AU2004200192A1 - Purine inhibitors of cyclin dependent kinase 2 and 1k-Aa - Google Patents

Description

P/00/01Il Regulation 3.2 AuSTRALIA Patents Act 1990 COMPLETE SPECIFICATION STANDARD PATENT Invention Title: Purine inhibitors of cyclin dependent kinase 2 and lk-Aa The following statement is a full description of this invention, including the best method of performing it known to us: Ireenills carter ~mitn I~eadIe Melbourne\004421 518 Printed 16 January 2004 (13:47) page 2 Freehills Carter Smith Beadle Melbourne\004421818 Printed 16 January 2004 (13:47) page 2 Title: PURINE INHIBITORS OF CYCLIN DEPENDENT KINASE 2 and IrB-a BACKGROUND OF THE INVENTION Field of the Invention This invention concerns 2,6,9-trisubstituted purines that have been discovered to be selective inhibitors of cell cycle kinases and, as such, the compounds are inhibitors of cell proliferation. The 2,6,9-trisubstituted purines are useful in for example in treating autoimmune diseases, e.g. rheumatoid arthritis, lupus, type I diabetes, multiple sclerosis, etc., in treating cancer, cardiovascular disease, such as restenosis, host vs graft disease, gout, polycystic kidney disease and other proliferative diseases whose pathogenesis involves abnormal cell proliferation.

This invention also concerns 2,6,9-trisubstituted purines that have been discovered to be potent and specific inhibitors of IKB-at kinase which prevents signal induced NF-KB activation and cytokine synthesis in vitro and in vivo. Such inhibitors are expected to inhibit the synthesis of cytokines and adhesion proteins whose synthesis is transcriptionally regulated by NF-KB. Proinflammatory cytokines such as IL-1, 1L-6, TNF and adhesion proteins (e.g.

ICAM, VCAM and selections) belong to this class of molecules and have been implicated in the pathogenesis of inflammatory diseases. Thus a potent inhibitor of IicB-a kinase is useful in the clinical management of diseases where NF-KB activation is required for disease induction.

Description of the Art In the past few years, advances in molecular and cellular biology have contributed to our understanding of the mechanisms of cell proliferation and of specific events that occur during progression of cells through mitosis. "Progress in Cell Cycle Research" Vol 1, Eds. L.

Meijer, S. Guidet and H.Y.L. Tung; Plenum Press, New York, 1995. These studies have shown that progression through the cell cycle is controlled by a family of serine/threonine kinases called cyclin dependent kinases. These enzymes contain a catalytic protein called cyclin dependent kinase (CDK) that uses ATP as a substrate and a regulatory protein called cyclin. Different cyclin-CDK combinations control events such as growth, DNA replication and cell division. One key member of the CDK family of enzymes is CDK2.

CDK2 activity has been shown to be essential for mammalian cell cycle progression at the G1/S boundary. Microinjection of antibodies directed against CDK2 blocks the progression of human diploid fibroblasts into the S phase of the cell cycle. Expression of a CDK2 dominant negative mutant in human osteosarcoma cells has a similar effect. Together, these studies indicate that inhibition of cellular CDK2 activity will prevent progression of cells through the mitotic cycle and induce growth arrest prior to the S phase. Consistent with this view, in vitro studies with olomoucine (2-(hydroxye l hylamino)-6benzylamino-9-methylpurine), have shown that it is a specific inhibitor of CDK2 with an IC 50 of approximately 2.1 gg/ml J. Vesely, et al.; Eur. J.Biochem 224, 771-786 (1994), L. Meijer "Chemical Inhibitors of Cyclin-Dependent Kinases" pp 351-356 in "Progress in Cell Cycle Research Vol 1, Eds. L. Meijer, S. Guidet and H.Y.L. Tung; Plenum Press, New York, 1995.

In vivo studies using mammalian cells in culture have shown that olomoucine inhibits cell proliferation at an approximate concentration of 50 gg/ml.

In this invention, we have developed several compounds whose biological activity is considerably more potent than olomoucine. In vivo studies using mammalian cells indicate that some of the disclosed compounds inhibit cell proliferation at concentrations that are significantly lower than olomoucine.

Recently an IKB-a kinase activity has been described in the cytoplasm of stimulated human umbilical vein endothelial cells (Bennett et al (1996) J. Biol.Chem 271, 19680- 19688). Some of the compounds of this invention have been identified as potent and specific inhibitors of IKB-Ca kinase which prevents signal induced NF-KB activation and cytokine synthesis in vitro and in vivo. The activation of the heterodimeric transcription factor NF-rB is a complex process. In unstimulated cells, the NF-KB (p50/p65) heterodimer is located in the cytosol where it is complexed with an inhibitory subunit IKBa, IKB-a, binds to NF-rB thus masking its nuclear localization signal and preventing translocation to the nucleus. Upon stimulation of cells with a variety of signals (e.g.

lipopolysaccharide) IKB-a is rapidly phosphorylated, uniquitinated and degraded by the proteasome. Degradation of IKB-a., allows the translocation of NF-KB to the nucleus where it activates transcription of a number of inflammatory response genes.

These observations suggest that IKB-c kinase is an attractive target for the identification of inhibitors that may be useful in the treatment of inflammatory diseases where NF-KB activation is required for disease induction.

Reference to any prior art in the specification is not, and should not be taken as, an acknowledgment or any form of suggestion that this prior art forms part of the common general knowledge in Australia or any other jurisdiction.

4 SUMMARY OF THE INVENTION It is an object of this invention to provide 2,6,9-trisubstituted purine compounds, which inhibit the cyclin dependent kinase 2.

It is another object of this invention to provide 2,6,9-trisubstituted purine compounds which are useful for inhibiting cell proliferation.

This invention also constitutes a pharmaceutical composition, which comprises a 2,6,9-trisubstituted purine compound and a pharmaceutically acceptable carrier.

