NZ727649B2 - Radiolabelled derivatives of a 2-amino-6-fluoro-n-[5-fluoro-pyridin-3-yl]- pyrazolo[1,5-a]pyrimidin-3-carboxamide compound useful as atr kinase inhibitor, the preparation of said compound and different solid forms thereof - Google Patents
Radiolabelled derivatives of a 2-amino-6-fluoro-n-[5-fluoro-pyridin-3-yl]- pyrazolo[1,5-a]pyrimidin-3-carboxamide compound useful as atr kinase inhibitor, the preparation of said compound and different solid forms thereof Download PDFInfo
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- NZ727649B2 NZ727649B2 NZ727649A NZ72764915A NZ727649B2 NZ 727649 B2 NZ727649 B2 NZ 727649B2 NZ 727649 A NZ727649 A NZ 727649A NZ 72764915 A NZ72764915 A NZ 72764915A NZ 727649 B2 NZ727649 B2 NZ 727649B2
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- 125000002147 dimethylamino group Chemical group [H]C([H])([H])N(*)C([H])([H])[H] 0.000 description 1
- UXGNZZKBCMGWAZ-UHFFFAOYSA-N dimethylformamide DMF Chemical compound CN(C)C=O.CN(C)C=O UXGNZZKBCMGWAZ-UHFFFAOYSA-N 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 238000000132 electrospray ionisation Methods 0.000 description 1
- 239000003480 eluent Substances 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 125000002534 ethynyl group Chemical group [H]C#C* 0.000 description 1
- 239000012065 filter cake Substances 0.000 description 1
- 238000003818 flash chromatography Methods 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 229910052731 fluorine Inorganic materials 0.000 description 1
- 230000001738 genotoxic Effects 0.000 description 1
- 231100000024 genotoxic Toxicity 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 125000000262 haloalkenyl group Chemical group 0.000 description 1
- 125000004438 haloalkoxy group Chemical group 0.000 description 1
- 125000001188 haloalkyl group Chemical group 0.000 description 1
- 125000005114 heteroarylalkoxy group Chemical group 0.000 description 1
- 125000004446 heteroarylalkyl group Chemical group 0.000 description 1
- 239000008079 hexane Substances 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 125000002962 imidazol-1-yl group Chemical group [*]N1C([H])=NC([H])=C1[H] 0.000 description 1
- 238000010820 immunofluorescence microscopy Methods 0.000 description 1
- 125000003387 indolinyl group Chemical group N1(CCC2=CC=CC=C12)* 0.000 description 1
- 125000001041 indolyl group Chemical group 0.000 description 1
- 229910010272 inorganic material Inorganic materials 0.000 description 1
- 239000011147 inorganic material Substances 0.000 description 1
- 229910052740 iodine Inorganic materials 0.000 description 1
- 239000011630 iodine Substances 0.000 description 1
- 125000002183 isoquinolinyl group Chemical group C1(=NC=CC2=CC=CC=C12)* 0.000 description 1
- 125000001786 isothiazolyl group Chemical group 0.000 description 1
- 230000003902 lesions Effects 0.000 description 1
- UBJFKNSINUCEAL-UHFFFAOYSA-N lithium;2-methylpropane Chemical compound [Li+].C[C-](C)C UBJFKNSINUCEAL-UHFFFAOYSA-N 0.000 description 1
- WGOPGODQLGJZGL-UHFFFAOYSA-N lithium;butane Chemical compound [Li+].CC[CH-]C WGOPGODQLGJZGL-UHFFFAOYSA-N 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000001404 mediated Effects 0.000 description 1
- 125000004370 n-butenyl group Chemical group [H]\C([H])=C(/[H])C([H])([H])C([H])([H])* 0.000 description 1
- 239000012299 nitrogen atmosphere Substances 0.000 description 1
- 239000012044 organic layer Substances 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 125000001715 oxadiazolyl group Chemical group 0.000 description 1
- KJIFKLIQANRMOU-UHFFFAOYSA-N oxidanium;4-methylbenzenesulfonate Chemical compound O.CC1=CC=C(S(O)(=O)=O)C=C1 KJIFKLIQANRMOU-UHFFFAOYSA-N 0.000 description 1
- 201000002528 pancreatic cancer Diseases 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 125000006340 pentafluoro ethyl group Chemical group FC(F)(F)C(F)(F)* 0.000 description 1
- 230000000737 periodic Effects 0.000 description 1
- 239000003208 petroleum Substances 0.000 description 1
- 239000008194 pharmaceutical composition Substances 0.000 description 1
- 125000004483 piperidin-3-yl group Chemical group N1CC(CCC1)* 0.000 description 1
- 125000004482 piperidin-4-yl group Chemical group N1CCC(CC1)* 0.000 description 1
- ZLMJMSJWJFRBEC-UHFFFAOYSA-N potassium Chemical compound [K] ZLMJMSJWJFRBEC-UHFFFAOYSA-N 0.000 description 1
- 229910052700 potassium Inorganic materials 0.000 description 1
- 239000011591 potassium Substances 0.000 description 1
- BNXGMSOVZHRHKF-UHFFFAOYSA-N prop-2-enyl 2-amino-6-fluoropyrazolo[1,5-a]pyrimidine-3-carboxylate Chemical compound C1=C(F)C=NC2=C(C(=O)OCC=C)C(N)=NN21 BNXGMSOVZHRHKF-UHFFFAOYSA-N 0.000 description 1
- NSETWVJZUWGCKE-UHFFFAOYSA-N propylphosphonic acid Chemical compound CCCP(O)(O)=O NSETWVJZUWGCKE-UHFFFAOYSA-N 0.000 description 1
- 238000000425 proton nuclear magnetic resonance spectrum Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 125000000561 purinyl group Chemical group N1=C(N=C2N=CNC2=C1)* 0.000 description 1
- 125000003373 pyrazinyl group Chemical group 0.000 description 1
- 125000003226 pyrazolyl group Chemical group 0.000 description 1
- 125000002206 pyridazin-3-yl group Chemical group [H]C1=C([H])C([H])=C(*)N=N1 0.000 description 1
- 125000002098 pyridazinyl group Chemical group 0.000 description 1
- UBQKCCHYAOITMY-UHFFFAOYSA-N pyridin-2-ol Chemical compound OC1=CC=CC=N1 UBQKCCHYAOITMY-UHFFFAOYSA-N 0.000 description 1
- VTGOHKSTWXHQJK-UHFFFAOYSA-N pyrimidin-2-ol Chemical compound OC1=NC=CC=N1 VTGOHKSTWXHQJK-UHFFFAOYSA-N 0.000 description 1
- 125000000246 pyrimidin-2-yl group Chemical group [H]C1=NC(*)=NC([H])=C1[H] 0.000 description 1
- 125000004527 pyrimidin-4-yl group Chemical group N1=CN=C(C=C1)* 0.000 description 1
- 125000004528 pyrimidin-5-yl group Chemical group N1=CN=CC(=C1)* 0.000 description 1
- 125000000719 pyrrolidinyl group Chemical group 0.000 description 1
- 125000002943 quinolinyl group Chemical group N1=C(C=CC2=CC=CC=C12)* 0.000 description 1
- 239000002534 radiation-sensitizing agent Substances 0.000 description 1
- 238000001953 recrystallisation Methods 0.000 description 1
- 230000002829 reduced Effects 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 125000002914 sec-butyl group Chemical group [H]C([H])([H])C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 231100000489 sensitizer Toxicity 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 239000002002 slurry Substances 0.000 description 1
- 239000001187 sodium carbonate Substances 0.000 description 1
- 235000019345 sodium thiosulphate Nutrition 0.000 description 1
- 239000012453 solvate Substances 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000007858 starting material Substances 0.000 description 1
- LSNNMFCWUKXFEE-UHFFFAOYSA-N sulfonic acid Chemical compound OS(O)=O LSNNMFCWUKXFEE-UHFFFAOYSA-N 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- QJKDSHHDJJUUNY-UHFFFAOYSA-N tert-butyl N-(5-fluoropyridin-2-yl)carbamate Chemical compound CC(C)(C)OC(=O)NC1=CC=C(F)C=N1 QJKDSHHDJJUUNY-UHFFFAOYSA-N 0.000 description 1
- ODGCEQLVLXJUCC-UHFFFAOYSA-N tetrafluoroborate Chemical compound F[B-](F)(F)F ODGCEQLVLXJUCC-UHFFFAOYSA-N 0.000 description 1
- WHRNULOCNSKMGB-UHFFFAOYSA-N tetrahydrofuran THF Chemical compound C1CCOC1.C1CCOC1 WHRNULOCNSKMGB-UHFFFAOYSA-N 0.000 description 1
- 125000003039 tetrahydroisoquinolinyl group Chemical group C1(NCCC2=CC=CC=C12)* 0.000 description 1
- 125000000147 tetrahydroquinolinyl group Chemical group N1(CCCC2=CC=CC=C12)* 0.000 description 1
- 125000003831 tetrazolyl group Chemical group 0.000 description 1
- 125000001544 thienyl group Chemical group 0.000 description 1
- 238000006478 transmetalation reaction Methods 0.000 description 1
- 125000004306 triazinyl group Chemical group 0.000 description 1
- 125000001425 triazolyl group Chemical group 0.000 description 1
- 229910000404 tripotassium phosphate Inorganic materials 0.000 description 1
- 229910052722 tritium Inorganic materials 0.000 description 1
- YZCKVEUIGOORGS-NJFSPNSNSA-N tritium Chemical compound [3H] YZCKVEUIGOORGS-NJFSPNSNSA-N 0.000 description 1
- 238000001946 ultra-performance liquid chromatography-mass spectrometry Methods 0.000 description 1
- 125000000391 vinyl group Chemical group [H]C([*])=C([H])[H] 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
- A61K31/4427—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems
- A61K31/4439—Non condensed pyridines; Hydrogenated derivatives thereof containing further heterocyclic ring systems containing a five-membered ring with nitrogen as a ring hetero atom, e.g. omeprazole
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- 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
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
- C07B2200/00—Indexing scheme relating to specific properties of organic compounds
- C07B2200/13—Crystalline forms, e.g. polymorphs
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
- C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
- C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
- C07D487/04—Ortho-condensed systems
Abstract
The present invention relates to the process of producing compounds of formula (I-1), useful as inhibitors of ATR protein kinase. The invention relates to preparing the compounds of this invention; intermediates for the preparation of the compounds of this invention; and solid forms of the compounds of this invention. of this invention.
Description
/032879
RADIOLABELLED DERIVATIVES OF A 2—AMINO—6—FLUORO—N—[5—FLUORO—PYRIDIN—3—YL]—
PYRAZOLO[1,5—A]PYRIMIDINCARBOXAMIDE COMPOUND USEFUL AS ATR KINASE
INHIBITOR, THE PREPARATION OF SAID COMPOUND AND DIFFERENT
SOLID FORMS THEREOF
OUND OF THE INVENTION
ATR (“ATM and Rad3 related”) kinase is a protein kinase involved in cellular
responses to DNA damage. ATR kinase acts with ATM (“ataxia telangiectasia mutated”)
kinase and many other ns to regulate a cell’s response to DNA damage, commonly
referred to as the DNA Damage se (“DDR”). The DDR stimulates DNA repair,
promotes survival and stalls cell cycle progression by ting cell cycle checkpoints,
which provide time for repair. Without the DDR, cells are much more sensitive to DNA
damage and readily die from DNA lesions induced by endogenous cellular processes such as
DNA replication or exogenous DNA damaging agents commonly used in cancer therapy.
Healthy cells can rely on a host of different proteins for DNA repair including the
DDR kinase ATR. In some cases these proteins can compensate for one another by activating
functionally redundant DNA repair processes. On the contrary, many cancer cells harbour
defects in some of their DNA repair processes, such as ATM signaling, and therefore display
a r reliance on their remaining intact DNA repair ns which include ATR.
