspension may be formulated according to techniques known in the art using suitable dispersing or wetting agents (such as, for example, Tween 80) and suspending agents.
  • 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.
  • the acceptable vehicles and solvents that may be employed are mannitol, water, Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oil may be employed including synthetic mono- or diglycerides.
  • Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically- acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions.
  • These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant.
  • compositions of this invention may be administered in the form of suppositories for rectal administration.
  • These compositions can be prepared by mixing a compound of this invention with a suitable non-irritating excipient which is solid at room temperature but liquid at the rectal temperature and therefore will melt in the rectum to release the active components.
  • suitable non-irritating excipient include, but are not limited to, cocoa butter, beeswax and polyethylene glycols.
  • compositions of this invention may be administered by nasal aerosol or inhalation.
  • Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other solubilizing or dispersing agents known in the art. See, e.g.: Rabinowitz, JD and Zaffaroni, AC, US Patent 6,803,031, assigned to Alexza Molecular Delivery Corporation.
  • Topical administration of the pharmaceutical compositions of this invention is especially useful when the desired treatment involves areas or organs readily accessible by topical application.
  • the pharmaceutical composition should be formulated with a suitable ointment containing the active components suspended or dissolved in a carrier.
  • Carriers for topical administration of the compounds of this invention include, but are not limited to, mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene polyoxypropylene compound, emulsifying wax, and water.
  • the pharmaceutical composition can be formulated with a suitable lotion or cream containing the active compound suspended or dissolved in a carrier.
  • suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol, and water.
  • the pharmaceutical compositions of this invention may also be topically applied to the lower intestinal tract by rectal suppository formulation or in a suitable enema formulation. Topically-transdermal patches and iontophoretic administration are also included in this invention.
  • Application of the subject therapeutics may be local, so as to be administered at the site of interest.
  • Various techniques can be used for providing the subject compositions at the site of interest, such as injection, use of catheters, trocars, projectiles, pluronic gel, stents, sustained drug release polymers or other device which provides for internal access.
  • the compounds of this invention may be incorporated into compositions for coating an implantable medical device, such as prostheses, artificial valves, vascular grafts, stents, or catheters.
  • an implantable medical device such as prostheses, artificial valves, vascular grafts, stents, or catheters.
  • Suitable coatings and the general preparation of coated implantable devices are known in the art and are exemplified in US Patents 6,099,562; 5,886,026; and 5,304,121.
  • the coatings are typically biocompatible polymeric materials such as a hydrogel polymer, polymethyldisiloxane, polycaprolactone, polyethylene glycol, polylactic acid, ethylene vinyl acetate, and mixtures thereof.
  • the coatings may optionally be further covered by a suitable topcoat of fluorosilicone, polysaccharides,
  • Coatings for invasive devices are to be included within the definition of pharmaceutically acceptable carrier, adjuvant or vehicle, as those terms are used herein.
  • the invention provides a method of coating an implantable medical device comprising the step of contacting said device with the coating composition described above. It will be obvious to those skilled in the art that the coating of the device will occur prior to implantation into a mammal.
  • the invention provides a method of impregnating an implantable drug release device comprising the step of contacting said drug release device with a compound or composition of this invention.
  • Implantable drug release devices include, but are not limited to, biodegradable polymer capsules or bullets, non-degradable, diffusible polymer capsules and biodegradable polymer wafers.
  • the invention provides an implantable medical device coated with a compound or a composition comprising a compound of this invention, such that said compound is therapeutically active.
  • the invention provides an implantable drug release device impregnated with or containing a compound or a composition comprising a compound of this invention, such that said compound is released from said device and is therapeutically active.
  • composition of this invention may be painted onto the organ, or a composition of this invention may be applied in any other convenient way.
  • the compound of the inveniton comprises between 28 and 68% (w/w) of the composition.
  • magnesium stearate and microcrystalline cellulose comprise about 2% (w/w) of the composition.
  • a composition of this invention further comprises a second therapeutic agent.
  • the second therapeutic agent may be selected from any compound or therapeutic agent known to have or that demonstrates advantageous properties when administered with a compound having the same mechanism of action as pentoxifylline.
  • Such agents include those indicated as being useful in combination with pentoxifylline, including but not limited to, those described in WO 1997019686, EP 0640342, WO 2003013568, WO 2001032156, WO 2006035418, and WO 1996005838.
  • the second therapeutic agent is an agent useful in the treatment or prevention of a disease or condition selected from peripheral obstructive vascular disease; glomerulonephritis; nephrotic syndrome; nonalcoholic steatohepatitis; Leishmaniasis; cirrhosis; liver failure; Duchenne's muscular dystrophy; late radiation induced injuries; radiation induced lymphedema; radiation-associated necrosis; alcoholic hepatitis; radiation-associated fibrosis; necrotizing enterocolitis in premature neonates; diabetic nephropathy, hypertension-induced renal failure, and other chronic kidney disease; Focal Segmental Glomerulosclerosis; pulmonary sarcoidosis; recurrent aphthous stomatitis; chronic breast pain in breast cancer patients; brain and central nervous system tumors; malnutrition-inflammation- cachexia syndrome; interleukin-1 mediated disease; graft versus host reaction and other allograft reactions; diet-induced fatty liver conditions, ather
  • the compounds of this invention can also be used to control intraocular pressure or to stabilize auto-regulation of cerebral blood flow in subjects who require such control as determined by medical examination.
  • the second therapeutic agent is selected from a- tocopherol and hydroxyurea.
  • the second therapeutic agent is useful in the treatment of diabetes or an associated disorder, and is selected from insulin or insulin analogues, glucagon- like-peptide-1 (GLP-1) receptor agonists, sulfonylurea agents, biguanide agents, alpha-glucosidase inhibitors, PPAR agonists, meglitinide agents, dipeptidyl-peptidase (DPP) IV inhibitors, other phosphodiesterase (PDE1, PDE5, PDE9, PDE10 or PDE1) inhibitors, amylin agonists, CoEnzyme A inhibitors, and antiobesity agents.
  • GLP-1 glucagon- like-peptide-1
  • sulfonylurea agents biguanide agents
  • alpha-glucosidase inhibitors PPAR agonists
  • meglitinide agents alpha-glucosidase inhibitors
  • DPP dipeptidyl-peptidase
  • PDE1 phosphodiesterase
  • insulin examples include, but are not limited to Humulin® (human insulin, rDNA origin), Novolin® (human insulin, rDNA origin), Velosulin® BR (human buffered regular insulin, rDNA origin), Exubera® (human insulin, inhaled), and other forms of inhaled insulin, for instance, as delivered by
  • insulin analogues include, but are not limited to, novarapid, insulin detemir, insulin lispro, insulin glargine, insulin zinc suspension and Lys-Pro insulin.
  • Glucagon-Like -Peptide- 1 receptor agonists include, but are not limited to BIM-51077 (CAS-No. 275371-94-3), EXENATIDE (CAS-No. 141758-74-9), CJC-1131 (CAS-No. 532951 -64-7), LIRAGLUTIDE (CAS-No. 20656-20-2) and ZP-10 (CAS-No. 320367-13-3).
  • sulfonylurea agents include, but are not limited to, TOLBUTAMIDE (CAS- No. 000064-77-7), TOLAZAMIDE (CAS-No. 001156-19- 0), GLIPIZIDE (CAS-No. 029094-61-9), CARBUT AMIDE (CAS-No. 000339-43- 5), GLISOXEPIDE (CAS-No. 025046-79-1), GLISENTIDE (CAS-No. 032797-92- 5), GLIBORNURIDE (CAS-No. 026944-48-9), GLIBENCLAMIDE (CAS-NO. 010238-21 -8), GLIQUIDONE (CAS-No. 033342-05-1), GLIMEPIRIDE (CAS-No. 093479-97-1) and GLICLAZIDE (CAS-No. 021187-98-4).
  • a specific example of a biguanide agent includes, but is not limited to METFORMIN (CAS-No. 000657-24-9).
  • alpha-glucosidase-inhibitors include, but are not limited to ACARBOSE (Cas-No. 056180-94-0), MIGLITOL (CAS-No. 072432-03-2) and VOGLIBOSE (CAS-No. 083480-29-9).
  • PPAR-agonists include, but are not limited to
  • MURAGLITAZAR (CAS-No. 331741 -94-7), ROSIGLITAZONE (CAS-NO.
  • Preferred PPAR- agonists are ROSGLITAZONE and
  • meglitinide agents include, but are not limited to REPAGLINIDE (CAS-No. 135062-02-1 ), NATEGLINIDE (CAS-No. 105816-04- 4) and MITIGLINIDE (CAS-No. 145375-43-5).
  • DPP IV inhibitors include, but are not limited to SITAGLIPTIN (CAS-No. 486460-32-6), SAXAGLIPTIN (CAS-No. 361442-04-8), VILDAGLIPTIN (CAS-No. 274901 -16-5), DENAGLIPTIN (CAS-No. 483369-58- 0), P32/98 (CAS-No. 251572-70-0) and NVP-DPP-728 (CAS-No. 247016-69-9).
  • SITAGLIPTIN CAS-No. 486460-32-6
  • SAXAGLIPTIN CAS-No. 361442-04-8
  • VILDAGLIPTIN CAS-No. 274901 -16-5
  • DENAGLIPTIN CAS-No. 483369-58- 0
  • P32/98 CAS-No. 251572-70-0
  • NVP-DPP-728 CAS-No. 247016-69-9.
  • PDE5 inhibitors include, but are not limited to
  • SILDENAFIL CAS-No. 139755-83-2
  • VARDENAFIL CAS-No. 224785-90-4
  • TADALAFIL CAS-No. 171596-29-5
  • PDE1, PDE9, PDE10 or PDE11 inhibitors which may be usefully employed according to the present invention can be found, for example, in US20020160939, WO2003037432,
  • amylin agonist includes, but is not limited to PRAMLINITIDE (CAS-No. 151126-32-8).
  • Coenzyme A inhibitor includes, but is not limited to ETOMOXIR (CAS- No. 082258-36-4).
  • anti-obesity drugs include, but are not limited to HMR- 1426 (CAS-No. 262376-75-0), CETILISTAT (CAS-No. 282526-98-1) and
  • SIBUTRAMINE (CAS-No. 106650-56-0).
  • the invention provides separate dosage forms of a compound of this invention and one or more of any of the above-described second therapeutic agents, wherein the compound and second therapeutic agent are associated with one another.
  • association with one another means that the separate dosage forms are packaged together or otherwise attached to one another such that it is readily apparent that the separate dosage forms are intended to be sold and administered together (within less than 24 hours of one another, consecutively or simultaneously).
  • the compound of the present invention is present in an effective amount.
  • the term is a pharmaceutical composition of the invention.
  • effective amount refers to an amount which, when administered in a proper dosing regimen, is sufficient to treat (therapeutically or prophylactically) the target disorder. For example, and effective amount is sufficient to reduce or ameliorate the severity, duration or progression of the disorder being treated, prevent the advancement of the disorder being treated, cause the regression of the disorder being treated, or enhance or improve the prophylactic or therapeutic effect(s) of another therapy.
  • an effective amount of a compound of this invention is in the range of 20 mg to 2000 mg per treatment. In more specific embodiments the amount is in the range of 40 mg to 1000 mg, or in the range of 100 mg to 800 mg, or more specifically in the range of 200 mg to 400 mg per treatment. Treatment typically is administered from one to three times daily.
  • Effective doses will also vary, as recognized by those skilled in the art, depending on the diseases treated, the severity of the disease, the route of administration, the sex, age and general health condition of the patient, excipient usage, the possibility of co-usage with other therapeutic treatments such as use of other agents and the judgment of the treating physician. For example, guidance for selecting an effective dose can be determined by reference to the prescribing information for pentoxifylline.
  • compositions that comprise a second therapeutic agent an effective amount of the second therapeutic agent is between about 20% and 100% of the dosage normally utilized in a monotherapy regime using just that agent.
  • an effective amount is between about 70% and 100% of the normal monotherapeutic dose.
  • the normal monotherapeutic dosages of these second therapeutic agents are well known in the art. See, e.g., Wells et al, eds.,
  • the invention provides a method of inhibiting the activity of phosphodiesterase (PDE) in a cell, comprising contacting a cell with one or more compounds of Formula A, Al, I, II, B, C, or D.
  • PDE phosphodiesterase
  • pentoxifylline is known to suppress the production of a number of other biological agents such as interleukin-1 (IL-1), IL-6, IL-12, TNF-alpha, fibrinogen, and various growth factors.
  • IL-1 interleukin-1
  • IL-6 interleukin-6
  • IL-12 TNF-alpha
  • fibrinogen various growth factors.
  • the invention provides a method of suppressing the production of interleukin-1 (IL-1), IL-6, IL-12, TNF-alpha, fibrinogen, and various growth factors in a cell, comprising contacting a cell with one or more compounds of Formula A, Al, I, II, B, C, or D.
  • the invention provides a method of treating a disease in a patient in need thereof that is beneficially treated by pentoxifylline comprising the step of administering to said patient an effective amount of a compound of Formula A, Al, I, II, B, C, or D or a pharmaceutical composition comprising a compound of Formula A, Al, I, II, B, C, or D and a pharmaceutically acceptable carrier.
  • Such diseases include, but are not limited to, peripheral obstructive vascular disease; glomerulonephritis; nephrotic syndrome; nonalcoholic steatohepatitis; Leishmaniasis; cirrhosis; liver failure; Duchenne's muscular dystrophy; late radiation induced injuries; radiation induced lymphedema; radiation-associated necrosis; alcoholic hepatitis; radiation-associated fibrosis; necrotizing enterocolitis in premature neonates; diabetic nephropathy, hypertension-induced renal failure, and other chronic kidney disease; Focal
  • Segmental Glomerulosclerosis pulmonary sarcoidosis; recurrent aphthous stomatitis; chronic breast pain in breast cancer patients; brain and central nervous system tumors; malnutrition-inflammation-cachexia syndrome; interleukin-1 mediated disease; graft versus host reaction and other allograft reactions; diet- induced fatty liver conditions, atheromatous lesions, fatty liver degeneration and other diet-induced high fat or alcohol-induced tissue-degenerative conditions;
  • human immunodeficiency virus type 1 HIV-1
  • other human retroviral infections multiple sclerosis; cancer; fibroproliferative diseases; fungal infection; drug-induced nephrotoxicity; collagenous colitis and other diseases and/or conditions characterized by elevated levels of platelet derived growth factor (PDGF) or other inflammatory cytokines; endometriosis; optic neuropathy and CNS impairments associated with acquired immunodeficiency syndrome (AIDS), immune disorder diseases, or multiple sclerosis; autoimmune disease; upper respiratory viral infection; depression; urinary incontinence; irritable bowel syndrome; septic shock; Alzheimers Dementia; neuropathic pain; dysuria; retinal or optic nerve damage; peptic ulcer; insulin-dependent diabetes; non-insulin-dependent diabetes; diabetic nephropathy; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension; hyperlipidemia;
  • hyperuricemia gout
  • hypercoagulability acute alcoholic hepatitis
  • olfaction disorders olfaction disorders
  • patent ductus arteriosus olfaction disorders
  • inflammation or injury associated with neutrophil chemotaxis and/or degranulation e.g., neutrophil chemotaxis and/or degranulation.
  • the compounds of Formula A, Al, I, II, B, C, or D can also be used to control intraocular pressure or to stabilize auto-regulation of cerebral blood flow in subjects who require such control as determined by medical examination.
  • the method of this invention is used to treat a disease or condition in a patient in need thereof selected from intermittent claudication on the basis of chronic occlusive arterial disease of the limbs and other peripheral obstructive vascular diseases; glomerulonephritis; Focal Segmental Glomerulosclerosis; nephrotic syndrome; nonalcoholic steatohepatitis;
  • Leishmaniasis Leishmaniasis; cirrhosis; liver failure; Duchenne's muscular dystrophy; late radiation induced injuries; radiation induced lymphedema; alcoholic hepatitis;
  • the method of this invention is used to treat diabetic nephropathy, hypertensive nephropathy or intermittent claudication on the basis of chronic occlusive arterial disease of the limbs. In one specific aspect of this embodiment, the method of this invention is used to treat diabetic nephropathy.
  • the method of this invention is used to treat a disease or condition in a patient in need thereof selected from intermittent claudication on the basis of chronic occlusive arterial disease of the limbs.
  • the method of this invention is used to treat chronic kidney disease.
  • the chronic kidney disease may be selected from glomerulonephritis, focal segmental glomerulosclerosis, nephrotic syndrome, reflux uropathy, or polycystic kidney disease.
  • the method of this invention is used to treat chronic disease of the liver.
  • the chronic disease of the liver may be selected from nonalcoholic steatohepatitis, fatty liver degeneration or other diet-induced high fat or alcohol-induced tissue-degenerative conditions, cirrhosis, liver failure, or alcoholic hepatitis.
  • the method of this invention is used to a diabetes-related disease or condition.
  • This disease may be selected from insulin resistance, retinopathy, diabetic ulcers, radiation-associated necrosis, acute kidney failure or drug-induced nephrotoxicity.
  • the method of this invention is used to treat a patient suffering from cystic fibrosis, including those patients suffering from chronic Pseudomonas bronchitis.
  • the method of this invention is used to aid in wound healing.
  • types of wounds that may be treated include venous ulcers, diabetic ulcers and pressure ulcers.
  • the method of this invention is used to treat a disease or condition in a patient in need thereof selected from insulin dependent diabetes; non-insulin dependent diabetes; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension;
  • hyperlipidemia hyperuricemia; gout; and hypercoagulability.
  • the method of this invention is used to treat a disease or condition in a patient in need thereof wherein the disease or condition is selected from anemia, Graves disease, retinal vein occlusion, lupus nephritis, macular degeneration, myelodysplasia, pruritis of HIV origin, pulmonary hypertension, retinal artery occlusion, intestinal inflammation, ischemic optic neuropathy, acute pancreatitis, sickle cell anemia and beta thalassemia.
  • the disease or condition is selected from anemia, Graves disease, retinal vein occlusion, lupus nephritis, macular degeneration, myelodysplasia, pruritis of HIV origin, pulmonary hypertension, retinal artery occlusion, intestinal inflammation, ischemic optic neuropathy, acute pancreatitis, sickle cell anemia and beta thalassemia.
  • Methods delineated herein also include those wherein the patient is identified as in need of a particular stated treatment. Identifying a patient in need of such treatment can be in the judgment of a patient or a health care professional and can be subjective (e.g. opinion) or objective (e.g. measurable by a test or diagnostic method).
  • any of the above methods of treatment comprises the further step of co-administering to the patient one or more second therapeutic agents.
  • the choice of second therapeutic agent may be made from any second therapeutic agent known to be useful for co-administration with pentoxifylline.
  • the choice of second therapeutic agent is also dependent upon the particular disease or condition to be treated. Examples of second therapeutic agents that may be employed in the methods of this invention are those set forth above for use in combination
  • compositions comprising a compound of this invention and a second therapeutic agent.
  • the combination therapies of this invention include coadministering a compound of Formula A, Al, I, II, B, C, or D and a second therapeutic agent for treatment of the following conditions (with the particular second therapeutic agent indicated in parentheses following the indication): late radiation induced injuries (a-tocopherol), radiation-induced fibrosis (a-tocopherol), radiation induced lymphedema (a-tocopherol), chronic breast pain in breast cancer patients (a-tocopherol), type 2 diabetic nephropathy (captopril), malnutrition- inflammation-cachexia syndrome (oral nutritional supplement, such as Nepro; and oral anti-inflammatory module, such as Oxepa); and brain and central nervous system tumors (radiation therapy and hydroxyurea).
  • a-tocopherol late radiation induced injuries
  • a-tocopherol radiation-induced fibrosis
  • a-tocopherol radiation induced lymphedema
  • chronic breast pain in breast cancer patients a-tocopherol
  • the combination therapies of this invention also include co-administering a compound of Formula A, Al, I, II, B, C, or D and a second therapeutic agent for treatment of insulin dependent diabetes; non-insulin dependent diabetes; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension; hyperlipidemia; hyperuricemia; gout; and hypercoagulability.
  • co-administered means that the second therapeutic agent may be administered together with a compound of this invention as part of a single dosage form (such as a composition of this invention comprising a compound of the invention and an second therapeutic agent as described above) or as separate, multiple dosage forms.
  • the additional agent may be administered prior to, consecutively with, or following the administration of a compound of this invention.
  • both the compounds of this invention and the second therapeutic agent(s) are administered by conventional methods.
  • composition of this invention comprising both a compound of the invention and a second therapeutic agent, to a patient does not preclude the separate administration of that same therapeutic agent, any other second therapeutic agent or any compound of this invention to said patient at another time during a course of treatment.
  • the effective amount of the compound of this invention is less than its effective amount would be where the second therapeutic agent is not administered. In another embodiment, the effective amount of the second therapeutic agent is less than its effective amount would be where the compound of this invention is not administered. In this way, undesired side effects associated with high doses of either agent may be minimized. Other potential advantages (including without limitation improved dosing regimens and/or reduced drug cost) will be apparent to those of skill in the art.
  • the invention provides the use of a compound of Formula A, Al, I, II, B, C, or D alone or together with one or more of the above- described second therapeutic agents in the manufacture of a medicament, either as a single composition or as separate dosage forms, for treatment or prevention in a patient of a disease, disorder or symptom set forth above.
  • Another aspect of the invention is a compound of Formula A, Al, I, II, B, C, or D for use in the treatment or prevention in a patient of a disease, disorder or symptom thereof delineated herein.
  • Step 1 3-Methyl-7-(methyl- j -lH-purine-2,6(3HJH)-dione (51).
  • a suspension of 3-methylxanthine 50 (5.0 g, 30.1 mmol, 1 equiv) and powdered K 2 C0 3 (5.0 g, 36.0 mmol, 1.2 equiv) in DMF (95 mL) was heated to 60 °C and iodomethane-d3 (Cambridge Isotopes, 99.5 atom% D, 2.2 mL, 36.0 mmol, 1.2 equiv) was added via syringe. The resulting mixture was heated at 80 °C for 5 hours (h).
  • the reaction mixture was cooled to room temperature (rt) and the DMF was evaporated under reduced pressure.
  • the crude residue was dissolved in 5% aqueous NaOH (50 mL), resulting in a dull yellow solution.
  • the aqueous solution was washed with CH 2 Cl 2 three times (500 mL total).
  • the aqueous layer was acidified to pH 5 with acetic acid (6 mL), resulting in formation of a tan precipitate.
  • the mixture was cooled in an ice-water bath, and the solids were filtered and washed with cold water.
  • the solid was dried in a vacuum oven to give 2.9 g of 51 as a tan solid.
  • the filtrate was concentrated to approximately 25 mL and a second crop (0.70 g) of 51 was collected by filtration. The total yield of 51 was 3.6 g.
  • the crude material was used without further purification.
  • Step 2 3-Methyl-7-(methyl- ⁇ -l-(5-oxohexyn-lH-purine-2,6(3HJH)- dione (Compound 100).
  • Crude 51 (1.50 g, 8.2 mmol, 1 equiv) and powdered K 2 CO 3 (2.28 g, 16.4 mmol, 2 equiv) were suspended in DMF (30 mL) and heated to 50 °C.
  • To the resulting tan suspension was added 6-chloro-2-hexanone (52, 1.2 mL, 9.0 mmol, 1.1 equiv) and the reaction temperature was raised to 130 °C.
  • Example 2 Synthesis of 8- 3-methyl-7-(methyl- j)-l-(6- j-4- 7-5- oxohexyl)-lH-purine-2,6(3H,7H)-dione (Compound 409). 8- ⁇ -3-methyl-7-(methyl- j )-l-(6- j -4-(i7-5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 409).
  • the H/D exchange reaction to convert the CH3C(0)CH 2 functional group in 100 to the CD3C(0)CD 2 functional group in 409 may be generally applied under analogous conditions to convert compounds having one or more hydrogens alpha to the carbonyl group to compounds having one or more deuteriums in place of the corresponding one or more hydrogens.
  • successive H/D exchange reactions may be performed as needed to further increase the amount of deuterium incorporation.
  • any excess D 2 0 at the end of a second H/D exchange reaction in a given batch run may be used in a first H/D exchange reaction in a subsequent batch run; and any excess D 2 0 at the end of a third H/D exchange reaction in a given batch run may be used in a second H/D exchange reaction in a subsequent batch run.
  • the Table below shows an exemplary deuteration on three separate batches of a compound having a CH 3 C(0)CH 2 functional group and indicates the deuteration agent (either >99% D 2 0 or an aqueous phase obtained from a later deuteration cycle of another batch) that may be used according to this invention.
  • the following table shows the percentage of deuterium incorporation at each of the methyl(CO), (CO)methylene, and the imidazole ring methine carbon for successive batch runs of 50 kg of pentoxifylline in each batch:
  • Exchange 4 is optional. In the Batch 3 run shown in the table, Exchange 4 was performed at half- volume to ensure high deuterium incorporation in Batch 3.
  • Step 1 3 ,7-Di(methyl-(ij)-lH-purine-2,6(3H,7H -dione (55).
  • the reaction mixture was cooled to room temperature, diluted with additional toluene (50 mL) and filtered through Celite to remove any unreacted starting material. The filtrate was evaporated to dryness under reduced pressure to produce 54 as a white solid (4.1 g).
  • Step 2 3,7-Di(methyl- -l-(5-oxohexyn-lH-purine-2,6(3H,7H)-dione (Compound 101).
  • a suspension of crude 55 (2.50 g, 13.4 mmol, 1.0 equiv) and powdered K 2 C0 3 (2.20 g, 16 mmol, 1.2 equiv) in DMF (50 mL) was heated to 60 °C.
  • To the resulting tan suspension was added 6-chloro-2-hexanone 52 (2.0 mL, 14.8 mmol, 1.1 equiv) and the mixture was heated to 140 °C. Heating was continued at 140 °C for 4 hours during which time the suspension became finer and darker in color.
  • Step 1 5-(3-Methyl-7-(methyl- ⁇ -2,3.6J-tetrahvdro-lH-purin-l-vn-N- methoxy-N-methylpentanamide (57).
  • a suspension of 51 (1.50 g, 8.2 mmol, 1.0 equiv, see Example 1 for preparation) and powdered K 2 CO 3 (1.80 g, 12.9 mmol, 1.6 equiv) in DMF (40 mL) was heated to 60 °C.
  • 5-Bromo-N-methoxy-N- methylpentanamide 56 (2.21 g, 9.8 mmol, 1.2 equiv, prepared as outlined in Org.
  • Step 2 3-Methyl-7-(methyl-(ij)-l-(6,6,6-(ij-5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 99).
  • a suspension of 57 (0.72 g, 2.2 mmol, 1.0 equiv) in THF (20 mL) was cooled to 2 °C and 1M CD 3 MgI in ether (2.4 mL, 2.4 mmol, 1.1 equiv, Aldrich >99 atom%> D) was added drop-wise via syringe at a rate to maintain the temperature below 5 °C. During the addition, the mixture became a fine, slightly yellow suspension.
  • reaction mixture was warmed to room temperature and was stirred for 3 hours.
  • the mixture was cooled to 2 °C and an additional portion of CD 3 MgI solution (0.4 mL, 0.4 mmol) was added.
  • the mixture was allowed to warm to room temperature and was stirred an additional 3 hours.
  • the reaction was quenched with IN HC1 (4 mL) and diluted with H 2 0 (10 mL) resulting in a slightly yellow solution that was extracted with CH 2 C1 2 (3X, 200 mL).
  • the combined organic extracts were dried over Na 2 S0 4 , filtered, and concentrated under reduced pressure to a yellow oil.
  • the crude product was purified using an Analogix automated chromatography system eluting with 100% CH 2 C1 2 for 8 minutes and then a gradient of 0-5% MeOH/ CH 2 C1 2 over 40 minutes.
  • the desired product elutes first at about 22 minutes, followed by unreacted starting material.
  • Fractions containing the desired product were concentrated under reduced pressure to a yellow oil that solidified upon standing.
  • the solid was triturated with hexane (25 mL) and collected via vacuum filtration to give 0.33 g (53%>) of Compound 99 as a white solid, mp 93.7 - 94.4 °C.
  • Fractions containing unreacted starting material were also collected and concentrated to give 0.21 g of 57 as a clear, colorless oil.
  • Example 7 Chiral Separation of (i?)-8- -l-(4,4,6,6,6- -5-Hydroxyhexyl)- 3-methyl-7-(methyl- ⁇ -lH-purine-2,6(3HJH)-dione (Compound 419(R) and (6 ⁇ - 8- ⁇ -l-(4 ⁇ ,6,6,6-(i -5-Hvdroxyhexyl)-3-methyl-7-(methyl-(ij)-lH-purine- 2,6(3H,7H)-dione (Compound 419(S)).
  • Enantiomeric separation was achieved using a Waters HPLC system equipped with a preparative Daicel Chiralpak AD column (20 X 250 mm).
  • the mobile phase was 80% hexane and 20% iPrOH along with 0.1%
  • Example 9 Chiral Separation of (i?)-8- l-(4A5,6,6,6-A-5- Hvdroxyhexyl)-3 -methyl-7-(methyl- ⁇ i j )- lH-purine-2,6(3H,7H)-dione (Compound 435(R) and ( ⁇ -8- -l-(4,4,5,6,6,6-A-5-Hvdroxyhexyn-3-methyl-7-(methyl- j - lH-purine-2,6(3HJH)-dione (Compound 435(S)V Separation of Enantiomers of Compound 435.
  • HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 ⁇ C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 254 nm): retention time: 3.25 min; 99.8% purity.
  • Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min;
  • HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 ⁇ C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 254 nm): retention time: 3.25 min; 99.5% purity.
  • Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min;
  • the sample was eluted with 20% iPrOH/hexanes (henceforth, with 0.1% diethylamine as co-eluent) while ramping up from a flow rate of 15 mL/min to 18 mL/min. Over the next 15 minutes, the sample was eluted at a flow rate of 18 mL/min with a gradient of 20% to 25%
  • the reaction mixture was diluted with 18 mL of water and the solution was heated to 25-30°C.
  • KH 2 P0 4 (0.85 g) was added to the mixture and the pH was adjusted to 7 with 4M KOH solution.
  • To the resulting mixture was added (2.5 g, 8.8 mmoles) of Compound 407.
  • a solution of NAD (15 mg), GDH (2.5 mg), KRED 101 (25 mg) in 12.5 mL of 0.1 KH 2 P0 4 buffer was added. The resulting solution was stirred at 25-30°C.
  • the pH of the reaction mixture was maintained between 6 and 7 by adding 4M KOH solution drop-wise as needed. HPLC monitoring of the reaction indicated that the reaction was complete after 12 hours with 99.97A% conversion by HPLC analysis.
  • Chiral HPLC analysis (method: Chiralpak AD-H 25 cm column - isocratic method 75% n-heptane/ 25% isopropanol for 25 min at 1.25 mL/min; wavelength: 274 nm): retention time: 17.56 min (major enantiomer); 15.5 min (expected for minor enantiomer): >99.95% ee purity.
  • the reaction mixture was diluted with 18ml mL of water and the solution was heated to 25-30°C.
  • KH 2 P0 4 (0.85 g) was added to the mixture and the pH was adjusted to 7 with 4M KOH solution.
  • a solution of NADP (15 mg), GDH (2.5 mg), CRED A311-NADP (25 mg) in 12.5 mL of 0.1 KH 2 P0 4 buffer was added.
  • the resulting solution was stirred at 25-30°C.
  • the pH of the reaction mixture was maintained between 6 and 7 by adding 4M KOH solution drop-wise.
  • the reaction was monitored by HPLC and was complete after 12 hours with 99.97% conversion by HPLC.
  • Example 16 Chiral Separation of (i?)-l-(5- ⁇ -5-Hydroxyhexyl)-3-methyl-7- (methyl- j -lH-purine-2,6(3H,7H -dione (Compound 13KR) and (S)- 5-d,-5- Hvdroxyhexyl)-3 -methyl-7-(methyl- ⁇ i j )- 1 H-purine-2,6(3H,7H)-dione (Compound 13KS
  • Example 17 Synthesis of ( ⁇ ) l-(4A6A6-d 5-hydroxyhexylV3 J-dimethyl-8- d-lH-purine-2,6(3H,7H)-dione (Compound 421). heme 21. Preparation of Compounds 421, 421(R) and 421(5).
  • Example 18 Chiral Separation of (i?)-l-(4A6,6,6-d 5-hydroxyhexyl)-3,7- dimethyl-8-d-lH-purine-2,6(3HJH)-dione (Compound 42KR) and (S)-l- (4 ⁇ ,6.6.6-d ⁇ -5-hvdroxyhexyn-3J-dimethyl-8-d-lH-purine-2,6(3H.7H)-dione (Compound 42K5
  • a solution of NADP (15 mg, 3 wt%), GDH (3 mg, 0.6 wt%), ALMAC CRED A311 -NADP (30 mg, 6 wt%) in 0.1 M KH 2 PO 4 buffer was added and maintained reaction temperature 25-30°C.
  • a compound of the invention may be further converted to a compound of the invention having the same structure except for having a group by treating with a suitable base and a proton source, such as water.
  • HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 ⁇ C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95%> ACN; Wavelength: 305 nm): retention time: 3.37 min; 99.5% purity.
  • Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.20 min (expected for R enantiomer); 28.78 min (S enantiomer); >99% ee purity.
  • Step 1 Compound 407. Pentoxifylline (58; 1 mol equiv) was combined with toluene (20 volumes). To the mixture was added D 2 0 (1.5 volumes) and potassium carbonate (0.25 equiv) and the mixture was heated to reflux (ca. 87°C) for 3-4 hrs. The mixture was cooled to 40-50°C and the aqueous layer was removed. To the remaining toluene solution was added D 2 0 (1.5 volumes) and potassium carbonate (0.25 equiv) and the mixture was heated to reflux (ca. 87°C) for 3-4 hrs. The mixture was cooled to 40-50°C and the aqueous layer was removed.
  • reaction mixture was a very thin milky suspension.
  • the aqueous layer was then extracted with EtOAc (2 x 5 vol) without any problems (no further slime or emulsion was observed).
  • the combined organic extracts were concentrated to ca. 10 volume and polish filtered to remove a small amount of the inorganic solids.