This invention further constitutes a method for inhibiting cell proliferation, which comprises administering to a mammal in need thereof an effective amount of a 2,6,9trisubstituted purine compound.

In one embodiment, this invention is A 2,6,9-trisubstituted purine composition of matter having the following formula: x

R

(I)

wherein R, is halogen or wherein X NH, 0, S, S(0 2 In the composition, is alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl, and alkynyl, each having one to 20 carbon 'atoms, which alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl, and alkynyl, are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, CF 3 heteroaryl, heterocyclyl, R, SR 20

S(O)R

1 S0 2

R

21

SO

2

NR

2

R

23 S0 2

NR

20 00R 1

SO.,NR'

0 00NR 20 RY3,

SONR

20 00 2

R

2 1, NR 20 R23, NR 20 C0R 2

NR

20 C0 2

R

1

NR

20

CONR

2 3, N(R' 0

)C(NR

2

)NHIRY,

NR

20 SO.R", OR 20

OCONR

2 0

R

23 000NR 20 S0 2

R

1 000NR 0

R

23 ON, C0 2

R

2 0 CONR 2 3, C0NR 2 1S0 2 ,R and C0R 2 1. Also in the composition, R 2 is a hydrogen or hydrocarbon selected from the group alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkvnyl,are optionally substituted with from 1 to 3 sub stituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R 22

S(O)R

1 S0 2

R

2 1 S0 2

NR

2 0R 2 1, S0 2

NR

20 C0R 2 1 SO2NR 20

CONR

20 R.2, SO2NR 20

CO

2 R 2 1

NR

2 0RnI, NR 20 00R 21

NR

2 'C0 2

R

2

WROCONR&R

2

N(R

20 )C(NR 20

)NHR

3

NR

20 S0 2

R

21 OR 20

OGONR

20 R21, 000NR 20 S0 2

R

2 1

OCONR

20 R23, ON, CO.R 20

CONR

20 R21, CONR 20 SO,R 2 1 and C0R 2 Further, in the compositions, R, is a halogen, hydroxyl, thio, ailkoxy, alkylthio, alkyl, -NRR, or a component having the formula: 6 where m=1-3, o=1,3, y=carbonyl, -NRR,, hydroxyl, thiol, alkoxy, alcyithiol; R, and R, are each independently hydrogen, OR 2 0

NR

20

R

23 or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which ailkyl, acyl, heterocyclyl, aryl, heteroaryi, aralkyl, heteroarylalcyl, alkenyl,and allcynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R 1 2

SR

20

S(O)R

21 SO2R 21 S02NRWORD, So, NR 20

COR

2 1 SON 0

CONROR

2 3

SON

2 0R

N

R20 RR, NR 0 0 21

NR

2 0 C 21

NR

20 C0NR 2

R

23

N(R

2 0 )C(NjR 20 )NI-1R 23

R

0 0R' OR 20 0C0NR 20

R

23

OCONR

20 SOR 1, OCONR 2 0pR 23

,CN

C0 2

R

20 C0NR 2

R

23

CON

20 SO,R 2 1 and C0R 20 R 20 is a member selected from the group consisting of H, ailkyl, C 2 4 5 alkenyl, C, alkynyl, heterocyclyl, aryl, and heteroaryl, which ailkyl, alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl or heteroaryl amiide, CN, 0-C.

6 ailkyl, CF 3 aryl, and heteroaryl; R 2 1 is a member selected from the group consisting of C,4, 5 alkyl, 5 alkenyl, C., alcynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkynyl, aryl, heterocyclyl, and heteroaryl are optionally substituted with 1 to 3 substi tuents independently selected from the group of halo, heterocyclyl, aryl, heteroaryl, CF 3 CN, OR 20 SR 2 1, N(R 0 2

S(O)R

22

SO,R

22

SO

2

N(R

0

SO,NR

20 C0R 2 S0 2

NV

20 C0 2 R1 2

SO

2

NR

2

CONR

2 2

N(R

20

NR

20 C0R 22

NR?

0 C0 2

R

22

NR

2

CON(R

20 2

NR

2 0

C(NR

20

)NHR

3 C0R 20 C0 2

R

20

CON(R

20 2

CONR

20 SOR 2 NR 2 0

SOR

22 S0 2

NR

2 0C0 2 R1 2

OR

20 0C0NR 0 S0 2

R?

2

OC(O)R

0

C(O)OCH.,OC(O)R

20 and OCON(R 20 2 and each optional heteroaryl, aryl, and heterocyclyl 7 substituent is optionally substituted with halo, alkyl, CF, amino, mono- or di- alkylamino, alkyl or aryl or heteroaryl amide, NCOR 2 2

NR

2 OSOR", COR 2 0

COR

20

CON(R

2 0 2

NR

2 0

CON(R

20 2

OC(O)R

2 0 OC(O)N(R2 0 2

SR

2 0 S(0)R 22

SO

2

R

2 2

SON(R

2 0 2 CN, or OR 2 0

R

2 2 is a member selected from the group consisting of C.i,5 alkyl, alkenyl, C2.i, alkynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl or heteroaryi amide, CN, alkyl, CF 3 aryl, and heteroaryl; and R" is R 1 or H.