In addition, many cancer cells express activated oncogenes or lack key tumour
suppressors, and this can make these cancer cells prone to dysregulated phases of DNA
replication which in turn cause DNA damage. ATR has been implicated as a critical
component of the DDR in se to ted DNA replication. As a result, these cancer
cells are more dependent on ATR activity for al than healthy cells. Accordingly, ATR
inhibitors may be useful for cancer treatment, either used alone or in combination with DNA
damaging agents, because they shut down a DNA repair mechanism that is more important
for cellular survival in many cancer cells than in healthy normal cells.
In fact, disruption ofATR function (e. g. by gene deletion) has been shown to
promote cancer cell death both in the absence and presence of DNA damaging agents. This
suggests that ATR inhibitors may be effective both as single agents and as potent sensitizers
to radiotherapy or genotoxic chemotherapy.
For all of these reasons, there is a need for the development of potent and selective
ATR inhibitors for the treatment of cancer, either as single agents or as ation therapies
with radiotherapy or xic chemotherapy. Furthermore, it would be desirable to have a
synthetic route to ATR tors that is amenable to large-scale synthesis and improves upon
currently known s.
ATR peptide can be expressed and ed using a y of methods known in the
literature (m e.g., Kacmaz et al, PNAS 99: 10, pp6673—6678, May 14, 2002;w
Kumagai et al. Cill 124, pp943 -955, March 10, 2006; Unsal—Kacmaz et al. Molecular and
Cellular Biology, Feb 2004, p1292-1300; and Hall-Jackson et al. Oncogene 1999, 18, 6707-
6713).
BRIEF DESCRIPTION OF THE FIGURES
FIGURE 1a: XRPD Compound I-I ous free base
FIGURE 2a: TGA Compound I-1 anhydrous free base
FIGURE 3a: DSC Compound I-l anhydrous free base
FIGLRE 1b: XRPD Compound I-l hydrate
FIGLRE 2b: TGA Compound I-1 hydrate
FIGURE 3b: DSC Compound I-1 hydrate
FIGURE 1c: XRPD nd I-1 tartaric acid
FIGURE 2c: TGA Compound I-1 tartaric acid
FIGLRE 3c: DSC Compound I-1 tartaric acid
SUMMARY OF THE INVENTION
The present invention relates to solid forms of ATR inhibitors as well as deuterated
ATR inhibitors. The present invention also relates to processes and intermediates for
preparing an aminopyrazolopyrimidine compound useful as a potent inhibitor ofATR kinase.
Amino-pyrazolopyrimidine derivatives are useful as ATR inhibitors and are also useful for
preparing ATR inhibitors.
One aspect of the ion provides a process for preparing compound 1-1:
Another aspect of the present invention comprises a compound of formula I-A:
WO 87451
TH2 /
/X1\X2/X3\ \
\N / H
/ x‘—YY56 WW,“ N/ \Y7
F Y2
or a pharmaceutically acceptable salt or derivative thereof, wherein:
each Y1, Y2, Y3, Y4, Y5, Y6, and Y7 is independently hydrogen or deuterium; provided at least
one of Y1, Y2, Y3, Y4, Y5, Y6, and Y7 is deuterium;
each X1, X2, and X4 is independently selected from 12C or 13C; and
x3 is independently selected from —1ZC(O)- 0r -13C(O)-.
Yet another aspect of the invention provides solid forms of a compound of formula
1-1:
Other aspects of the invention are set forth herein.
DETAILED PTION OF THE INVENTION
Processes
Another aspect of the t invention ses a process for preparing a
compound of formula I-1:
sing the step of reacting the compound of formula 6b:
NH2 0
/ /N:N
NN / O’N
g\ //N
F CI
with a nd of formula 11:
H2N F
/ N/
:/ . 2 HCI
under suitable conditions to form an amide bond.
Suitable conditions for forming the amide bond comprises reacting the compound of
a 6b with the substituted 3-amino pyridine 11 in the presence of a solvent and an
organic base. In one embodiment, the solvent can be selected from NMP, DMF or anisole
(preferred). In another embodiment, the organic base is an aliphatic amine independently
selected from triethylamine or DIPEA (preferred).
Still other embodiments of the present invention comprises a process for preparing
the compound of formula 11:
H2N F
/ N/
N=/ . 2 HCI
by reacting the compound of a 9:
O N\
iokN l /
with a compound of formula 10:
under suitable metal catalysed cross-coupling conditions to form an adduct
containing a protected amine group; and
ting the resulting adduct to suitable deprotection conditions.
Suitable metal catalysed cross—coupling conditions include a metal catalyst, a
suitable solvent, and a suitable base. In some embodiments, the metal catalyst is a palladium
catalyst. Examples of suitable palladium catalysts e, but are not limited to,
PdClz(PPh3)2, Pd(Ph3)4, and PdClz(dppf) (wherein each Ph is phenyl, and dppf is 1,1—
bis(diphenylphosphino)ferrocene). Suitable bases include, but are not limited to, ium
phosphate, K2C03, tBuOK and Na2C03. Suitable solvents include, but are not limited to,
DME, tetrahydrofuran, toluene, and ethanol.
Suitable deprotection conditions for removing the protecting group comprises
ng the protected species in the presence of a strong acid, such as HCl (preferred), HBr,
sulfuric acid or trifluoroacetic acid.
Another embodiment provides a s for preparing a compound of formula 9:
by ng the compound of formula 8:
7LOJKNO l
under suitable halogenation conditions.
Suitable halogenation conditions comprises reacting compound 8 in an aprotic
t, in the presence of a strong base, and an electrophilic source of halogen. In one
embodiment, the t can be selected from DCM, diethylether or THF (preferred). In
another embodiment, the strong base is ed from tert—BuLi, sec-BuLi or n—BuLi
(preferred). In yet another embodiment, the electrophilic species used to introduce the
n atom can, for example, be selected from I2 (preferred), CF31, diiodoethane, Brz, CBr4.
Still other embodiments of the present invention provides a process for preparing a
compound of formula 8:
under suitable conditions to generate a protected amine group.
Suitable conditions for introducing the protecting group ses reacting the
amino species 7 in an aprotic solvent, in the presence of B0020. Such on can be
conducted in the presence of a base. In one embodiment, the solvent can be selected from
diethylether or THF (preferred). In another ment, the strong base can be selected from
DMAP, n-BuLi, LHMDS or NaHMDS (preferred).
Deuterated Compounds
Isotopes can be introduced on compound 1-1 by selecting building blocks that
contain the isotopic atoms (either commercial or that can be prepared ing to the
literature) and engaging them into a sequence similar to the novel and inventive s
reported for the unlabelled material (described above).
Another aspect of the present invention provides a nd of Formula I-A:
NH2 /
x1 x3
N/ \xz/ \N \
\N4/< :3 /Y5
r1wN N/XK—YG
F Y2
or a pharmaceutically acceptable salt or derivative thereof, wherein:
each Y1, Y2, Y3, Y4, Y5, Y6, and Y7 is independently hydrogen or ium; provided at least
one of Y1, Y2, Y3, Y4, Y5, Y6, and Y7 is deuterium;
each X1, X2, and X4 is independently selected from 12C or 13C; and
X3 is independently selected from —12C(O)- or -13C(O)-.
The following labelled building blocks, which can be used in the synthetic route for
ing the compound of a I-A, are all commercially available:
0 1, 2-Di13C-2—cyanoacetic acid;
0 1-13C—2-cyano(13C)acetic acid ethyl ester;
0 2-13C—2-cyano(13C)acetic acid ethyl ester;
0 1-(trideuteromethyl)-1H-imidazole;
0 2,4,5 -trideuter0(methyl)-1H—imidazole; and
o 2 ,4,5 utero(trideuteromethyl)-1H-imidazole.
Other ed building blocks, which can be used in the synthetic route for
preparing the compound of Formula I-A, are known to those skilled in the art. These may
include, but are not limited to, the following labelled building blocks:
- 2-cyano(13C)acetic acid; Triplett et al., J Labelled Comp Radiopharm, 1978, 14(1),
0 1-13Ccyanoacetic acid; Matsumoto et al., Heterocycles, 1985, 23(8), 2041;
- 2-13C—2-cyanoacetic acid; n et al., J Am Chem Soc, 1989, 111(9), 3319;
0 1-deutero(diethylamino)—2-fluoroacrylaldehyde; ki et al., Chem Lett, 1997,
(8), 739;
o 2-deutero-1—(methyl)-lH—imidazole; Torregrosa et al., Tetrahedron, 2005, 61(47),
11148—11155;
- 4,5-dideutero—1-(methyl)—1H—imidazole; Pavlik et al., J. Org. Chem, 1991, 56(22),
6313-6320;
0 4,5-dideutero(trideuteromethyl)-lH-imidazole; Mamer et al., Rapid
Communications in Mass Spectrometry, 2005, 19(12), 1771—1774;
- 2-tritio-1—(methyl)-1H—imidazole; Buncel et 211., Can. J. Chem., 1986, 64(6), 1240—
1245;
- 2,4,5-tritritio(methyl)—1H-imidazole; Grimmett, Scien of Synthesis, 2002, 325-528;
- 1-(13C-methyl)-1H-imidazole; Van Thuijl et al., Organic Mass ometry, 1973,
7(10), 1165-1172.
In one or more embodiments of the present invention, Y1, Y2, Y3, and Y4 are
independently selected from deuterium or hydrogen; and Y5, Y6, and Y7 are deuterium.
In some embodiments, Y1 and Y2 are independently selected from deuterium or
hydrogen; and Y3, Y4, Y5, Y6, and Y7 are deuterium.
In another embodiment, Y1, Y2, Y5, Y6, and Y7 are independently selected from
deuterium or hydrogen; and Y3 and Y4 are deuterium.
In other ments, Y1, Y3, and Y4 are ndently selected from deuterium or
hydrogen; and Y2, Y5, Y6, and Y7 are deuterium.
In still other embodiments, Y1, Y2, Y3, Y4, Y5, Y6, and Y7 are en; and X4 is
13C.
In yet another embodiment, Y1, Y2, Y3, Y4, Y5, Y6, and Y7 are hydrogen; and X1 and
X4 are 13C.
In some embodiments, Y1, Y2, Y3, Y4, Y5, Y6, and Y7 are hydrogen; and X3 is —
13C(0)—.
In another embodiment, Y1, Y3, Y4, Y5, Y6, and Y7 are hydrogen; Y2 is deuterium;
and X4 is 13C.
In other embodiments, Y1, Y2, Y3, and Y4 are hydrogen; Y5, Y6, and Y7 are
deuterium; and X1 is 13
In still other embodiments, Y1, Y3, Y4, Y5, Y6, and Y7 are hydrogen; Y2 is deuterium;
and X1 is 13 C.
In yet another ment, Y1, Y2, Y3, Y5, Y6, and Y7 are hydrogen; Y4 is
deuterium; and X1 is C.
In another embodiment, Y1 is hydrogen; Y2, Y3, Y4, Y5, Y6, and Y7 are deuterium;
X2 is 13C; and X3 is —13C(O)-.
In another example, the compounds of formula I-A of this invention are ented
in Table 1. It will be appreciated by those skilled in the art that the compounds of the present
invention may be represented in varying tautomeric forms.
PCT/U82015/032879
Table 1
1-2 1-3 1-4
1-6 1-7
1-8 1-10
1-12
Solid Forms
Another aspect of the present invention es a solid form of a compound of
formula I-1:
<\ N
/ /N—
wherein, the form is selected from the group consisting of Compound I-l anhydrous free
base, Compound I-1 hydrate, or Compound I-1 tartaric acid .