  • the filtrate was concentrated further to ca. 5 volumes and product solids were precipitated, n- heptane (8 vol) was added to the slurry (at 40-60°C) over 30 minutes. The slurry was stirred overnight at 20-25 °C and filtered. The filter cake was washed with n-heptane (2 x 1 vol).
  • the isolated product was a very high quality (100% purity by HPLC) and a single enantiomer (100/0 S/R% by chiral HPLC) from the main lot with 99.5% "D” incorporation at the methyl position by 1H NMR.
  • a solution of glucose (558 g, Aldrich lot # 088K0039) in buffer (2 L) was added in one portion followed by a solution of NAD (19.25 g, Spectrum lot # YA0655) in buffer (500 mL), and a solution of GDH (1.5 g, ALMAC lot # IM- 1311-131-1) in buffer (500 mL).
  • the initial reaction mixture was pH 6.98.
  • a mixture of Compound 407 in buffer (3 L) at 30 °C was added to the reaction mixture and the container was rinsed with buffer (1.6 L). The rinse was charged to the reactor.
  • the pH of the reaction mixture was 6.99.
  • the reaction mixture was warmed to 30°C and monitored by pH meter.
  • the reaction temperature was kept at 29.0 to 31.5°C and the pH of the reaction mixture was kept between pH 6.93 and pH 7.02 by adding 4M KOH solution drop-wise as needed.
  • the reaction was complete after 22 hours with 99.96% conversion as determined by HPLC.
  • the chiral HPLC analysis of the resulting product showed the chiral selectivity was 99.85%) to the desired S-alcohol.
  • Example 27 Chiral Separation of (R)-l -(4,4,5, 6,6,6-dfi-5-Hvdroxyhexyl)- 3,7-di(methyl-d ⁇ -lH-purine-2,6(3H,7H)-dione (Compound 434(R) and (6 ⁇ -1- (4,4,5,6,6,6-dfi-5-Hydroxyhexyl)-3,7-di(methyl-d -lH-purine-2,6(3H,7H)-dione (Compound 434(5)).
  • Example 29 Chiral Separation of (i?)-l-(4,4,5,6,6,6-dfi-5-hvdroxyhexyl)-3- methyl-7-methyl-d ⁇ -lH-purine-2,6(3H,7H)-dione (Compound 135(RV) and (S)-l- (4,4, 5,6,6, 6-dfi-5-hydroxyhexyl)-3 -methyl-7-methyl-d ⁇ -lH-purine-2,6(3H,7H - dione (Compound 135(5)).
  • HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 ⁇ CI 8-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 305 nm): retention time: 3.27 min; 99.8% purity.
  • Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.39 min (R enantiomer;
  • Example 32 Chiral Separation of (i?)-l-(5- ⁇ -5-Hydroxyhexyl)-3,7- dimethyl-lH-purine-2,6(3H,7H -dione (Compound 133(R) and (S)- 5-d L -5- Hvdroxyhexyl)-3,7-dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 133(5)).
  • Example 33 Synthesis of 3,7-dimethyl-l-(6,6,6-(ij-5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 157) and (S)-3 J-dimethyl-l-(6,6,6- ⁇ -5- hvdroxyhexyl)-lH-purine-2,6(3H,7H)-dione (Compound 156(5)).
  • Step 1 5-(3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purin-l-yl)-N- methoxy-N-methylpentanamide (61).
  • 50 (1.49 g, 8.27 mmol, 1.0 eq) and DMSO (40mL).
  • the mixture was stirred and heated to about 35 °C to dissolve all materials, and then NaH was added (60% dispersion in oil; 347 mg, 8.68 mmol, 1.05 eq) as a single portion.
  • the solid residue was suspended in MTBE (25 mL), the mixture was stirred at 50 °C for 1 hr, then at room temperature for another hour, followed by filtration via a medium-porosity funnel, an MTBE rinse (2 x 10 mL), and drying under vacuum.
  • the product was collected as 2.19 g (82%) of an off-white solid with an AUC purity of 99 A%.
  • Step 2 3,7-dimethyl-l-(6,6,6-(ij-5-oxohexyl)-lH-purine-2,6(3H,7H)-dione (Compound 157).
  • 61 (1.30 g, 4.01 mmol, 1.0 eq) and THF (45mL). The mixture was stirred and heated to about 45 °C to dissolve all materials, then was cooled to 0 °C (note: a solid precipitated, but less solid was present than prior to heating).
  • CD 3 MgI was added (1.0 M in Et 2 0, 8.82 mL, 2.2 eq) via syringe, at such a rate that the internal temperature did not rise above 5 °C. After the addition was complete, the mixture was stirred at 0 °C for 30 mins, then the cold bath was removed and the mixture was allowed to warm up to room temperature. The mixture was stirred at room temperature for 1.5 hrs, whereupon IPC analysis by HPLC indicated conversion was 80 A%. Further stirring at room temperature for 1.5 hrs afforded no additional conversion, as shown by another IPC analysis. The mixture was cooled to 0 °C and more CD 3 MgI (1.0 M in Et 2 0, 2.0 mL, 0.5 eq) was added via a syringe.
  • the mixture was warmed to room temperature overnight. The next day, IPC analysis by HPLC indicated that conversion was higher than 95 A%.
  • the reaction mixture was quenched with 0.5 N aqueous citric acid (40 mL) and extracted with MTBE (60 mL). The phases were separated and the organic layer was washed with water (20 mL), aq. satd. NaHC0 3 (20 mL), then water (20 mL). The organic layer was concentrated in a rotovap to afford crude product with an AUC purity at 91 A%. Further purification was carried out by slurrying in MTBE (5 mL) at room temperature overnight, followed by filtration and an MTBE rinse.
  • Step 3 (S)-3 ,7-dimethyl- 1 -(6,6,6- j-5-hydroxyhexyO- 1 H-purine-2,6(3H,7H)-dione (Compound 156).
  • Compound 157 563 mg, 2.00 mmol, 1.0 eq
  • D (+)-glucose 844 mg
  • 0.1 M KH 2 P0 4 11 mL
  • NAD 3.4 mg, 0.6 w %)
  • GDH 0.6 mg, 0.1 w %)
  • enzyme KRED-NADH 5.6 mg, 1.0 w %)
  • 0.1 N KH 2 P0 4 This solution was added to the reaction mixture.
  • the resulting cloudy solution was stirred at room temperature for 5 hrs, during which 4N aq. KOH was added dropwise to the reaction mixture to maintain its pH between 6 and 7, as measured by a pH meter. An aliquot was sampled and analyzed by HPLC and indicated greater than 99.5 A% conversion. Solid NaCl ( ⁇ 3 g) was added and the mixture stirred for 30 mins. EtOAc (10 mL) was added and the mixture was stirred for another 30 mins.
  • Example 34a Evaluation of Pharmacokinetics in Dogs Following Oral Administration. Comparison of Compound 409 and Pentoxifylline
  • Plasma samples were removed from dosed dogs at various time points and plasma isolated therefrom. The plasma samples were used for the determination of plasma drug levels by LC-MS/MS (liquid chromatography with tandem mass spectrometry) for estimating pharmacokinetic parameters.
  • LC-MS/MS liquid chromatography with tandem mass spectrometry
  • Compound 409 and pentoxifylline were dissolved separately in saline to a concentration of 4 mg/mL.
  • a 1 : 1 (v/v) mixture of the two solutions was prepared to yield a solution having a final concentration of 2 mg/mL of both Compound 409 and pentoxifylline.
  • Blood samples (1.5 - 2 mL) were collected via the femoral vein at 0 min (pre-dose), 15 min, 30 min, 45 min, 1 hr, 1.5 hr, 2 hr, 3 hr, 4 hr, 6 hr, 8 hr, 10 hr, 12 hr, 16 hr and 24 hr post-dose. Blood was stored on ice prior to centrifugation to obtain plasma samples. Centrifugation took place within 1 hour of blood collection to harvest plasma (maximum volume). The plasma was decanted immediately and frozen/stored at -70 °C until analysis.
  • Example 34a shows the results of the evaluation described in Example 34a.
  • the deuterated compound exhibited greater exposure in the dog plasma than pentoxifylline.
  • Example 34b Repeat Evaluation of Pharmacokinetics in Dogs Following Oral Administration. Comparison of Compound 409 and Pentoxifylline with Monitoring of Metabolites
  • Example 34a was repeated with additional monitoring of the pentoxifylline and Compound 409 metabolites.
  • Compound 409 and pentoxifylline were dissolved separately in saline to a concentration of 4.4 and 4 mg/mL respectively.
  • a 1 : 1 (v/v) mixture of the two solutions was prepared to yield a solution having a final concentration of 2.2 mg/mL of Compound 409 and 2 mg/mL pentoxifylline.
  • Post-dosing data analysis included adjustments to account for the 10% difference in dosing concentration between compound 409 and pentoxifylline.
  • Plasma samples were analyzed by LC-MS/MS for the presence of the administered com ound and its corresponding Ml metabolite:
  • FIGS. 1A and IB The results from each of the four dogs are shown in FIGS. 1A and IB.
  • the results from one of the four dogs (Dog H, FIG. lb) were inconsistent with that of the other three. That dog showed a 10-fold higher plasma concentration of each of the administered compounds and their respective metabolites at 5 minutes post- administration. In addition, that dog did not show a characteristic increase in plasma concentration of the administered compounds between 5 and 15 minutes post- administration. It was concluded that this dog was most likely improperly gavaged and that the compounds were probably administered through the trachea, rather than into the GI tract as would have been desired. Accordingly, the data from this dog was excluded from the analyses. The summary analysis of the three remaining dogs is shown in Table 9.

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Abstract

This invention relates to novel compounds that are substituted xanthine derivatives and pharmaceutically acceptable salts thereof. For example, this invention relates to novel substituted xanthine derivatives that are derivatives of pentoxifylline. This invention also provides compositions comprising one or more compounds of this invention and a carrier and the use of the disclosed compounds and compositions in methods of treating diseases and conditions for which pentoxifylline and related compounds are beneficial.

Description

SUBSTITUTED XANTHINE DERIVATIVES
RELATED APPLICATIONS
This application claims the benefit of priority of U.S. Patent Application Number 12/873,991, filed September 1, 2010 and U.S. Provisional Application Number 61/239,342, filed September 2, 2009, the entire teachings of which are incorporated herein by reference.
BACKGROUND OF THE INVENTION
Many current medicines suffer from poor absorption, distribution, metabolism and/or excretion (ADME) properties that prevent their wider use. Poor ADME properties are also a major reason for the failure of drug candidates in clinical trials. While formulation technologies and prodrug strategies can be employed in some cases to improve certain ADME properties, these approaches have failed to overcome the inherent ADME problems that exist for many drugs and drug candidates. One inherent problem is the rapid metabolism that causes a number of drugs, which otherwise would be highly effective in treating a disease, to be cleared too rapidly from the body. A possible solution to rapid drug clearance is frequent or high dosing to attain a sufficiently high plasma level of drug. This, however, introduces a number of potential treatment problems, such as poor patient compliance with the dosing regimen, side effects that become more acute with higher doses, and increased cost of treatment.
In some select cases, a metabolic inhibitor will be co-administered with an important drug that is rapidly cleared. Such is the case with the protease inhibitor class of drugs that are used to treat HIV infection. These drugs are typically co- dosed with ritonavir, an inhibitor of cytochrome P450 enzyme CYP3A4, the enzyme responsible for their metabolism. Ritonavir itself has side effects and it adds to the pill burden for HIV patients who must already take a combination of different drugs. Similarly, dextromethorphan which undergoes rapid CYP2D6 metabolism is being tested in combination with the CYP2D6 inhibitor quinidine for the treatment of pseudobulbar disease. In general, combining drugs with cytochrome P450 inhibitors is not a satisfactory strategy for decreasing drug clearance. The inhibition of a CYP enzyme activity can affect the metabolism and clearance of other drugs metabolized by that same enzyme. This can cause those other drugs to accumulate in the body to toxic levels.
A potentially attractive strategy, if it works, for improving a drug's metabolic properties is deuterium modification. In this approach, one attempts to slow the CYP-mediated metabolism of a drug by replacing one or more hydrogen atoms with deuterium atoms. Deuterium is a safe, stable, non-radioactive isotope of hydrogen. Deuterium forms stronger bonds with carbon than hydrogen does. In select cases, the increased bond strength imparted by deuterium can positively impact the ADME properties of a drug, creating the potential for improved drug efficacy, safety, and tolerability. At the same time, because the size and shape of deuterium are essentially identical to hydrogen, replacement of hydrogen by deuterium would not be expected to affect the biochemical potency and selectivity of the drug as compared to the original chemical entity that contains only hydrogen.
Over the past 35 years, the effects of deuterium substitution on the rate of metabolism have been reported for a very small percentage of approved drugs (see, e.g., Blake, MI et al, J Pharm Sci, 1975, 64:367-91; Foster, AB, Adv Drug Res 1985, 14: 1-40 ("Foster"); Kushner, DJ et al, Can J Physiol Pharmacol 1999, 79-88; Fisher, MB et al, Curr Opin Drug Discov Devel, 2006, 9: 101-09 ("Fisher")). The results have been variable and unpredictable. For some compounds deuteration caused decreased metabolic clearance in vivo. For others, there was no change in metabolism. Still others demonstrated decreased metabolic clearance. The variability in deuterium effects has also led experts to question or dismiss deuterium modification as a viable drug design strategy for inhibiting adverse metabolism. (See Foster at p. 35 and Fisher at p. 101).
The effects of deuterium modification on a drug's metabolic properties are not predictable even when deuterium atoms are incorporated at known sites of metabolism. Only by actually preparing and testing a deuterated drug can one determine if and how the rate of metabolism will differ from that of its undeuterated counterpart. Many drugs have multiple sites where metabolism is possible. The site(s) where deuterium substitution is required and the extent of deuteration necessary to see an effect on metabolism, if any, will be different for each drug.
SUMMARY OF THE INVENTION
This invention relates to novel compounds that are substituted xanthine derivatives and pharmaceutically acceptable salts thereof. For example, this invention relates to novel substituted xanthine derivatives that are structurally related to pentoxifylline. This invention also provides compositions comprising one or more compounds of this invention and a carrier and the use of the disclosed compounds and compositions in methods of treating diseases and conditions for which pentoxifylline and related compounds are beneficial.
BRIEF DESCRIPTION OF THE DRAWINGS
FIGS. 1 A and IB depict the serum levels of a compound of this invention, pentoxifylline and certain of their respective metabolites in four individual dogs following oral administration of a combination of pentoxifylline and that compound of this invention.
FIG. 2 depicts the time course of the production of the specific metabolites measured in FIG. 3 following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-Ml with rat whole blood.
FIG. 3 depicts the relative amount of specific metabolites produced following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-Ml with rat whole blood.
FIG. 4 depicts the time course of the production of the specific metabolites measured in FIG. 5 following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-M1 with human liver microsomes.
FIG. 5 depicts the relative amount of specific metabolites produced following incubation of various compounds of this invention, pentoxifylline, (S)-Ml and (R)-Ml with human liver microsomes DETAILED DESCRIPTION OF THE INVENTION
The terms "ameliorate" and "treat" are used interchangeably and include both therapeutic and prophylactic treatment. Both terms mean decrease, suppress, attenuate, diminish, arrest, or stabilize the development or progression of a disease (e.g., a disease or disorder delineated herein), lessen the severity of the disease or improve the symptoms associated with the disease.
"Disease" means any condition or disorder that damages or interferes with the normal function of a cell, tissue, or organ.
It will be recognized that some variation of natural isotopic abundance occurs in a synthesized compound depending upon the origin of chemical materials used in the synthesis. Thus, a preparation of pentoxifylline will inherently contain small amounts of deuterated isotopologues. The concentration of naturally abundant stable hydrogen and carbon isotopes, notwithstanding this variation, is small and immaterial as compared to the degree of stable isotopic substitution of compounds of this invention. See, for instance, Wada E et al, Seikagaku, 1994, 66: 15; Gannes LZ et al, Comp Biochem Physiol Mol Integr Physiol, 1998, 119: 725. In a compound of this invention, when a particular position is designated as having deuterium, it is understood that the abundance of deuterium at that position is substantially greater than the natural abundance of deuterium, which is 0.015%. A position designated as having deuterium typically has a minimum isotopic enrichment factor of at least 3340 (50.1% deuterium incorporation) at each atom designated as deuterium in said compound.
The term "isotopic enrichment factor" as used herein means the ratio between the isotopic abundance and the natural abundance of a specified isotope.
In other embodiments, a compound of this invention has an isotopic enrichment factor for each designated deuterium atom of at least 3500 (52.5% deuterium incorporation at each designated deuterium atom), at least 4000 (60% deuterium incorporation), at least 4500 (67.5% deuterium incorporation), at least 5000 (75%o deuterium), at least 5500 (82.5% deuterium incorporation), at least 6000 (90%) deuterium incorporation), at least 6333.3 (95%> deuterium incorporation), at least 6466.7 (97% deuterium incorporation), at least 6600 (99% deuterium incorporation), or at least 6633.3 (99.5%> deuterium incorporation).
In the compounds of this invention any atom not specifically designated as a particular isotope is meant to represent any stable isotope of that atom. Unless otherwise stated, when a position is designated specifically as "H" or "hydrogen", the position is understood to have hydrogen at its natural abundance isotopic composition. Also unless otherwise stated, when a position is designated
specifically as "D" or "deuterium", the position is understood to have deuterium at an abundance that is at least 3340 times greater than the natural abundance of deuterium, which is 0.015% (i.e., at least 50.1% incorporation of deuterium).
The term "isotopologue" refers to a species that differs from a specific compound of this invention only in the isotopic composition thereof.
The term "compound," when referring to a compound of this invention, refers to a collection of molecules having an identical chemical structure, except that there may be isotopic variation among the constituent atoms of the molecules. Thus, it will be clear to those of skill in the art that a compound represented by a particular chemical structure containing indicated deuterium atoms, will also contain lesser amounts of isotopologues having hydrogen atoms at one or more of the designated deuterium positions in that structure. The relative amount of such isotopologues in a compound of this invention will depend upon a number of factors including the isotopic purity of deuterated reagents used to make the compound and the efficiency of incorporation of deuterium in the various synthesis steps used to prepare the compound. However, as set forth above, the relative amount of such isotopologues in toto will be less than 49.9% of the compound.
The invention also provides salts of the compounds of the invention. A salt of a compound of this invention is formed between an acid and a basic group of the compound, such as an amino functional group, or a base and an acidic group of the compound, such as a carboxyl functional group. According to another embodiment, the compound is a pharmaceutically acceptable acid addition salt.
The term "pharmaceutically acceptable," as used herein, refers to a component that is, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and other mammals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio. A "pharmaceutically acceptable salt" means any non-toxic salt that, upon administration to a recipient, is capable of providing, either directly or indirectly, a compound of this invention. A "pharmaceutically acceptable counterion" is an ionic portion of a salt that is not toxic when released from the salt upon administration to a recipient. Acids commonly employed to form pharmaceutically acceptable salts include inorganic acids such as hydrogen sulfide, hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid and phosphoric acid, as well as organic acids such as para-toluenesulfonic acid, salicylic acid, tartaric acid, bitartaric acid, ascorbic acid, maleic acid, besylic acid, fumaric acid, gluconic acid, glucuronic acid, formic acid, glutamic acid, methanesulfonic acid, ethanesulfonic acid,
benzenesulfonic acid, lactic acid, oxalic acid, para-bromophenylsulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid and acetic acid, as well as related inorganic and organic acids. Such pharmaceutically acceptable salts thus include sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate,
monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caprate, heptanoate, propiolate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, butyne-l,4-dioate, hexyne-l,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate, terephthalate, sulfonate, xylene sulfonate,
phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, β-hydroxybutyrate, glycolate, maleate, tartrate, methanesulfonate, propanesulfonate, naphthalene- 1- sulfonate, naphthalene-2-sulfonate, mandelate and other salts. In one embodiment, pharmaceutically acceptable acid addition salts include those formed with mineral acids such as hydrochloric acid and hydrobromic acid, and especially those formed with organic acids such as maleic acid.
The invention also includes solvates and hydrates of the compound of the invention. As used herein, the term "hydrate" means a compound which further includes a stoichiometric or non-stoichiometric amount of water bound by non- covalent intermolecular forces. As used herein, the term "solvate" means a compound which further includes a stoichiometric or non-stoichiometric amount of solvent such as water, acetone, ethanol, methanol, dichloromethane, 2-propanol, or the like, bound by non-covalent intermolecular forces.
1 2
It is understood that the carbon atom that bears substituents Y and Y in
1 2 3
Formulae A, Al, I and B can be chiral in some instances (when Y , Y and R are different from one another) and in other instances it can be achiral (when at least two
1 2 3 1 ofY1 Y" and lc are the same). This carbon atom (i.e., the carbon atom bearing Y and Y ) is indicated by an "*" in Formulae A, Al, I and B. As such, chiral compounds of this invention can exist as either individual enantiomers, or as racemic or scalemic mixtures of enantiomers. Accordingly, a compound of the present invention will include racemic and scalemic enantiomeric mixtures, as well as individual respective stereoisomers that are substantially free from another possible stereoisomer. The term "substantially free of other stereoisomers" as used herein means less than 25% of other stereoisomers, preferably less than 10% of other stereoisomers, more preferably less than 5% of other stereoisomers and most preferably less than 2% of other stereoisomers, or less than "X"% of other stereoisomers (wherein X is a number between 0 and 100, inclusive) are present. Methods of obtaining or synthesizing an individual enantiomer for a given compound are well known in the art and may be applied as practicable to final compounds or to starting material or intermediates.
Unless otherwise indicated, when a disclosed compound is named or depicted by a structure without specifying the stereochemistry and has one or more chiral centers, it is understood to represent all possible stereoisomers of the compound.
The term "stable compounds," as used herein, refers to compounds which possess stability sufficient to allow for their manufacture and which maintain the integrity of the compound for a sufficient period of time to be useful for the purposes detailed herein (e.g., formulation into therapeutic products, intermediates for use in production of therapeutic compounds, isolatable or storable intermediate compounds, treating a disease or condition responsive to therapeutic agents).
"D" refers to deuterium. "Stereoisomer" refers to both enantiomers and diastereomers. "Tert", " 1 ", and "t-" each refer to tertiary. "US" refers to the United States of America.
As used herein the term "alkylene" means a straight or branched chain divalent hydrocarbon radical, preferably having from one to six carbon atoms (Ci_6alkylene). In some embodiments, the alkylene group has from one to four carbon atoms (Ci_4alkylene). Examples of "alkylene" as used herein include, but are not limited to, methylene (-CH2-), ethylene (-CH2CH2-), propylene (-CH2CH2CH2-), and branched versions thereof such as (-CH(CH3)-), -CH2CH(CH3)- and the like.
"Halo" means chloro, bromo, fluoro, or iodo.
"Alkyl" means an aliphatic hydrocarbon group which may be straight or branched having 1 to 15 carbon atoms in the chain. Preferred alkyl groups have 1 to 12 carbon atoms in the chain, and more preferably 1 to 6 carbon atoms. Branched means that one or more lower alkyl groups such as methyl, ethyl or propyl are attached to a linear alkyl chain. "Lower alkyl" means about 1 to about 4 carbon atoms in the chain which may be straight or branched. Exemplary alkyl groups include methyl, fluoromethyl, difluoromethyl, trifluoromethyl, cyclopropylmethyl, cyclopentylmethyl, ethyl, n-propyl, i-propyl, n-butyl, t-butyl, n-pentyl, 3-pentyl, heptyl, octyl, nonyl, decyl and dodecyl; preferred are methyl, difluoromethyl and i- propyl. Alkyl groups may be optionally substituted with one or more groups selected from halo, cyano, hydroxyl, carboxy, alkoxy, alkoxycarbonyl, oxo, amino, alkylamino, dialkylamino, cycloheteroalkyl, alkylcycloheteroalkyl, aryl, alkylaryl, heteroaryl, and alkylheteroaryl. Typically any alkyl or alkoxy moiety of the alkyl substituent group has 1 to 6 carbon atoms.
"Aryl" means an aromatic carbocyclic radical containing 6 to 10 carbon atoms. Exemplary aryl groups include phenyl or naphthyl. Aryl groups may be optionally substituted with one or more groups which may be the same or different, and which are selected from alkyl, aryl, aralkyl, alkoxy, aryloxy, aralkyloxy, halo, and nitro.
Typically any alkyl or alkoxy moiety of the aryl substituent group has 1 to 6 carbon atoms.
"Heteroaryl" means a 5- to a 10-membered aromatic monocyclic or multicyclic hydrocarbon ring system in which one or more of the carbon atoms in the ring system is or are element(s) other than carbon, for example nitrogen, oxygen or sulfur. Heteroaryl groups may be optionally substituted with one or more groups which may be the same or different, and which are selected from alkyl, aryl, aralkyl, alkoxy, aryloxy, aralkyloxy, halo, and nitro. Exemplary heteroaryl groups include pyrazinyl, furanyl, thienyl, pyridyl, pyrimidinyl, isoxazolyl, isothiazolyl, pyridazinyl, 1,2,4-triazinyl, quinolinyl, and isoquinolinyl.
"Aralkyl" means an aryl-alkyl group in which the aryl and alkyl components are as previously described. Preferred aralkyls contain a lower alkyl moiety.
Exemplary aralkyl groups include benzyl and 2-phenethyl.
"Heteroaralkyl" means a heteroaryl-alkyl group in which the heteroaryl and alkyl components are as previously described.
"Cycloalkyl" means a non-aromatic mono-, multicyclic, or bridged ring system of 3 to 10 carbon atoms. The cycloalkyl group is optionally substituted by one or more halo, or alkyl. Exemplary monocyclic cycloalkyl rings include cyclopentyl, fluorocyclopentyl, cyclohexyl and cycloheptyl.
"Heterocycloalkyl" means a non-aromatic mono-, bi- or tricyclic, or bridged hydrocarbon ring system in which one or more of the atoms in the ring system is or are element(s) other than carbon, for example nitrogen, oxygen or sulfur. Preferred heterocycloalkyl groups contain rings with a ring size of 3-6 ring atoms. Exemplary heterocycloalkyl groups pyrrolidine, piperidine, tetrahydropyran, tetrahydrofuran, tetrahydrothiopyran, and tetrahydrothiofuran.
"Cycloalkylalkyl" means a group in which the cycloalkyl and alkyl components are as previously described.
"Heteroycloalkylalkyl" means a group in which the cycloalkyl and alkyl components are as previously described.
The term "optionally substituted with deuterium" means that one or more hydrogen atoms in the referenced moiety or compound may be replaced with a corresponding number of deuterium atoms.
Throughout this specification, a variable may be referred to generally
1 2 3
(e.g., "each R") or may be referred to specifically (e.g., R , R , R , etc.). Unless otherwise indicated, when a variable is referred to generally, it is meant to include all specific embodiments of that particular variable.
THERAPEUTIC COMPOUNDS
The resent invention provides a compound of Formula A:
Figure imgf000010_0001
(A), or a pharmaceutically acceptable salt thereof, wherein:
R 1 and R 2" are each independently selected from hydrogen, -(Ci-C4)alkyl, or - (Ci-C4)alkylene-0-(Ci-C2)alkyl, wherein the alkyl and alkylene groups at each instance are independently and optionally substituted with deuterium;
R3 is selected from -CH3, -CH2D, -CHD2 and -CD3;
R4 is n-butylene optionally substituted with deuterium;
R5 is selected from hydrogen, deuterium, alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, heterocycloalkylalkyl, aryl, and heteroaryl, wherein each of the alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, heterocycloalkylalkyl, aryl, and heteroaryl is optionally substituted and wherein one or more hydrogen atoms in the alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, heterocycloalkylalkyl, aryl, or heteroaryl or optional substituent thereof is optionally replaced with a corresponding number of deuterium atoms; and
either (a)Y 1 and Y 2 are each fluorine, or are taken together with the carbon to which they are bound to form C=0 or (b) Y1 is selected from fluorine and OH; and Y is selected from hydrogen, deuterium, -CH , -CH2D, -CHD2 and -CD ;
with the provisos that:
when Y 1 and Y 2 are taken together with the carbon to which they are bound to form C=0, then at least one of R1, R2, R3, R4, and R5 bears at least one deuterium atom; and
when Y1 is OH and Y2 is hydrogen or CH3, then at least one of R1, R2, R3, R4 and R5 bears at least one deuterium atom.
In another embodiment, the compound of Formula A is other than the following:
Figure imgf000011_0001
In another embodiment of Formula A, when R 1 and R 2 are each methyl optionally substituted with deuterium and R5 is hydrogen or deuterium, then either: (i) Y1 is fiuoro; or (ii) Y1 is OH, and Y2 is selected from -CH3, -CH2D, -CHD2 and -CD . In one aspect of this embodiment, the compound is not
Figure imgf000011_0002
2 1 2 3
In a more specific aspect of this embodiment, at least one of Y\ R , R\ R and R4 bears at least one deuterium atom.
In still another embodiment of Formula A, R 1 and R 2 are each methyl optionally substituted with deuterium; R5 is hydrogen or deuterium; and either: (a)
Y 1 and Y 2" are taken together with the carbon atom to which they are bound to form
=0, or (b) Y 1 is -OH and Y 2 is selected from hydrogen and deuterium, with the provisos that:
when Y 1 and Y 2 are taken together with the carbon to which they are bound to form C=0, then at least one of R1, R2, R3, R4, and R5 bears at least one deuterium atom; and
when Y1 is OH, then at least one of Y2, R1, R2, R3, R4 and R5 bears at least one deuterium atom.
In another embodiment of Formula A, R5 is D, the compound having
Formula Al :
Figure imgf000012_0001
(Al), or a salt thereof, wherein R1, R2, R3, R4,
1 2
Y and Y" are as defined for Formula A.
In one aspect of Formula Al , R 1 and R 2 are each independently selected from -CH3, -CH2D, -CHD2 and -CD3; R3 is selected from -CH3, -CH2D, -CHD2 and -CD3; R4 is selected from -(CH2)4-, -(CD2)4-,†- (CD2)3CH2, and†-CD2(CH2)3-, wherein
4 1 2
"†" represents the portion of the R moiety bound to C(Y )(Y ) in the compound; and either (a) Y 1 is OH and Y2 is selected from hydrogen and deuterium; or (b) Y 1 and Y are taken together with the carbon to which they are attached to form C=0.
In a more specific aspect of Formula A 1, R 1 and R 2 are each independently selected from -CH3 and -CD3; R3 is selected from -CH3 and -CD3; R4 is selected from -(CH2)4- and†-CD2(CH2)3-; and either (a) Y1 is OH and Y2 is selected from hydrogen and deuterium; or (b) Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0.
1 2
In another aspect of Formula A 1, R and R are each independently selected from -CH3 and -CD3; R3 is selected from -CH3 and -CD3; R4 is selected from
-(CH2)4- and†-CD2(CH2)3-; and Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0.
In another embodiment, the present invention provides a compound of Formula A, wherein R5 is hydrogen, the compound having Formula I:
Figure imgf000013_0001
(I), or a salt thereof, wherein:
R 1 and R 2" are each independently selected from hydrogen, -(Ci-C4)alkyl, or - (Ci-C4)alkylene-0-(Ci-C2)alkyl, wherein the alkyl and alkylene groups at each instance are independently and optionally substituted with deuterium;
R3 is selected from -CH3, -CH2D, -CHD2 and -CD3;
R4 is n-butylene optionally substituted with deuterium; and
either (a) Y 1 and Y 2 are each fluorine, or taken together with the carbon to which they are attached, form C=0; or (b) Y1 is selected from fluorine and OH; and Y is selected from hydrogen, deuterium, -CH , -CH2D, -CHD2 and -CD , with the provisos that:
when Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0, at least one of R1, R2, R3 and R4 bears at least one deuterium atom; and
when Y 1 is OH and Y2 is hydrogen or -CH3, then at least one of R 1 , R2 , R 3 and R4 bears at least one deuterium atom.
In a more specific embodiment of Formula I, R 1 and R 2 are each
independently selected from -CH , -CH2D, -CHD2 and -CD ; R is selected from -CH3, -CH2D, -CHD2 and -CD3; R4 is selected from -(CH2)4-, -(CD2)4-,†- (CD2) CH2, and†-CD2(CH2) -, wherein "†" represents the portion of the R4 moiety
1 2 1 2
bound to C(Y )(Y ) in the compound; and either: Y is OH and Y is selected from hydrogen and deuterium; or Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0.
1 2
In another aspect of Formula I, R and R are each independently selected from -CH and -CD ; R3 is selected from -CH and -CD ; R4 is selected from
-(CH2)4- and†-CD2(CH2) -; and either: Y 1 is OH and Y 2 is selected from hydrogen and deuterium; or Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0.
1 2
In another aspect of Formula I, R and R are each independently selected from -CH and -CD ; R3 is selected from -CH and -CD ; R4 is selected from
-(CH2)4- and†-CD2(CH2)3-; and Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0. In another embodiment, in any of the aspects set forth above, the compound of Formula I is other than the following:
Figure imgf000014_0001
, or
In yet another embodiment, in any of the aspects set forth above, the compound of Formula I is other than the following:
Figure imgf000014_0002
In yet another embodiment, in any of the aspects set forth above, the compound of Formula I is other than the following:
Figure imgf000014_0003
Another embodiment of the present invention provides a compound of Formula II:
Figure imgf000014_0004
R 1 and R 2" are each independently selected from hydrogen, -(Ci-C4)alkyl, or - (Ci-C4)alkylene-0-(Ci-C2)alkyl, wherein the alkyl and alkylene groups at each instance are independently and optionally substituted with deuterium;
R3 is selected from -CH3, -CH2D, -CHD2 and -CD3;
R4 is n-butylene optionally substituted with deuterium; and
2 3 4
wherein at least one of R , R and R bears at least one deuterium atom. One embodiment relates to a compound of Formula A, A 1 , 1, or II, wherein
2 3
R" and RJ are each independently selected from -CH3, -CH2D, -CHD2 and -CD3.
Another embodiment relates to a compound of Formula A, A 1 , 1, or II,
2 3
wherein R and R are each independently selected from -CH3, and -CD3.
