U.S. patent application number 10/464430 was filed with the patent office on 2004-03-11 for processes for preparing substituted pyrimidines.
Invention is credited to Charrier, Jean-Damien, Kay, David, Mazzei, Francesca, Miller, Andrew.
Application Number | 20040049032 10/464430 |
Document ID | / |
Family ID | 30003168 |
Filed Date | 2004-03-11 |
United States Patent
Application |
20040049032 |
Kind Code |
A1 |
Charrier, Jean-Damien ; et
al. |
March 11, 2004 |
Processes for preparing substituted pyrimidines
Abstract
The present invention provides a facile process for the
preparation of tri- and tetra-substituted pyrimidines. The process
is useful for preparing inhibitors of protein kinases, especially
Aurora kinase. These inhibitors are useful for treating or
lessening the severity of Aurora-mediated diseases or
conditions.
Inventors: |
Charrier, Jean-Damien;
(Grove Wantage, GB) ; Mazzei, Francesca;
(Abindgon, GB) ; Kay, David; (Wiltshire, GB)
; Miller, Andrew; (Upton, GB) |
Correspondence
Address: |
VERTEX PHARMACEUTICALS INC.
130 WAVERLY STREET
CAMBRIDGE
MA
02139-4242
US
|
Family ID: |
30003168 |
Appl. No.: |
10/464430 |
Filed: |
June 18, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60390658 |
Jun 20, 2002 |
|
|
|
60411609 |
Sep 18, 2002 |
|
|
|
Current U.S.
Class: |
544/60 ; 544/122;
544/295; 544/316; 544/323; 544/324 |
Current CPC
Class: |
A61P 1/16 20180101; A61P
9/00 20180101; C07D 453/02 20130101; A61P 5/00 20180101; A61P 19/00
20180101; A61P 25/18 20180101; A61P 37/08 20180101; C07D 403/12
20130101; A61P 37/04 20180101; A61P 11/06 20180101; C07D 401/14
20130101; A61P 35/00 20180101; A61P 37/06 20180101; A61P 43/00
20180101; A61P 37/00 20180101; C07D 403/14 20130101; A61P 7/00
20180101; A61P 19/08 20180101; A61P 3/00 20180101; A61P 35/02
20180101; A61P 25/00 20180101; A61P 29/00 20180101; A61P 25/28
20180101; A61P 13/08 20180101; C07D 239/34 20130101 |
Class at
Publication: |
544/060 ;
544/122; 544/295; 544/316; 544/323; 544/324 |
International
Class: |
C07D 417/02; C07D
413/02; C07D 43/02 |
Claims
We claim:
1. A method for preparing a compound of formula I: 200wherein: Q
and T are each independently selected from oxygen, sulfur or N(R);
each R is independently selected from hydrogen or an optionally
substituted C.sub.1-6 aliphatic group, wherein: two R bound to the
same nitrogen atom are optionally taken together with the nitrogen
to form an optionally substituted 3-7 membered monocyclic or 8-10
membered bicyclic saturated, partially unsaturated, or fully
unsaturated ring having 0-3 heteroatoms, in addition to the
nitrogen bound thereto, independently selected from nitrogen,
oxygen, or sulfur; R.sup.x is U-R.sup.5; R.sup.5 is selected from
halogen, NO.sub.2, CN, R, or Ar; each U is independently selected
from a valence bond or a C.sub.1-4 alkylidene chain, wherein: up to
two methylene units of U are optionally and independently replaced
by --O--, --S--, --SO--, --SO.sub.2--, --N(R)SO.sub.2--,
--SO.sub.2N(R)--, --N(R)--, --C(O)--, --CO.sub.2--, --N(R)C(O)--,
--N(R)C(O)O--, --N(R)CON(R)--, --N(R)SO.sub.2N(R)--, --N(R)N(R)--,
--C(O)N(R)--, --OC(O)N(R)--, --C(R).dbd.NN(R)--, or
--C(R).dbd.N--O--; each Ar is independently selected from an
optionally substituted ring selected from a 3-7 membered monocyclic
or an 8-10 membered bicyclic saturated, partially unsaturated, or
fully unsaturated ring having 0-4 heteroatoms independently
selected from nitrogen, oxygen, or sulfur; R.sup.y is
--N(R.sup.1).sub.2, --OR.sup.1, or --SR.sup.1; each R.sup.1 is
independently selected from R or a 3-8 membered monocyclic, an 8-10
membered bicyclic, or a 10-12 membered tricyclic saturated,
partially unsaturated, or fully unsaturated ring having 0-4
heteroatoms independently selected from nitrogen, oxygen, or
sulfur, and wherein: each R.sup.1 is optionally and independently
substituted by up to four substituents independently selected from
R.sup.2; each R.sup.2 is independently selected from --R.sup.3,
--OR.sup.3, --SR.sup.3, --CN, --NO.sub.2, oxo, halogen,
--N(R.sup.3).sub.2, --C(O)R.sup.3, --OC(O)R.sup.3,
--CO.sub.2R.sup.3, --SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or --NR.sup.3CO.sub.2(R.sup.3);
each R.sup.3 is independently selected from R or Ar; R.sup.z1 is
selected from a C.sub.1-6 aliphatic group or a 3-8 membered
monocyclic, an 8-10 membered bicyclic, or a 10-12 membered
tricyclic saturated, partially unsaturated, or fully unsaturated
ring having 0-4 heteroatoms independently selected from oxygen,
nitrogen or sulfur, wherein: R.sup.z1 is substituted with 0-4
independently selected R.sup.2 groups; R.sup.z2 is C.sub.1-6
aliphatic group or a 3-8 membered monocyclic or an 8-10 membered
bicyclic saturated, partially unsaturated, or fully unsaturated
ring having 0-4 heteroatoms independently selected from nitrogen,
oxygen or sulfur, wherein: R.sup.z2 is substituted by 0-4
substituents independently selected from oxo or U-R.sup.5; said
process comprising the step of combining a compound of formula II
and a compound of formula R.sup.y--H in a suitable medium:
201wherein: said suitable medium comprises: i) a suitable solvent;
and ii) optionally, a suitable base; and L.sup.3 is a suitable
leaving group.
2. The method according to claim 1, wherein said compound of
formula II is prepared by combining a compound of formula III with
a compound of formula R.sup.z1-Q-H in a suitable medium:
202wherein: said suitable medium comprises: i) a suitable solvent;
and ii) optionally, a suitable base; and L.sup.2 is a suitable
leaving group.
3. The method according to claim 2, wherein said compound of
formula III is prepared by combining a compound of formula IV with
a compound of formula R.sup.z2-T-H in a suitable medium:
203wherein: said suitable medium comprises: i) a suitable solvent;
and ii) optionally, a suitable base; and L.sup.1 is a suitable
leaving group.
4. The method according to claim 1, wherein L.sup.3 is selected
from halogen, optionally substituted arylsulfonyl, or optionally
substituted alkylsulphonyl.
5. The method according to claim 4, wherein L.sup.3 is fluoro,
chloro, bromo, iodo, paratoluenesulfonyl, methanesulfonyl,
paranitrophenylsulfonyl, parabromophenylsulfonyl, or
trifluoromethanesulfonate.
6. The method according to claim 5, wherein L.sup.3 is chloro or
iodo.
7. The method according to claim 2, wherein L.sup.2 is halogen,
optionally substituted arylsulfonyl, or optionally substituted
alkylsulphonyl.
8. The method according to claim 7, wherein L.sup.2 is fluoro,
chloro, bromo, iodo, paratoluenesulfonate, methanesulfonate,
paranitrophenylsulfonyl, parabromophenylsulfonyl, or
trifluoromethanesulfonate.
9. The method according to claim 8, wherein L.sup.2 is chloro or
Iodo.
10. The method according to claim 3, wherein L.sup.1 is halogen,
optionally substituted arylsulfonyl, or optionally substituted
alkylsulphonyl.
11. The method according to claim 10, wherein L.sup.1 is optionally
substituted alkylsulfonyl.
12. The method according to claim 11, wherein L.sup.1 is
methanesulfonyl.
13. The method according to claim 1, wherein Q is N(R).
14. The method according to claim 1, wherein T is oxygen or
sulfur.
15. The method according to claim 14, wherein T is sulfur.
16. The method according to claim 1, wherein R.sup.y is --OR.sup.1
or --N(R.sup.1).sub.2.
17. The method according to claim 16, wherein R.sup.y is
--N(R.sup.1).sub.2, and wherein: R.sup.1 is selected from R or a
3-7 membered monocyclic or an 8-10 membered bicyclic saturated,
partially unsaturated, or fully unsaturated ring having 0-4
heteroatoms independently selected from nitrogen, oxygen, or
sulfur, or: each R.sup.1 is R such that the two R on the same
nitrogen atom are taken together to form an optionally substituted
4-7 membered saturated ring having up to two additional heteroatoms
independently selected from nitrogen, oxygen, or sulfur, and
wherein: each R.sup.1 is optionally and independently substituted
by up to four substituents selected from --R.sup.3, --OR.sup.3,
--SR.sup.3, --CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2,
--C(O)R.sup.3, --OC(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.- 3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or
--NR.sup.3CO.sub.2(R.sup.3).
18. The method according to claim 17, wherein R.sup.y is
N(R.sup.1).sub.2, wherein: each R.sup.1 is independently selected
from R, wherein R is hydrogen or an optionally substituted
C.sub.1-4 aliphatic group.
19. The method according to claim 17, wherein R.sup.y is
N(R.sup.1).sub.2 wherein: each R.sup.1 is R such that the two R
groups are taken together to form an optionally substituted 4-7
membered saturated ring having up to two additional heteroatoms
independently selected from nitrogen, oxygen, or sulfur.
20. The method according to claim 19, wherein R.sup.y is selected
from pyrrolidin-1-yl, piperidin-1-yl, morpholin-4-yl,
thiomorpholin-4-yl, piperazin-1-yl, diazepanyl, or
tetrahydroisoquinolinyl, wherein each ring is optionally
substituted with one or two groups independently selected from
methyl, ethyl, methylsulfonyl, (CH.sub.2).sub.2SO.sub.2CH.sub.3,
cyclopropyl, CH.sub.2cyclopropyl, (CH.sub.2).sub.2OH,
CO.sub.2t-butyl, CH.sub.2phenyl, phenyl, NH.sub.2, NH(CH.sub.3),
N(CH.sub.3).sub.2, (CH.sub.2).sub.2NH.sub.2,
(CH.sub.2).sub.2morpholin-4-yl, (CH.sub.2).sub.2N(CH.sub.3).sub.2,
isopropyl, propyl, t-butyl, (CH.sub.2).sub.2CN, or
(CH.sub.2).sub.2C(O)morpholin-4-yl.
21. The method according to claim 1, wherein R.sup.z1 is a 3-7
membered monocyclic or an 8-10 membered bicyclic saturated,
partially unsaturated, or fully unsaturated ring having 0-4
heteroatoms independently selected from oxygen, nitrogen or sulfur,
wherein said ring is optionally and independently substituted by up
to three substituents selected from --R.sup.3, --OR.sup.3,
--SR.sup.3, --CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2,
--C(O)R.sup.3, --OC(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.- 3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or --NR.sup.3CO.sub.2R.sup.3.
22. The method according to claim 21, wherein R.sup.z1 is a 5-6
membered fully unsaturated ring having 1-3 heteroatoms
independently selected from nitrogen, oxygen, or sulfur, wherein
said ring is optionally and independently substituted by up to
three substituents selected from --R.sup.3, --OR.sup.3, --SR.sup.3,
--CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2, --C(O)R.sup.3,
--OC(O)R.sup.3, --CO.sub.2R.sup.3, --SO.sub.2R.sup.3,
--SO.sub.2N(R.sup.3).sub.2, --N(R.sup.3)SO.sub.2R.sup.- 3,
--C(O)NR(R.sup.3), --C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or --NR.sup.3CO.sub.2R.sup.3.
23. The method according to claim 22, wherein R.sup.z1 is an
optionally substituted ring selected from pyrazole or any one of
the following 5-6 membered rings: 204
24. The method according to claim 23, wherein R.sup.z1 is a
pyrazole ring having up to two substituents independently selected
from --N(R.sup.3).sub.2, --OR.sup.3, or a C.sub.1-C.sub.4 aliphatic
group.
25. The method according to claim 24, wherein R.sup.z1 is a
pyrazole optionally substituted with one substituent selected from
methyl, ethyl, propyl, isopropyl, t-butyl, cyclopropyl, or
phenyl.
26. The method according to claim 1, wherein R.sup.z2 is an
optionally substituted ring selected from a 5-6 membered monocyclic
or an 8-10 membered bicyclic saturated, partially unsaturated, or
fully unsaturated ring having 0-4 heteroatoms independently
selected from nitrogen, oxygen or sulfur, wherein said ring is
optionally substituted by up to three substituents independently
selected from halogen, --CN, --NO.sub.2, --C(O)R.sup.3,
--CO.sub.2R.sup.3, --C(O)NR(R.sup.3), --NR.sup.3C(O)R.sup.3,
--N(R.sup.3).sub.2, --N(R.sup.3)SO.sub.2R.sup.3,
--NR.sup.3C(O)N(R.sup.3).sub.2, or --NR.sup.3CO.sub.2R.sup.3.
27. The method according to claim 26, wherein: R.sup.z2 is selected
from phenyl, imidazolyl, pyrazolyl, pyridyl, pyridazinyl,
pyrazinyl, naphthyl, tetrahydronaphthyl, benzimidazolyl,
benzthiazolyl, quinolinyl, quinazolinyl, benzodioxinyl,
isobenzofuran, indanyl, indolyl, indolinyl, indazolyl, or
isoquinolinyl, wherein: R.sup.z2 is optionally substituted with up
to three substituents independently selected from --Cl, --Br, --F,
--CN, --CF.sub.3, --COOH, --CONHMe, --CONHEt, --NH.sub.2, --NHAc,
--NHSO.sub.2Me, --NHSO.sub.2Et, --NHSO.sub.2(n-propyl),
--NHSO.sub.2(isopropyl), --NHCOEt, --NHCOCH.sub.2NHCH.sub.3,
--NHCOCH.sub.2N(CO.sub.2t-Bu)CH.sub.3,
--NHCOCH.sub.2N(CH.sub.3).sub.2,
--NHCOCH.sub.2CH.sub.2N(CH.sub.3).sub.2,
--NHCOCH.sub.2CH.sub.2CH.sub.2N(- CH.sub.3).sub.2,
--NHCO(cyclopropyl), --NHCO(isopropyl), --NHCO(isobutyl),
--NHCOCH.sub.2(morpholin-4-yl),
--NHCOCH.sub.2CH.sub.2(morpholin-4-yl),
--NHCOCH.sub.2CH.sub.2CH.sub.2(morpholin-4-yl),
--NHCO.sub.2(t-butyl), --NH(cyclohexyl), --NHMe, --NMe.sub.2, --OH,
--OMe, methyl, ethyl, cyclopropyl, isopropyl, or t-butyl.
28. The method according to claim 27, wherein R.sup.z2 has one
substituent selected from --NR.sup.3C(O)R.sup.3, wherein: each
R.sup.3 is independently selected from R or Ar, and wherein: R is
hydrogen or an optionally substituted C.sub.1-4 aliphatic
group.
29. The method according to claim 1 wherein said suitable solvent
is a protic solvent, a halogenated hydrocarbon, an ether, an
aromatic hydrocarbon, a polar or a non-polar aprotic solvent, or
any mixtures thereof.
30. The method according to claim 29, wherein said solvent is a
C.sub.1-5 straight or branched alkyl alcohol, ether, or a polar or
non-polar aprotic solvent.
31. The method according to claim 1 wherein said suitable base is
selected from an organic amine, an alkaline earth metal carbonate,
an alkaline earth metal hydride, or an alkaline earth metal
hydroxide.
32. The method according to claim 31, wherein said suitable base is
selected from a trialkyl amine, sodium carbonate, potassium
carbonate, sodium hydride, potassium hydride, sodium hydroxide, or
potassium hydroxide.
33. The method according to claim 1, wherein said method is used to
prepare a compound selected from the following Table 1 and Table 2
compounds:
6TABLE 1 No. V- Structure 1 205 2 206 3 207 4 208 5 209 6 210 7 211
8 212 9 213 10 214 11 215 12 216 13 217 14 218 15 219 16 220 17 221
18 222 19 223 20 224
7TABLE 2 225 I-1 226 I-2 227 I-3 228 I-4 229 I-5 230 I-6 231 I-7
232 I-8 233 I-9 234 I-10 235 I-11 236 I-12 237 I-13 238 I-14 239
I-15 240 I-16 241 I-17 242 I-18 243 I-19 244 I-20 245 I-21 246 I-22
247 I-23 248 I-24 249 I-25 250 I-26 251 I-27 252 I-28 253 I-29 254
I-30 255 I-31 256 I-32 257 I-33 258 I-34 259 I-35 260 I-36 261 I-37
262 I-38 263 I-39 264 I-40 265 I-41 266 I-42 267 I-43 268 I-44 269
I-45 270 I-46 271 I-47 272 I-48 273 I-49 274 I-50 275 I-51 276 I-52
277 I-53 278 I-54 279 I-55 280 I-56 281 I-57 282 I-58 283 I-59 284
I-60 and 285 I-61.
34. A compound of formula V: 286or a pharmaceutically acceptable
derivative or salt thereof, wherein: R.sup.5 is selected from
hydrogen or C.sub.1-4 aliphatic; R.sup.6 is selected from C.sub.1-3
aliphatic; and R.sup.7 is selected from Cab aliphatic.
35. The compound according to claim 34, wherein R.sup.5 is selected
from hydrogen, methyl, ethyl, t-butyl, or isopropyl.
36. The compound according to claim 35, wherein R.sup.6 is selected
from methyl, ethyl, or cyclopropyl.
37. The compound according to claim 36, wherein R.sup.7 is selected
from methyl, ethyl, t-butyl, or cyclopropyl.
38. A compound selected from the following Table 1 compounds:
8TABLE 1 No. V- Structure 1 287 2 288 3 289 4 290 5 291 6 292 7 293
8 294 9 295 10 296 11 297 12 298 13 299 14 300 15 301 16 302 17 303
18 304 19 305 20 306
39. A compound selected from the following Table 3 compounds:
9TABLE 3 V-1 i 307 V-1 ii 308 V-1 iii 309 V-1 iv 310 V-1 v 311 V-1
vi 312 V-1 vii 313 V-1 viii 314 V-1 ix 315 V-1 x 316 V-1 xi 317
V-20 i 318
40. A composition comprising a compound according to claim 34, and
a pharmaceutically acceptable carrier, adjuvant, or vehicle.
41. The composition according to claim 40, additionally comprising
an anti-proliferative agent or a chemotherapeutic agent.
42. A method of inhibiting Aurora-1 in a biological sample,
comprising contacting said sample with: (a) a compound according to
claim 34; or (b) a composition according to claim 40.
43. A method of inhibiting Aurora-2 in a biological sample,
comprising contacting said sample with: (a) a compound according to
claim 34; or (b) a composition according to claim 40.
44. A method of inhibiting Aurora-3 in a biological sample,
comprising contacting said sample with: (a) a compound according to
claim 34; or (b) a composition according to claim 40.
45. A method of inhibiting FLT-3 in a biological sample, comprising
contacting said sample with: (a) a compound according to claim 34;
or (b) a composition according to claim 40.
46. A method of inhibiting Aurora-1 in a patient, comprising
administering to said patient: (a) a compound according to claim
34; or (b) a composition according to claim 40.
47. A method of inhibiting Aurora-2 in a patient, comprising
administering to said patient: (a) a compound according to claim
34; or (b) a composition according to claim 40.
48. A method of inhibiting Aurora-3 in a patient, comprising
administering to said patient: (a) a compound according to claim
34; or (b) a composition according to claim 40.
49. A method of inhibiting FLT-3 in a patient, comprising
administering to said patient: (a) a compound according to claim
34; or (b) a composition according to claim 40.