Compound I-1 ousfree base
In some aspects of the present inventions, the solid form is Compound I-1 anhydrous
free base. In another aspect of the present invention, the solid form is crystalline Compound
I-1 anhydrous free base. In some embodiments, the solid form is characterized by one or
more peaks expressed in 2—theta :: 0.2 at about 9.9, 12.8, 15.4, 17.0, 23.1, 27.8, 29.0, and 30.1
degrees in an X—Ray powder diffraction pattern ed using Cu K alpha radiation. In other
embodiments, the solid form is characterized as having an X-ray powder diffraction pattern
ntially the same as that shown in Figure la.
Compound I—I hydrate
In some s of the present invention, the solid form is Compound I-l e.
In another aspect of the t invention, the solid form is crystalline Compound I-1
hydrate. In other embodiments, the crystalline Compound I-1 hydrate has a Compound I-1 to
water ratio of 1:3. In still other embodiments, Compound I-1 hydrate is characterized by a
weight loss of from about 12.6% in a temperature range from about 40°C and about 100°C.
In some embodiments, the solid form is characterized by one or more peaks expressed in 2-
theta :: 0.2 at about 27.5, 20.6, and 9.7 degrees in an X-Ray powder diffraction pattern
obtained using Cu K alpha radiation. In yet other embodiments, the solid form is
characterized as having an X-ray powder diffraction pattern substantially the same as that
shown in Figure 1b.
Compound I—I ic acid
In some aspects of the present invention, the solid form is Compound I-1 tartaric
acid. In another aspect of the present invention, the solid form is lline Compound I-1
tartaric acid. In other embodiments, the crystalline Compound I-1 tartaric acid has a
Compound I-1 to ic acid ratio of 1:1. In some embodiments, the solid form is
2015/032879
characterized by one or more peaks expressed in 2-theta i 0.2 at about 7.1, 18.3, and 13.2
degrees in an X—Ray powder diffraction pattern obtained using Cu K alpha radiation. In yet
other embodiments, the solid form is characterized as having an X-ray powder diffraction
pattern substantially the same as that shown in Figure 1c.
For purposes of this application, it will be understood that the terms embodiment,
example, and aspect are used interchangeably.
Compounds of this invention include those described lly herein, and are
further illustrated by the classes, subclasses, and species disclosed herein. As used herein, the
following definitions shall apply unless otherwise indicated. For purposes of this invention,
the al elements are identified in accordance with the Periodic Table of the Elements,
CAS version, Handbook of Chemistry and s, 75th Ed. Additionally, l principles
of organic try are described in “Organic Chemistry”, Thomas l, sity
Science Books, Sausalito: 1999, and “March’s Advanced Organic Chemistry”, 5th Ed., Ed.:
Smith, MB. and March, J., John Wiley & Sons, New York: 2001, the entire contents of
which are hereby incorporated by reference.
As described herein, a specified number range of atoms includes any integer n.
For example, a group having from 1—4 atoms could have 1, 2, 3, or 4 atoms.
As described herein, compounds of the ion may optionally be substituted with
one or more substituents, such as are illustrated generally herein, or as exemplified by
particular classes, subclasses, and species of the invention. It will be appreciated that the
phrase “optionally substituted” is used interchangeably with the phrase ituted or
unsubstituted.” In general, the term “substituted”, whether preceded by the term “optionally”
or not, refers to the replacement of hydrogen radicals in a given structure with the radical of a
specified substituent. Unless otherwise indicated, an optionally substituted group may have a
substituent at each substitutable position of the group, and when more than one position in
any given structure may be substituted with more than one substituent selected from a
specified group, the substituent may be either the same or different at every position.
Combinations of substituents envisioned by this invention are preferably those that result in
the formation of stable or chemically feasible compounds.
Unless otherwise indicated, a substituent connected by a bond drawn from the center
of a ring means that the substituent can be bonded to any on in the ring. In example i
below, for instance, 1‘” can be bonded to any position on the pyridyl ring. For bicyclic rings,
a bond drawn h both rings indicates that the tuent can be bonded from any
position of the ic ring. In example ii below, for instance, JW can be bonded to the 5—
membered ring (on the nitrogen atom, for instance), and to the ered ring.
/ NA '
—|—(JW)0.5 5
\ §_<NJ\//
N H
i ii
The term “stable”, as used herein, refers to compounds that are not substantially
altered when subjected to conditions to allow for their production, detection, ry,
purification, and use for one or more of the purposes disclosed . In some embodiments,
a stable compound or chemically feasible compound is one that is not substantially altered
when kept at a temperature of 40°C or less, in the absence of moisture or other chemically
reactive conditions, for at least a week.
The term “aliphatic” or “aliphatic group”, as used herein, means a straight-chain
(i.e., unbranched), branched, or cyclic, substituted or unsubstituted hydrocarbon chain that is
tely saturated or that contains one or more units of unsaturation that has a single point
of attachment to the rest of the molecule.
Unless otherwise specified, aliphatic groups contain 1-20 aliphatic carbon atoms. In
some embodiments, tic groups n 1-10 aliphatic carbon atoms. In other
embodiments, aliphatic groups contain 1—8 tic carbon atoms. In still other
embodiments, aliphatic groups contain 1—6 aliphatic carbon atoms, and in yet other
embodiments aliphatic groups contain 1—4 aliphatic carbon atoms. Aliphatic groups may be
linear or branched, substituted or unsubstituted alkyl, alkenyl, or alkynyl groups. Specific
examples include, but are not limited to, methyl, ethyl, isopropyl, n-propyl, sec-butyl, vinyl,
n—butenyl, ethynyl, and tert—butyl. Aliphatic groups may also be cyclic, or have a combination
of linear or branched and cyclic groups. Examples of such types of aliphatic groups e,
but are not limited to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclohexenyl, -CH2-
cyclopropyl, CHZCHZCH(CH3)—cyclohexyl.
The term “cycloaliphatic” (or “carbocycle” or “carbocyclyl”) refers to a clic
C3-C8 hydrocarbon or bicyclic C8—C1; hydrocarbon that is completely saturated or that
contains one or more units of unsaturation, but which is not aromatic, that has a single point
of attachment to the rest of the molecule wherein any individual ring in said bicyclic ring
system has 3-7 members. Examples of cycloaliphatic groups include, but are not limited to,
cycloalkyl and cycloalkenyl . Specific examples include, but are not limited to,
cyclohexyl, cyclopropyl, and cyclobutyl.
The term “heterocycle”, “heterocyclyl”, or “heterocyclic” as used herein means non—
aromatic, monocyclic, bicyclic, or tricyclic ring systems in which one or more ring members
are an independently selected heteroatom. In some embodiments, the “heterocycle”,
“heterocyclyl”, or “heterocyclic” group has three to fourteen ring members in which one or
more ring members is a heteroatom ndently selected from oxygen, , nitrogen, or
phosphorus, and each ring in the system contains 3 to 7 ring members.
Examples of heterocycles include, but are not limited to, 3-1H—benzimidazolone,
3 —( l -alkyl)-benzimidazol-2—one, 2-tetrahydrofuranyl, 3—tetrahydrofuranyl, 2—
tetrahydrothiophenyl, 3-tetrahydrothiophenyl, holino, 3 -morpholino, 4-morpholino, 2—
rpholino, 3-thiomorpholino, 4—thiomorpholino, l—pyrrolidinyl, 2-pyrrolidinyl, 3—
pyrrolidinyl, l-tetrahydropiperazinyl, 2-tetrahydropiperazinyl, 3-tetrahydropiperazinyl, l—
dinyl, 2-piperidinyl, ridinyl, l-pyrazolinyl, 3—pyrazolinyl, 4-pyrazolinyl, 5-
pyrazolinyl, l—piperidinyl, 2—piperidinyl, 3—piperidinyl, 4—piperidinyl, 2—thiazolidinyl, 3—
thiazolidinyl, 4-thiazolidinyl, l-imidazolidinyl, 2-imidazolidinyl, 4-imidazolidinyl, 5-
imidazolidinyl, indolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, benzothiolane,
benzodithiane, and l,3-dihydro-imidazol—2-one.
Cyclic groups, (e. g. cycloaliphatic and heterocycles), can be linearly fused, bridged,
or spirocyclic.
The term “heteroatom” means one or more of , sulfur, nitrogen, orus,
or silicon (including, any ed form of nitrogen, sulfur, phosphorus, or silicon; the
quaternized form of any basic nitrogen or; a substitutable nitrogen of a heterocyclic ring, for
example N (as in hydro—2H—pyrrolyl), NH (as in pyrrolidinyl) or NR+ (as in N—
substituted pyrrolidinyl)).
The term "unsaturated", as used , means that a moiety has one or more units of
unsaturation. As would be known by one of skill in the art, unsaturated groups can be
partially unsaturated or fully unsaturated. Examples of partially unsaturated groups include,
but are not limited to, butene, cyclohexene, and ydropyridine. Fully unsaturated groups
can be aromatic, anti—aromatic, or non—aromatic. Examples of fully unsaturated groups
include, but are not limited to, phenyl, cyclooctatetraene, pyridyl, thienyl, and l-
methylpyridin-2(lH)-one.
The term “alkoxy”, or “thioalkyl”, as used herein, refers to an alkyl group, as
previously defined, attached through an oxygen (“alkoxy”) or sulfur (“thioalkyl”) atom.
The terms “haloalkyl”, “haloalkenyl”, “haloaliphatic”, and “haloalkoxy” mean alkyl,
alkenyl or alkoxy, as the case may be, substituted with one or more halogen atoms. This term
includes perfluorinated alkyl groups, such as —CF3 and -CF2CF3.
The terms “halogen”, “halo”, and “hal” mean F, Cl, Br, or I.
The term “aryl” used alone or as part of a larger moiety as in lkyl”,
“arylalkoxy”, or “aryloxyalkyl”, refers to monocyclic, bicyclic, and tricyclic ring systems
having a total of five to fourteen ring s, wherein at least one ring in the system is
ic and wherein each ring in the system contains 3 to 7 ring s. The term “aryl”
may be used interchangeably with the term “aryl ring”.
The term “heteroaryl”, used alone or as part of a larger moiety as in
“heteroarylalkyl” or “heteroarylalkoxy”, refers to monocyclic, bicyclic, and lic ring
systems having a total of five to fourteen ring members, wherein at least one ring in the
system is aromatic, at least one ring in the system contains one or more heteroatoms, and
wherein each ring in the system contains 3 to 7 ring members. The term “heteroaryl” may be
used interchangeably with the term oaryl ring” or the term “heteroaromatic”. es
of heteroaryl rings include, but are not limited to, 2-furanyl, 3-furanyl, N-imidazolyl, 2-
imidazolyl, 4-imidazolyl, 5—imidazolyl, benzimidazolyl, 3—isoxazolyl, 4-isoxazolyl, 5—
isoxazolyl, olyl, 4-oxazolyl, 5-oxazolyl, olyl, 2-pyrrolyl, 3-pyrrolyl, 2-pyridyl,
3—pyridyl, 4-pyridyl, 2—pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl, pyridazinyl (e.g., 3—
pyridazinyl), 2—thiazolyl, zolyl, 5—thiazolyl, tetrazolyl (e.g., 5—tetrazolyl), triazolyl (e. g.,
2—triazolyl and 5—triazolyl), 2—thieny1, nyl, benzofuryl, benzothiophenyl, indolyl (e.g., 2-
indolyl), pyrazolyl (e.g., 2—pyrazolyl), isothiazolyl, 1,2,3—oxadiazolyl, oxadiazolyl,
1,2,4-oxadiazolyl, 1,2,3-triazolyl, 1,2,3-thiadiazolyl, 1,3,4-thiadiazolyl, 1,2,5-thiadiazolyl,
purinyl, pyrazinyl, triazinyl, quinolinyl (e.g., 2-quinolinyl, 3-quinolinyl, 4-quinolinyl),
and isoquinolinyl (e.g., l-isoquinolinyl, 3—isoquinolinyl, or 4-isoquinolinyl).