There are some limitations to the scope of the compositions of this invention. When Y is carbonyl, Y and R' 4 together may be a single oxygen atom, R 4 and together may be a single oxygen atom, R 4 and R 5 may together be a single oxygen atom. When R 3 is 2hydroxyethylamino and R 2 is methyl, is not amino, 3-methyl-2-butenylamino, benzylamino, or m-hydroxybenzylamino when R 3 is not 2-hydroxyethylamino. When R, is isopropyl, is not benzylamino, m-hydroxybenzylamino, or 3-methylbutylamino. When

R

3 is 2-hydroxyethylamino and R, is 2-hydroxyethyl, is not benzylamino and when R 3 is selected from the group consisting of 2-methyl-2-hydroxypropylamino, and 2dimethylaminoethylamino, and when R, is methyl, then is not benzylamino In another embodiment, this invention is a method for inhibiting cell proliferation in mammals comprising administering a therapeutically effective amount of the composition of claim 1 to the mammal. The method is useful for treating cell proliferation disorders such as rheumatoid arthritis, lupus, type I diabetes, multiple sclerosis, cancer, restenosis, host graft disease, and gout.

8 In yet another embodiment, this invention is a pharmaceutical composition of matter comprising the composition above in an admixture with one or more pharmaceutical excipients.

In still another embodiment, this invention is a composition useful for treating fungal infections (fungi) in humans, animals, and in plants.

DESCRIPTION OF THE FIGURE Figure 1 is a plot of the mean neointimal area of a rat carotid artery treated with a saline vehicle and treated with compound 3 prepared according to Example 2 wherein the unshaded bar represents the untreated section of the carotid artery and the shaded bar represents the treated section of the carotid artery.

It will be understood that the term "comprises" or its grammatical variants as used herein is equivalent to the term "includes" and is not to be taken as excluding the presence of other elements or features.

DESCRIPTON OF THE CURRENT EMBODIMN The present invention relates to a 2,6,9-trisubstituted purine compound having the following formulas: Rl-.% where: R, is halogen or wherein X NH, 0, S, S(0 2

R'

1 is ailkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl, and alkynyl, each having one to 20 carbon atoms, which ailkyl, heterocyclyl, aryl, heteroaryl, arailcyl, heteroarylalkyl, alkenyl, and alkynyl, are optionally substituted with from I to 3 substituents independently selected from the group consisting of halo, ar-yl, CF 3 heteroaryl, heterocyclyl, R 22

SR

20

S(O)R

21 S02R 21

SONR

0

R

3 ,I So NR 20 C0R 2 1

SONR

20 C0NR 2

OR

2 1,

SO,NR

2 0

,R

2

NR

20

R

23

NR

20 00R 21

NR

20 C0 2

R

2

NR

20

CONR

2

"R

23

N(R

2 0 )c(NR 20

)NHR

2

NR

20 S0 2

R

21 OR 2 0

OCONR

20 R23, 0C0NR 20 S0 2

R

1 0C0N 20

R

2 3 CN, C0 2 R 2 1, C0NR 2

R

23

CONR

2 0 1SOR 2 1 and C0R 0 R, is a hydrogen or hydrocarbon selected from the group alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalcyl, alkenyl,and alkynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl., SR 20 S(O)R 2 S0 2

R

2 1

SO

2

NR

2 0

R

2 3, S &,NR 2 0 C0R 1

SO

2

NR

2

OCONR

2

&R

23

SON

2

C

2 2

,NR

20 R21, NR 20

CO

21

NR

2

C

2 2 1

NR

20 c0NR 20 R 23

N(R

20

)C(NR

20

)NIR

2 3

NR

20 SOR 21 OR 20 0C0NR 0

R

23 0C0NR 2 1S0 2 R 2 1 0C0NR 2

R

2 3 CN, C0 2 R 20 C0NR 20

R

23

CONR

20

SO'R

2 1 and C0R 2 1

R

3 is a halogen, hydroxyl, thio, alcoxy, alkylthio, alkyl, -NRR., or a component having the formula: Yj n where m=1-3, n=1-3, o=1,3, y=carbonyl, -NR 4

R.

5 hydroxyl, thiol, alkoxy, alkyithiol; R, and R, are each independently hydrogen, OR 2 0

NR

2 ,R23, or a hydrocarbon selected fromn the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkcynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and allcynyl,are optionally substituted with from I to 3 substituents independently selected from the group consisting of halo,. aryl, 12 heteroaryl, heterocyclyl, R" 2

SR

20

SOW)R

2 S0 2

R

21 S0NlR S0N 2 0R S0 2

NR'

0 C0NR 20

R

3 S0 2

NR

20 C0 2

R

21

NR

2

&R

23

NR

20 COR 2 1

NR

20 C0 2

R

2 1

NR

20 CoiR2OR 23

N(R

20

)C(NR

20 )NH;lR 23

NR

20 S0 2

R

2 1 OR 20

OCONR

20 R OOONR 2 0 SO2R 21 OCONR 20 'R 2

CN,

C0 2

R

20

CONR

20 R13, C0NR 20

SO

2 R 2 and C0R 2 0 with the proviso that when Y is carbonyl, Y and together may be a single oxygen atom, R 4 and together may be a single oxygen atom, and may together be a single oxygen atom, and wherein when R 3 is 2hydxoxyethyl amino and R, is methyl, R 1 is not amino, 3-methyl-2-butenylamino, benzylamnino, or m-hydroxybenzylamino, when R 3 is not 2-hydroxyethylamino, when R, is isopropyl, is not benzylamino, m-hydroxybenzylamino, or 3-methylbutylamino, when

R

3 is 2-hydroxyethylamino and is 2-hydroxyethyl, R, is not benzylamino and when R 3 is selected from the group consisting of 2-methyl-2-hydroxypropylamino, and 2dimethylaminoethylamino, and when R 2 is methyl, then R 1 is not benzylamino.

In the compositions, R 2 1 is a member selected from the group consisting of H, alkyl, alkenyl, alkcynyl, heterocyclyl, aryl, and heteroaryl, which ailcyl, alkenyl,, alkynyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylanino, alkyl or aryl or heteroaryl amide, ON, 0-01. alkyl, OF 3 aryl, and heteroaryl.