50. A method of inhibiting Aurora-1, Aurora-2, Aurora-3, and FLT-3
in a patient, comprising administering to said patient: (a) a
compound according to claim 34; or (b) a composition according to
claim 40.
51. A method of treating cancer in a patient comprising the step of
administering to said patient a composition according to claim
40.
52. The method according to claim 51 comprising the step of
administering to said patient an additional chemotherapeutic or
anti-proliferative agent.
53. The method according to claim 51, wherein said cancer is
selected from melanoma, lymphoma, neuroblastoma, leukemia, or a
cancer selected from colon, breast, lung, kidney, ovary,
pancreatic, renal, CNS, cervical, prostate, or cancer of the
gastric tract.
54. The method according to claim 51, wherein said cancer is
selected from acute-myelogenous leukemia (AML), acute lymphocytic
leukemia (ALL), mastocytosis or gastrointestinal stromal tumor
(GIST).
55. A method of treating or lessening the severity of a cancer in a
patient comprising the step of disrupting mitosis of the cancer
cells by inhibiting Aurora protein kinase with a compound according
to claim 34.
56. The method according to claim 55, comprising the step of
administering to said patient a composition according to claim 40.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to U.S. Provisional Patent
Application No. 60/390,658 filed Jun. 20, 2002 and U.S. Provisional
Patent Application No. 60/411,609 filed Sep. 18, 2002, the contents
of which are incorporated herein by reference.
FIELD OF THE INVENTION
[0002] The present invention provides a facile process for the
preparation of substituted pyrimidines. The process is useful for
preparing inhibitors of protein kinases, especially of FLT-3 and
the Aurora-family kinases, serine/threonine protein kinases. The
present invention also relates to inhibitors of FLT-3, Aurora-1,
Aurora-2, and Aurora-3 protein kinases, and compositions
thereof.
BACKGROUND OF THE INVENTION
[0003] The search for new therapeutic agents has been greatly aided
in recent years by a better understanding of the structure of
enzymes and other biomolecules associated with target diseases. One
important class of enzymes that has been the subject of extensive
study is protein kinases.
[0004] Protein kinases mediate intracellular signal transduction.
They do this by effecting a phosphoryl transfer from a nucleoside
triphosphate to a protein acceptor that is involved in a signaling
pathway. There are a number of kinases and pathways through which
extracellular and other stimuli cause a variety of cellular
responses to occur inside the cell. Examples of such stimuli
include environmental and chemical stress signals (e.g., osmotic
shock, heat shock, ultraviolet radiation, bacterial endotoxin, and
H.sub.2O.sub.2), cytokines (e.g., interleukin-1 (IL-1) and tumor
necrosis factor alpha (TNF-.alpha.)), and growth factors (e.g.,
granulocyte macrophage-colony-stimulating factor (GM-CSF), and
fibroblast growth factor (FGF)). An extracellular stimulus may
affect one or more cellular responses related to cell growth,
migration, differentiation, secretion of hormones, activation of
transcription factors, muscle contraction, glucose metabolism,
control of protein synthesis and regulation of cell cycle.
[0005] Many diseases are associated with abnormal cellular
responses triggered by protein kinase-mediated events. These
diseases include autoimmune diseases, inflammatory diseases, bone
diseases, metabolic diseases, neurological and neurodegenerative
diseases, cancer, cardiovascular diseases, allergies and asthma,
Alzheimer's disease and hormone-related diseases. Accordingly,
there has been a substantial effort in medicinal chemistry to find
protein kinase inhibitors that are effective as therapeutic
agents.
[0006] The Aurora family of serine/threonine kinases is essential
for cell proliferation [Bischoff, J. R. & Plowman, G. D. (The
Aurora/Ipl1p kinase family: regulators of chromosome segregation
and cytokinesis) Trends in Cell Biology 9, 454-459 (1999); Giet, R.
and Prigent, C. (Aurora/Ipl1p-related kinases, a new oncogenic
family of mitotic serine-threonine kinases) Journal of Cell Science
112, 3591-3601 (1999); Nigg, E. A. (Mitotic kinases as regulators
of cell division and its checkpoints) Nat. Rev. Mol. Cell Biol. 2,
21-32 (2001); Adams, R. R, Carmena, M., and Earnshaw, W. C.
(Chromosomal passengers and the (aurora) ABCs of mitosis) Trends in
Cell Biology 11, 49-54 (2001)]. Inhibitors of the Aurora kinase
family therefore have the potential to block growth of all tumour
types.
[0007] The three known mammalian family members, Aurora-A ("1"), B
("2") and C ("3"), are highly homologous proteins responsible for
chromosome segregation, mitotic spindle function and cytokinesis.
Aurora expression is low or undetectable in resting cells, with
expression and activity peaking during the G2 and mitotic phases in
cycling cells. In mammalian cells proposed substrates for Aurora
include histone H3, a protein involved in chromosome condensation,
and CENP-A, myosin II regulatory light chain, protein phosphatase
1, TPX2, all of which are required for cell division.
[0008] Since its discovery in 1997 the mammalian Aurora kinase
family has been closely linked to tumorigenesis. The most
compelling evidence for this is that over-expression of Aurora-A
transforms rodent fibroblasts (Bischoff, J. R., et al. A homologue
of Drosophila aurora kinase is oncogenic and amplified in human
colorectal cancers. EMBO J. 17, 3052-3065 (1998)). Cells with
elevated levels of this kinase contain multiple centrosomes and
multipolar spindles, and rapidly become aneuploid. The oncogenic
activity of Aurora kinases is likely to be linked to the generation
of such genetic instability. Indeed, a correlation between
amplification of the aurora-A locus and chromosomal instability in
mammary and gastric tumours has been observed. (Miyoshi, Y., Iwao,
K., Egawa, C., and Noguchi, S. Association of centrosomal kinase
STK15/BTAK mRNA expression with chromosomal instability in human
breast cancers. Int. J. Cancer 92, 370-373 (2001). (Sakakura, C. et
al. Tumor-amplified kinase BTAK is amplified and overexpressed in
gastric cancers with possible involvement in aneuploid formation.
British Journal of Cancer 84, 824-831 (2001)). The Aurora kinases
have been reported to be over-expressed in a wide range of human
tumours. Elevated expression of Aurora-A has been detected in over
50% of colorectal (Bischoff, J. R., et al. A homologue of
Drosophila aurora kinase is oncogenic and amplified in human
colorectal cancers. EMBO J. 17, 3052-3065 (1998)) (Takahashi, T.,
et al. Centrosomal kinases, HsAIRk1 and HsAIRK3, are overexpressed
in primary colorectal cancers. Jpn. J. Cancer Res. 91, 1007-1014
(2000)). ovarian (Gritsko, T. M. et al. Activation and
overexpression of centrosome kinase BTAK/Aurora-A in human ovarian
cancer. Clinical Cancer Research 9, 1420-1426 (2003)), and gastric
tumors (Sakakura, C. et al. Tumor-amplified kinase BTAK is
amplified and overexpressed in gastric cancers with possible
involvement in aneuploid formation. British Journal of Cancer 84,
824-831 (2001)), and in 94% of invasive duct adenocarcinomas of the
breast (Tanaka, T., et al. Centrosomal kinase AIK1 is overexpressed
in invasive ductal carcinoma of the breast. Cancer Research. 59,
2041-2044 (1999)). High levels of Aurora-A have also been reported
in renal, cervical, neuroblastoma, melanoma, lymphoma, pancreatic
and prostate tumour cell lines. (Bischoff, J. R., et al. A
homologue of Drosophila aurora kinase is oncogenic and amplified in
human colorectal cancers. EMBO J. 17, 3052-3065 (1998) (Kimura, M.,
Matsuda, Y., Yoshioka, T., and Okano, Y. Cell cycle-dependent
expression and centrosomal localization of a third human
Aurora/Ipl1-related protein kinase, AIK3. Journal of Biological
Chemistry 274, 7334-7340 (1999))(Zhou et al. Tumour amplifiec
kinase STK15/BTAK induces centrosome amplification, aneuploidy and
transformation Nature Genetics 20: 189-193 (1998))(Li et al.
Overexpression of oncogenic STK15/BTAK/Aurora-A kinase in human
pancreatic cancer Clin Cancer Res. 9(3):991-7 (2003)).
Amplification/overexpression of Aurora-A is observed in human
bladder cancers and amplification of Aurora-A is associated with
aneuploidy and aggressive clinical behaviour (Sen S. et al
Amplification/overexpression of a mitotic kinase gene in human
bladder cancer J Natl Cancer Inst. 94(17): 1320-9 (2002)).
Moreover, amplification of the aurora-A locus (20q13) correlates
with poor prognosis for patients with node-negative breast cancer
(Isola, J. J., et al. Genetic aberrations detected by comparative
genomic hybridization predict outcome in node-negative breast
cancer. American Journal of Pathology 147, 905-911 (1995)).
Aurora-B is highly expressed in multiple human tumour cell lines,
including leukemic cells (Katayama et al. Human AIM-1: cDNA cloning
and reduced expression during endomitosis in megakaryocyte-lineage
cells. Gene 244:1-7)). Levels of this enzyme increase as a function
of Duke's stage in primary colorectal cancers (Katayama, H. et al.
Mitotic kinase expression and colorectal cancer progression.
Journal of the National Cancer Institute 91, 1160-1162 (1999)).
Aurora-C, which is normally only found in germ cells, is also
over-expressed in a high percentage of primary colorectal cancers
and in a variety of tumour cell lines including cervical
adenocarinoma and breast carcinoma cells (Kimura, M., Matsuda, Y.,
Yoshioka, T., and Okano, Y. Cell cycle-dependent expression and
centrosomal localization of a third human Aurora/Ipl1-related
protein kinase, AIK3. Journal of Biological Chemistry 274,
7334-7340 (1999). (Takahashi, T., et al. Centrosomal kinases,
HsAIRk1 and HsAIRK3, are overexpressed in primary colorectal
cancers. Jpn. J. Cancer Res. 91, 1007-1014 (2000)).
[0009] Based on the known function of the Aurora kinases,
inhibition of their activity should disrupt mitosis leading to cell
cycle arrest. In vivo, an Aurora inhibitor therefore slows tumor
growth and induces regression.
[0010] Elevated levels of all Aurora family members are observed in
a wide variety of tumour cell lines. Aurora kinases are
over-expressed in many human tumors and this is reported to be
associated with chromosomal instability in mammary tumors (Miyoshi
et al 2001 92, 370-373).
[0011] Aurora-2 is highly expressed in multiple human tumor cell
lines and levels increase as a function of Duke's stage in primary
colorectal cancers [Katayama, H. et al. (Mitotic kinase expression
and colorectal cancer progression) Journal of the National Cancer
Institute 91, 1160-1162 (1999)]. Aurora-2 plays a role in
controlling the accurate segregation of chromosomes during mitosis.
Misregulation of the cell cycle can lead to cellular proliferation
and other abnormalities. In human colon cancer tissue, the Aurora-2
protein has been found to be over expressed [Bischoff et al., EMBO
J., 17, 3052-3065 (1998); Schumacher et al., J. Cell Biol., 143,
1635-1646 (1998); Kimura et al., J. Biol. Chem., 272, 13766-13771
(1997)]. Aurora-2 is over-expressed in the majority of transformed
cells. Bischoff et al found high levels of Aurora-2 in 96% of cell
lines derived from lung, colon, renal, melanoma and breast tumors
(Bischoff et al EMBO J. 1998 17, 3052-3065). Two extensive studies
show elevated Aurora-2 in 54% and 68% (Bishoff et al EMBO J. 1998
17, 3052-3065)(Takahashi et al 2000 Jpn J Cancer Res. 91,
1007-1014) of colorectal tumours and in 94% of invasive duct
adenocarcinomas of the breast (Tanaka et al 1999 59,
2041-2044).
[0012] Aurora-1 expression is elevated in cell lines derived from
tumors of the colon, breast, lung, melanoma, kidney, ovary,
pancreas, CNS, gastric tract and leukemias (Tatsuka et al 1998 58,
4811-4816).
[0013] High levels of Aurora-3 have been detected in several tumour
cell lines, although it is restricted to testis in normal tissues
(Kimura et al 1999 274, 7334-7340). Over-expression of Aurora-3 in
a high percentage (c. 50%) of colorectal cancers has also been
documented (Takahashi et al 2000 Jpn J Cancer Res. 91, 1007-1014).
In contrast, the Aurora family is expressed at a low level in the
majority of normal tissues, the exceptions being tissues with a
high proportion of dividing cells such as the thymus and testis
(Bischoff et al EMBO J. 1998 17, 3052-3065).
[0014] For further review of the role Aurora kinases play in
proliferative disorders, see Bischoff, J. R. & Plowman, G. D.
(The Aurora/Ipl1p kinase family:regulators of chromosome
segregation and cytokinesis) Trends in Cell Biology 9, 454-459
(1999); Giet, R. and Prigent, C. (Aurora/Ipl1p-related kinases, a
new oncogenic family of mitotic serine-threonine kinases) Journal
of Cell Science 112, 3591-3601 (1999); Nigg, E. A. (Mitotic kinases
as regulators of cell division and its checkpoints) Nat. Rev. Mol.
Cell Biol. 2, 21-32 (2001); Adams, R. R, Carmena, M., and Earnshaw,
W. C. (Chromosomal passengers and the (aurora) ABCs of mitosis)
Trends in Cell Biology 11, 49-54 (2001); and Dutertre, S.,
Descamps, S., & Prigent, P. (On the role of aurora-A in
centrosome function) Oncogene 21, 6175-6183 (2002).
[0015] The type III receptor tyrosine kinase, Flt3, plays an
important role in the maintenance, growth and development of
hematopoietic and non-hematopoietic cells. [Scheijen, B, Griffin J
D, Oncogene, 2002, 21, 3314-3333 and Reilly, J T, British Journal
of Haematology, 2002, 116, 744-757]. FLT-3 regulates maintenance of
stem cell/early progenitor pools as well the development of mature
lymphoid and myeloid cells [Lyman, S, Jacobsen, S, Blood, 1998, 91,
1101-1134]. FLT-3 contains an intrinsic kinase domain that is
activated upon ligand-mediated dimerization of the receptors. Upon
activation, the kinase domain induces autophosphorylation of the
receptor as well as the phosphorylation of various cytoplasmic
proteins that help propogate the activation signal leading to
growth, differentiation and survival. Some of the downstream
regulators of FLT-3 receptor signaling include, PLC.gamma.,
P13-kinase, Grb-2, SHIP and Src related kinases [Scheijen, B,
Griffin J D, Oncogene, 2002, 21, 3314-3333]. FLT-3 kinase plays a
role in a variety of hematopoietic and non-hematopoietic
malignancies. Mutations that induce ligand independent activation
of FLT-3 have been implicated in acute-myelogenous leukemia (AML),
acute lymphocytic leukemia (ALL), mastocytosis and gastrointestinal
stromal tumor (GIST). These mutations include single amino acid
changes in the kinase domain or internal tandem duplications, point
mutations or in-frame deletions of the juxtamembrane region of the
receptors. In addition to activating mutations, ligand dependent
(autocrine or paracrine) stimulation of over-expressed wild-type
FLT-3 contributes to the malignant phenotype [Scheijen, B, Griffin
J D, Oncogene, 2002, 21, 3314-3333]. See also Sawyer, C. I.
(Finding the next Gleevec: FLT3 targeted kinase inhibitor therapy
for acute myeloid leukaemia) Cancer Cell. 1, 413-415 (2002).
[0016] Tri- or tetra-substituted pyrimidine derivatives useful as
kinase inhibitors are known in the art. Typically, these pyrimidine
derivatives are 2,4,6- or 2,4,5,6-substituted, as shown below:
1
[0017] Known methods for preparing such pyrimidine derivatives have
many synthetic drawbacks such as lacking the ability to
regioselectively introduce substituents at the 2-, 4-, or
6-position in high yields. See M. Botta, Nucleosides Nucleotides,
13, 8, 1994, 1769-78; M. Ban, Bioorg. Med. Chem., 6, 7, 1998,
1057-68; Y. Fellahi, Eur. J. Med. Chem. Chim. Ther., 31, 1, 1996,
77-82; T. J. Delia, J. Het. Chem., 35, 2, 1998, 269-74; H. Uchel,
Tetraheron Lett., 36, 52, 1995, 9457-60; and Y. Nezu, Pestic. Sci.,
47, 2, 1996, 115-24.
[0018] There is a need for a synthetic process that can be readily
used to prepare the tri- or tetra-substituted pyrimidine
derivatives on a large scale. There is also a need for a process
that employs minimal steps and utilizes readily available starting
materials and simple reaction media. Ideally, such a process will
be easy to scale up, if need be, and will be inexpensive. There is
also a need for a process that does not lead to regioisomeric
intermediate mixtures that must be separated by, e.g.,
chromatographic methods. Such separations reduce the overall
yields.
[0019] It would be desirable to have a synthetic process to produce
tri- or tetra-substituted pyrimidine derivatives that possesses the
above advantages and thereby improves upon the currently available
processes.
SUMMARY OF THE INVENTION
[0020] The present invention provides a process for preparing a
compound of formula I: 2
[0021] wherein:
[0022] Q and T are each independently selected from oxygen, sulfur
or N(R);
[0023] each R is independently selected from hydrogen or an
optionally substituted C.sub.1-6 aliphatic group, wherein:
[0024] two R bound to the same nitrogen atom are optionally taken
together with the nitrogen to form an optionally substituted 3-7
membered monocyclic or 8-10 membered bicyclic saturated, partially
unsaturated, or fully unsaturated ring having 0-3 heteroatoms, in
addition to the nitrogen bound thereto, independently selected from
nitrogen, oxygen, or sulfur;
[0025] R.sup.x is U-R.sup.5;
[0026] R.sup.5 is selected from halogen, NO.sub.2, CN, R, or
Ar;
[0027] each U is independently selected from a valence bond or a
C.sub.1-4 alkylidene chain, wherein:
[0028] up to two methylene units of U are optionally and
independently replaced by --O--, --S--, --SO--, --SO.sub.2--,
--N(R)SO.sub.2--, --SO.sub.2N(R)--, --N(R)--, --C(O)--,
--CO.sub.2--, --N(R)C(O)--, --N(R)C(O)O--, --N(R)CON(R)--,
--N(R)SO.sub.2N(R)--, --N(R)N(R)--, --C(O)N(R)--, --OC(O)N(R)--,
--C(R).dbd.NN(R)--, or --C(R).dbd.N--O--;
[0029] each Ar is independently selected from an optionally
substituted ring selected from a 3-7 membered monocyclic or an 8-10
membered bicyclic saturated, partially unsaturated, or fully
unsaturated ring having 0-4 heteroatoms independently selected from
nitrogen, oxygen, or sulfur;
[0030] R.sup.y is --N(R.sup.1).sub.2, --OR.sup.1, or
--SR.sup.1;
[0031] each R.sup.1 is independently selected from R or a 3-8
membered monocyclic, an 8-10 membered bicyclic, or a 10-12 membered
tricyclic saturated, partially unsaturated, or fully unsaturated
ring having 0-4 heteroatoms independently selected from nitrogen,
oxygen, or sulfur, and wherein:
[0032] each R.sup.1 is optionally and independently substituted by
up to four substituents independently selected from R.sup.2;
[0033] each R.sup.2 is independently selected from --R.sup.3,
--OR.sup.3, --SR.sup.3, --CN, --NO.sub.2, oxo, halogen,
--N(R.sup.3).sub.2, --C(O)R.sup.3, --OC(O)R.sup.3,
--CO.sub.2R.sup.3, --SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or
--NR.sup.3CO.sub.2(R.sup.3);
[0034] each R.sup.3 is independently selected from R or Ar;
[0035] R.sup.z1 is selected from a C.sub.1-6 aliphatic group or a
3-8 membered monocyclic, an 8-10 membered bicyclic, or a 10-12
membered tricyclic saturated, partially unsaturated, or fully
unsaturated ring having 0-4 heteroatoms independently selected from
oxygen, nitrogen or sulfur, wherein:
[0036] R.sup.z1 is substituted with 0-4 independently selected
R.sup.2 groups;
[0037] R.sup.z2 is C.sub.1-6 aliphatic group or a 3-8 membered
monocyclic or an 8-10 membered bicyclic saturated, partially
unsaturated, or fully unsaturated ring having 0-4 heteroatoms
independently selected from nitrogen, oxygen or sulfur,
wherein:
[0038] R.sup.z2 is substituted by 0-4 substituents independently
selected from oxo or U-R.sup.5;
[0039] said process comprising the step of combining a compound of
formula II and a compound of formula R.sup.y--H in a suitable
medium: 3
[0040] wherein:
[0041] said suitable medium comprises:
[0042] i) a suitable solvent; and
[0043] ii) optionally, a suitable base; and
[0044] L.sup.3 is a suitable leaving group.