It shall be understood that the term “heteroaryl” includes n types of heteroaryl
rings that exist in equilibrium between two different forms. More specifically, for example,
species such hydropyridine and pyridinone (and likewise hydroxypyrimidine and
pyrimidinone) are meant to be encompassed within the definition of “heteroaryl.”
\ \
l ‘— l
/N NH
OH 0
The term “protecting group” and “protective group” as used , are
interchangeable and refer to an agent used to temporarily block one or more desired
functional groups in a compound with multiple reactive sites. In certain embodiments, a
protecting group has one or more, or ably all, of the following characteristics: a) is
added selectively to a onal group in good yield to give a ted substrate that is b)
stable to reactions occurring at one or more of the other reactive sites; and c) is selectively
ble in good yield by reagents that do not attack the rated, deprotected functional
group. As would be understood by one skilled in the art, in some cases, the reagents do not
attack other reactive groups in the compound. In other cases, the reagents may also react
with other reactive groups in the compound. Examples of protecting groups are detailed in
Greene, T.W., Wuts, P. G in “Protective Groups in Organic Synthesis”, Third Edition, John
Wiley & Sons, New York: 1999 (and other editions of the book), the entire contents of
which are hereby incorporated by reference. The term “nitrogen protecting group”, as used
herein, refers to an agent used to temporarily block one or more desired nitrogen reactive
sites in a multifunctional nd. Preferred nitrogen protecting groups also possess the
characteristics exemplified for a protecting group above, and certain exemplary nitrogen
protecting groups are also detailed in Chapter 7 in Greene, T.W., Wuts, P. G in “Protective
Groups in Organic Synthesis”, Third Edition, John Wiley & Sons, New York: 1999, the
entire contents of which are hereby incorporated by reference.
In some embodiments, a methylene unit of an alkyl or aliphatic chain is ally
replaced with another atom or group. Examples of such atoms or groups include, but are not
limited to, nitrogen, oxygen, sulfur, -C(O)-, CN)-, )-, -C(=NOR)—, —SO-,
and -SOz-. These atoms or groups can be combined to form larger groups. Examples of such
larger groups include, but are not limited to, -OC(O)—, —C(O)CO-, —COz—, —C(O)NR-, —C(=NCN
), -NRCO-, -NRC(O)O-, -SOZNR—, -NRSOz-, -NRC(O)NR-, -OC(O)NR—,
and NR-, wherein R is, for example, H or C1_6aliphatic. It should be understood that
these groups can be bonded to the methylene units of the aliphatic chain via single, double, or
triple bonds. An example of an optional replacement (nitrogen atom in this case) that is
bonded to the aliphatic chain via a double bond would be —CH2CH=N-CH3. In some cases,
2015/032879
especially on the terminal end, an optional replacement can be bonded to the aliphatic group
Via a triple bond. One example of this would be CHZCHZCHZCEN. It should be understood
that in this situation, the terminal nitrogen is not bonded to another atom.
It should also be understood that, the term “methylene unit” can also refer to
branched or substituted methylene units. For example, in an isopropyl moiety [-CH(CH3)2], a
nitrogen atom (e.g. NR) replacing the first recited “methylene unit” would result in
dimethylamine [-N(CH3)2]. In instances such as these, one of skill in the art would
understand that the nitrogen atom will not have any additional atoms bonded to it, and the
“R” from “NR” would be absent in this case.
Unless otherwise indicated, the optional replacements form a chemically stable
compound. Optional replacements can occur both within the chain and/or at either end of the
chain; i.e., both at the point of ment and/or also at the terminal end. Two optional
replacements can also be adjacent to each other within a chain so long as it results in a
ally stable compound. For example, a C3 aliphatic can be ally replaced by 2
en atoms to form —C—NEN. The al replacements can also tely replace all
of the carbon atoms in a chain. For example, a C3 aliphatic can be ally replaced
by -NR—, -C(O)—, and —NR- to form -NRC(O)NR- (a urea).
Unless otherwise indicated, if the replacement occurs at the terminal end, the
replacement atom is bound to a hydrogen atom on the terminal end. For e, if a
ene unit of —CH2CH2CH3 were ally replaced with —O—, the ing compound
could be -OCH2CH3, -CH20CH3, or -CH2CH20H. It should be understood that if the
terminal atom does not contain any free valence electrons, then a hydrogen atom is not
required at the terminal end (e.g., -CH2CH2CH=O or -CH2CH2CEN).
The term "cross—coupling reaction", as used herein, refers to a reaction in which a
carbon-carbon bond is formed with the aid of a metal catalyst. Usually, one of the carbon
atoms is bonded to a functional group (a "cross-coupling group") while the other carbon atom
is bonded to a halogen. Examples of cross coupling reactions include, but are not limited to,
Suzuki couplings, Stille couplings, and Negishi couplings.
The term "cross—coupling group", as used herein, refers to a functional group capable
of reacting with another functional group (e. g., halo) in a cross coupling reaction to form a
carbon—carbon ("C-C") bond. In some embodiments, the C—C bond is formed between two
aromatic groups.
The term "cross ng condition", as used herein, refers to the chemical
ions (e.g., temperature, length of time of reaction, volume of solvent required) required
in order to enable the cross coupling reaction to occur.
Examples of cross—coupling groups and their respective cross—coupling conditions
e, but are not limited to, boronic acids and boronic esters with Suzuki coupling
conditions, SnBu3 (Bu: butyl) with Stille coupling conditions, and ZnX (X: halogen) with
Negishi coupling conditions.
All three of these coupling conditions typically involve the use of a st, a
suitable t, and optionally a base. Suzuki coupling ions involve the use of a
palladium catalyst and a suitable solvent. Examples of suitable palladium catalysts include,
but are not limited to, PdClz(PPh3)2, Pd(Ph3)4, and PdClz(dppf) (wherein each Ph is ,
and dppf is 1,1-bis(diphenylphosphino)ferrocene). Suitable bases include, but are not limited
to, K2CO3 and NaZCO3. Suitable solvents include, but are not limited to, tetrahydrofuran,
e, and ethanol.
Stille coupling conditions e the use of a catalyst (usually palladium, but
mes nickel), a suitable solvent, and other optional reagents. Examples of suitable
catalysts include, but are not limited to, PPh3)2, Pd(Ph3)4, and PdClz(dppf). Suitable
solvents include, but are not limited to, tetrahydrofuran, toluene, and dimethylformamide.
Negishi ng conditions involve the use of a catalyst (palladium or nickel) and a
suitable solvent. Examples of suitable catalysts include, but are not limited to Pd2(dba)3,
Ni(PPh3)2C12, PdClz(PPh3)2, and Pd(Ph3)4 (where “dba” is
tris(dibenzylideneacetone)dipalladium). Suitable solvents include, but are not limited to,
tetrahydrofuran, toluene, and ylformamide.
Suzuki, Stille, and Negishi ions are known to one skilled in the art and are
described in more detail in a variety of references, including "March's Advanced Organic
Chemistry".
As would be understood by one skilled in the art, cross-coupling groups are formed
from coupling group precursors. A ng group precursor is a reagent or group of reagents
used to form a cross—coupling group. Examples include, but are not limited to,
bis(pinacolato)diborane for the formation of boronate esters, trimethylborates for the
formation of boronic acids, Bu3SnCl for the ion of stannanes, and ZnClz for the
formation zincates in Negishi coupling reactions. Examples of suitable coupling group
formation conditions include, but are not limited to, making boronic esters via palladium-
mediated catalysis; making boronic acids by hydrolyzing boronic esters; making nes
via a two step process: 1) n metal ge followed by 2) transmetallation with
Bu3SnCl and making zincates via a two step process: 1) halogen metal exchange followed by
2) addition of ZnClz.
Unless otherwise indicated, structures depicted herein are also meant to include all
isomeric (e.g., enantiomeric, diastereomeric, geometric, conformational, and rotational)
forms of the structure. For example, the R and S configurations for each asymmetric center,
(Z) and (E) double bond isomers, and (Z) and (E) conformational isomers are included in this
invention. As would be understood to one d in the art, a substituent can freely rotate
around any rotatable bonds. For example, a substituent drawn as also represents
Therefore, single stereochemical isomers as well as enantiomeric, reomeric,
ric, conformational, and rotational mixtures of the present compounds are within the
scope of the invention.
Unless otherwise indicated, all tautomeric forms of the compounds of the ion
are within the scope of the invention.
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 on is
understood to have hydrogen at its natural abundance isotopic composition. Also unless
otherwise , when a position is designated specifically as "D" or "deuterium", the
position is understood to have ium 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).
"D" and "d" both refer to deuterium.
Additionally, unless otherwise indicated, structures depicted herein are also meant to
include compounds that differ only in the presence of one or more isotopically enriched
atoms. For example, compounds having the present structures except for the replacement of
hydrogen by deuterium or tritium, or the ement of a carbon by a 13C— or 14C—enriched
carbon are within the scope of this invention. Such compounds are useful, for example, as
ical tools or probes in biological assays.
As used herein “crystalline” refers to a solid that has a specific ement and/or
conformation of the molecules in the l lattice.
As used herein the term “amorphous” refers to solid forms that consist of disordered
arrangements of molecules and do not s a distinguishable crystal lattice.
As used herein, the term “solvate” refers to a crystalline solid adduct containing
either stoichiometric or nonstoichiometric amounts of a solvent orated within the
crystal structure. If the incorporated t is water, such adduct is refered to as a “hydrate”.
Abbreviations
The following abbreviations are used:
DMSO dimethyl sulfoxide
DCM dichloromethane
ATP adenosine triphosphate
TFA trifluoroacetic acid
1HNMR proton nuclear magnetic resonance
HPLC high performance liquid chromatography
LCMS liquid chromatography-mass spectrometry
Rt retention time
XRPD X-Ray Powder Diffraction
DSC Differential scanning calorimetry
TGA Thermogravimetric analysis
RT room temperature
NMP N-methyl—2-pyrrolidone
Bp g point
DMF dimethylformamide
PTSA p—Toluenesulfonic acid
DIPEA N,N-diisopropylethylamine
HOBT hydroxybenzotriazole
HATU l- [B is(dimethylamino)methylene] - l H— l ,2 ,3 -triazolo[4,5 -b]pyridinium 3—oxid
hexafluorophosphate
TBTU benzotriazole-l-yl)-1 3 -tetramethyluronium tetrafluoroborate
, 1,3 ,
T3P Propylphosphonic anhydride
COMU l- [( l -(Cyano—2-ethoxyoxoethylideneaminooxy)—dimethylamino-
morpholino)]uroniumhexafluorophosphate
TCTU [(6—chlorobenzotriazol—1—yl)oxy—(dimethylamino)methylene]—dimethyl—
ammonium tetrafluoroborate
HBTU otriazole-N,N,N ’ ,N ’ -tetramethyl-uronium-hexafluoro-phosphate
DME Dimethoxyethane
THF tetrahydrofuran
TMEDA tetramethylethylenediamine
NaHMDS sodium hexamethyldisilazane
LHMDS Lithium bis(trimethylsilyl)amide
Processes
Processes and compounds described herein are useful for producing ATR inhibitors
that contain an aminopyrazolopyrimidine core. The general synthetic procedures shown in
schemes herein are useful for generating a wide array of chemical species which can be used
in the manufacture of ceutical compounds.
SCHEMEA
NH2 0
O NH2 O /
O anion NC ,AII pyrazole dine N\ / 0’All
NCQLOAII condensation_ 0 formation N" / O’A” formation N
H” N
HZN CCI3 g\ //
1 F
2 3
amide bond
NH2 NH2
0 0
N=N NH2 0 /N
' I
/ activated ester N‘ /
amide bond
N‘ \
/ OH formation / O’N / formation N/ N F
N N / H
—. \ _. \N
[>\ N s\ N \\ / N/
// // J <\ /N 2/
F F
6a-b |_1
Compounds of this invention can be synthesised according to methods similar to the
one depicted in Scheme A.