Also in the compositions, R 1 is a member selected from the group consisting of alkyl, 02.,5 alkenyl, C2.1, alkcynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkcynyl, aryl, heterocyclyl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from the group of halo, heterocyclyl, aryl, heteroaryl, OF 3 CN, OR" 0

N(R

2 0 2

S(O)R

22 SOOR22, SOXNR 2 S0 2

NR

20 00R 2 2 S0 2

NR

20 C0 2 R 1 2

SO

2

NR

20 C0N(R 2 0 2

N(R

20 2

NR

20 00R 2

NR

2 0

O

2 R22, NR? 0 00N(R 20 2

NR

20

C(NR

2 O)NKM3, 13 CopR 2 1, C0R 0

O(R

2 0

CONTR

2 0 1SO,R 22

NR

20

SO

3

R

22 SON 2

OR

22

OR

20

OCONR

2 0

SO,R

22 OC(O)R 20

C(O)OCHOO(O)R

0 and OCON(R 2 and each optional heteroaryl, aryl, and heterocyclyl substituent is optionally substituted with halo, alkyl, OF 3 amino, mono- or di- alkylamino, alkyl or aryl or heteroaryl amide, NCOR 22

NR

20 SO,R 22 C0R 0

COR

20

CON(R

0 2

NR

20 C0NR 0 2

OC(O)R

0

OC(O)N(R

20 2

SR

20

S(O)R

2 S0 2 R 22

SO

2 N(R 2 0 2 ON, or OR 20 In the compositions, R 2 2 is a member selected from the group consisting of C, alkyl, 02.15 alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl or heteroaryl amide, ON, alkyl, OF3, aryl, and heteroaryl, and is R or H.

is preferably an aryl, substituted aryl, each having 6 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, OF3, aryl, R22, NR 20 R?3' NR 20 C0R 21 OR 20 ON; an aralkyl, substituted arailcyl, each having 6-8 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, OF3, aryl, R 22

NR?

0

R

2 3

NR

20 C0R 21

OR

20 ON; -CH,-phenyl wherein the phenyl ring is optionally substituted with from I to 2 substituents independently selected from the group consisting of halo, OF3, R 22

OR"

0 ON; substituted pyridylalkyl; unsubstituted pyridylalkyl; pyraidyl; substituted pyridyl; and benzyl substituted with a halogen, alkoxy, phenyl, pyridyl or nitro group. In some compositions, is preferably OH 2 aryl or 0112 substituted aryl. Most preferably, is selected from 3-methylthiophenyl, 4methylthiophenyl, 4-phenylbenzyl, 4-methoxybenzyl, 4-biphenyl, 3-methoxybenzyl, 4-(2thienyl)benzyl, 4-(4-methyl)phenylbenzyl, 4-(4-trifluoromethyl)phenylbenzyl, 4-(4nitrilo)phenylbenzyl, 4-(2-pyridinyl)benzyl, piperonyl, 3-methoxbenzyl, 4-chlorobenzyl, and 4-nitrobenzyl.

R

2 is preferably hydrogen or a hydrocarbon selected from the group substituted lower alkyl, cycloallcyl, substituted cycloalcyl each having one to 6 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, R 22 NR'cOR 2

OR

20 substitUted alkyl, cycloalkyl, substituted cycloalkyl each having one to 6 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group i0 consisting of halo, R 22

NR

2 and OR 20 More preferably, R2 is isopropyl.

R

3 is preferably -NR 4 R, wherein R, and R, are each selected from the group consisting of hydrogen, ailkyl, heterocyclyl, acyl, aryl, heteroaryl, aralkyl, heteroaralkyl, alkyl alkenyl, alkyl alkynyl, each having one to 20 carbon atoms, which ailkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarvlalkyl,are optionally substituted with from 1 to 3 substituients independently selected from the group consisting of halo, aryl, heteroaryl,heterocyclyl, R 22

SR

20

S(O)R

2 1

SOR

21

SONR

0

R

2 3 Nle&R2 NR 20 C0R 1

NR

20 C0 2

R

1

NR

20

CONR

20 R21,

NR'

0

SO,R

1

OR

20 CN, CO,R 0

CONR

20 R'3, and COR 2 Q. In some compositions, and R, are preferably each H, -CH 2

CH

2 OH -CHR'CHOH, or -CH,CHR'OH, -CH.,CH,NH,,

CHR'CH.NH

2

-CH,CHR'NR

2 wherein R' is hydrogen or alkyl having from I to 6 carbon atoms.

R"

0 is preferably a member selected from the group consisting of H, C,.

8 allcyl.

R

2 is preferably a member selected from the group consisting of C, 3 alkyl, which alkyl is optionally substituted with 1 to 2 substituents independently selected from the group of halo,