DESCRIPTION OF THE INVENTION
[0045] The present invention provides a process for preparing a
compound of formula I: 4
[0046] wherein:
[0047] Q and T are each independently selected from oxygen, sulfur
or N(R);
[0048] each R is independently selected from hydrogen or an
optionally substituted C.sub.1-6 aliphatic group, wherein:
[0049] two R bound to the same nitrogen atom are optionally taken
together with the nitrogen to form an optionally substituted 3-7
membered monocyclic or 8-10 membered bicyclic saturated, partially
unsaturated, or fully unsaturated ring having 0-3 heteroatoms, in
addition to the nitrogen bound thereto, independently selected from
nitrogen, oxygen, or sulfur;
[0050] R.sup.x is U-R.sup.5;
[0051] R.sup.5 is selected from halogen, NO.sub.2, CN, R, or
Ar;
[0052] each U is independently selected from a valence bond or a
C.sub.1-4 alkylidene chain, wherein:
[0053] up to two methylene units of U are optionally and
independently replaced by --O--, --S--, --SO--, --SO.sub.2--,
--N(R)SO.sub.2--, --SO.sub.2N(R)--, --N(R)--, --C(O)--,
--CO.sub.2--, --N(R)C(O)--, --N(R)C(O)O--, --N(R)CON(R)--,
--N(R)SO.sub.2N(R)--, --N(R)N(R)--, --C(O)N(R)--, --OC(O)N(R)--,
--C(R).dbd.NN(R)--, or --C(R).dbd.N--O--;
[0054] each Ar is independently selected from an optionally
substituted ring selected from a 3-7 membered monocyclic or an 8-10
membered bicyclic saturated, partially unsaturated, or fully
unsaturated ring having 0-4 heteroatoms independently selected from
nitrogen, oxygen, or sulfur;
[0055] R.sup.y is --N(R.sup.1).sub.2, --OR.sup.1, or
--SR.sup.1;
[0056] each R.sup.1 is independently selected from R or a 3-8
membered monocyclic, an 8-10 membered bicyclic, or a 10-12 membered
tricyclic saturated, partially unsaturated, or fully unsaturated
ring having 0-4 heteroatoms independently selected from nitrogen,
oxygen, or sulfur, and wherein:
[0057] each R.sup.1 is optionally and independently substituted by
up to four substituents independently selected from R.sup.2;
[0058] each R.sup.2 is independently selected from --R, --OR.sup.3,
--SR.sup.3, --CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2,
--C(O)R.sup.3, --OC(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or
--NR.sup.3CO.sub.2(R.sup.3);
[0059] each R.sup.3 is independently selected from R or Ar;
[0060] R.sup.z1 is selected from a C.sub.1-6 aliphatic group or a
3-8 membered monocyclic, an 8-10 membered bicyclic, or a 10-12
membered tricyclic saturated, partially unsaturated, or fully
unsaturated ring having 0-4 heteroatoms independently selected from
oxygen, nitrogen or sulfur, wherein:
[0061] R.sup.z1 is substituted with 0-4 independently selected
R.sup.2 groups;
[0062] R.sup.z2 is C.sub.1-6 aliphatic group or a 3-8 membered
monocyclic or an 8-10 membered bicyclic saturated, partially
unsaturated, or fully unsaturated ring having 0-4 heteroatoms
independently selected from nitrogen, oxygen or sulfur,
wherein:
[0063] R.sup.z2 is substituted by 0-4 substituents independently
selected from oxo or U-R.sup.5;
[0064] said process comprising the step of combining a compound of
formula II and a compound of formula R.sup.y--H in a suitable
medium: 5
[0065] wherein:
[0066] said suitable medium comprises:
[0067] i) a suitable solvent; and
[0068] ii) optionally, a suitable base; and
[0069] L is a suitable leaving group.
[0070] According to another embodiment, a compound of formula II is
prepared by combining a compound of formula III with a compound of
formula R.sup.z1-Q-H in a suitable medium: 6
[0071] wherein:
[0072] said suitable medium comprises:
[0073] i) a suitable solvent; and
[0074] ii) optionally, a suitable base; and
[0075] L is a suitable leaving group.
[0076] According to yet another embodiment, a compound of formula
III is prepared by combining a compound of formula IV with a
compound of formula R.sup.z2-T-H in a suitable medium: 7
[0077] wherein:
[0078] said suitable medium comprises:
[0079] i) a suitable solvent; and
[0080] ii) optionally, a suitable base; and
[0081] L.sup.1 is a suitable leaving group.
[0082] A suitable solvent is a solvent or a solvent mixture that,
in combination with the combined compounds, may facilitate the
progress of the reaction therebetween. The suitable solvent may
solubilize one or more of the reaction components, or,
alternatively, the suitable solvent may facilitate the agitation of
a suspension of one or more of the reaction components. Examples of
suitable solvents useful in the present invention are a protic
solvent, a halogenated hydrocarbon, an ether, an aromatic
hydrocarbon, a polar or a non-polar aprotic solvent, or any
mixtures thereof. These and other such suitable solvents are well
known in the art, e.g., see, "Advanced Organic Chemistry", Jerry
March, 4.sup.th edition, John Wiley and Sons, N.Y. (1992).
[0083] Preferably the suitable solvent is a C.sub.1-7 straight or
branched alkyl alcohol, ether, or a polar or non-polar aprotic
solvent.
[0084] For the reaction between a compound of formula II and a
compound R.sup.y--H, a more preferred suitable solvent is selected
from ethanol, isopropanol, t-butanol, n-butanol or
tetrahydrofuran.
[0085] For the reaction between a compound of formula III and a
compound R.sup.z1-Q-H, a more preferred suitable solvent is
selected from ethanol, isopropanol, t-butanol, n-butanol,
N,N-dimethylformamide, dimethylsulfoxide, or tetrahydrofuran.
[0086] For the reaction between a compound of formula IV and a
compound R.sup.z2-T-H, a more preferred suitable solvent is
selected from N,N-dimethylformamide, dimethylsulfoxide, or
tetrahydrofuran.
[0087] According to an alternate embodiment, the suitable solvent
is R.sup.y--H. Thus, in such an embodiment, the reagent R.sup.y--H
acts, in part, as a suitable solvent in combination with a compound
of formula II, and also acts, in part, as a reagent and reacts with
the compound of formula II to produce compound of formula I.
[0088] According to another alternative embodiment, the suitable
solvent is R.sup.z1-Q-H. Thus, in such an embodiment, the reagent
R.sup.z1-Q-H acts, in part, as a suitable solvent in combination
with a compound of formula III, and also acts, in part, as a
reagent and reacts with the compound of formula III to produce
compound of formula II.
[0089] According to another alternative embodiment, the suitable
solvent is R.sup.z2_T-H. Thus, in such an embodiment, the reagent
R.sup.z2-T-H acts, in part, as a suitable solvent in combination
with a compound of formula IV, and also acts, in part, as a reagent
and reacts with the compound of formula IV to produce compound of
formula III.
[0090] A suitable base is a chemical entity that has the ability to
be a proton acceptor. Examples include organic amines, alkaline
earth metal carbonates, alkaline earth metal hydrides, and alkaline
earth metal hydroxides. These and other such suitable bases are
well known in the art, e.g., see, "Advanced Organic Chemistry,"
Jerry March, 4.sup.th Ed., pp. 248-253, John Wiley and Sons, N.Y.
(1992). Preferred suitable bases include trialkyl amines, sodium
carbonate, potassium carbonate, sodium hydride, potassium hydride,
sodium hydroxide, or potassium hydroxide. More preferably, the
suitable base is diisopropylethylamine or triethylamine.
[0091] A suitable leaving group is a chemical group that is readily
displaced by a desired incoming chemical moiety. Thus, the choice
of the specific suitable leaving group is predicated upon its
ability to be readily displaced by the incoming chemical moiety
R.sup.y in R.sup.y--H, R.sup.z1-Q in R.sup.z1-Q-H, or R.sup.z2-T in
R.sup.z2-T-H. Suitable leaving groups are well known in the art,
e.g., see, "Advanced Organic Chemistry," Jerry March, 4.sup.th Ed.,
pp. 351-357, John Wiley and Sons, N.Y. (1992). Such leaving groups
include, but are not limited to, halogen, alkoxy, sulphonyloxy,
optionally substituted alkylsulphonyl, optionally substituted
alkenylsulfonyl, optionally substituted arylsulfonyl, and diazonium
moieties. Examples of suitable leaving groups include chloro, iodo,
bromo, fluoro, methanesulfonyl (mesyl), tosyl, triflate,
nitro-phenylsulfonyl (nosyl), and bromo-phenylsulfonyl
(brosyl).
[0092] For example, in the process of preparing a compound of
formula I, L.sup.3 is displaced by incoming moiety R.sup.y of
R.sup.y--H. Thus, if R.sup.y--H is e.g., a piperazine, then L.sup.3
is a leaving group that is readily displaced by the --NH-- moiety
in piperazine.
[0093] Preferred L.sup.3 leaving groups are selected from halogen,
optionally substituted arylsulfonyl, or optionally substituted
alkylsulphonyl. More preferably, L.sup.3 is chloro, iodo, or
methanesulfonyl. Most preferably, L.sup.3 is chloro.
[0094] For example, in the process of preparing a compound of
formula II, L.sup.2 is displaced by incoming moiety R.sup.z1-Q of
R.sup.z1-Q-H. Thus, if R.sup.z1-Q-H is, e.g., 3-aminopyrazole, then
L.sup.2 is a leaving group that is readily displaced by the
3-aminopyrazole.
[0095] Preferred L.sup.2 leaving groups are selected from halogen,
optionally substituted arylsulfonyl, or optionally substituted
alkylsulphonyl. More preferably, L.sup.3 is chloro, iodo, or
fluoro. Most preferably, L.sup.3 is chloro.
[0096] For example, in the process of preparing a compound of
formula III, L.sup.1 is displaced by incoming moiety R.sup.z2-T of
R.sup.z2-T-H. Thus, if R.sup.z2-T is e.g., an optionally
substituted arylthiol, then L.sup.1 is a leaving group that is
readily displaced by the thio group in the optionally substituted
arylthiol.
[0097] Preferred L.sup.1 leaving groups are selected from halogen,
optionally substituted arylsulfonyl, or optionally substituted
alkylsulphonyl. More preferably, L.sup.3 is chloro, iodo, or
methanesulfonyl. Most preferably, L.sup.3 is methanesulfonyl.
[0098] According to an alternate embodiment, the suitable leaving
group may be generated in situ within the reaction medium. For
example, L.sup.3 in a compound of formula II may be generated in
situ from a precursor of that compound of formula II wherein said
precursor contains a group readily replaced by L.sup.3 in situ. In
a specific illustration of such a replacement, said precursor of a
compound of formula II contains a group (for example, a chloro
group or hydroxyl group) which is replaced in situ by L.sup.3, such
as an iodo group. The source of the iodo group may be, e.g., sodium
iodide. Accordingly, L.sup.2 and L.sup.1 may also be formed in situ
in an analogous manner. Such an in situ generation of a suitable
leaving group is well known in the art, e.g., see, "Advanced
Organic Chemistry," Jerry March, pp. 430-431, 4.sup.th Ed., John
Wiley and Sons, N.Y. (1992).
[0099] According to yet another alternative embodiment, an anion of
any of R.sup.y in R.sup.y--H, R.sup.z1-Q in R.sup.z1-Q-H, or
R.sup.z2-T in R.sup.z2-T-H may be formed prior to addition to the
reaction medium. The preparation of said anion is well known to one
of skill in the art. For example, when T is oxygen, the anion of
R.sup.z2-T-H is readily formed by treating R.sup.z2-T-H with a
base, such as sodium hydride. This oxygen anion may then be
combined with the compound of formula IV to form a compound of
formula III.
[0100] According to another embodiment, the reactions described
herein are performed at a temperature less than or equal to the
reflux temperature of the reaction medium. According to another
embodiment, said reaction medium has a temperature less than the
boiling point of said suitable solvent or at a temperature attained
by refluxing said suitable solvent in said reaction medium. In
another embodiment, said reaction medium has a temperature between
about 0.degree. C. and about 190.degree. C. According to yet
another embodiment, said reaction medium has a temperature between
about 40.degree. C. and about 120.degree. C. According to another
aspect of the present invention, said reaction medium has a
temperature between about 70.degree. C. and about 115.degree.
C.
[0101] As used herein, the following definitions shall apply unless
otherwise indicated.
[0102] The term "Aurora" refers to any isoform of the Aurora family
of protein kinases, including Aurora-1, Aurora-2, and Aurora-3. The
term "Aurora" also refers to isoforms of the Aurora family of
protein kinases known as Aurora-A, Aurora-B, and Aurora-C.
[0103] The phrase "optionally substituted" is used interchangeably
with the phrase "substituted or unsubstituted." Unless otherwise
indicated, an optionally substituted group may have a substituent
at each substitutable position of the group, and each substitution
is independent of the other.
[0104] The term "aliphatic" or "aliphatic group" as used herein
means a straight-chain or branched C.sub.1-C.sub.8 hydrocarbon
chain that is completely saturated or that contains one or more
units of unsaturation, or a monocyclic C.sub.3-C.sub.8 hydrocarbon
or bicyclic C.sub.8-C.sub.12 hydrocarbon that is completely
saturated or that contains one or more units of unsaturation, but
which is not aromatic (also referred to herein as "carbocycle" or
"cycloalkyl"), 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. For example, suitable aliphatic groups
include, but are not limited to, linear or branched or alkyl,
alkenyl, alkynyl groups and hybrids thereof such as
(cycloalkyl)alkyl, (cycloalkenyl)alkyl or (cycloalkyl)alkenyl.
[0105] The terms "alkyl", "alkoxy", "hydroxyalkyl", "alkoxyalkyl",
and "alkoxycarbonyl", used alone or as part of a larger moiety
include both straight and branched chains containing one to twelve
carbon atoms. The terms "alkenyl" and "alkynyl" used alone or as
part of a larger moiety shall include both straight and branched
chains containing two to twelve carbon atoms.
[0106] The term "heteroatom" means nitrogen, oxygen, or sulfur and
includes any oxidized form of nitrogen and sulfur, and the
quaternized form of any basic nitrogen. Also the term "nitrogen"
includes a substitutable nitrogen of a heterocyclic ring. As an
example, in a saturated or partially unsaturated ring having 0-3
heteroatoms selected from oxygen, sulfur or nitrogen, the nitrogen
may be N (as in 3,4-dihydro-2H-pyrrolyl), NH (as in pyrrolidinyl)
or NR.sup.+ (as in N-substituted pyrrolidinyl).
[0107] The term "aryl" or "aryl ring" refers to a monocyclic,
bicyclic, or tricyclic ring systems having a total of five to
fourteen ring carbon atoms, wherein at least one ring is aromatic
and wherein each ring in the system contains three to seven ring
members. The term "aryl" may be used interchangeably with the term
"aryl ring." Examples include phenyl, indanyl, 1-naphthyl,
2-naphthyl, 1-anthracyl, 2-anthracyl and bicyclo
[2.2.2]oct-3-yl.
[0108] More preferred ring sizes for aryl rings are as set forth
below for the various preferred embodiments of compounds of formula
I.
[0109] The term "aryl" used alone or as part of a larger moiety as
in "aralkyl", "aralkoxy", or "aryloxyalkyl", refers to monocyclic,
bicyclic and tricyclic ring systems having a total of five to
fourteen ring members, wherein at least one ring in the system is
aromatic and wherein each ring in the system contains 3 to 7 ring
members. The term "aryl" may be used interchangeably with the term
"aryl ring". The term "aryl" also refers to heteroaryl ring systems
as defined hereinbelow.
[0110] The term "heterocycle", "heterocyclyl", or "heterocyclic" as
used herein means non-aromatic, monocyclic, bicyclic or tricyclic
ring systems having five to fourteen ring members in which one or
more ring members is a heteroatom, wherein each ring in the system
contains 3 to 7 ring members.
[0111] The term "heteroaryl", used alone or as part of a larger
moiety as in "heteroaralkyl" or "heteroarylalkoxy", refers to
monocyclic, bicyclic and tricyclic 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 "heteroaryl ring" or the term "heteroaromatic".
[0112] An aryl (including aralkyl, aralkoxy, aryloxyalkyl and the
like) or heteroaryl (including heteroaralkyl and heteroarylalkoxy
and the like) group may contain one or more substituents. Suitable
substituents on the unsaturated carbon atom of an aryl, heteroaryl,
aralkyl, or heteroaralkyl group are selected from halogen,
--R.degree., --OR.degree., --SR.degree., 1,2-methylene-dioxy,
1,2-ethylenedioxy, phenyl (Ph) optionally substituted with
R.degree., --O(Ph) optionally substituted with R.degree.,
--CH.sub.2(Ph) optionally substituted with R.degree.,
--CH.sub.2CH.sub.2(Ph), optionally substituted with R.degree.,
--NO.sub.2, --CN, --N(R.degree.).sub.2, --NR.degree.C(O)R.degree.,
--NR.degree.C(O)N(R.degree.).sub.2, --NR.degree.CO.sub.2R.degree.,
--NR.degree. NR.degree.C(O)R.degree.,
--NR.degree.NR.degree.C(O)N(R.degre- e.).sub.2,
--NR.degree.NR.degree.CO.sub.2RO, --C(O)C(O)R.degree.,
--C(O)CH.sub.2C(O)R.degree., --CO.sub.2R, --C(O)R.degree.,
--C(O)N(R.).sub.2, --OC(O)N(R.degree.).sub.2,
--S(O).sub.2R.degree., --SO.sub.2N(R.degree.).sub.2,
--S(O)R.degree., --NR.degree.SO.sub.2N(R.de- gree.).sub.2,
--NR.degree.SO.sub.2R.degree., --C(.dbd.S)N(R.degree.).sub.2- ,
--C(.dbd.NH)--N(R.degree.).sub.2, or
--(CH.sub.2).sub.yNHC(O)R.degree., wherein each R.degree. is
independently selected from hydrogen, optionally substituted
C.sub.1-6 aliphatic, an unsubstituted 5-6 membered heteroaryl or
heterocyclic ring, phenyl, --O(Ph), or --CH.sub.2(Ph). Optional
substituents on the aliphatic group of R.degree. are selected from
NH.sub.2, NH(C.sub.1-4 aliphatic), N(C.sub.1-4 aliphatic).sub.2,
halogen, C.sub.1-4 aliphatic, OH, O(C.sub.1-4 aliphatic), NO.sub.2,
CN, CO.sub.2H, CO.sub.2(C.sub.1-4 aliphatic), O(halo C.sub.1-4
aliphatic), or halo C.sub.1-4 aliphatic.
[0113] An aliphatic group or a non-aromatic heterocyclic ring may
contain one or more substituents. Suitable substituents on the
saturated carbon of an aliphatic group or of a non-aromatic
heterocyclic ring are selected from those listed above for the
unsaturated carbon of an aryl or heteroaryl group and the
following: .dbd.O, .dbd.S, .dbd.NNHR, .dbd.NN(R*).sub.2,
.dbd.NNHC(O)R*, .dbd.NNHCO.sub.2(alkyl), .dbd.NNHSO.sub.2(alkyl),
or .dbd.NR*, where each R* is independently selected from hydrogen
or an optionally substituted C.sub.1-6 aliphatic. Optional
substituents on the aliphatic group of R* are selected from
NH.sub.2, NH(C.sub.1-4 aliphatic), N(C.sub.1-4 aliphatic).sub.2,
halogen, C.sub.1-4 aliphatic, OH, O(C.sub.1-4 aliphatic), NO.sub.2,
CN, CO.sub.2H, CO.sub.2(C.sub.1-4 aliphatic), O(halo C.sub.1-4
aliphatic), or halo(C.sub.1-4 aliphatic).
