Title of Invention

A 6-AMINO-2-PHENYL SUBSTITUTED PYRAZINE COMPOUND.

Abstract A 6-amino-2-phenyl substituted pyrazine compound of the general formula: or pharmaceutically acceptable salts, hydrates, solvates, crystal forms of diastereomers thereof is described.
Full Text A 6-AMIN0-2-PHENYL SUBSTITUTED PYRAZINE COMPOUNDS
FIELD OF THE INVENTION
The present invention relates to the field of inhibitors of protein kinases.
BACKGROUND OF THE INVENTION
Protein kinases are a family of enzymes that catalyse the phosphorylation of
specific residues in proteins. In general protein kinases fall into several groups; those
which preferentially phosphorylate serine and/or threonine residues, those which
preferentially phosphorylate tyrosine residues and those which phosphorylate both
tyrosine and Ser/Thr residues. Protein kinases are therefore key elements in signal
transduction pathways responsible for transducing extracellular signals/ including the
action of cytokines on their receptors, to the nuclei, triggering various biological
events. The many roles of protein kinases in normal cell physiology include cell cycle
control and cell growth, differentiation, apoptosis, cell mobility and mitogenesis.
Protein kinases include members of the Protein Tyrosine Kinase family (PTKs),
which in turn can be divided into the cytoplasmic PTKs (CTKs) and the receptor PTKs
(RTKs). The cytoplasmic PTKs include the SRC family, (including: BLK; FGR; FYN;
HCK; LCK; LYN; SRC;YES and YRK); the BRK Family (including: BRK; FRK, SAD;
and SRM); the CSK family (including: CSK and CTK); the BTK family, (including BTK;
ITK; TEC MKK2 and TXK), the Janus kinase family, (including: JAK1, JAK2, JAK3 and
Tyk2), the FAK family (including, FAK and PYK2); the Fes family (including FES and
FER), the ZAP70 family (including ZAP70 and SYK); the ACK family (including ACK1
and ACK2); and the Abl family (including ABL and ARG). The RTK family includes
the EGF-Receptor family (including, EGFR,HER2, HER3 and HER4); the Insulin
Receptor family (including INS-R and IGF1-R); the FDGF-Receptor family (including
PDGFRa, PDGFRß, CSFIR, KIT, FLK2); the VEGF-Receptor family (including; FLT1,
FLK1 and FLT4);the FGF-Receptor family (including FGFR1, FGFR2, FGFR3 and
FGFR4); me CCK4 family (including CCK4); the MET family (including MET and
RON); the TRK family (including TRKA, TRKB, and TRKC ); the AXL family .
(including AXL, MER, and SKY); the ITE/TEK family (including TIE and TIE2/TEK);
the EPH family (including EPHA1, EPHA2, EPHA3, EPHA4, EPHA5, EPHA6,
EPHA7, EPHA8, EPHB1, EPHB2, EPHB3, EPHB4, EPHB5, EPHB6); the RYK family
(including RYK); the MCK family (including MCK and TYRO10); the EOS family
(including ROS); the RET family (including RET); the LTK family (including LTK and
ALK); the ROR family (including ROR1 and ROR2); The Musk family (including
Musk); the LMR family including LMR3, LMR2 and LMR3); and the SuKTKlO6 family
(including SuRTK106).
Similarly, the serine /threonine specific kinases comprise a number of distinct
sub-families, including; the extracellular signal regulated kinases, (p42/ERK2 and
p44/ERKI); c-Jun NH2-terminal kinase (JNK); cAMP-responsive element-binding
protein kinases (CREBK); the cydin dependent kinases (CDKs); cAMP-dependent
lanase (CAPK); mitogen-acaVatad protein kinase-activated protein kinase (MAPK and
its relatives); stress-activated protein kinase p38/SAPK2; mitogen-and stress-activated
kinase (MSK); protein kinases, PKA, PKB and PKC inter alia.
Additionally, the genomes of a number of pathogenic organisms possess genes
encoding protein kinases. For example, the malarial parasite Flasmodiuzn faldparum
and viruses such as HPV and Hepatitis viruses appear to bear kinase related genes.
Inappropriately high protein kinase activity has been implicated in many
diseases resulting from abnormal cellular function. This might arise either directly or
indirectly, for example by failure of the proper control mechanisms for the kinase,
related for example to mutation, over-expression or inappropriate activation of the
enzyme; or by over- or under-production of cytokines or growth factors also .
participating in the transduction of signals upstream or downstream of the kinase. In
all of these instances, selective inhibition of the action of the kinase might be expected
to have a beneficial effect Diseases where aberrant kinase activity has been implicated
include: diabetes; restenosis; atherosclerosis; fibrosis of the liver and kidney; ocular
diseases; myelo- and lymphoproliferative disorders; cancer such as prostate cancer,
colon cancer, breast cancer, head and neck cancer, leukemia and lymphoma; and,
auto-immune diseases such as Atopic Dermatitis, Asthma, rheumatoid arthritis,.
Crohn's disease, psoriasis, Crouzon syndrome, achondroplasia, and thanatophoric
dysplasia.
SUMMARY OF THE INVENTION
The present inventors have found that a group of compounds based upon a
disubstituted pvrazine scaffold are inhibitors of protein kinases.
This invention is therefore directed to compounds that potentially modulate
Protein Kinasc signal transduciion by affecting the enzymatic activity of RTKs, CTKs
and/orSTKs, thereby interfering with the signals transduced by such proteins. More
particularly, the present invention is directed to compounds which modulate RTK,
CTK and/ or 5TK mediated signal transduction pathways as a therapeutic approach to
cure many kinds of tumor.
Accordingly/ in a first aspect the present invention consists in a compound of
the general f onnula

or pharmaceutically acceptable salts, hydrates, solvates, crystal forms or diastereomers
thereof, wherein:
R1 is H,C1-4 alkyl
Q is a bond, or C1-4 alkyl
A is aryl, hetaryl optionally substituted with 0-3 substituents independently
chosen from halogen, C1-4 alkyl CH2F, CHF, CF3, CN, aryl hetaryl, OCF, OC1-4
alkyl, OC2-5alkyINR4R5, Oaryl, Ohetaryl, CO2R4, CONR4R5, nitro, NR4R5, C1-4
aIkyl NR4R5, NR6C1-4alkylNR4R5, NRCOR5, NH6CONR4R5, NMSO2RS; and
R4, R5 are each independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1-4 alkyl
cyclohetalkyl, aryl, hetaryl C1-4alkyl aryl, C1-4 alkyl hetaryl, or may be joined to
form an optionally substituted 3-8 membered ring optionally containing an
atom selected from O, S, NR7; and R6 is selected from H, C1-4 alky]; and R7 is
selected from H, C1-4 alkyl, aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl.
R2 is 0-2 substituents independently selected from halogen, C1-4alkyl, OH, OC1-4
alkyl, CH2F, CHF2, CF2, OCF2, CN, C1-4alkylNR8R9, OC1-4alkylNR8R9, CO2RS,
CONR8R9, NR8R9, NRBCOR9, NR10CONR8R9, NR8SO2R9; and R8, R9 are
each independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1-4 alkyl cyclohetalkyl,
aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl, or may be joined to form an
optionally substituted 3-8 membered ring optionally containing an atom
selected from O, S, NR11; and R10 is selected from H, C1-4 alkyl, aryl or hetaryl;
and R11 is selected from H, C1-4 alkyl, aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl
hetaryl.
Y is halogen, OH, NR12R13, NR12COR13, NR12CONR13, N12SO2R13; and
R12, and R13 are each independently H, CH2F, CHF2, CF2, CN, C1-4
alkyl, C1-4 alkyl cydoalkyl, C1-4 alkyl cyclohetalkyl, or may be joined to form an optionally
substituted 3-6 membered ring optionally containing an atom selected from O,
S, NR14 and R14 is selected from H, C1-4 alkyl.
n=0-4
W is selected from K, C1-4alkyl, C1-4alkenyl; where C1-4alkyl or C1-4alkenyl may
be optionally substituted with C1-4alkyl, OH, OC1-4alkyl, NR1SR16; and R15,
and R16 are each independently H, C1-4alkyl, C1-4 alkyl cycloalkyt C1-4 alkyi
cyclohetalkyl, or may be joined to form an optionally substituted 3-8 membered
ring optionally containing an atom selected from O, S, NR17 and R17 is selected
from H,C1-4 alkyl;
wherein when Y is OH or NHCOGH3 then R2 is 1-2 subsiituents and wherein, when Y
is NH2 and R2 b absent then Y is in the para position.
In a second aspect the present invention consists in a composition comprising a
carrier and at least one compound of the first aspect of the invention.
In a third aspect the present invention consists in a method of treating a protein
kinase-associated disease state, the method comprising administering a
"therapeutically effective amount of at least one compound of the first aspect of the
invention or a therapeutically effective amount of a composition of the second aspect
of the invention.
BRIEF DESCRIPTION OF THE ACCOMPANYING DRAWINGS
Figure 1: Dose-response curves for R and S enantiomers of representative
2-(a-methyl benzylamino)-pyrazine possessing inhibitory activity in Tel-Jak2
transformed cell line
Figure 2: Kinase Inhibition Activity Of Representative 2-(a-methyl
benzylamino)-pyrazines. Kinase assays were performed as described in the methods
section. Figute 2A, CYC10124 (exemplified as Chemistry 141 in tabJe 1) exhibits
potent inhibitory activity against the c-KIT, TIE2 and ABL protein kinases. Figure 2B,
CYC10268 (exemplified as Chemistry 268 in table 1) exhibits potent inhibitory
activity against c-FMS. Figure 2C, CYC10119 (exemplified as Chemistry 136 in table
1) exhibits potent inhibitory activity against JAK2 and c-KTT.
DETAILED DESCRIPTION
This invention is directed to compounds- that potentially modulate Protein
Kinase signal transduction by affecting the enzymatic activity of RTKs, CTK8 and/ or
STKs, thereby interfering with the signals transduced by such proteins. More
particularly/ the present invention is directed to compounds which modulate RTK,
CTK and/or STK mediated signal transduction pathways as a therapeutic approach to
cure many kinds of tumor.
Accordingly, in a first aspect the present invention consists in a compound of
the general formula