Another embodiment relates to a compound of Formula A, A 1 , 1, or II, wherein R1 is selected from hydrogen, (Ci-C3)alkyl, and (Ci-C2)alkylene-0(Ci- C2)alkyl.
Another embodiment relates to a compound of Formula A, A 1 , 1, or II, wherein R1 is hydrogen, -CH3, -CD3, -CH2CH2CH3, -CD2CH2CH3, -CD2CD2CH3, - CD2CD2CD3, -CH2OCH2CH3, -CH2OCD2CH3, -CH2OCD2CD3, -CD2OCH2CH3, - CD2OCD2CH3, or -CD2OCD2CD3.
Another embodiment relates to a compound of Formula A, wherein R5 is selected from hydrogen, deuterium, alkyl, cycloalkyl, heterocycloalkyl,
cycloalkylalkyl, and heterocycloalkylalkyl, wherein each of alkyl, cycloalkyl, heterocycloalkyl, cycloalkylalkyl, and heterocycloalkylalkyl may be optionally substituted.
In other embodiments of Formula A, Al or I:
a) each methylene unit in R4 is selected from -CH2- and -CD2-; more
4 †
specifically R is selected from -(CH2)4-, -(CD2)4-, '-CD2(CH2)3- and
†-(CD2)3CH2-, wherein "†" represents the point where R4 is attached to
C(Yl)(Y2) in the compound;
b) when Y1 is F, Y2 is selected from hydrogen, -CH3, -CH2D, -CHD2 and -CD3; or
1 2
c) when Y is F, Y is fluorine; or
1 2 2 3 1 d) when Y and Y are not the same and Y and R are not the same and Y and
Y1 Νγ2
3 R3^R4- R are not the same, the stereochemistry at "*" is represented by: * ; or
1 2 2 3 1 e) when Y and Y are not the same and Y and R are not the same and Y and γ1 γ2
3 R3 R4-^
R are not the same, the stereochemistry at "*" is represented by: * .
1 2 3
In other embodiments of Formula A, Al or I, R is -CD3; R and R are
1 2 4
-CH3; Y and ΎΔ are taken together to form C=0; and R is selected from -(CH2)4-, -(CD2)4-,†-CD2(CH2)3- and†-(CD2)3CH2-.
1 2 3
In other embodiments of Formula A, Al or I, R is -CD3; R and R are
1 2 4
-CH3; Y and ΎΔ are taken together to form C=0; and R is selected from -(CH2)4-, and -(CD2)4-.
1 2 3
In other embodiments of Formula A, Al or I, R is -CD3; R and R are -
4 1 2
CH3; R is -(CH2)4-; Y is fluoro; and Y is selected from deuterium, -CH2D, -CHD2 and -CD3.
1 2 3
In other embodiments of Formula A, Al or I, R is -CD3; R and R are - CH3; R4 is -(CH2)4-; Y1 is fluoro; and Y2 is fluorine.
1 2 3
In other embodiments of Formula A or A 1 , R is -CD3; R and R are -CH3; R4 is -(CH2)4-; R5 is deuterium; Y1 is fluoro; and Y2 is selected from deuterium, -CH2D, -CHD2 and -CD3.
1 2 3
In other embodiments of Formula A or A 1 , R is -CD3; R and R are -CH3; R4 is -(CH2)4-; R5 is deuterium; Y1 is fluoro; and Y2 is fluorine.
1 2
In other embodiments of Formula A, Al or I, Y is F; Y is selected from hydrogen; R3 is -CH3; and R4 is -(CH2)4-.
1 2 3
In other embodiments of Formula A, Al or I, Y is F; Y is fluorine; R is - CH3; and R4 is -(CH2)4-.One embodiment provides a compound of Formula B:
Figure imgf000016_0001
B, or a pharmaceutically
1 2
acceptable salt thereof, wherein each of R and R is independently selected from
5 3
-CH3 and -CD3; R is hydrogen or deuterium; each Z is hydrogen or deuterium; each Z4 is hydrogen or deuterium; each Z5 is hydrogen or deuterium; and either (a)
1 2 1 2
Y is OH, and Y is hydrogen or deuterium, or (b) Y and Y are taken together with the carbon to which they are attached to form C=0.
One embodiment provides a compound of Formula B, wherein each Z3, Z4 5 1 2 5 and Z is hydrogen. In one aspect, R and R are each -CD3. In another aspect R is deuterium. In another aspect, Y 1 and Y 2 are taken together with the carbon to which
1 2 they are attached to form C=0. In still another aspect, Y and is OH, and Y is hydrogen or deuterium.
Another embodiment provides a compound of Formula B, wherein each Z , Z4 and Z5 is deuterium. In one aspect, R1 and R2 are each -CD3. In another aspect R5 is deuterium. In another aspect, Y 1 and Y 2 are taken together with the carbon to which they are attached to form C=0. In still another aspect, Y 1 and is OH, and Y 2 is hydrogen or deuterium.
Yet another embodiment provides a compound of Formula B, wherein R1 and R2 are each -CD3. In one aspect, R5 is deuterium. In another aspect, each Z , Z and Z5 is hydrogen and R5 is deuterium. In another aspect, each Z3, Z4 and Z5 is deuterium and R5 is deuterium.
A further embodiment provides a compound of Formula B, wherein Y1 and Y are taken together with the carbon to which they are attached to form C=0. In one aspect, R5 is deuterium. In another aspect, each Z3, Z4 and Z5 is hydrogen and R5 is deuterium. In another aspect, each Z3, Z4 and Z5 is deuterium and R5 is deuterium. In another aspect, R 1 and R2 are each 1
-CD3. In another aspect, R and R2 are each R 5 is deuterium. In another aspect, R 1
-CD3 and and R2 are each -CD3, and each Z3, Z4 and Z5 is deuterium. In another aspect, R1 and R2 are each -CD3, each Z3, Z4 and Z5 is deuterium and R5 is deuterium. In another aspect, R1 and R2 are each -CD3, and each Z3, Z4 and Z5 is hydrogen. In another aspect, R1 and R2 are each -CD3, each Z3, Z4 and Z5 is hydrogen and R5 is deuterium
A still further embodiment provides a compound of Formula B, Y1 and is
OH, and Y 2 is hydrogen or deuterium. In one aspect, R 5 is deuterium. In another aspect, each Z3, Z4 and Z5 is hydrogen and R5 is deuterium. In another aspect, each Z3, Z4 and Z5 is deuterium and R5 is deuterium. In another aspect, R1 and R2 are each R 1 and R2 are each 5
-CD3. In another aspect, -CD3 and R is deuterium. In another aspect, R1 and R2 are each -CD3, and each Z3, Z4 and Z5 is deuterium. In another aspect, R1 and R2 are each -CD3, each Z3, Z4 and Z5 is deuterium and R5 is deuterium. In another aspect, R1 and R2 are each -CD3, and each Z3, Z4 and Z5 is hydrogen. In another aspect, R1 and R2 are each -CD3, each Z3, Z4 and Z5 is hydrogen and R5 is deuterium
Another embodiment provides a compound of Formula B, wherein R5 is deuterium.
Another embodiment provides a compound of Formula B, wherein R5 is deuterium, Z3, Z4 and Z5 is hydrogen and R1 is -CD3.
Specific examples of compounds of Formula A, Al , I, or II include those shown in Tables 1-6 (below) or pharmaceutically acceptable salts thereof, wherein
† 4 1 2
"'" represents the portion of the R moiety bound to C(Y )(Y ) in the compound. In the tables, compounds designated as "(7?)" or "(5)" refer to the stereochemistry at the carbon bearing the Y1 substituent. Compounds lacking either designation and
1 2
containing a chiral carbon atom bound to Y and Y are intended to represent a racemic mixture of enantiomers.
Table 1 : Examples of Specific Compounds of Formula I. Deuterated and/or
Fluorinated Analogs of Pentoxifylline and its Metabolites.
Figure imgf000018_0001
Figure imgf000019_0001
Compound R R R R Y Y
155(R) CD3 CH3 CD3 (CH2)4 (R)OH H
156 CH3 CH3 CD3 (CH2)4 OH H
156(5) CH3 CH3 CD3 (CH2)4 (S)OH H
156(R) CH3 CH3 CD3 (CH2)4 (R)OH H
157 CH3 CH3 CD3 (CH2)4 taken together as =0
Table 1 above shows examples of specific compounds of Formula I. These examples are deuterated and/or fluorinated analogs of pentoxifylline and its metabolites.
1 2
Table 2: Examples of Specific Compounds of Formula I Where R is H and Y is
CH3 or CD3.
Figure imgf000020_0001
Table 2 above shows examples of specific compounds of Formula I where R is H and Y is CH3 or CD3. These compounds include deuterated and fluorinated analogs of Albifylline (HWA-138). Albifylline has been studied for uses that are associated with pentoxifylline.
Table 3: Specific Examples of Formula I Where R1 is -CH2-0-CH2CH3 Optionally
Substituted with Deuterium.
Figure imgf000020_0002
Compound R R R R Y Y
259 CD2OCH2CH3 CH3 CD3 (CH2)4 OH CD3
260 CH2OCH2CH3 CH3 CD3 (CH2)4 OH CD3
261 CD2OCD2CD3 CD3 CD3 TCD2(CH2)3 OH CD3
262 CD2OCH2CH3 CD3 CD3 TCD2(CH2)3 OH CD3
263 CH2OCH2CH3 CD3 CD3 TCD2(CH2)3 OH CD3
264 CD2OCD2CD3 CH3 CD3 TCD2(CH2)3 OH CD3
265 CD2OCH2CH3 CH3 CD3 TCD2(CH2)3 OH CD3
266 CH2OCH2CH3 CH3 CD3 TCD2(CH2)3 OH CD3
267 CD2OCD2CD3 CD3 CD3 (CD2)4 OH CD3
268 CD2OCH2CH3 CD3 CD3 (CD2)4 OH CD3
269 CH2OCH2CH3 CD3 CD3 (CD2)4 OH CD3
270 CD2OCD2CD3 CH3 CD3 (CD2)4 OH CD3
271 CD2OCH2CH3 CH3 CD3 (CD2)4 OH CD3
272 CH2OCH2CH3 CH3 CD3 (CD2)4 OH CD3
273 CD2OCD2CD3 CD3 CH3 (CH2)4 F CH3
274 CD2OCH2CH3 CD3 CH3 (CH2)4 F CH3
275 CH2OCH2CH3 CD3 CH3 (CH2)4 F CH3
276 CD2OCD2CD3 CH3 CH3 (CH2)4 F CH3
277 CD2OCH2CH3 CH3 CH3 (CH2)4 F CH3
278 CH2OCH2CH3 CH3 CH3 (CH2)4 F CH3
279 CD2OCD2CD3 CD3 CD3 (CD2)4 F CD3
280 CD2OCH2CH3 CD3 CD3 (CD2)4 F CD3
281 CH2OCH2CH3 CD3 CD3 (CD2)4 F CD3
282 CD2OCD2CD3 CH3 CD3 (CD2)4 F CD3
283 CD2OCH2CH3 CH3 CD3 (CD2)4 F CD3
284 CH2OCH2CH3 CH3 CD3 (CD2)4 F CD3
Table 3 above shows examples of specific compounds of Formula I where R is -CH2-0-CH2CH3, optionally substituted with deuterium. In these examples, Y1 is OH or F and Y is CH3 or CD3. These compounds include deuterated and fluorinated analogs of torbafylline (HWA-448). Torbafylline has been studied for the treatment of depression, urinary incontinence, irritable bowel syndrome and multiple sclerosis.
Table 4: Specific Examples of Formula I Where R1 is -CH2CH2CH3 Optionally
Substituted With Deuterium and Y1 is OH or F.
Figure imgf000021_0001
Compound R R R R Y Y
306 CD2CH2CH3 CD3 CD3 (CH2)4 OH CD3
307 CH2CH2CH3 CD3 CD3 (CH2)4 OH CD3
308 CD2CD2CD3 CH3 CD3 (CH2)4 OH CD3
309 CD2CH2CH3 CH3 CD3 (CH2)4 OH CD3
310 CH2CH2CH3 CH3 CD3 (CH2)4 OH CD3
311 CD2CD2CD3 CD3 CD3 TCD2(CH2)3 OH CD3
312 CD2CH2CH3 CD3 CD3 TCD2(CH2)3 OH CD3
313 CH2CH2CH3 CD3 CD3 TCD2(CH2)3 OH CD3
314 CD2CD2CD3 CH3 CD3 TCD2(CH2)3 OH CD3
315 CD2CH2CH3 CH3 CD3 TCD2(CH2)3 OH CD3
316 CH2CH2CH3 CH3 CD3 TCD2(CH2)3 OH CD3
317 CD2CD2CD3 CD3 CD3 (CD2)4 OH CD3
318 CD2CH2CH3 CD3 CD3 (CD2)4 OH CD3
319 CH2CH2CH3 CD3 CD3 (CD2)4 OH CD3
320 CD2CD2CD3 CH3 CD3 (CD2)4 OH CD3
321 CD2CH2CH3 CH3 CD3 (CD2)4 OH CD3
322 CH2CH2CH3 CH3 CD3 (CD2)4 OH CD3
323 CD2CD2CD3 CD3 CH3 (CH2)4 F CH3
324 CD2CH2CH3 CD3 CH3 (CH2)4 F CH3
325 CH2CH2CH3 CD3 CH3 (CH2)4 F CH3
326 CD2CD2CD3 CH3 CH3 (CH2)4 F CH3
327 CD2CH2CH3 CH3 CH3 (CH2)4 F CH3
328 CH2CH2CH3 CH3 CH3 (CH2)4 F CH3
329 CD2CD2CD3 CD3 CD3 (CD2)4 F CD3
330 CD2CH2CH3 CD3 CD3 (CD2)4 F CD3
331 CH2CH2CH3 CD3 CD3 (CD2)4 F CD3
332 CD2CD2CD3 CH3 CD3 (CD2)4 F CD3
333 CD2CH2CH3 CH3 CD3 (CD2)4 F CD3
334 CH2CH2CH3 CH3 CD3 (CD2)4 F CD3
Table 4 above shows examples of specific compounds of Formula I where R is -CH2CH2CH3 optionally substituted with deuterium. In these examples, Y1 is OH or F and Y is CH3 or CD3. These compounds include deuterated and fluorinated analogs of A-802715. A-802715 has been studied for the treatment of septic shock and inhibition of effects of allograft reaction.
Table 5 : Specific Examples of Formula I where R1 is -CH2CH2CH3 Optionally
1 2
Substituted With Deuterium and Y and Y Are Taken Together As = O
Figure imgf000022_0001
Compound R R R R Y1 Y2
354 CD2CH2CH3 CH3 CH3 (CH2)4 taken together as =0
355 CD2CD2CD3 CD3 CD3 (CH2)4 taken together as =0
356 CD2CH2CH3 CD3 CD3 (CH2)4 taken together as =0
357 CH2CH2CH3 CD3 CD3 (CH2)4 taken together as =0
358 CD2CD2CD3 CH3 CD3 (CH2)4 taken together as =0
359 CD2CH2CH3 CH3 CD3 (CH2)4 taken together as =0
360 CH2CH2CH3 CH3 CD3 (CH2)4 taken together as =0
361 CD2CD2CD3 CD3 CD3 TCD2(CH2)3 taken together as =0
362 CD2CH2CH3 CD3 CD3 TCD2(CH2)3 taken together as =0
363 CH2CH2CH3 CD3 CD3 TCD2(CH2)3 taken together as =0
364 CD2CD2CD3 CH3 CD3 TCD2(CH2)3 taken together as =0
365 CD2CH2CH3 CH3 CD3 TCD2(CH2)3 taken together as =0
366 CH2CH2CH3 CH3 CD3 TCD2(CH2)3 taken together as =0
367 CD2CD2CD3 CD3 CD3 (CD2)4 taken together as =0
368 CD2CH2CH3 CD3 CD3 (CD2)4 taken together as =0
369 CH2CH2CH3 CD3 CD3 (CD2)4 taken together as =0
370 CD2CD2CD3 CH3 CD3 (CD2)4 taken together as =0
371 CD2CH2CH3 CH3 CD3 (CD2)4 taken together as =0
372 CH2CH2CH3 CH3 CD3 (CD2)4 taken together as =0
Table 5 above shows examples of specific compounds of Formula I where R is -CH2CH2CH3 optionally substituted with deuterium. In these examples, Y1 and Y are taken together with their intervening carbon to form a carbonyl. These compounds include deuterated analogs of propentofylline. Propentofylline has been studied for the treatment of Alzheimer's disease, neuropathic pain, traumatic brain injury, dysuria, retinal or optic nerve head damage, and peptic ulcers. It has also been studied for controlling intraocular pressure, stabilization of auto-regulation of cerebral blood flow and inhibition of effects of allograft reaction.
Table 6: Examples of Specific Compounds of Formula A. Deuterated and/or
Fluorinated Analogs of Pentoxifylline and its Metabolites where R5 is D
Compound R R R R R5 Y Y
410 CD3 CHs CDs T(CD2)sCH2 D Taken together as =0
411 CHs CDs CDs TCD2(CH2)s D Taken together as =0
412 CHs CDs CDs T(CD2)sCH2 D Taken together as =0
413 CDs CDs CDs TCD2(CH2)s D Taken together as =0
414 CDs CDs CDs T(CD2)sCH2 D Taken together as =0
415 CDs CDs CHs (CH2)4 D OH H
416 CDs CHs CHs (CH2)4 D OH H
417 CHs CDs CHs (CH2)4 D OH H
418 CDs CDs CDs TCD2(CH2)s D OH H
418(R) CDs CDs CDs TCD2(CH2)s D (R)OH H
418(5) CDs CDs CDs TCD2(CH2)s D (5)0H H
419 CDs CHs CDs TCD2(CH2)s D OH H
419(R) CDs CHs CDs TCD2(CH2)s D (R)OH H
419(5) CDs CHs CDs TCD2(CH2)s D (5)OH H
420 CHs CDs CDs TCD2(CH2)s D OH H
421 CHs CHs CDs TCD2(CH2)s D OH H
421(R) CHs CHs CDs TCD2(CH2)s D (R)OH H
421(5) CHs CHs CDs TCD2(CH2)s D (5)0H H
422 CDs CDs CDs (CD2)4 D OH H
423 CDs CHs CDs (CD2)4 D OH H
424 CHs CDs CDs (CD2)4 D OH H
425 CHs CHs CDs (CD2)4 D OH H
426 CDs CDs CDs T(CD2)sCH2 D OH H
427 CDs CHs CDs T(CD2)sCH2 D OH H
428 CHs CDs CDs T(CD2)sCH2 D OH H
429 CHs CHs CDs T(CD2)sCH2 D OH H
430 CDs CDs CHs (CH2)4 D OH D
431 CDs CHs CHs (CH2)4 D OH D
432 CHs CDs CHs (CH2)4 D OH D
433 CHs CHs CHs (CH2)4 D OH D
434 CDs CDs CDs TCD2(CH2)s D OH D
434(R) CDs CDs CDs TCD2(CH2)s D (R)OH D
434(5) CDs CDs CDs TCD2(CH2)s D (5)0H D
435 CDs CHs CDs TCD2(CH2)s D OH D
435(R) CDs CHs CDs TCD2(CH2)s D (R)OH D
435(5) CDs CHs CDs TCD2(CH2)s D (5)0H D
436 CHs CDs CDs TCD2(CH2)s D OH D
437(R) CHs CHs CDs TCD2(CH2)s D (R)OH D
437(5) CHs CHs CDs TCD2(CH2)s D (5)0H D
437 CHs CHs CDs TCD2(CH2)s D OH D
438 CDs CDs CDs (CD2)4 D OH D
439 CDs CHs CDs (CD2)4 D OH D
440 CHs CDs CDs (CD2)4 D OH D
441 CHs CHs CDs (CD2)4 D OH D
442 CDs CDs CDs T(CD2)sCH2 D OH D
443 CDs CHs CDs T(CD2)sCH2 D OH D
444 CHs CDs CDs T(CD2)sCH2 D OH D Compound R R R R R5 Y Y
445 CH3 CH3 CD3 T(CD2)3CH2 D OH D
446 CD3 CD3 CH3 (CH2)4 D F H
447 CH3 CH3 CH3 (CH2)4 D F H
448 CD3 CH3 CH3 (CH2)4 D F H
449 CH3 CD3 CH3 (CH2)4 D F H
450 CD3 CD3 CH3 (CH2)4 D F F
451 CD3 CH3 CH3 (CH2)4 D F F
452 CH3 CD3 CH3 (CH2)4 D F F
453 CH3 CH3 CH3 (CH2)4 D F F
Table 6 above shows examples of specific compounds of Formula A. These examples are deuterated and/or fluorinated analogs of pentoxifylline and its metabolites where R5 is deuterium.
In one aspect of the above embodiment, the compound is not any one of Compounds 100, 116, or 149.
Examples of specific compounds of this invention include the following:
Figure imgf000025_0001
Figure imgf000025_0002
Figure imgf000025_0003
Figure imgf000025_0004
Figure imgf000026_0001
Figure imgf000026_0002
The present invention provides a compound of Formula C
Figure imgf000026_0003
C
or a pharmaceutically acceptable salt thereof, wherein R is selected from
2 1 2
-CH and -CD ; R is selected from -CH3 and -CD3; one of Y and Y is -OH; and the other of Y 1 and Y 2 is deuterium or hydrogen^
provided that if R1 is CD3 and R2 is CH3, then Y2 is -OH.
One embodiment provides a compound of Formula C, wherein R1 is -CH3. One embodiment provides a compound of Formula C, wherein R1 is -CD3. One embodiment provides a compound of Formula C, wherein R is -CH3. One embodiment provides a compound of Formula C, wherein R is -CD3. One embodiment provides a compound of Formula C, wherein R1 is -CH3 and R 2 is -CH3. In one aspect of this embodiment, Y 1 is -OH. In another aspect, Y2 is OH.
One embodiment provides a compound of Formula C, or a pharmaceutically acceptable salt thereof, wherein the compound has the structure:
Figure imgf000026_0004
In one aspect of this embodiment, Y is deuterium. In another aspect, Y is hydrogen. One embodiment provides a compound of Formula C, or a pharmaceutically acce table salt thereof, wherein the compound has the structure:
Figure imgf000027_0001
In one aspect of this embodiment, Y 2 is deuterium. In another aspect, Y 2 is hydrogen.
Examples of the compounds of the formula C include the following compounds and pharmaceutically acceptable salts thereof:
Figure imgf000027_0002
Figure imgf000027_0003
The present invention also provides a compound of Formula D:
Figure imgf000028_0001
D
or a pharmaceutically acceptable salt thereof, wherein R is selected from
2 1 2
-CH3 and -CD3; R is selected from -CH3 and -CD3; one of Y and Y is -OH; and the other of Y 1 and Y 2 is deuterium or hydrogen;
with the proviso that
(i) if either one of Y 1 and Y2 is deuterium, then R 2 is CD3; and
(ii) if either one of Y 1 and Y2 is hydrogen, then R 1 is CH3.
One embodiment provides a compound of Formula D, wherein R1 is -CH3. One embodiment provides a compound of Formula D, wherein R1 is -CD3. One embodiment provides a compound of Formula D, wherein R is -CH3. One embodiment provides a compound of Formula D, wherein R is -CD3. One embodiment provides a compound of Formula D, wherein R1 is -CH3 and R 2 is -CH3. In one aspect of this embodiment, Y 1 is -OH. In another aspect, Y2 is OH.
One embodiment provides a compound of Formula D, or a pharmaceutically acce table salt thereof, wherein the compound has the structure:
Figure imgf000028_0002
In one aspect of this embodiment, Y is deuterium. In another aspect, Y is hydrogen.
One embodiment provides a compound of Formula D, or a pharmaceutically acce table salt thereof, wherein the compound has the structure:
Figure imgf000028_0003
In one aspect of this embodiment, Y 2 is deuterium. In another aspect, Y 2 is hydrogen.
Examples of the compounds of the formula D include the following compounds and pharmaceutically acceptable salts thereof:
Figure imgf000029_0001
In another set of embodiments, any atom not designated as deuterium in any of the embodiments set forth above is present at its natural isotopic abundance.
The synthesis of compounds of this invention can be achieved by synthetic chemists of ordinary skill. Relevant procedures and intermediates are disclosed, for instance in Sidzhakova, D et al., Farmatsiya, (Sofia, Bulgaria) 1988, 38(4): 1-5; Davis, PJ et al, Xenobiotica, 1985, 15(12): 1001-10; Akgun, H et al, J Pharm Sci, 2001, 26(2): 67-71; German Patent publication DD 274334; Czech Patent Nos. CS 237719, CS201558; PCT patent publication WO9531450; and in Japanese Patent publication Nos. JP58150594, JP58134092, JP58038284, JP57200391, JP57098284, JP57085387, JP57062278, JP57080385, JP57056481, JP57024385, JP57011981, JP57024386, JP57024382, JP56077279, JP56032477, JP56007785, JP56010188, JP56010187, JP55122779, and JP55076876.
Such methods can be carried out utilizing corresponding deuterated and optionally, other isotope-containing reagents and/or intermediates to synthesize the compounds delineated herein, or invoking standard synthetic protocols known in the art for introducing isotopic atoms to a chemical structure.
EXEMPLARY SYNTHESIS
Methods for synthesizing compounds of Formula I are depicted
following schemes. Scheme 1A. Synthesis of Compounds of Formula I
Figure imgf000030_0001
10 11
Formula I
As depicted in Scheme 1 A, deuterated compound 10 is alkylated with deuterated intermediate 11 (wherein X is chloride, bromide or iodide) in the presence of potassium carbonate to afford compounds of Formula I. Alternatively, sodium hydroxide in aqueous methanol may be employed to afford compounds of Formula I according to the methods of US Patent 4289776.
Scheme IB. Preparation of Compounds Where Y1 = OH From Compounds of Formula II
Figure imgf000030_0002
Formula II
As depicted in Scheme IB, compounds of Formula II can be used to make compounds where Y1 is OH. Thus, compounds of Formula II are reduced with either sodium borohydride or sodium borodeuteride (commercially available at 99 atom %D) according to the general method of European Patent publication 0330031 to form compounds wherein Y 1 is OH and Y 2 is hydrogen or deuterium. The enantiomeric alcohol products may be separated, for example through the method of Nicklasson, M et al, Chirality, 2002, 14(8): 643-652. In an alternate method, enzymatic reduction affords an enantiomerically-enriched alcohol product using the methods disclosed in Pekala, E et al., Acta Poloniae Pharmaceutica, 2007, 64(2): 109-113, or in Pekala, E et al, Biotech J, 2007, 2(4): 492-496. 1 2
Stereoselective preparation of compounds where C(Y )(Y ) is C(H)OH or
1 2
C(D)OH from corresponding compounds where C(Y )(Y ) is C=Q may be carried out in the presence of a ketoreductase or carbonyl reductase^Compounds of the
1 2
invention where C(Y )(Y ) is C(H)OH or C(D)OH may be prepared
1 2
stereoselective^ from corresponding compounds where C(Y )(Y ) is C=0 by treating with a hydride source or a deuteride source in the presence of a
ketoreductase or carbonyl reductase at an appropriate pH with an enantiomeric excess of at least 80%. The ketoreductase or carbonyl reductase that favors formation of a compound wherein the stereochemistry at the C(H)OH or C(D)OH group is (S) may be, for example, any one of ALMAC Carbonyl Reductases CRED A131 , CRED A801 , CRED A901 , CRED A251 , or CRED A271 (each
commercially available from ALMAC Group Ltd, Craigavon, England), any one of CODEXIS Ketoreductases KRED-119, KRED-137, KRED-148, KRED-169, KRED-174, KRED-NADH 101 , KRED-NADH 102, KRED-NADHl 12, or KRED- NADH 126 (each commercially available from Codexis Inc., Redwood City, CA), or SYNCORE Ketoreductases ES-KRED-121 , ES-KRED-128, ES-KRED-130, ES- KRED-142, ES-KRED-175, ES-KRED-169, or ES-KRED-171 (each commercially available from Syncore Labs, Shanghai, China). In one aspect of, the enzyme is selected from CRED A131 , CRED A251 , KRED-NADH 101 , KRED-NADH 102, KRED-NADH 1 12, KRED-NADH 126, ES-KRED-121 , ES-KRED-128, ES- KRED-130, ES-KRED-142, ES-KRED-169, or ES-KRED-171. In a more specific aspect, the enzyme is selected from CRED A131 , CRED A251 , and KRED-NADH 101. The ketoreductase or carbonyl reductase that favors formation of a compound wherein the stereochemistry at the C(H)OH or C(D)OH group is (R) may be, for example, any one of KRED-NADP-1 18, CRED A601-NADP, CRED A291-NADP, CRED A31 1-NADP. KRED- AD-1 10, ES-KRED-120, ES CRED-I 31 , CRED A 1G1-NADP.
The amount of ketoreductase or carbonyl reductase used in the reaction ranges from 0.05 wt% to 10 wt% as a percentage of the weight of the substrate, such as 0.5 wt% to 5 wt%. In one embodiment, the amount of enzyme is between 1.0 wt% and 2.0 wt%. In a more specific aspect, the amount of enzyme is about 1.0 wt%.
The hydride source is a compound or mixture that is capable of providing a hydride anion or a synthon of a hydride anion. The deuteride source is a compound or mixture that is capable of providing a deuteride anion or a synthon of a deuteride anion. A hydride or deuteride source comprises a catalytic co-factor and optionally, a co-factor regeneration system. A catalytic co-factor used with the ketone reductase or carbonyl reductase in the process of this invention is selected from NAD, NADP, NADH, NADPH, NAD2H and NADP2H. The choice of co-factor may be based upon (a) the presence or absence of a co-factor regeneration system; (b) the requirement for a hydride versus a deuteride source; and (c) an appropriate pH to perform the method according to the present invention means buffer conditions that maintain the pH at between 6.0 and 7.5 throughout the reaction. In one embodiment, the pH of the reaction was maintained at between 6.5 and 7.3. In another embodiment, the pH of the reaction was maintained between 6.0 and 7.0. Typically dropwise addition of KOH is used to maintain the desired pH because the enzymatic reaction generates acid. In one aspect, the pH of the reaction is maintained between 6.90 and 7.05. The process may be performed at a temperature of about 20° C to 37° C. In one aspect of this embodiment, the temperature is about 29° C to 32° C. The process may be performed over a time period of about 12 hours to about 24 hours. In one embodiment, the time period is about 24 hours to about 40 hours. In one embodiment, the time period is about 40 hours to about 72 hours. In one embodiment, the time period is a time period sufficient for less than about 5% of
1 2
the initial amount of the compound wherein C(Y )(Y ) is C=Q to be present.
Synthesis of Compound 10
Referring to Scheme 1 A, compounds that can be used as compound 10 to make compounds of Formula I are known and include, but are not limited to, the following: theobromine (wherein R 1 and R 2 are CH3) which is commercially
1 2 1 available. Isotopologues of 10 wherein: (a) R is -CD3 and R is -CH3; (b) R is -
2 1 2
CH3 and R" is -CD3; and (c) R and R" are -CD3 are all known. See Benchekroun, Y et al, J Chromatogr B, 1977, 688: 245; Ribon, B et al, Coll INSERM, 1988, 164: 268; and Horning, MG et al, Proc Int Conf Stable Isot 2nd, 1976, 41-54. 3-Methyl-
1 2
7-propylxanthine, wherein R is n-propyl and R is -CH3, is commercially available.
Compound 10 wherein R 1 is 2
CH2OCH3 and R is C¾ is also known. See German patent application DE 3942872A1. Scheme 2. Synthesis of Compound 10
Figure imgf000033_0001
1. NaOH, H2Q
2. HCl, H20
Figure imgf000033_0002
HCOOH
Figure imgf000033_0003
A synthesis of compound 10 is depicted in Scheme 2 starting with commercially-available N-nitroso-N-methylurea. Treatment with appropriately deuterated amine 12 in water affords N-alkylurea 13 following the methods of Boivin, JL et al., Canadian Journal of Chemistry, 1951, 29: 478-81. Urea 13 may be treated with 2-cyanoacetic acid and acetic anhydride to provide cyanoacetamide derivative 14, which is treated first with aqueous NaOH and then with aqueous HCl to provide cyclized pyrimidinedione 15 according to the methods of Dubey, PK et al., Indian Journal of Heterocyclic Chemistry, 2005, 14(4): 301-306. Alternatively, cyanoacetamide 14 may be treated with trimethylsilylchloride and
hexamethyldisilazane to afford the cyclized product 15 via the methods of Fulle, F et al, Heterocycles, 2000, 53(2): 347-352.
Following the methods of Merlos, M et al., European Journal of Medicinal Chemistry, 1990, 25(8): 653-8, treatment of pyrimidinedione 15 with sodium nitrite in acetic acid, and then by ammonium hydroxide and sodium dithionite, yields compound 16, which is treated with formic acid to provide purine derivative 17. Following the methods disclosed by Rybar, A et al, in Czech patent application CS 263595B1, alkylation of 17 with appropriately deuterated electrophile 18 (X is chloro, bromo, or iodo) in the presence of potassium carbonate and optionally in the presence of additives such as NaBr, KBr, Nal, KI, or iodine, affords compound 10.
Referring to Scheme 2, useful deuterated amine reagents 12 include, but are not limited to, commercially-available compounds such as n-propyl-d7-amine, or known compounds such as l-propan-l,l-d2-amine (Moritz, F et al, Organic Mass
Spectrometry, 1993, 28(3): 207-15). Useful deuterated urea reagents 13 may include, but are not limited to, commercially-available compounds such as N-
O O
°3 ^ΝΗ2 D3 ^- D2
methyl-d3-urea H , or methylurea-d6 D
Useful deuterated electrophiles 18 may include, but are not limited to,
commercially-available compounds such as iodomethane-d3, or bromomethane-d3, or l-bromopropane-d7, or l-bromopropane-l,l-d2, or known compounds such as
(chloromethoxy-d2)-ethane (Williams, AG, WO 2002059070A1), or
bromomethoxymethane-d2 (Van der Veken, BJ et al., Journal of Raman
Spectroscopy, 1992, 23(4): 205-23, or (bromomethoxy-d2)-methane-d3 (Van der
Veken, BJ et al, Journal of Raman Spectroscopy, 1992, 23(4): 205-23. The commercially available deuterated intermediates 12, 13 and 18 mentioned above are available having an isotopic purity of at least 98 atom % D.