[0114] Optional substituents on the nitrogen of a non-aromatic
heterocyclic ring are selected from --R.sup.+, --N(R.sup.+).sub.2,
--C(O)R.sup.+, --CO.sub.2R.sup.+, --C(O)C(O)R.sup.+,
--C(O)CH.sub.2C(O)R.sup.+, --SO.sub.2R.sup.+,
--SO.sub.2N(R.sup.+).sub.2, --C(.dbd.S)N(R.sup.+).sub.2,
--C(.dbd.NH)--N(R.sup.+).sub.2, or --NR.sup.+SO.sub.2R.sup.+;
wherein R.sup.+ is hydrogen, an optionally substituted C.sub.1-6
aliphatic, optionally substituted phenyl, optionally substituted
--O(Ph), optionally substituted --CH.sub.2(Ph), optionally
substituted --CH.sub.2CH.sub.2(Ph), or an unsubstituted 5-6
membered heteroaryl or heterocyclic ring. Optional substituents on
the aliphatic group or the phenyl ring of R+are selected from
NH.sub.2, NH(C.sub.1-4 aliphatic), N(C.sub.1-4 aliphatic).sub.2,
halogen, C.sub.1-4 aliphatic, OH, O(C.sub.1-4 aliphatic), NO.sub.2,
CN, CO.sub.2H, CO.sub.2(C.sub.1-4 aliphatic), O(halo C.sub.1-4
aliphatic), or halo(C.sub.1-4 aliphatic).
[0115] The term "alkylidene chain" refers to a straight or branched
carbon chain that may be fully saturated or have one or more units
of unsaturation and has two points of attachment to the rest of the
molecule.
[0116] A combination of substituents or variables is permissible
only if such a combination results in a stable or chemically
feasible compound. A stable compound or chemically feasible
compound is one that is not substantially altered when kept at a
temperature of 40.degree. C. or less, in the absence of moisture or
other chemically reactive conditions, for at least a week.
[0117] It will be apparent to one skilled in the art that certain
compounds of this invention may exist in tautomeric forms, all such
tautomeric forms of the compounds being within the scope of the
invention.
[0118] Unless otherwise stated, structures depicted herein are also
meant to include all stereochemical forms of the structure; i.e.,
the R and S configurations for each asymmetric center. Therefore,
single stereochemical isomers as well as enantiomeric and
diastereomeric mixtures of the present compounds are within the
scope of the invention. Unless otherwise stated, 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 a hydrogen by a deuterium or tritium, or the
replacement of a carbon by a .sup.13C- or .sup.14C-enriched carbon
are within the scope of this invention. Such compounds are useful,
for example, as analytical tools or probes in biological
assays.
[0119] According to another embodiment, Q of formula I is NH,
oxygen, or sulfur.
[0120] According to a preferred embodiment, Q of formula I is NR.
More preferably, Q of formula I is NH.
[0121] According to another preferred embodiment, T of formula I is
oxygen or sulfur. More preferably, T of formula I is sulfur.
[0122] According to another embodiment, T of formula I is oxygen
and the anion of R.sup.z2-T-H is formed prior to combing with a
compound of formula IV to form a compound of formula III.
[0123] According to another embodiment, R.sup.x of formula I is
U-R.sup.5, wherein U is a valence bond, --O--, or --NR--, and
R.sup.5 is R or Ar.
[0124] According to another preferred embodiment R.sup.x of formula
I is selected from R, Ar, or --N(R).sub.2. More preferably, R.sup.x
of formula I is hydrogen.
[0125] According to another preferred embodiment R.sup.y of formula
I is selected from --OR.sup.1 or --N(R.sup.1).sub.2.
[0126] According to another embodiment, R.sup.y of formula I is
selected from N(R.sup.1).sub.2 wherein each R.sup.1 is
independently selected from R or a 3-7 membered monocyclic or an
8-10 membered bicyclic saturated, partially unsaturated, or fully
unsaturated ring having 0-4 heteroatoms independently selected from
nitrogen, oxygen, or sulfur. Preferred substituents R.sup.1 are
selected from --OR.sup.3, --SR.sup.3, --CN, --NO.sub.2, oxo,
halogen, --N(R.sup.3).sub.2, --C(O)R.sup.3, or a 3-6 membered
aromatic or non-aromatic ring having zero to two heteroatoms
independently selected from nitrogen, oxygen, or sulfur. More
preferred substituents on R.sup.1 are 5-6 membered non-aromatic
rings having 1-2 heteroatoms independently selected from nitrogen,
oxygen, or sulfur. Most preferred substituents on the R.sup.1
C.sub.1-4 aliphatic group are NH(CH.sub.3), NH.sub.2, OH,
OCH.sub.3, morpholinyl, piperidinyl, piperazinyl, pyrrolidinyl, and
thiomorpholinyl.
[0127] According to another preferred embodiment, R.sup.y of
formula I is selected from N(R.sup.1).sub.2 wherein each R.sup.1 is
R such that the two R groups are taken together to form an
optionally substituted 4-7 membered non-aromatic ring having up to
two additional heteroatoms independently selected from nitrogen,
oxygen, or sulfur. Preferred substituents on said ring are selected
from --R.sup.3, --OR.sup.3, --SR.sup.3, --CN, --NO.sub.2, oxo,
halogen, --N(R.sup.3).sub.2, --C(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, or a 3-6 membered aromatic or non-aromatic ring
having zero to two heteroatoms independently selected from
nitrogen, oxygen, or sulfur. More preferred substituents said ring
are selected from optionally substituted C.sub.1-4 aliphatic,
NH.sub.2, NH(C.sub.1-4 aliphatic), N(C.sub.1-4 aliphatic).sub.2,
optionally substituted phenyl, CO.sub.2(C.sub.1-4 aliphatic), or
SO.sub.2(C.sub.1-4 aliphatic). Most preferred substituents on said
ring are selected from methyl, ethyl, methylsulfonyl,
(CH.sub.2).sub.2SO.sub.2CH.sub.3, cyclopropyl, CH.sub.2cyclopropyl,
(CH.sub.2).sub.2OH, CO.sub.2t-butyl, CH.sub.2phenyl, phenyl,
NH.sub.2, NH(CH.sub.3), N(CH.sub.3).sub.2,
(CH.sub.2).sub.2NH.sub.2, (CH.sub.2).sub.2morpholin-4-yl,
(CH.sub.2).sub.2N(CH.sub.3).sub.2, isopropyl, propyl, t-butyl,
(CH.sub.2).sub.2CN, or (CH.sub.2).sub.2C(O)morpholin-4-yl.
[0128] Most preferably, R.sup.y of formula I is pyrrolidin-1-yl,
piperidinl-yl, morpholin-4-yl, thiomorpholin-4-yl, piperazin-1-yl,
diazepanyl, or tetrahydroisoquinolinyl, wherein each ring is
optionally substituted with one or two groups independently
selected from methyl, ethyl, methylsulfonyl,
(CH.sub.2).sub.2SO.sub.2CH.sub.3, cyclopropyl, CH.sub.2cyclopropyl,
(CH.sub.2).sub.2OH, CO.sub.2t-butyl, CH.sub.2phenyl, phenyl,
NH.sub.2, NH(CH.sub.3), N(CH.sub.3).sub.2,
(CH.sub.2).sub.2NH.sub.2, (CH.sub.2).sub.2morpholin-4-yl,
(CH.sub.2).sub.2N(CH.sub.3).sub.2, isopropyl, propyl, t-butyl,
(CH.sub.2).sub.2CN, or (CH.sub.2).sub.2C(O)morpholin-4-yl.
[0129] According to another embodiment R.sup.z1 of formula I is a
3-7 membered monocyclic or an 8-10 membered bicyclic saturated,
partially unsaturated, or fully unsaturated ring having 0-4
heteroatoms independently selected from oxygen, nitrogen or sulfur,
wherein said ring is optionally and independently substituted by up
to three substituents selected from --R.sup.3, --OR.sup.3,
--SR.sup.3, --CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2,
--C(O)R.sup.3, --OC(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).sub.2, or --NR.sup.3CO.sub.2R.sup.3.
[0130] According to another embodiment, R.sup.z1 of formula I is a
5-6 membered monocyclic or an 8-10 membered bicyclic saturated,
partially unsaturated, or fully unsaturated ring having 1-4
heteroatoms independently selected from oxygen, nitrogen or sulfur,
wherein said ring is optionally and independently substituted by up
to three substituents selected from --R.sup.3, --OR.sup.3,
--SR.sup.3, --CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2,
--C(O)R.sup.3, --OC(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).sub.2, or --NR.sup.3CO.sub.2R.sup.3.
[0131] According to a more preferred embodiment, R.sup.z1 of
formula I is a five or six membered fully unsaturated ring having
1-3 heteroatoms independently selected from nitrogen, oxygen, or
sulfur, wherein said ring is optionally and independently
substituted by up to three substituents selected from --R.sup.3,
--OR.sup.3, --SR.sup.3, --CN, --NO.sub.2, oxo, halogen,
--N(R.sup.3).sub.2, --C(O)R.sup.3, --OC(O)R.sup.3,
--CO.sub.2R.sup.3--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).- sub.2, or --NR.sup.3CO.sub.2R.sup.3.
[0132] Preferred R.sup.z1 rings of formula I are optionally
substituted rings selected from pyrazole or any one of the
following 5-6 membered rings: 8
[0133] Most preferably, R.sup.z1 of formula I is a pyrazole ring
having up to three substituents as defined above.
[0134] According to another preferred embodiment R.sup.z1 of
formula I has up to two substituents, wherein said substituents are
as set forth above. More preferably, R.sup.z1 of formula I has one
substituent, wherein said substituent is as setsforth above.
[0135] Preferred substituents on the R.sup.z1 moiety of formula I
are --N(R.sup.3).sub.2, --OR.sup.3, Ar, or an optionally
substituted C.sub.1-C.sub.4 a liphatic group, wherein Ar is an
optionally substituted 5-6 membered saturated, partially
unsaturated, or fully unsaturated ring having 0-4 heteroatoms
independently selected from nitrogen, oxygen, or sulfur. An even
more preferred substituents on the R.sup.z1 moiety of formula I is
a C.sub.1-C.sub.4 aliphatic group. Most preferred substituents on
the R.sup.z1 moiety of formula I are selected from methyl, ethyl,
propyl, isopropyl, t-butyl, cyclopropyl, or phenyl.
[0136] According to another embodiment R.sup.z1 of formula I is a
C.sub.1-6 aliphatic group substituted with 0-4 R.sup.2 groups.
Preferably, R.sup.z1 is substituted with 0-3 R.sup.2 groups,
wherein each R.sup.2 is independently selected from R.sup.3, oxo,
halogen, N(R.sup.3).sub.2, CN, or CO.sub.2R.sup.3.
[0137] According to a preferred embodiment, R.sup.z2 of formula I
is a 5-6 membered monocyclic or an 8-10 membered bicyclic
saturated, partially unsaturated, or fully unsaturated ring having
0-4 heteroatoms independently selected from nitrogen, oxygen or
sulfur, wherein said ring is optionally substituted by up to three
substituents independently selected from --R.sup.3, --OR.sup.3,
--SR.sup.3, --CN, --NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2,
--C(O)R.sup.3, --OC(O)R.sup.3, --CO.sub.2R.sup.3,
--SO.sub.2R.sup.3, --SO.sub.2N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--C(O)N(R.sup.3).sub.2, --OC(O)NR(R.sup.3),
--OC(O)N(R.sup.3).sub.2, --NR.sup.3C(O)R.sup.3,
--NR.sup.3C(O)N(R.sup.3).sub.2, or --NR.sup.3CO.sub.2R.sup.3.
[0138] More preferably, R.sup.z2 of formula I is selected from an
optionally substituted ring selected from a 5-6 membered monocyclic
or an 9-10 membered bicyclic saturated, partially unsaturated, or
fully unsaturated ring having 0-4 heteroatoms independently
selected from nitrogen, oxygen or sulfur; wherein said ring is
optionally substituted by up to three substituents independently
selected as set forth above. Most preferably, R.sup.z2 of formula I
is selected from phenyl, imidazolyl, pyrazolyl, pyridyl,
pyridazinyl, pyrazinyl, naphthyl, tetrahydronaphthyl,
benzimidazolyl, benzthiazolyl, quinolinyl, quinazolinyl,
benzodioxinyl, isobenzofuran, indanyl, indolyl, indolinyl,
indazolyl, or isoquinolinyl, wherein the R.sup.z2 moiety of formula
I is optionally and independently substituted with up to three
substituents as set forth above.
[0139] Preferred substituents on R.sup.z2 of formula I, when
present, are independently selected from halogen, --CN, --NO.sub.2,
--C(O)R.sup.3, --CO.sub.2R.sup.3, --C(O)NR(R.sup.3),
--NR.sup.3C(O)R.sup.3, --N(R.sup.3).sub.2,
--N(R.sup.3)SO.sub.2R.sup.3, --NR.sup.3C(O)N(R.sup.3)- .sub.2, or
--NR.sup.3CO.sub.2R.sup.3. More preferred substituents on the
R.sup.z2 moiety of formula I are independently selected from --Cl,
--Br, --F, .sub.7CN, --CF.sub.3, --COOH, --CONHMe, --CONHEt,
--NH.sub.2, --NHAc, --NHSO.sub.2Me, --NHSO.sub.2Et,
--NHSO.sub.2(n-propyl), --NHSO.sub.2(isopropyl), --NHCOEt,
--NHCOCH.sub.2NHCH.sub.3, --NHCOCH.sub.2N(CO.sub.2t-Bu)CH.sub.3,
--NHCOCH.sub.2N(CH.sub.3).sub.2,
--NHCOCH.sub.2CH.sub.2N(CH.sub.3).sub.2,
--NHCOCH.sub.2CH.sub.2CH.sub.2N(- CH.sub.3).sub.2,
--NHCO(cyclopropyl), --NHCO(isopropyl), --NHCO(isobutyl),
--NHCOCH.sub.2(morpholin-4-yl),
--NHCOCH.sub.2CH.sub.2(morpholin-4-yl),
--NHCOCH.sub.2CH.sub.2CH.sub.2(morpholin-4-yl),
--NHCO.sub.2(t-butyl), --NH(cyclohexyl), --NHMe, --NMe.sub.2, --OH,
--OMe, methyl, ethyl, cyclopropyl, isopropyl, or t-butyl.
[0140] According to another preferred embodiment R.sup.z2 of
formula I has up to two substituents, wherein said substituents are
as set forth above. More preferably, R.sup.z2 of formula I has one
substituent, wherein said substituent is as set forth above. Most
preferably, R of formula I has one substituent selected from
--NR.sup.3C(O)R.sup.3, wherein each R.sup.3 is independently
selected from R or Ar and wherein R is hydrogen or an optionally
substituted C.sub.1-4 aliphatic group.
[0141] According to another embodiment, R.sup.z2 of formula I is
C.sub.1-6 aliphatic group substituted with 0-3 groups independently
selected from halogen, oxo, --CN, --NO.sub.2, --C(O)R.sup.3,
--CO.sub.2R.sup.3, --C(O)NR(R.sup.3), --NR.sup.3C(O)R.sup.3,
--N(R.sup.3).sub.2, --N(R.sup.3)SO.sub.2R.sup.3,
--NR.sup.3C(O)N(R.sup.3).sub.2, or --NR.sup.3CO.sub.2R.sup.3. More
preferably, R of formula I is a C.sub.1-4 aliphatic group
substituted with 0-3 groups independently selected from halogen,
--CN, --NO.sub.2, --C(O)R.sup.3, --CO.sub.2R.sup.3,
--N(R.sup.3).sub.2, or --NR.sup.3CO.sub.2R.sup.3.
[0142] Preferred embodiments of R.sup.x, T, Q, R.sup.z1, and
R.sup.z2 in formula II are as set forth for these moieties in
formula I.
[0143] Preferred embodiments of the R.sup.y moiety of R.sup.y--H
are as set forth for the R.sup.y group in formula I.
[0144] Preferred embodiments of R.sup.x, L.sup.3, T, and R.sup.z2
in formula III are as set forth for these moieties in formula
I.
[0145] Preferred embodiments of the Q and R.sup.z1 moieties of
R.sup.z1-Q-H are as set forth for these moieties in formula I.
[0146] Preferred embodiments R.sup.x, L.sup.3, L.sup.2 and Q in
formula IV are as set forth for these moieties in formula I.
[0147] Preferred embodiments of R.sup.z2 and T in R.sup.z2-T-H are
as set forth for these moieties in formula I.
[0148] Preferably R.sup.x in the processes of the present invention
is other than a suitable leaving group.
[0149] Preferred compounds of formula I, prepared using the
processes of the present invention, have formula I': 9
[0150] or a pharmaceutically acceptable derivative or salt thereof,
wherein R.sup.1 and R.sup.3 are as defined above.
[0151] Preferred R.sup.1 groups of formula I' are independently
selected from R, wherein R is hydrogen or an optionally substituted
C.sub.1-4 aliphatic group. Preferred substituents on the C.sub.1-4
aliphatic group of the R.sup.1 moiety of formula I' are selected
from --OR.sup.3, --SR.sup.3, --CN, --NO.sub.2, oxo, halogen,
--N(R.sup.3).sub.2, --C(O)R.sup.3, or a 3-6 membered aromatic or
non-aromatic ring having zero to two heteroatoms independently
selected from nitrogen, oxygen, or sulfur. More preferred
substituents on the C.sub.1-4 aliphatic group of the R.sup.1 moiety
of formula I' are 5-6 membered non-aromatic rings having 1-2
heteroatoms independently selected from nitrogen, oxygen, or
sulfur. Most preferred substituents on the R.sup.1 C.sub.1-4
aliphatic group of the R.sup.1 moiety of formula I' are
NH(CH.sub.3), NH.sub.2, OH, OCH.sub.3, morpholinyl, piperidinyl,
piperazinyl, pyrrolidinyl, and thiomorpholinyl.
[0152] According to another preferred embodiment, each R.sup.1 of
formula I' is R such that the two R groups are taken together to
form an optionally substituted 4-7 membered non-aromatic ring
having up to two additional heteroatoms independently selected from
nitrogen, oxygen, or sulfur. Preferred substituents on said ring
are selected from --R.sup.3, --OR.sup.3, --SR.sup.3, --CN,
--NO.sub.2, oxo, halogen, --N(R.sup.3).sub.2, --C(O)R.sup.3,
--CO.sub.2R.sup.3, --SO.sub.2R.sup.3, or a 3-6 membered aromatic or
non-aromatic ring having zero to two heteroatoms independently
selected from nitrogen, oxygen, or sulfur. More preferred
substituents said ring are selected from optionally substituted
C.sub.1-4 aliphatic, NH.sub.2, NH(C.sub.1-4 aliphatic), N(C.sub.1-4
aliphatic).sub.2, optionally substituted phenyl, CO.sub.2(C.sub.1-4
aliphatic), or SO.sub.2(C.sub.1-4 aliphatic). Most preferred
substituents on said ring are selected from methyl, ethyl,
methylsulfonyl, (CH.sub.2).sub.2SO.sub.2CH.sub.3, cyclopropyl,
CH.sub.2cyclopropyl, (CH.sub.2).sub.2OH, CO.sub.2t-butyl,
CH.sub.2phenyl, phenyl, NH.sub.2, NH(CH.sub.3), N(CH.sub.3).sub.2,
(CH.sub.2).sub.2NH.sub.2, (CH.sub.2).sub.2morpholin-4-yl,
(CH.sub.2).sub.2N(CH.sub.3).sub.2, isopropyl, propyl, t-butyl,
(CH.sub.2).sub.2CN, or (CH.sub.2).sub.2C(O)morpholin-4-yl.