Step 1
The anion of commercially available allyl cyanoacetate 1 can react with, e.g.,
trichloroacetonitrile to provide intermediate 2. In the anion condensation step, the anion of
commercially available allyl cetate 1 can be generated with a base such as ium
acetate in an appropriate solvent such as an alcohol (e.g., isopropylalcohol). The anion then
reacts with trichloroacetonitrile at room temperature.
$19;
Intermediate 2 then reacts with hydrazine to form the diaminopyrazole 3. In the
pyrazole formation step, intermediate 2 is reacted with hydrazine (or its hydrate) in an aprotic
solvent, such as DMF, to provide the diaminopyrazole 3. The reaction occurs under basic
conditions (e.g., in the presence of potassium e or AcONa) with heating (e.g.,z 110°C)
to ensure complete cyclisation.
Step 3
Intermediate 3 can further be condensed with a dielectrophilic coupling partner to
form the pyrimidine 4. In the dine formation step, intermediate 3 is reacted with a 1,3—
dielectrophilic species (e.g., a 1,3-dialdehyde or a 3—(dialkylamino)-prop-2—enal) in various
types of solvents (e.g., DMF or DMSO/water) to furnish the bicyclic cores 4. When one or
two of the electrophilic centers is protected/masked (e. g., aldehyde masked as a ketal),
introduction of a sulfonic acid (e.g., PTSA) is required to liberate the reactive onal
group.
Step 4
Deprotection, e.g, via hydrolysis, of the allyl ester leads to the carboxylic acids 5. In
the deprotection step, compound 4 is ted to hydrolytic conditions that are known to
those skilled in the art. For example, ent of 4 with phenylsilane or 4-
methylbenzenesulfinate in the ce of a catalytic amount of palladium (e.g., 3)4)
leads to the formation of the ponding carboxylic acid 5. Alternatively, compounds 4
could be treated with aqueous alkali (e.g., NaOH, LiOH, or KOH) to produce acids 5.
Step 5
In the activated ester formation step, the carboxylic acids 5 are reacted with amide
coupling agents known to those skilled in the art. Suitable amide coupling partners include,
but are not limited to TBTU, TCTU, HATU, T3P, and COMU. When the coupling agent is
chosen appropriately, the ons can proceed rapidly (~lhr.) at room temperature in the
presence of an c base such as an aliphatic amine (e. g., triethylamine, DIPEA) to
provide the activated esters 6a-b. For example, when the amide coupling agents TBTU [J=H]
or =Cl] are used, nds 6a-b are obtained y by filtration of the reaction
mixture.
Formation of the activated esters 6a-b prior to the amide bond formation to prepare
LA is generally preferred, although a direct conversion of 5 into the compounds of formula I-
A of this invention is also le. Alternative activated esters can also be utilised (isolated
or formed in situ) and will be known to those skilled in the art (e. g., using TBTU, TCTU,
HATU, T3P, COMU coupling agents).
Step 6
In the amide bond formation step, activated esters 6a-b can react with substituted 3—
aminopyridine 11 to provide compound I-1 of this invention. The reaction conditions for the
amide coupling are in a solvent (e.g. anisole, NMP, pyridine, DMF, etc ...) with heating (e. g.,
2 90°C).
Alternatively, the two steps described above can be combined: carboxylic acid 5 can
be used as starting points for the amide bond formation, the activated esters being generated
in situ, using the same amide couplings agents as those described above. nds of this
invention are ed in a similar manner to the one described above.
PREPARATIONS AND EXAMPLES
All commercially available solvents and reagents were used as received. Microwave
reactions were carried out using a CEM Discovery microwave. Flash Chromatography, e. g.,
was carried out on an ISCO© CombiflashR CompanionTM system eluting with a 0 to 100%
EtOAc/petroleum ether nt. Other methods known in the art were also utilized to
perform Flash Chromotography. Samples were applied pre—absorbed on silica. Where stated,
supercritical fluid tography (SFC) was performed on a Berger Minigram SFC
machine. All 1H NMR spectra were recorded using a Bruker Avance III 500 ment at
500 MHz. MS samples were analyzed on a Waters SQD mass spectrometer with electrospray
ionization operating in ve and negative ion mode. Samples were introduced into the
mass spectrometer using chromatography. All final products had a purity 295%, unless
specified otherwise in the experimental details. HPLC purity was measured on a Waters
Acquity UPLC system with a Waters SQD MS instrument ed with a Waters UPLC
BEH C8 1.7 pm, 2.1 x 50 mm column and a Vanguard BEH C8 1.7 pm, 2.1 x 5 mm guard
column.
As used herein, the term "Rt(min)" refers to the HPLC retention time, in minutes,
associated with the compound. Unless otherwise indicated, the HPLC methods utilized to
obtain the reported retention times are as described below:
HPLC Method B
Instrument: Waters Acquity UPLC-MS;
Column: Waters UPLC BEH C8 1.7 pm, 2.1 x 50 mm with Vanguard BEH C8 1.7 um, 2.1 x
mm guard column;
Column temperature: 45°C;
Mobile Phase A: lOmM ammonium formate in water2acetonitrile 95:5, pH 9;
Mobile Phase B: acetonitrile;
ion: 210-400 nm;
nt: 0-0.40 min: 2% B, 0.40-4.85 min: 2% B to 98% B, 4.85-4.90 min: 98% B to 2% B,
4.90-5.00 min: hold at 2% B;
Flow rate: 0.6 mL/minute.
ation 1: Ally] amino-1H-pyrazole—4—carboxylate
NH2 0
Nil/(0”/N /
Step I: allyl 3-amino-4, 4, 4-trichlorocyanobutenoate 2
To a solution of KOAc (589.4 g, 6.006 mol) in isopropanol (3 L) was added allyl
cyanoacetate (429.4 g, 403.2 mL, 3.432 mol) and the reaction mixture was cooled to 5°C.
Trichloroacetonitrile (495.5 g, 3.432 mol) was added in 50 mL portions, maintaining
temperature below 15°C. The reaction mixture was then allowed to warm to 20°C and stirred
for 3 hr. Water (~4 L) was added to dissolve the inorganic materials and precipitate out the
desired product. The mixture was stirred for 20 minutes and the solid was isolated by
filtration under vacuum. This solid was filtered, washed with water (2 x 0.5 L) and dried in a
vacuum oven overnight at 40°C to afford allyl 3-amino-4,4,4-trichlorocyanobut-2—enoate 2
as an off—white powder (787 g, 85%).
Step 2: Allyl 3,5-diamino-IH-pyrazole—4-carb0xylate 3
To a suspension of allyl 3-amino—4,4,4-trichlorocyano—but-2—enoate 2 (619 g,
2.297 mol) and KOAc (676.3 g, 6.891 mol) in DMF (2.476 L) at 0°C was slowly added
ine hydrate (172.5 g, 167.6 mL, 3.446 mol) over 15 min. The reaction mixture was
then stirred at ambient ature for 2 hr., at which stage 1H NMR shows complete
consumption of the starting material. Reaction mixture was then heated overnight at 110°C
before being allowed to cool to ambient and stirred for another 48hr. The e was
filtered h a sintered glass funnel to remove the precipitated solid and the filtrate was
evaporated under reduced pressure to give a thick liquid. DCM (approx 2 L) was added, and
the mixture d again to remove additional solids that have precipitated. The filtrate was
d through a 1 kg silica gel plug (gradient of OH as an eluent), and the solvent
was removed to afford an orange solid which was suspended in acetonitrile and heated at
about 70°C until all the solid went into solution, at which point the solution was allowed to
cool to ambient temperature, then to 2°C. The precipitate that formed was isolated by
filtration under vacuum, washed with chilled MeCN (~50 mL) and dried to constant mass in a
vacuum oven to furnish the title compound as an ite powder (171.2 g, 41%).
Preparation 23: 1H-benzo[d][1,2,3]triazol—1-yl 2-amin0fluoropyrazolo[1,5-
a]pyrimidine-3—carboxylate
Step 1: allyl 2-amin0-6—flu0r0-pymzol0fl,5-ajpyrimidinecarb0xylate 4
To a suspension of allyl 3,5-diamino-lH-pyrazolecarboxylate 3 (42.72 g, 234.5
mmol) in DMSO (270.8 mL) / Water (270.8 mL), was added p-TsOH hydrate (46.72 g, 245.6
mmol) and 3-(diisopropylamino)-2—fluoro-prop-2—enal (described in Tetrahedron Letters,
33(3), 357-60; 1992) (38.69 g, 223.3 mmol). The reaction mixture was heated to 100°C for
3hr. during which time a solid slowly precipitated out of solution. The orange suspension was
allowed to cool down to RT overnight. The solid was d, washed with water and dried
under vacuum to give allyl 2-aminofluoro-pyrazolo[l,5-a]pyrimidine—3 -carboxylate 4 as a
sand solid (45.05 g, 85% yield).
Step 2: 2-amin0-6flu0r0-pyrazolofl,5-a]pyrimidinecarb0xylic acid 5
] To a suspension of allyl 2—amino—6-fluoro—pyrazolo[1,5-a]pyrimidine—3—carboxylate
4 (45 g, 190.5 mmol) in DCM (1.35 L) was added phenylsilane (41.23 g, 46.96 mL, 381.0
mmol), followed by Pd(PPh3)4 (8.805 g, 7.620 mmol). The reaction was stirred at room
ature for 2hr. 30min. The reaction e was filtered and the solid was washed with
DCM to give a light yellow solid ). This solid was triturated further in DCM (225 mL)
at RT for 45 min, then filtered and dried overnight under vacuum to provide 2—amino—6—
pyrazolo[1,5-a]pyrimidine—3-carboxylic acid 5 as a light yellow solid (3 7.77g, 100%
yield).
In an alternative method, 4-methylbenzenesulfinate (anhydrous, 1.2 eqv, 22.6g,
127mmol) was suspended in dry DMSO (20 vol, 500ml). The stirred e was warmed to
°C under a en atmosphere. Upon complete dissolution Pd(PPh3)4 (2 mol%, 2.4g, 2.1
mmol) was added. The mixture was stirred for 10 min at 25—30°C after which time a turbid
yellow solution was present. Allyl 2-aminofluoro-pyrazolo[ l ,5-a]pyrimidine
carboxylate 4 (25g, 105.8mmol) was added portionwise, maintaining the temperature at 25—
°C. Once addition was complete the cloudy solution was stirred until the reaction was
complete by HPLC (2—3 hrs). A heavy precipitate formed after 15 minutes post addition of
the substrate. The mixture became thicker as the reaction proceeded. The reaction mixture
was diluted with water (125 ml) and 2M HCl (66 ml) was added slowly, maintaining the
temperature at 25-3 0°C. The slurry was stirred for 30 minutes, then filtered. The filtration
was slow . The resulting solid was washed with water, then dried on the sinter. The
solid was slurried in DCM (8 vol) for 1hr. The solid was filtered (rapid filtration) and washed
with DCM. The solid was re-slurried in chloroform (8 vol) for 1 hr. The acid was filtered and
WO 87451
dried on the sinter. It was further dried in a vacuum oven at 50°C for 24 hrs. The t 5
was obtained as an off-white solid (18.6g, 85%); 1H NMR (500 MHz, DMSO-d6) 5 12.14
(1H, brs), 9.31 (1H, dd), 8.69 (1H, m), 6.47 (2H, brS); 19F NMR (500 MHz, DMSO—d6) 5 -
153.65; MS (ES+) 197.1.