OF

3 ON, OR 20

SR

20

N(R

20 2

R

22 is preferably a member selected from the group consisting of 0 1 3 alkyl, aryl, heteroaryl which alkyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, ailkyl, mono- or dialkylamino, alkyl or aryl, ON, O-CIalkyl, and CF 3 Preferred 2,6,9-trisubstuted purine compositions of this invention include hydroxyethyl)[9-(methylethyl)-6-( {[4-(trifluoromethyl)phenyl~rnethyl I amino)purin-2yl]aniino ethan- 1 -ol, {((2S)oxolan-2-yl)methyl](6- [(4-fluorophenyl)methyl] amino)} -9- (methylethyl)purin-2-yl)anmjne, [((2R)oxolan-2-yl)methyl)(6- fluorophenyl)methyl] amino)} -9-(methylethyl)purin-2-yI)amine, (2-aminoethyl)(6- dichlorophenyl)methyflamino -9-(methylethyl)purin-2-yl)amine, (2-aminoethyl)[6-( chloro-3-(trifluoromethyl)phenyl~methyl) amino)-9-(methylethyl)purin-2-yljamine, chlorophenyl)methyl]amnino -9-(methylethyl)purin-2-yl)anino]-3-methylbutanamide, (2amino-2-methylpropyl)(6- [(4-chlorophenyl)methyllamino -9-(methylethyl)purin-2yl)an-ine, 3 -(2-[bis(2-hydroxyethyl)aminoj-6- f [4-chlorophenyl)methyl] amino) purin-9yl)butan-2 -one, [(4-chlorophenyl)methyl]axnno -9-(methylethyl)purin-2-yl)amino]-3-methylbutan- 1 ol, {2-[(2-aminoethyl)amino]-9-(methylethyl)purih-6yl) am aino)methyl]benzenesulfonamide, 2-[(2-hydroxyethyl)(6- rnethoxyphenyl)methyl] amino)} -9-(methylethyl)purin-2-yl)amiino]ethan- I -ol, 2(2 hydroxyethyl) methylethyl)-6-[(4-phenylphenyl)anino]purin-2-yl} amino)ethan-l1-ol, 16 [(2-amino-2-propyl)amino] -9-(methylethyl)purin-6-yl} [(4-hlorophenyl)methyl]amine, {2-[(2-aminoethyl)amino]-9-( methylethyl)purin-6-yl }[(4-chlorophenyl)methyl]amine, [(2-aminopropyl)arnino]-9-( methylethyl)purin-6-yl }[(4-chlorophenyl)methyl~amine and 2-[(2-aminoethyl)(6- [(4-chlorophenyl)methyl]amino -9-(methylethyl)purin-2yl)am-inojethan- 1 -ol.

There are some limitations to the scope of R 1 R, and R 3 As mentioned above, when R 3 is 2-hydroxyethylamino and R 2 is methyl, R 1 cannot be amino, 3 -methyl-2-butenyl amino, benzylamino, or m-hydroxybenzyl-amino. When R, is 2-hydroxyethylamino and R 2 is isopropyl, R 1 cannot be benzylamino, mhydroxybenzylamino, or 3-methylbutylamino. When R 3 is 2-hydroxyethylamnino and R, is 2-hydroxyetbyl, R 1 cannot be benzylamino. When R 3 is 2-propanol-2-methylamino or 2-dimethylaminoethylamino and R2 is methyl, R 1 cannot be benzylaxnino.

The following are definitions for certain terms used herein.

"Halo" or "Halogen" alone or in combination means all halogens, that is, chioro (Cl), fluoro bromo iodo "Hydroxyl" refers to the group -OH.

"Thiol" or "mercapto" refers to the group -SH.

"Alkyl" alone or in combination means an alkane-derived radical containing from I to preferably 1 to 15, carbon atoms (unless specifically defined). It is a straight chain alkyl, branched alkyl or cycloalkyl. Preferably, straight or branched alkyl groups containing from 1 more preferably 1 to 8, even more preferably 1-6, yet more preferably 1-4 and most preferably 1-2, carbon atoms, such as methyl, ethyl, propyl, isopropyl, butyl, t-butyl and the like. The term "lower ailkyl" is used herein to describe the straight chain ailkyl groups described immediately above. Preferably, cycloalkyl groups are monocyclic, bicyclic or tricyclic ring systems of 3-8, more preferably 3-6, ring members per ring, such as cyclopropyl, cyclopentyl, cyclohexyl, adamantyl and the like. Alkyl also includes a straight chain or branched alkyl group that contains or is interrupted by a cycloalkyl portion. The straight chain or branched alkyl group is attached at any available point to produce a stable compound.

Examples of this include, but are not limited to, 4-(isopropyl)-cyclohexylethyl or 2-methylcyclopropylpentyl. A substituted alkyl is a straight chain alkyl, branched alkyl, or cycloalkyl group defined previously, independently substituted with 1 to 3 groups or substituents of halo, hydroxy, alkoxy, alkylthio, alkylsulfinyl, alkylsulfonyl, acyloxy, aryloxy, heteroaryloxy, amino optionally mono- or di-substituted with alkyl, aryl or heteroaryl groups, amidino, urea optionally substituted with alkyl, aryl, heteroaryl or heterocyclyl groups, aminosulfonyl optionally N-mono- or N,N-di-substituted with alkyl, aryl or heteroaryl groups, alkylsulfonylamino, arylsulfonylamino, heteroarylsulfonylamino, alkylcarbonylamino, arylcarbonylamino, heteroarylcarbonylamino, or the like.

"Alkenyl" alone or in combination means a straight, branched, or cyclic hydrocarbon containing 2-20, preferably 2-17, more preferably 2-10, even more preferably 2-8, most preferably 2-4, carbon atoms and at least one, preferably 1-3, more preferably 1-2, most preferably one, carbon to carbon double bond. In the case of a cycloalkyl group, conjugation of more than one carbon to carbon double bond is not such as to confer aromaticity to the ring.

Carbon to carbon double bonds may be either contained within a cycloalkyl portion, with the exception of cyclopropyl, or within a straight chain or branched portion. Examples of alkenyl groups include ethenyl, propenyl, isopropenyl, butenyl, cyclohexenyl, cyclohexenylalkyl and the like. A substituted alkenyl is the straight chain alkenyl, branched alkenyl or cycloalkenyl 18 group defined previously, independently substituted with 1 to 3 groups or substituents of halo, hydroxy, alkoxy, alkylthio, alkylsulfinyl, alkylsulfonyl, acyloxy, aryloxy, heteroaryloxy, amino optionally mono- or di-substituted with alkyl, aryl or heteroaryl groups, amidino, urea optionally substituted with alkyl, aryl, heteroaryl or heterocyclyl groups, aminosulfonyl optionally N-mono- or N,N-di-substituted with alkyl, aryl or heteroaryl groups, alkylsulfonylamino, arylsulfonylamino, heteroarylsulfonylamino, alkylcarbonylamino, arylcarbonylamino, heteroarylcarbonylamino, carboxy, alkoxycarbonyl, aryloxycarbonyl, heteroaryloxycarbonyl, or the like attached at any available point to produce a stable compound.