[0153] More preferably, the ring formed by N(R.sup.1).sub.2 of
formula I' is pyrrolidinyl, piperidinyl, morpholin-4-yl,
thiomorpholin-4-yl, piperazin-1-yl, diazepanyl, or
tetrahydroisoquinolinyl, wherein each ring is optionally
substituted with one or two groups independently selected from
methyl, ethyl, methylsulfonyl, (CH.sub.2).sub.2SO.sub.2CH.sub.3,
cyclopropyl, CH.sub.2cyclopropyl, (CH.sub.2).sub.2OH,
CO.sub.2t-butyl, CH.sub.2phenyl, phenyl, NH.sub.2, NH(CH.sub.3),
N(CH.sub.3).sub.2, (CH.sub.2).sub.2NH.sub.2,
(CH.sub.2).sub.2morpholin-4-yl, (CH.sub.2).sub.2N(CH.sub.3).sub.2,
isopropyl, propyl, t-butyl, (CH.sub.2).sub.2CN, or
(CH.sub.2).sub.2C(O)morpholin-4-yl.
[0154] More preferred compounds within compounds of formula I
prepared using the processes of the present invention have formula
V: 10
[0155] or a pharmaceutically acceptable derivative or salt thereof,
wherein:
[0156] R.sup.5 is selected from hydrogen or C.sub.1-4
aliphatic;
[0157] R.sup.6 is selected from C.sub.1-3 aliphatic; and
[0158] R.sup.7 is selected from C.sub.1-4 aliphatic.
[0159] Preferred R.sup.5 groups of formula V are selected from
hydrogen, methyl, ethyl, t-butyl, propyl, cyclopropyl,
cyclopropylmethyl, or isopropyl. More preferred R.sup.5 groups of
formula V are selected from hydrogen or methyl. Most preferably
R.sup.5 of formula V is methyl.
[0160] Preferred R.sup.6 groups of formula V are selected from
methyl, ethyl, or cyclopropyl. More preferred R.sup.6 groups of
formula V are methyl of cyclopropyl. Most preferably, R.sup.6 of
formula V is methyl.
[0161] Preferred R.sup.7 groups of formula V are selected from
methyl, ethyl, t-butyl, or cyclopropyl. More preferred R.sup.7
groups of formula V are selected from ethyl or cyclopropyl. Most
preferably, R.sup.7 of formula V is cyclopropyl.
[0162] According to another embodiment, the present invention
relates to a compound of formula V: 11
[0163] or a pharmaceutically acceptable derivative or salt thereof,
wherein:
[0164] R.sup.5 is selected from hydrogen or C.sub.1-4
aliphatic;
[0165] R.sup.6 is selected from C.sub.1-3 aliphatic; and
[0166] R.sup.7 is selected from C.sub.1-4 aliphatic; provided that
said compound is other than
N-{4-[4-(4-methyl-piperazin-1-yl)-6-(5-methyl-2H-p-
yrazol-3-ylamino)-pyrimidin-2-ylsulfanyl]-phenyl}-propionamide.
[0167] Preferred R.sup.5 groups of formula V are selected from
hydrogen, methyl, ethyl, t-butyl, or isopropyl. More preferred
R.sup.5 groups of formula V are selected from hydrogen or methyl.
Most preferably R.sup.5 of formula V is methyl.
[0168] Preferred R.sup.6 groups of formula V are selected from
methyl, ethyl, or cyclopropyl. More preferred R groups of formula V
are methyl of cyclopropyl. Most preferably, R.sup.6 of formula V is
methyl.
[0169] Preferred R.sup.7 groups of formula V are selected from
methyl, ethyl, t-butyl, or cyclopropyl. More preferred R groups of
formula V are selected from ethyl or cyclopropyl. Most preferably,
R.sup.7 of formula V is cyclopropyl.
[0170] Compounds of formula V fall within the genus of compounds
described in PCT publication WO 02/057259. However, applicants have
discovered that the present compounds have surprising and
unexpectedly increased potency as inhibitors of Aurora protein
kinase and/or FLT-3 protein kinase.
[0171] Exemplary structures of formula V are set forth in Table 1
below.
1TABLE 1 No. V- Structure 1 12 2 13 3 14 4 15 5 16 6 17 7 18 8 19 9
20 10 21 11 22 12 23 13 24 14 25 15 26 16 27 17 28 18 29 19 30 20
31
[0172] Other exemplary compounds of formula I prepared according to
the processes of the present invention are set forth in Table 2
below.
2TABLE 2 Exemplary Compounds of Formula I 32 I-1 33 I-2 34 I-3 35
I-4 36 I-5 37 I-6 38 I-7 39 I-8 40 I-9 41 I-10 42 I-11 43 I-12 44
I-13 45 I-14 46 I-15 47 I-16 48 I-17 49 I-18 50 I-19 51 I-20 52
I-21 53 I-22 54 I-23 55 I-24 56 I-25 57 I-26 58 I-27 59 I-28 60
I-29 61 I-30 62 I-31 63 I-32 64 I-33 65 I-34 66 I-35 67 I-36 68
I-37 69 I-38 70 I-39 71 I-40 72 I-41 73 I-42 74 I-43 75 I-44 76
I-45 77 I-46 78 I-47 79 I-48 80 I-49 81 I-50 82 I-51 83 I-52 84
I-53 85 I-54 86 I-55 87 I-56 88 I-57 89 I-58 90 I-59 91 I-60 92
I-61
[0173] Preferably the processes of the present invention are used
to prepare a compound selected from Tables 1 and 2. More preferably
the processes of the present invention are used to prepare a
compound selected from Table 1.
[0174] According to an alternate embodiment, the present invention
provides a compound of formula II, formula III, or formula IV:
93
[0175] or a pharmaceutically acceptable salt thereof, wherein
R.sup.x, R.sup.y, L.sup.1, L.sup.2, L.sup.3, T, R.sup.z2, and Q,
and the preferred embodiments thereof, are as defined above.
[0176] According to a preferred embodiment, the present invention
provides an intermediate of formula II.
[0177] According to another preferred embodiment, the present
invention provides an intermediate of formula III.
[0178] According to yet another preferred embodiment, the present
invention provides an intermediate of formula IV.
[0179] According to another embodiment, the invention provides a
composition comprising a compound of this invention or a
pharmaceutically acceptable derivative thereof and a
pharmaceutically acceptable carrier, adjuvant, or vehicle. The
amount of compound in the compositions of this invention is such
that is effective to detectably inhibit a protein kinase,
particularly Aurora and/or FLT-3 kinase, in a biological sample or
in a patient. Preferably the composition of this invention is
formulated for administration to a patient in need of such
composition. Most preferably, the composition of this invention is
formulated for oral administration to a patient.
[0180] The term "patient", as used herein, means an animal,
preferably a mammal, and most preferably a human.
[0181] The term "pharmaceutically acceptable carrier, adjuvant, or
vehicle" refers to a non-toxic carrier, adjuvant, or vehicle that
does not destroy the pharmacological activity of the compound with
which it is formulated. Pharmaceutically acceptable carriers,
adjuvants or vehicles that may be used in the compositions of this
invention include, but are not limited to, ion exchangers, alumina,
aluminum stearate, lecithin, serum proteins, such as human serum
albumin, buffer substances such as phosphates, glycine, sorbic
acid, potassium sorbate, partial glyceride mixtures of saturated
vegetable fatty acids, water, salts or electrolytes, such as
protamine sulfate, disodium hydrogen phosphate, potassium hydrogen
phosphate, sodium chloride, zinc salts, colloidal silica, magnesium
trisilicate, polyvinyl pyrrolidone, cellulose-based substances,
polyethylene glycol, sodium carboxymethylcellulose, polyacrylates,
waxes, polyethylene-polyoxypropylene-block polymers, polyethylene
glycol and wool fat.
[0182] The term "detectably inhibit", as used herein means a
measurable change in protein kinase activity between a sample
comprising said composition and protein kinase and an equivalent
sample comprising protein kinase in the absence of said
composition.
[0183] A "pharmaceutically acceptable derivative or salt" means any
non-toxic salt, ester, salt of an ester or other derivative of a
compound of this invention that, upon administration to a
recipient, is capable of providing, either directly or indirectly,
a compound of this invention or an inhibitorily active metabolite
or residue thereof. As used herein, the term "inhibitorily active
metabolite or residue thereof" means that a metabolite or residue
thereof is also an inhibitor of Aurora and/or FLT-3 protein
kinase.
[0184] According to another embodiment, the present invention
provides processes for preparing a pharmaceutically acceptable salt
of compound of formula I, I', or V comprising the step of
converting a compound of formula I, I', or V prepared according to
the processes of the present invention into the desired
pharmaceutically acceptable salt. Such conversions are well known
in the art. See, generally, "Advanced Organic Chemistry," Jerry
March, 4.sup.th Ed., John Wiley and Sons, N.Y. (1992).
[0185] Pharmaceutically acceptable salts of the compounds of this
invention include those derived from pharmaceutically acceptable
inorganic and organic acids and bases. Examples of suitable acid
salts include acetate, adipate, alginate, aspartate, benzoate,
benzenesulfonate, bisulfate, butyrate, citrate, camphorate,
camphorsulfonate, cyclopentanepropionate, digluconate,
dodecylsulfate, ethanesulfonate, formate, fumarate,
glucoheptanoate, glycerophosphate, glycolate, hemisulfate,
heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide,
2-hydroxyethanesulfonate, lactate, maleate, malonate,
methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate,
oxalate, palmoate, pectinate, persulfate, 3-phenylpropionate,
phosphate, picrate, pivalate, propionate, salicylate, succinate,
sulfate, tartrate, thiocyanate, tosylate and undecanoate. Other
acids, such as oxalic, while not in themselves pharmaceutically
acceptable, may be employed in the preparation of salts useful as
intermediates in obtaining the compounds of the invention and their
pharmaceutically acceptable acid addition salts.
[0186] Salts derived from appropriate bases include alkali metal
(e.g., sodium and potassium), alkaline earth metal (e.g.,
magnesium), ammonium and N.sup.+(C.sub.1-4 alkyl).sub.4 salts. This
invention also envisions the quaternization of any basic
nitrogen-containing groups of the compounds disclosed herein. Water
or oil-soluble or dispersible products may be obtained by such
quaternization.
[0187] Table 3 below sets forth representative salts of compounds
of Formula V of the present invention.
3TABLE 3 Representative Salts of Compounds of Formula V V-1 i 94
V-1 ii 95 V-1 iii 96 V-1 iv 97 V-1 v 98 V-1 vi 99 V-1 vii 100 V-1
viii 101 V-1 ix 102 V-1 x 103 V-1 xi 104 V-20 i 105 V-20 ii 106
[0188] The compositions of the present invention may be
administered orally, parenterally, by inhalation spray, topically,
rectally, nasally, buccally, vaginally or via an implanted
reservoir. The term "parenteral" as used herein includes
subcutaneous, intravenous, intramuscular, intra-articular,
intra-synovial, intrasternal, intrathecal, intrahepatic,
intralesional and intracranial injection or infusion techniques.
Preferably, the compositions are administered orally,
intraperitoneally or intravenously. Sterile injectable forms of the
compositions of this invention may be aqueous or oleaginous
suspension. These suspensions may be formulated according to
techniques known in the art using suitable dispersing or wetting
agents and suspending agents. The sterile injectable preparation
may also be a sterile injectable solution or suspension in a
non-toxic parenterally-acceptable diluent or solvent, for example
as a solution in 1,3-butanediol. Among the acceptable vehicles and
solvents that may be employed are water, Ringer's solution and
isotonic sodium chloride solution. In addition, sterile, fixed oils
are conventionally employed as a solvent or suspending medium.
[0189] For this purpose, any bland fixed oil may be employed
including synthetic mono- or di-glycerides. Fatty acids, such as
oleic acid and its glyceride derivatives are useful in the
preparation of injectables, as are natural
pharmaceutically-acceptable oils, such as olive oil or castor oil,
especially in their polyoxyethylated versions. These oil solutions
or suspensions may also contain a long-chain alcohol diluent or
dispersant, such as carboxymethyl cellulose or similar dispersing
agents that are commonly used in the formulation of
pharmaceutically acceptable dosage forms including emulsions and
suspensions. Other commonly used surfactants, such as Tweens, Spans
and other emulsifying agents or bioavailability enhancers which are
commonly used in the manufacture of pharmaceutically acceptable
solid, liquid, or other dosage forms may also be used for the
purposes of formulation.
[0190] The pharmaceutically acceptable compositions of this
invention may be orally administered in any orally acceptable
dosage form including, but not limited to, capsules, tablets,
aqueous suspensions or solutions. In the case of tablets for oral
use, carriers commonly used include lactose and corn starch.
Lubricating agents, such as magnesium stearate, are also typically
added. For oral administration in a capsule form, useful diluents
include lactose and dried cornstarch. When aqueous suspensions are
required for oral use, the active ingredient is combined with
emulsifying and suspending agents. If desired, certain sweetening,
flavoring or coloring agents may also be added.
[0191] Alternatively, the pharmaceutically acceptable compositions
of this invention may be administered in the form of suppositories
for rectal administration. These can be prepared by mixing the
agent with a suitable non-irritating excipient that is solid at
room temperature but liquid at rectal temperature and therefore
will melt in the rectum to release the drug. Such materials include
cocoa butter, beeswax and polyethylene glycols.
[0192] The pharmaceutically acceptable compositions of this
invention may also be administered topically, especially when the
target of treatment includes areas or organs readily accessible by
topical application, including diseases of the eye, the skin, or
the lower intestinal tract. Suitable topical formulations are
readily prepared for each of these areas or organs.
[0193] Topical application for the lower intestinal tract can be
effected in a rectal suppository formulation (see above) or in a
suitable enema formulation. Topically-transdermal patches may also
be used.
[0194] For topical applications, the pharmaceutically acceptable
compositions may be formulated in a suitable ointment containing
the active component suspended or dissolved in one or more
carriers. Carriers for topical administration of the compounds of
this invention include, but are not limited to, mineral oil, liquid
petrolatum, white petrolatum, propylene glycol, polyoxyethylene,
polyoxypropylene compound, emulsifying wax and water.
Alternatively, the pharmaceutically acceptable compositions can be
formulated in a suitable lotion or cream containing the active
components suspended or dissolved in one or more pharmaceutically
acceptable carriers. Suitable carriers include, but are not limited
to, mineral oil, sorbitan monostearate, polysorbate 60, cetyl
esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and
water.
[0195] For ophthalmic use, the pharmaceutically acceptable
compositions may be formulated as micronized suspensions in
isotonic, pH adjusted sterile saline, or, preferably, as solutions
in isotonic, pH adjusted sterile saline, either with or without a
preservative such as benzylalkonium chloride. Alternatively, for
ophthalmic uses, the pharmaceutically acceptable compositions may
be formulated in an ointment such as petrolatum.
[0196] The pharmaceutically acceptable compositions of this
invention may also be administered by nasal aerosol or inhalation.
Such compositions are prepared according to techniques well-known
in the art of pharmaceutical formulation and may be prepared as
solutions in saline, employing benzyl alcohol or other suitable
preservatives, absorption promoters to enhance bioavailability,
fluorocarbons, and/or other conventional solubilizing or dispersing
agents.
[0197] Most preferably, the pharmaceutically acceptable
compositions of this invention are formulated for oral
administration.
[0198] The amount of the compounds of the present invention that
may be combined with the carrier materials to produce a composition
in a single dosage form will vary depending upon the host treated,
the particular mode of administration. Preferably, the compositions
should be formulated so that a dosage of between 0.01-100 mg/kg
body weight/day of the inhibitor can be administered to a patient
receiving these compositions.
[0199] It should also be understood that a specific dosage and
treatment regimen for any particular patient will depend upon a
variety of factors, including the activity of the specific compound
employed, the age, body weight, general health, sex, diet, time of
administration, rate of excretion, drug combination, and the
judgment of the treating physician and the severity of the
particular disease being treated. The amount of a compound of the
present invention in the composition will also depend upon the
particular compound in the composition.
[0200] Depending upon the particular condition, or disease, to be
treated or prevented, additional therapeutic agents, which are
normally administered to treat or prevent that condition, may also
be present in the compositions of this invention. As used herein,
additional therapeutic agents that are normally administered to
treat or prevent a particular disease, or condition, are known as
"appropriate for the disease, or condition, being treated".
[0201] For example, chemotherapeutic agents or other
anti-proliferative agents may be combined with the compounds of
this invention to treat proliferative diseases and cancer. Examples
of known chemotherapeutic agents include, but are not limited to,
Gleevec.TM., adriamycin, dexamethasone, vincristine,
cyclophosphamide, fluorouracil, topotecan, taxol, interferons, and
platinum derivatives. Other examples of agents the inhibitors of
this invention may also be combined with include, without
limitation: treatments for Alzheimer's Disease such as Aricept.RTM.
and Excelono; treatments for Parkinson's Disease such as
L-DOPA/carbidopa, entacapone, ropinrole, pramipexole,
bromocriptine, pergolide, trihexephendyl, and amantadine; agents
for treating Multiple Sclerosis (MS) such as beta interferon (e.g.,
Avonex.RTM. and Rebifo), Copaxone.RTM., and mitoxantrone;
treatments for asthma such as albuterol and Singulair agents for
treating schizophrenia such as zyprexa, risperdal, seroquel, and
haloperidol; anti-inflammatory agents such as corticosteroids, TNF
blockers, IL-1 RA, azathioprine, cyclophosphamide, and
sulfasalazine; immunomodulatory and immunosuppressive agents such
as cyclosporin, tacrolimus, rapamycin, mycophenolate mofetil,
interferons, corticosteroids, cyclophophamide, azathioprine, and
sulfasalazine; neurotrophic factors such as acetyl cholinesterase
inhibitors, MAO inhibitors, interferons, anti-convulsants, ion
channel blockers, riluzole, and anti-Parkinsonian agents; agents
for treating cardiovascular disease such as beta-blockers, ACE
inhibitors, diuretics, nitrates, calcium channel blockers, and
statins; agents for treating liver disease such as corticosteroids,
cholestyramine, interferons, and anti-viral agents; agents for
treating blood disorders such as corticosteroids, anti-leukemic
agents, and growth factors; and agents for treating
immunodeficiency disorders such as gamma globulin.
[0202] Further examples of chemotherapeutic agents or other
anti-proliferative agents that may be combined with the compounds
of the present invention to treat proliferative diseases and cancer
include, but are not limited to, For example, other therapies or
anticancer agents that may be used in combination with the
inventive anticancer agents of the present invention include
surgery, radiotherapy (in but a few examples, gamma-radiation,
neutron beam radiotherapy, electron beam radiotherapy, proton
therapy, brachytherapy, and systemic radioactive isotopes, to name
a few), endocrine therapy, biologic response modifiers
(interferons, interleukins, and tumor necrosis factor (TNF) to name
a few), hyperthermia and cryotherapy, agents to attenuate any
adverse effects (e.g., antiemetics), and other approved
chemotherapeutic drugs, including, but not limited to, alkylating
drugs (mechlorethamine, chlorambucil, Cyclophosphamide, Melphalan,
Ifosfamide), antimetabolites (Methotrexate), purine antagonists and
pyrimidine antagonists (6-Mercaptopurine, 5-Fluorouracil,
Cytarabile, Gemcitabine), spindle poisons (Vinblastine,
Vincristine, Vinorelbine, Paclitaxel), podophyllotoxins (Etoposide,
Irinotecan, Topotecan), antibiotics (Doxorubicin, Bleomycin,
Mitomycin), nitrosoureas (Carmustine, Lomustine), inorganic ions
(Cisplatin, Carboplatin), enzymes (Asparaginase), and hormones
(Tamoxifen, Leuprolide, Flutamide, and Megestrol), Gleevec.TM.,
adriamycin, dexamethasone, and cyclophosphamide. For a more
comprehensive discussion of updated cancer therapies see,
http:/Hwww.nci.nih.gov/, a list of the FDA approved oncology drugs
at http://www.fda.gov/cder/cancer/druglistframe.htm, and The Merck
Manual, Seventeenth Ed. 1999, the entire contents of which are
hereby incorporated by reference.