or pharmaceutically acceptable salts, hydrates, solvates, crystal forms or diastereomers
thereof, wherein:
R1 is H, C1-4 alkyl
Q is a bond, or C1-4 alkyl
A is aryl, hetaryl optionally substituted with 0-3 substituents independently
chosen from halogen, C1-4 alkyl, CH2F, CHF2, CF2, CN, aryl, hetaryl, OCFj, OC1-4
alkyl, OC2-5alkylNR4R5, Oaryl, Ohetaryl, CO2R4, CONR4R5, nitro, NR4R5, C1-4
alkylNR4R5, NR6C1-4alkylNR4R5, NR4COR5, NR6CONR4R5, MR4SO2R5; and
R4, R5 are each independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1-4 alkyl
cydohetalkyl aryl, hetaryl C1-4alkyl aryl, C1-4 alky] hetaryl, or may be joined to
form an optionally substituted 3-8 membered ring optionally containing an
atom selected from O, S, NR7; and R6 is selected from H, C1-4 alkyl; arid R7 is
selected from H, C1-4 alkyl, aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl.
R2 is 0-2 substituents independently selected from halogen, C1-4alkyl, PH, OC1-4
alkyl, CH2F, CHF2, CF2, OOF3, CN, C1-4alkylNR8R9, OC1-4alkylNR8R9l CO2RS,
CONRSR9, NR8R9, NR8COR9, NR10CONR8R9, NR8SO2R9; and RS, R9 are
each independently H, C1-4 alkyl, C1-4 alkyl cydoalkyl C1-4 alkyl cyclohetalkyL
aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl or may be joined to form an
optionally substituted 3-8 membered ring optionally containing an atom
selected from O, S, NR11; and R10 is selected from H, C1-4 alkyl, aryl or hetaryl;
and Rll is selected from H, C1-4 alkyl/ aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl
hetaryl.
Y is halogen, OH, NR12R13, NR12CORI3, NR12CONR13, Nl2SO2R13; and
R12, and R13 are each independently H, CH1F, CHF2, CF3, CN, C1-4 alkyl, C1-4
alkyl cydoalkyl, C1-4 alkyl cyclohetalkyl, or may be joined to form an optionally
substituted 3-6 membered ring optionally containing an atom selected! from 0,
S, NR14 and R14 is selected from H. C1-4 alkyl.
n = 0-4
W is selected from H, C1-4alkyl, C1-4alkenyl; where C1-4alkyl or C2-6alkenyl may
be optionally substituted with C1-4alkyl, OH OC1-4alkyl, NR15R16; and; R15,
and R16 are each independently H, C1-4 alkyl, C1-4 alkyl cydoalkyl. C1-4 alkyl
cydohetalkyl or may be joined to form an optionally substituted 3-8 membered
ring optionally containing an atom selected from O, S, NR17 and R17 is selected
from H C1-4 alkyl;
wherein when Y is OH or NHCOCH3 then R2 is 1-2 substituents and wherein when Y
is NH2 and R2 is absent then Y is in the para position.
In the above description it will be appreciated that:
C1-4 alkyl means a straight or branched alkyl chain
Aryl means unsubstituted or optionally substituted phenyl or naphthyl.
Hetaryl means an unsubstituted or optionally substituted 5- or 6-mernbered
heteroaromatic ring containing one or more heteroatoms selected from O, N, S.
Cycloalkyl means a 3-8 membered saturated ring
Cydohetalkyl means a 3-8 membered saturated ring containing 1-3
heteroatoms selected from O, S, NR18, where R18 is H C1-4 alkyl aryl hetaryl.
In a further preferred embodiment the compound is selected from compounds of the
general formula II.
or pharmaceutically acceptable salts, hydrates, solvates, crystal forms or diastereomers
thereof, wherein:
Rl is H,C1-4 alkyl
A is aryl, hetaryl optionally substituted with 0-3 substituents independently
chosen from halogen, C1-4 alkyl, CH2F, CHF2, CF3, CN, aryl, hetaryl OCR3, OC1-4
alkyl, OC2-5alkylNR4R5, Oaryl Ohetaryl CO2R4, CONR4R5, NR4R5, C1-4
alkylNR4R5, NR6C1-4alkylNR4R5, NR4COR5, NR6CONR4R3, NR4SCO2R5; and
R4 R5 are each independently H C1-4 alkyl, C1-4 alkyl cydoalkyl, C1-4 alkyl
cydohetalkyl, aryl hetaryl C1-4alkyl aryl, C1-4 alkyl hetaryl or may be joined to
form an optionally substituted 3-8 membered ring optionally containing an
atom selected from O, S, NR7; and R6 is selected from H, C1-4 alkyl; and R7 is
selected from H, C1-4 alkyl aryl hetaryl C1-4 alkyl aryl C1-4 alkyl hetaryl.
R2 is 0-2 substituents independently selected from halogen, C1-4alkyl,OH, OC1-4
alkyl, CH2F, CHF2, CF3, OCF3, CN, C1-4alkylNRSR9, OC1-4alkylNR8R9,, CO2R8,
CONR8R9, NR8R9, NR8COR9, NR10CONR8R9, NRSSO2R9; and R8, R9 are
each independently H, C1-4 alkyl C1-4 alkyl cycloalkyl, C1-4 alkyl cyclohetalkyl,
aryl, hetaryl, C1-4 alkkyl aryl, C1-4 alkyl hetaryl, or may be joined to form an
optionally substituted 3-8 membered ring optionally containing an atom
selected from 0,5, NR11; and RJO is selected from H, C1-4 alkyl, aryl or hetaryl;
and R11 is selected from H, C1-4 alkyl, aryl hetaryl, C1-4 alkyl aryl, C1-4 alkyl
hetaryl.
Y is halogen, OH, NR12R13, NR12COR13, NR12CONR13, N12SO2R13; and
R12, and R13 are each independently H, CH2F, CHF1, CF3, CN, C1-4 alkyl C1-4
alkyl cydoalkyl, C1-4 alkyl cyclohetalkyl, or may be joined to form an optionally
substituted 3-6 membered ring optionally containing an atom selected from 0,
S, NR14 and R14 is selected from H, C1-4 alkyl.
n = 0-4
W is selected from H, C1-4alkyl, C2-6alkenyl; where C1-4alkyl or C2-6alkenyl may
be optionally substituted with C1-4 alkyl, OH, OC1-4alkyl, NR15R16; and R15,
and R16 are each independently H, C1-4 alkyl C1-4 alkyl cydoalkyl, C1-4 alkyl
cydohetalkyl, or may be joined to form an optionally substituted 3-8 membered
ring optionally containing an atom selected from O, S, NR17 and R17 is selected
from H,C1-4 alkyl;
wherein when Y is OH or NHCOCH3, then R2 is 1-2 substituents and wherein when Y
is NH2 and R2 is absent then Y is in the para position.
In the above description it will be appreciated that:
C1-4 alkyl means a straight or branched alkyl chain
Aryl means unsubstituted or optionally substituted phenyl or naphthyl.
Hetaryl means an unsubstituted or optionally substituted 5- or 6-memmbered
heteroaromatic ring containing one or more heteroatoms selected from O, N, S.
Cycloalkyl means a 3-8 membered saturated ring
Cyclohetalkyl means a 3-8 membered saturated ring containing 1-3
heteroatoms selected from O, S, NR18, where R18 is H, C1-4 alkyl, aryl, hetaryl.
The compounds of this invention include all confbrmational isomers (eg. cis
and trans isomers). The compounds of the present invention have asymmetric centers
and therefore exist in different enantiomeric and diastereomeric forms. This invention
relates to the use of all optical isomers and stereoisomers of the compounds of the
present invention, and mixtures thereof, and to all pharmaceutical compositions and
methods of treatment that may employ or contain them. The compounds of formula I
may also exist as tautonvers. This invention relates to the use of all such tautomers and
mixtures thereof.
This invention also encompasses pharmaceutical compositions containing
prodrugs of compounds of the formula I. This invention also encompasses methods of
treating or preventing disorders that can be treated or prevented by the inhibition of
protein kinases, such as JAK comprising administering prodrugs of compounds of the
formula I. Compounds of formula I having free amino, amido, hydroxy or carboxylic
groups can be converted into prodrugs. Prodrugs include compounds wherein an
amino acid residue, or a polypeptide chain of two or more (eg, two, three or fbur)
amino acid residues which are covalently joined through peptide bonds to free amino,
hydroxy and carboxylic acid groups of compounds of formula I. The amino add
residues include the 20 naturally occurring amino adds commonly designated by
three letter symbols and also include, 4-hydroxyproline, hydroxylysine, demosine,
isodemosine, 3-methylhistidine, norvlin, beta-alanine, gamma-aminobutyric acid,
dtrulline, homocysteine, homoserine, omithine and methioine sulfone. Prodrugs also
include compounds wherein carbonates, carbamates, amides and alkyl esters which
are covalently bonded to the above substituents of formula I through the carbomyl
carbon prodrug sidechain. Prodrugs also indude phosphate derivatives of
compounds of formula I (such as adds, salts of adds, or esters) joined through a
phosphorus-oxygen bond to a free hydroxyl of compounds of formula I.
In a still further preferred embodiment the compound possesses ^chirality at
the chiral carbon bearing W, where W is C1-4 alkyl. The compound can be used as a
purified isomer or as a mixture of any ratio of isomers. It is however preferred that
the mixture comprises at least 70%, 80%, 90%, 95%, or 99% of the preferred isomer.
In a still further preferred embodiment the compound is selected from the
compounds set out in Table 1.
In a second aspect the present invention consists in a composition comprising a
carrier and at least one compound of the first aspect of the invention.
In a third aspect the present invention consists in a method of treating a protein
kinase-assocdated disease state, the method comprising administering a
therapeutically effective amount of at least one compound of the first aspect of the
invention or a therapeutically effective amount of a composition of the second aspect
of the invention.
In a preferred embodiment, the disease state involves a receptor tyrosine
kinase selected from the group consisting of EGF, HER2, HER3, HER4, IR,IGF-IR,
IRR, PDGFR-alpha., PDGFRbeta., CSHR C-Kit, C-fms,FIk-lR, FIk4, KDR/FIk-1, Flt-1,
FGFR-1R, FGFR-2R, FGFR-3R and FGFR-4R.
In another preferred embodiment the disease state involves a cellular tyrosine
kinase selected from the group consisting of Src, Frk, Btk, Csk, Abl, ZAP70, Fes/Fps,
Fak, Ack, Yes, Fyn, Lya Lck, BIk, Hck, Fgr and Yrk.
In a further preferred embodiment the disease state involves a tyrosine kinase
selected from the group consisting of JAK1, JAK2, JAK3 and TYK2.
In a yet further preferred embodiment, the disease state involves a
serine/threonine kinase selected from the group consisting of ERK2, c-Jun, p38
MAPK, PKA, PKB, PKC, a cyclin-dependent kinase, CDK1, CDK2, CDK3, CDK4,
CDK5, CDK6, CDK7, CDK8, CDK9, CDK10, and CDK11.
In a preferred embodiment of the present invention the disease state is selected
from the group consisting of Atopy, such as Allergic Asthma, Atopic Dermatitis
(Eczema), and Allergic Rhinitis; Cell Mediated Hypersensitivity, such as Allergic
Contact Dermatitis and Hypersensitivity Pneumonias; Rheumatic Diseases, such as
Systemic Lupus Erythematosus (SUE), Rheumatoid Arthritis, Juvenile Arthritis,
Sjogren's Syndrome, Sdetoderma, Polymyositis, Ankylosirtg Spondylitis, Psoriatic
Arthritis; Other autoimmune diseases such as Type I diabetes, autoimmune thyroid
disorders, and Alzheimer's disease; Viral Diseases, such as Epstein Barr Virns (EBV),
Hepatitis B, Hepatitis C, HIV, HTLV1, Varicella-Zoster Virus (VZV), Human
Papilloma Virus (HFV), Cancer, such as Leukemia, Lymphoma and Prostate Cancer.
In one embodiment the method of the invention is used in the treatment of
sarcomas, carcinomas and/or leukemias. Exemplary disorders for which the!subject
method can be used alone or as part of a treatment regimen include: fibrosarcoma,
myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma,
angiosarcoma, endotheliosarcorna, lymphangiosarcoma,
lymphangioendotheliosarcoma, synovioma, mesorhetioxna, Ewing's tumor,
leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast
cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma,
adenocarecioma, sweat gland carcinoma, sebaceous gland carcinoma, papillary
cardnoma, papillary adenocardnomas, cystadenocardnoma, medullary cardnoma,
bronchogenic carcinoma, renal cell cardnoma, hepatoma, bile duct carcinoma,
choriocardnoma, seminoma, embryonal carcinoma, Wilms tumor, cervical cancer/
testicular tumor, lung cardnoma, small cell lung cardnoma, bladder carcinoma,
epithelial cardnoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma,
ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma,
meningioma, melanoma, neuroblastoma, and retinoblastoma.
In certain embodiments, the method of the invention is be vised to treat
disorders such as carcinomas forming from tissue of the breast, prostate/ kidney,
bladder or colon.
In other embodiments, the method of the invention is used to treat hyperplastic
or neoplastic disorders arising in adipose tissue, such as adipose cell tumors, e.g.,
lipamas, fibrolipomas, lipoblastomas, lipomatosis, hibemomas, hexnangiomas and/or
Iipbsarcomas.
As used herein the term "protein kinase-associated disease state" refers to those
disorders which result from aberrant protein kinase activity, in particular JAK activity
and/or which are alleviated by inhibition of one or more of these enzymes.
In further aspects the present invention provides the use of the compounds
described in the preparation of medicaments for the treatment of protein kinase-
assodated disease states including JAK-assodated disease states.
As used herein the term "JAK", "JAK kinase" or "JAK family" refers to: protein
tyrosine kinases which possess the characterizing features of JAK1, JAK2, JAK3 and
TYK as described herein.
The present invention provides pharmaceutical compositions comprising at
least one of the compounds of the formula I or II capable of treating a protein kinase-
assodated disorder in an amount effective therefor, and a phannaceutically acceptable
vehicle or diluent The compositions of the present invention may contain other
therapeutic agents as described below, and may be formulated, for example, by
employing conventional solid or liquid vehicles or diluents, as well as pharmaceutical
additives of a type appropriate to the mode of desired administration (for example,
excipients, binders, preservatives, stabilizers, flavors, etc) according to techniques
such as those well known in the art of pharmaceutical formulation.
The compounds of the formula I or IT may be administered by any suitable
means, for example/ orally, such as in the form of tablets, capsules, granules or
powders; sublingually; buccally; parenterally, such as by subcutaneous, intravenous,
intramuscular, or intradstemal injection or infusion techniques (e.g., as sterile
injectable aqueous or non-aqueous solutions or suspensions); nasally such as by
inhalation spray; topically, such as in the form of a cream or ointment; or radially such
as in the form of suppositories; in dosage unit formulations containing non-toxic,
phannaceutically acceptable vehicles or diluents. The compounds may, for example,
be administered in a form suitable for immediate release or extended release.
Immediate release or extended release may be achieved by the use of suitable
pharmaceutical compositions comprising the present compounds, or, particularly in
the case of extended release, by title use of devices such as subcutaneous implants or
osmotic pumps.
In addition to primates, such as humans, a variety of other mammals can be
treated according to the method of the present invention. For instance, mammals
induding, but not limited to, cows, sheep, goats, horses, dogs, cats, guinea pigs, rats or
other bovine/ ovine, equine, canine, feline, rodent or murine species can be treated.
However, the method can also be practiced in other species, such as avian species (e.g.,
chickens).
Diseases and conditions associated with inflammation and infection can be
treated using the method of the present invention. In a preferred embodiment the
disease or condition is one in which the actions of eosinophils and/or lymphocytes are
to be inhibited or promoted, in order to modulate the inflammatory response.
The subjects treated in the above methods, in whom which JAK inhibition is
desired, are mammals, including, but not limited to, cows, sheep, goats, horses, dogs,
cats, guinea pigs, rats or other bovine, ovine, equine, canine, feline, rodent of murine
species, and preferably a human being, male or female.
The term "therapeutically effective amount" means the amount of the subject
composition that will elicit the biological or medical response of a tissue, system,
animal or human that is being sought by the researcher, veterinarian, medical doctor
or other clinician.
The term "composition" as used herein is intended to encompass a product
comprising the specified ingredients in the specified amounts, as well as any product
which results, directly or indirectly, from combination of the specified ingredients in
the specified amounts. By "phannaceutically acceptable" it is meant the carrier,
diluent or exripient must be compatible with the other ingredients of the formulation
and not deleterious to the recipient thereof.
The terms "administration of and or "administering a" compound should be
understood to mean providing a compound of the invention to the individual in need
of treatment.
The pharmaceutical compositions for the administration of the compounds of
this invention may conveniently be presented in dosage unit form and may be
prepared by any of the methods well known in the art of pharmacy. All methods
include the step of bringing the active ingredient into association with the carrier
which constitutes one or more accessory ingredients. In general, the pharmaceutical
compositions are prepared by uniformly and intimately bringing the active ingredient