Step 3: IH—benzo[d][1,2, 3]lriazol—I-yZ 2-amin0-6—flu0r0pyrazolo[I,5-a]pyrimidine
carboxylate 62
To a suspension of 2-amino-6—fluoro-pyrazolo[1,5-a]pyrimidinecarboxylic acid 5
(20 g, 102.0 mmol) in chloroform (300 mL) was added Et3N (11.35 g, 15.63 mL, 112.2
mmol). The suspension was stirred for ~ 5mins and then (benzotriazol-l-yloxydimethylamino-methylene
)—dimethy1—ammonium Boron Tetrafluoride was added (32.75 g,
102.0 mmol). The suspension was heated to 60°C for 1hr. before the thick suspension was
allowed to cool down to RT. The resulting suspension was filtered, washed with chloroform
(200 mL) and dried under vacuum overnight to afford the title nd 63 as a light yellow
powder (32.5 g, 88%).
Preparation 2b: (6-chlorobenzotriazol-I-yD-Z-amin0fluor0-pyrazololl,5-alpyrimidine-
3-carb0xylate 6b
[\‘I/ / OO,NIN:N
F CI
In a 2.5 L three-necked flask equipped with stirrer bar, condenser, nitrogen line and
Hanna temperature probe was charged 2-aminofluoro-pyrazolo[l,5-a]pyrimidine—3-
carboxylic acid 5 (60 g, 305.9 mmol), chloroform (900.0 mL) and triethylamine (32.44 g,
44.68 mL, 320.6 mmol). lorobenzotriazolyl)oxy-(dimethylamino)methylene]-
dimethyl-ammonium (Boron Tetrafluoride 1011 (1)) (87.00 g, 244.7 mmol) was added
portionwise over 5 mins (internal dropped from 22.7 to 215°C on complete on).
Mixture heated at 60°C (internal temp) for 2hr., still a cream suspension. Mixture cooled to
room temperature then solid collected by filtration, washed well with chloroform (until
te runs essentially colourless) and dried by suction to leave product 6b as a cream solid
(82.2g, 77% . 1H NMR (500 MHz, DMSO-d6) 5 9.55 (dd, 1H), 8.91 (d, 1H), 8.22 (dd,
1H), 8.09 (dd, 1H), 7.57 (dd, 1H) and 6.87 (s, 2H). MS (ES+) 348.1.
] In an alternative method, o—6-fluoropyrazolo[1,5-a]pyrimidine—3-carboxylic
acid 5 (30g, 153 mmol) was slurried in acetonitrile (540ml). Triethylamine (22.5ml,
153mmol) was added, followed by [(6-chlorobenzotriazol-1y1)oxy-
(dimethylamino)methylene]-dimethy1ammonium tetrafluoroborate (TCTU, 54.4g, 153mm01).
The mixture was stirred at room temperature for 2 hrs. The product was isolated by filtration—
the filter cake was washed with acetonitrile (2x60ml). The product was obtained as a brown
solid , 93%); 1H NMR (500 MHz, DMSO'd6) 6 9.55 (dd, 1H), 8.91 (d, 1H), 8.22 (dd,
1H), 8.09 (dd, 1H), 7.57 (dd, 1H) and 6.87 (s, 2H); 19F NMR (500 MHZ, DMSO—d6) 5 —
150.1; MS (ES+) 348.1.
Exam 1e 1: S nthesis of 2-amin0flu0r0-N- 5-flu0r0 l-meth midazol—S-
N B00201.15eq. N
\ O \
I l
HZN F NaHMDS 2.15 eq. 0 H F
7 a
.248
i) n-BuLi 2.5 eq.
TMEDA 2.5 eq. f4N—
THF NV N
°C to 20°C N |
71 i 1‘ 1°
.N /.
/ 2
o N F .2 HCI
ii) |21.5eq., I Pd(PPh3)4 0.1 / N’
-20 to -30°c. K3PO4 3 eq. N=/
71% 9 Reflux 11
48hn
ii) 4M HCI in dioxane
H2N F 2.HC]
NH2 / N/
0 NH2 0 N
=N /
/ , N=/ 11
N / 0’N N
\ / /
N \
\ /N \ /N /
Anisole
CI NJ
F F
95°C (intemal)
48hr
6b 50% 1-1
Step I: tert—butyl (5-fluor0pyridinyl)carbamate
In a 50 L jacketed vessel was added THF (2.5 L), 5-fluoropyridinamine 1 (500 g,
4.460 mol) then additional THF (5 L). To this d mixture was added a solution of tert-
butoxycarbonyl tert—butyl carbonate (1.119 kg, 5.129 mol) in THF (2.5 L), pumped in via a
vacuum line. The line was then rinsed with THF (1 L) in to the reaction vessel. The reaction
temperature was cooled to 0°C before NaHMDS (4.794 L of 2 M in THF, 9.589 mol) was
added in 12 x 400 mL portions (approx. 5°C exotherm after each addition, dosing ued
once internal cooled to 0°C). Addition was completed after 1 hr. The internal temperature
was raised to 5°C and stirred at this temperature for 1 hr. The reaction was carefully
quenched by slow addition of a saturated ammonium de aqueous solution (1 L)
(exothermic). The internal was raised to 10°C and onal saturated ammonium chloride
aqueous solution (3 L) was added. The internal was raised to 25°C and the reaction mixture
was extracted with EtOAc (1 x 5 L then 1 x 2.5 L). The ed organic layers were
washed with water (1 x 5.5 L then 1 x 3 L) then with brine (3 L).
The organic phase was concentrated in vacuo to a total volume of approx. 6 L, dried
), filtered through filter paper and concentrated in vacuo (on a rotary evaporator,
40°C bath temp) until product crystallised out (approx. 2 L of solvent remaining). Heptane
(2.5 L) was added and the mixture rotated on a rotary evaporator at 40°C. The solution was
concentrated in vacuo (on a rotary evaporator, 40°C bath temp) to remove more EtOAc until
the product crystallised out of solution. The mixture was then left to cool and stand at
ambient temperature overnight. The solid was collected by filtration h Whatman N9 1
filter paper, washed with heptane until filtrate ran essentially colourless. The solid was dried
for approx. 5 hr. to leave crop 1 of product as an off white solid, 382.51 g.
The mother liquor was concentrated slowly in vacuo (on a rotary evaporator, 40°C
bath temp) until a solid crystallised out. The mixture was left to stand at ambient overnight
and the solid collected by filtration, washed with heptane and dried by n to leave crop 2
of product 8 as an off white solid, 203.5 g. The process was repeated on the mother liquor to
give crop 3 as an off white solid, 178.7 g. Total yield of product, 764.71 g, 81%. 1H NMR
(500 MHz, DMSO-d6) 5 9.86 (s, 1H), 8.44 (s, 1H), 8.17 (d, J = 2.6 Hz, 1H), 7.83 (d, J = 11.6
Hz, 1H), 3.30 (s, 1H). MS (ES+) 213.0.
Step 2: ulyl (5-fluor0i0d0pyridinyZ)carbamale
7LOJLO |N\
N F
In a 50 L jacket vessel was added THF (2.5 L), tert—butyl N—(5-fluoro
pyridyl)carbamate 8 (400 g, 1.885 mol) in THF (2.5 L), onal THF (3 L) and N,N,N',N'-
tetramethylethane-1,2—diamine (547.6 g, 711.2 mL, 4.712 mol). The reaction mixture was
cooled to —28°C (internal temperature), then n—BuLi (1.885 L of 2.5 M in hexanes, 4.712 mol)
was added via canula at such a rate as to keep internal temperature below —20°C (i.e., over 2
hr.). On complete addition, the reaction mixture was stirred at between -30 and -20°C
(internal temperature) for a further 50 mins. Solid molecular iodine (765.5 g, 3.016 mol) was
slowly added in 12 roughly equal portions over 1 hr. (approx. 2/3°C delayed exotherm after
each portion added) keeping the internal temperature below -20°C. On complete addition of
iodine, the reaction mixture was stirred at -30°C (internal temperature) for a further 45 mins.
The reaction was then ed by the slow addition of a saturated ammonium
chloride aqueous solution (2 L) (exothermic). Water (2 L) was then added and the reaction
mixture warmed to 20°C (internal temperature) and left to stand overnight. To the reaction
mixture was added EtOAc (5 L) and stirring continued for 10 mins. The s phase was
removed then a ted sodium thiosulfate aqueous solution (2 L) was added to the organic
phase, stirred vigorously for 10 mins. Additional EtOAc (2.5 L) and water (2 L) was added
and ng continued for 10 mins. The aqueous phase was removed and the c phase
washed further with a saturated sodium thiosulfate aqueous solution (2 L) and water (1 x 2 L
then 1 x 2.5 L) and then brine (2 L). The organic phase was concentrated in vacuo (rotary
evaporator) to such a volume that the product started to crystallise out to give a thick
suspension. The mixture was left to stand at room ature overnight.
The solid was collected by ion, washed with minimal EtOAc (a few d
mL) then washed well with heptane, dried by suction for 3hr. to leave crop 1 of t 9 as a
white solid, 311.99 g. The mother liquor was concentrated in vacuo (rotary evaporator) to
dryness leaving a dark green solid. (approx 200 g) which was dissolved in EtOAc (750 mL)
by heating under reflux. Activated carbon (20 g) was then added and the mixture stirred
under reflux for 10 mins. The mixture was filtered through filter paper then concentrated
slowly on rotary evaporator until a thick sion . The resulting solid was collected
by filtration, washed with minimal EtOAc then heptane, dried by suction then in a vacuum
oven at 40°C for 2 hr., leaving crop 2 as a white solid, 103.9 g. The mother liquor was
concentrated again until a thick suspension . The solid was collected by filtration,
washed with heptane and dried by suction in vacuo (rotary evaporator) then in a vacuum oven
at 40°C for a few hours to leave product crop 3 as a white solid, 39.4 g. Total yield = 455.29
g, 71%. 1H NMR (500 MHz, DMSO-d6) 6 8.98 (s, 1H), 8.27 (dd, J = 1.2, 0.6 Hz, 2H), 1.47
(s, 9H). MS (ES+) 338.9.
Step 3: 5-flu0r0(I—methyl—IH—z'midazol—5—pryridinamine dihydrochloride
.2HCI
To a degassed (3 x vacuum/nitrogen cycles) mixture of tert—butyl N—(5-fluoro—4-
iodo-3—pyridyl)carbamate 9 (190 g, 561.9 mmol), 1—methyl(4,4,5,5-tetramethyl-1,3,2-
dioxaborolanyl)imidazole 10 (175.4 g, 842.8 mmol) and ium phosphate (226.0 g,
1.686 mol) in DME (2.28 L) was added Pd(PPh3)4 (64.93 g, 56.19 mmol). The reaction vessel
was again flushed with nitrogen via /nitrogen cycles (3x). The e was heated
under reflux and under a nitrogen atmosphere for 48 hr. The mixture was cooled to room
temperature then passed through a pad of celite, rinsing through with EtOAc until e
almost colourless (approx. 1.5 L). The filtrate was concentrated in vacuo to leave a sticky
brown solid, 339.7 g.
The crude product was dissolved in dioxane (950 mL) and methanol (431.1 mL) and
the solution cooled on ice bath (internal of 10°C), HCl (4 M in 1,4-dioxane) (842.8 mL of 4
M, 3.371 mol) was then added in 8 roughly equal ns over 20 mins. (approximately 3 to
4°C exotherm observed on each addition). On complete addition, the mixture was warmed to
40°C and stirred at this temperature for 3 hr., then left to cool to room temperature overnight
with stirring. The solid was collected by filtration, washed with 1,4-dioxane and dried under
vacuum for 1 hr. to leave product 11 as a sand/brown solid (107.9 g, 72% yield). 1H NMR
(500 MHz, Deuterium Oxide) 6 9.09 (s, 1H), 8.24 (s, 1H), 8.15 (br s, 1H), 7.91 — 7.90 (1H, br
s), 7.88 (m, 1H), 3.85 (s, 3H). MS (ES+) 193.1.