Synthesis of Intermediate lla-i/5 (cf. Scheme 1A)
Scheme 3. Synthesis of Intermediate lla-ώ
Figure imgf000034_0001
Figure imgf000034_0002
21 11a-t 5
An approach to the preparation of compound lla-i 5 (cf. Scheme 1A)
3 4 † 1 2
(wherein R is CD3; R is '-CD2(CH2)3-, and Y and Y are taken together to form =0), is depicted in Scheme 3. Thus, methyllithium is added to commercially- available delta-valerolactone 19 according to the procedure of Zhang, Q et al., Tetrahedron, 2006, 62(50): 11627-11634 to afford ketone 20. Treatment of 20 with TFA-i / (99 atom %D) in D20 (99 atom %D) under microwave conditions provides deuterated ketone 21 according to the general method of Fodor-Csorba K, Tet Lett, 2002, 43: 3789-3792. The alcohol moiety in 21 is converted to the chloride upon treatment with triphenylphosphine and carbon tetrachloride to yield chloride lla-i/5 following the general procedures of Clement, J-L, Org Biomol Chem, 2003, 1 : 1591-1597.
Scheme 4. Synthesis of Intermediates lla-f/g and lla-d
Figure imgf000035_0001
22 23
Figure imgf000035_0002
Scheme 4 depicts a synthesis of compound lla-i 9 and compound lla-i 77. Thus, commercially-available 4-phenylbutyric acid 22 may be heated in D20 (99 atom %D) in the presence of Pd/C and hydrogen gas to afford deuterated acid 23 according to the general methods of Esaki, et al, Chem Eur J, 2007, 13: 4052-4063. Addition of deuterated methyllithium in the presence of trimethylsilyl chloride provides ketone 24, according to the general method of Porta, A et al., J Org Chem, 2005, 70(12): 4876-4878. Ketone 24 is converted to acetal 25 by treatment with D2SO4 (99 atom %D) and commercially-available ethyleneglycol-^ (99 atom %D). Treatment of 25 with NaI04 and RuCl3 according to the general method of Gamier, J-M et al., Tetrahedron: Asymmetry, 2007, 18(12): 1434-1442 provides carboxylic acid 26. Reduction with either L1AIH4 or LiAlD4 (98 atom %D) provides the alcohols (not shown), which are then chlorinated using either phosphorus oxychloride or triphenylphosphine and N-chlorosuccinimide (Naidu, SV et al, Tet Lett, 2007, 48(13): 2279-2282), followed by acetal cleavage with D2S04
(Heathcock, CH et al, J Org Chem, 1995, 60(5): 1 120-30) to provides chlorides lla-i 9 and lla-i n, respectively.
Scheme 4a. Synthesis of Intermediates llb-(R)
Figure imgf000036_0001
27 28 11 b-(R)
Scheme 4b. Synthesis of Chloride llb-(S)
Figure imgf000036_0002
11 b-(S)
Schemes 4a and 4b depict the synthesis of specific enantiomers of chlorides
1 2
llb-(R) (wherein Y is fluorine; Y is selected from hydrogen and deuterium; and
1 2 the compound is in the (R) configuration) and llb-(S) (wherein Y is fluorine; Y is selected from hydrogen and deuterium; and the compound is in the (S)
configuration). In Scheme 4a, a deuterated (or nondeuterated) benzyl-protected alcohol 27, such as known [[[(5R)-5-fluorohexyl]oxy]methyl]-benzene (PCT publication WO2000031003) is deprotected by hydrogenation in the presence of Pd/C to provide alcohol 28. The alcohol is chlorinated with thionyl chloride according to the general procedure of Lacan, G et al., J Label Compd Radiopharm, 2005, 48(9): 635-643 to afford chloride llb-(R).
In Scheme 4b, a deuterated (or nondeuterated) alcohol 29, such as known (S)-(+)-5-fluorohexanol (Riswoko, A et al, Enantiomer, 2002, 7(1): 33-39) is chlorinated to afford chloride llb-(S).
Scheme 5. Synthesis of Intermediates 11c and lie
Figure imgf000037_0001
Scheme 5 depicts a synthesis of other intermediates 11c and lie. Thus, following the methods of either Kutner, Andrzej et al., Journal of Organic
Chemistry, 1988, 53(15): 3450-7, or of Larsen, SD et al, Journal of Medicinal Chemistry, 1994, 37(15): 2343-51, compounds 30 or 31 (wherein X is a halide) may be treated with deuterated Grignard reagent 32 to afford intermediate 11c wherein
3 2 1
RJ and are the same, Y is OH, and X is a halide. Treatment with
diethylaminosulfur trifluoride (DAST) in dichloromethane or toluene provides
3 2 1
intermediate lie wherein R and Y are the same, Y is F, and X is a halide according to the general procedures of either Karst, NA et al, Organic Letters, 2003, 5(25): 4839-4842, or of Kiso, M et al, Carbohydrate Research, 1988, 177: 51-67.
Commercially available halides can be used to make compounds 11 as disclosed in Scheme 5. For example, commercially-available 5-chlorovaleryl chloride, or commercially-available 5-bromovaleryl chloride, or commercially- available ethyl 5-bromovalerate, may be useful as reagents 30 or 31. Referring again to Scheme 5, use of commercially-available methyl-<¾-magnesium iodide as
3 2
Grignard reagent 32 affords electrophile 11 wherein R and Y are simultaneously CD3.
Scheme 6. Synthesis of Intermediate lie (X=Br)
Figure imgf000037_0002
CSA PPh3, NBS
ΌΤΗΡ MeOH ΌΗ benzene Br
35 36 11e (X=Br)
Scheme 6 depicts an alternate synthesis of intermediate lie, wherein R and the same and X=Br. Thus, according to the procedures of Hester, JB et al,
Journal of Medicinal Chemistry, 2001 , 44(7): 1099-1 1 15, commercially-available 4-chloro-l-butanol is protected via treatment with 3,4-dihydro-2H-pyran (DHP) and camphorsulfonic acid (CSA) to provide chloride 33. Generation of the
corresponding Grignard reagent with magnesium, followed by addition of acetone (R3 = Y2 = CH3) or acetone-de (Y2 = R3 = CD3), affords alcohol 34. Fluorination with diethylaminosulfur trifluoride (DAST) in CH2Cl2provides fluoride 35.
Deprotection with CSA in MeOH provides alcohol 36, and treatment with N- bromosuccinimide and triphenyl phosphine affords intermediate lie.
Scheme 7. Alternative Synthesis of Intermediate lie (X=Br)
EtCL DHP, CSA, Et20 LiAID4 HOv
OH ΌΤΗΡ ΌΤΗΡ
O or Et,0 D D
37 DHP, TsOH,
r, CH2CI2 38 39
Figure imgf000038_0001
3 2
Scheme 7 depicts the synthesis of intermediate lie wherein R and Y are the same and X=Br. Thus, commercially-available 4-hydroxy-butanoic acid ethyl ester 37 is treated with DHP and CSA, or with DHP, TsOH, and pyridine to provide ester 38. Reduction with L1AID4 affords deuterated alcohol 39, which is treated with either triphenyl phosphine in CC14 (Sabitha, G et al., Tetrahedron Letters, 2006, (volume date 2007), 48(2): 313-315) or with methanesulfonyl chloride, lithium chloride, and 2,6-lutidine in DMF (Blaszykowski, C et al, Organic Letters, 2004, 6(21): 3771-3774) to afford chloride 40. Following the same methods as in Scheme 6, chloride 40 may be converted to lie. Scheme 8. Synthesis of Intermediate lle-fifs (X=Br)
Figure imgf000039_0001
41 42 11e- 8
Scheme 8 depicts the synthesis of intermediate lle-i n R 3 and Y 2
8 wherei are the same and X=Br. Thus, commercially-available THF-<¾ 41 may be treated with DCl and ZnCl2 according to the general methods of Yang, A et al., Huagong Shikan, 2002, 16(3): 37-39 to afford known chloride 42 (Aiken, Rudolf-Giesbert, WO 2003080598A1). Following the same methods as in Scheme 6, chloride 42 may be converted to l l -dg.
Scheme 9. Synthesis of Intermediate llc-fifs (X=Br)
Figure imgf000039_0002
43 44 11c-d8
Scheme 9 depicts the synthesis of intermediate llc-i 3 Y 2
8 wherein R and are the same and X=Br. Thus, known carboxylic acid 43 (Lompa-Krzymien, L et al., Proc. Int. Conf. Stable Isot. 2nd, 1976, Meeting Date 1975, 574-8) is treated with either diazomethane (according to the general method of Garrido, NM et al., Molecules, 2006, 11(6): 435-443.) or with trimethylsilyl chloride and methanol-<i/ (according to the general method of Doussineau, T et al, Synlett, 2004, (10): 1735- 1738) to provide methyl ester 44. As in Scheme 5, treatment of the ester with deuterated Grignard reagent 45 affords intermediate llc-i s- For example, use of commercially-available methyl-d3-magnesium iodide as Grignard reagent 45 affords
3 2
llc-i 8 wherein R and Y are simultaneously CD3.
Scheme 10. Synthesis of Intermediate llc-< ?.
Figure imgf000039_0003
11c-d2 3 2
Scheme 10 depicts a preparation of llc-i/2, wherein R and Y are the same.
Thus, known deuterated ester 46 (Feldman, KS et al, Journal of Organic Chemistry, 2000, 65(25): 8659-8668) is treated with carbon tetrabromide and
triphenylphosphine (Brueckner, AM et al., European Journal of Organic Chemistry, 2003, (18): 3555-3561) to afford ester 47 wherein X is bromide, or is treated with methanesulfonyl chloride and triethylamine, followed by lithium chloride and DMF (Sagi, K et al, Bioorganic & Medicinal Chemistry, 2005, 13(5): 1487-1496) to afford ester 47 wherein X is chloride. As in Scheme 5, treatment of ester 47 with deuterated Grignard reagent 48 affords llc-i 2. For example, use of commercially- available methyl-d3 -magnesium iodide as Grignard reagent 48 affords 11c- wherein R 3 and Y 2
di are simultaneously CD3.
Additional known chlorides that may be utilized as reagent 11 in Scheme 1 A include:
l-chloro-5,5-difluoro-hexane (Rybczynski, PJ et al, J Med Chemistry, 2004, 47(1): 196-209); l-chloro-5-fluorohexane (Chambers, RD et al., Tetrahedron, 2006, 62(30): 7162-7167); 6-chloro-2-hexanol (European Patent publication 0412596); (S)-6-chloro-2-hexanol (Keinan, E et al, J Am Chem Soc, 1986, 108(12): 3474- 3480); commercially-available (R)-6-chloro-2-hexanol; commercially available 6- chloro-2-hexanone; known 6-chloro-2-methylhexan-2-ol (Kutner, A et al., Journal of Organic Chemistry, 1988, 53(15): 3450-7); known 6-bromo-2-methylhexan-2-ol (Kutner, A et al, Journal of Organic Chemistry, 1988, 53(15): 3450-7); known 1- bromo-5-fluoro-5-methylhexane (Hester, JB et al., Journal of Medicinal Chemistry, 2001, 44(7): 1099-1115).
Scheme 11. Synthesis of Compounds of Formula Al
Figure imgf000040_0001
Formula I Formula A1
Scheme 11 depicts the synthesis of a compound of Formula Al . Thus, a compound of Formula I is treated with potassium carbonate in D20 to effect a hydrogen-to-deuterium exchange reaction, providing a compound of Formula Al . One skilled in the art will appreciate that additional hydrogen-to-deuterium exchange reactions may also occur elsewhere in the molecule.
Scheme 12. Alternative S nthesis of Compounds of Formula Al
Figure imgf000041_0001
10 50 Formula A1
An alternative synthesis of a compound of Formula Al is depicted in Scheme 12. Thus, intermediate 10 (cf. Scheme 1 A) is treated with potassium carbonate in D20 to effect a hydrogen-to-deuterium exchange reaction, providing compound 50 as either the N-D or N-H species. Alkylation with intermediate 11 in the presence of potassium carbonate affords compounds of Formula Al .
Scheme 12b. Alternative Synthesis of Compounds of Formula I.
Figure imgf000041_0002
An alternative synthesis of compounds of Formula I is depicted in Scheme 12b. Thus, compounds of Formula Al are treated with potassium carbonate in water to effect a deuterium-to-hydrogen exchange, which affords other compounds of
1 2
Formula I. Preferably, in the method of Scheme 12b, either (a) Y and Y are each
1 2
fluorine; or (b) Y is selected from fluorine and OH; and Y is selected from hydrogen, deuterium, -CH3, -CH2D, -CHD2 and -CD3.
A number of novel intermediates can be used to prepare compounds of Formula A. Thus, the invention also provides such a compound which is selected from the following:
Figure imgf000042_0001
Figure imgf000042_0002
Figure imgf000042_0003
Figure imgf000042_0004
Figure imgf000042_0005
Figure imgf000043_0001
1 m
Figure imgf000043_0002
Compounds a-d above may be prepared as generally described in Org. Lett., 2005, 7: 1427-1429 using appropriately-deuterated starting materials. Compounds e- o may be prepared from the appropriate bromides listed above by reference to Scheme 15 shown below.
Certain xanthine intermediates useful for this invention are also novel. Thus, the invention provides a deuterated xanthine intermediate III:
Figure imgf000043_0003
III, where W is hydrogen or deuterium, and each of R1 and R is independently selected from hydrogen, deuterium, Ci_3 alkyl optionally substituted with deuterium, and Ci_3 alkoxyalkyl optionally substituted with deuterium. Examples of R 1 and R 2 Ci_3 alkyl include -CH3, -CD3, -CH2CH2CH3, and -CD2CD2CD3. Examples of Ci_3 alkoxyalkyl include -CH2OCH2CH3, - CD2OCH2CH3, -CD2OCD2CH3, and -CD2OCD2CD3.
Specific examples of formula III include the following: CH,
Ο'" "Ν ' ' Ν Ο'" ΊΜ ' " N O'" ΊΜ ' " N
CH3 CD3 CD3
III-a, Ill-b, III-c,
Figure imgf000044_0001
Ill-d, Ill-e, Ill-f,
Figure imgf000044_0002
ni-g, Ill-h, Ill-i,
Figure imgf000044_0003
ni-j, Ill-k, and III-l
In each of the above examples of formula III, W is hydrogen. In a set of
corresponding examples, W is deuterium. Salts of compounds of Formula III are also useful, including salts that are known to be useful with respect to known xanthines. Examples of useful salts include, but are not limited to, the lithium salt, sodium salt, potassium salt, and cesium salt. An example of a particularly useful salt is the potassium salt.
The specific approaches and compounds shown above are not intended to be limiting. The chemical structures in the schemes herein depict variables that are hereby defined commensurately with chemical group definitions (moieties, atoms, etc.) of the corresponding position in the compound formulae herein, whether
1 2
identified by the same variable name (i.e., R , R , R 3 , etc.) or not. The suitability of a chemical group in a compound structure for use in the synthesis of another compound is within the knowledge of one of ordinary skill in the art.
Additional methods of synthesizing compounds of this invention and their synthetic precursors, including those within routes not explicitly shown in schemes herein, are within the means of chemists of ordinary skill in the art. Synthetic chemistry transformations and protecting group methodologies (protection and deprotection) useful in synthesizing the applicable compounds are known in the art and include, for example, those described in Larock R, Comprehensive Organic Transformations, VCH Publishers (1989); Greene TW et al, Protective Groups in Organic Synthesis, 3rd Ed., John Wiley and Sons (1999); Fieser L et al, Fieser and Fieser's Reagents or Organic Synthesis, John Wiley and Sons (1994); and Paquette L, ed.,
Encyclopedia of Reagents for Organic Synthesis, John Wiley and Sons (1995) and subsequent editions thereof.
Combinations of substituents and variables envisioned by this invention are only those that result in the formation of stable compounds.
COMPOSITIONS
The invention also provides pyrogen-free compositions comprising an effective amount of a compound of this invention or pharmaceutically acceptable salts thereof; and an acceptable carrier. Preferably, a composition of this invention is formulated for pharmaceutical use ("a pharmaceutical composition"), wherein the carrier is a pharmaceutically acceptable carrier. The carrier(s) are "acceptable" in the sense of being compatible with the other ingredients of the formulation and, in the case of a pharmaceutically acceptable carrier, not deleterious to the recipient thereof in an amount used in the medicament.
Pharmaceutically acceptable carriers, adjuvants and vehicles that may be used in the pharmaceutical compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, microcrystalline cellulose, cellulose- based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene -polyoxypropylene-block polymers, polyethylene glycol and wool fat.
If required, the solubility and bioavailability of the compounds of the present invention in pharmaceutical compositions may be enhanced by methods well-known in the art. One method includes the use of lipid excipients in the formulation. See "Oral Lipid-Based Formulations: Enhancing the Bioavailability of Poorly Water- Soluble Drugs (Drugs and the Pharmaceutical Sciences)," David J. Hauss, ed.
Informa Healthcare, 2007; and "Role of Lipid Excipients in Modifying Oral and Parenteral Drug Delivery: Basic Principles and Biological Examples," Kishor M. Wasan, ed. Wiley-Interscience, 2006.
Another known method of enhancing bioavailability is the use of an amorphous form of a compound of this invention optionally formulated with a poloxamer, such as LUTROL™ and PLURONIC™ (BASF Corporation), or block copolymers of ethylene oxide and propylene oxide. See United States patent 7,014,866; and United States patent publications 20060094744 and 20060079502.
The pharmaceutical compositions of the invention include those suitable for oral, rectal, nasal, topical (including buccal and sublingual), vaginal or parenteral (including subcutaneous, intramuscular, intravenous and intradermal)
administration. In certain embodiments, the compound of the formulae herein is administered transdermally (e.g., using a transdermal patch or iontophoretic techniques). Other formulations may conveniently be presented in unit dosage form, e.g., tablets, sustained release capsules, and in liposomes, and may be prepared by any methods well known in the art of pharmacy. See, for example, Remington's Pharmaceutical Sciences, Mack Publishing Company, Philadelphia, PA (17th ed. 1985).
Such preparative methods include the step of bringing into association with the molecule to be administered ingredients such as the carrier that constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active ingredients with liquid carriers, liposomes or finely divided solid carriers, or both, and then, if necessary, shaping the product.
In certain embodiments, the compound is administered orally. Compositions of the present invention suitable for oral administration may be presented as discrete units such as capsules, sachets, or tablets each containing a predetermined amount of the active ingredient; a powder or granules; a solution or a suspension in an aqueous liquid or a non-aqueous liquid; an oil-in-water liquid emulsion; a water-in-oil liquid emulsion; packed in liposomes; or as a bolus, etc. Soft gelatin capsules can be useful for containing such suspensions, which may beneficially increase the rate of compound absorption.
In the case of tablets for oral use, carriers that are commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried cornstarch. When aqueous suspensions are administered orally, the active ingredient is combined with emulsifying and suspending agents. If desired, certain sweetening and/or flavoring and/or coloring agents may be added.
Compositions suitable for oral administration include lozenges comprising the ingredients in a flavored basis, usually sucrose and acacia or tragacanth; and pastilles comprising the active ingredient in an inert basis such as gelatin and glycerin, or sucrose and acacia.
Compositions suitable for parenteral administration include aqueous and non-aqueous sterile injection solutions which may contain anti-oxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampules and vials, and may be stored in a freeze dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets.
Such injection solutions may be in the form, for example, of a sterile injectable aqueous or oleaginous suspension. This suspension may be formulated according to techniques known in the art using suitable dispersing or wetting agents (such as, for example, Tween 80) and suspending agents. 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. Among the acceptable vehicles and solvents that may be employed are mannitol, water, Ringer's solution and isotonic sodium chloride solution. 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. Fatty acids, such as oleic acid and its glyceride derivatives are useful in the preparation of injectables, as are natural pharmaceutically- acceptable oils, such as olive oil or castor oil, especially in their polyoxyethylated versions. These oil solutions or suspensions may also contain a long-chain alcohol diluent or dispersant.
The pharmaceutical compositions of this invention may be administered in the form of suppositories for rectal administration. These compositions can be prepared by mixing a compound of this invention with a suitable non-irritating excipient which is solid at room temperature but liquid at the rectal temperature and therefore will melt in the rectum to release the active components. Such materials include, but are not limited to, cocoa butter, beeswax and polyethylene glycols.
The pharmaceutical compositions of this invention may be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well-known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and/or other solubilizing or dispersing agents known in the art. See, e.g.: Rabinowitz, JD and Zaffaroni, AC, US Patent 6,803,031, assigned to Alexza Molecular Delivery Corporation.
Topical administration of the pharmaceutical compositions of this invention is especially useful when the desired treatment involves areas or organs readily accessible by topical application. For topical application topically to the skin, the pharmaceutical composition should be formulated with a suitable ointment containing the active components suspended or dissolved in a carrier. Carriers for topical administration of the compounds of this invention include, but are not limited to, mineral oil, liquid petroleum, white petroleum, propylene glycol, polyoxyethylene polyoxypropylene compound, emulsifying wax, and water.
Alternatively, the pharmaceutical composition can be formulated with a suitable lotion or cream containing the active compound suspended or dissolved in a carrier. Suitable carriers include, but are not limited to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol, and water. The pharmaceutical compositions of this invention may also be topically applied to the lower intestinal tract by rectal suppository formulation or in a suitable enema formulation. Topically-transdermal patches and iontophoretic administration are also included in this invention.
Application of the subject therapeutics may be local, so as to be administered at the site of interest. Various techniques can be used for providing the subject compositions at the site of interest, such as injection, use of catheters, trocars, projectiles, pluronic gel, stents, sustained drug release polymers or other device which provides for internal access.
Thus, according to yet another embodiment, the compounds of this invention may be incorporated into compositions for coating an implantable medical device, such as prostheses, artificial valves, vascular grafts, stents, or catheters. Suitable coatings and the general preparation of coated implantable devices are known in the art and are exemplified in US Patents 6,099,562; 5,886,026; and 5,304,121. The coatings are typically biocompatible polymeric materials such as a hydrogel polymer, polymethyldisiloxane, polycaprolactone, polyethylene glycol, polylactic acid, ethylene vinyl acetate, and mixtures thereof. The coatings may optionally be further covered by a suitable topcoat of fluorosilicone, polysaccharides,
polyethylene glycol, phospholipids or combinations thereof to impart controlled release characteristics in the composition. Coatings for invasive devices are to be included within the definition of pharmaceutically acceptable carrier, adjuvant or vehicle, as those terms are used herein.
According to another embodiment, the invention provides a method of coating an implantable medical device comprising the step of contacting said device with the coating composition described above. It will be obvious to those skilled in the art that the coating of the device will occur prior to implantation into a mammal.
According to another embodiment, the invention provides a method of impregnating an implantable drug release device comprising the step of contacting said drug release device with a compound or composition of this invention.
Implantable drug release devices include, but are not limited to, biodegradable polymer capsules or bullets, non-degradable, diffusible polymer capsules and biodegradable polymer wafers.
According to another embodiment, the invention provides an implantable medical device coated with a compound or a composition comprising a compound of this invention, such that said compound is therapeutically active. According to another embodiment, the invention provides an implantable drug release device impregnated with or containing a compound or a composition comprising a compound of this invention, such that said compound is released from said device and is therapeutically active.
Where an organ or tissue is accessible because of removal from the patient, such organ or tissue may be bathed in a medium containing a composition of this invention, a composition of this invention may be painted onto the organ, or a composition of this invention may be applied in any other convenient way.
In another embodiment, the compound of the inveniton comprises between 28 and 68% (w/w) of the composition. In this embodiment, magnesium stearate and microcrystalline cellulose comprise about 2% (w/w) of the composition.
In another embodiment, a composition of this invention further comprises a second therapeutic agent. The second therapeutic agent may be selected from any compound or therapeutic agent known to have or that demonstrates advantageous properties when administered with a compound having the same mechanism of action as pentoxifylline. Such agents include those indicated as being useful in combination with pentoxifylline, including but not limited to, those described in WO 1997019686, EP 0640342, WO 2003013568, WO 2001032156, WO 2006035418, and WO 1996005838.
Preferably, the second therapeutic agent is an agent useful in the treatment or prevention of a disease or condition selected from peripheral obstructive vascular disease; glomerulonephritis; nephrotic syndrome; nonalcoholic steatohepatitis; Leishmaniasis; cirrhosis; liver failure; Duchenne's muscular dystrophy; late radiation induced injuries; radiation induced lymphedema; radiation-associated necrosis; alcoholic hepatitis; radiation-associated fibrosis; necrotizing enterocolitis in premature neonates; diabetic nephropathy, hypertension-induced renal failure, and other chronic kidney disease; Focal Segmental Glomerulosclerosis; pulmonary sarcoidosis; recurrent aphthous stomatitis; chronic breast pain in breast cancer patients; brain and central nervous system tumors; malnutrition-inflammation- cachexia syndrome; interleukin-1 mediated disease; graft versus host reaction and other allograft reactions; diet-induced fatty liver conditions, atheromatous lesions, fatty liver degeneration and other diet-induced high fat or alcohol-induced tissue- degenerative conditions; human immunodeficiency virus type 1 (HIV-1) and other human retroviral infections; multiple sclerosis; cancer; fibroproliferative diseases; fungal infection; drug-induced nephrotoxicity; collagenous colitis and other diseases and/or conditions characterized by elevated levels of platelet derived growth factor (PDGF) or other inflammatory cytokines; endometriosis; optic neuropathy and CNS impairments associated with acquired immunodeficiency syndrome (AIDS), immune disorder diseases, or multiple sclerosis; autoimmune disease; upper respiratory viral infection; depression; urinary incontinence; irritable bowel syndrome; septic shock; Alzheimers Dementia; neuropathic pain; dysuria; retinal or optic nerve damage; peptic ulcer; insulin-dependent diabetes; non-insulin-dependent diabetes; diabetic nephropathy; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension; hyperlipidemia;
hyperuricemia; gout; hypercoagulability; and inflammation or injury associated with neutrophil chemotaxis and/or degranulation. The compounds of this invention can also be used to control intraocular pressure or to stabilize auto-regulation of cerebral blood flow in subjects who require such control as determined by medical examination.
In one embodiment, the second therapeutic agent is selected from a- tocopherol and hydroxyurea.
In another embodiment, the second therapeutic agent is useful in the treatment of diabetes or an associated disorder, and is selected from insulin or insulin analogues, glucagon- like-peptide-1 (GLP-1) receptor agonists, sulfonylurea agents, biguanide agents, alpha-glucosidase inhibitors, PPAR agonists, meglitinide agents, dipeptidyl-peptidase (DPP) IV inhibitors, other phosphodiesterase (PDE1, PDE5, PDE9, PDE10 or PDE1) inhibitors, amylin agonists, CoEnzyme A inhibitors, and antiobesity agents.
Specific examples of insulin include, but are not limited to Humulin® (human insulin, rDNA origin), Novolin® (human insulin, rDNA origin), Velosulin® BR (human buffered regular insulin, rDNA origin), Exubera® (human insulin, inhaled), and other forms of inhaled insulin, for instance, as delivered by
Mannkind's "Technosphere Insulin System".
Specific examples of insulin analogues include, but are not limited to, novarapid, insulin detemir, insulin lispro, insulin glargine, insulin zinc suspension and Lys-Pro insulin.
Specific examples of Glucagon-Like -Peptide- 1 receptor agonists include, but are not limited to BIM-51077 (CAS-No. 275371-94-3), EXENATIDE (CAS-No. 141758-74-9), CJC-1131 (CAS-No. 532951 -64-7), LIRAGLUTIDE (CAS-No. 20656-20-2) and ZP-10 (CAS-No. 320367-13-3).
Specific examples of sulfonylurea agents include, but are not limited to, TOLBUTAMIDE (CAS- No. 000064-77-7), TOLAZAMIDE (CAS-No. 001156-19- 0), GLIPIZIDE (CAS-No. 029094-61-9), CARBUT AMIDE (CAS-No. 000339-43- 5), GLISOXEPIDE (CAS-No. 025046-79-1), GLISENTIDE (CAS-No. 032797-92- 5), GLIBORNURIDE (CAS-No. 026944-48-9), GLIBENCLAMIDE (CAS-NO. 010238-21 -8), GLIQUIDONE (CAS-No. 033342-05-1), GLIMEPIRIDE (CAS-No. 093479-97-1) and GLICLAZIDE (CAS-No. 021187-98-4).
A specific example of a biguanide agent includes, but is not limited to METFORMIN (CAS-No. 000657-24-9).
Specific examples of alpha-glucosidase-inhibitors include, but are not limited to ACARBOSE (Cas-No. 056180-94-0), MIGLITOL (CAS-No. 072432-03-2) and VOGLIBOSE (CAS-No. 083480-29-9).
Specific examples of PPAR-agonists include, but are not limited to
MURAGLITAZAR (CAS-No. 331741 -94-7), ROSIGLITAZONE (CAS-NO.
122320-73-4), PIOGLITAZONE (CAS-No.111025-46-8), RAGAGLITAZAR (CAS-NO. 222834-30-2), FARGLITAZAR (CAS-No. 196808-45-4),
TESAGLITAZAR (CAS- No. 251565-85-2), NAVEGLITAZAR (CAS-No.
476436-68-7), NETOGLITAZONE (CAS-NO. 161600-01 -7), RIVOGLITAZONE (CAS-NO. 185428-18-6), K-l 11 (CAS-No. 221564-97-2), GW-677954 (CAS-No. 622402-24-8), FK-614 (CAS-No 193012-35-0) and (-)-Halofenate (CAS-No.
024136-23-0). Preferred PPAR- agonists are ROSGLITAZONE and
PIOGLITAZONE.
Specific examples of meglitinide agents include, but are not limited to REPAGLINIDE (CAS-No. 135062-02-1 ), NATEGLINIDE (CAS-No. 105816-04- 4) and MITIGLINIDE (CAS-No. 145375-43-5).
Specific examples of DPP IV inhibitors include, but are not limited to SITAGLIPTIN (CAS-No. 486460-32-6), SAXAGLIPTIN (CAS-No. 361442-04-8), VILDAGLIPTIN (CAS-No. 274901 -16-5), DENAGLIPTIN (CAS-No. 483369-58- 0), P32/98 (CAS-No. 251572-70-0) and NVP-DPP-728 (CAS-No. 247016-69-9).
Specific examples of PDE5 inhibitors include, but are not limited to
SILDENAFIL (CAS-No. 139755-83-2), VARDENAFIL (CAS-No. 224785-90-4) and TADALAFIL (CAS-No. 171596-29-5). Examples of PDE1, PDE9, PDE10 or PDE11 inhibitors which may be usefully employed according to the present invention can be found, for example, in US20020160939, WO2003037432,
US2004220186, WO2005/003129, WO2005012485, WO2005120514 and
WO03077949.
A specific example of an amylin agonist includes, but is not limited to PRAMLINITIDE (CAS-No. 151126-32-8).
A specific example of a Coenzyme A inhibitor includes, but is not limited to ETOMOXIR (CAS- No. 082258-36-4).
Specific examples of anti-obesity drugs include, but are not limited to HMR- 1426 (CAS-No. 262376-75-0), CETILISTAT (CAS-No. 282526-98-1) and
SIBUTRAMINE (CAS-No. 106650-56-0).
In another embodiment, the invention provides separate dosage forms of a compound of this invention and one or more of any of the above-described second therapeutic agents, wherein the compound and second therapeutic agent are associated with one another. The term "associated with one another" as used herein means that the separate dosage forms are packaged together or otherwise attached to one another such that it is readily apparent that the separate dosage forms are intended to be sold and administered together (within less than 24 hours of one another, consecutively or simultaneously).
In the pharmaceutical compositions of the invention, the compound of the present invention is present in an effective amount. As used herein, the term
"effective amount" refers to an amount which, when administered in a proper dosing regimen, is sufficient to treat (therapeutically or prophylactically) the target disorder. For example, and effective amount is sufficient to reduce or ameliorate the severity, duration or progression of the disorder being treated, prevent the advancement of the disorder being treated, cause the regression of the disorder being treated, or enhance or improve the prophylactic or therapeutic effect(s) of another therapy.
The interrelationship of dosages for animals and humans (based on milligrams per meter squared of body surface) is described in Freireich et al., Cancer Chemother. Rep, 1966, 50: 219. Body surface area may be determined
approximately from height and weight of the patient. See, e.g., Scientific Tables, Geigy Pharmaceuticals, Ardsley, N.Y., 1970, 537.
In one embodiment, an effective amount of a compound of this invention is in the range of 20 mg to 2000 mg per treatment. In more specific embodiments the amount is in the range of 40 mg to 1000 mg, or in the range of 100 mg to 800 mg, or more specifically in the range of 200 mg to 400 mg per treatment. Treatment typically is administered from one to three times daily.
Effective doses will also vary, as recognized by those skilled in the art, depending on the diseases treated, the severity of the disease, the route of administration, the sex, age and general health condition of the patient, excipient usage, the possibility of co-usage with other therapeutic treatments such as use of other agents and the judgment of the treating physician. For example, guidance for selecting an effective dose can be determined by reference to the prescribing information for pentoxifylline.
For pharmaceutical compositions that comprise a second therapeutic agent, an effective amount of the second therapeutic agent is between about 20% and 100% of the dosage normally utilized in a monotherapy regime using just that agent.
Preferably, an effective amount is between about 70% and 100% of the normal monotherapeutic dose. The normal monotherapeutic dosages of these second therapeutic agents are well known in the art. See, e.g., Wells et al, eds.,
Pharmacotherapy Handbook, 2nd Edition, Appleton and Lange, Stamford, Conn. (2000); PDR Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, Deluxe Edition, Tarascon Publishing, Loma Linda, Calif. (2000), each of which references are incorporated herein by reference in their entirety.
It is expected that some of the second therapeutic agents referenced above will act synergistically with the compounds of this invention. When this occurs, it will allow the effective dosage of the second therapeutic agent and/or the compound of this invention to be reduced from that required in a monotherapy. This has the advantage of minimizing toxic side effects of either the second therapeutic agent of a compound of this invention, synergistic improvements in efficacy, improved ease of administration or use and/or reduced overall expense of compound preparation or formulation.