into association with a liquid carrier or a finely divided solid carrier or both/ and then,
if necessary, shaping the product into the desired formulation. In the pharmaceutical
composition the active object compound is included in an amount sufficient to
produce the desired effect upon the process or condition of diseases. As used herein/
the term "composition" is intended to encompass a product comprising the specified
ingredients in the specified amounts/ as well as any product which results, directly or
indirectly, from combination of the specified ingredients in the specified amounts.
The pharmaceutical compositions containing the active ingredient may be in a
form suitable for oral use, for example, as tablets/ troches, lozenges, aqueous or oily
suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or
syrups or elixirs. Compositions intended for oral use may be prepared according to
any method known to the art for the manufacture of pharmaceutical compositions and
such compositions may contain one or more agents selected from the group consisting
of sweetening agents, flavoring agents, coloring agents and preserving agents in order
to provide pharmaceutically elegant and palatable preparations. Tablets contain the
active ingredient in admixture with non-toxic pharmaceutically acceptable excipients
which are suitable for the manufacture of tablets. These exdpients may be for
example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium
phosphate or sodium phosphate; granulating and disintegrating agents, for example,
corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia, and
lubricating agents, for example magnesium stearate, stearic acid or talc The tablets
may be uncoated or they may be coated by known techniques to delay disintegration
and absorption in the gastrointestinal tract and thereby provide a sustained action
over a longer period. For example/ a time delay material such as glyceryl
monostearate or glyceryl distearate may be employed. They may also be coated to
form osmotic therapeutic tablets for control release.
Formulations for oral use may also be presented as hard gelatin capsules
wherein the active ingredient is mixed with an inert solid diluent for example,
calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein
the active ingredient is mixed with water or an oil medium, for example peanut oil,
liquid paraffin, or olive oil.
Aqueous suspensions contain the active materials in admixture with excipients
suitable for the manufacture of aqueous suspensions. Such excipients are suspending
agents, for example sodium carboxymethylcellulose, methylcellulose, hydroxy-
propylmethylcellulose, sodium alginate, polyvinyl-pyrrolidone, gum tragacanth and
gum acacia; dispersing or wetting agents may be a naturally-occurring phosphatide,
for example lecithin, or condensation products of an alkylene oxide with fatty adds,
for example polyoxyethylene stearate, or condensation products of ethylene oxide
with long chain aliphatic alcohols, for example heptadecaethyleneoxycetanol, or
condensation products of ethylene oxide with partial esters derived from fatty acids
and a hexitol such as polyoxyethylene sorbitol monooleate, or condensation products
of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides/
for example polyethylene sorbitan monooleate. The aqueous suspensions may also
contain one or more preservatives, for example ethyl, or n-propyl, p-hydroxybenzoate,
one or more coloring agents, one or more flavoring agents, and one or more
sweetening agents, such as sucrose or saccharin.
Oily suspensions may be formulated by suspending the active ingredient in a
vegetable oil, for example arachis oil, olive oil, sesame oil or coconut oil, or in a
mineral oil such as liquid paraffin. The oily suspensions may contain a thickening
agent, for example beeswax hard paraffin or cetyl alcohol. Sweetening agents such as
those set forth above, and flavoring agents may be added to provide a palatable oral
preparation. These compositions may be preserved by the addition of an anti-oxidant
such as ascorbic add.
Dispersible powders and granules suitable for preparation of an aqueous
suspension by the addition of water provide the active ingredient in admixture with a
dispersing or wetting agent, suspending agent and one or more preservatives.
Suitable dispersing or wetting agents and suspending agents are exemplified by those
already mentioned above. Additional excipients, for example sweetening, flavoring
and coloring agents, may also be present.
The pharmaceutical compositions of the invention may also be in the form of
oil-in-water emulsions. The oily phase may be a vegetable oil, for example olive oil or
arachis oil, or a mineral oil, for example liquid paraffin or mixtures of these. Suitable
emulsifying agents may be naturally- occurring gums, for example gum acacia or gum
tragacanth, naturally-occurring phosphatides, for example soy bean, lecithin, and
esters or partial esters derived from fatty acids and hexitol anhydrides, for example
sorbitan monooleate, and condensation products of the said partial esters with
ethylene oxide, for example polyoxyethylene sorbitan monooleate. The emulsions
may also contain sweetening and flavoring agents.
Syrups and elixirs may be formulated with sweetening agents, for example
glycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a
demulcent a preservative and flavoring and coloring agents.
The pharmaceutical compositions may be in the form of a sterile injectable
aqueous or oleagenous suspension. This suspension may be formulated according to
the known art using those suitable dispersing or wetting agents and suspending
agents which have been mentioned above. The sterile injectable preparation may also
be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable
diluent or solvent for example as a solution in 13-butane diol. Among the acceptable
vehicles and solvents that maybe employed are water, Singer'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. In addition, fatty acids such as
oleic acid find use in the preparation of injectables.
The compounds of the present invention may also be administered in the form
of suppositories for rectal administration of the drug. These compositions can be
prepared by mixing the drug with a suitable non-irritating excipient which is solid at
ordinary temperatures but liquid at the rectal temperature and will therefore melt in
the rectum to release the drug. Such materials are cocoa butter and polyethylene
glycols.
For topical use, creams, ointments, jellies, solutions or suspensions/ etc,
containing the compounds of the present invention are employed. (For purposes of
this application, topical application shall include mouthwashes and gargles.)
The compounds of the present invention can also be administered in the form
of liposomes. As is known in the art liposomes are generally derived from
phospholipids or other lipid substances. Liposomes are formed by mono or
multamellar hydrated liquid crystals that are dispersed in an aqueous medium. Any
non-toxic, physiologically acceptable and metabolisable lipid capable of forming
liposomes can be used. The .present compositions in iiposome form can contain, in
addition to a compound of the present invention, stabilisers, preservatives, excipients
and the like. The preferred lipids are the phospholipids and phosphatidyl cholines,
both natural and synthetic. Methods to form liposomes are known in the art
The pharmaceutical composition and method of the present invention may
further comprise other therapeutically active compounds as noted herein which are
-usually applied in the treatment of the above mentioned pathological conditions.
Selection of the appropriate agents for use in combination therapy may be made by
one of ordinary skill in the art according to conventional pharmaceutical principles.
The combination of therapeutic agents may act synergistically to effect the treatment
or prevention of the various disorders described above. Using this approaclvone may
be able to achieve therapeutic efficacy with lower dosages of each agent thus reducing
the potential for adverse side effects.
Examples of other therapeutic agents include the following:
cyclosporins (e.g., cydosporin A), CTLA4-Ig, antibodies such as ICAM-3, anti-IL-2
receptor (Anti-Tac), anti-CD45RB, anti-CD2, anti-CD3 (OKT-3), anti-CD4, anti-CD80,
anti-CD86, agents blocking the interaction between CD40 and gp39, such a$ antibodies
specific for CD40 and/ or gp39 (i.e., CD154), fusion proteins constructed from CD40
and gp39 (CD401g and CD8gp39), inhibitors, such as nuclear translocation inhibitors,
of NF-kappa 6 function, such as deoxyspergualin (DSG), cholesterol biosynthesis
inhibitors such as HMG CoA reductase inhibitors (lovastatin and simvastatin),
non-steroidal antiinflammatory drugs (NSAIDs) such as ibuprofen, aspirin, .
acetaminophen and cyclooxygenase inhibitors such as rofecoxib, steroids such as
prednisolone or dexamethasone, gold compounds, antiproliferative agents such as
methotrexate, FK506 (tacrolimus, Frograf), mycophenolate mofeal, cytotoxic drugs
such as azathioprine, VF-16, etoposide, fludarabine, cisplatin and cyclophosphamide,
TNF-a inhibitors such as tenidap, anti-TNF antibodies or soluble TNF receptor, and
rapamycin (sirolimu6 or Rapamune) or derivatives thereof.
When other therapeutic agents are employed in combination with the
compounds of the present invention they may be used for example in amounts as
noted in the Physician Desk Reference (PDR) or as otherwise determined by one of
ordinary skill in the art.
In the treatment or prevention of conditions which require protein kinase
inhibition an appropriate dosage level will generally be about 0.01 to 500 mg per kg
patient body weight per day which can be administered in single or multiple doses.
Preferably/ the dosage level will be about 0.1 to about 250 mg/kg per day; more
preferably about 0.5 to about 100 mg/kg per day. A suitable dosage level may be
about 0.01 to 250 mg/kg per day, about 0.05 to 100 mg/kg per day, or about 0.1 to 50
mg/kg per day. Within this range the dosage may be 0.05 to 0.5,0..5 to 5 or 5 to 50
mg/kg per day. For oral administration, the compositions are preferably provided in
the form of tablets containing 1.0 to 1000 milligrams of the active ingredient
particularly 1.0,5.0,10.0,15.0. 20.0,25.0,50.0,75.0,100.0,150.0,200.0,250.0,300.0,
400.0,500.0,600.0,750.0, 800.0,900.0, and 1000.0 milligrams of the active ingredient for
the symptomatic adjustment of the dosage to the patient to be treated. The
compounds may be administered on a regimen of 1 to 4 times per day, preferably once
or twice per day.
It will be understood, however, that the specific dose level and frequency of
dosage for any particular patient may be varied and will depend upon a variety of
factors including the activity of the specific compound employed, the metabolic
stability and length of action of that compound, the age, body weight general health,
sex, diet mode and time of administration, rate of excretion, drug combination, the
severity of the particular condition, and the host undergoing therapy.
Throughout this specification the word "comprise", or variations such as
"comprises" or "comprising", will be understood to imply the inclusion of a stated
element integer or step, or group of elements, integers or steps, but not the exclusion
of any other element integer or step, or group of elements, integers or steps.
All publications mentioned in this specification are herein incorporated by
reference.
Any discussion of documents, acts, materials, devices, articles or the like which
has been included in the present specification is solely for the purpose of providing a
context for the present invention. It is not to be taken as an admission that any or all
of these matters form part of the prior art base or were common general knowledge in
the field relevant to the present invention as it existed in Australia before the priority
date of each claim of this application.
In order that the nature of the present invention may be more clearly
understood preferred forms thereof will now be described by reference to the
following non-limiting Examples.
MATERIALS AND METHODS:
Compound Synthesis
Compounds are generally prepared in a 2-step process starring from 2,6-
dichloropyrazine.
The first step is a nucleophilic aromatic substitution to generate a monoamino-
monohalo intermediate. (Scheme 1).