Step 4: 2—aminoflu0r0-N-(5—flu0r0(I-methyl—1H-imidazol—5-pryridin
pryrazolofl, 5-a]pyrimidirzecarb0xamide (Compound I-l)
~250 /N
/ \/
N H
$\_// F
A mixture of 5—fluoro-4—(3-methylimidazol-4—yl)pyridin—3-amine ochloride 11
(8.006 g, 30.2 mmol) and (6—chlorobenzotriazol—1—yl)2—amino—6—fluoro—pyrazolo[1,5—
a]pyrimidine—3-carboxylate 6b (10 g, 28.76 mmol) was suspended in anisole (100 mL). To
this suspension was added DIPEA (8.177 g, 11.02 mL, 63.27 mmol) and the mixture was
heated at 95°C (internal ature) for 44 hr. then allowed to cool to room temperature
overnight. The solid was collected by filtration, washed with minimal anisole (approx 20
mL), dried under vacuum for 1 hr., then the solid dried in a vacuum oven at 45°C (internal
ature) for 2 hr. to leave product as a light yellow solid, 7.8 g. This solid was ded
in water (78 mL) and MeCN (117 mL) and TFA (2.4 g, 1.62 mL, 1 eq.) was added. The
reaction mixture was stirred at room temperature for 10 mins. then filtered through filter
paper, washed through with small amount of water. The filtrate was basified to pH = 8 by
addition of 2 M sodium carbonate whilst stirring. The solid was collected by filtration,
washed with water then dried under vacuum for 1hr. The solid was then dried in vacuum
oven at 45°C (internal temperature) overnight leaving t I-1 as a pale yellow solid, 5.29
g. 1H NMR (500 MHz, DMSO—d6) 6 9.68 (s, 2H), 9.42 (dd, J = 4.8, 2.5 Hz, 1H), 8.46 (s, 1H),
8.31 (d, J = 2.5 Hz, 1H), 8.07 (s, 1H), 7.25 (d, J = 1.0 Hz, 1H), 6.71 (s, 2H), 3.46 (s, 3H). MS
(ES+) 371.0.
Compound Analytical Data
H NMR (500 MHz, DMSO—d6) 5 9.68 (s, 2H), 09.42 (dd, J
= 4.8, 2.5 Hz, 1H), 8.46 (s, 1H), 8.31 (d, J = 2.5 Hz, 1H),
I-l 371.0 1.80
8.07 (s, 1H), 7.25 (d, J = 1.0 Hz, 1H), 6.71(s, 2H), 3.46 (s,
Solid Forms of Compound I-1
Compound I-1 has been prepared in various solid forms, including anhydrous forms.
The solid forms of the present invention are useful in the manufacture of ments for the
treatment of . One embodiment provides use of a solid form described herein for
treating cancer. In some embodiments, the cancer is triple negative breast cancer, pancreatic
cancer, small cell lung cancer, colorectal cancer, ovarian cancer, or non-small cell lung
cancer. r embodiment provides a pharmaceutical composition comprising a solid form
described herein and a pharmaceutically acceptable carrier.
Applicants describe herein a novel solid form of Compound I-l. The name and
stoichiometry of the solid form is provided in Table 2 below:
Table 2
Com-oundl-1 anh drous free base
Compound I-1 e
Example 4 Compound I-1 tartaric acid
e 2: Compound I-1 [anhydrous free base]
Compound I-l anhydrous free base can be ed according to the methods
described in Example 1, Step 4.
XRPD of Compound I-l [anhydrous free base]
The XRPD pattern of compound I-1 anhydrous free base was recorded at room
ature in reflection mode using a PANaZyticaZ diffractometer equipped with an
Empyrean tube source and a P1Xcel 1D detector (PANaZyticaZ, The Netherlands). The X—
ray generator was operating at a voltage of 45 kV and a current of 40 mA. The powder
sample was placed in a silicon . The data were recorded over the range of 3°—39° 2
theta with a step size of 0.0130 and a dwell time of 0.5s per step. Figure 1a shows the X-ray
powder diffractogram of the sample which is characteristic of crystalline drug substance.
Representative XRPD peaks from Compound I-1 anhydrous free base:
8.704
9.8727
12.7565 .
4* 15.4224 31.96
* 5 29.04
6 17.4518 6.14
.5734
21.2755
9 5.54
23.0565
24.3907
27.8453
28.9558
.1162
31.7775
32.2508
33.04
36.5878
37.6243
Thermo Analysis of Compound I-1 (anhydrous free base)
] A thermogravimetric analysis of compound I-l anhydrous free base was performed
to determine the percent weight loss as a function of ature using the Discovery TGA
(TA Instruments Trios). A sample (2.84mg) was added to a pre—tared aluminum pan and
heated from ambient temperature to 400°C at 10°C/min. The TGA results seen in Figure 2a
show very little observed weight loss prior to melting or thermal ation. From ambient
temperature to 261°C, the weight loss is 0.60%. The onset temperature of
melting/degradation is 299°C.
Differential Scanning Calorimetm of Compound I-l [anhydrous free base]
Differential scanning calorimetry of compound I-1 anhydrous free base was
measured using the TA Instrument DSC Q2000. A sample (1.71 mg) was weighed in a
pinholed hermetic aluminum pan and heated from ambient temperature to 400°C at
°C/min. The DSC results seen in Figure 3a show a single melting endotherm at 302°C
(onset).
Example 3: Compound I-1 [hydrate]
] Compound I-1 anhydrous free base, ed according to the methods described in
Example 1, Step 4, was slurried in water or organic solvent water mixtures to produce
Compound I-1 hydrate.
XRPD of Compound I-l [hydrate]
The XRPD pattern of Compound I-1 hydrate was recorded at room temperature in
reflection mode using a PANaZytical diffractometer equipped with an Empyrean tube source
and a PIXcel lD detector (PANaZyticaZ, The Netherlands). The X—ray generator was
operating at a voltage of 45 kV and a current of 40 mA. The powder sample was placed in a
n holder. The data were over the range of 3°—39° 2 theta with a step size of 0.013° and a
dwell time of 0.5s per step. Figure lb shows the X-ray powder diffractogram of the sample
which is characteristic of crystalline drug substance.
Representative XRPD peaks from nd I-1 hydrate:
Thermo Analysis of Compound I-1 [hydrate]
A thermal gravimetric analysis (TGA) of Compound I-1 hydrate was performed to
determine the percent weight loss as a function of ature using the Discovery TGA (TA
Instruments Trios). A sample (4.74 mg) was added to a pre-tared aluminum pan and heated
from t ature to 400°C at 10°C/min. The TGA results seen in Figure 2b show a
large weight loss of 12.6% below 100°C. This weight loss corresponds to approximately 3
molar equivalents of water. The subsequent weight loss above 250°C is a result of melting
and degradation.
Differential Scanning Calorimetfl of nd I-1 [hydrate]
Differential scanning calorimetry (DSC) of Compound I-1 hydrate was measured
using the TA Instrument DSC Q2000. A sample (2.78 mg) was d in a pinholed
aluminum hermetic pan and heated from ambient temperature to 370°C at 10°C/min. The
DSC results seen in Figure 3b show a broad desolvation endotherm below 100°C followed by
a exotherm recrystallization to Compound I-1 anhydrous free base between 100-150°C.The
erm peak between 300-305°C indicates the melting of nd I-1 anhydrous free
base.
e 4: Compound I-1 ric acid]
Compound I-l anhydrous free base, prepared according to the methods described in
Example 1, Step 4, was slurried with tartaric acid and ethanol to produce Compound I-1
tartaric acid.
XRPD of Compound I-l [tartaric acid]
The XRPD n of Compound I-1 tartaric acid form was recorded at room
temperature in reflection mode using a PANaZytz‘caZ diffractometer equipped with an
Empyrean tube source and a P1Xcel 1D detector (PANalytz'caZ, The Netherlands). The X-ray
generator was operating at a voltage of 45 kV and a current of 40 mA. The powder sample
was placed in a silicon holder. The data were over the range of 4.50-39O 2 theta with a step
size of 0.013° and a dwell time of 299.6s per step. Figure 1c shows the X—ray powder
diffractogram of the sample which is characteristic of crystalline drug substance.
entative XRPD peaks from Compound I-1 tartaric acid:
Thermo is of Compound I-1 [tartaric acid]
A thermal gravimetric is (TGA) of Compound I-1 tartaric acid form was
performed to determine the percent weight loss as a function of temperature using the
Discovery TGA (TA Instruments . A sample (3.35 mg) was added to a pre-tared
aluminum pan and heated from ambient temperature to 330°C at 10°C/min. The TGA results
seen in Figure 20 show three step weight losses of 12.4%, 12.6%, and 8.5% between 150-
330°C.
Differential Scanning metfl of Compound I-l [tartaric acid]
] Differential scanning calorimetry (DSC) of Compound I-1 tartaric acid was
measured using the TA Instrument DSC Q2000. A sample (1.08 mg) was weighed in a
pinholed aluminum hermetic pan and heated from ambient temperature to 350°C at
°C/min. The DSC results seen in Figure 3c show the first 2 exotherm peaks between 200—
275°C corresponding to the first 2 step weight losses in TGA, and the last endoterm peak
above 275°C corresponding to the last step weight loss in TGA.
Example 5: Cellular ATR Inhibition Assay:
] Compounds can be ed for their ability to inhibit intracellular ATR using an
immunofluorescence microscopy assay to detect phosphorylation of the ATR substrate
histone H2AX in hydroxyurea treated cells. HT29 cells are plated at 14,000 cells per well in
96-well black imaging plates (BD 353219) in McCoy’s 5A media (Sigma M8403)
supplemented with 10% foetal bovine serum (JRH Biosciences ,
Penicillin/Streptomycin on diluted 1:100 (Sigma P7539), and 2mM L-glumtamine
(Sigma G7513), and allowed to adhere overnight at 37°C in 5% C02. Compounds are then
added to the cell media from a final concentration of 25uM in 3-fold serial dilutions and the
cells are incubated at 37°C in 5% C02. After 15min, hydroxyurea (Sigma H8627) is added to
a final concentration of 2mM.
After 45min of treatment with hydroxyurea, the cells are washed in PBS, fixed for
10min in 4% formaldehyde diluted in PBS (Polysciences Inc 18814), washed in 0.2% Tween-
in PBS (wash buffer), and permeabilised for 10min in 0.5% Triton X—100 in PBS, all at
room ature. The cells are then washed once in wash buffer and blocked for 30min at
room temperature in 10% goat serum (Sigma G9023) diluted in wash buffer (block buffer).
To detect H2AX phosphorylation levels, the cells are then ted for 1h at room
temperature in primary antibody (mouse monoclonal anti-phosphorylated histone H2AX
Ser139 antibody; Upstate 05-636) d 1:250 in block buffer. The cells are then washed
five times in wash buffer before incubation for 1h at room temperature in the dark in a
mixture of secondary antibody (goat anti-mouse Alexa Fluor 488 conjugated antibody;
Invitrogen A11029) and t stain (Invitrogen H3570); diluted 1:500 and 115000,
tively, in wash buffer. The cells are then washed five times in wash buffer and finally
100ul PBS is added to each well before imaging.
Cells are imaged for Alexa Fluor 488 and Hoechst intensity using the BD Pathway
855 Bioimager and Attovision software (BD Biosciences, n 1.6/855) to quantify
phosphorylated H2AX Ser139 and DNA staining, respectively. The percentage of
orylated H2AX—positive nuclei in a e of 9 images at 20x magnification is then
calculated for each well using BD Image Data Explorer software (BD Biosciences Version
2.2.15). Phosphorylated H2AX-positive nuclei are defined as Hoechst-positive regions of
interest containing Alexa Fluor 488 ity at 1.75—fold the average Alexa Fluor 488
intensity in cells not treated with hydroxyurea. The percentage of H2AX positive nuclei is
finally plotted against concentration for each compound and IC50s for intracellular ATR
inhibition are determined using Prism software (GraphPad Prism version 3.00x for
Macintosh, GraphPad Software, San Diego California, USA).