"Alkynyl" alone or in combination means a straight or branched hydrocarbon containing 2-20, preferably 2-17, more preferably 2-10, even more preferably 2-8, most preferably 2-4, carbon atoms containing at least one, preferably one, carbon to carbon triple bond. Examples of alkynyl groups include ethynyl, propynyl, butynyl and the like. A substituted alkynyl refers to the straight chain alkynyl or branched alkenyl defined previously, independently substituted with 1 to 3 groups or substituents of halo, hydroxy, alkoxy, alkylthio, alkylsulfinyl, alkylsulfonyl, acyloxy, aryloxy, heteroaryloxy, amino optionally mono- or disubstituted with alkyl, aryl or heteroaryl groups, amidino, urea optionally substituted with alkyl, aryl, heteroaryl or heterocyclyl groups, aminosulfonyl optionally N-mono- or N,N-disubstituted with alkyl, aryl or heteroaryl groups, alkylsulfonylamino, arylsulfonylamino, heteroarylsulfonylamino, alkylcarbonylamino, arylcarbonylamino, heteroarylcarbonylamino, or the like attached at any available point to produce a stable compound.

"Alkyl alkenyl" refers to a group where R is lower alkyl, or substituted lower alkyl, R'"'may independently be hydrogen, halogen, lower alkyl, 19 substituted lower alkyl, acyl, aryl, substituted aryl, hetaryl, or substituted hetaryl as defined below.

"Alkyl alkynyl" refers to a groups -RC=CR' where R is lower alkyl or substituted lower alkyl, R' is hydrogen, lower alkyl, substituted lower alkyl, acyl, aryl, substituted aryl, hetaryl, or substituted hetaryl as defined below.

"Alkoxy" denotes the group -OR, where R is lower alkyl, substituted lower alkyl, acyl, aryl, substituted aryl, aralkyl, substituted aralkyl, heteroalkyl, heteroarylalkyl, cycloalkyl, substituted cycloalkyl, cycloheteroalkyl, or substituted cycloheteroalkyl as defined.

"Alkylthio" denotes the group -SR, 2 where R is lower alkyl, substituted lower alkyl, aryl, substituted aryl, aralkyl or substituted aralkyl as defined herein.

"Acyl" denotes groups where R is hydrogen, lower alkyl substituted lower alkyl, aryl, substituted aryl and the like as defined herein.

"Aryloxy" denotes groups -OAr, where Ar is an aryl, substituted aryl, heteroaryl, or substituted heteroaryl group as defined herein.

"Amino" denotes the group NRR', where R and R' may independently by hydrogen, lower alkyl, substituted lower alkyl, aryl, substituted aryl, hetaryl, or substituted hetaryl as defined herein or acyl.

"Amido" denotes the group -C(O)NRR', where R and R' may independently by hydrogen, lower alkyl, substituted lower alkyl, aryl, substituted aryl, hetaryl, substituted hetaryl as defined herein.

"Carboxyl" denotes the group -C(O)OR, where R is hydrogen, lower alkyl, substituted lower alkyl, aryl, substituted aryl, hetaryl, and substituted hetaryl as defined herein.

"Aryl" alone or in combination means phenyl or naphthyl optionally carbocyclic fused with a cycloalkyl of preferably 5-7, more preferably 5-6, ring members and/or optionally substituted with 1 to 3 groups or substituents of halo, hydroxy, alkoxy, alkylthio, alkylsulfinyl, alkylsulfonyl, acyloxy, aryloxy, heteroaryloxy, amino optionally mono- or disubstituted with alkyl, aryl or heteroaryl groups, amidino, urea optionally substituted with alkyl, aryl, heteroaryl or heterocyclyl groups, aminosulfonyl optionally N-mono- or N,N-disubstituted with alkyl, aryl or heteroaryl groups, alkylsulfonylamino, arylsulfonylamino, heteroarylsulfonylamino, alkylcarbonylamino, arylcarbonylamino, heteroarylcarbonylamino, or the like.

"Substituted aryl" refers to aryl optionally substituted with one or more functional groups, halogen, lower alkyl, lower alkoxy, alkylthio, acetylene, amino, amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Heterocycle" refers to a saturated, unsaturated, or aromatic carbocyclic group having a single ring morpholino, pyridyl or furyl) or multiple condensed rings naphthpyridyl, quinoxalyl, quinolinyl, indolizinyl or benzo[b]thienyl) and having at least one hetero atom, such as N, 0 or S, within the ring, which can optionally be unsubstituted or substituted with, halogen, lower alkyl, lower alkoxy, alkylthio, acetylene, amino, amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Heteroaryl" alone or in combination means a monocyclic aromatic ring structure containing 5 or 6 ring atoms, or a bicyclic aromatic group having 8 to 10 atoms, containing one or more, preferably 1-4, more preferably 1-3, even more preferably 1-2, heteroatoms 21 independently selected from the group 0, S, and N, and optionally substituted with 1 to 3 groups or substituents of halo, hydroxy, alkoxy, alkylthio, alkylsulfinyl, alkylsulfonyl, acyloxy, aryloxy, heteroaryloxy, amino optionally mono- or di-substituted with alkyl, aryl or heteroaryl groups, amidino, urea optionally substituted with alkyl, aryl, heteroaryl or heterocyclyl groups, aminosulfonyl optionally N-mono- or N,N-di-substituted with alkyl, aryl or heteroaryl groups, alkylsulfonylamino, arylsulfonylamino, heteroarylsulfonylamino, alkylcarbonylamino, arylcarbonylamino, heteroarylcarbonylamino, or the like. Heteroaryl is also intended to include oxidized S or N, such as sulfinyl, sulfonyl and N-oxide of a tertiary ring nitrogen. A carbon or nitrogen atom is the point of attachment of the heteroaryl ring structure such that a stable aromatic ring is retained. Examples of heteroaryl groups are pyridinyl, pyridazinyl, pyrazinyl, quinazolinyl, purinyl, indolyl, quinolinyl, pyrimidinyl, pyrrolyl, oxazolyl, thiazolyl, thienyl, isoxazolyl, oxathiadiazolyl, isothiazolyl, tetrazolyl, imidazolyl, triazinyl, furanyl, benzofuryl, indolyl and the like. A substituted heteroaryl contains a substituent attached at an available carbon or nitrogen to produce a stable compound.