The nucleophilic aromatic substitution is typically carried out by addition of a
primary amine to the di-halogenated heterocyle in a solvent such as ethanol,
isopropanol, tert-butanol, dioxane, THF, DMF, toluene or xylene. The reaction is
typically performed at elevated temperature in the presence of excess amine or a non-
nudeophilic base such as triemylanune or diisopropylethylamme, or an inorganic base
such as potassium carbonate or sodium carbonate.
Alternatively, the amino substituent may be introduced through a transition
metal catalysed amination reactioa Typical catalysts for such transformations include
Pd(OAc)2/P(t-Bu)3, Pd2{dba)3/BINAP and Pd(OAc)2 BINAP. These reactions are
typically out in solvents such as toluene or dioxane, in the presence of bases such as
caesium carbonate or sodium or potassium tert-butoxide at temperatures ranging
from room temperature to reflux.
The amines employed in the first step of the synthesis of these compounds are
obtained commercially or are prepared using methods well known to those skilled in
the art Of particular interest are a-methylbenzylamines which may be prepared
through reduction of oximes (Scheme 2). Typical reductants include lithium
aluminium hydride, hydrogen gas in the presence of palladium on charcoal catalyst,
Zn in the presence of hydrochloric add, sodium borohydride in the presence of a
Lewis acid such as TiCl2, ZrCl4, NiCl2 and MoO3, or sodium borohydride in
conjunction with Amberlyst H15 ion exchange resin and lid.

a-Methyl benzylamines of high optical purity may be prepared from chiral de
methyl benzyl alcohols using methods well known to those skilled in the art Such
methods include derivatisation of the hydroxyl as a mesylate or tosylate and
displacement with a nitrogen nucleophile, such as phthalimide or azide which is then
converted to the primary amine using conventional synthetic methods; or,
displacement of the hydroxyl with a suitable nitrogen nucleophile under Mitsunobu
conditions. The chiral a-methyl benzyl alcohols may be obtained through chiral
reduction of the corresponding ketones. Chiral reducing methods are now well
known in organic chemistry and include enzymatic processes, asymmetric
hydrogenation procedures and chiral oxazaborolidines.
The second step of the synthesis typically involves a palladium mediated cross-
coupling of the monoaxnino-manochloro intermediate with a suitably fundionalified
coupling partner. Typical coupling partners are borornic adds (Suzuki coupling: see
for example Miyaura, N. and Suzuki Chem Her. 1995, 952457) or stannanes (Stille
coupling: see for example Stille, J.K., Angew. Chew., Int Ed. Engl., 1986,25,508)
(Scheme 3).

The Suzuki coupling is the preferred coupling method and is typically
performed in a solvent such as DME, THF, DMF, ethanol, propanol, toluene, or 1,4-
dioxane in the presence of a base such as potassium carbonate, lithium hydroxide,
caesium carbonate, sodium hydroxide, potassium fluoride or potassium phosphate.
The reaction maybe carried out at elevated temperatures and the palladium catalyst
employed may be selected from [Pd(PPh3)4], Pd(OAc)2 [PdCl2(dppf)l. Pd2(dba)3/P(t-
Bu)3.
The products formed from this reaction sequence may be further derivatised
using techniques well-known to those skilled in the art Alternatively, derivatisation
of the mono-amino mono-chloropyrazine may be undertaken prior to displacement of
the 6-chloro subsntuent This derivatisation typically involves functionality originally
present on the amine species and employs methods well known to those skilled in the
art.
Representative syntheses are reported below.
Example 1

A solution of R-a-methylbenzylamine (057g, 4.7mmol) and 2,6-
dichloropyrazine (0.6388g, 429inmol) in dioxane (2.5 mL) was heated at reflux under
N2 for 48 hours. The solvent was removed and the product crystallised front toluene-
hexane(0.82g,82%).
1H-n.m.r. (CDa3) d 1.58 (d, J= 6.6HZ, 3H, CH3), 4.88 (m, 1H, CH), 5-07 (d, 1H,
NH), 724-7.36 (m, 5H Ap-H), 7.61 (8,1H, pyraz-H), 7.79 (s, 1H pyraz-H).

Under a nitrogen atmosphere a mixture of 6-chloro-N-{(1R)-1-
phenyIethyI]pyrazin-2-amine (0.611g, 2-61mmol), 4-(4,4,4,5-tetxamethyl-1,3,2-
dioxaborolan-2-yl)phenol (0.785g, 3-14mmol),
tetrakids(triphenylpho&phine)palladium(0) (030g, 0.26mmol) and toluene (3rnL) was
treated with 2M aqueous sodium carbonate solution {1.6mL, 2.6mmol). The resulting-
mixture was stirred vigorously whilst being heated under reflux for 24 hours. Once
cool ethyl acetate was added and the mixture dried (MgSO4) and filtered. Removal of
solvent in vacua then yielded crude product, which was purified by column
chromatography using dichloromethane:diethyl ether (99:1? 90:10) as eluenr (0.619g,
74%).
1H-n.m.r. (CDCl3) 81.72 (d, 3H, J=6.9 Hz, CH9), 4.06 (s, 3H, OCH3), 4.90 (m,
1H, CH), 5.75 (br s, 1H, NH), 6.98 (d, 1H, J= 8.1 Hz, ArH), 7.26-7.46 (rn, 7H, Ar-H),
7.97 (s, 1H, pyraz-H), 8.20 (s, 1H, pyraz-H).
Example 3

In a procedure analogous to Example 1, reaction of R-a-methylbernzylamine
(1.0g, 6.6mmol) and 2,6-dichloropyrazine (0.440g, 2.95mmol) furnished the product
(517mg, 67%).
1H-n.m.r. (CDCl3) d 1.59 (d, J=6.9Hz, 3H, CH) 3.81 (sy 3H OCH3), 4.87 (m, 1H,
CH), 5.47 (br s,1H, NH), 6.79-7.30 (m, 4H Ar-H), 7.66 (s, 1H, pyraz-H), 7.79 (s, 1H,
pyraz-H).
Example 4

In a procedure analogous to Example 2, reaction of 2-(R-a-methyl-3-methoxy-
benzylamino)-6-chloro-pyrazlne (137.2mg, 0.52 mmol) and 2-methoxy-4-(4,45,5-
tetramethyl-1,3,2-dioxaborolan-2-yl)phenol (143mg, 0.57mmol) furnished the product
(32mg, 18%).
1H-n.m.r. (CDCl3) d1.61 (d, J=6.6Hz, 3H, CH3), 3.79 {s, 3H OCH3), 3.94 (s, 3H.
OCH3), 4.94 (m, 1H, CH), 5.02 (d, J=6Hz, 1H, NH), 6.04 (br s, 1H, OH), 6.77-7.48 (m 7H.
Ar-H), 7.69 (s, 1H, pyraz-H), 8.23 (s, 1H, pyraz-H)
m/z(ES)352(M++H).
Example 5

In a procedure analogous to Example 1, reaction of R-a-methylbenzylamine
(1.0g, 6.6mmol) and 2,6-dichloropyrazine (0.4355g, 2.92mmol) furnished the product
(0.72g, 93%).
1H-n-m.r. (CDCl3) d1.56 (d, 3H, J= 6.9Hz, CH3), 3.80 (s, 3H, OCH3), 4.84 (m, 1H,
CH), 525 (br s, 1H, NH), 6.88 (AA'XX', 2H, Ar-H), 7.28 (AA'XX', 2H, Ar-H). 7.64 (s,
1H, pyraz-H), 7.78 (s, 1H, pyraz-H).
Example 6

In a procedure analogous to Example 2, reaction of 2-{R-a-methyl-4-methoxy-
benzy]amino)-6-chloro-pyrazine (127.1mg, 0.48mmol) and 2-methoxy-4-(4,4,5,5-
tetramethy-1,3,2-dioxaborolan-2-yl)phenol (145mg, 058mmol) furnished the product
(59.5mg,35%).
1H-n.m.r. (CDCls) d1.59 (d, 3H, J= 6.6 Hz, CH,), 3.79 (s, 3H, OCH,), 3.95 (s, 3H
OCH3), 4.97 (m, 2H, CH and NH), 5.95 (br s, 1H OH), 6.87 (AA'XX', 2H, ArH), 6.97 (d,
1H, J= 8.1 Hz, ArH), 732 (AA'XX', 2H, Ar-H), 7.46 (m, 2H, ArH), 7.66 (s, 1H, pyraz-
H),8.22(s,1H,pyraz-H}.
m/z(ES)352{M++H).
Example 7

In a procedure analogous to Example 1, reaction of (1R)-1,2,3,4-
tetrahydronaphthalen-1-amine (441mg, 3.Qmmol) and 2,6-dichloropyrazine (0.4055g,
2.72mmol) furnished the product (521mg, 74%).
1H-n.m.r. (CDCl3) d1.89 (m, 2H, CH2OH2Ar), 1.97 (nv 1H, H-
CHCH2CH2Ar)2.08 (m, 1H, HC-H-CH2CH2Ar), 2.83 (m, 2H, CH2Ar2)/ 4.94 (br s, 1H,
NH) 5.15 (m, 1H, CH), 7.12-731 (m, 4H, Ar-H), 7.76 (s, 1H, pyraz-H), 7.81 (s, 1H,
pyraz-H).
Example 8

In a procedure analogous to Example 2, reaction of 6-chloro-N-{(1R)-1,2,3,4-
tetrahydronaphthalen-l-yl]pyrazin-2-amine (139mg, 0536mmol) and 2-methoxy-4-
(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenol (147mg, 039mmol) furnished the
product (87mg, 47%).
1H-n.m.r. (CDCl3) d 1.91 (m, 2H, CJH2Ar), 2.09 (m, 2H, CH2CH2CH2Ar),2-85
(m, 2H, CH2Ar), 3.96 (s, 3H, OCH3), 4.87 (d,/= 7.SHz, 1H, NH), 5.28 (m, 1H, CH), 6.04
(br s, 1H, OH), 6.9B-7.73 (m, 7H, Ar-H), 7.79 (s, 1H, pyraz-H), 8.26 (s, 1H, pyraz-H).
m/z(ES)348(M++H).
Example 9