[0203] The amount of additional therapeutic agent present in the
compositions of this invention will be no more than the amount that
would normally be administered in a composition comprising that
therapeutic agent as the only active agent. Preferably the amount
of additional therapeutic agent in the presently disclosed
compositions will range from about 50% to 100% of the amount
normally present in a composition comprising that agent as the only
therapeutically active agent.
[0204] According to another embodiment, the invention relates to a
method of inhibiting Aurora-1, Aurora-2, Aurora-3, and/or FLT-3
kinase activity in a biological sample comprising the step of
contacting said biological sample with a compound of formula V, or
a composition comprising said compound.
[0205] The term "biological sample", as used herein, includes,
without limitation, cell cultures or extracts thereof; biopsied
material obtained from a mammal or extracts thereof; and blood,
saliva, urine, feces, semen, tears, or other body fluids or
extracts thereof.
[0206] Inhibition of Aurora-1, Aurora-2, Aurora-3, and/or FLT-3
kinase activity in a biological sample is useful for a variety of
purposes that are known to one of skill in the art. Examples of
such purposes include, but are not limited to, blood transfusion,
organ-transplantation, biological specimen storage, and biological
assays.
[0207] According to another embodiment, the invention relates to a
method of inhibiting Aurora-1 kinase activity in a patient
comprising the step of administering to sail patient a compound of
formula V, or a composition comprising said compound.
[0208] According to another embodiment, the invention relates to a
method of inhibiting Aurora-2 kinase activity in a patient
comprising the step of administering to said patient a compound of
formula V, or a composition comprising said compound.
[0209] According to another embodiment, the invention relates to a
method of inhibiting Aurora-3 kinase activity in a patient
comprising the step of administering to said patient a compound of
formula V, or a composition comprising said compound.
[0210] According to another embodiment, the invention relates to a
method of inhibiting FLT-3 kinase activity in a patient comprising
the step of administering to said patient a compound of formula V,
or a composition comprising said compound.
[0211] According to another embodiment, the invention relates to a
method of inhibiting Aurora-1, Aurora-2, Aurora-3, and FLT-3 kinase
activity in a patient comprising the step of administering to said
patient a compound of formula V, or a composition comprising said
compound.
[0212] According to another embodiment, the invention provides a
method for treating or lessening the severity of an Aurora-mediated
disease or condition in a patient comprising the step of
administering to said patient a compound of formula V, or
composition comprising said compound.
[0213] The term "Aurora-mediated disease", as used herein, means
any disease or other deleterious condition or disease in which an
Aurora family protein kinase is known to play a role. Such diseases
or conditions include, without limitation, melanoma, leukemia, or a
cancer selected from colon, breast, gastric, ovarian, cervical,
melanoma, renal, prostate, lymphoma, neuroblastoma, pancreatic,
leukemia and bladder.
[0214] According to another embodiment, the present invention
relates to a method of treating cancer in a patient, comprising the
step of administering to said patient a compound of formula V or
composition thereof.
[0215] According to another embodiment, the present invention
relates to a method of treating melanoma, lymphoma, neuroblastoma,
leukemia, or a cancer selected from colon, breast, lung, kidney,
ovary, pancreatic, renal, CNS, cervical, prostate, or cancer of the
gastric tract in a patient, comprising the step of administering to
said patient a compound of formula V or composition thereof.
[0216] According to another embodiment, the present invention
relates to a method of treating acute-myelogenous leukemia (ANL),
acute lymphocytic leukemia (ALL), mastocytosis or gastrointestinal
stromal tumor (GIST) in a patient, comprising the step of
administering to said patient a compound of formula V or
composition thereof.
[0217] Another aspect of the present invention relates to the
disruption of mitosis of cancer cells in a patient, comprising the
step of administering to said patient a compound of formula V or
composition thereof.
[0218] According to another embodiment, the present invention
relates to a method of treating or lessening the severity of a
cancer in a patient comprising the step of disrupting mitosis of
the cancer cells by inhibiting Aurora-1, Aurora-2, and/or Aurora-3
with a compound of formula V or composition thereof.
[0219] In an alternate embodiment, the methods of this invention
that utilize compositions that do not contain an additional
therapeutic agent, comprise the additional step of separately
administering to said patient an additional therapeutic agent. When
these additional therapeutic agents are administered separately
they may be administered to the patient prior to, sequentially with
or following administration of the compositions of this
invention.
[0220] In order that the invention described herein may be more
fully understood, the following examples are set forth. It should
be understood that these examples are for illustrative purposes
only and are not to be construed as limiting this invention in any
manner.
EXAMPLES
[0221] 107
Example 1
[0222] 4,6-Dichloropyrimidine-2-methylsulfone (A): Prepared by
methods substantially similar to those set forth in Koppell et al,
JOC, 26, 1961, 792, in the following manner. To a stirred solution
of 4,6-dichloro-2-(methylthio)pyrimidine (50 g, 0.26 mol) in
dichloromethane (1 L) at 0.degree. C. was added
meta-chloroperoxybenzoic acid (143.6 g, 0.64 mol) over a period of
20 minutes. The solution was allowed to warm to room temperature
and was stirred for 4 hours. The mixture was diluted with
dichloromethane (1.5 L) and then treated sequentially with 50%
Na.sub.2S.sub.2O.sub.3/NaHCO.sub.3 solution (2.times.200 ml), sat.
NaHCO.sub.3 solution (4.times.300 ml), and brine (200 ml) then
dried (MgSO.sub.4). The solvent was removed in vacuo to afford an
off-white solid which was redissolved in EtOAc (1L) and treated
sequentially with sat. NaHCO.sub.3 solution (3.times.300 ml), and
brine (100 ml) then dried (MgSO.sub.4). The solvent was removed in
vacuo to afford the title compound (A) as a white solid (55.6 g,
96% yield). .sup.1H NMR CDCl.sub.3 .delta. 3.40 (3H, s, CH3), 7.75
(1H, s. ArH).
Example 2
[0223] Cyclopropane carboxylic acid
[4-(4,6-dichloro-pyrimidin-2-ylsulphan- yl)-phenyl]-amide (C): A
suspension of compound A (10 g, 44.04 mmol) and cyclopropane
carboxylic acid (4-mercapto-phenyl)-amide (B, 8.51 g, 44.04 mmol)
in t-butanol (300 ml) was degassed by evacuation, then flushing
with nitrogen. The mixture was stirred at 90.degree. C under
nitrogen atmosphere for 1 hour then the solvent was removed in
vacuo. The residue was dissolved in ethyl acetate (600 ml) and
washed with an aqueous solution of potassium carbonate and sodium
chloride. The organic extract was dried over magnesium sulphate,
concentrated to a low volume and allowed to crystallize. The
product C was collected as colourless crystals, (11.15 g, 74%).
.sup.1H-NMR DMSO-d.sup.6, .delta. 0.82-0.89 (4H, m), 1.80-1.88 (1H,
m), 7.55 (2H, d), 7.70-7.76 (3H, m), 10.49 (1H, s); M+H, 340.
Example 3
[0224] Cyclopropane carboxylic
acid{4-[4-chloro-6-(5-methyl-2H-pyrazol-3-y-
lamino)-pyrimidin-2-ylsulphanyl]-phenyl} amide (D): A mixture of
compound C (1.0 g, 2.94 mmol)and 3-amino-5-methylpyrazole (314 mg,
3.23 mmol) in dimethylformamide (6 ml) was treated with
diisopropylethylamine (0.614 ml, 3.53 mmol) and sodium iodide (530
mg, 3.53 mmol). The mixture was stirred under nitrogen at
85.degree. for 4 hours, cooled to room temperature and diluted with
ethyl acetate. The solution was washed with water (.times.4), dried
over magnesium sulphate and concentrated to 5 ml to afford, upon
crystallization and harvesting of colourless crystals, the title
compound D (920 mg, 78%). .sup.1H-NMR DMSO-d.sup.6, .delta.
0.80-0.87 (4H, m), 1.77-1.85 (1H, m), 1.92 (1H, s), 5.24 (1H, br
s), 6.47 (1H, br s), 7.55 (2H, d), 7.70-7.80 (2H, m), 10.24 (1H,
s), 10.47 (1H, s), 11.92 (1H, s).
Example 4
[0225] Cyclopropane carboxylic acid
{4-[4-(4-methyl-piperazin-1-yl)-6-(5-m-
ethyl-2H-pyrazol-3-ylamino)-pyrimidin-2-ylsulphanyl]-phenyl}-amide
(V-1): Compound D (2.373 g, 5.92 mmol) was treated with
N-methylpiperazine (10 ml) and the mixture stirred at 110.degree.
for 2 hours. The excess N-methylpiperazine was removed in vacuo
then the residue was dissolved in ethyl acetate, washed with
aqueous sodium bicarbonate solution, dried over magnesium sulphate,
and concentrated. The residue was crystallised from methanol to
give colourless crystals of desired product V-1 (1.82 g, 66%),
.sup.1H-NMR DMSO-d.sup.6, .delta. 0.81 (4H, d), 1.79 (1H, m), 2.01
(3H, s), 2.18 (3H, s), 2.30 (4H, m), 3.35 (masked signal), 5.42
(1H, s), 6.02 (1H, br s), 7.47 (2H, d), 7.69 (2H, d), 9.22 (1H, s),
10.39 (1H, s), 11.69 (1H, s).
Example 5
[0226] 108
[0227]
N-{4-[4-(5-Methyl-2H-pyrazol-3-ylmethyl)-6-(4-propyl-piperazin-1-yl-
)-pyrimidin-2-ylsulfanyl]-phenyl}-propionamide (V-5): Ethane
carboxylic acid
{4-[4-chloro-6-(5-methyl-2H-pyrazol-3-ylamino)-pyrimidin-2-ylsulphan-
yl]-phenyl} amide (119 mg, 0.306 mmol, prepared by methods
analogous to those set forth in Examples 1, 2, and 3) in n-BuOH (5
mL) was treated with N-propylpiperazine dihydrobromide (887 mg,
3.06 mmol) followed by diisopropylethylamine (1.066 mL, 6.12 mmol).
The resulting mixture was stirred at 110.degree. for 20 hours. The
solvent was removed under reduced pressure, and the residue was
purified using preparative HPLC to afford the title compound.
.sup.1H NMR (DMSO): .delta. 1.10 (3H, t), 2.05 (3H, s), 2.35 (2H,
d), 3.30 (4H, s), 3.70 (4H, s), 5.45 (1H, s), 6.05 (1H, br s), 7.45
(2H, d), 7.70 (2H, d), 9.20 (1H, s), 10.05 (1H, s), 11.70 (1H, br
s).
Example 6
[0228] 109
[0229] N-[4-(4,6-Dichloro-pyrimidin-2-yloxy)-phenyl]-acetamide:_A
solution of 4-acetamidophenol (666 mg, 4.40 mmol) in anhydrous THF
(40 ml), stirring at ambient temperature, was treated with a 60%
dispersion of sodium hydride in mineral oil (176 mg, 4.40 mmol).
The reaction mixture was then allowed to stir for 30 minutes at
ambient temperature before
4,6-dichloro-2-methanesulfonyl-pyrimidine (1.0 g, 4.40 mmol) was
added. The reaction was then allowed to stir for a further 3 hours
before the reaction was diluted with saturated aqueous NH.sub.4Cl
and EtOAc. The organic layer was separated, washed with saturated
aqueous NaCl and dried over sodium sulfate then concentrated in
vacuo. The residue was purified by column chromatography (Silica
Gel, MeOH:CH.sub.2Cl.sub.2, 5:95) yield the title compound 1.25 g,
(95%) as a solid. .sup.1H-NMR (400 MHz, DMSO-d.sub.6): .delta. 2.06
(3H, s), 7.18 (2H, d, J=8.5 Hz), 7.62 (2H, d, J=8.5 Hz), 10.05 (1H,
s), LC-MS: ES+=298.16, ES-=296.18).
Example 7
[0230] Cyclopropanecarboxylic acid
{4-[4-(4-methyl-4-oxy-piperazin-1-yl)-6-
-(5-methyl-2H-pyrazol-3-ylamino)-pyrimidin-2-ylsulfanyl]-phenyl}-amide
(V-19): Compound V-1 (1 g, 2.1 mmol) was suspended in
dichloromethane (20 mL), cooled to 0.degree. C. and treated with a
dichloromethane solution of mCPBA in 10 aliquots at 10 minute
intervals (each aliquot consisting of 100 mg, 0.44 mmol in 1 ml
DCM). Each time an aliquot was added the solution turned brown and
gradually returned to a yellow colour as the mCPBA was consumed.
Once all the starting material had been consumed, the solvent was
removed in vacuo and the resulting orange oil was purified by
preparative HPLC to give the title compound as an off-white solid
(69 mg, 7%); .sup.1H NMR (DMSO-d.sub.6): 0.85-0.91 (4H, m), 1.90
(1H, m), 2.10 (3H, s), 3.10-3.17 (2H, m), 3.25 (3H, s), 3.50-3.66
(4H, m), 3.98 (2H, d), 5.50 (1H, s), 6.11 (1H, br s), 7.56 (2H, d),
7.80 (2H, d), 9.42 (1H, s), 10.50 (1H, s), 11.82 (1H, br s).
Example 8
[0231] Cyclopropane carboxylic acid
{4-[4-(4-methyl-piperazin-1-yl)-6-(5-m-
ethyl-2H-pyrazol-3-ylamino)-pyrimidin-2-ylsulphanyl]-phenyl}-amide
methanesulfonate (V-1ii): Compound V-1 (515 mg, 1.11 mmol) was
suspended in ethanol (80 mL) and heated to reflux. To the clear
solution was added methanesulfonic acid (106 mg, 1.11 mmol) and the
reaction mixture was refluxed for a further 10 minutes. The mixture
was allowed to cool to room temperature and the solvent was
evaporated until a precipitate began to form. The mixture was then
cooled to 0.degree. C. and the resulting precipitate collected by
filtration before being dried under vacuum to afford the title
compound as a white solid (290 mg, 47%); .sup.1H NMR (400 MHz,
DMSO-d.sub.6) 0.81-0.82 (4H, d), 1.82 (1H, m), 2.36 (6H, s), 2.83
(3H, d), 3.03-3.12 (4H, m), 3.40-3.47 (2H, m), 3.79 (br s, OH),
4.14-4.18 (2H, m), 5.50 (1H, s), 6.05 (1H, s), 7.49 (2H, d), 7.72
(2H, d), 9.61 (1H, s), 10.41 (1H, br s), 10.80 (1H, s)
Example 9
[0232] The following compounds set forth in Table 4 below were
prepared according to the processes of the present invention and by
methods substantially similar to those set forth in Examples 1-8
above. The characterization data for these compounds is summarized
in Table 4 below and includes .sup.1H NMR, melting point (m.p.),
and mass spectral (MS) data.
[0233] Unless otherwise indicated each annotated .sup.1H NMR, set
forth in Table 4, was obtained at 400 MHz in deuterated
dimethylsulfoxide (dmso-d.sub.6).