"Heterocyclyl" alone or in combination means a non-aromatic cycloalkyl group having from 5 to 10 atoms in which from 1 to 3 carbon atoms in the ring are replaced by heteroatoms of 0, S or N, and are optionally benzo fused or fused heteroaryl of 5-6 ring members and/or are optionally substituted as in the case of cycloalkyl. Heterocycyl is also intended to include oxidized S or N, such as sulfinyl, sulfonyl and N-oxide of a tertiary ring nitrogen. The point of attachment is at a carbon or nitrogen atom. Examples of heterocyclyl groups are tetrahydrofuranyl, dihydropyridinyl, piperidinyl, pyrrolidinyl, piperazinyl, dihydrobenzofuryl, dihydroindolyl, and the like. A substituted hetercyclyl contains a substituent nitrogen attached at an available carbon or nitrogen to produce a stable compound.

"Substituted heteroaryl" refers to a heterocycle optionally mono or poly substituted with one or more functional groups, halogen, lower alkyl, lower alkoxy, alkylthio, acetylene, amino, amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Aralkyl" refers to the group -R-Ar where Ar is an aryl group and R is lower alkyl or substituted lower alkyl group. Aryl groups can optionally be unsubstituted or substituted with, halogen, lower alkyl, alkoxy, alkylthio, acetylene, amino, amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Heteroalkyl" refers to the group -R-Het where Het is a heterocycle group and R is a lower alkyl group. Heteroalkyl groups can optionally be unsubstituted or substituted with halogen, lower alkyl, lower alkoxy, alkylthio, acetylene, amino, amido, carboxyl, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Heteroarylalkyl" refers to the group -R-HetAr where HetAr is an heteroaryl group and R lower alkyl or substituted lower alkyl. Heteroarylalkyl groups can optionally be unsubstituted or substituted with, halogen, lower alkyl, substituted lower alkyl, alkoxy, alkylthio, acetylene, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Cycloalkyl" refers to a divalent cyclic or polycyclic alkyl group containing 3 to carbon atoms.

"Substituted cycloalkyl" refers to a cycloalkyl group comprising one or more substituents with, halogen, lower alkyl, substituted lower alkyl, alkoxy, alkylthio, acetylene, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Cycloheteroalkyl" refers to a cycloalkyl group wherein one or more of the ring carbon atoms is replaced with a heteroatom N, 0, S or P).

"Substituted cycloheteroalkyl" refers to a cycloheteroalkyl group as herein defined which contains one or more substituents, such as halogen, lower alkyl, lower alkoxy, alkylthio, acetylene, amino, amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Alkyl cycloalkyl" denotes the group -R-cycloalkyl where cycloalkyl is a cycloalkyl group and R is a lower alkyl or substituted lower alkyl. Cycloalkyl groups can optionally be unsubstituted or substituted with e.g. halogen, lower alkyl, lower alkoxy, alkylthio, acetylene, amino,; amido, carboxyl, hydroxyl, aryl, aryloxy, heterocycle, -substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Alkyl cycloheteroalkyl" denotes the group -R-cycloheteroalkyl where R is a lower alkyl or substituted lower alkyl. Cycloheteroalkyl groups can optionally be unsubstituted orsubstituted with e.g. halogen, lower alkyl, lower alkoxy, alkylthio, amino, amido, carboxyl, acetylene, hydroxyl, aryl, aryloxy, heterocycle, substituted heterocycle, hetaryl, substituted hetaryl, nitro, cyano, thiol, sulfamido and the like.

"Pharmacologically acceptable salt" a salt prepared by conventional means, and are well known by those skilled in the art. The "pharmacologically acceptable salts" include basic salts of inorganic and organic acids, including but not limited to hydrochloric acid, 24 hydrobromic acid, sulphuric acid, phosphoric acid, methanesulphonic acid, ethanesulfonic acid, malic acid, acetic acid, oxalic acid, tartaric acid, citric acid, lactic acid, fumaric acid, succinic acid, maleic acid, salicylic acid, benzoic acid, phenylacetic acid, mandelic acid and the like. When compounds of formula I-Ef include an acidic function such as a carboxy group, then suitable pharmaceutically acceptable cation pairs for the carboxy group are well known to those skilled in the art and include alkaline, alkaline earth, ammonium, quaternary ammonium cations and the like. For additional examples of the types of compounds that are "pharmacologically acceptable salts," see Berge et al, J. Pharm. Sci. 66, 1 (1977).

If the final 2,6,9-trisubstituted purine compound of this invention contains a basic group, then an acid addition salt of the composition may be prepared. Acid addition salts of the compounds of this invention are prepared in a standard manner in a suitable solvent from the parent compound and an excess of acid, such as, but not limited to, hydrochloric, hydrobromic, sulfuric, phosphoric, acetic, maleic, succinic, citric or methanesulfonic. The hydrochloric salt form is especially useful.

If the final 2,6,9-trisubstituted purine compound contains an acidic group, then cationic salts of the composition may be prepared. Typically the acidic parent compound is treated with an excess of an alkaline reagent, such as, but not limited to, hydroxide, carbonate or alkoxide, containing the appropriate cation such as but not limited to, Na, K Ca" 2 and NH Certain of the compounds form inner salts or zwitterions which are also acceptable.