In a procedure analogous to Example 1, reaction of (lR)-2,3-dihydro-1H-inden-
1-ylamine (1.0g, 7.6mmol) and 2,6-dichloropyrazine (0-452g, 3.04mmol) furnished the
product (673.8mg, 90%).
1H-n.m.r (OX33) d1-91 (m, 1H, H-CHCH2Ar), 2.68 (m, 1H, HC-U-CHCH2Ar),
3.00 (m, 2H CH2Ar), 5.03 (br s, 1H, NH), 5.45 (m, 1H, CH), 7.18-733 (m, 4H,. Ar-H),
7.82 (br s, 2H, 2xpyraz-H).
Example 10

In a procedure analogous to Example 2, reaction of 6-chloro-N-((1R)-2,3-
dihydro-lH-inden-l-yi]pyrazin2-amine (136.8mg, 0-56mmol) and 2-methoxy-4-
(4,4,5,5-tetramethyl-1,2-dioxaborolan-2-yl)phenol (153mg, X0.61mmol) furnished the
product (130mg, 70%).
1H-n.m.r. (CDCl) d 2.00 (m, 1H, HC-H-CH2Ar), 2.71 (m, 1H, H-CHCH2Ar)3.01
(m, 2H, CH2Ar), 3.96 (s, 3H, OCH3), 4.90 (d, J= 7.8Hz, 1H, NH), 5.57 (m, 1H, CH), 6.06
(br s, 1H, OH), 6.98-7.82 (m, 7H, Ar-H), 7.85 (s, 1H, pyraz-H), 829 (s, 1H pyraz-H);
m/z(ES)334(M++H).
Example 11

In a procedure analogous to Example 1, reaction of a-(R)-4-
dimethylbenzylamine (250mg, 1.85mmol) and 2,6-dichloropyrazine (0.251g, 1.67mmol)
furnished the product (199.5mg, 48%).
1H-n.m.r. (CDCl3) d1.56 (d, 3H, J= 6.9H2, CH3), 2.33 (s, 3H. CH3), 4.84 (m, 1H,
CH), 5.05 (br s, 1H, NH), 7.15 (AA'XX', 2H, Ar-H), 72.4 (AA'XX', 2H, Ar-H), 7.60 (s,
1H, pyraz-H), 7.78 (s, 1H, pyraz-H).
Example 12

In a procedure analogous to Example 2, reaction of 6-chIoro-N-[(1R)-1-(4-
methylphenyl)ethyl]pyrazin-2-amine (56.8mg, 0.229mmol) and 2-methoxy-4-(4,4,5,5-
tetramethyl-1,3,2-dioxaborolan-2--yl)phenol 63mg, 0.25mmol) furnished the product
(5mg, 6%).
1H-n-m-r. (CDCl3) d1.60 {df 3H, J= 6.6 Hz, CH3), 2.33 (s, 3H, CH3), 3.95 (s, 3H,
OCH3), 4.96 (m, 2H, CH and NH), 5.89 (br s, 1H, OH), 6.97 (d, 1H,/= 8.4 Hz, ArH),
7.14 (AA'XX',2H, ArH),, 7.30 (AA'XX', 2H, Ar-H), 7.42-7.48 (m, 2H, Ar-H).. 7.67 (s, 1H,
pyraz-H), 8.62 (s, 1H,. pyraz-H).
m/z(ES)336(M++H).
Example 13

In a procedure analogous to Example 1, reaction of 5-a-methylbenzylamine
(568.8mg, 4.72mmol) and 2,6-dichloropyrazine (0.6388g, 4.29mmol) furnished the
product (821mg, 82%).
1H-n.m.r. (CDCl3) d 1.58 (d, J=6.6Hz, 3H, CH,), 4.88 (m, 1H, CH), 5.07 (d, 1H,
NH), 7.24-7.36 (m, 5H, Ar-H), 7.61 (s, 1H, pyraz-H), 7.79 (s, 1H, pyraz-H).
Example 14

In a procedure analogous to Example 2, reaction of 6-chloro-N-[(1S)-l-
phenylethyl]pyrazin-2-amine (7173mg, 3.07mmol) and 2-methoxy-4-(4,4,5,5-
tetramethyl-lA2-dioxaboroJan-2-yl)phcnol (845mg, 3.38mmol) furnished the product
(689mg, 70%).
1H-n.m.r. (CDCl3) d1.63 (d, 3H,/= 6.6 Hz, CH,), 3.95 (s, 3H, OCH3), 4.99 (m,
2H, CH+NH), 5.74 (br s, 1H, OH), 6.97 (d, 1H, J= 8.1 Hz, Ar-H), 7.24-7.46 (m, 7H, Ar-
H), 7.69 (s, 1H, pyraz-H), 8.23 (s, 1H, pyraz-H).
Example 15

In a procedure analogous to Example 1, reaction of S-a-ethylbenzylamine
(558mg, 4.21mmol) and 2,6-dichloropyrazine (570 mg, 3.82mmol) furnished the
product (655mg, 73%).
1H-n.m.r. (OXls) S 0.96 (t, 3H CH3), 1.90 (m, 2H, CH2), 4.59 (m, 1H, CH), 5.12
(d, 1H. NH), 7.24-7.37 (m, 5H, Ar-H), 7.60 (s, 1H, pyraz-H), 7.78 (s, 1H. p)Taz-H).
Example 16

In a procedure analogous to Example 2, reaction of 6-chloro-N-[(1S)-1-
phenylpropyI]pyrazin-2-amine (135mg, 0.57mmol) and 2-methoxy-4-(4,45,5-
tetramethyl-1,3,2-dioxaborolan-2-yl)phenol (158mg, 0.63mmol) furnished the product
(87mg, 45%).
1H-n.m.r. (CDCl3) d1.00 (t 3H, J= 7.5 Hz, CH5), 1.94 (dq, 2H, /= 7.5 Ha. CH2),
3.96 (S, 3H, OCH3), 4.71 (dt, 1H, J= 75 Hz, CH), 5.00 (br s, 1H, NH), 5.75 (br s, 1H,
OH), 6.97 (d, 1H J= 8.4 Hz, ArH), 724 (m, 1H, ArH), 7.30-7.47 (m, 6H, ArH), 7.67 (s,
1H, pyraz-H), 8.21 (s, 1H, pyraz-H).
m/z(ES)336(M++H).
Example 17

In a procedure analogous to Example 1, reaction of (2R)-2-amino-2-
phenylethanol (420mg, 3.1mmol) and 2,6-dichloropyrazme (415mg, 2.79mmol)
furnished the product (261mg, 37%).
1H-n.m.r. (CDCl3) d 0.91 (d, 1H, OH), 3.97 (m, 2H, CH2), 4.94 (m. 1H, CH), 5.56
(d, 1H, NH), 7.30-7,44 (m, 5H, Ar-H), 7.70 (s, 1H, pyraz-H), 7.81 (s, 1H, pyraz-H).
Example 18

In a procedure analogous to Example Z reaction of (2R)-2-[(6-chloropyrazin-2-
yl)amino]-2-phenyleihanol (137mg, 0.55 mmol) and 2-methoxy-4-(4,4,5,5-tetramethyl-
1,32-dioxaborolan-2-yl)phenol (151mg, 0.60mmol) furnished the product (70mg, 38%).
1H-n.m.r. (CDCl3) d1.16 (s, 1H OH), 382 (s, 3H, OCH3), 3.90 (m, 2H, CH2), 4.92
{m, 1H, CH), 5.50 (br s, 1H, NH), 6.87 (d, 1H,J= 9 Hz, ArH), 7.15-7.66 (m, 8H ArH),
8.14 (s, 1H, pyraz-H).
m/2 (ES) 338 (M++H).
Example 19

In a procedure analogous to Example 1, reaction of 4-methoxy-a-(S)-
methylbenzylamine (0.70mg; 4.6mmol) and 2,6-dichloropyrazine (0.62S9g, 4.20mmol)
furnished the product (873mg, 79%).
'H-rrnur. (CDCl3) d 1.56 (d, 3H, J= 6.9Hz, CH3), 3.80 (s, 3H, OCH,), 4.84 (m, 1H,
CH), 5.01 (br s, 1H NH), 6.88 (AA'XX', 2H, Ar-H), 7.28 (AA'XX', 2H, Ar-H), 7.61 (s,
1H, pyraz-H), 7.79 (s, 1H, pyraz-H).
Example 20

In a procedure analogous to Example 2, reaction of 6-chloro-N-[(1S)-1-(4-
methoxyphenyl)ethyl]pyrazin-2-arnme (149.4mg, 0.57znmol) and 2-methoxy-4-(4,4,5,5-
tetramethyl-1,3,2-dioxaborolan-2-yl)phenol (156mg, 0.62mmol) furnished the product
(71mg,35%).
1H-n.m.r.. (CDCl3) d1.59 (d, 3H, J= 6.6 Hz, CH3), 3.79 (s, 3H, OCH3), 3.95 (s, 3H,
OCHj), 4.95 (m, 2H. CH and NH), 5.98 (br s, 1H, OH), 6.87 (AA'XX', 2H, ArH), 6.97 (d,
lH, J= 8.1 Hz, ArH), 7.33 (AA'XX', 2H, Ar-H), 7.43-7.49 (nv 2H, ArH), 7.66. (s, 1H.
pyraz-H), 8.22 (s, 1H, pyraz-H).
m/z(ES)352(M++H).
Example 22

A mixture 2,6-dichloropyrazine (0.671 mmol) and 3-picolylamine (2.014 mmol)
in xylene (25 ml) was refluxed overnight. The residue obtained after evaporation of
the solvent was suspended between CH2Cl2 (100 ml) end water (100 ml)- The organic
layer was separated and the aqueous layer was extracted with CH2Cl2 (3 x 50 ml). The
combined organic extracts were washed with brine (1 x 100 ml), dried (Na2SO4 and
the solvent removed in vacua The residue was then purified by column
chromatography eluting with a hexane:ethyl acetate gradient mixture to afford the
desired product (93%).
1H-n.m.r. (CDC13) 54.61 (d, J= 5.7 Hz, 2 H, NCH2), 5.29 (s, broad, 1H, NH), 7.27
(m, 1H, pyrid.-H), 730 (m, 1H, pyrid.-HX 7.71 (d, J= 7.8 Hz, 1H, pyrid.-H), 7.85 (s, 1H,
pyrid.-H), 8.54 (s, broad, 1H, pyraz.-H), 8.61 (s, broad, 1H, pyraz.-H).
Example 22

A mixture of 6-chloro-N-(pyridin-3-ylmethyl)pyrazin-2-amine (49mg, 0.22
mmol), 2-methoxy-4-(4,45,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenol (52ms, 020
mmol), (PPh3)4Pd (23mg, 0.020 mmol) and a Na2CO3 solution (022 mmol of a 2 M
solution) in toluene (10 ml) was heated under reflux overnight. After removal of the
solventsm the residue was dissolved in CH2C12 (150 ml), dried (Na2SO4), filtered and the
CH2Cl2 removed in vacuo. The residue was purified by column chromatography, ,
eluting with a n-hexane:ethyl acetate gradient mixture to obtain the desired product
(62mg, 75%).
1H-n.m.r. (CDCl3) d3.94 (br s, 3H, CH3), 4.70 (d, 2H, J= 6.0 Hz, CH2), 5.01 (br s,
1H, NH), 5.83 (br s, 1H, OH), 6.98 (d, 1H, J= 8.7Hz, ArH), 7.29 (m, 1H, Ar-H), 7.48 (m,
2H, ArH), 7.73 (br d, lH, J= 8.7 Hz, ArH), 7.83 (s, 1H, pyraz-H), 830 (s, 1H, pyraz-H),
8.54 (m, 1H, ArH), 8.70 (s, 1H, ArH).
m/z(ES)309(M++H).
Example 23