] The compounds described herein can also be tested according to other methods
known in the art (@ Sarkaria et a1, “Inhibition ofATM and ATR Kinase Activities by the
Radiosensitizing Agent, Caffeine: Cancer Research 59: 4375—5382 (1999); Hickson et al,
“Identification and Characterization of a Novel and Specific Inhibitor of the Ataxia-
Telangiectasia Mutated Kinase ATM” Cancer Research 64: 9152—9159 (2004); Kim et a1,
“Substrate Specificities and Identification of Putative Substrates of ATM Kinase Family
Members” The Journal ofBiological Chemistry, 274(53): 37538-37543 (1999); and Chiang
et a1, mination of the catalytic activities of mTOR and other s of the
phosphoinositidekinase-re1ated kinase family” Methods Mol. Biol. 281: 125-41 (2004)).
Example 6: ATR Inhibition Assay:
] Compounds can be screened for their ability to inhibit ATR kinase using a
radioactive—phosphate incorporation assay. Assays are d out in a mixture of 50mM
Tris/HCl (pH 7.5), 10mM MgClz and 1mM DTT. Final substrate trations are lOuM
[y-33P]ATP (3mCi 33P ATP/mmol ATP, Perkin Elmer) and 800 [1M target peptide
(ASELPASQPQPFSAKKK).
] Assays are carried out at 25°C in the presence of 5 nM full-length ATR. An assay
stock buffer solution is prepared containing all of the reagents listed above, with the
ion of ATP and the test compound of interest. 13.5 uL of the stock solution is placed
in a 96 well plate followed by addition of 2 uL of DMSO stock containing serial dilutions of
the test compound (typically starting from a final concentration of 15 “M with 3-fold serial
dilutions) in duplicate (final DMSO concentration 7%). The plate is pre—incubated for 10
minutes at 25°C and the on initiated by on of 15 uL [y-33P]ATP (final
concentration 10 uM).
The reaction is stopped after 24 hours by the addition of 30uL 0.1M phosphoric acid
containing 2mM ATP. A creen phosphocellulose filter 96-well plate (Millipore, Cat
no. MAPHNOB50) is pretreated with 100uL 0.2M phosphoric acid prior to the addition of
45 [1L of the stopped assay mixture. The plate is washed with 5 x 200[1L 0.2M phosphoric
acid. After drying, 100 uL Optiphase ‘SuperMix’ liquid scintillation cocktail (Perkin Elmer)
is added to the well prior to scintillation counting (1450 Microbeta Liquid Scintillation
Counter, Wallac).
After removing mean background values for all of the data , Ki(app) data are
calculated from non—linear regression analysis of the initial rate data using the Prism software
package (GraphPad Prism version 3.0cx for Macintosh, GraphPad Software, San Diego
California, USA).
In general, the nds of the present invention are ive for inhibiting ATR.
Compound I-1 inhibits ATR at Ki values below 1 pM.
Example 7: Cisplatin Sensitization Assay
Compounds can be screened for their ability to sensitize HCT116 colorectal cancer
cells to Cisplatin using a 96h cell Viability (MTS) assay. HCT116 cells, which possess a
defect in ATM signaling to Cisplatin (fl, Kim et al.; Oncogene 4 ; w,
Takemura et al.; JBC 28130814 (2006)) are plated at 470 cells per well in 96—well
polystyrene plates (Costar 3596) in 150p] of McCoy’s 5A media (Sigma M8403)
supplemented with 10% foetal bovine serum (JRH Biosciences 12003),
llin/Streptomycin solution diluted 1:100 (Sigma P7539), and 2mM L-glumtamine
(Sigma G7513), and allowed to adhere overnight at 37°C in 5% C02. Compounds and
Cisplatin are then both added simultaneously to the cell media in 2-fold serial dilutions from
a top final concentration of IOuM as a full matrix of concentrations in a final cell volume of
200111, and the cells are then incubated at 37°C in 5% C02. After 96h, 40ul of MTS reagent
(Promega G358a) is added to each well and the cells are incubated for 1h at 37°C in 5% CO2.
Finally, absorbance is measured at 490nm using a SpectraMax Plus 384 reader (Molecular
Devices) and the concentration of compound ed to reduce the IC50 of Cisplatin alone
by at least 3—fold (to 1 decimal place) can be ed.
In general, the compounds of the present invention are effective for izing
cancer cells to Cisplatin. nd I-1 have Cisplatin sensitization values of < 0.2 uM.
Example 8: Single Agent HCT116 Activity
nds can be ed for single agent activity against HCT116 colorectal
cancer cells using a 96h cell viability (MTS) assay. HCT116 are plated at 470 cells per well
in 96—well polystyrene plates (Costar 3596) in 150g] of McCoy’s 5A media (Sigma M8403)
supplemented with 10% foetal bovine serum (JRH ences 12003), Penicillin/
Streptomycin solution diluted 1:100 (Sigma P7539), and 2mM tamine (Sigma
G7513), and allowed to adhere overnight at 37°C in 5% CO2. Compounds are then added to
the cell media in 2—fold serial ons from a top final concentration of lOuM as a full
matrix of concentrations in a final cell volume of 200111, and the cells are then incubated at
37°C in 5% CO2. After 96h, 40ul of MTS reagent (Promega G35 8a) is added to each well
and the cells are incubated for 1h at 37°C in 5% CO2. Finally, absorbance is measured at
490nm using a SpectraMax Plus 384 reader (Molecular Devices) and IC50 values can be
calculated.
Example 9: ATR-complex Inhibition Assay
Compounds were screened for their ability to inhibit ATR kinase, in the presence of
partner proteins ATRIP, CLK2 and TopBPl, using a radioactive-phosphate incorporation
assay. Assays were carried out in a mixture of 50 mM Tris/HCl (pH 7.5), 10 mM MgCl2 and
1 mM DTT. Final ate concentrations were 10 “M ]ATP (3.5 “Ci 33P ATP/nmol
ATP, Perkin Elmer, Massachusetts, USA) and 800 “M target peptide
(ASELPASQPQPFSAKKK, Isca Biochemicals, Cambridgeshire, UK).
Assays were carried out at 25°C in the presence of 4 nM full-length ATR, 40 nM
full-length ATRIP, 40 nM full-length CLK2 and 600 nM TopBPl(A89l-Sl 105). An enzyme
stock buffer solution was prepared containing all of the reagents listed above, with the
exception of target peptide, ATP and the test nd of interest. This enzyme stock was
pre—incubated for 30 minutes at 25°C. 8.5 ML of the enzyme stock solution was placed in a
l plate followed by addition of 5 p1 of target peptide and 2 “L ofDMSO stock
containing serial dilutions of the test compound (typically starting from a final concentration
of 1.5 uM with 2.5-fold serial dilutions) in duplicate (final DMSO concentration 7%). The
plate was pre-incubated for 10 minutes at 25°C and the on initiated by addition of 15 [JL
[g—33P]ATP (final concentration 10 uM).
The reaction was d after 20 hours by the addition of 30 uL 0.3 M phosphoric
acid containing 2 mM ATP. A phosphocellulose filter 96-well plate (Multiscreen HTS
MAPHNOBSO, Merck—Millipore, Massachusetts, USA) was pretreated with 100 [1L 0.1 M
phosphoric acid prior to the on of 45 [LL of the stopped assay mixture. The plate was
washed with 5 x 200 ML 0.1 M oric acid. After drying, 50 [LL ase ‘SuperMix’
liquid scintillation cocktail n Elmer, Massachusetts, USA) was added to the well prior
to scintillation counting (Wallac 1450 eta Liquid Scintillation Counter, Perkin Elmer,
Massachusetts, USA).
After removing mean background values for all of the data points, Ki(app) data were
calculated from non—linear regression analysis of the initial rate data using the Prism software
package (GraphPad Prism version 6.00 for Macintosh, GraphPad Software Inc., San Diego,
USA).
While we have described a number of ments of this invention, it is apparent
that our basic examples may be altered to provide other embodiments that utilize the
compounds, methods, and processes of this invention. Therefore, it will be appreciated that
the scope of this invention is to be defined by the appended claims rather than by the specific
embodiments that have been represented by way of example herein.
Claims (20)
1. A process for preparing a compound of formula I-1: O N N H F F N comprising a step of reacting the compound of formula 6b: with a compound of formula 11: under suitable conditions to form an amide bond.
2. The process of claim 1, further comprising a step of ing the nd of formula 11: comprising reacting a compound of formula 9: wherein X is a halogen; with a compound of a 10: under suitable metal zed coupling conditions to form an adduct containing a protected amine group; and subjecting the resulting adduct to suitable deprotection conditions.
3. The process of claim 2, further comprising a step of preparing a compound of formula wherein X is a halogen; comprising ng the compound of formula 8: under suitable halogenation conditions.
4. The process of claim 3, further comprising a step of preparing a compound of formula comprising reacting a compound of formula 7: under suitable conditions to generate a protected amine group.
5. The process of claim 1, wherein the step of reacting a compound of formula 6b with a compound of formula 11 occurs in the presence of a solvent and an organic base.
6. The process of claim 5, wherein the solvent is selected from NMP, DMF or anisole.
7. The process of claim 5, wherein the organic base is an aliphatic amine.
8. The process of claim 7, wherein the aliphatic amine is selected from triethylamine or DIPEA.
9. The process of claim 2, wherein suitable metal catalyzed cross-coupling conditions comprise a metal catalyst, a suitable solvent, and a suitable base.
10. The s of claim 9, wherein the metal st is a palladium catalyst.
11. The s of claim 10, wherein palladium catalyst is selected from PdCl2(PPh3)2, Pd(Ph3)4, and PdCl2(dppf).
12. The process of claim 9, wherein the le base comprises one or more of potassium phosphate, K2CO3, tBuOK and .
13. The process of claim 9, wherein the suitable solvent comprises one or more of DME, tetrahydrofuran, toluene, and ethanol.
14. The process of claim 1, further sing a step of treating nd I-1 under suitable conditions to provide crystalline Compound I-1 ous free base.
15. The process of claim 14, wherein the crystalline Compound I-1 anhydrous free base is characterized by having one or more peaks expressed in 2-theta ± 0.2 at about 9.9, 12.8, 15.4, 17.0, 23.1, 27.8, 29.0, and 30.1 degrees in an X-Ray powder diffraction pattern obtained using Cu K alpha radiation.
16. The process of claim 14, further comprising a step of treating crystalline nd I- 1 anhydrous free base under suitable conditions to provide crystalline Compound I-1 hydrate.
17. The process of claim 16, n the crystalline Compound I-1 hydrate is characterized by having one or more peaks expressed in 2-theta ± 0.2 at about 27.5, 20.6, and 9.7 degrees in an X-Ray powder diffraction pattern ed using Cu K alpha radiation.
18. The process of claim 14, further comprising a step of treating crystalline Compound I- 1 anhydrous free base under le conditions to provide crystalline Compound I-1 tartaric acid.
19. The process of claim 18, wherein the crystalline Compound I-1 tartaric acid is characterized by having one or more peaks expressed in 2-theta ± 0.2 at about 7.1, 18.3, and 13.2 degrees in an X-Ray powder diffraction pattern obtained using Cu K alpha radiation.
20. The s of claim 1, substantially as herein described with reference to any one of the Examples and/or
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NZ764150A NZ764150B2 (en) | 2015-05-28 | Radiolabelled derivatives of a 2-amino-6-fluoro-n-[5-fluoro-pyridin-3-yl]- pyrazolo[1,5-a]pyrimidin-3-carboxamide compound useful as ATR kinase inhibitor, the preparation of said compound and different solid forms thereof | |
NZ764151A NZ764151B2 (en) | 2015-05-28 | Radiolabelled derivatives of a 2-amino-6-fluoro-n-[5-fluoro-pyridin-3-yl]- pyrazolo[1,5-a]pyrimidin-3-carboxamide compound useful as atr kinase inhibitor, the preparation of said compound and different solid forms thereof |
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