4TABLE 4 Characterization Data for Representative Compounds Mass
No. Structure m.p. .sup.1H-NMR Spec V-2 110 -- 1.09(3H, t),
2.00(3H, s), 2.34(2H, q), 2.72(4H, m), 3.3(masked signal), 5.42(1H,
s), 5.98(1H, s), 7.47(2H, d), 7.69(2H, d), 9.20(1H, s), 10.07(1H,
s), 11.69(1H, s) ES+439.4 ES-437.4 V-3 111 178-181 C. 1.24(9H, s),
1.98(3H, s), 2.68-2.70(4H, m), 3.31(4H, masked signal), 5.35(1H,
s), 5.96(1H, br s), 7.47(2H, d), 7.79(2H, d), 9.20(1H, s), 9.33(1H,
s), 11.66(1H, s) ES+467.35 ES-465.38 V-4 112 -- 0.97(6H, d),
1.10(3H, t), 2.00(3H, s), 2.35(2H, q), 2.45(4H, br s), 2.65(1H, br
s), 3.35(4H, br s), 5.40(1H, s), 6.00(1H, br s), 7.50(2H, d),
7.70(2H, d), 9.20(1H, s), 10.10(1H, s), 11.70(1H, br s) ES+481.4
V-6 113 -- 1.01(3H, t), 1.09(3H, t), 2.00(3H, s), 2.31-2.37(8H, m),
3.35(masked signal), 5.42(1H, s), 6.01(1H, br s), 7.47(2H, d),
7.70(2H, d), 9.22(1H, s), 10.07(1H, s), 11.69(1H, s) ES+467.3
ES-465.4 V-7 114 -- 0.80-0.82(4H, m), 1.81(1H, m), 2.01(3H, s),
2.68(4H, m), 3.1-3.5(5H, m), 5.43(1H, s), 5.99(1H, br s), 7.47(2H,
d), 7.69(2H, d), 9.21(1H, s), 10.40(1H, s), 11.70(1H, s) ES+451.3
ES-449.4 V-8 115 -- 0.08(2H, m), 0.46(2H, m), 0.84(1H, m), 1.09(3H,
t), 2.00(3H, s), 2.19(2H, d), 2.34(2H, q), 2.44(4H, m), 3.35(masked
signal), 5.41(1H, s), 6.00(1H, br s), 7.47(2H, d), 7.70(2H, d),
9.23(1H, s), 10.08(1H, s), 11.66(1H, s) # ES+493.4 ES-491.4 V-9 116
-- 0.33(2H, m), 0.42(2H, m), 1.09(3H, t), 1.62(1H, m), 2.00(3H, s),
2.34(2H, q), 2.53(4H, m), 3.32(masked signal), 5.42(1H, s),
6.00(1H, br s), 7.47(2H, d), 7.70(2H, d), 9.22(1H, s), 10.07(1H,
s), 11.69(1H, s) ES+479.4 ES-477.4 V-10 117 161-163 1.01(9H, s),
1.09(3H, t), 2.00(3H, s), 2.34(2H, q), 2.5(masked signal),
3.36(masked signal), 5.42(1H, s), 5.98(1H, brs), 7.47(2H, d),
7.70(2H, d), 9.21(1H, s), 10.08(1H, s), 11.69(1H, s) ES+495.4
ES-493.4 V-11 118 -- 0.80-0.82(4H, m), 1.01(3H, t), 1.81(1H, m),
2.01(3H, s), 2.32-2.37(6H, m), 3.35(masked signal), 5.43(1H, s),
6.01(1H, br s), 7.47(2H, d), 7.69(2H, d), 9.23(1H, s), 10.40(1H,
s), 11.69(1H, s), ES+479.3 ES-477.4 V-12 119 -- 0.48-0.59(2H, m),
1.75-1.87(2H, m), 1.08(3H, t, J=7.5 Hz), 1.61-1.75(1H, m), 2.32(2H,
q, J=7.5 Hz), 2.61-2.71(4H, m), 3.20-3.30(4H, m), 5.47(1H, s),
6.10(1H, brs), 7.47(2H, d, J=8.4 Hz), 7.70(2H, d, J=8.4 Hz), #
9.20(1H, brs), 10.13(1H, s), 11.74(1H, brs) ES+465.34 ES-463.37
V-13 120 -- 0.81-0.82(4H, m), 1.01(3H, t), 1.05(3H, t), 1.81(1H,
m), 2.26-2.38(8H, m), 3.35(masked signal), 5.44(1H, s), 6.03(1H, br
s), 7.47(2H, d), 7.70(2H, d), 9.25(1H, s), 10.39(1H, s), 11.74(1H,
s) ES+493.4 ES-491.4 V-14 121 -- 0.54(2H, m), 0.79-0.82(6H, m),
1.01(3H, t), 1.69(1H, m), 1.82(1H, m), 2.32-2.36(6H, m),
3.35(masked signal), 5.45(1H, s), 6.07(1H, br s), 7.47(2H, d),
7.70(2H, d), 9.23(1H, s), 10.38(1H, s), 11.70(1H, s) ES+505.4
ES-503.4 V-15 122 -- 0.49-0.59(2H, m), 0.76-0.85(2H, m), 1.08(3H,
t, J=7.5 Hz), 1.63-1.72(1H, m), 2.19(3H, s), 2.23-2.38(6H, m),
3.30-3.43(4H, m), 5.50(1H, s), 6.15(1H, brs), 7.48(2H, d, J=8.6
Hz), 7.70(2H, d, J=8.6 Hz), 9.23(1H, brs), # 10.04(1H, s),
11.71(1H, brs). ES+479.34 ES-477.37 V-16 123 -- 2.02(3H, s),
2.07(3H, s), 2.18(3H, s), 2.30(4H, m), 3.35(masked signal),
5.44(1H, s), 6.03(1H, br s), 7.47(2H, d), 7.67(2H, d), 9.23(1H, s),
10.14(1H, s), 11.71(1H, s) ES+439.3 ES-437.4 V-17 124 -- 1.23(9H,
s), 1.97(3H, s), 2.20(3H, s), 2.30-2.33(4H, m), 3.31(4H, masked
signal), 5.37(1H, s), 5.96(1H, br s), 7.47(2H, d), 7.79(2H, d),
9.24(1H, s), 9.38(1H, s), 11.67(1H, brs) ES+481.4 ES-479.4 V-18 125
-- 0.76-0.91(4H, m), 1.00-1.18(3H, m), 1.76-1.86(1H, m), 2.18(3H,
s), 2.22-2.43(6H, m), 3.3-3.4(4H obscured), 5.46(1H, s), 6.08(1H,
br s), 7.49(2H, d), 7.72(2H, d), 9.30(1H, s), 10.40(1H, s),
11.72(1H, s) ES+479.3 ES-477.3 V-20 126 137.5-138.9 1.10(3H, t),
2.00(2H, s), 2.18(3H, s), 2.28-2.36(5H, m), 2.98(2H, br s),
3.32(4H, m), 5.40(1H, s), 6.05(1H, br s), 7.45(2H, d), 7.70(2H, d),
9.20(1H, s), 10.10(1H, s), 11.70(1H, s) MS+453.5 (M +H)+ I-1 127
238-239 1.10(3H, t), 2.00(3H, s), 2.45(2H, q), 3.65(4H, s),
5.45(1H, s), 6.05(1H, br s), 7.50(2H, d), 7.80(2H, d), 9.25(1H, s),
10.05(1H, s), 11.70(1H, br s) MS 440.3 (M +H)+ I-2 128 -- 1.09(3H,
t), 2.00(3H, s), 2.34(2H, q), 2.59(2H, m), 3.04(2H, m), 3.3(masked
signal), 5.39(1H, s), 5.77(1H, br s), 6.85(1H, s), 7.47(2H, d),
7.69(2H, d), 9.07(1H, s), 10.07(1H, s), 11.63(1H, br s) ES+413.3
ES-411.4 I-3 129 -- 1.09(3H, t), 1.35-1.37(2H, m), 1.44-1.46(4H,
m), 2.03(3H, s), 2.26(6H, m), 2.33(2H, q), 3.13(2H, m), 5.45(1H,
s), 5.84(1H, br s), 6.75(1H, br s), 7.46(2H, d), 7.68(2H, d),
9.05(1H, s), 10.05(1H, s), 11.65(1H, brs) ES+481.3 ES-479.4 I-4 130
-- 2.01(3H, s), 2.07(3H, s), 2.34-2.44(12H, m), 3.3(masked signal),
3.55(4H, m), 5.43(1H, s), 6.02(1H, br s), 7.47(2H, d), 7.67(2H, d),
9.22(1H, s), 10.14(1H, s), 11.70(1H, s) ES+538.3 ES-536.4 I-5 131
-- 1.06(3H, t), 1.98(3H, s), 2.20(3H, s), 2.31(2H, q), 2.50(masked
signal), 2.84(3H, s), 3.38(2H, m), 5.42(1H, s), 5.77(1H, s),
7.45(2H, d), 7.65(2H, d), 9.00(1H, s), 10.18(1H, s), 11.68(1H, br
s) ES+441.3 ES-439.4 I-6 132 -- 1.10(3H, t), 1.45(4H, s), 1.60(2H,
s), 2.00(3H, s), 2.35(2H, q), 3.35(4H, s), 5.40(1H, s), 6.05(1H, br
s), 7.50(2H, d), 7.70(2H, d), 9.15(1H, s), 10.05(1H, s), 11.80(1H,
br s) ES+438.3 I-7 133 -- 1.10(3H, t), 1.70(2H, s), 2.05(3H, s),
2.35(2H, q), 2.70(2H, s), 2.75(2H, s), 3.45(2H, s), 5.50(1H, s),
6.00(1H, br s), 7.50(2H, d), 7.70(2H, d), 9.15(1H, s), 10.10(1H,
s), 11.70(1H, br s) ES+453.3 I-8 134 -- 1.10(3H, t), 2.05(3H, s),
2.15(6H, s), 2.3-2.4(4H, m), 3.15(2H, s), 5.40(1H, s), 5.85(1H, br
s), 6.75(1H, br s), 7.45(2H, d), 7.70(2H, d), 9.05(1H, br s),
10.10(1H, s), 11.65(1H, brs) ES+441.3 I-9 135 -- 1.09(3H, t),
1.78(2H, m), 2.03(3H, s), 2.22(3H, s), 2.33(2H, q), 2.41(4H, m),
3.3(masked signal), 3.50(2H, m), 5.48(1H, s), 5.97(1H, br s),
7.46(2H, d), 7.68(2H, d), 9.14(1H, s), 10.06(1H, s), 11.70(1H, s)
ES+467.4 ES-465.4 I-10 136 -- 1.09(3H, t), 1.94(3H, s), 2.20(3H,
s), 2.30-2.38(6H, m), 3.42(4H, m), 6.94(1H, s), 7.49(2H, d),
7.69(2H, d), 7.95(1H, s), 9.27(1H, s), 10.07(1H, s) ES+470.2
ES-468.3 I-11 137 -- 1.10(3H, t), 2.05(3H, s), 2.35(2H, d),
3.30(4H, s), 3.70(4H, s), 5.45(1H, s), 6.05(1H, br s), 7.45(2H, d),
7.70(2H, d), 9.20(1H, s), 10.05(1H, s), 11.70(1H, br s) ES+456.2
I-12 138 -- 1.09(3H, t), 2.00(3H, s), 2.25-2.41(8H, m),
3.35(partially masked signal), 3.51(2H, m), 4.45(1H, m), 5.42(1H,
s), 6.00(1H, br s), 7.47(2H, d), 7.70(2H, d), 9.22(1H, s),
10.08(1H, s), 11.70(1H, s) ES+483.4 ES-481.4 I-13 139 -- 1.10(3H,
t), 1.23(2H, q), 1.37(9H, s), 1.70(2H, d), 2.00(3H, s), 2.35(2H,
q), 2.83(2H, t), 3.47(1H, m), 3.95(2H, d), 5.45(1H, s), 6.05(1H, br
s), 6.85(1H, d), 7.50(2H, d), 7.70(2H, d), 9.20(1H, s), 10.10(1H,
s), 11.70(1H, br s) ES+553.4 I-14 140 -- 0.46-0.58(2H, m),
0.78-0.89(2H, m), 1.08(3H, t, J=7.5 Hz), 1.62-1.72(1H, m),
2.21-2.43(6H, m), 3.23-3.40(4H, m), 3.50(2H, s), 5.48(1H, s),
6.10(1H, brs), 7.19-7.36(5H, m), 7.46(2H, d, J=8.5 Hz), 7.68(2H, d,
J=8.5 Hz), # 9.21(1H, s), 10.03(1H, s), 11.70(1H, brs) ES+555.34
ES-553.40 I-15 141 -- 1.07(3H, t), 1.12-1.22(2H, m), 1.70(2H, d),
2.02(3H, s), 2.35(2H, q), 2.80-2.90(3H, m), 3.95(2H, d), 5.45(1H,
s), 6.00(1H, br s), 7.45(2H, d), 7.70(2H, d), 9.20(1H, s),
10.15(1H, s), 11.75(1H, br s) ES+453.3 I-16 142 -- 1.00(3H, d),
1.08(3H, t), 2.00(3H, s), 2.35(2H, q), 2.55-2.90(3H, m),
3.65-4.25(5H, m), 5.45(1H, s), 6.00(1H, br s), 7.45(2H, d),
7.70(2H, d), 9.25(1H, br s), 10.20(1H, s), 11.70(1H, br s) ES+453.3
I-17 143 -- 0.47-0.55(2H, m), 0.72-0.81(2H, m), 1.08(3H, t, J=7.5
Hz), 1.41(9H, s), 1.62-1.73(1H, m), 2.32(2H, q, J=7.5 Hz),
3.30-3.41(8H, m), 5.48(1H, s), 6.10(1H, brs), 7.47(2H, d, J=8.5
Hz), 7.70(2H, d, J=8.5 Hz), 9.29(1H, brs), 10.05(1H, s), #
11.74(1H, brs). ES+565.33 ES-563.36 I-18 144 -- 1.10(3H, t),
2.01(3H, s), 2.35(2H, q), 3.16-3.18(4H, m), 3.52-3.54(4H, m),
5.43(1H, s), 6.08(1H, br s), 6.891(1H, t), 6.97(2H, d), 7.23(2H,
t), 7.49(2H, d), 7.71(2H, d), 9.28(1H, s), 10.09(1H, s), 11.72(1H,
s) ES+515.3 ES-513.4 I-19 145 151-152 0.95(6H, s), 1.10(3H, t),
2.05(3H, s), 2.20(2H, t), 2.35(2H, q), 2.60(2H, br s), 3.80(2H, br
s), 5.50(1H, s), 6.05(1H, br s), 7.50(2H, d), 7.70(2H, d), 9.15(1H,
s), 10.05(1H, s), 11.70(1H, br s) ES+467.3 I-20 146 159-160
1.10(3H, t), 1.75(1H, br s), 2.00(3H, s), 2.30-2.40(3H, m),
2.65(1H, m), 3.25-3.45(3H, m), 3.60(1H, br s), 5.45(1H, s),
5.80(1H, br s), 7.50(2H, d), 7.70(2H, d), 9.15(1H, br s), 10.05(1H,
s), 11.70(1H, br s) ES+439.3 I-21 147 -- 0.50-0.58(2H, m),
0.78-0.85(2H, m), 0.90(3H, d, J=6.1 Hz), 0.95-1.05(2H, m), 1.09(3H,
t, J=7.6 Hz), 1.51-1.64(3H, m), 1.66-1.75(1H, m), 2.32(2H, q, J=7.5
Hz), 2.66-2.78(2H, m), 3.96-4.08(2H, m), 5.48(1H, s), # 6.16(1H,
brs), 7.48(2H, q, J=8.6 Hz), 7.69(2H, d, J=8.6 Hz), 9.18(1H, brs),
10.04(1H, s), 11.74(1H, brs) ES+478.37 ES-476.39 I-22 148 --
0.80-0.81(4H, m), 1.23-1.38(6H, m), 1.82(1H, m), 2.04(3H, s),
2.34(6H, m), 3.17(2H, m), 5.47(1H, s), 5.86(1H, br s), 6.80(1H, br
s), 7.46(2H, d), 7.69(2H, d), 9.07(1H, s), 10.41(1H, s), 11.65(1H,
br s) ES+493.4 ES-491.4 I-23 149 -- 0.80-0.82(4H, m), 1.81(1H, m),
2.04(3H, s), 2.28(6H, m), 3.15(2H, m), 3.53(4H, m), 5.48(1H, s),
5.89(1H, br s), 6.81(1H, br s), 7.46(2H, d), 7.68(2H, d), 9.06(1H,
s), 10.38(1H, s), 11.66(1H, brs) ES+495.4 ES-493.4 I-24 150 --
1.10(3H, t), 1.47(2H, q), 1.90(2H, d), 2.03(3H, s), 2.35(2H, q),
2.85(2H, br s), 3.23(2H, d), 5.45(1H, s), 5.90(1H, br s), 7.05(1H,
d), 7.50(2H, d), 7.70(2H, d), 8.30(1H, br s), 8.55(1H, br s),
9.10(1H, s), 10.10(1H, s), 11.70(1H, br s) ES+453.3 I-25 151 --
0.83(4H, m), 1.82(1H, m), 2.22(3H, s), 2.89(4H, m), 3.33(4H, m)
(masked), 5.81(1H, s), 6.24(1H, br s), 7.36(1H, s), 7.48(2H, d),
7.65(2H, d), 9.32(1H, br s), 10.35(1H, s), 12.10(1H, br s) ES-449.4
ES+451.3 I-26 152 -- 0.81-0.83(4H, m), 1.81(1H, m), 3.29-3.31(4H,
m), 3.59-3.61(4H, m), 5.82(1H, s), 6.22(1H, br s), 7.36(1H, s),
7.48(2H, d), 7.64(2H, d), 9.38(1H, s), 10.37(1H, s), 12.10(1H, s)
ES+438.3 ES-436.4 I-27 153 -- 0.8-0.83(4H, m), 1.81(1H, m),
2.37-2.41(6H, m), 3.3(masked signal), 3.50(2H, m), 4.44(1H, s),
5.81(1H, s), 6.23(1H, br s), 7.36(1H, s), 7.47(2H, d), 7.65(2H, d),
9.32(1H, s), 10.38(1H, s), 12.10(1H, s) ES+481.3 ES-479.4 I-28 154
-- 0.81-0.83(4H, m), 0.96(6H, d), 1.81(1H, m), 2.41-2.43(4H, m),
2.65(1H, m), 3.3(masked signal), 5.82(1H, s), 6.24(1H, br s),
7.36(1H, s), 7.47(2H, d), 7.65(2H, d), 9.31(1H, s), 10.37(1H, s),
12.10(1H, s) ES+479.3 ES-477.4 I-29 155 220-222 -- M + H 479 I-30
156 -- 0.77-0.88(4H, m), 1.28-1.55(6H, m), 1.76-1.88(1H, m),
2.12-2.43(6H, m), 3.05-3.17(2H, m), 5.81(1H, brs), 6.04(1H, brs),
6.84(1H, brs), 7.39(1H, brs), 7.47(2H, d, J=8.6 Hz), 7.65(2H, d,
J=8.6 Hz), 9.12(1H, brs), 10.33(1H, s), 12.06(1H, brs) # ES+479.35
ES-477.41 I-31 157 -- 0.78-0.89(4H, m), 1.59-1.86(3H, m),
2.18-2.26(3H, m), 2.38-2.52(2H, m), 2.70-2.83(2H, m), 3.28-3.55(4H,
m), 5.88(1H, s), 6.15(1H, brs), 7.39(1H, s), 7.47(2H, d, J=8.6 Hz),
7.63(2H, d, J=8.6 Hz), 9.25(1H, brs), 10.35(1H, s), 12.11(1H, brs)
# ES+465.34 ES-463.41 I-32 158 -- 0.72-0.90(4H, m), 1.31-1.54(3H,
m), 2.20-2.35(2H, m), 2.57-2.75(3H, m), 3.12-3.50(2H, m), 5.80(1H,
s), 6.22(1H, brs), 7.38(1H, brs), 7.47(2H, d, J=8.6 Hz), 7.64(2H,
d, J=8.6 Hz), 9.29(1H, s), 10.36(1H, s), 12.08(1H, brs) # ES+437.3
ES-435.37 I-33 159 -- 1.15(3H, t, J=7.5 Hz), 2.19(3H, s),
2.25-2.40(6H, m), 3.30-3.40(4H, m), 5.80(1H, s), 6.25(1H, brs),
7.38(1H, s), 7.48(2H, d, J=8.6 Hz), 7.66(2H, d, J=8.6 Hz), 9.32(1H,
s), 10.06(1H, s), 12.12(1H, brs). ES+439.34 ES-437.39 I-34 160 --
1.10(3H, t, J=7.5 Hz), 2.36(2H, q, J=7.5 Hz), 3.25-3.40(4H, m),
3.55-3.69(4H, m), 5.80(1H, s), 6.21(1H, brs), 7.32(1H, brs),
7.47(2H, d, J=8.6 Hz), 7.65(2H, d, J=8.6 Hz), 9.38(1H, s),
10.04(1H, s), 12.10(1H, brs). ES+426.29 ES-424.38 I-35 161 Shrinks
140.degree. C. Melts 280-282.degree. C. 0.81-0.82(4H, m), 1.81(1H,
m), 2.08(6H, s), 2.33(2H, br s), 3.10-3.12(2H, m), 5.81(1H, s),
6.03(1H, br s), 6.79(1H, br s), 7.38(1H, s), 7.47(2H, d), 7.64(2H,
d), 9.12(1H, br s), 10.34(1H, s), 12.05(1H, br s) # ES+439.40
ES-437.24 I-36 162 mpt Shrinks 130.degree. C. Melts 209-212.degree.