In a procedure analogous to Example 21, reaction of N-methyl benzylamine
and 2,6-dichloropyrazine furnished the product (70%).
JH-n.m.r. (CDCl3) d3.11 (s, 3 H, NCH3), 4.78 (s, 2H, ArCH2N), 7.24 (d, J= 6.9 Hz,
2 H, ArH), 7.37-758 (m, 4H, ArH), 7.81 (s, 1H, pyraz.-H), 7.88 (s, 1H, pyraz.-H).
Example 24

In a procedure analogous to Example 22, reaction of N-benzyl-6-chloro-N-
methylpyrazin-2-amine and 2-methoxy-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-
yl)phenol furnished the product (51%).
1H-n.mr. (CDCl3) d 3.20 (br 6,3H, NCH,), 3.91 (s, 3H, OCH3), 4.89 (s, 2H, CH2),
5.83 (br s, 1H, OH), 6.98 (d, 1H, J= 8.1 Hz, ArH), 7.27 (m, SH, Ar-H), 7.53 (m, 2H,
ArH), 7.93 (s, 1H, pyraz.-H), 6.28 (s, 1H, pyraz.-H).
m/z(ES)322(M++H).
Example 25

In a procedure analogous to Example 21, reaction of tetrahydroisoquinoline,
and 2,6-dichloropyrazine furnished the product (95%).
1H-n.m.r. (CDCl3) d2.99 (t J= 5.7 Hz, 2H, ArCH2CH2N), 3.86 (t, J= 5.7 Hz, 2H,
ArCH2CH2N), 4.73 (s, 2H, ArCH2N), 7.27-7.19 (m, 4H ArH), 7.82 (s, 1H, pyraz.-H),
8.01 (s, 1H, pyraz.-H).
Example 26

In a procedure analogous to Example 22, reaction of 2-(6-chloropyrazin-2-yl)-
1,2,3,4-tetrahydroisoquinoline and 2-methoxy-4-(4,4,5,5-tetramethyyl-1,3,2-
dioxaborolart-2-yl)phenol furnished the product (44%).
1H-n.m.r. (CDCl3,) d 3.03 (m, 2H, CHj), 3.96 (m, 2H, CH2), 4.01 (s, 3H, OCH3),
4.83 (s, 2H, CH2), 5.S7 (br s, 1H, OH), 7.04 (m, 1H, ArH), 7.21 (m, 3H, Ar-H), 7.56 (nv
2H, ArH), 8.07 (br s, 1H, pyraz.-H), 8.28 (br s, 1H, pyraz.-H).
m/z(ES)374(M+H+K)+.
Example 27

In a procedure analogous to Example 21, reaction of 3,4-dichlorooenzylamine
and 2,6-dichloropyrazme furnished the product (89%).
1H-n.m.r. (CDCl3) d4.55 (d, J= 6 Hz, 2H, NCH,), 5.01 (s, broad, 1H, NH), 7.18
(dd,J= 2.1,2.1 Hz, 1H, ArH), 720 (dd,J= 2.1, Zl Hz, 1H, ArH), 7.45-7.41 {m, 2H,
ArH), 7.77 (s, 1H, pyraz.-H), 7.86 (s, 1H, pyraz.-H).
Example 28

In a procedure analogous to Example 22, reaction of 6-chloro-N-(3,4-
dichlorobenzyl)pyrazin-2-amine and 2-methoxy-4-(4,4,5,5-tetramethyl-1,3,2-
dioxaborolan-2-y])phenol furnished the product (57%).
1'H-n.m.r. (CDCl3) d 3.93 (s, 3H, CH3), 4-62 (d, 2H, J= 6.0 Hz, CH2), 5.01 (br s,
1H, NH), 5.79 (br s, 1H, OH), 6.98 (d, 1H, J= 8.7 Hz, ArH), 7.45 (m, 4H, ArH), 7.68 (m,
1H, ArH), 7.95 (s, 1H, pyraz.-H), 8.29 (s, 1H, pyraz.-H).
m/z(ES)376(M+).
Example 29

In a procedure analogous to Example 21, reaction of 3,5-dimethoxybenzylamine
and 2,6-dichloropyrazine furnished the product (91%).
1H-n.m.r. (CDCl3) d 3.78 (s, 6H, OCH3), 4.49 (d, J= 5.4 Hz, 2H, NCH3), 5.12 (br s,
1H, NH), 6.39 (t, J= 2.1 Hz, 1H, ArH), 6.50 (d, J= 2.1 Hz, 2H, ArH), 7.75 (s, 1H pyraz.-
H),7.82{s, 1H,pyraz.-H).
Example 30

In a procedure analogous to Example 22, reaction of 6-chloro-N-(3,5-
dimeihoxybeirzyl)pyrazin-2-amine and 2-methoxy-4(4,4,5,5-tetramethyl-1,3,2-
dioxaborolan-2-yl)phenol furnished the product (88%).
1H-n.m.r. (as mesylate salt) (d6-DM5O) d 239 (s, 3H, CH2SO3), 3.69 (s, 6H
OCH3), 3.80 (6,3H, OCH3), 4.51 (s, 2H. CH2), 6.36 (d, 1H, J= 2.1 Hz, ArH), 6.57 (d, 2H J
m 2.1 Hz, ArH), 6.83 (d, 1H, J= 8,1 Hz, ArH), 754 (xny 2H, ArH), 7.87 (s, 1H, pyraz.-H),
8.29(s,1H,pyraz.-H).
m/2(ES)368(M++H).
Example 31

In a procedure analogous to Example 21, reaction of furfurylamine and 2,6-
dichloropyrazine furnished the product (98%).
1H-n.m.r. (CDCl3) d4.57 (d, J= 5.7 Hz, 2H, NCH2), 5.01 (S, broad, 1H, NH), 6.30
(d, J= 3.3 Hz, 1H, furanyl-H), 6.35-6.33 (m, 2H, furanyl-H), 7.81 (s, 1H, pyraz.-H), 7.84
(s, 1H, pyraz.-H).
Example 32

In a procedure analogous to Example 2, reaction of 6-chloro-N-(2-
furyImethyl)pyrazin-2-amine and 2-methoxy-4-(4,4,5,5-tetramethyl-1,3,2-
dioxaborolan-2-yl)phenol furnished the product (92%).
1H-n.mr. (as mediate salt) (d6-DMSO) 5 238 (s, 3H, CH3SO3), 3.84 (s,3H,
OCH3), 459 (s, 2H, CH3), 6.33 (s, 1H, ArH), 6.38 (s, 1H, ArH), 6.87 (d, 2H, J= 8.1 Hz,
ArH), 752 (m, 3H, ArH), 7.86 (br s, 1H, pyraz.-H), 8.30 (br s, 1H, pyraz.-H).
m/z(ES)298(M++H).
Example 33

A solution of 4-bromo-2-chIorophenol (246mg, 1.18mmol),
bis(pinacolato)dicoron( 332mg, 1.3mmol), [1,1'-
bis(diphenylphosphino)ierrocenelpalladium(n) chloride (26mg, 0.035mmol) and
potassium acetate (222mg, 2-26mmol) in dry methanol (4mL) was degassed and
heated at 65°C for 24h. After cooling, the reaction mixture was diluted with ether and
filtered through Celite. The solvent was removed under reduced pressure and the
residue purified by chromatography using dichloromethane-hexane (90:10) as eluant.
The boronate thus obtained (50mg) was reacted with 6-chloro-N-[(1S)-1-
phenylethyl]pyrazin-2-amine (50mg, 02mmol) under conditions analogous to those of
example 2, to furnish the pure product after chromatography eluting with
dichloromethane-ether (90:10) (44mg, 68%).
1H-n.m.r. S 81.59 (d, 3H,/-6.0 Hz, CH,), 4.88 (m, 1H, CH), 5.08 (br s, 1H, NH),
5.69 (br s, 1H. NH), 7.07 (d. 1H, J= 8.5 Hz, ArH), 7.27-7.36 (m, 6H, Ar-H), 7.4S (d, 1H,
J= 1.5Hz, ArH), 7.62 (s, 1H, pyraz-H), 7.80 (s, 1H, pyraz-H).
Example 34

A mixture of 6-chloro-N-t(1S)-1-phenylethyl]pyrazin-2-amine (1.10g,
4.71mmol), 4-(4,45,5-tetramethyl-1,3,2-dioxaborolan-2-yl) aniline (1.10g, 5.02mmol),
(PPh3),Pd (580mg, 05mmol) and a Na2CQ, solution (2.6ml, 2M solution) in toluene (20
ml) was heated under reflux for 40 h. Upon cooling, the mixture was diluted with
water (30mL) and the product extracted with ethyl acetate (3 x 40ml). The organic
layers were combined, washed with brine (30ml), dried (Na2SO4)/ and the solvent
removed in vacuo. The residue was purified by column chromatography, eluting with
a hexane-ethyl acetate (2:3) to furnish the desired product from the polar fractions
(0.86g,63%).
1H-n.m.r. (CDC1,) 81.57 (d, 3H,J=- 62Hz, CH3), 3.80 (br s, 2H, NH2), 4.92-4.99
(rn, 2H, CH + NH), 6.69 (d, 2H J= 6.7H2, ArH), 7.21-7.40 (m, 5H, ArH), 7.72 (d, 2H, J=
6.7HZ, ArH), 7.57 (s, 1H pyraz.-H), 8.16 (s, 1H pyraz.-H).
m/z(ES)291(M++H)
Example 35

A solution of the amide (40mg, 0.12mmol) in THF (5mL) was treated with solid
LiAIH4 (38mg, 1mmol), and the mixture stirred at RT for 4h. The reaction was then
treated sequentially with HjO (5ml), 2M NaOH (5ml) and water (10ml) and the
resulting suspension then extracted with ethyl acetate (3 x 15ml). The combined
organic layers were dried (Na2SO4) and concentrated under reduced pressure. The
crude product thus obtained was purified by column chromatography using ethyl
acetate-hexane (3:1) as eluant to give the product as a colorless solid (22mg, 58%).
lH-n.m.r (CDCl3) 81.25 (t, 3H,J= 7.0Bz, CH3), 1.57 (d, 3H,J= 6.8Hz, CH3),
3.18 (q, 2H, J= 7.0HZ, CH2), 3.74 (br s. 1H, NH), 4.85-5.01 (m, 2H, CH + NH), 6.59-6.63
(m, 2H, ArH), 7.21-7.40 (m, 5H, ArH), 7.54 (s, 1H pyraz.-H), 7.73-7.77 (m, 2H, ArH),
8.16(s,1H,pyraz.-H).
m/z(E5)319(M++H)
Example 36

To a stirred solution of 6-(4-ammophenyl)N-[(15)-1-phenylethyl]pyrazin-2-
amine (58mg, 0.2mmol) in dry THF (3mL) was added triethylamine (70µL, 0.5mmol).
The solutions was cooled to 0°C and xnethanesulphonyl chloride (18.6µL, 0.24mmol)
was added dropwise. The mixture was allowed to warm to RT and stirred overnight
before dilution with water (15mL). The product was extracted into ethyl acetate (2 x
15mL) and the combined extracts washed with 10% aqueous NazCO3 and brine, and
then dried (Na2SO4). The solvent was removed under reduced pressure and the
product purified by chromatography eluting with ethyl acetate-hexane (3:2) to afford
the product as a pale yellow solid (54mg, 73%).
1H-n.m.r. (CDCl3) d1.59 (d, 3H, J= 6.2Hz, CH,), 3.01 (s, 3H, CH3), 4.96-5.01 (m,
2H, CH + NH), 6.52 (br s, 1H, NHSOJ, 722-7.40 (m, 7H, ArH), 7.70 (s, 1H, pyraz.-H),
7.85-7.89 (m, 2H, ArH), 8.20 (s, 1H, pyraz.-H).
m/z(ES)369(M++H).
Example 37