METHODS OF TREATMENT
In one embodiment, the invention provides a method of inhibiting the activity of phosphodiesterase (PDE) in a cell, comprising contacting a cell with one or more compounds of Formula A, Al, I, II, B, C, or D.
In addition to its PDE inhibitory activity, pentoxifylline is known to suppress the production of a number of other biological agents such as interleukin-1 (IL-1), IL-6, IL-12, TNF-alpha, fibrinogen, and various growth factors. Accordingly, in another embodiment, the invention provides a method of suppressing the production of interleukin-1 (IL-1), IL-6, IL-12, TNF-alpha, fibrinogen, and various growth factors in a cell, comprising contacting a cell with one or more compounds of Formula A, Al, I, II, B, C, or D.
According to another embodiment, the invention provides a method of treating a disease in a patient in need thereof that is beneficially treated by pentoxifylline comprising the step of administering to said patient an effective amount of a compound of Formula A, Al, I, II, B, C, or D or a pharmaceutical composition comprising a compound of Formula A, Al, I, II, B, C, or D and a pharmaceutically acceptable carrier.
Such diseases are well known in the art and are disclosed in, but not limited to the following patents and published applications: WO 1988004928, EP 0493682, US 5112827, EP 0484785, WO 1997019686, WO 2003013568, WO 2001032156, WO 1992007566, WO 1998055110, WO 2005023193, US 4975432, WO
1993018770, EP 0490181, and WO 1996005836. Such diseases include, but are not limited to, peripheral obstructive vascular disease; glomerulonephritis; nephrotic syndrome; nonalcoholic steatohepatitis; Leishmaniasis; cirrhosis; liver failure; Duchenne's muscular dystrophy; late radiation induced injuries; radiation induced lymphedema; radiation-associated necrosis; alcoholic hepatitis; radiation-associated fibrosis; necrotizing enterocolitis in premature neonates; diabetic nephropathy, hypertension-induced renal failure, and other chronic kidney disease; Focal
Segmental Glomerulosclerosis; pulmonary sarcoidosis; recurrent aphthous stomatitis; chronic breast pain in breast cancer patients; brain and central nervous system tumors; malnutrition-inflammation-cachexia syndrome; interleukin-1 mediated disease; graft versus host reaction and other allograft reactions; diet- induced fatty liver conditions, atheromatous lesions, fatty liver degeneration and other diet-induced high fat or alcohol-induced tissue-degenerative conditions;
human immunodeficiency virus type 1 (HIV-1) and other human retroviral infections; multiple sclerosis; cancer; fibroproliferative diseases; fungal infection; drug-induced nephrotoxicity; collagenous colitis and other diseases and/or conditions characterized by elevated levels of platelet derived growth factor (PDGF) or other inflammatory cytokines; endometriosis; optic neuropathy and CNS impairments associated with acquired immunodeficiency syndrome (AIDS), immune disorder diseases, or multiple sclerosis; autoimmune disease; upper respiratory viral infection; depression; urinary incontinence; irritable bowel syndrome; septic shock; Alzheimers Dementia; neuropathic pain; dysuria; retinal or optic nerve damage; peptic ulcer; insulin-dependent diabetes; non-insulin-dependent diabetes; diabetic nephropathy; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension; hyperlipidemia;
hyperuricemia; gout; hypercoagulability; acute alcoholic hepatitis; olfaction disorders; patent ductus arteriosus; and inflammation or injury associated with neutrophil chemotaxis and/or degranulation.
The compounds of Formula A, Al, I, II, B, C, or D can also be used to control intraocular pressure or to stabilize auto-regulation of cerebral blood flow in subjects who require such control as determined by medical examination.
In one particular embodiment, the method of this invention is used to treat a disease or condition in a patient in need thereof selected from intermittent claudication on the basis of chronic occlusive arterial disease of the limbs and other peripheral obstructive vascular diseases; glomerulonephritis; Focal Segmental Glomerulosclerosis; nephrotic syndrome; nonalcoholic steatohepatitis;
Leishmaniasis; cirrhosis; liver failure; Duchenne's muscular dystrophy; late radiation induced injuries; radiation induced lymphedema; alcoholic hepatitis;
radiation-induced fibrosis; necrotizing enterocolitis in premature neonates; diabetic nephropathy, hypertension-induced renal failure and other chronic kidney diseases; pulmonary sarcoidosis; recurrent aphthous stomatitis; chronic breast pain in breast cancer patients; brain and central nervous system tumors; obesity; acute alcoholic hepatitis; olfaction disorders; endometriosis-associated infertility; malnutrition- inflammation-cachexia syndrome; and patent ductus arteriosus.
In one embodiment, the method of this invention is used to treat diabetic nephropathy, hypertensive nephropathy or intermittent claudication on the basis of chronic occlusive arterial disease of the limbs. In one specific aspect of this embodiment, the method of this invention is used to treat diabetic nephropathy.
In another particular embodiment, the method of this invention is used to treat a disease or condition in a patient in need thereof selected from intermittent claudication on the basis of chronic occlusive arterial disease of the limbs.
In one embodiment, the method of this invention is used to treat chronic kidney disease. The chronic kidney disease may be selected from glomerulonephritis, focal segmental glomerulosclerosis, nephrotic syndrome, reflux uropathy, or polycystic kidney disease.
In one embodiment, the method of this invention is used to treat chronic disease of the liver. The chronic disease of the liver may be selected from nonalcoholic steatohepatitis, fatty liver degeneration or other diet-induced high fat or alcohol-induced tissue-degenerative conditions, cirrhosis, liver failure, or alcoholic hepatitis.
In one embodiment, the method of this invention is used to a diabetes-related disease or condition. This disease may be selected from insulin resistance, retinopathy, diabetic ulcers, radiation-associated necrosis, acute kidney failure or drug-induced nephrotoxicity.
In one embodiment, the method of this invention is used to treat a patient suffering from cystic fibrosis, including those patients suffering from chronic Pseudomonas bronchitis.
In one embodiment, the method of this invention is used to aid in wound healing. Examples of types of wounds that may be treated include venous ulcers, diabetic ulcers and pressure ulcers.
In another particular embodiment, the method of this invention is used to treat a disease or condition in a patient in need thereof selected from insulin dependent diabetes; non-insulin dependent diabetes; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension;
hyperlipidemia; hyperuricemia; gout; and hypercoagulability.
In one embodiment, the method of this invention is used to treat a disease or condition in a patient in need thereof wherein the disease or condition is selected from anemia, Graves disease, retinal vein occlusion, lupus nephritis, macular degeneration, myelodysplasia, pruritis of HIV origin, pulmonary hypertension, retinal artery occlusion, intestinal inflammation, ischemic optic neuropathy, acute pancreatitis, sickle cell anemia and beta thalassemia.
Methods delineated herein also include those wherein the patient is identified as in need of a particular stated treatment. Identifying a patient in need of such treatment can be in the judgment of a patient or a health care professional and can be subjective (e.g. opinion) or objective (e.g. measurable by a test or diagnostic method).
In another embodiment, any of the above methods of treatment comprises the further step of co-administering to the patient one or more second therapeutic agents. The choice of second therapeutic agent may be made from any second therapeutic agent known to be useful for co-administration with pentoxifylline. The choice of second therapeutic agent is also dependent upon the particular disease or condition to be treated. Examples of second therapeutic agents that may be employed in the methods of this invention are those set forth above for use in combination
compositions comprising a compound of this invention and a second therapeutic agent.
In particular, the combination therapies of this invention include coadministering a compound of Formula A, Al, I, II, B, C, or D and a second therapeutic agent for treatment of the following conditions (with the particular second therapeutic agent indicated in parentheses following the indication): late radiation induced injuries (a-tocopherol), radiation-induced fibrosis (a-tocopherol), radiation induced lymphedema (a-tocopherol), chronic breast pain in breast cancer patients (a-tocopherol), type 2 diabetic nephropathy (captopril), malnutrition- inflammation-cachexia syndrome (oral nutritional supplement, such as Nepro; and oral anti-inflammatory module, such as Oxepa); and brain and central nervous system tumors (radiation therapy and hydroxyurea).
The combination therapies of this invention also include co-administering a compound of Formula A, Al, I, II, B, C, or D and a second therapeutic agent for treatment of insulin dependent diabetes; non-insulin dependent diabetes; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension; hyperlipidemia; hyperuricemia; gout; and hypercoagulability.
The term "co-administered" as used herein means that the second therapeutic agent may be administered together with a compound of this invention as part of a single dosage form (such as a composition of this invention comprising a compound of the invention and an second therapeutic agent as described above) or as separate, multiple dosage forms. Alternatively, the additional agent may be administered prior to, consecutively with, or following the administration of a compound of this invention. In such combination therapy treatment, both the compounds of this invention and the second therapeutic agent(s) are administered by conventional methods. The administration of a composition of this invention, comprising both a compound of the invention and a second therapeutic agent, to a patient does not preclude the separate administration of that same therapeutic agent, any other second therapeutic agent or any compound of this invention to said patient at another time during a course of treatment.
Effective amounts of these second therapeutic agents are well known to those skilled in the art and guidance for dosing may be found in patents and published patent applications referenced herein, as well as in Wells et al, eds.,
Pharmacotherapy Handbook, 2nd Edition, Appleton and Lange, Stamford, Conn. (2000); PDR Pharmacopoeia, Tarascon Pocket Pharmacopoeia 2000, Deluxe Edition, Tarascon Publishing, Loma Linda, Calif. (2000), and other medical texts. However, it is well within the skilled artisan's purview to determine the second therapeutic agent's optimal effective-amount range.
In one embodiment of the invention, where a second therapeutic agent is administered to a subject, the effective amount of the compound of this invention is less than its effective amount would be where the second therapeutic agent is not administered. In another embodiment, the effective amount of the second therapeutic agent is less than its effective amount would be where the compound of this invention is not administered. In this way, undesired side effects associated with high doses of either agent may be minimized. Other potential advantages (including without limitation improved dosing regimens and/or reduced drug cost) will be apparent to those of skill in the art.
In yet another aspect, the invention provides the use of a compound of Formula A, Al, I, II, B, C, or D alone or together with one or more of the above- described second therapeutic agents in the manufacture of a medicament, either as a single composition or as separate dosage forms, for treatment or prevention in a patient of a disease, disorder or symptom set forth above. Another aspect of the invention is a compound of Formula A, Al, I, II, B, C, or D for use in the treatment or prevention in a patient of a disease, disorder or symptom thereof delineated herein.
SYNTHETIC EXAMPLES
The synthetic examples below provide detailed procedures for making certain compounds of this invention. It will be apparent to one skilled in the art that further compounds of this invention may be prepared through the use of other reagents or intermediates by reference to these procedures and the schemes described above. The prepared compounds were analyzed by NMR, mass spectrometry, and/or elemental analysis as indicated. 1HNMR were taken on a 300 MHz instrument, which was useful for determining deuterium incorporation. Unless otherwise stated, the absence of an NMR signal as noted in the examples below indicates a level of deuterium incorporation that is at least 90%.
Example 1. Synthesis of 3-Methyl-7-(methyl-<ij)- 1 -(5-oxohexyD- 1H- purine-2,6(3H,7H)-dione (Compound 100).
Scheme 13. Preparation of Compounds 100 and 409.
Figure imgf000060_0001
Step 1. 3-Methyl-7-(methyl- j -lH-purine-2,6(3HJH)-dione (51). A suspension of 3-methylxanthine 50 (5.0 g, 30.1 mmol, 1 equiv) and powdered K2C03 (5.0 g, 36.0 mmol, 1.2 equiv) in DMF (95 mL) was heated to 60 °C and iodomethane-d3 (Cambridge Isotopes, 99.5 atom% D, 2.2 mL, 36.0 mmol, 1.2 equiv) was added via syringe. The resulting mixture was heated at 80 °C for 5 hours (h). The reaction mixture was cooled to room temperature (rt) and the DMF was evaporated under reduced pressure. The crude residue was dissolved in 5% aqueous NaOH (50 mL), resulting in a dull yellow solution. The aqueous solution was washed with CH2Cl2three times (500 mL total). The aqueous layer was acidified to pH 5 with acetic acid (6 mL), resulting in formation of a tan precipitate. The mixture was cooled in an ice-water bath, and the solids were filtered and washed with cold water. The solid was dried in a vacuum oven to give 2.9 g of 51 as a tan solid. The filtrate was concentrated to approximately 25 mL and a second crop (0.70 g) of 51 was collected by filtration. The total yield of 51 was 3.6 g. The crude material was used without further purification.
Step 2. 3-Methyl-7-(methyl- ^-l-(5-oxohexyn-lH-purine-2,6(3HJH)- dione (Compound 100). Crude 51 (1.50 g, 8.2 mmol, 1 equiv) and powdered K2CO3 (2.28 g, 16.4 mmol, 2 equiv) were suspended in DMF (30 mL) and heated to 50 °C. To the resulting tan suspension was added 6-chloro-2-hexanone (52, 1.2 mL, 9.0 mmol, 1.1 equiv) and the reaction temperature was raised to 130 °C. Heating was continued at 130 °C for 2 h, during which time the suspension became finer and darker in color. The reaction mixture was cooled to rt and DMF was evaporated under reduced pressure. The residual tan paste was suspended in EtOAc (250 mL) and filtered to remove insoluble material. The filtrate was concentrated under reduced pressure resulting in a yellow oil. The crude product was purified using an Analogix chromatography system eluting with 100% EtOAc (10 minutes) followed by a gradient of 0 to 25% MeOH/EtOAc over 50 minutes (min). Product fractions were concentrated under reduced pressure to give a slightly yellow oil that solidified after standing for several minutes. The solid was triturated with heptanes (100 mL) and filtered to give 2.00 g of 100 as an off-white solid, mp 101.8-103.0 °C. 1H- NMR (300 MHz, CDC13): δ 1.64-1.68 (m, 4H), 2.15 (s, 3H), 2.51 (t, J = 7.0, 2H), 3.57 (s, 3H), 4.01 (t, J = 7.0, 2H), 7.52 (s, 1H). 13C-NMR (75 MHz, CDCI3): δ 20.95, 27.41, 29.69, 29.98, 40.80, 43.18, 107.63, 141.41, 148.75, 151.45, 155.26, 208.80. HPLC (method: 20 mm CI 8-RP column - gradient method 2 to 95% ACN + 0.1% formic acid in 3.3 min with 1.7 min hold at 95% ACN; Wavelength: 254 nm): retention time: 2.54 min; 98.5% purity. MS (M+H): 282.0. Elemental Analysis (CisHisDs^Os): Calculated: C=55.50, H=6.45, N=19.92. Found:
C=55.58, H=6.48, N=19.76.
Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 2. Synthesis of 8- 3-methyl-7-(methyl- j)-l-(6- j-4- 7-5- oxohexyl)-lH-purine-2,6(3H,7H)-dione (Compound 409). 8-^-3-methyl-7-(methyl- j)-l-(6- j-4-(i7-5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 409). A suspension of 100 (1.80 g, 6.4 mmol, 1 equiv) and powdered K2CO3 (0.23 g, 1.7 mmol, 0.25 equiv) in D20 (Cambridge Isotope Labs, 99 atom% D) (45 mL) was stirred under reflux conditions for 24 h during which time the suspension became a slightly yellow solution. The reaction mixture was cooled to rt, saturated with sodium chloride, and extracted four times with dichloromethane (400 mL total). The combined organic solution was dried over Na2S04, filtered, and evaporated under reduced pressure to provide 1.7 g of a slightly yellow oil that solidified upon standing. The crude material was re-subjected to the hydrogen/deuterium exchange conditions described above with fresh K2CO3 and D20. After an identical workup, the off-white solid was triturated with hexanes (100 mL) and filtered to give 1.61 g of 409 as an off white solid, mp 99.6-99.8 °C. 1H-NMR (300 MHz, CDCI3): δ 1.64-1.69 (m, 4H), 3.57 (s, 3H), 4.01 (t, J = 7.0, 2H). 13C-NMR (75 MHz, CDC13): δ 21.05, 27.61, 29.90, 41.02, 107.83, 148.99, 151.69, 155.50, 209.28. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη CI 8-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN; Wavelength: 254 nm): retention time: 3.26 min; 98% purity. MS (M+H): 288.3. Elemental Analysis (CoHpDpN^): Calculated:
C=54.35, H=6.31, N=19.50. Found: C=54.36, H=6.32, N=19.10.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a singlet at around 2.15 ppm indicating an absence of methyl ketone hydrogens; a triplet at around 2.51 ppm indicating an absence of methylene ketone hydrogens; and a singlet at around 7.52 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
The H/D exchange reaction to convert the CH3C(0)CH2 functional group in 100 to the CD3C(0)CD2 functional group in 409 may be generally applied under analogous conditions to convert compounds having one or more hydrogens alpha to the carbonyl group to compounds having one or more deuteriums in place of the corresponding one or more hydrogens. In one embodiment, successive H/D exchange reactions may be performed as needed to further increase the amount of deuterium incorporation. In one aspect of this embodiment, any excess D20 at the end of a second H/D exchange reaction in a given batch run may be used in a first H/D exchange reaction in a subsequent batch run; and any excess D20 at the end of a third H/D exchange reaction in a given batch run may be used in a second H/D exchange reaction in a subsequent batch run.
The Table below shows an exemplary deuteration on three separate batches of a compound having a CH3C(0)CH2 functional group and indicates the deuteration agent (either >99% D20 or an aqueous phase obtained from a later deuteration cycle of another batch) that may be used according to this invention.
Figure imgf000063_0002
of pentoxifylline
Figure imgf000063_0001
to the structure below
Figure imgf000064_0001
, may be effected in successive batches. The following table shows the percentage of deuterium incorporation at each of the methyl(CO), (CO)methylene, and the imidazole ring methine carbon for successive batch runs of 50 kg of pentoxifylline in each batch:
(CO)methy- methine
Batch 1 lene methyl(CO) carbon
Exchange 1 88.6 88.8 13.8
Exchange 2 97.8 98.0 26.3
Exchange 3 99.2 99.2 37.4
Batch 2
Exchange 1 (Rl-2) 82.3 83.8 6.4
Exchange 2 (Rl-3) 96.7 96.8 12.4
Exchange 3 98.9 98.9 26.4
Batch 3
Exchange 1 (R2-2) 78.0 80.8 5.8
Exchange 2 (R2-2) 95.1 95.6 11.6
Exchange 3 98.9 99.0 27.1
Exchange 4* 99.3 99.4 32.6
Rl-2: recycled from Batch 1, Exchange 2
Rl-3: recycled from Batch 1, Exchange 3
R2-2: recycled from Batch 2, Exchange 2
R2-3 : recycled from Batch 2, Exchange 3
In one embodiment, Exchange 4 is optional. In the Batch 3 run shown in the table, Exchange 4 was performed at half- volume to ensure high deuterium incorporation in Batch 3.
Example 3. Synthesis of 3,7-Di(methyl-(i2 -l-(5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 101).
Scheme 14. Preparation of Compounds 101 and 413.
Figure imgf000065_0001
Step 1. 3 ,7-Di(methyl-(ij)-lH-purine-2,6(3H,7H -dione (55). A suspension of xanthine 53 (2.00 g, 13.2 mmol, 1.0 equiv) and hexamethyldisilazane (32 mL) in toluene (60 mL) was heated to reflux and stirred for 4 days. The reaction mixture was cooled to room temperature, diluted with additional toluene (50 mL) and filtered through Celite to remove any unreacted starting material. The filtrate was evaporated to dryness under reduced pressure to produce 54 as a white solid (4.1 g). A portion of this material (3.00 g) was placed in a 100 mL sealed tube reaction vessel, followed by the addition of toluene (60 mL) and CD3I (4 mL, Cambridge Isotopes, 99.5 atom% D). The reaction mixture was heated in a 120 °C oil bath and stirred for 24 hours, during which time the reaction mixture turned yellow and a solid formed. The reaction mixture was cooled to room temperature, resulting in the entire reaction mixture solidifying to a yellow solid. The mixture was diluted with acetone (30 mL) and MeOH (5 mL) and filtered under a stream of N2. The solids were washed with acetone (100 mL) which removed the yellow color to afford an off- white solid. The solid was dried on the filter under a stream of N2 to give a mixture of 55 and monoalkylated side product, 7-(methyl-<¾)-xanthine in a roughly 1 : 1 ratio. Total mass recovery was 2.6 g (42% crude yield). Due to the poor solubility of this mixture, it was carried forward without further purification.
Step 2. 3,7-Di(methyl- -l-(5-oxohexyn-lH-purine-2,6(3H,7H)-dione (Compound 101). A suspension of crude 55 (2.50 g, 13.4 mmol, 1.0 equiv) and powdered K2C03 (2.20 g, 16 mmol, 1.2 equiv) in DMF (50 mL) was heated to 60 °C. To the resulting tan suspension was added 6-chloro-2-hexanone 52 (2.0 mL, 14.8 mmol, 1.1 equiv) and the mixture was heated to 140 °C. Heating was continued at 140 °C for 4 hours during which time the suspension became finer and darker in color. The reaction mixture was cooled to room temperature and the DMF was evaporated under reduced pressure. The resulting tan paste was suspended in 1 : 1 dichloromethane/ethyl acetate (200 mL) and filtered to remove insoluble material. The filtrate was concentrated under reduced pressure giving a yellowish-brown oil (3.0 g). This crude reaction product was adsorbed onto silica gel and dry- loaded onto a silica gel column packed with 100% dichloromethane. The column was eluted with a gradient of 0-5% MeOH/dichloromethane. Fractions containing product were concentrated under reduced pressure to give 0.75 g of a yellow oil. LCMS showed the material to be about 90% pure. The yellow oil was further purified using an Analogix chromatography system eluting initially with 60% EtO Ac/heptanes followed by a gradient of 60-100% EtO Ac/heptanes over 20 min. The desired product eluted at about 20 minutes. Fractions containing product were concentrated under reduced pressure to give 0.55 g (16%>) of Compound 101 as a slightly yellow oil which solidified upon standing. 1H-NMR (300 MHz, CDC13): δ 1.64-1.69 (m, 4H), 2.15 (s, 3H), 2.51 (t, J = 7.0, 2H), 4.02 (t, J = 7.0, 2H), 7.51 (s, 1H). 13C-NMR (75 MHz, CDC13): δ 20.97, 27.43, 29.97, 40.80, 43.19, 107.64, 141.40, 148.78, 151.48, 155.29, 208.77. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1%> formic acid; Wavelength: 305 nm): retention time: 3.24 min; 98.6% purity. MS (M+H): 285.3, (M+Na): 307.2.
Elemental Analysis (Ci3Hi2D6N403): Calculated: C=54.92, H=6.38, N=19.71. Found: C=54.90, H=6.40, N=19.50.
Notable in the 1H-NMR spectrum above was the absence of a singlet at around 3.57 ppm indicating an absence of N-methyl hydrogens at the 3 position of the purine ring. Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 4. Synthesis of 8- -3.7-Di(methyl-^Vl-(4.4.6.6.6-^-5- oxohexyl)-lH-purine-2,6(3H,7H)-dione (Compound 413).
8- -3J-Di(methyl- j -l-(4- 2-6-^-5-oxohexyn-lH-purine-2,6(3HJH)- dione (Compound 413). A suspension of Compound 101 (0.60 g, 2.1 mmol, 1.0 equiv) and powdered K2C03 (0.10 g, 0.72 mmol, 0.30 equiv) in D20 (15 mL, Cambridge Isotopes, 99 atom% D) was heated and stirred at reflux for 16 hours during which time the suspension became a slightly yellow solution. The reaction mixture was cooled to room temperature, saturated with sodium chloride, and extracted four times with dichloromethane (200 mL). The combined organic extracts were dried over Na2S04, filtered, and concentrated under reduced pressure to provide 0.53 g of a slightly yellow oil that solidified upon standing. The crude reaction product was re-subjected to the above reaction conditions with fresh powdered K2C03 and D20. After an identical workup, the off-white solid was triturated with hexanes (50 mL) and filtered to give 0.45 g (74%) of Compound 413 as an off-white solid, mp 99.2-99.3 °C. 1H-NMR (300 MHz, CDC13): δ 1.64-1.71 (m, 4H), 4.01 (t, J = 7.0, 2H). 13C-NMR (75 MHz, CDC13): δ 20.85, 27.41, 40.81, 107.63, 148.80, 151.50, 155.31, 209.09. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with a 4 minute hold at 95% ACN + 0.1% formic acid; Wavelength: 254 nm): retention time: 3.25 min; 98.7% purity. MS (M+H): 291.3, (M+Na): 313.2. Elemental Analysis (Ci3H6Di2N403): Calculated: C=53.78, H=6.25, N=19.30. Found: C=53.76, H=6.39, N=19.11.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a singlet at around 2.15 ppm indicating an absence of methyl ketone hydrogens; a triplet at around 2.51 ppm indicating an absence of methylene ketone hydrogens; a singlet around 3.57 ppm indicating an absence of N-methyl hydrogens at the 3 position on the purine ring; and a singlet at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 5. Synthesis of 3-Methyl-7-(methyl- j)-l-(6,6,6- j-5-oxohexyl)- lH-purine-2,6(3H,7H)-dione (Compound 99). Scheme 15. Preparation of Compound 99.
Figure imgf000068_0001
Figure imgf000068_0002
Step 1. 5-(3-Methyl-7-(methyl- ^-2,3.6J-tetrahvdro-lH-purin-l-vn-N- methoxy-N-methylpentanamide (57). A suspension of 51 (1.50 g, 8.2 mmol, 1.0 equiv, see Example 1 for preparation) and powdered K2CO3 (1.80 g, 12.9 mmol, 1.6 equiv) in DMF (40 mL) was heated to 60 °C. 5-Bromo-N-methoxy-N- methylpentanamide 56 (2.21 g, 9.8 mmol, 1.2 equiv, prepared as outlined in Org. Lett., 2005, 7: 1427-1429) was added and the mixture was heated at 110 °C for 4 hours during which time the suspended solid became finer and tan in color. The reaction mixture was cooled to room temperature and DMF was evaporated under reduced pressure. The resulting tan paste was suspended in 1 : 1
Figure imgf000068_0003
acetate (250 mL) and the suspension was filtered to remove insoluble material. The filtrate was concentrated under reduced pressure to a yellow oil. This crude reaction product was purified using an Analogix automated chromatography system eluting with 100% CH2CI2 for 8 minutes followed by a gradient of 0-5% MeOH/ CH2C12 over 40 minutes. The desired product eluted at approximately 24 minutes. Fractions containing product were concentrated under reduced pressure to a slightly yellow oil. 1H NMR of the oil indicated it contained approximately 10% unreacted 51. A second purification on an Analogix automated chromatography system eluting with 100% CH2CI2 for 10 minutes followed by a gradient of 0-5% MeOH/ CH2C12 over 50 minutes allowed for removal of the impurity. Fractions containing product were concentrated under reduced pressure to a slightly yellow oil that crystallized as an off-white solid on standing. The solid was triturated with heptanes (100 mL) and filtered to give 1.29 g (49%) of 57 as an off-white solid.
Step 2. 3-Methyl-7-(methyl-(ij)-l-(6,6,6-(ij-5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 99). A suspension of 57 (0.72 g, 2.2 mmol, 1.0 equiv) in THF (20 mL) was cooled to 2 °C and 1M CD3MgI in ether (2.4 mL, 2.4 mmol, 1.1 equiv, Aldrich >99 atom%> D) was added drop-wise via syringe at a rate to maintain the temperature below 5 °C. During the addition, the mixture became a fine, slightly yellow suspension. When addition was complete, the reaction mixture was warmed to room temperature and was stirred for 3 hours. The mixture was cooled to 2 °C and an additional portion of CD3MgI solution (0.4 mL, 0.4 mmol) was added. The mixture was allowed to warm to room temperature and was stirred an additional 3 hours. The reaction was quenched with IN HC1 (4 mL) and diluted with H20 (10 mL) resulting in a slightly yellow solution that was extracted with CH2C12 (3X, 200 mL). The combined organic extracts were dried over Na2S04, filtered, and concentrated under reduced pressure to a yellow oil. The crude product was purified using an Analogix automated chromatography system eluting with 100% CH2C12 for 8 minutes and then a gradient of 0-5% MeOH/ CH2C12 over 40 minutes. The desired product elutes first at about 22 minutes, followed by unreacted starting material. Fractions containing the desired product were concentrated under reduced pressure to a yellow oil that solidified upon standing. The solid was triturated with hexane (25 mL) and collected via vacuum filtration to give 0.33 g (53%>) of Compound 99 as a white solid, mp 93.7 - 94.4 °C. Fractions containing unreacted starting material were also collected and concentrated to give 0.21 g of 57 as a clear, colorless oil. The recovered material was re-subjected to the above alkylation reaction to give, after workup and purification, an additional 0.06 g (33%, 62% overall based on total starting material) of Compound 99, mp 93.3 - 94.0 °C. 1H-NMR (300 MHz, CDC13): δ 1.64-1.68 (m, 4H), 2.50 (t, J = 7.0, 2H), 3.58 (s, 3H), 4.02 (t, J = 7.0, 2H), 7.51 (s, 1H). 13C-NMR (75 MHz, CDC13): δ 21.16, 27.65, 29.91, 41.03, 43.41, 107.87, 141.62, 149.00, 151.69, 155.50, 209.12. HPLC
(method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1%) formic acid; Wavelength: 305 nm): retention time: 3.24 min; 99.0%> purity. MS (M+H): 285.3, (M+Na): 307.2. Elemental Analysis (Ci3Hi2D6N403): Calculated: C=54.92, H=6.38, N=19.71. Found: C=54.85, H=6.36, N=19.49.
Notable in the 1H-NMR spectrum above was the absence of a singlet at around 2.15 ppm indicating an absence of methyl ketone hydrogens. Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 6. Synthesis of (±)8-^-l-(4,4,6,6,6- -5-Hvdroxyhexyl)-3- methyl-7-(methyl-(ij)-lH-purine-2,6(3H,7H -dione (Compound 419).
Scheme 16. Preparation of Compounds 419, 419(R), and 419(5).
Figure imgf000070_0001
Chiral HPLC
Separation
Figure imgf000070_0002
(±)8-^-l-(4,4,6,6,6-(i -5-Hydroxyhexyl)-3-methyl-7-(methyl-(ij)-lH-purine- 2,6(3H,7H)-dione (Compound 419). Compound 409 (0.50 g, 1.7 mmol, 1.0 equiv, see Example 2) was dissolved in EtOD (13 mL, Aldrich 99.5 atom% D) and NaBH4 (0.07 g, 1.9 mmol, 1.1 equiv) was added. An increase in temperature from 24 to 28 °C was observed. The reaction was stirred 2 hours at room temperature, then was quenched by the addition of D20 (30 mL, Cambridge Isotope Labs, 99 atom% D). A white suspension formed that was extracted with MTBE (4X, 200 mL total). The combined organic extracts were dried over Na2S04, filtered, and concentrated under reduced pressure to a clear, colorless oil (0.45 g). The crude product was purified by silica gel chromatography eluting first with 1% MeOH/ CH2C12 followed by a gradient of 1-5% MeOH/ CH2C12. Fractions containing product were concentrated under reduced pressure to give (0.41 g, 83 %) of Compound 419 as a clear colorless oil that solidified on standing.
Example 7. Chiral Separation of (i?)-8- -l-(4,4,6,6,6- -5-Hydroxyhexyl)- 3-methyl-7-(methyl- ^-lH-purine-2,6(3HJH)-dione (Compound 419(R) and (6^- 8-^-l-(4^,6,6,6-(i -5-Hvdroxyhexyl)-3-methyl-7-(methyl-(ij)-lH-purine- 2,6(3H,7H)-dione (Compound 419(S)).
Separation of Enantiomers of Compound 419. Compound 419 obtained from Example 6 above (0.38 g) was dissolved in a minimal amount of iPrOH (6 mL, HPLC grade, heating required) and diluted with hexane (4 mL, HPLC grade).
Enantiomeric separation was achieved using a Waters HPLC system equipped with a preparative Daicel Chiralpak AD column (20 X 250 mm). For the first minute of the run, the mobile phase was 80% hexane and 20% iPrOH along with 0.1%
diethylamine. After the first minute a gradient to 75% hexane and 25% iPrOH along with 0.1%) diethylamine over 15 minutes was used, followed by holding at this solvent ratio for 17 minutes at a flow rate of 18 mL/min. This method resulted in baseline separation with 419(R) eluting first (21.0 min), followed by 419(5) (24.1 min). Fractions containing each enantiomer were concentrated under reduced pressure to give 0.16 g each of 419(R) (mp 107.8-108.8 °C) and 419(5) (mp 108.3- 108.4 °C) as off-white solids.
A). (R)-8-dL- 1 -(4,4,6,6,6- -5-Hvdroxyhexyn-3-methyl-7-(methyl- j - 1H- purine-2,6(3H,7H)-dione (Compound 419(R)V 1H-NMR (300 MHz, CDC13): δ 1.36-1.50 (m, 2H), 1.60-1.74 (m, 3H), 3.58 (s, 3H), 3.80 (s, 1H), 4.02 (t, J = 7.3, 2H). 13C-NMPv (75 MHz, CDC13): δ 22.70, 27.86, 29.71 , 41.14, 67.66, 107.66, 148.78, 151.54, 155.40. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95%> ACN + 0.1%> formic acid; Wavelength: 254 nm): retention time: 3.26 min; 99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min; Wavelength: 254 nm): retention time: 27.51 min (major enantiomer); 31.19 min (expected for minor enantiomer): >99.9% ee purity. MS (M+H): 290.1, (M+Na): 312.3. Elemental Analysis (C13H11D9N4O3): Calculated: C=53.97, H=6.97, N=19.36. Found: C=54.39, H=7.11, N=18.98.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a peak at around 1.19 ppm indicating an absence of methyl hydrogens alpha to the hydroxyl group; and a singlet at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a multiplet at 1.36-1.50 ppm and a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence a peak at 1.51 ppm corresponding to the presence or absence of methylene hydrogens alpha to the hydroxyl group and of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
B . (^-8- -l-(4,4,6,6,6- .-5-Hvdroxyhexyn-3-methyl-7-(methyl- ^-lH- purine-2,6(3HJH)-dione (Compound 419(5 1H-NMR (300 MHz, CDC13): δ 1.41- 1.48 (m, 2H), 1.64-1.72 (m, 3H), 3.58 (s, 3H), 3.79 (s, 1H), 4.02 (t, J = 7.4, 2H). 13C-NMR (75 MHz, CDCI3): δ 22.70, 27.86, 29.71, 41.15, 67.66, 107.67, 148.78, 151.54, 155.41. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη CI 8-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 254 nm): retention time: 3.26 min; 99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min; Wavelength: 254 nm): retention time: 31.19 min (major enantiomer); 27.51 min (expected for minor enantiomer): >99.9% ee purity. MS (M+H): 290.1, (M+Na): 312.3. Elemental Analysis (C13H11D9N4O3): Calculated: C=53.97, H=6.97, N=19.36. Found: C=54.35, H=7.28, N=18.75.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a peak at around 1.19 ppm indicating an absence of methyl hydrogens alpha to the hydroxyl group; and a singlet at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a multiplet at 1.36-1.50 ppm and a triplet at 4.01 ppm in the above 1H-NMR spectrum, determination of the presence or absence a peak at 1.51 ppm corresponding to the presence or absence of methylene hydrogens alpha to the hydroxyl group and of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 8. Synthesis of (±)8- y_-l-(4,4,5,6,6,6-^-5-Hydroxyhexyl)-3- methyl-7-(methyl-(ij)-lH-purine-2,6(3H,7H)-dione (Compound 435).