C. 0.81-0.82(4H, m), 1.80(1H, m), 2.24(6H, m), 3.10-3.15(2H, m),
3.51-3.53(4H, m), 5.84(1H, br s), 6.05(1H, br s), 6.87(1H, br s),
7.41(1H, s), 7.48(2H d), 7.66(2H, d), 9.13(1H, br s), 10.35(1H, s),
12.07(1H, br s) # ES+481.34 ES-479.39 I-37 163 131-132
0.80-0.85(4H, m), 1.82(1H, quin), 2.40-2.45(4H, m), 2.58(2H, t),
2.70(2H, t), 3.33-3.38(4H, m), 5.85(1H, s), 6.30(1H, br s),
7.40(1H, s), 7.50(2H, d), 7.70(2H, d), 9.35(1H, s), 10.40(1H, s),
12.10(1H, br s) ES+490.3 I-38 164 Shrinks 220.degree. C. mpt
greater than 340.degree. C. 0.80-0.82(4H, m), 1.63(4H, m), 1.63(1H,
m), 2.33(6H, m), 3.10-3.13(2H, m), 5.82(1H, s), 5.99(1H, br s),
6.87(1H, br s), 7.38(1H, s), 7.46(2H, d), 7.65(2H, d), 9.17(1H, br
s), 10.37(1H, s), 12.07(1H, br s) # ES+465.39 ES-463.31 I-39 165 --
0.80-0.83(4H, m), 1.15-2.02(5H, m), 2.22-2.47(1H, 2m),
2.63-2.79(1H, 2m), 2.91-3.62(6H, m), 4.03-4.53(1H, 2m), 5.80(1H,
s), 6.15 and 6.24(1H, 2br s), 7.35(1H, s), 7.46-4.49(2H, d),
7.65-7.69(2H, 2d), 9.32 and 9.37(1H, 2s), 10.48 and 10.49(1H, 2s),
12.09(1H, br s) # ES+0 477.3 ES-475.4 I-40 166 -- 0.80-0.83(4H, m),
1.15-2.02(5H, m), 2.22-2.47(1H, 2m), 2.63-2.79(1H, 2m),
2.91-3.62(6H, m), 4.03-4.53(1H, 2m), 5.80(1H, s), 6.15 and 6.24(1H,
2br s), 7.35(1H, s), 7.46-4.49(2H, d), 7.65-7.69(2H, 2d), 9.32 and
9.37(1H, 2s), 10.48 and 10.49(1H, 2s), 12.09(1H, br s) # ES+477.3
ES-475.4 I-41 167 -- 0.79-0.89(4H, m), 1.80-1.89(1H, m),
2.65-2.73(1H, m), 2.90-2.99(1H, m), 4.49(2H, s), 5.86(1H, brs),
6.30(1H, brs), 6.95-7.20(4H, m), 7.40(1H, s), 7.50(2H, d, J=8.6
Hz), 7.69(2H, d, J=8.6 Hz), 9.36(1H, s), 10.40(1H, s), 12.15(1H,
brs) # ES+484.36 ES-482.37 I-42 168 168-169 0.80-0.88(4H, m),
1.82(1H, m), 2.88(3H, s), 3.13(4H, br s), 3.48(4H, br s), 5.82(1H,
s), 6.27(1H, br s), 7.40(1H, s), 7.50(2H, d), 7.68(2H, d), 9.41(1H,
s), 10.40(1H, s), 12.15(1H, br s) ES+515.3 I-43 169 -- 0.86(7H, m),
1.43(2H, m), 1.80(1H, m), 2.23(2H, t), 2.33(4H, m), 3.31(4H, m),
(masked), 5.81(1H, s), 6.23(1H, br s), 7.36(1H, s), 7.48(2H, d),
7.65(2H, d), 9.31(1H, s), 10.35(1H, s), 12.15(1H, br s) ES-463.4
ES+465.3 I-44 170 -- 0.83(4H, m), 1.82(1H, m), 2.22(3H, s),
2.89(4H, m), 3.33(4H, m)(masked), 5.81(1H, s), 6.24(1H, br s),
7.36(1H, s), 7.48(2H, d), 7.65(2H, d), 9.32(1H, br s), 10.35(1H,
s), 12.10(1H, br s) ES-477.5 ES+479.4 I-45 171 154-155
0.80-0.84(4H, m), 1.80(1H, quin), 2.40-2.43(4H, m), 2.72(2H, t),
3.03(3H, s), 3.28-3.35(6H, m), 5.80(1H, s), 6.25(1H, br s),
7.40(1H, s), 7.50(2H, d), 7.65(2H, d), 9.35(1H, s), 10.40(1H, s),
12.10(1H, br s) ES+543.3 I-46 172 160-161 0.80-0.85(4H, m),
1.06(3H, d), 1.80(1H, quin), 2.67(1H, br s), 3.65(1H, m), 4.05(1H,
br s), 5.85(1H, s), 6.25(1H, br s), 7.40(1H, s), 7.50(2H, d),
7.65(2H, d), 9.30(1H, br s), 10.35(1H, s), 12.10(1H, br s) ES+451.3
I-47 173 158-159 0.80-0.85(4H, m), 1.82(1H, quin), 2.35-2.45(4H,
m), 3.17(2H, br s), 3.22-3.26(2H, m), 3.42-3.45(2H, m),
3.50-3.58(6H, m), 5.85(1H, s), 6.25(1H, br s), 7.40(1H, s),
7.50(2H, d), 7.70(2H, d), 9.35(1H, br s), 10.40(1H, s), 12.10(1H,
br s) ES+564.3 I-48 174 -- 0.78-0.82(4H, m), 1.79(1H, m), 2.36(4H,
m), 3.3(masked signal), 3.48(2H, s), 5.81(1H, s), 6.19(1H, br s),
7.24-7.35(6H, m), 7.47(2H, d), 7.63(2H, d), 9.33(1H, s), 10.34(1H,
s), 12.09(1H, s) ES+527.4 ES-525.4 I-49 175 -- 0.80-0.81(4H, m),
1.80(1H, m), 2.00(3H, s), 2.36-2.38(4H, m), 3.3(masked signal),
3.49(2H, s), 5.42(1H, s), 5.99(1H, br s), 7.25-7.35(5H, m),
7.47(2H, d), 7.69(2H, d), 9.23(1H, s), 10.39(1H, s), 11.69(1H, s)
ES+541.4 ES-539.4 I-50 176 -- 0.80(4H, m), 0.93(6H, d), 1.82(1H,
m), 2.20(2H, t), 2.58(2H, m), 3.79(2H, m), 5.87(1H, s), 6.23(1H, br
s), 7.40(1H, s), 7.47(2H, d), 7.74(2H, d), 9.27(1H, br s),
10.35(1H, s), 12.11(1H, brs) ES-463.5 ES+465.4 I-51 177 --
1.10(.English Pound.h, t, J=7.5 Hz), 2.36(2H, q, J=7.5 Hz),
3.32(3H, s), 5.80(1H, brs), 6.05(1H, brs), 7.12-7.45(6H, m),
7.49(2H, d, J=8.6 Hz), 7.71(2H, d, J=8.6 Hz), 9.48(1H, brs),
10.11(1H, s), 12.05(1H, brs). ES+446.31 ES-444.34 I-52 178 --
0.75-0.89(4H, m), 0.89-1.03(6H, m), 1.74-1.88(1H, m), 3.15 3.29(4H,
m), 5.89(1H, brs), 6.18(1H, brs), 7.42(1H, brs), 7.47(2H, d, J=8.6
Hz), 7.63(2H, d, J=8.6 Hz), 9.19(1H, brs), 10.34(1H, s), 12.10(1H,
brs). ES+424.34 ES-422.35 I-53 179 167-169 0.81-0.83(4H, m),
1.00(9H, s), 1.81(1H, m), 2.47(4H, m), 3.14(4H, m), 5.82(1H, s),
6.20(1H, br s), 7.36(1H, s), 7.47(2H, d), 7.65(2H, d), 9.32(1H, s),
10.37(1H, s), 12.09(1H, s) ES+493.4 ES-491.4 I-54 180 -- 2.21(3H,
s), 2.27-2.40(4H, m), 3.31-3.50(4H, m), 5.90(1H, s), 6.31(1H, brs),
7.10-7.25(3H, m), 7.35-7.50(3H, m), 9.38(1H, s), 12.14(1H,
brs). ES+352.28 ES-350.32 I-55 181 -- 2.05(3H, s), 2.19(3H, s),
2.26-2.39(4H, m), 3.36-3.46(4H, m), 5.95(1H, brs), 6.37(1H, brs),
7.06(2H, d, J=8.9 Hz), 7.45(1H, brs), 7.56(2H, d, J=8.9 Hz),
9.30(1H, brs), 9.95(1H, s), 12.12(1H, s). ES+409.31 ES-407.37 I-56
182 drkns 250 277-9 ES+485.3 I-57 183 -- 1.34(3H, t, J=7.1 Hz),
3.33-3.42(4H, m), 3.59-3.68(4H, m), 4.32(2H, q, J=7.1 Hz), 5.94(1H,
s), 6.40(1H, brs), 7.29(2H, d, J=8.7 Hz), 7.49(1H, brs), 7.99(2H,
d, J=8.7 Hz), 9.50(1H, s), 12.20(1H, brs). ES+411.30 ES-409.37 I-58
184 -- 3.30-3.39(4H, m), 3.60-3.65(4H, m), 5.89(1H, s), 6.25(1H,
brs), 7.15-7.50(9H, m), 9.40(1H, brs), 12.12(1H, s). ES+415.32
ES-413.37 I-59 185 -- 2.05(3H, s), 3.25-3.45(4H, m), 3.59-3.70(4H,
m), 5.94(1H, s), 6.35(1H, brs), 7.07(2H, d, J=8.9 Hz), 7.46(1H,
brs), 7.58(2H, d, J=8.9 Hz), 9.40(1H, s), 9.98(1H, s), 12.13(1H,
brs). ES+396.32 ES-394.38 I-60 186 -- 0.38-0.48(2H, m),
0.79-0.89(2H, m), 1.64-1.73(1H, m), 2.04(3H, s), 3.34-3.40(4H, m),
3.61-3.69(4H, m), 5.46(1H, s), 6.10(1H, brs), 7.05(2H, d, J=8.9
Hz), 7.61(2H, d, J=8.9 Hz), 9.34(1H, s), 9.99(1H, s), 11.85(1H, br
s). ES+436.36 ES-434.41 I-61 187 238-239 0.85(4H, s), 1.80(1H, m),
2.00(3H, s), 3.35(4H, s), 3.60(4H, s), 5.43(1H, s), 6.00(1H, br s),
7.50(2H, d), 7.70(2H, d), 9.30(1H, s), 10.40(1H, s), 11.70(1H, br
s) ES+452.2 V-1i 188 -- 0.81(4H, d), 1.83(1H, m), 2.02(3H, s),
2.77(3H, s), 2.90-3.17(4H, m), 4.09-4.33(4H, m), 5.46(1H, s),
6.06(1H, s), 7.47(2H, d), 7.72(2H, d), 9.35(1H, s), 1.45(1H, s),
10.62(1H, s), 11.72(1H, s) -- V-1iii 189 -- 0.81-0.83(4H, d),
1.81(1H, m), 2.04(3H, s), 2.82-2.83(3H, m), 3.08-3.11(4H, m),
3.42-3.47(4H, m), 4.14-4.17(br m, OH), 5.49(1H, s), 6.04(1H, s),
7.48(2H, d), 7.71(2H, d), 9.53(1H, s), 9.64(1H, s), 10.39(1H, s) --
V-1iv 190 -- 0.82(4H, d), 1.80(1H, m), 2.02(3H, s), 2.45(3H, s),
2.69(br s, OH), 3.01(2H, s), 3.38-3.47(8H, m), 5.45(1H, s),
6.05(1H, s), 7.47(2H, d), 7.70(2H, d), 9.25(1H, s), 10.36(1H, s) --
V-1v 191 -- 0.80-0.82(4H, m), 1.81(1H, m), 2.02(3H, s), 2.21(3H,
s), 2.34-2.36(4H, m), 3.36-3.38(masked signal for 4H + OH),
5.45(1H, s), 6.04(1H, s), 6.61(1H, s), 7.47(2H, d), 7.69(2H, d),
9.18(1H, s), 10.36(1H, s) -- V-1vi 192 -- 0.80-0.82(4H, d),
1.81(1H, m), 2.02(3H, s), 2.21(3H, s), 2.33-2.36(4H, m), 2.41(4H,
s), 3.30-3.45(masked signal, 4H, m), 4.19(1H, br s), 5.45(1H, s),
6.03(1H, br s), 7.47(2H, d), 7.69(2H, d), 9.18(1H, s), 10.35(1H,
s), 11.70(1H, br s) -- V-1vii 193 -- 0.81-0.83(4H, d), 1.81(1H, m),
2.02(3H, s), 2.80(3H, s), 3.11-3.45(masked signal, 8H, m), 5.45(1H,
s), 6.07(3H, s), 7.48(2H, d), 7.71(2H, d), 9.36(1H, s), 10.38(1H,
s), 11.75(1H, br s) -- V-1viii 194 -- 0.81-0.82(4H, d), 1.81(1H,
m), 2.02(3H, s), 2.27(3H, s), 2.43(4H, m), 3.38-3.47(masked signal,
4H, m), 4.20(2H, s), 5.45(1H, s), 6.04(1H, br s), 7.47(2H, d),
7.69(2H, d), 9.20(1H, s), 10.36(1H, s) -- V-1ix 195 --
0.81-0.83(4H, d), 1.81(1H, m), 2.02(3H, s), 2.82(3H, s),
3.03-3.13(4H, m), 3.36-3.75(masked signal, 6H, m), 4.12-4.15(2H,
m), 5.45(1H, s), 6.05(1H, s), 7.48(2H, d), 7.71(2H, d), 9.37(1H,
s), 9.61(1H, brs), 10.38(1H, s) -- V-1x 196 -- 0.81-0.82(4H, d),
1.81(1H, m), 2.02(3H, s), 2.40(3H, s), 2.54-2.68(8H, m),
3.40-3.45(masked signal, 4H, m), 4.32(1H, br s), 5.45(1H, s),
6.05(1H, br s), 7.47(2H, d), 7.69(2H, d), 9.24(1H, s), 10.36(1H, s)
-- V-1xi 197 -- 0.80-0.82(4H, d), 1.80(1H, m), 2.02(3H, s),
2.31(3H, s), 2.50(masked signal, 4H), 3.36-3.47(4H, m), 4.88(br m,
OH), 5.45(1H, s), 6.04(1H, s), 7.47(2H, d), 7.69(2H, d), 9.22(1H,
s), 10.36(1H, s) -- V-20 i 198 1.09(3H, t), 2.00(3H, s), 2.38(2H,
q), 2.77(3H, s), 3.00(2H, m), 3.18(2H, m), 3.40(2H, d), 4.10(2H,
d), 5.41(1H, s), 6.06(1H, br s), 7.48(2H, d), 7.73(2H, d), 9.42(1H,
s), 10.15(1H, s), 10.64(1H, br s), 11.77(1H, br s) ES-451.4
ES+453.4 (M +H)+ V-20 ii 199 1.09(3H, t), 2.0(3H, s), 2.35(5H, m),
2.81(3H, s), 3.09(4H, m), 3.44(2H, d), 4.12(2H, d), 5.41(1H, s),
6.02(1H, br s), 7.48(2H, d), 7.73(2H, d), 9.44(1H, s), 9.70(1H, br
s), 10.10(1H, s), 11.80(1H, br s) ES-451.4 ES+453.4 (M +H)+
Biological Assays
[0234] The activity of the compounds of this invention as kinase
inhibitors may be assayed in vitro, in vivo or in a cell line. In
vitro assays include assays that determine inhibition of either the
kinase activity or ATPase activity of activated Aurora and/or FLT-3
enzyme. Alternate in vitro assays quantitate the ability of the
inhibitor to bind to Aurora and/or FLT-3 and may be measured either
by radiolabelling the inhibitor prior to binding, isolating the
inhibitor/Aurora and/or inhibitor/FLT-3 complex and determining the
amount of radiolabel bound, or by running a competition experiment
where new compounds are incubated with Aurora and/or FLT-3 bound to
known radioligands. One may use any type or isoform of Aurora,
depending upon which Aurora type or isoform is to be inhibited. The
details of the conditions used for the enzymatic assays are set
forth in the Examples hereinbelow.
Example 10
K.sub.i Determination for the Inhibition of Aurora
[0235] Compounds were screened in the following manner for their
ability to inhibit Aurora using a standard coupled enzyme assay
(Fox et al (1998) Protein Sci 7, 2249). To an assay stock buffer
solution containing 0.1M HEPES 7.5, 10 mM MgCl.sub.2, 1 mM DTT, 25
mM NaCl, 2.5 mM phosphoenolpyruvate, 300 mM NADH, 30 mg/ml pyruvate
kinase, 10 mg/ml lactate dehydrogenase, 40 mM ATP, and 800 .mu.M
peptide (LRRASLG, American Peptide, Sunnyvale, Calif.) was added a
DMSO solution of a compound of the present invention to a final
concentration of 30 .mu.M. The resulting mixture was incubated at
30.degree. C. for 10 minutes. The reaction was initiated by the
addition of 10 .mu.L of Aurora stock solution to give a final
concentration of 70 nM in the assay. The rates of reaction were
obtained by monitoring absorbance at 340 nm over a 5 minute read
time at 30.degree. C. using a BioRad Ultramark plate reader
(Hercules, Calif.). The K.sub.i values were determined from the
rate data as a function of inhibitor concentration.
[0236] Compounds of formula V of the present invention were found
to be inhibitors of Aurora-1, Aurora-2, and Aurora-3.
Example 11
K.sub.i Determination for the Inhibition of FLT-3
[0237] Compounds were screened for their ability to inhibit FLT-3
activity using a radiometric filter-binding assay. This assay
monitors the .sup.33P incorporation into a substrate poly(Glu, Tyr)
4:1 (pE4Y). Reactions were carried out in a solution containing 100
mM HEPES (pH 7.5), 10 mM MgCl.sub.2, 25 mM NaCl, 1 mM DTT, 0.01%
BSA and 2.5% DMSO. Final substrate concentrations in the assay were
90 .mu.M ATP and 0.5 mg/ml pE4Y (both from Sigma Chemicals, St
Louis, Mo.). The final concentration of a compound of the present
invention is generally between 0.01 and 5 .mu.M. Typically, a
12-point titration was conducted by preparing serial dilutions from
10 mM DMSO stock of test compound. Reactions were carried out at
room temperature.
[0238] Two assay solutions were prepared. Solution 1 contains 100
mM HEPES (pH 7.5), 10 mM MgCl.sub.2, 25 mM NaCl, 1 mg/ml pE4Y and
180 .mu.M ATP(containing 0.3 .mu.Ci of [.gamma.-.sup.33P]ATP for
each reaction). Solution 2 contains 100 mM HEPES (pH 7.5), 10 mM
MgCl.sub.2, 25 mM NaCl, 2 mM DTT, 0.02% BSA and 3 nM FLT-3. The
assay was run on a 96 well plate by mixing 50 .mu.l each of
Solution 1 and 2.5 ml of the compounds of the present invention.
The reaction was initiated with Solution 2. After incubation for 20
minutes at room temperature, the reaction was stopped with 50 .mu.l
of 20% TCA containing 0.4 mM of ATP. All of the reaction volume was
then transferred to a filter plate and washed with 5% TCA by a
Harvester 9600 from TOMTEC (Hamden, Conn.). The amount of .sup.33P
incorporation into pE4y was analyzed by a Packard Top Count
Microplate Scintillation Counter (Meriden, Conn.). The data was
fitted using Prism software to get an IC.sub.50 or K.sub.i.
[0239] Compounds of formula V of the present invention were found
to be inhibitors of FLT-3.
Example 12
IC.sub.50 Determination for the Inhibition of Aurora in a Colo205
Cellular Assay
[0240] Compounds were also assayed for the inhibition of cell
proliferation. In this assay, a complete media was prepared by
adding 10% fetal bovine serum, L-glutamine and
penicillin/streptomycin solution to RPMI 1640 medium (Sigma). Colon
cancer cells (COLO-205 cell line) were added to a 96 well plate at
a seeding density of 1.25.times.104 cells/well/150 .mu.L. A
solution of test compound was prepared in complete media by serial
dilution, the test compound solution (50 .mu.L) was added to each
per well.
[0241] Each plate contained a series of wells in which only
complete media (200 .mu.L) was added to form a control group in
order to measure maximal proliferation. A vehicle control group was
also added to each plate. The plates were incubated at 37.degree.
C. for 2 days. A stock solution of .sup.3H-thymidine (1 mCi/mL,
Amersham Phamacia UK) was diluted to 20 .mu.Ci/mL in RPMI medium
then 25 .mu.L of this solution was added to each well. The plates
were further incubated at 37.degree. C. for 3 hours then harvested
and analyzed for .sup.3H-thymidine uptake using a liquid
scintillation counter.
[0242] Compounds of formula V of the present invention were found
to be inhibitors of proliferation of Colo205 cancer cells.
Example 13
Measurement of Cell Proliferation in a Panel of Tumour and Normal
Cell Types: .sup.3H thymidine Incorporation Assay
[0243] The .sup.3H thymidine incorporation assay was chosen as a
well characterized method of determining cell proliferation. Cells
from normal tissues and a wide variety of different tumour types
were chosen for analysis. Many of the tumour cells were selected
because they express high levels of Aurora proteins (e.g. MCF-7,
PC3, A375, A549) (See section 5.3.5 and Bischoff et al EMBO J. 1998
17, 3052-3065) and/or are able to form tumours in nude mice or rats
(e.g. HCT116, MCF-7 and MDA-MB-231).
[0244] Logarithmically growing cells were incubated with compound
for 96 hours. To measure cell proliferation, 3 hours prior to the
end of the experiment 0.5 .mu.Ci of .sup.3H thymidine was added to
each well. Cells were then harvested, washed and the incorporated
radioactivity counted on a Wallac microplate beta-counter. To
determine the inhibition of proliferation, cpm were plotted versus
compound concentration, and the IC.sub.50 graphically
determined.
[0245] Table 5 below sets forth the cell lines utilized in the
above described cell proliferation assay. For each cell line, the
inhibition of cell proliferation and .sup.3H thymidine
incorporation (96 hour time-point) was determined.
5TABLE 5 Cell Lines Origin Cell line Colorectal adenocarcinoma
HCT-116 Colorectal adenocarcinoma LS174T Leukemia HL60 Mammary
gland adenocarcinoma MDA-MB-231 Mammary gland adenocarcinoma
ZR-75-1 Mammary gland adenocarcinoma MCF-7 Prostate adenocarcinoma
PC3 Pancreatic MIA-Pa-Ca-2 Melanoma A375 Primary PHA-stimulated
human lymphocytes PHA blasts
[0246] While a number of embodiments of this invention have been
described, it is apparent that the basic examples may be altered to
provide other embodiments which utilize the compounds and methods
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 which have been represented by way
of example.
* * * * *
References