In a method analogous to that reported in example 39, reaction of 6-(4-
aminophenyl)-N-[(1S)-1-phenylethyl]pyrazin-2-amine (58mg, 0.2mmol) and
cyclopropanecarbonyl chloride (25mg, 0.24mmol) furnished the pure product after
chromatographic purification using ethyl acetate-hexane (3:2) (46mg, 64%).
1H-n.m.r. (CDCl3) d0.82-0.88 (m, 2H, CH2), 1.05-1.10 (m. 2H, CH2), 1.49-1.60 (m,
4H, CH, CH3), 4.91-4.9 (m,2H + NH), 7.23-7.40 (m, 5H, ArH), 7.56 (AA'XX', 2H.
ArH), 7.65 (s, 1H, pyraz.-H), 7.85 (AA'XX', 2H, ArH), 8.21 (s, 1H pyraz.-H).
m/z (ES) 359 (M++H).
Example 38

To a solution of hydroxylamine hydrochloride (3.44g) in water (20 ml) was
added NaOH (20 %, 30 ml). The ketone (5g, 41 mmol) was added at once and the
resulting mixture was stirred at RT until TLC showed no ketone remained. The
solvents were distilled off in vacuo and the residue extracted with CH2Cl2 (3 x 100 ml)
and dried (Na2SO4). After filtration and removal of the solvent the crude ketoxime
was recrystaliised from CH2Cl2/n-hexane.
1H-n.m.r. (CDCl3) d2.31 (s, 3H, CH3) 7.33 (dd, J= 4.8, 4-8 Hz, 1H, ArH), 7.97
(ddd, J= 8.1,1.8,1.8Hz, 1H, ArH), 8.61 (dd,/- 5.1,1.8 Hz, 1H, ArH), 8.96 (d, J=1.8
Hz, 1H, ArH), 10.62 (s, 1H, OH).
Example 39

A mixture of the ketone (2.0g, 13mmol), hydroxylamine hydrochloride (0.98g,
14mmol), NaOH (10%, 4ml), water (6.2ml) and EtOH (25 ml) was heated under reflux
for 2 hours- Upon cooling in ice, the ketoxime precipitated and was collected by
suction filtration. The crude product was recrystallised from CH2Cl2/n-hexane
(1.88g, 86%).
1H-n.m.r. (CDCl3) d 228 (s, 3H, CH3), 7.51 (s, 4H, ArH), 8.67 (s, 1H, OH).
Example 40

A mixture of the ketoxime (1g, 6mmol) and LiAIH4 (0.27g) in anhydrous THF
(100 ml) was heated at reflux under dry N2 overnight The reaction mixture was
cooled in ice-water and carefully quenched with H2O (60ml). The mixture was
allowed to stir at RT (or half an hour, after which time it was filtered through Celite®.
The inorganic salts were washed with EtOAc (3 x 100 ml). The filtrate was
concentrated under reduced pressure, diluted with 2M HCl (50ml) and the aqueous
phase washed with Et2O (2 x 70ml). The aqueous phase was basified with 40%
aqueous NaOH and the product extracted with Et2O (3 x 50ml). The combined organic
layers were washed with brine (50ml) and dried (MgSO4). The solvents were removed
in vacuo to afford the pure amine (0.65g, 71%).
1H-n.m.r. (CDCl3) d1.38 (d, J= 6.6 Hz, 3H, CH-CH3), 1.63 (br s,2H NH2), 4.13-
4.06 (m, 1H, CH-CH3), 7.23-7.18 (m, 3 H, ArH), 7.35 (s, 1H, ArH).
Example 41

To a mixture of the ketoxime (4.S5g, 36 mmol) and Zn powder (12g) at 0°C was
slowly added, with vigorous stirring, concentrated HC1 (50 ml). When the initial
vigorous reaction had subsided, the mixture was heated under reflux until TLC
showed all the ketoxime had been consumed. After cooling to RT, the strongly acidic
mixture was extracted with CH2Cl2 (2 x 75 ml). The reaction mixture was then made
strongly basic with 50% KOH solution. After removal of the solvent the residue was
extracted with boiling MeOH (4 x 100 ml). The MeOH was distilled off to leave the
crude amine which was used in the ensuing reactions without further purification.
1H-n.m.r. (CDCl3) d1.07 {d, J= 6.6 Hz, 3 H, CH3), 137 (br s, 2H, NHj), 3.84 (q, J=
4.6Hz, 1H, CH-CH3), 6.93 (dd, J= 7.8,48 Yiz, 1H, ArH), 7.38 (ddd, J= 7.8,2.1,1.5 Hz,
1H, ArH), 8.15 (dd,J= 4.8,15 Hz, 1H, ArH), 8.27 (d, J= 2.1 Hz, 1H, ArH).
SCREENING
Establishment of TEL:JAK cell lines
The coding region encompassing nudeotides 1-487 of TELwas amplified by
PCR using the oligonudeotides 5TEL (5' -GGA GGA TCC TGA TCT CTC TCG CTG
TGA GAC-3') and 3TEL (5'-AGGC GTCGAC TTC TTC TTC ATG GTTCTG-3') and
U937 mRNA as template. A BamH I site was present into the 5TEL Primer, a Sal I site
was incorporated into the 3TEL primer. The regions encompassing the kinase domains
of JAK2 (nudeotides 2994-3914;; JAK2F 5VACGC GTC GAC GGT GCC TTT GAA
GAC CGG GAT-3'; JAK2R 5'-ATA GTT TAG CGGCCG CTC AGA ATG AAG GTC
ATT T-3') and JAK3 (nudeotides 2520-3469; JAK3F 5'-GAA GTCGAC TAT GCC
TGC CAA GAC CCC ACG ATC TT-3' ; JAK3R 5'-GGA TCTAGA CTA TGA AAA
GGA CAG GGA GTG GTG TTT -3' ) were generated by PCR using Taq DNA
Polymerase (Gibco/BRL) and U937 mRNA as template. A Sall site was incorporated
into the forward primer of JAK2 and JAK3, a Not I site was incorporated into the
JAK2 reverse primer and a Xba I site was added to the reverse primer of JAK3.
A TEL/Jak2 fusion was generated by digestion of the TELPCR product with
BamH I /Sal I, digestion of the JAK2 PCR product with Sal 1/ Not I followed by
ligation and subdontng into the mammalian expression Vector pTRE 2 (Clontedi)
digested with BamH I-Not I (pTELJAK2). For JAK3 Sal 1/ Not I deaved kinase domain
PCR product was ligated with BamH I /Sal I cleaved TELproduct followed by ligation
into BamH I/Not I deaved pTRE2 (pTELJAKS).
The growth factor dependent myelomonocytic cell line BaF3 bearing the pTET-
of ff plasmid (Clontech) was transfected with either pTELJAK2 or pTELJAK3 and me
cells selected for factor independent growth. BaF 3 wild type cells were cultured in
DMEM10% FCS, 10% WEHI3B conditioned medium. BaF3 TELJAK cells were
cultured in DMEM 10% Tet-System Approved FBS (without WEHI 3B conditioned
medium).
Cellular assays were performed as follows:
Cell suspensions were prepared by harvesting cells from culture. (Cells used in this
test should be in later log phase growth and high viability.) Cells were diluted in correct
growth medium to l.lx final concentration (from 50000 cell/mL to 200,000 cell/mL,
depending on cell line).
Compounds to be tested were added (10µL, 10X final concentration) to a flat bottom
96-we!l plate. The cellular suspension (90µL per well) was added, and the plate incubated
for 40 hr at 37 °C, 5% CO2. MTT (20 µL per well, 5mg/mL in PBS) was added and the
plates were returned to the incubator for a further 6 hours. Lysis buffer (100 µL per well,
10% SDS, 0.01N HC1) was added and the plate stored in the incubator overnight. The plate
was then read at 590 nm.
Kinase assays were performed either in a 96 well capture based ELISA assayor in 384 well
Optiplates (Packard) using an Alphascrecn Protein Tyrosine Kinase kit. In either casse
using approximately 1.5 mg of affinity purified PTK domain in the presence of 50mM
HEPES,pH 7.5, 10mMMgC12, I50mMNaCI and lOmM-1mM ATP. The biotimylated
substrate biotin?EGPWLEEEEEAYGWMDF?NH2 (final concentration 5mM) was used as
substrate. In the ELISA assay lynxsine phosphorylation was quanlitated following transfer to
an avidin coated ELISA plate using pcroxidase linked anti-phospho-tyrosinc antibody
PY20. In the Alphascreen assay, Alphascreen phosphotyrosine acceptor beads followed by
streplavidin donor beads were added under subdued light. The ELISA plates were read on a
RM.G Fluorostar, the Alphascreen plates were read on a Packard Fusion Alpha. Inhibitors
were added to the assays fifteen minutes prior to the addition of ATP. Inhibitors were added
in aqueous DMSO, with DMSO concentrations never exceeding 1%."
Results
The activity of a range of compounds is shown in Table 1. Compounds that
exhibited a capacity to inhibit 50% of cell growth at a concentration of 50µM (measured
under standard conditions, see Methods), are designated as "+". In Table 1 a reference to
"NT" means "not tested,"
It will be appreciated by persons skilled in the art that numerous variations
and/ or modifications may be made to the invention as shown in the specific
embodiments without departing from the spirit or scope of the invention as broadly
described. The present embodiments are, therefore, to be considered in all respects as
illustrative and not restrictive.
Table 1: 2-amino-6-carba-disubstituted pyrazine and 2-amino-6-carba-
disubstituted pyridine possessing growth inhibitory activity (>50%) in transformed
cell lines (Tel-Jak2 and Tel-Jak3) at 50µM

References
Spiotto MT, and Chung TD. (2000) STAT3 mediates IL-6-induced growth inhibition in
the human prostate cancer cell line LNCaP. Prostate42 88-98
WE CLAIM:
1. A 6-amino-2-phenyl substituted pyrazine compound of the general formula

or phatrnaceutically acceptable salts, hydrates, solvates, crystal forms or
diastereomers thereof, wherein:
R1 is H,C1-4 alkyl
Q is a bond, or C1-4 alkyl
A is atryl, fietaryl optionally substituted with 0-3 substiluents independently chosen
from halogen. C1-4 alkyl, CH2F, CHF2, CF3, CN7 aryl, hctaryl, OCR,, OC1-4alkyl,
OC2-5 alkylNR4R5, Oaryl, Ohetaryl, CO2R4, CONR4R5, nitro, NR4R5, C1-4
alkylNR4R5, NR6C1-4 alkylNR4R5, NR4COR5, NR6CONR4R5, NR4SO2R5; and
R4, R5 ace each independently H, C1-4 alkyl, C1-4 alky! cycloalkyl, C1-4 alkyl
cycloheralkyl, aryl, hetaryl, C1-4alkyl aryl, C1-4 alkyl hctaryl, or may be joined to
form an optionally substituted 3-8 membered ring optionally containing an atom
selected from O, S, NR7; and R6 is selected from H, C1-4 alkyl; and R7 is selected
from H, C1-4. alkyl, aryl, hctaryl, C1-4 alky] aryl C1-4 alkyl hetaryl.
R2 is 0-2 substiments independently selected from halogen, C1-4alkyl, OH, OC1-4
alkyl, CH2F. CI-IF2, CF3) OCF3, CN, C1-4alkylNR8R9, OC1-4alkylNR8R9, CO2RS,
CONR8R9, NR8R9, NR8COR9, NR10CONR8R9, NR8SOZR9; and R8, R9 are each
independentlyH. C1-4 alkyl. C1-4 alkyl cycloalkyl, C1-4. alkyl cyclohetalkyl, aryl,
hciaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl, or may be joined to form an optionally
substituted 3-8 membered ring, option ally containing an atom selected from O, S,
NR11; and R10 is selected from H, C1-4 alky], aryl or heuiryl; and R11 is selected
from II, C1-4 alkyl, aryl, hetaryl, C1-4 alky! aryl, C1-4 alky] hetaryl.
Y is halogen, OH, NR12R13, NR12COR13, NR12CONR13, N12SO2R13; and R12.
and RI3 are each independently II, CH3F, CHF2 cycloalkyl, C1-4 alky] cyclohetalkyl, or may be joined to form an optionally
substituted 3-6 membcrcd ring optionally containing an atom selected from O, S.
NR14 and R14 is selected from H, C1-4 alkyl.
n = 0-4
W is selected from H, C1-4alkyl, C2-6alkenyl; where C1-4alky 1 or C2-6alkenyl may be
optionally substituted with C1-4alkyl, OH, OC1-4allkyl, NRI5R16; and R15, and RI6
are each independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1-4 alkyl cyclohetalkyl,
or may be joined to form an optionally substituted 3-8 membered ring optionally
containing an atom selected from O, S, NR17 and R17 is selected from H, C1-4 alkyl;
wherein when Y is OH or NHCOCH3 then R2 is 1-2 substituents and wherein when
Y is NH2 and R2 is absent then Y is in the para position,
2. A 6-amino-2-phenyl substituted pyrazine compound as claimed in claim 1 selected
from compounds of the general formula II