Scheme 17. Preparation of Compounds 435, 435(R), and 435fS .
Figure imgf000073_0001
(± 8- -l-(4,4,5.6.6.6-A-5-Hvdroxyhexyn-3-methyl-7-(methyl- j -lH- purine-2,6(3H,7H)-dione (Compound 435). To a solution of Compound 409 (0.50 g, 1.7 mmol, 1.0 equiv) in EtOD (13 mL, Aldrich 99.5 atom% D) was added NaBD4 (0.08 g, 1.9 mmol, 1.1 equiv, Cambridge Isotope Labs, 99 atom% D). An increase in temperature from 24 to 27 °C was observed. The reaction was stirred 2 hours at room temperature then was quenched by the addition of of D20 (30 mL) (Cambridge Isotope, 99 atom% D). A white suspension formed that was extracted with MTBE (4X, 200 mL total). The combined organic extracts were dried over Na2S04, filtered, and concentrated under reduced pressure to a clear, colorless oil (0.45 g). The crude product was purified by silica gel chromatography eluting first with 1% MeOH/ CH2C12 followed by a gradient of 1 -5% MeOH/ CH2C12. Fractions containing product were concentrated under reduced pressure to give 0.40 g (81 %) of Compound 435 as a clear colorless oil that solidified on standing.
Example 9. Chiral Separation of (i?)-8- l-(4A5,6,6,6-A-5- Hvdroxyhexyl)-3 -methyl-7-(methyl-<ij)- lH-purine-2,6(3H,7H)-dione (Compound 435(R) and (^-8- -l-(4,4,5,6,6,6-A-5-Hvdroxyhexyn-3-methyl-7-(methyl- j - lH-purine-2,6(3HJH)-dione (Compound 435(S)V Separation of Enantiomers of Compound 435. Compound 435 obtained from Example 8 above (0.32 g) was dissolved in a minimal amount of iPrOH (5 mL, HPLC grade, heating was required) and diluted with hexane (4 mL, HPLC grade). Enantiomer separation was achieved using a Waters HPLC system equipped with a preparative Daicel Chiralpak AD column (20 X 250 mm). For the first minute of the run, the mobile phase was 80% hexane and 20% iPrOH along with 0.1%
diethylamine. After the first minute a gradient to 75% hexane and 25% iPrOH along with 0.1%) diethylamine over 15 minutes was used, followed by holding at this solvent ratio for 17 minutes at a flow rate of 18 mL/min. This method resulted in baseline separation with Compound 435(R) eluting first (21.9 min), followed by Compound 435(5) (25.2 min). Fractions containing each enantiomer were concentrated under reduced pressure to give 0.12 g each of 435(R) (mp 108.0-108.1 °C) and 435(5) (mp 107.6- 107.7 °C) as off-white solids.
A). (i? -8- -l-(4,4,5,6,6,6-A-5-Hvdroxyhexyn-3-methyl-7-(methyl- ^-lH- purine-2,6(3H,7H)-dione (Compound 435(R)V 1H-NMR (300 MHz, CDC13): δ 1.40- 1.48 (m, 3H), 1.66-1.70 (m, 2H), 3.58 (s, 3H), 4.02 (t, J = 7.5, 2H). 13C-NMR (75 MHz, CDC13): δ 22.66, 27.86, 29.71 , 41.15, 107.67, 148.80, 151.54, 155.41. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 254 nm): retention time: 3.25 min; 99.8% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min;
Wavelength: 254 nm): retention time: 27.24 min (major enantiomer); 31.1 1 min (expected for minor enantiomer): >99.9%> ee purity. MS (M+H): 291.3, (M+Na): 313.2. Elemental Analysis (Ci3HioDioN403): Calculated: C=53.78, H=6.94, N=19.30. Found: C=54.01 , H=7.07, N=18.90.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a peak at around 1.19 ppm indicating an absence of methyl hydrogens alpha to the hydroxyl group; a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position; and a singlet at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a multiplet at 1.36-1.50 ppm and a triplet at 4.01 ppm in the above 1H- NMR spectrum, determination of the presence or absence a peak at 1.51 ppm corresponding to the presence or absence of methylene hydrogens alpha to the hydroxyl group and of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
B . (^-8- -l-(4,4,5,6,6,6-A-5-Hvdroxyhexyn-3-methyl-7-(methyl- ^-lH- purine-2,6(3HJH)-dione (Compound 435(5 1H-NMR (300 MHz, CDC13): δ 1.41- 1.48 (m, 3H), 1.62-1.72 (m, 2H), 3.58 (s, 3H), 4.03 (t, J = 7.4, 2H). 13C-NMR (75 MHz, CDC13): δ 22.69, 27.90, 29.70, 41.17, 107.69, 148.82, 151.58, 155.43. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 254 nm): retention time: 3.25 min; 99.5% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min;
Wavelength: 254 nm): retention time: 31.11 min (major enantiomer); 27.24 min (expected for minor enantiomer): >99.9%> ee purity. MS (M+H): 291.3, (M+Na): 313.2. Elemental Analysis (Ci3HioDioN403): Calculated: C=53.78, H=6.94, N=19.30. Found: C=54.01, H=7.11, N=18.78.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a peak at around 1.19 ppm indicating an absence of methyl hydrogens alpha to the hydroxyl group; a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position; and a singlet at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a multiplet at 1.36-1.50 ppm and a triplet at 4.01 ppm in the above 1H- NMR spectrum, determination of the presence or absence a peak at 1.51 ppm corresponding to the presence or absence of methylene hydrogens alpha to the hydroxyl group and of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 10. Synthesis of 8-( -3,7-Dimethyl-l-(4,4,6,6,6-( 5-oxohexyl)- lH-purine-2,6(3HJH)-dione (Compound 407). Scheme 18. Preparation of Compounds 407. 437. 437(R). and 437(S).
Figure imgf000076_0001
8- -3.7-Dimethyl-l -(4.4.6.6.6-^-5-oxohexyn-lH-purine-2.6(3H.7H)-dione (Compound 407). A mixture of commercially-available 58 (7.95 g, 28.6 mmol) and potassium carbonate (990 mg, 7.2 mmol) in D20 (195 mL, Cambridge Isotopes, 99.9 atom% D) was heated to reflux for 24 hours. The suspended solid dissolved gradually giving a yellow solution. The solution was cooled to approximately 40 °C and was concentrated under reduced pressure to a tan solid. The solid was dissolved in D20 (195 mL) and the solution was heated to reflux for another 24 hours. The solution was cooled to room temperature and concentrated under reduced pressure to a tan solid. Ethyl acetate (200 mL) was added and the mixture was stirred 0.5 hours at approximately 40 °C. The insoluble materials were filtered off and the filtrate was concentrated under reduced pressure to a pale yellow solid, which was triturated with MTBE (40 mL) to give 7.5 g (93%) of Compound 407 as an off-white solid. 1H-NMR (300 MHz, CDC13): δ 1.64-1.68 (m, 4H), 3.57 (s, 3H), 3.99 (s, 3H), 3.99- 4.04 (m, 2H). 13C-NMR (75 MHz, CDC13): δ 20.84, 27.40, 29.69, 33.57, 40.81 , 107.62, 148.77, 151.48, 155.28, 209.07. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid;
Wavelength: 305 nm): retention time: 3.24 min; 99.9% purity. MS (M+H): 285.3, (M+Na): 307.2. Elemental Analysis (Ci3Hi2D6N403): Calculated: C=54.92, H=6.38, N=19.71. Found: C=54.89, H=6.38, N=19.70.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a singlet at around 2.15 ppm indicating an absence of methyl ketone hydrogens; a triplet at around 2.51 ppm indicating an absence of methylene ketone hydrogens; and a singlet at around 7.52 ppm indicating an absence of hydrogen at the number 8 position on the purine ring.
Example 11. Synthesis of (± 8-< ,-l-(4A5A6.6-<^-5-HvdroxyhexylV3.7- dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 437).
(±)8-^-l-(4,4,5,6,6,6-( -5-Hydroxyhexyl)-3,7-dimethyl-lH-purine- 2,6(3H,7H)-dione (Compound 437). Sodium borodeuteride (1.06 g, 25.3 mmol, Cambridge Isotopes, 99 atom% D) was added to a suspension of 407 (6.5 g, 22.9 mmol) in ethanol-di (65 mL, Aldrich, 99.5 atom% D) at 0 °C. The mixture was warmed to room temperature and stirred until a clear solution had developed (approximately 1 hour). The reaction was quenched with a saturated solution of ammonium chloride-d4 (Cambridge Isotopes, 98 atom% D) in D20 (8 mL,
Cambridge Isotope, 99.9 atom% D), ethanol-di was evaporated under reduced pressure and the residue was extracted with EtOAc (160 mL). The organic phase was washed with D20 (20 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure to give 4.8 g (73%) of Compound 437 as a pale yellow solid.
Figure imgf000077_0001
Hvdroxyhexyl)-3,7-dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 437(RV) and (^-8- -l-(4,4,5,6,6,6-A-5-Hvdroxyhexyn-3J-dimethyl-lH-purine-2,6(3H,7H - dione (Compound 437(5)).
Separation of Enantiomers of Compound 437. Compound 437 obtained from Example 1 1 above (1.60 g) was dissolved in iPrOH (20 mL, HPLC grade, heating required). Enantiomeric separation was achieved using a Waters HPLC system equipped with a preparative Chiralpak AD column (20 x 250 mm Daicel, 10 μΜ) with a preparative Chiralpak AD guard column (20 x 50 mm Daicel, 10 μΜ) preceding it. For the first minute of the run, the sample was eluted with 20% iPrOH/hexanes (henceforth, with 0.1% diethylamine as co-eluent) while ramping up from a flow rate of 15 mL/min to 18 mL/min. Over the next 15 minutes, the sample was eluted at a flow rate of 18 mL/min with a gradient of 20% to 25%
iPrOH/hexanes. For the next 19 minutes the sample was eluted at a flow rate of 18 mL/min with 25% iPrOH/hexanes. Over the next 0.5 minutes, the sample was eluted at a flow rate of 18 mL/min with a gradient of 25% to 20% iPrOH/hexanes. For the next 4.5 minutes, the sample was eluted at a flow rate of 18 mL/min with 20% iPrOH/hexanes. This elution method resulted in baseline separation of Compound 437(R) eluting first (retention time approximately 29 min) and Compound 437(5) eluting second (retention time approximately 33 min). Fractions containing each enantiomer were collected and concentrated under reduced pressure to give 340 mg of 437(R) (mp 1 12.0-1 14.5 °C) and 375 mg of 437(5) (mp 1 1 1.9-1 12.3 °C) as off- white solids. [Note: only 1.0 g of 437 was injected from the solution prepared above.]
A. (i? -8-( -l-(4.4.5.6.6.6-A-5-Hvdroxyhexyn-3.7-dimethyl-lH-purine- 2,6(3FL7HVdione (Compound 437(R)V 1H-NMR (300 MHz, CDC13): δ 1.36-1.50 (m, 2H), 1.54 (s, 1H), 1.64-1.74 (m, 2H), 3.58 (s, 3H), 3.99 (s, 3H), 4.00-4.05 (m, 2H). 13C-NMR (75 MHz, CDC13): δ 22.66, 27.86, 29.70, 33.59, 41.14, 107.65, 148.76, 151.52, 155.40. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95%> ACN + 0.1%> formic acid; Wavelength: 305 nm): retention time: 3.28 min; 99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.20 min (major enantiomer); 28.39 min (expected for minor enantiomer): >99.9% ee purity. MS (M+H): 288.3, (M+Na): 310.2. Elemental Analysis (Ci3Hi3D7N403): Calculated: C=54.34, H=7.02, N=19.50. Found: C=54.32, H=7.23, N=19.35.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a peak at around 1.19 ppm indicating an absence of methyl hydrogens alpha to the hydroxyl group; a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position; and a singlet peak at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a multiplet at 1.36-1.50 ppm in the above 1H-NMR spectrum, determination of the presence or absence a peak at 1.51 ppm corresponding to the presence or absence of methylene hydrogens alpha to the hydroxyl group was not possible.
B. (t^-8- -l-(4,4,5.6.6.6-A-5-Hvdroxyhexyn-3J-dimethyl-lH-purine- 2.6(3H.7H)-dione (Compound 437(5)). 1H-NMR (300 MHz, CDC13): δ 1.38-1.48 (m, 2H), 1.55 (s, 1H), 1.64-1.72 (m, 2H), 3.58 (s, 3H), 3.99 (s, 3H), 4.00-4.05 (m, 2H). 13C-NMR (75 MHz, CDC13): δ 22.65, 27.84, 29.71, 33.59, 41.13, 107.64, 148.75, 151.52, 155.39. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1%> formic acid; Wavelength: 305 nm): retention time: 3.27 min; 99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min; Wavelength: 254 nm): retention time: 28.39 min (major enantiomer); 25.20 min (expected for minor enantiomer): >99.9% ee purity. MS (M+H): 288.3, (M+Na): 310.2. Elemental Analysis (CisHisD^Os): Calculated: C=54.34, H=7.02, N=19.50. Found: C=54.33, H=7.30, N=19.36.
Notable in the 1H-NMR spectrum above was the absence of the following peaks: a peak at around 1.19 ppm indicating an absence of methyl hydrogens alpha to the hydroxyl group; a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position; and a singlet peak at around 7.51 ppm indicating an absence of hydrogen at the number 8 position on the purine ring. Due to the presence of a multiplet at 1.36-1.50 ppm in the above 1H-NMR spectrum, determination of the presence or absence a peak at 1.51 ppm corresponding to the presence or absence of methylene hydrogens alpha to the hydroxyl group was not possible.
Example 13. Alternative synthesis of (y>-8- -l-(4,4,5,6,6,6-A-5- Hydroxyhexyl)-3,7-dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 437(5)).
Scheme 19. Preparation of Compound 437(5).
Figure imgf000080_0001
r
59 60
Figure imgf000080_0002
437(S)
(6 -8-(i^-l-(4,4,5,6,6,6-(i^-5-Hydroxyhexyl)-3 ,7-dimethyl-lH-purine- 2.6(3H.7HVdione (Compound 437(5 D-Glucono-5-lactone 59 (5 g, 28.09 mmoles) was added in one portion to ice-cold water (35 mL, 0-3°C) and stirred for 10 min. A freshly prepared, ice-cold solution of NaBD4 (0.294 g, 7.02 mmoles, 99%D) in 10 mL of water was added slowly over 10 min. The reaction is slightly exothermic (2° to 10°C) and the pH of the reaction is 7.42. Stirring was continued for 30 min, with the temperature maintained at 0-3°C by cooling. Acetic acid (0.32 mL, 5.61 mmoles) was added and stirring was continued for another 30 min.
The reaction mixture was diluted with 18 mL of water and the solution was heated to 25-30°C. KH2P04 (0.85 g) was added to the mixture and the pH was adjusted to 7 with 4M KOH solution. To the resulting mixture was added (2.5 g, 8.8 mmoles) of Compound 407. A solution of NAD (15 mg), GDH (2.5 mg), KRED 101 (25 mg) in 12.5 mL of 0.1 KH2P04 buffer was added. The resulting solution was stirred at 25-30°C. The pH of the reaction mixture was maintained between 6 and 7 by adding 4M KOH solution drop-wise as needed. HPLC monitoring of the reaction indicated that the reaction was complete after 12 hours with 99.97A% conversion by HPLC analysis. Sodium chloride (12.5 g) was added and stirred for 30 min. The mixture was extracted with ethyl acetate (3 x 25 mL). The organic layer was separated, filtered through a celite pad and concentrated to a small volume (~ 5 vol). Product solids precipitated during the concentration. The slurry was heated at 40-60 °C and heptanes (20 mL) was added over 10 minutes. The slurry was stirred overnight at 20-25 °C and filtered. The wet cake dried at 50 °C for 12 hours to afford Compound 437(5) as a white solid. (2.12 g, 85% yield). The product purity was determined to be >99.5A% by HPLC analysis. A single enantiomer was observed by chiral HPLC analysis. The deuterium incorporation at the methine 5 position was -95% D. HPLC (method: Waters Symmetry 4.6 x 50 mm 3.5 μιη CI 8 column - gradient method: 15% MeOH + 85% 0.1% formic acid in water for 5 min (1.25 mL/min), ramp to 80% MeOH + 20% 0.1% formic acid in water over 5 min, ramp to 15% MeOH + 85% 0.1% formic acid in water over 6 s followed by a 3.9 min hold at 15% MeOH + 85% 0.1% formic acid in water; wavelength: 274 nm): >99.5% purity. Chiral HPLC analysis (method: Chiralpak AD-H 25 cm column - isocratic method 75% n-heptane/ 25% isopropanol for 25 min at 1.25 mL/min; wavelength: 274 nm): retention time: 17.56 min (major enantiomer); 15.5 min (expected for minor enantiomer): >99.95% ee purity.
Example 14. Alternative synthesis of (i?)-8-^-l-(4,4,5,6,6,6-^-5- Hvdroxyhexyl)-3,7-dimethyl-lH- urine-2,6(3H,7H)-dione (Compound 437(RV).
Figure imgf000081_0001
437(R)
(i?)-8-(i -l-(4,4,5,6,6,6-( -5-Hydroxyhexyl)-3,7-dimethyl-lH-purine- 2.6(3H.7HVdione (Compound 437(R)V
Preparation of compound 437(R) from compound 407:
(S)-%-di-l-(4,4, 5,6,6, 6-A-5-Hvdroxyhexyl)-3,7-dimethyl-lH-purine- 2,6(3HJH -dione (Compound 437(R)V D-Glucono-5-lactone 59 (5 g, 28.09 mmoles) was added in one portion to ice-cold water (35 mL, 0-3°C) and stirred for 10 min. A freshly prepared, ice-cold solution of NaBD4 (0.294 g, 7.02 mmoles, 99%D) in 10 mL of water was added slowly over 10 min. The reaction is slightly exothermic (2° to 10°C) and the pH of the reaction is 7.42. Stirring was continued for 30 min, with the temperature maintained at 0-3°C by cooling. Acetic acid (0.32 mL, 5.61 mmoles) was added and stirring was continued for another 30 min.
The reaction mixture was diluted with 18ml mL of water and the solution was heated to 25-30°C. KH2P04 (0.85 g) was added to the mixture and the pH was adjusted to 7 with 4M KOH solution. To this was added 2.5 g (8.8 mmoles) of 407. A solution of NADP (15 mg), GDH (2.5 mg), CRED A311-NADP (25 mg) in 12.5 mL of 0.1 KH2P04 buffer was added. The resulting solution was stirred at 25-30°C. The pH of the reaction mixture was maintained between 6 and 7 by adding 4M KOH solution drop-wise. The reaction was monitored by HPLC and was complete after 12 hours with 99.97% conversion by HPLC. Sodium chloride (12.5 g) was added and stirred for 30 min. The mixture was extracted with ethyl acetate (3 x 25 mL). The organic layer was separated, filtered through celite pad and concentrated to a small volume (~ 5 vol) and product solids were precipitated. Heptanes (20 mL) were added to the slurry (at 40-60°C) over 10 minutes. The slurry was stirred overnight at 20-25°C and filtered. The wet cake was dried at 50°C for 12 hours to afford 437(R) as a white solid. (2.12 g, 85% yield). The isolated product purity was >99.95%> by HPLC and as a single enantiomer by chiral HPLC.
Example 15. Synthesis of (±)1 -(5-dr-5 -Hydroxyhexyl)-3 -methyl-7-(methyl- (ij)-lH-purine-2,6(3H,7H)-dione (Compound 131).
Scheme 20. Preparation of Compounds 131, 13KR), and 131(5).
Figure imgf000082_0001
(± l-(5- -5-Hvdroxyhexyn-3-methyl-7-(methyl- ^-lH-purine-2,6(3H,7H - dione (Compound 131). Following the same general method as for the synthesis of Compound 437 above, Compound 100 (see Example 1) was treated with NaBD4 in EtOH to afford Compound 131.
Example 16. Chiral Separation of (i?)-l-(5-^-5-Hydroxyhexyl)-3-methyl-7- (methyl- j -lH-purine-2,6(3H,7H -dione (Compound 13KR) and (S)- 5-d,-5- Hvdroxyhexyl)-3 -methyl-7-(methyl-<ij)- 1 H-purine-2,6(3H,7H)-dione (Compound 13KS
Separation of Enantiomers of Compound 131. A portion of racemic Compound 131 obtained from Example 15 above was separated in the same manner as racemic Compound 437 above, to afford separated enantiomers Compound
131(R) (mp 1 12.2-1 12.7 °C) (210 mg) and Compound 131(5) (mp 1 12.0-1 12.1 °C) (220 mg).
A. (R)- 1 -(5 - -5 -Hydroxyhexyl)-3 -methyl-7-(methyl-(ij)- 1 H-purine- 2,6(3H,7H -dione (Compound 13UR) . 1H-NMR (300 MHz, CDC13): δ 1.19 (s, 3H), 1.39-1.56 (m, 5H), 1.64-1.74 (m, 2H), 3.58 (s, 3H), 4.03 (t, J=7.3, 2H), 7.51 (s, 1H). 13C-NMR (75 MHz, CDC13): δ 22.87, 23.40, 27.89, 29.71 , 38.64, 41.13, 107.68, 141.40, 148.76, 151.52, 155.39. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid;
Wavelength: 305 nm): retention time: 3.29 min; 99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.14 min (major enantiomer); 28.51 min (expected for minor enantiomer): >99.9% ee purity. MS (M+H): 285.3, (M+Na): 307.2. Elemental Analysis (CisHi^^Os): Calculated: C=54.92, H=7.09, N=19.71. Found:
C=54.67, H=7.04, N=19.35.
Notable in the 1H-NMR spectrum above was the absence of a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position. Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum,
determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
B . (S)- 1 -(5 -di-5 -Hvdroxyhexyl)-3 -methyl-7-(methyl-<ij)- 1 H-purine- 2,6(3H,7H -dione (Compound 13 S)). 1H-NMR (300 MHz, CDCI3): δ 1.18 (s, 3H), 1.39-1.55 (m, 5H), 1.67-1.72 (m, 2H), 3.58 (s, 3H), 4.03 (t, J=7.3, 2H), 7.51 (s, 1H). 13C-NMR (75 MHz, CDC13): δ 23.10, 23.63, 28.12, 29.94, 38.87, 41.36, 107.91 , 141.63, 148.99, 151.75, 155.62. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid;
Wavelength: 305 nm): retention time: 3.29 min; 99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.01% diethylamine for 40 min at 1.00 mL/min; Wavelength: 254 nm): retention time: 28.51 min (major enantiomer); 25.14 min (expected for minor enantiomer): >99.9% ee purity. MS (M+H): 285.3, (M+Na): 307.2. Elemental Analysis (Ci3Hi6D4N403): Calculated: C=54.92, H=7.09, N=19.71. Found:
C=54.65, H=7.04, N=19.32.
Notable in the 1H-NMR spectrum above was the absence of a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position. Due to the presence of a triplet at 4.01 ppm in the above 1H-NMR spectrum,
determination of the presence or absence of a singlet peak at around 3.99 ppm corresponding to the presence or absence of hydrogens on the N-methyl group at the 7 position (R1) of the purine ring was not possible.
Example 17. Synthesis of (±) l-(4A6A6-d 5-hydroxyhexylV3 J-dimethyl-8- d-lH-purine-2,6(3H,7H)-dione (Compound 421). heme 21. Preparation of Compounds 421, 421(R) and 421(5).
Figure imgf000084_0001
purine-2,6(3H,7H)-dione (Compound 421). Following the same general method as for the synthesis of Compound 437 in Example 11 above, Compound 407 (see Example 10) was treated with NaBH4 in EtOD and extracted with CH2CI2 to afford Compound 421.
Example 18. Chiral Separation of (i?)-l-(4A6,6,6-d 5-hydroxyhexyl)-3,7- dimethyl-8-d-lH-purine-2,6(3HJH)-dione (Compound 42KR) and (S)-l- (4^,6.6.6-d^-5-hvdroxyhexyn-3J-dimethyl-8-d-lH-purine-2,6(3H.7H)-dione (Compound 42K5
Separation of Enantiomers 421(R) and 421(S) from (±)Compound 421. A portion of racemic Compound 421 obtained as described above was separated in the same manner as racemic Compound 437 (see Example 12) to afford separated enantiomers Compound 421(R) (560 mg) and Compound 421(5) (520 mg).
A. (i? -l-(4,4,6.6.6-ds-5-hvdroxyhexyl -3.7-dimethyl-8-d-lH-purine- 2,6(3H,7H)-dione (Compound 42UR) . 1H-NMR (300 MHz, CDC13): δ 1.41-1.48 (m, 2H), 1.64-1.72 (m, 3H), 3.58 (s, 3H), 3.79 (s, 1H), 3.99 (s, 3H), 4.03 (t, J=7.3, 2H). 13C-NMR (75 MHz, CDC13): δ 22.69, 27.84, 29.72, 33.60, 41.14, 67.62, 107.64, 148.74, 151.51 , 155.38. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 254 nm): retention time: 3.33 min; >99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 24.77 min (R enantiomer); 28.16 min (expected for S enantiomer); >99.9% ee purity. MS (M+H- H20): 269.1; (M+H): 287.1 ; (M+Na): 309.3. Elemental Analysis (Ci3Hi4D6N403): Calculated: C=54.53, H=7.04, N=19.57. Found: C=54.44, H=7.18, N=19.32.
B. (t^-l-(4,4,6,6,6-d^-5-hvdroxyhexyl -3,7-dimethyl-8-d-lH-purine- 2,6(3H,7H)-dione (Compound 42 S)). 1H-NMR (300 MHz, CDC13): δ 1.37-1.48 (m, 2H), 1.64-1.74 (m, 3H), 3.58 (s, 3H), 3.79 (s, 1H), 3.99 (s, 3H), 4.03 (t, J=7.4, 2H). 13C-NMR (75 MHz, CDC13): δ 22.70, 27.84, 29.71 , 33.60, 41.14, 67.61 , 107.64, 148.74, 151.51 , 155.38. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C 18-RP column - gradient method 5-95%> ACN + 0.1%> formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 254 nm): retention time: 3.34 min; >99.9% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 28.16 min (S enantiomer); 24.77 min (expected for R enantiomer); >99.9% ee purity. MS (M+H- H20): 269.1; (M+H): 287.1 ; (M+Na): 309.3. Elemental Analysis (Ci3Hi4D6N403): Calculated: C=54.53, H=7.04, N=19.57. Found: C=54.54, H=7.18, N=19.31. Notable in the 1H-NMR spectrum of both 42l(R) and 42l(S) was the absence of a peak at around 7.51 ppm, indicating an absence of hydrogen at the 2- position on the imidazole ring system.
Alternative preparation of 421(7?):
Figure imgf000086_0001
421 (R)
Preparation of Compound 421(Λ) from Compound 407 Using CRED A131
A lOOmL 3 -necked RB flask equipped with a heating mantle, a J-Kem thermocouple, magnetic stir bar, a reflux condenser, and a pH probe was charged with 407 (500 mg, 1.75 mmol), D(+) Glucose (750 mg, 1.5 wt) in 10 mL buffer (0.1M KH2PO4, pH = 7.0) and heated to 25-30°C. A solution of NADP (15 mg, 3 wt%), GDH (3 mg, 0.6 wt%), ALMAC CRED A311 -NADP (30 mg, 6 wt%) in 0.1 M KH2PO4 buffer was added and maintained reaction temperature 25-30°C. To this added lmL of methyl-t-butyl ether (MTBE). The pH of the reaction mixture was maintained between 6 and 7 adding 4M KOH solution drop-wise. The reaction was monitored by HPLC and was complete after 29 hours with 99.87A% conversion by HPLC. Sodium chloride (2.5 g, 5 wt) was added and stirred for 20 min. The reaction mixture was extracted with ethyl acetate (3 x 15 mL). The organic layer was separated, filtered through celite pad and concentrated to a small volume (~ 5 vol) and product solids were precipitated. Heptanes (5 mL) was added to the slurry (at 40-60°C) over 5 minutes. The slurry was stirred at 20-25°C and filtered. The wet cake was dried at 50°C for 12 hours to afford 421(R) as a white solid. (0.422 g, 84% yield). The isolated product purity was >99.5%> by HPLC and single enantiomer by chiral HPLC.
Example 19. Synthesis of (±)-l-(4A5A6.6-dfi-5-Hvdroxyhexyl)-3 J-dimethyl- lH-purine-2,6(3H,7H)-dione (Compound 137). heme 22. Preparation of Compound 137.
Figure imgf000087_0001
Synthesis of (±)-l-(4,4,5,6,6,6-dfi-5-Hvdroxyhexyl)-3,7-dimethyl-lH-purine- 2,6(3H,7H)-dione (Compound 137). Compound 437 (560 mg, approximately 2 mmol, see Example 11) was stirred with K2CO3 (270 mg, 2 mmol) in water (10 mL). The mixture was heated at 120-130 °C to give a clear solution and was heated overnight. The solution was extracted with CH2CI2 (1 x 50 mL, 2 x 20 mL) and the CH2CI2 solution was dried (Na2S04) and filtered. After removal of solvent, the solid was stirred with K2CO3 (140 mg, 1 mmol) in water (10 mL) and was heated overnight as above to ensure complete deuterium-to-hydrogen exchange. After extraction with CH2CI2 (1 x 50 mL, 2 x 20 mL), the CH2CI2 solution was dried (Na2S04), filtered and concentrated. The crude product was purified by
chromatography on silica gel eluting with 2-3% MeOH/CE^Cb to give 480 mg (86%) of 137.
HPLC (method: Zorbax 4.6x50 mm SB-Aq 3.5 μιη column - gradient method 2- 98% ACN + 0.1% formic acid in 6.0 min with MSD in ESI positive mode; 0.63 mL/min; Wavelength: 254 nm): retention time: 2.51 min; 98.7% purity. MS (M+H): 287.1; (M+Na): 309.0.
In general, any compound of the invention A having a group
Figure imgf000087_0002
may be further converted to a compound of the invention having the same structure except for having a group
Figure imgf000087_0003
by treating with a suitable base and a proton source, such as water.
Example 20. Synthesis of (^n-^ASAe.e-dfi-S-Hvdroxyhexyn-SJ- dimethyl-lH-purine^.enHJHl-dione (Compound 137(R)V
Scheme 23. Pre aration of Compound 137(R).
Figure imgf000088_0001
437(R) 137(R)
Synthesis of (i?)-l-(4,4,5,6,6,6-dfi-5-Hydroxyhexyl)-3,7-dimethyl-lH-purine- 2,6(3H,7H)-dione (Compound 137(R)V A solution of 437(R) (650 mg, 2.26 mmol, see Example 12) and K2CO3 (320 mg, 2.3 mmol) in water (40 mL) was heated at 1 10 °C (bath temperature) for 26 hours. The solution was concentrated to dryness, redissolved in water (30 mL) and heated to 100 °C for a further 6 hours. After cooling to ambient temperature the solution was extracted with CH2CI2 (4 x 50 mL). The organic solution was dried (Na2S04), filtered, concentrated, then dried under vacuum to afford 565 mg of 137(R) as an off-white solid.
1H-NMR (300 MHz, CDC13): δ 1.38-1.48 (m, 2H), 1.64-1.72 (m, 3H), 3.58 (s, 3H), 3.99 (d, J=0.5, 3H), 4.02 (t, J=7.4, 2H), 7.51 (d, J=0.6, 1H). 13C-NMR (75 MHz, CDCI3): δ 22.65, 27.84, 29.71 , 33.61 , 41.13, 107.67, 141.43, 148.73, 151.50, 155.37. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 305 nm): retention time: 3.30 min; >99.9% purity. MS (M+H-H2O): 269.4; (M+H): 287.1 ; (M+Na): 309.3. Elemental Analysis (Ci3Hi4D6N403): Calculated: C=54.53, H=7.04, N=19.57. Found: C=54.43, H=6.93, N=19.44.
Alternative synthesis of (R)-l-(4,4,5,6,6,6-dfi-5-Hydroxyhexyl)-3,7- dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 137(RV). In a 3-L 3 -necked RB flask, Compound 437(i?) (100 g) was charged followed by water (1.0 L) and K2CO3 (0.25 equiv). The reaction mixture was heated to 80±5°C and monitored by 1H NMR. The reaction was complete after 24 hours and worked up after 65 hours. The resulting product was extracted with three times with EtOAc and the solid products from the three extractions combined and re-dissolved in 5 volumes of EtOAc at 60- 65°C. n-heptane (5.5 vol.) was added at 60-65°C over 15 minutes and cooled to 20°C over night (16 hrs). The slurry was filtered and the wet cake was washed with n-heptane (2x1 vol. to afford product Compound 137(i?) after drying at 40-50°C. A total of 92.4 g of Compound 137(R) was isolated. HPLC purity was 99.92% (AUC) and chiral selectivity was 100% to "S" enantiomer. The 1H NMR analysis showed 99.2%) of "H" at the 8-position in the 3,4,5,7-tetrahydro-lH-purine-2,6-dione ring and 99.4% of "D" at the methyl position.
Example 21. Synthesis of (S)- l -(4.4.5.6.6.6-d^-5-Hvdroxyhexyn-3.7- dimethyl-lH-purine-2,6(3HJH)-dione (Compound 137(5)).
Figure imgf000089_0001
Following the same general method as for the synthesis of Compound
137(R) in Example 20 above, a portion of Compound 437(5) (see Example 12) was converted to 310 mg of Compound 137(5).
1H-NMR (300 MHz, CDC13): δ 1.36-1.45 (m, 2H), 1.62 (s, 1H), 1.64-1.74 (m, 2H), 3.58 (s, 3H), 3.99 (s, 3H), 4.02 (t, J=7.3, 2H), 7.50 (s, 1H). 13C-NMR (75 MHz, CDCI3): δ 23.05, 28.24, 30.07, 33.95, 41.49, 107.92, 141.57, 148.93, 151.68, 155.53. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 305 nm): retention time: 3.34 min; 99.6%> purity. MS (M+H-H20): 269.1 ; (M+H): 287.1 ; (M+Na): 309.3. Elemental Analysis (Ci3Hi4D6N403): Calculated: C=54.53, H=7.04, N=19.57. Found: C=54.71 , H=7.28, N=19.53.
Notable in the 1H-NMR spectrum of 137(5) was the absence of a peak at around 3.80 ppm, indicating an absence of hydrogen at the methinyl hydroxyl position.
Example 22. Synthesis of (± -l-(4.4.6.6.6-ds-5-Hvdroxyhexyl -3.7- dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 121).
Figure imgf000090_0001
Following the same general method as for the synthesis of Compound 137 in Example 19 above, a portion of Compound 421 (see Example 17) was converted to 2.1 g of Compound 121.
1H-NMR (300 MHz, CDC13): δ 1.41-1.48 (m, 2H), 1.64-1.72 (m, 2H), 1.85 (bs, 1H), 3.58 (s, 3H), 3.79 (s, 1H), 3.99 (d, J=0.5, 3H), 4.02 (t, J=7.3, 2H), 7.52 (d, J=0.6, 1H). 13C-NMR (75 MHz, CDC13): δ 22.69, 27.82, 29.70, 33.61 , 41.14, 67.55, 107.66, 141.44, 148.72, 151.49, 155.35. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95%> ACN; Wavelength: 305 nm): retention time: 3.31 min; 99.3% purity. MS (M+H-H20): 268.2; (M+H): 286.2; (M+Na): 308.1. Elemental Analysis (Ci3Hi5D5N403): Calculated: C=54.72, H=7.07, N=19.64. Found: C=54.75, H=6.85, N=19.54.
Example 23. i?-l-(4,4,6,6,6-d^-5-Hydroxyhexyl)-3,7-dimethyl-lH-purine-
2 6(3H,7H)-dione (Compound 12UR)).
Figure imgf000090_0002
Following the same general method as for the synthesis of Compound
137(R) in Example 20 above, a portion of Compound 421(R) (see Example 18) was converted to 1.3 g of Compound 121(R).
1H-NMR (300 MHz, CDC13): δ 1.37-1.48 (m, 2H), 1.64-1.73 (m, 2H), 1.72 (bs, 0.5H), 3.58 (s, 3H), 3.79 (s, 1H), 3.99 (s, 3H), 4.00 (t, J=7.5, 2H), 7.51 (d, J=0.6, 1H). 13C-NMR (75 MHz, CDC13): δ 22.67, 27.83, 29.67, 33.57, 41.12, 67.60, 107.66, 141.40, 148.75, 151.51 , 155.37. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 4.5 minutes (1.0 mL/min) with 1.5 minute hold at 95% CAN (1.5 mL/min);
Wavelength: 305 nm): retention time: 3.29 min; 99.7% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.20 min (R enantiomer); 28.78 min (expected for S enantiomer); >99% ee purity. MS (M+H-H20): 268.2; (M+H): 286.2; (M+Na): 308.1.
Example 24. 6'-l-(4,4,6,6,6-d -5-Hydroxyhexyl)-3,7-dimethyl-lH-purine-
2 6(3H.7H -dione (Compound 121f5)V
Figure imgf000091_0001
Following the same general method as for the synthesis of Compound
137(R) in Example 20 above, a portion of Compound 421(5) (see Example 18) was converted to 590 mg of Compound 121(5).
1H-NMR (300 MHz, CDC13): δ 1.37-1.48 (m, 2H), 1.64-1.73 (m, 2H), 1.86 (bs, 0.5H), 3.58 (s, 3H), 3.79 (s, 1H), 3.99 (d, J=0.6, 3H), 4.02 (t, J=7.4, 2H), 7.52 (d, J=0.7, 1H). 13C-NMR (75 MHz, CDC13): δ 22.70, 27.84, 29.71 , 33.62, 41.14, 67.59, 107.67, 141.43, 148.73, 151.50, 155.37. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C 18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95%> ACN; Wavelength: 305 nm): retention time: 3.37 min; 99.5% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.20 min (expected for R enantiomer); 28.78 min (S enantiomer); >99% ee purity. MS (M+H- H20): 268.2; (M+H): 286.2; (M+Na): 308.1. Elemental Analysis (Ci3Hi5D5N403): Calculated: C=54.72, H=7.07, N=19.64. Found: C=54.77, H=7.13, N=19.59.
Alternatively, Compound 121(5) is synthesized from pentoxifylline (58) in a two step method according to Scheme 22a:
Scheme 24: Alternative Preparation of 6'-l-(4,4,6,6,6-d_s-5-Hydroxyhexyl)-3,7- dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 121f5)V
Figure imgf000092_0001
58 Compound 407
Figure imgf000092_0002
Compound 421(S) Compound 121(S)
Step 1. Compound 407. Pentoxifylline (58; 1 mol equiv) was combined with toluene (20 volumes). To the mixture was added D20 (1.5 volumes) and potassium carbonate (0.25 equiv) and the mixture was heated to reflux (ca. 87°C) for 3-4 hrs. The mixture was cooled to 40-50°C and the aqueous layer was removed. To the remaining toluene solution was added D20 (1.5 volumes) and potassium carbonate (0.25 equiv) and the mixture was heated to reflux (ca. 87°C) for 3-4 hrs. The mixture was cooled to 40-50°C and the aqueous layer was removed. To the remaining toluene solution was added D20 (1.5 volumes) and potassium carbonate (0.25 equiv) and the mixture was heated to reflux (ca. 87°C) for 3-4 hrs. The mixture was cooled to 40-50°C and the aqueous layer was removed. The organic layer was concentrated to ca. 5 volumes below 45°C, was cooled to 20-25°C and then heptane (1 volume) was added, followed by stirring at 20-25°C for 30 min. The slurry was filtered and washed with heptane, followed by drying in vacuo at 40- 50°C to a constant weight. The yield of Compound 407 was approximately 90%. Step 2. Compound 421(5). A 3-necked 12-L RB flask equipped with a heating mantle, a J-Kem thermocouple, a mechanical stirrer, and a pH probe was charged with glucose (547.5g, Aldrich lot # 088K0039) followed by 3.47 L of 0.1M
KH2PO4, pH = 7.0 ("Buffer"; 9.5 vol). The reaction mixture was stirred to dissolve all solids. A mixture of Compound 407 (365 g) in Buffer (2.92 L) was added and the container was rinsed with Buffer (1.28 L). The rinse was added to the reactor.
Initially, the reaction mixture was a very thin milky suspension. A solution of KRED-NADH- 101 (3.65 g, CODEXIS lot # 1021908WW), NAD (2.19 g,
SPECTRUM lot # YA0655), GDH (365 mg, CODEXIS lot # 22016700017) in Buffer (1.46 L) was charged to the reactor. The container was rinsed with Buffer (2 x 0.91 L) and the rinses were added to the reactor. The reaction mixture was warmed to 20-30°C and monitored by a pH meter. The reaction mixture turned clear after 30 minutes. The pH of the reaction mixture was maintained between 6.50 and 6.90 by adding 4M KOH solution drop-wise as needed. The reaction was monitored by HPLC and was complete after 5 hours with 99.97% conversion by HPLC. The reaction mixture was stirred at 20-25°C overnight and warmed to 30°C for the workup.
Sodium chloride (1.825 kg) was added to the reaction mixture and dissolved completely after stirring for 15 minutes. The batch was extracted with EtOAc (10 vol). The organic phase contained a thin solid gel, which collapsed into a slimy separate phase between the aqueous and organic layers immediately when agitated slightly. The slime could be retained on a paper filter but formed a thin impermeable layer that prevented flow through the filter. It was observed on a sample that a small amount of filter aid (celite) easily adsorbed the slime. The aqueous layer was charged back to the reactor and extracted with EtOAc (10 vol). Filter aid (100 g) was charged to the reactor to absorb the slime. The batch was filtered (less than one hour) and the organic layer was collected. The aqueous layer was then extracted with EtOAc (2 x 5 vol) without any problems (no further slime or emulsion was observed). The combined organic extracts were concentrated to ca. 10 volume and polish filtered to remove a small amount of the inorganic solids. The filtrate was concentrated further to ca. 5 volumes and product solids were precipitated, n- heptane (8 vol) was added to the slurry (at 40-60°C) over 30 minutes. The slurry was stirred overnight at 20-25 °C and filtered. The filter cake was washed with n-heptane (2 x 1 vol). The wet cake (370 g) was dried at 40-50°C over the weekend to afford Compound 421(5) as a white solid (332.0 g, 90.0% yield). The filtrate was concentrated followed by precipitation with heptane to afford a second crop of Compound 421(5) (7.1 g, 1.9% yield). In order to check the mass balance of the product, the aqueous layer was extracted again with EtOAc (10 vol) and afforded only 4.8 g of Compound 421(5) (1.3 % yield) of product as a white solid. The combined mother liquor was concentrated to afford 2.0 g of Compound 421(5) as a yellow solid (0.5% yield). The isolated product was a very high quality (100% purity by HPLC) and a single enantiomer (100/0 S/R% by chiral HPLC) from the main lot with 99.5% "D" incorporation at the methyl position by 1H NMR.
Step 2 (alternative procedure):
[1] A 12-L 3 -necked RB flask equipped with a heating mantle, a J-Kem thermocouple, a mechanical stirrer, a reflux condenser, and a pH probe was charged with CRED A131 (9.5 g, ALMAC lot # IM-1311-061-1) and 2 L of buffer solution (0.1M KH2P04, pH = 7.0, same as below). The reaction mixture was stirred to dissolve all solids. A solution of glucose (558 g, Aldrich lot # 088K0039) in buffer (2 L) was added in one portion followed by a solution of NAD (19.25 g, Spectrum lot # YA0655) in buffer (500 mL), and a solution of GDH (1.5 g, ALMAC lot # IM- 1311-131-1) in buffer (500 mL). The initial reaction mixture was pH 6.98. A mixture of Compound 407 in buffer (3 L) at 30 °C was added to the reaction mixture and the container was rinsed with buffer (1.6 L). The rinse was charged to the reactor. The pH of the reaction mixture was 6.99. The reaction mixture was warmed to 30°C and monitored by pH meter. The reaction temperature was kept at 29.0 to 31.5°C and the pH of the reaction mixture was kept between pH 6.93 and pH 7.02 by adding 4M KOH solution drop-wise as needed. The reaction was complete after 22 hours with 99.96% conversion as determined by HPLC. The chiral HPLC analysis of the resulting product showed the chiral selectivity was 99.85%) to the desired S-alcohol.
[2] The reaction mixture was mixed with NaCl (2 kg) and extracted with EtOAc (1 x 4 L and 3 x 2 L). During the first extraction, a rag layer was formed and the reaction mixture was filtered through a celite pad. No further issues with phase separation were encountered after the filtration. The combined organic extracts were concentrated to about 1.5 L at 50-60°C and n-heptane (2 L) was added to precipitate the solids. The slurry was cooled to 20° C and filtered. The flask was rinsed with filtrate to complete the transfer. The filter cake was washed with n-heptane (2 x 500 mL) and dried over the weekend at 40-50° C to afford Compound 421(5) (366 g, 94% yield). The product was analyzed by HPLC (99.95% purity), chiral HPLC (99.88/0.12 S/R), and 1H NMR (99.5% "D" incorporation at the methyl position). Step 3. Compound 121(5). In a 3 -L 3 -necked RB flask, Compound 421(5) (100 g) was charged followed by water (1.0 L) and K2CO3 (0.25 equiv). The reaction mixture was heated to 80±5°C and monitored by 1H NMR. The reaction was complete after 24 hours and worked up after 65 hours. The resulting product was extracted with three times with EtOAc and the solid products from the three extractions combined and re-dissolved in 5 volumes of EtOAc at 60-65°C. n-heptane (5.5 vol.) was added at 60-65°C over 15 minutes and cooled to 20°C over night (16 hrs). The slurry was filtered and the wet cake was washed with n-heptane (2x1 vol. to afford product Compound 121(5) after drying at 40-50°C. A total of 92.4 g of Compound 121(5) was isolated. HPLC purity was 99.92% (AUC) and chiral selectivity was 100% to "S" enantiomer. The 1H NMR analysis showed 99.2% of "H" at the 8-position in the 3,4,5,7-tetrahydro-lH-purine-2,6-dione ring and 99.4% of "D" at the methyl position.
Example 25. Synthesis of 3,7-Dimethyl-l-(4,4,6,6,6-d 5-oxohexyl)-lH- purine-2,6(3H,7H)-dione (Compound 107).
Scheme 25. Pre aration of Compound 107.
Figure imgf000095_0001
Synthesis of 3,7-Dimethyl-l-(4,4,6,6,6-ds-5-oxohexyl)-lH-purine- 2,6(3H.7H)-dione (Compound 107). Compound 121 (0.49 g, 1.72 mmol, see Example 22) and N-methylmorpholine N-oxide "NMO" (301 mg, 2.58 mmol) were dissolved in CH2CI2 (20 mL). Tetrapropylammonium perruthenate "TPAP" (27 mg, 0.086 mmol) was added and the solution was stirred for 2.5 hours at ambient temperature. TLC (EtOAc) showed the reaction was complete. The reaction was concentrated and purified by silica gel chromatography eluting with EtOAc. The material was dried in a vacuum oven (50 °C) for 4 hours to afford 400 mg (82%) of Compound 107. The material was further purified by crystallization
(EtO Ac/heptane) to give 320 mg of 107. NMR and LCMS analysis indicated no loss of deuterium.
1H-NMR (300 MHz, CDCI3): δ 1.64-1.70 (m, 4H), 3.57 (s, 3H), 3.99 (d, J=0.6, 3H), 4.01-4.04 (m, 2H), 7.51 (d, J=0.6, 1H). 13C-NMR (75 MHz, CDCI3): δ 20.82, 27.38, 29.69, 33.61, 40.80, 107.75, 141.42, 148.76, 151.46, 155.26. HPLC (method:
Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 305 nm): retention time: 3.28 min; >99.9% purity. MS (M+H): 284.1 ; (M+Na): 306.0.
Example 26. Synthesis of (±) l-(4.4.5.6.6.6-dg-5-Hvdroxyhexyl)-3.7- di(methyl-dQ-lH-purine-2,6(3H,7H)-dione (Compound 434).
Scheme 26. Preparation of Compounds 434, 434(R) and 434(5).
Figure imgf000096_0001
Chir
sep
Figure imgf000096_0002
purine-2,6(3H,7H)-dione (Compound 434). Following the same general method as for the synthesis of Compound 437 in Example 1 1 above, a portion of Compound 413 (see Example 4) was treated with NaBD4 in EtOD to and extracted with CH2CI2 afford 190 mg of Compound 434.
Example 27. Chiral Separation of (R)-l -(4,4,5, 6,6,6-dfi-5-Hvdroxyhexyl)- 3,7-di(methyl-d^-lH-purine-2,6(3H,7H)-dione (Compound 434(R) and (6^-1- (4,4,5,6,6,6-dfi-5-Hydroxyhexyl)-3,7-di(methyl-d -lH-purine-2,6(3H,7H)-dione (Compound 434(5)).
Separation of Enantiomers of Compound 434. A portion of racemic Compound 434 obtained as described above was separated in the same manner as racemic Compound 437 (see Example 12) to afford separated enantiomers
Compound 434(R) (72 mg) and Compound 434(5) (74 mg).
A. (i? -l-(4,4,5.6.6.6-dfi-5-Hvdroxyhexyn-3.7-di(methyl-d^-lH-purine- 2,6(3H,7H)-dione (Compound 434(R)V 1H-NMR (300 MHz, CDC13): δ 1.34-1.52 (m, 2H), 1.59-1.76 (m, 3H), 4.02 (t, J=7.3, 2H). 13C-NMR (75 MHz, CDCI3): 5 22.65, 27.84, 41.12, 107.64, 151.52, 155.40. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 254 nm): retention time: 3.29 min; 99.5% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 24.34 min (R enantiomer); 28.82 min (expected for S enantiomer); >99% ee purity. MS (M+H- H20): 276.3; (M+H): 294.3; (M+Na): 316.2.
B. (t^-l-(4,4,5,6,6,6-dfi-5-Hvdroxyhexyn-3J-di(methyl-d -lH-purine- 2,6(3H,7H)-dione (Compound 434(S)V 1H-NMR (300 MHz, CDC13): δ 1.36-1.50 (m, 2H), 1.64-1.76 (m, 3H), 4.02 (t, J=7.5, 2H). 13C-NMR (75 MHz, CDC13):
δ 22.65, 27.84, 41.12, 151.52, 155.40. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 254 nm): retention time: 3.29 min; 99.4% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 24.34 min (expected for R enantiomer); 28.82 min (S enantiomer); >99% ee purity. MS (M+H- H20): 276.3; (M+H): 294.3; (M+Na): 316.2.
Example 28. Synthesis of (±)-! -(4,4,5, 6,6,6-dfi-5-hydroxyhexyl)-3-methyl- 7-methyl-d3-lH-purine-2,6(3H,7H)-dione (Compound 135).
Scheme 27. Pre aration of Compounds 135, 135(R) and 135(5).
Figure imgf000097_0001
Figure imgf000097_0002
Synthesis of (±)-! -(4,4,5, 6,6,6-dfi-5-hvdroxyhexyl)-3 -methyl-7-methyl-cU- lH-purine-2,6(3H,7H)-dione (Compound 135). Following the same general method as for the synthesis of Compound 137 in Example 19 above, a portion of Compound 435 (see Example 8) was converted to 0.99 g of Compound 135.
Example 29. Chiral Separation of (i?)-l-(4,4,5,6,6,6-dfi-5-hvdroxyhexyl)-3- methyl-7-methyl-d^-lH-purine-2,6(3H,7H)-dione (Compound 135(RV) and (S)-l- (4,4, 5,6,6, 6-dfi-5-hydroxyhexyl)-3 -methyl-7-methyl-d^-lH-purine-2,6(3H,7H - dione (Compound 135(5)).
Separation of Enantiomers of Compound 135. A portion of racemic
Compound 135 obtained as described above was separated in the same manner as racemic Compound 437 (see Example 12) to afford separated enantiomers
Compound 135(R) (352 mg) and Compound 135(5) (343 mg).
A. (R)- 1 -(4,4,5 ,6,6,6-dfi-5-hvdroxyhexyl)-3 -methyl-7-methyl-d^- lH-purine- 2,6(3H,7H)-dione (Compound 135(R)). 1H-NMR (300 MHz, CDC13): δ 1.41-1.48 (m, 2H), 1.64-1.74 (m, 3H), 3.58 (s, 3H), 4.02 (t, J=7.4, 2H), 7.50 (s, 1H). 13C- NMR (75 MHz, CDC13): δ 22.65, 27.84, 29.68, 41.12, 107.67, 141.38, 148.76, 151.52, 155.37. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη CI 8-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 305 nm): retention time: 3.27 min; 99.6% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.21 min (R enantiomer); 28.42 min (expected for S enantiomer); >99.5% ee purity. MS (M+H-H20): 272.1 ; (M+H): 290.1 ; (M+Na): 312.3. Elemental Analysis (C13H11D9N4O3): Calculated: C=53.97, H=6.97, N=19.36. Found: C=53.83, H=6.98, N=19.30.
B. (6)-l-(4,4,5,6,6,6-dfi-5-hydroxyhexyl)-3 -methyl-7-methyl-d3-lH-purine- 2,6(3H,7H)-dione (Compound 135(5)). 1H-NMR (300 MHz, CDCI3): δ 1.38-1.48 (m, 2H), 1.64-1.74 (m, 3H), 3.58 (s, 3H), 4.02 (t, J=7.4, 2H), 7.50 (s, 1H). 13C- NMR (75 MHz, CDCI3): δ 22.64, 27.84, 29.68, 41.12, 107.67, 141.38, 148.76, 151.52, 155.37. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη CI 8-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 minutes (1.0 mL/min) with 4 minute hold at 95% ACN; Wavelength: 305 nm): retention time: 3.27 min; 99.8% purity. Chiral HPLC (method: Chiralpak AD 25 cm column - isocratic method 78% hexane/ 22% isopropanol/0.1% diethylamine for 40 minutes at 1.00 mL/min; Wavelength: 254 nm): retention time: 25.39 min (R enantiomer;
minor species); 28.42 min (S enantiomer; major species); 99.1% ee purity. MS (M+H-H20): 272.1 ; (M+H): 290.1 ; (M+Na): 312.3. Elemental Analysis
(C13H11D9N4O3): Calculated: C=53.97, H=6.97, N=19.36. Found: C=53.93, H=7.03, N=19.29.
Example 30. Synthesis of (±) l-(5-Hvdroxyhexyl)-3-methyl-7-methyl-(iJ-lH- .
Figure imgf000099_0001
Following the same general method as for the synthesis of Compound 437 in Example 1 1 above, Compound 100 (see Example 1) was treated with NaBH4 in EtOH and extracted with CH2CI2 to afford Compound 116.
MS (M+H-H2O): 266.1 ; (M+H): 284.1 ; (M+Na): 306.0.
Figure imgf000099_0002
purine^^OH.JFD-dione (Compound 133) and of the (R) and (S) enantiomers of Compound 133.
Scheme 28. Preparation of Compounds 133, 133(61 and 133(i?).
Figure imgf000099_0003
(±) 1 -(5-<i/-5-Hvdroxyhexyl)-3 ,7-dimethyl- 1 H-purine-2,6(3H,7H)-dione (Compound 133). Following the same general method as for the synthesis of Compound 437 (see Example 11), commercially available 58 was treated with NaBD4 in EtOH to afford Compound 133). 1H-NMR (300 MHz, CDC13): δ 1.18 (s, 3H), 1.39-1.55 (m, 3H), 1.61-1.64 (m, 1H), 1.66-1.76 (m, 2H), 3.58 (s, 3H), 3.99 (s, 3H), 4.02 (t, J= 7.4, 2H), 7.51 (s, 1H). 13C-NMR (75 MHz, CDC13): δ 22.87, 23.37, 27.89, 29.67, 33.57, 38.64, 41.12, 67.11, 67.40, 67.69, 107.67, 141.40, 148.79, 151.51, 155.36. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 305 nm): retention time: 3.32 min; >99% purity. MS (M+H): 282.0. Elemental Analysis
(C13H19DN4O3): Calculated: C=55.50, H=7.17, N=19.92. Found: C=55.4, H=7.34, N=19.72.
Example 32. Chiral Separation of (i?)-l-(5-^-5-Hydroxyhexyl)-3,7- dimethyl-lH-purine-2,6(3H,7H -dione (Compound 133(R) and (S)- 5-dL-5- Hvdroxyhexyl)-3,7-dimethyl-lH-purine-2,6(3H,7H)-dione (Compound 133(5)).
Separation of Enantiomers of Compound 133. A portion of racemic
Compound 133 obtained from Example 31 above was separated in the same manner as racemic Compound 437 (see Example 12), to afford separated enantiomers.
Compound 133( f) (mp 112.9-113.1 °C) (290 mg) and Compound 133(S) (mp 112.1-112.2 °C) (302 mg).
A. (i? -l-(5- -5-Hvdroxyhexyn-3J-dimethyl-lH-purine-2,6(3H,7H -dione (Compound 133(R)V 1H-NMR (300 MHz, CDCI3): δ 1.18 (s, 3H), 1.40-1.54 (m, 3H), 1.61 (s, 1H), 1.65-1.72 (m, 2H), 3.58 (s, 3H), 3.99 (s, 3H), 4.03 (t, J= 7.5, 2H), 7.50 (s, 1H). 13C-NMR (75 MHz, CDCI3): δ 23.47, 24.00, 28.50, 30.30, 34.19, 39.25, 41.73, 142.01. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18- RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95%> ACN + 0.1%> formic acid; Wavelength: 305 nm): retention time: 3.31 min; >99% purity. MS (M+H): 282.0. Elemental Analysis (Ci3Hi9DN403): Calculated: C=55.50, H=7.17, N=19.92. Found: C=55.73, H=7.02, N=19.83. Notable in the 1H-NMR spectrum above was the absence of a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position.
B . (S)- 5 -dj-5 -Hvdroxyhexyn-3 ,7-dimethyl- 1 H-purine-2,6(3 HJHVdione (Compound 133(5)). 1H-NMR (300 MHz, CDC13): δ 1.19 (s, 3H), 1.39-1.56 (m, 3H), 1.65-1.74 (m, 3H), 3.58 (s, 3H), 3.99 (t, J=7.3, 2H), 4.03 (t, J= 7.4, 2H), 7.51 (s, 1H). 13C-NMR (75 MHz, CDC13): δ 22.86, 23.38, 27.89, 29.67, 33.57, 38.64, 41.11, 141.40, 148.76, 151.51, 155.37. HPLC (method: Waters Atlantis T3 2.1 x 50 mm 3 μιη C18-RP column - gradient method 5-95% ACN + 0.1% formic acid in 14 min (1.0 mL/min) with 4 min hold at 95% ACN + 0.1% formic acid; Wavelength: 305 nm): retention time: 3.30 min; >99% purity. MS (M+H): 282.3. Elemental Analysis (C13H19DN4O3): Calculated: C=55.50, H=7.17, N=19.92. Found:
C=55.51, H=7.10, N=19.72.
Notable in the 1H-NMR spectrum above was the absence of a peak at around 3.80 ppm indicating an absence of hydrogen at the methinyl hydroxyl position.
Example 33. Synthesis of 3,7-dimethyl-l-(6,6,6-(ij-5-oxohexyl)-lH-purine- 2,6(3H,7H)-dione (Compound 157) and (S)-3 J-dimethyl-l-(6,6,6-^-5- hvdroxyhexyl)-lH-purine-2,6(3H,7H)-dione (Compound 156(5)).
Scheme 29. Preparation of Compounds 157 and 156(5).
Figure imgf000101_0001
157
156(S)
Step 1. 5-(3,7-dimethyl-2,6-dioxo-2,3,6,7-tetrahydro-lH-purin-l-yl)-N- methoxy-N-methylpentanamide (61). To a 100 mL round-bottom flask equipped with a magnetic stirrer and thermocouple was added 50 (1.49 g, 8.27 mmol, 1.0 eq) and DMSO (40mL). The mixture was stirred and heated to about 35 °C to dissolve all materials, and then NaH was added (60% dispersion in oil; 347 mg, 8.68 mmol, 1.05 eq) as a single portion. The mixture was heated to 50 °C and stirred at 50 °C for 30 mins (note: stirring became difficult due to formation of a pasty mixture), then cooled to room temperature. To the mixture was then added a solution of crude bromide 56 (1.95 g, 8.68 mmol, 1.05 eq) in DMSO (5 mL) via syringe. The mixture was stirred at room temperature overnight. It became a clear yellow solution. The solution was diluted with copious amount of water (200 mL), and then extracted with CH2CI2 (3 x 100 mL). The combined CH2Cl2layers were washed with water (2 x 100 mL). A solid residue was obtained after removing the organic volatiles in a rotovap. The solid residue was suspended in MTBE (25 mL), the mixture was stirred at 50 °C for 1 hr, then at room temperature for another hour, followed by filtration via a medium-porosity funnel, an MTBE rinse (2 x 10 mL), and drying under vacuum. The product was collected as 2.19 g (82%) of an off-white solid with an AUC purity of 99 A%.
Step 2. 3,7-dimethyl-l-(6,6,6-(ij-5-oxohexyl)-lH-purine-2,6(3H,7H)-dione (Compound 157). To a 300 mL, 3 -neck round-bottom flask equipped with a magnetic stirrer and thermocouple was added 61 (1.30 g, 4.01 mmol, 1.0 eq) and THF (45mL). The mixture was stirred and heated to about 45 °C to dissolve all materials, then was cooled to 0 °C (note: a solid precipitated, but less solid was present than prior to heating). CD3MgI was added (1.0 M in Et20, 8.82 mL, 2.2 eq) via syringe, at such a rate that the internal temperature did not rise above 5 °C. After the addition was complete, the mixture was stirred at 0 °C for 30 mins, then the cold bath was removed and the mixture was allowed to warm up to room temperature. The mixture was stirred at room temperature for 1.5 hrs, whereupon IPC analysis by HPLC indicated conversion was 80 A%. Further stirring at room temperature for 1.5 hrs afforded no additional conversion, as shown by another IPC analysis. The mixture was cooled to 0 °C and more CD3MgI (1.0 M in Et20, 2.0 mL, 0.5 eq) was added via a syringe. The mixture was warmed to room temperature overnight. The next day, IPC analysis by HPLC indicated that conversion was higher than 95 A%. The reaction mixture was quenched with 0.5 N aqueous citric acid (40 mL) and extracted with MTBE (60 mL). The phases were separated and the organic layer was washed with water (20 mL), aq. satd. NaHC03 (20 mL), then water (20 mL). The organic layer was concentrated in a rotovap to afford crude product with an AUC purity at 91 A%. Further purification was carried out by slurrying in MTBE (5 mL) at room temperature overnight, followed by filtration and an MTBE rinse. The desired product was obtained as 0.92 g (81%) of a white solid with an AUC purity of - 95 A%. 1H NMR: δ 1.61-1.71 (m, 4H), 2.50 (t, J = 7.1, 2H), 3.57 (s, 3H), 3.98 (s, 3H), 4.01 (t, J= 7.1, 2H), 7.50 (s, 1H).
Notable in the 1H-NMR spectrum above was the absence of a singlet at around 2.15 ppm, indicating an absence of methyl ketone hydrogens.
Step 3. (S)-3 ,7-dimethyl- 1 -(6,6,6- j-5-hydroxyhexyO- 1 H-purine-2,6(3H,7H)-dione (Compound 156). To a 100 mL, 3 -neck round-bottom flask equipped with a magnetic stirrer and thermocouple was added Compound 157 (563 mg, 2.00 mmol, 1.0 eq), D (+)-glucose (844 mg), and 0.1 M KH2P04 (11 mL). The mixture was stirred at room temperature. NAD (3.4 mg, 0.6 w %), GDH (0.6 mg, 0.1 w %), and enzyme KRED-NADH (5.6 mg, 1.0 w %) were dissolved in 0.1 N KH2P04. This solution was added to the reaction mixture. The resulting cloudy solution was stirred at room temperature for 5 hrs, during which 4N aq. KOH was added dropwise to the reaction mixture to maintain its pH between 6 and 7, as measured by a pH meter. An aliquot was sampled and analyzed by HPLC and indicated greater than 99.5 A% conversion. Solid NaCl (~ 3 g) was added and the mixture stirred for 30 mins. EtOAc (10 mL) was added and the mixture was stirred for another 30 mins. The mixture was filtered through a pad of wet Celite to remove a gel-like substance, and the wet filter cake was rinsed with EtOAc (2 x 10 mL). The filtrate was collected and the phases were separated. The aqueous layer was washed with EtOAc (2 x 60 mL). The combined organic layers were concentrated to dryness in a rotovap. 527 mg of crude product were collected, affording a mass balance of 93%. The residue was purified by flash chromatography (silica gel, eluent MeOH/ CH2C12, gradient 2 - 20 % MeOH) to afford 398 mg (70%)of the desired product as a white solid with an AUC purity of 99 A%. 1H NMR: δ 1.45-1.57 (m, 4H), 1.63- 1.74 (m, 2H), 3.57 (s, 3H), 3.76-3.83 (m, 1H), 3.98 (s, 3H), 4.02 (t, J= 7.3, 2H), 7.50 (s, 1H).
Notable in the 1H-NMR spectrum above was the absence of a peak at around 1.19 ppm, indicating an absence of methyl hydrogens alpha to the hydroxyl group. Chiral HPLC analysis (method: Chiralpak AD-H 25 cm column - isocratic method 75% n-heptane/ 25% isopropanol for 25 min at 1.25 mL/min; wavelength: 274 nm): retention time: 17.5 min (major enantiomer); 15.5 min (expected for minor enantiomer): >99.95% ee purity.
BIOLOGICAL EVALUATION
Example 34a. Evaluation of Pharmacokinetics in Dogs Following Oral Administration. Comparison of Compound 409 and Pentoxifylline
Metabolism of the title compounds were studied following oral
administration to male beagle dogs. Blood samples were removed from dosed dogs at various time points and plasma isolated therefrom. The plasma samples were used for the determination of plasma drug levels by LC-MS/MS (liquid chromatography with tandem mass spectrometry) for estimating pharmacokinetic parameters.
Compound 409 and pentoxifylline were dissolved separately in saline to a concentration of 4 mg/mL. A 1 : 1 (v/v) mixture of the two solutions was prepared to yield a solution having a final concentration of 2 mg/mL of both Compound 409 and pentoxifylline.
Two male beagle dogs were fasted overnight and then orally dosed via gavage with 2.5 mg/kg of Compound 409 and pentoxifylline using the mixture described above. Blood samples (1.5 - 2 mL) were collected via the femoral vein at 0 min (pre-dose), 15 min, 30 min, 45 min, 1 hr, 1.5 hr, 2 hr, 3 hr, 4 hr, 6 hr, 8 hr, 10 hr, 12 hr, 16 hr and 24 hr post-dose. Blood was stored on ice prior to centrifugation to obtain plasma samples. Centrifugation took place within 1 hour of blood collection to harvest plasma (maximum volume). The plasma was decanted immediately and frozen/stored at -70 °C until analysis.