or pharmaccutically acceptable salts, hydrates, solvates, crystal forms or
diastereomers thereof, wherein:
R1 is H, C1-4 alkyl
A is aryl, hetaryl optionally substituted with 0-3 substituents independently chosen
from halogen, C1-4 alkyl, CH2F, CHF2, CF3, CN. aryl. hetaryl, OCF3, OC1-4alkyl,
OC2-5alkylNR4R5, Oaryl, Ohetaryl, CO2R4, CONR4R5, NR4R5, C1-4 alkylNR4R5,
NR6C1-4alkylNR.4RS, NR4COR5, NR6CONR4R5, NR4SO2R5-, and R4, R5 are each
independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1.4 alkyl cyclohetalkyl, aryl,
hctaryl, C1-4alkyl aryl, C1-4 alkyl hetaryl, or may be joined to form an optionally
substituted 3-8 membcrcd ring optionally containing an atom selected from O, S,
NR7; and R6 is selected from H, C1-4 alkyl; and R7 is selected from H, C1-4 alkyl,
aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl.
R2 is 0-2 substituents independently selected from halogen, C1-4alk.yl, OH, OC1-4
alkyl, CH2F, CHF2, CF3, OCF3, CN, C1-4alkylNR8R9, OC,.(alkyINR8R9, CO2R8,
CONR8R9, NR8R9, NR8COR9, NR10CONR8R9, NR8SO2R9; and R8, R9 are each
independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1-4 alkyl cyclohetalkyl, aryl,
hetaryi, C1-4 alkyl aryl, C1-4 alkyl hetaryi, or may be joined to form an optionally
substituted 3-8 membered ring optionally containing an atom selected from O, S,
NR11; and RiO is selected from H, C1-4 alkyl, aryl or hetaryl; and R11 is selected
from H, C1-4 alkyl, aryl, hetaryl, C1-4 alkyl aryl, C1-4 alkyl hetaryl.
Y is halogen, OII, NRI2R13, NR12COR13, NR.12CONR13, N12SO2R13; and R12,
and R13 arc each independently H, CII2F, CHF2, CF3, CN, C1.4 alkyl, C1-4 alkyl
cycloalkyl, C1-4 alkyl cyclohctalkyl, or may be joined to form an optionally
substituted 3-6 membered ring optionally containing an atom selected from O, S,
NR14 and R14 is selected from H, C1-4 alkyl.
n = 0-4
W is selected from H7 C1-4alkyl, C2-6alkenyl; where C1-4alkyl or C2-6 alkenyl may be
optionally substituted wilh C1-4alkyl, Oil, OC1-4alkyl, NRI5R16; andR15, andR16
are each independently H, C1-4 alkyl, C1-4 alkyl cycloalkyl, C1-4 alkyl cyclohetalkyl,
ot may be joined to form an optionally substituted 3-8 membered ring, optionally
containing an atom selected from O, S, NR17 and R17 is selected from H, C1-4 alkyl;
wherein when Y is OH or NHCOCH3 then R2 is 1-2 substitucms and wherein when
Y is NH2 and R2 is absent then Y is in the para position.
3. A compound as claimed in claim 1 or claim 2 where W is C1-4 alkyl wherein the
compound possesses S chirality at the chiral carbon bearing W.
4. A compound as claimed in claim 3 wherein the compound is a mixture of Rand S
isomers and the mixture comprises at least 70% of the S isomer..
5. A compound as claimed in claim 4 wherein the compound comprises at least 80%
of the S isomer
6. A compound as claimed in claim 4 wherein the compound comprises at least 90%
of the S isomer
7. A compound as claimed in claim 4 wherein the compound comprises at least 95%
of the S isomer
8. A compound as claimed in claim 4 wherein the compound comprises at least 99%
of the S isomer.
9. A compound as claimed in claim 1 wherein the compound is selected from the
group consisting of:

10. A composition comprising a carrier and at least one compound as claimed in
anyone of claims Ho 9 .
11. A pharmaceutical composition comprising at least one of the compounds as
claimed in anyone of claims 1 to 9 capable of treating a protein kinase-associated
disease state in an amount effective therefor, and a pharmaceutically acceptable
vehicle or diluent.
12. A pharmaceutical composition as claimed in claim 11, wherein the disease state
involves a receptor tyrosine kinase selected from the group consisting of EGF,
HER2, HER3, HER4, IR, 1GF-IR, IRR, PDGFR.alpha., PDGFR.beta., CSFIR, C-
Kit, C-fms, Flk-IR, Flk4, KDR/Flk-1, FIt-I, FGFR-IR, FGFR-2R, FGFR-3R and
FGFR-4R.
13. A pharmaceutical composition as claimed in claim 11, wherein the disease state
involves a cellular tyrosine kinase selected from the group consisting of Src, Frk.
Btk, Csk, Abl, ZAP70, Fes/Fps, Fak, Ack, Yes. Fyn, Lyn, Lck, Blk. Hck, Fgr and
Yrk
14. A pharmaceutical composition as claimed in claim 11, wherein the disease state
involves a tyrosine kinase selected from the group consisting of JAK1, JAK2,
JAK3 andTYK2.
15. A pharmaceutical composition as claimed in claim 11, wherein the disease state
involves a serine/threonine kinase selected from the group consisting of ERK2. c-
Jun, p38MAPK, PKA, PKB, PKC, a cyclin-dependent kinase, CDK 1, CDK2.
CDK3, CDK4, CDK5, CDK6, CDK7, CDK8, CDK9, CDKIO, and CDK11.
16. - A pharmaceutical composition as claimed in claim 11, wherein the disease state is
selected from the group consisling of Atopy, such as Allergic Asthma, Atopic
Dermatitis (Eczema), and Allergic Rhinitis; Cell Mediated Hypersensitivily, such as
Afiergic Contact Dermatitis and Hypersensiti vity "Pneumonitis; Rheumatic Diseases,
such as Systemic Lupus Erythematosus (SLE), Rheumatoid Arthritis, Juvenile
Arthritis, Sjogren's Syndrome. Scleroderma, Polymyositis, Ankylosing Spondylitis,
Psoriatic Arthritis: Other autoimmune diseases such as Type I diabetes, autoimmune
thyroid disorders, and Alzheimer's disease; Viral Diseases, such as Epstein Barr
Virus (EBV), Hepatitis B, Hepatitis C, ILLV, HTLV 1, Varicella-ZosLer Virus
(VZV), Human Papilloma Vims (HPV), Cancer, sueh as Leukemia, Lymphoma and
Prostate Cancer.
17. A pharmaceutical composition as claimed in claim 11, wherein the protein kinase-
associated disease state is selected from the group consisting of one or more of
sarcomas, carcinomas and leukemias.
18 . A pharmaceutical composition as claimed in claim 17, wherein the protein kinase-
associated disease state is selected from the group consisting of one or more of
fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma,
chordoma, angiosarcoma, endothcliosarcormt, lymphangiosarcoma,
lymphangiocndotheliosarcoma, synovioma, rnesothclioma, Ewing's tumor,
Ieiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast
cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell
carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma,
papillary carcinoma, papillary adenocarcinomas, cystadenocardnoma, medullary
carcinoma, broncliogenic carcinoma, renal cell carcinoma, hepatoma, bile duct
carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor,
cervical cancer, tcsticular tumor, lung carcinoma, small cell lung carcinoma, bladder
carcinoma, epithelial carcinoma, glioma, astrooytoma, medulloblastoma,
craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic
neuroma, oligodendroglioma, meningioma, melanoma, neuroblastoma, and
rctinoblastoma.
19. A pharmaceutical composition as claimed in claim 11, wherein the protein kinase-
associated disease Slate is a carcinoma formed from the tissue of the breast,
prostate, kidney, bladder or colon,
20. A pharmaceutical composition as claimed in claim 11, wherein the protein kinase
associated disease state is a hyperplastic or neoplastic disorder arising in adipose
tissue.
21. A pharmaceutical composition as claimed in claim 20, wherein the hyperplastic or
neoplastic disorder is an adipose cell tumour.
22. A pharmaceutical composition as claimed in claim 21, wherein the adipose cell
tumour is one or more of lipoma, fibrolipoma, lipoblastoma, lipomatosis,
hibemoma, hemangioma and liposarcoma
A 6-amino-2-phenyl substituted pyrazine compound of the general formula:
or pharmaceutically acceptable salts, hydrates, solvates, crystal forms of diastereomers
thereof is described.

Documents:

1738-KOLNP-2004-CORRESPONDENCE.pdf

1738-KOLNP-2004-FORM 27-1.1.pdf

1738-KOLNP-2004-FORM 27.pdf

1738-kolnp-2004-granted-abstract.pdf

1738-kolnp-2004-granted-assignment.pdf

1738-kolnp-2004-granted-claims.pdf

1738-kolnp-2004-granted-correspondence.pdf

1738-kolnp-2004-granted-description (complete).pdf

1738-kolnp-2004-granted-drawings.pdf

1738-kolnp-2004-granted-examination report.pdf

1738-kolnp-2004-granted-form 1.pdf

1738-kolnp-2004-granted-form 18.pdf

1738-kolnp-2004-granted-form 3.pdf

1738-kolnp-2004-granted-form 5.pdf

1738-kolnp-2004-granted-pa.pdf

1738-kolnp-2004-granted-reply to examination report.pdf

1738-kolnp-2004-granted-specification.pdf

1738-KOLNP-2004-OTHER PATENT DOCUMENT.pdf


Patent Number 222897
Indian Patent Application Number 1738/KOLNP/2004
PG Journal Number 35/2008
Publication Date 29-Aug-2008
Grant Date 27-Aug-2008
Date of Filing 17-Nov-2004
Name of Patentee CYTOPIA PTY LTD.
Applicant Address LEVEL 5, BAKER HEART RESEARCH INSTITUTE BUILDING, MELBOURNE, VICTORIA
Inventors:
# Inventor's Name Inventor's Address
1 BURNS CHRISTOPHER JOHN 3 BROWNING STREET, SEDDON, VICTORIA 3011
2 BU XIANYONG 1 DAIRY COURT, POSANNA AUSTRALIAN EAST, VICTORIA 3084
3 WILKS ANDREW FREDERICK 6 MACFARLANE LANE, SOUTH YARRA, VICTORIA 3141
PCT International Classification Number C07D 241/20, 405/12
PCT International Application Number PCT/AU03/00629
PCT International Filing date 2003-05-23
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 PS 2515 2002-05-23 Australia