Table 8. Plasma Levels of Compound 409 vs Pentoxifylline in Dogs (Example 29a)
Figure imgf000104_0001
a) % Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
Table 8 shows the results of the evaluation described in Example 34a. The average Cmax and average AUC for Compound 409, a deuterated version of pentoxifylline, were significantly greater than for pentoxifylline. The deuterated compound exhibited greater exposure in the dog plasma than pentoxifylline. Example 34b. Repeat Evaluation of Pharmacokinetics in Dogs Following Oral Administration. Comparison of Compound 409 and Pentoxifylline with Monitoring of Metabolites
Example 34a was repeated with additional monitoring of the pentoxifylline and Compound 409 metabolites. In this experiment Compound 409 and pentoxifylline were dissolved separately in saline to a concentration of 4.4 and 4 mg/mL respectively. A 1 : 1 (v/v) mixture of the two solutions was prepared to yield a solution having a final concentration of 2.2 mg/mL of Compound 409 and 2 mg/mL pentoxifylline. Post-dosing data analysis included adjustments to account for the 10% difference in dosing concentration between compound 409 and pentoxifylline.
Four beagle dogs (2-3 years of age, and weighed 5 to 8 kg) were fasted overnight and then orally dosed via gavage with 2.75 mg/kg Compound 409 and 2.5 mg/kg pentoxifylline using the mixture described above. Blood samples
(approximately lmL) were collected via femoral vein at 0 min (pre-dose), 5 min, 15 min, 30 min, 45 min, 1 fir, 1.5 hr, 2 hr, 3 hr, 4 hr, and 6 hr post-dose. Blood was stored on ice prior to centrifugation to obtain plasma samples. Centrifugation took place within 15 minutes of blood collection to harvest plasma (maximum volume). The plasma was decanted immediately and frozen/stored at -20 °C until analysis.
Plasma samples were analyzed by LC-MS/MS for the presence of the administered com ound and its corresponding Ml metabolite:
Figure imgf000105_0001
pentoxifylline Ml
Figure imgf000106_0001
Compound 409 (administered) Compound 419 (Ml metabolite)
The results from each of the four dogs are shown in FIGS. 1A and IB. The results from one of the four dogs (Dog H, FIG. lb) were inconsistent with that of the other three. That dog showed a 10-fold higher plasma concentration of each of the administered compounds and their respective metabolites at 5 minutes post- administration. In addition, that dog did not show a characteristic increase in plasma concentration of the administered compounds between 5 and 15 minutes post- administration. It was concluded that this dog was most likely improperly gavaged and that the compounds were probably administered through the trachea, rather than into the GI tract as would have been desired. Accordingly, the data from this dog was excluded from the analyses. The summary analysis of the three remaining dogs is shown in Table 9.
Table 9. Plasma Levels of Compound 409 vs Pentoxifylline in Dogs (Example 34b)
Figure imgf000106_0002
a) The dosing concentration of compound 409 was 10% higher than that for pentoxifylline and thus the numbers reported here reflect the adjustment for that 10%) increase.
b) % Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
As can be seen in Table 9, higher levels of Compound 409 in terms of C, and AUC were observed when compared to pentoxifylline co-dosed at the same level. FIG. 1 demonstrates that Compound 409 was more slowly cleared from the plasma than pentoxifylline in the three dogs that were orally dosed. FIG. la and lb demonstrate that Compound 409 was more slowly cleared from the plasma than pentoxifylline in the three dogs that were orally dosed. FIGS, la and lb also show that overall systemic exposure to Compound 419 (the deuterated Ml metabolite of 409) following dosing of Compound 409 was greater than that of the Ml metabolite following dosing of pentoxifylline.
Example 34c. Evaluation of Pharmacokinetics in Dogs Following Oral
Administration. Comparison of Compound 413 and Pentoxifylline.
This study was similar to those described in Examples 34a and 34b, except that Compound 413 was evaluated. Four male beagle dogs were orally dosed by gavage with a mixture containing 2 mg/mL each of pentoxifylline and Compound 413 in saline. Blood samples were taken as in Example 34b.
Table 10. Plasma Levels of Compound 413 vs Pentoxifylline in Dogs (Example
Figure imgf000107_0001
Figure imgf000107_0002
a) % Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
The results of this study are summarized in Table 10 above. The table depicts the plasma levels of Compound 413 compared to pentoxifylline following oral dosing. Higher levels of Compound 413 in terms of Cmax and AUC were observed when compared to pentoxifylline co-dosed at the same level.
Example 35. Evaluation of the Stability of Compounds in Rat Whole Blood. Comparison of Compounds 409, 435(5), 435(R) and Pentoxifylline and its M-l Metabolites.
This study was performed to evaluate the stability of the title compounds in rat whole blood. Because the ketone (or keto-compound; either pentoxifylline or 409) and its corresponding M-l alcohol metabolite interconvert, levels of these components were measured after either the keto-compound was added to the blood or the M-l was added. In other words, in some tests the keto-compound was the starting test compound and in other tests an M-l metabolite was the starting test compound.
Fresh rat whole blood was obtained from ViviSource Laboratories, Waltham, MA. Stock solutions (7.5 millimolar (mM)) of test compounds were prepared in dimethyl sulfoxide (DMSO). The 7.5 mM stock solutions were diluted to 500 micromolar (μΜ) in acetonitrile (ACN). To 990 microliters (μί) of blood pre- warmed to 37 °C for 7 minutes was added 10 of 500 μΜ test compound to a final concentration of 5 μΜ. The test compounds were pentoxifylline, (5)-Ml metabolite of pentoxifylline, (i?)-Ml metabolite of pentoxifylline, Compound 409, Compound 435(5), and Compound 435(R). The latter two test compounds are deuterated (S)- Ml and (i?)-Ml metabolites, respectively, of Compound 409. The reaction mixture was incubated at 37 °C. Aliquots (50 μί) were removed at 0 min, 5 min, 15 min, 30 min, 1 hour and 2 hours following the addition of test compound and added to 96- well plates containing 150 μί of ice cold acetonitrile with an internal standard to stop the reaction. The plates were stored at -20 °C for 20 minutes after which 100 μΐ, of 50% acetonitrile/water was added to the wells of the plate prior to
centrifugation to pellet precipitated proteins. A 200-μί aliquot of each supernatant was transferred to another 96-well plate and analyzed by LC-MS/MS using an Applied Bio-systems API 4000 mass spectrometer for amounts of the administered compound and its specific metabolite listed in Table 1 1 below.
Table 1 1. Compound-Metabolite Pairs Analyzed in Rat Whole Blood. (Examples
Figure imgf000108_0001
a) Mass observed via LC-MS/MS. Stereochemistry presumed to be >95% (S) based on published pentoxifylline metabolism reports.
The results of this study are depicted in FIGS. 2 and 3. The time course of metabolite formation is shown in FIG. 2. The relative amount of metabolite formed, as shown in FIG. 3, was calculated based on the amount present at 2 hr relative to the earliest time point at which it was detected in the incubation mixture, 5 minutes for A and B, and 15 minutes for C.
As seen in FIG. 3, after approximately 2 hours the amount of (5)-Ml formed in rat whole blood incubated with pentoxifylline (Fig 3, column A) was similar to the amount of Compound 419(5) formed in rat whole blood incubated with
Compound 409 (Fig 3, column B). Thus, the deuterium substitution in Compound 409 had no discernable effect on the relative level of deuterated (5)-Ml metabolite (Compound 419(5)) formed as compared to the relative level of undeuterated (S)- Ml formed from undeuterated pentoxifylline.
For the reverse reaction, (5)-Ml to the keto-compound, deuteration did have a significant effect. Column C in FIG. 3 shows an appreciable amount of pentoxifylline present after addition of (5)-Ml . By contrast, 2 hours after addition of Compound 435 (5), Compound 409 was not detected (FIG. 3, column D). Under these conditions, the deuterium substitution in Compound 435 (5) impedes the conversion of this compound to the corresponding ketone. Such an effect is particularly beneficial for enhancing the plasma levels of the desired M-l metabolite.
No metabolism of (i?)-Ml to pentoxifylline was detected in this assay.
Similarly, Compound 409 was not detected after addition of Compound 435 (R) to the rat blood. Thus, no conclusions could be made concerning the effect of deuteration on the conversion of (i?)-Ml to pentoxifylline. FIG. 2 shows the time course of the specific metabolite produced during incubation of the administered compound with rat whole blood.
Example 36. Evaluation of Compound Stability in Human Liver Microsomes. Comparison of Compounds 409, 435(5), 435(R) and Pentoxifylline.
Example 36 is similar to Example 35 in design, except that human liver microsomes were used instead of rat whole blood to study the metabolism of the compounds. Table 1 1 above shows each pair of test compound and metabolite that was analyzed in this Example 36.
Human liver microsomes (20 mg/mL) were obtained from Xenotech, LLC (Lenexa, KS). β -nicotinamide adenine dinucleotide phosphate, reduced form (NADPH), magnesium chloride (MgCl2), and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich.
Stock solutions containing 7.5 mM of test compounds (pentoxifylline, (S)- Ml metabolite, (i?)-Ml metabolite, Compound 409, Compound 435(5), and
Compound 435(R)) were prepared in DMSO. The 7.5-mM stock solutions were diluted to 250 μΜ in acetonitrile (ACN). The human liver microsomes were diluted to 2.5 mg/mL in 0.1 M potassium phosphate buffer, pH 7.4, containing 3 mM MgCl2. The diluted microsomes were added to wells of a 96-well deep-well polypropylene plate in triplicate. 10 μΐ, οΐ the 250 μΜ test compound was added to the microsomes and the mixture was pre-warmed to 37 °C for 10 minutes.
Reactions were initiated by addition of pre-warmed NADPH solution. The final reaction volume was 0.5 mL and contained 2.0 mg/mL human liver microsomes, 5 μΜ test compound, and 2 mM NADPH in 0.1 M potassium phosphate buffer, pH 7.4, and 3 mM MgCl2. The reaction mixtures were incubated at 37 °C, and 50-μί aliquots were removed at 0, 5, 10, 20, and 30 minutes and added to shallow-well 96- well plates which contained 50 μί of ice-cold acetonitrile with internal standard to stop the reactions. The plates were stored at 4 °C for 20 minutes after which 100 μΐ^ of water was added to the wells of the plate before centrifugation to pellet precipitated proteins. Supernatants were transferred to another 96-well plate and analyzed for the amount of the administered compound and its specific metabolite (listed in Table 1 1 above) by LC-MS/MS using an Applied Bio-systems API 4000 mass spectrometer.
The results of this study are depicted in FIGS. 4 and 5. The time course of metabolite formation is shown in FIG. 4. The relative amount of metabolite formed, as shown in FIG. 5, was calculated based on the amount present at 30 minutes relative to the earliest time point at which it was detected in the incubation mixture, 0 minutes for A, B, C and E, 5 minutes for D, and 10 minutes for F. The amount of (iS)-Ml formed in human liver microsomes incubated with pentoxifylline (FIG. 5, column A) after 30 minutes was similar to the amount Compound 419(5) formed in human liver microsomes incubated with Compound 409 (FIG. 5, column B). Thus, deuteration of pentoxifylline as embodied by Compound 409 had no discernable effect on the relative level of deuterated (5)-Ml metabolite (Compound 419(5)) formed as compared to the relative level of undeuterated (S)-M1 formed from undeuterated pentoxifylline. These results in human liver microsomes were consistent with those seen using rat whole blood.
For the reverse reaction, (5)-Ml to the keto-compound, deuteration did have an appreciable effect. Column C in FIG. 5 shows a significant amount of pentoxifylline present 30 minutes after addition of (5)-Ml . By contrast, after addition of Compound 435 (5), the level of Compound 409 that was detected after 30 minutes was less than the level of (S)-M1 (Fig 5, column D). Approximately 30% more pentoxifylline was produced from (S)-M1 than Compound 409 produced from Compound 435 (5). Under these conditions, the deuterium substitution in Compound 435 (5) impedes the conversion of this compound to the corresponding ketone. While deuterium had a greater effect in rat blood, the results are consistent.
A dramatic deuterium effect on the metabolism of (i?)-Ml metabolite was observed in human liver microsomes. Deuteration of (i?)-Ml (Compound 435(R)) reduced by almost 5 -fold the amount of deuterated pentoxifylline formed
(Compound 409) after 30 minute incubation with human liver microsomes as compared to the amount of undeuterated pentoxifylline formed from undeuterated (i?)-Ml (comparing columns E and F in FIG. 5). FIG. 4 shows the time course of the specific metabolite produced during incubation of the administered compound with human liver microsomes.
Example 37. Pharmacokinetic Study in Rats of (5)-Ml and Compound 435(5) After Oral and Intravenous Dosing.
(iS)-Ml and Compound 435(5) (a deuterated form of (5)-Ml) were separately dissolved in saline at a concentration of 10 mg/mL. A 1 : 1 mixture of the two compounds was then prepared containing a final concentration of 5 mg/mL of each compound, which was used for intravenous administration. For oral administration the mixture was further diluted in saline to a final concentration of 1 mg/mL for each compound.
Three male Sprague-Dawley rats were used in each of the oral and intravenous studies. Animals were fasted overnight prior to administration of compounds. Intravenous administration was achieved by bolus injection of a single 5 mg/kg dose of the 1 : 1 combination into the cannulated jugular vein of the rats. Cannulation was achieved the day prior to dosing on rats that had been placed under anesthesia using ketamine (IM 30 mg/kg). Oral administration was achieved by oral gavage of a single 5 mg/kg dose. Blood samples (250 μί) were collected from the dosed rats at various times post-dosing (2 min, 5 min, 10 min, 20 min, 30 min, 1 hr, 2 hr, 3 hr, 4 hr, 5 hr, 6 hr) by retro-orbital sampling of the rats temporarily anesthetized with isoflurane. Blood samples were placed in tubes containing K2- EDTA and stored on ice until centrifuged. Within 30 minutes of collection, plasma was isolated by centrifugation. A 100-μΙ, aliquot was removed, mixed with 200 μΙ_, of acetonitrile and stored at -20 °C until further analysis by LC-MS/MS using an Applied Bio-systems API 4000 mass spectrometer.
Samples were analyzed for the presence of the administered compound, the corresponding ketone (pentoxifylline and Compound 409) and the corresponding M5 metabolite. Samples (10 μί) were injected into a Zorbax SB-C8 (Rapid
Resolution) column (2.1 x 30 mm, 3.5 μιη). The initial mobile phase condition was 100% A (10 mM ammonium acetate in water) and 0% B (methanol) with a flow rate at 0.5 mL/min. Mobile phase B was allowed to reach 55% in 3 minutes and from 55% to 90% in 1 minute before ramping back to 0% in another minute. The overall run time was 5 minutes. For pentoxifylline and its Ml and M5 metabolites, the precursor/product ion pairs were set at m/z 281/193 (Ml), m/z 279/181
(pentoxifylline), and m/z 267/221 (M5).
For Compound 435(5) and Compound 409 more than one ion pair was set up for to detect species that arose from loss of deuterium. It was found that some degree of deuterium loss occurs on those compounds of the invention, such as Compound 409, which have deuterium on the side chain at positions adjacent to the carbonyl carbon. This loss of deuterium appears to occur both in vivo and ex vivo by an unknown mechanism. The addition of acetonitrile to serum samples was used to stop any additional ex vivo deuterium loss prior to analysis. Typically, no more than 2 deuterium atoms were replaced by hydrogen. For Compound 435(5), there is a deuterium at the methinyl position which was lost upon oxidation to the keto- compound 409. Reduction of 409 to an Ml metabolite introduced a proton at the methinyl position. When serum from animals dosed with 435(5) were analyzed to
- I l l - quantitate administered compound and metabolites, compound species were included with one and two less side chain deuteriums in the total amounts (referred to hereinafter as the "-1D" and the "-2D" species). Thus, for Compound 435(5) and Compound 409 separate ion pairs were set up to detect the compound and its corresponding - ID and -2D species. For Compound 435(5) three ion pairs were detected: m/z 291/197, 290/197, and 189/197. For Compound 409 ion pairs of m/z 288/186, 287/186 and 286/186 were monitored. Inclusion of -ID and -2D species in the measurements of Compound 409 and Compound 435(5) more accurately quantitates the total active species and is reasonable based on what is known about the metabolism and activities of pentoxifylline and its M-l metabolites. Increased plasma exposure to Compound 409 or any M-l metabolites of 409 would be desirable. This includes the - ID and -2D species.
For the corres onding deuterated M5 metabolite (M5a):
Figure imgf000113_0001
(M5a), which has no deuterium on its acid side chain, only one ion pair was used at m/z 271/225. The internal standard for the analysis was indiplon.
Table 12. Pharmacokinetic Results After Oral Administration of 435(S) and (S)-Ml in Rats.
Figure imgf000113_0002
a) Mass observed via LC-MS/MS. Stereochemistry presumed to be >95%> (S) based on published pentoxifylline metabolism reports.
b) % Difference = [(deuterated species)-(nondeuterated species)] (100)/(nondeuterated species)
The results of the oral administration in rats are shown in Table 12. The deuterated Compound 435(5) demonstrated a significantly higher AUCo_∞ and Cmax than its undeuterated counterpart (5)-Ml . Because there is a significant serum interconversion between (5)-Ml and pentoxifylline and both species are
therapeutically active, we also quantitated AUCo-∞ and Cmax for (5)-Ml together with pentoxifylline, and for Compound 435(5) together with Compound 409.
Compound 435(5) together with Compound 409 demonstrated a significantly higher AUCo-oo and Cmax than did (5)-Ml together with pentoxifylline after the oral administration of (5)-Ml and 435(5) respectively.
The AUCo-oo was also measured for the M-5 and M5a metabolites arising from the oral administration of (5)-Ml and 435(5), respectively. The M-5 metabolite may be associated with toxicity in certain patients and is considered undesirable. Table 12 shows that oral administration of Compound 435(5) provides considerably less M5a compared to the level of M5 obtained after administration of non-deuterated (5)-Ml . The ratio of active species to M5 metabolite was much more favorable for the deuterated compounds than for the non-deuterated compounds. The ratio of (Compound 435(5) + Compound 409) to M5a was 7.0, which was much better than the ratio of 1.6 for ((5)-Ml + pentoxifylline) to M5.
Table 13. Pharmacokinetic Results After Intravenous Administration in Rats.
Figure imgf000114_0001
a) Mass observed via LC-MS/MS. Stereochemistry presumed to be >95%> (S) based on published pentoxifylline metabolism reports. b) % Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
Table 13 shows the results following intravenous administration in rats. The results for intravenous administration were similar to those for oral administration. Compound 435(5) had an average AUCo_∞ that was 1 10% greater than its undeuterated counterpart (5)-Ml after intravenous administration. Compound
435(5) together with Compound 409 had an average AUCo_∞ that was 79% greater than (iS)-Ml together with pentoxifylline after intravenous administration.
Intravenous administration of Compound 435(5) provides an amount of M5a metabolite that is 15% less than the amount of M5 metabolite than is provided by intravenous administration of (5)-Ml . The ratio of active species to the
corresponding M5 metabolite in rats that were intravenously administered
Compound 435(5) was 7.4 as compared to 3.5 for rats that were intravenously administered (S)-M 1.
Example 38. Pharmacokinetic Study of Pentoxifylline and Compound 435(S) in Chimps After Oral and Intravenous Dosing.
Pentoxifylline and Compound 435(5) were separately dissolved in warm (65 °C) saline at a concentration at 10 mg/mL. A 1 : 1 mixture of the two compounds was then prepared containing a final concentration of 5 mg/mL of each compound and the mixture was then sterile filtered through a 0.2-μιη filter.
Two chimps (one male and one female) were used in each of the oral and intravenous studies. Animals were fasted overnight prior to administration of compounds. All animals were sedated with ketamine (approximately 10 mg/kg) and/or telazol (approximately 5 mg/kg) prior to dosing. Intravenous administration was achieved by IV infusion of 75 mg of each compound (15 mL total dosing solution) over 10 minutes. Oral administration was achieved by oral gavage of a single 75 mg dose of each compound (15 mL total dosing solution). Blood samples (6 mL) were collected from the dosed chimps at various times prior to and after dosing. For intravenous administrations blood samples were collected at 0 min (preinfusion), 5 min, 9.5 min (immediately before the end of the infusion), then 6, 15, 30 and 45 min, and 1 , 2, 4, 6, 8, 10 and 12 hr after the infusion is stopped. For oral administrations, blood samples were collected at 0 min (predose), 15 and 30 min, and 1 , 1.5, 2, 4, 6, 8, 10 and 12 hr ostdose.
Blood samples were placed in tubes containing sodium heparin, mixed and stored on ice until centrifuged. Within 30 minutes of collection, plasma was isolated by centrifuging the blood samples and removing an aliquot (200 μί) of the resulting plasma. Each 200-μί aliquot of plasma was mixed with 400 acetonitrile and stored at -70 °C until further analysis by LC-MS/MS using an Applied Bio-systems API 4000 mass spectrometer.
The analysis of all samples by LC-MS/MS was performed as described above for the rat plasma samples in Example 37.
Table 14. Pharmacokinetic Results Following Oral Administration in Chimps.
Figure imgf000116_0001
a) Mass observed via LC-MS/MS. Stereochemistry presumed to be >95%> (S) based on published pentoxifylline metabolism reports.
b) %> Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
Table 14 shows the results of oral administration of 435(5) and
pentoxifylline in chimps. Following oral administration of a 1 : 1 combination of Compound 435(5) and pentoxifylline, both Compound 435(5) and its corresponding ketone Compound 409 demonstrated significantly higher average AUCo_∞ values than the corresponding undeuterated counterparts, S)-M1 and pentoxifylline. The average AUCo_∞ for Compound 435(5) together with Compound 409 was significantly higher than the average AUCo_∞ for (5)-Ml together with
pentoxifylline. In addition, the average AUCo-∞ for the undesired deuterated M-5 metabolite (M5a) was significantly lower than that of the undeuterated M-5.
Finally, the ratio of active species to M5 metabolite for the deuterated compounds {(435(5) + 409) : (deuterated M5)} was approximately 8-fold higher than the corresponding ratio for the undeuterated species {((S)-M1 + pentoxifylline) : M5} .
Table 15. Pharmacokinetic Results Following Intravenous Administration in Chimps.
Figure imgf000117_0001
on published pentoxifylline metabolism reports. b) % Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
Table 15 shows the results of intravenous administration of 435(5) and pentoxifylline in chimps. The results following intravenous administration showed favorable differentiation of the deuterated compounds, though not as pronounced as those observed following oral administration. Compared to administration of pentoxifylline, the amounts of active species produced from the administration of Compound 435(5) were between 40 and 57% higher, while the amounts of M5 metabolite produced decreased by between 60 and 65%. The ratio of active species to M5 metabolite in chimps that were intravenously administered Compound 435(5) was approximately 4-fold higher than in chimps administered pentoxifylline.
The above results show that compounds of this invention provide significantly greater plasma exposure of desired active species than the corresponding non-deuterated compounds. Moreover, deuterium substitution in the present compounds was shown to reduce levels of the M5 metabolite, which may be associated with intolerability in renally-impaired patients.
Example 39. Pharmacokinetic Study of Pentoxifylline and Compound 435(5*) in Chimps After Oral Dosing.
Pentoxifylline and Compound 435(5) were tested according to a protocol similar to the Oral Dosing protocol followed in Example 38, except that (a) the compounds were separately dissolved in water rather than saline; (b) the mixture of the two compounds was not sterile filtered; and (c) oral administration was achieved by
oral gavage of a single 65 mg dose of each compound (13 mL total dosing solution). Table 16. Pharmacokinetic Results Following Oral Administration in Chimps.
Figure imgf000118_0001
a) Mass observed via LC-MS/MS. Stereochemistry presumed to be >95%> (S) based on published pentoxifylline metabolism reports.
b) % Difference = [(deuterated species)-(nondeuterated
species)] (100)/(nondeuterated species)
Table 16 shows the results of oral administration of 435(5) and
pentoxifylline in chimps. Similarly to what was shown in Table 14, both Compound 435(5) and its corresponding ketone Compound 409 demonstrated significantly higher average AUCo-12 values than the corresponding undeuterated counterparts, (iS)-Ml and pentoxifylline, where AUCo-12 refers to the area under the curve for the 0- to 12-hour period. The average AUCo-12 for Compound 435(5) together with Compound 409 was significantly higher than the average AUCo-12 for (5)-Ml together with pentoxifylline.
The average AUCo-12 for the deuterated M-5 metabolite (M5a) was significantly lower than that of the undeuterated M-5.
Example 40. Pharmacokinetic Study of 435(5) and other representative compounds in Chimps After Oral Dosing.
435(5) and representative compounds 12US), 137(5), 421(5) and 437(5) were separately dissolved in warm (65 °C) water at a concentration [[of 10 mg/mL]]. The same protocol described for Example 39 was then followed for each
representative compound.
Table 17 i) - iv). Pharmacokinetic Results Following Oral Administration in
Chimps.
i)
Figure imgf000119_0001
a) Mass observed via LC-MS/MS. iii)
Figure imgf000120_0001
a) Mass observed via LC-MS/MS.
Table 17 i)-iv) shows the results of oral administration of 435(5) and of compounds 121(5), 137(5), 421(5) and 437(5), respectively, in chimps. For each chimp, the values of AUC0_i2 for each of 121(5), 137(5), 421(5) and 437(5) are comparable to the values of AUCo-12 for of 435(5). Similarly, the sum of AUCo-12 values for 435(5) and 409 is comparable to the sums of AUCo-12 values for each of 121(5), 137(5), 421(5) and 437(5) and their respective ketone metabolites 107, 107, 407 and 407.
Finally, comparable values were also found for 435(5) metabolite M5a and the corresponding metabolites of 121(5), 137(5), 421(5) and 437(5), i.e., M5, M5, M5b and M5b (shown below).
Figure imgf000121_0001
Without further description, it is believed that one of ordinary skill in the art can, using the preceding description and the illustrative examples, make and utilize the compounds of the present invention and practice the claimed methods. It should be understood that the foregoing discussion and examples merely present a detailed description of certain preferred embodiments. It will be apparent to those of ordinary skill in the art that various modifications and equivalents can be made without departing from the spirit and scope of the invention.

Claims

CLAIMS ; claimed is:
1. A compound of Formula C
Figure imgf000122_0001
C
or a pharmaceutically acceptable salt thereof, wherein R1 is selected from
2 1 2
-CH3 and -CD3; R is selected from -CH3 and -CD3; one of Y and Y is -OH; and the other of Y 1 and Y 2 is deuterium or hydrogen^
provided that if R1 is CD3 and R2 is CH3, then Y2 is -OH.
2. The compound of claim 1, wherein R1 is -CH3.
3. The compound of claim 1, wherein R1 is -CD3.
4. The compound of claim 1, 2 or 3 wherein R is -CH3.
5. The compound of claim 1, 2 or 3 wherein R is -CD3.
6. The compound of claim 1, wherein R 1 is -CH3 and R 2 is -CH3.
7. The compound of any one of claims 1 to 6, wherein the compound ha the structure:
Figure imgf000122_0002
or a pharmaceutically acceptable salt thereof.
8. The compound of claim 7, wherein Y1 is deuterium.
9. The compound of claim 7, wherein Y1 is hydrogen.
10. The compound of any one of claims 1 to 6, wherein the compound has the structure:
Figure imgf000123_0001
or a pharmaceutically acceptable salt thereof.
11. The compound of claim 10, wherein Y1 is deuterium.
12. The compound of claim 10, wherein Y1 is hydrogen.
13. The compound of claim 1, wherein the compound is selected from the group consisting of the following compounds
Figure imgf000123_0002
pharmaceutically acceptable salt thereof.
14. A compound of Formula D :
Figure imgf000124_0001
D
or a pharmaceutically acceptable salt thereof, wherein R is selected from
2 1 2
-CH and -CD ; R is selected from -CH and -CD ; one of Y and Y is -OH; and the other of Y 1 and Y 2 is deuterium or hydrogen;
with the proviso that
1 2 2
(i) if either one of Y and Y is deuterium, then R is CD3; and
(ii) if either one of Y 1 and Y2 is hydrogen, then R 1 is CH3.
15. The compound of claim 14, wherein R1 is -CH3.
16. The compound of claim 14, wherein R1 is -CD3.
17. The compound of claim 14, 15 or 16, wherein R is -CH3.
18. The compound of claim 14, 15 or 16, wherein R is -CD3.
19. The compound of claim 14, wherein R 1 is -CH3 and R 2 is -CH3.
20. The compound of any one of claims 14 to 19, wherein the compound has the structure
Figure imgf000124_0002
or a pharmaceutically acceptable salt thereof.
21. The compound of claim 20, wherein Y1 is deuterium.
22. The compound of claim 20, wherein Y1 is hydrogen.
23. The compound of any one of claims 14 to 19, wherein the compound has the structure
Figure imgf000125_0001
or a pharmaceutically acceptable salt thereof.
24. The compound of claim 23, wherein Y is deuterium.
25. The compound of claim 23, Y is hydrogen.
26. The compound of claim 14, wherein the compound is selected from the group consisting of the following compounds:
Figure imgf000125_0002
or a pharmaceutically acceptable salt thereof.
The compound of any of claims 1, 7, 10, 13, 14, 20, 23 or 26, wherein any atom not designated as deuterium in any of the embodiments set forth above is present at its natural isotopic abundance.
28. A pharmaceutical composition comprising a compound of claim 1 or 14 and a pharmaceutically acceptable carrier.
29. A method of treating a disease or condition in a patient in need
thereof, comprising administering to the patient an effective amount of a composition of claim 28, wherein the disease is selected from diabetic nephropathy, hypertensive nephropathy or intermittent claudication on the basis of chronic occlusive arterial disease of the limbs.
30. A method of treating chronic kidney disease in a patient in need
thereof, comprising administering to the patient an effective amount of a composition of claim 28.
31. The method of claim 30 wherein the chronic kidney disease is
glomerulonephritis, focal segmental glomerulosclerosis, nephrotic syndrome, reflux uropathy, or polycystic kidney disease.
32. A method of treating chronic disease of the liver in a patient in need thereof, comprising administering to the patient an effective amount of a composition of claim 28.
33. The method of claim 32 wherein the chronic disease of the liver is nonalcoholic steatohepatitis, fatty liver degeneration or other diet- induced high fat or alcohol-induced tissue-degenerative conditions, cirrhosis, liver failure, or alcoholic hepatitis.
34. A method of treating a diabetes-related disease or condition in a patient in need thereof, comprising administering to the patient an effective amount of a composition of claim 28, wherein the disease or condition is selected from insulin resistance, retinopathy, diabetic ulcers, radiation-associated necrosis, acute kidney failure or drug- induced nephrotoxicity.
35. A method of treating intermittent claudication in a patient in need thereof, comprising administering to the patient an effective amount of a composition of claim 28.
36. A method of treating chronic kidney disease in a patient in need
thereof, comprising administering to the patient an effective amount of a composition of claim 28.
37. A method of treating a disease or condition in a patient in need thereof, wherein the disease or condition is selected from insulin dependent diabetes; non-insulin dependent diabetes; metabolic syndrome; obesity; insulin resistance; dyslipidemia; pathological glucose tolerance; hypertension; hyperlipidemia; hyperuricemia; gout; and hypercoagulability, comprising administering to the patient an effective amount of a composition of claim 28.
A method of treating a disease or condition in a patient in need thereof, wherein the disease or condition is selected from anemia, Graves disease, retinal vein occlusion, lupus nephritis, macular degeneration, myelodysplasia, pruritis of HIV origin, pulmonary hypertension, retinal artery occlusion, intestinal inflammation, ischemic optic neuropathy, acute pancreatitis, sickle cell anemia and beta thalassemia, comprising administering to the patient an effective amount of a composition of claim 28.
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MX2012002705A MX2012002705A (en) 2009-09-02 2010-09-02 Substituted xanthine derivatives.
ES10814504.6T ES2542401T3 (en) 2009-09-02 2010-09-02 Xanthine substituted derivatives
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CA2771123A CA2771123C (en) 2009-09-02 2010-09-02 Substituted xanthine derivatives
JP2012528054A JP5292516B2 (en) 2009-09-02 2010-09-02 Substituted xanthine derivatives
EA201270339A EA023809B1 (en) 2009-09-02 2010-09-02 Deuterated xanthine derivatives and use thereof
DK10814504.6T DK2473053T3 (en) 2009-09-02 2010-09-02 SUBSTITUTED xanthine derivatives
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PL10814504T PL2473053T3 (en) 2009-09-02 2010-09-02 Substituted xanthine derivatives
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SI201030985T SI2473053T1 (en) 2009-09-02 2010-09-02 Substituted xanthine derivatives
ZA2012/01295A ZA201201295B (en) 2009-09-02 2012-02-21 Substituted xanthine derivatives
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SI2473053T1 (en) 2015-10-30
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EP2473053A1 (en) 2012-07-11
CA2771123C (en) 2017-11-14
BR112012008108B1 (en) 2019-10-22
ES2542401T3 (en) 2015-08-05
CA2771123A1 (en) 2011-03-10
JP2013503894A (en) 2013-02-04
JP5292516B2 (en) 2013-09-18
HUE026800T2 (en) 2016-07-28
EA023809B1 (en) 2016-07-29
EP2473053B1 (en) 2015-04-22
EA201270339A1 (en) 2012-09-28
HRP20150797T1 (en) 2015-09-11
PT2473053E (en) 2015-09-14
US20110053961A1 (en) 2011-03-03
CN102480968B (en) 2015-01-07
KR20120050509A (en) 2012-05-18
EP2963040A1 (en) 2016-01-06
PL2473053T3 (en) 2015-10-30
CN102480968A (en) 2012-05-30
AU2010289465B2 (en) 2016-03-10
CN104478881A (en) 2015-04-01
KR101591137B1 (en) 2016-02-02
IN2012DN01641A (en) 2015-06-05
ME02177B (en) 2015-10-20
ZA201201295B (en) 2012-10-31
BR112012008108A2 (en) 2015-09-22
DK2473053T3 (en) 2015-07-27
HK1170377A1 (en) 2013-03-01
AU2010289465A1 (en) 2012-03-01

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