The compounds of this invention are useful in inhibiting cell proliferation in mammals including humans. The 2,6,9-trisubstituted purines are useful in for example in treating autoimmune diseases, e.g. rheumatoid arthritis, lupus, type I diabetes, multiple sclerosis, cancer, cardiovascular disease such as restenosis following balloon angioplasty and atherectomy, restensosis following vascular modifying surgical procedures, host vs graft disease, gout, polycystic kidney disease and other proliferative diseases whose pathogenesis involves abnormal cell proliferation. In particular, the compositions of this invention are useful in treating cancers including cancers including lymphoyd neoplasm, cancer of the colon, breast cancer, ovarian cancer, pancreatic cancer, and cancers derived of endotherical cells.

The method of treatment comprises the administration parenterally, and orally, of an effective quantity of the chosen compound of this invention, preferably dispersed in a pharmaceutical carrier. Therapeutically useful amounts of the composition of this invention will generally range from about 0.01 to about 100 mg/kg, but will be readily determined by one skilled in the art depending upon the route of administration, and the age and condition of the patient. Therapeutically useful amounts of the composition of this invention may be administered from one to ten times daily or more for acute or chronic disease. No unacceptable toxicological effects are expected when compounds of the invention are administered in accordance with the present invention.

The compounds of this invention are also useful as antiinflammatory and antifungal agents. As such, the compositions of this invention are useful for treating antiinflammatory and fungal infections in humans, animals, and fungal infections in plants.

Pharmaceutical compositions including the compounds of this invention, and/or 26 derivatives thereof, may be formulated as solutions or lyophilized powders for parenteral administration. Powders may be reconstituted by addition of a suitable diluent or other pharmaceutically acceptable carrier prior to use. If used in liquid form the compositions of this invention are preferably incorporated into a buffered, isotonic, aqueous solution.

Examples of suitable diluents are normal isotonic saline solution, standard 5% dextrose in water and buffered sodium or ammonium acetate solution. Such liquid formulations are suitable for parenteral administration, but may also be used for oral administration.

It may be desirable to add excipients such as polyvinylpyrrolidinone, gelatin, hydroxycellulose, acaia, polyethylene glycol, mannitol, sodium chloride, sodium citrate or any other excipient known to one of skill in the art to pharmaceutical compositions including compounds of this invention. Alternatively, the pharmaceutical compounds may be encapsulated, tableted or prepared in an emulsion or syrup for oral administration.

Pharmaceutically acceptable solid or liquid carriers may be added to enhance or stabilize the composition, or to facilitate preparation of the composition. Liquid carriers include, but are not limited to syrup, peanut oil, olive oil, glycerin, saline, alcohols and water. Solid carriers include, but are not limited to, starch, lactose, calcium sulfate, dihydrate, teffa alba, magnesium stearate or stearic acid, talc, pectin, acacia, agar or gelatin. The carrier may also include a sustained release material such as, but not limited to, glyceryl monostearate or glyceryl distearate, alone or with a wax. The amount of solid carrier varies but, preferably, will be between about 20 mg to about 1 gram per dosage unit.

The pharmaceutical dosages are made using conventional techniques such as, but not limited to, milling, mixing, granulation, and compressing, when necessary, for tablet forms; or milling, mixing and filling for hard gelatin capsule forms. When a liquid carrier is used, 27 the preparation will be in the form of a syrup, elixir, emulsion or an aqueous or non-aqueous suspension. Such a liquid formulation may be administered directly or filled into a soft gelatin capsule.

The Examples which follow serve to illustrate this invention. The Examples are intended to in no way limit the scope of this invention, but are provided to show how to make and use the compounds of this invention. In the Examples, all temperatures are in degrees Centigrade. RT indicates room temperature.

EXAMPLE 1 The compounds of this invention are prepared by conventional methods of organic chemistry. The reaction sequence outlined in the synthesis scheme below is a general method useful for the synthesis of compounds of this invention. 2,6-dichloropurine is dissolved in butanol and the appropriate R, amine is added. After heating for several hours, the reaction mixture is cooled, and the compound 1 is obtained. To compound 1, is added, sodium hydride followed by R 2 and compound 2 is isolated. To compound 2, R 3 is added in solution with N-methylpyrrolidinone. The mixture is heated for an appropriate period followed by purification leading to the desired compound.

Ci R

R

3 Ct N N-methylpyrrolidinoe butanol.A 0., R2 R2 3 2 The following compound was prepared according to the method above.

Preparation of 2-chloro-6-(4-methoxybenzylamino) purine The 2,6-dichloropurine (4.06 g, 21.5 mmol) was suspended in n-butanol (150 ml) and the 4-methoxybenzylamine was added (3.4 ml, 26 mmol). The solution turned clear and then cloudy a few minutes later. The solution was heated at 120°C for 2 hr and then cooled. The n-butanol was evaporated followed by suspension of the residue in water and diethyl ether mixture. A solution of 2N NaOH (1.3ml, 26 mmol) was added and the solution stirred for min before filtration. The filtered precipitate was washed with water and a small portion of ether and then dried under vacuum. The residual liquor was left overnight and more crystals were collected the next day and washed with diethyl ether.

Preparation of 2-chloro-6-(4-methoxybenzylamino)-9-isopropylpurine (2) 2-chloro-6-(4-methoxybenzylamino) purine was suspended in dry DMF (5 ml) and treated with sodium hydride, 60% dispersion (82 mg, 2.06 mmol). The suspension was stirred for 30 min over which time it became a clear yellow/green solution. 2-Iodopropane (0.280 mL, 1.7 eq.) was added over 5 min and the resultant solution stirred for 2 days. Water was added and the solution and extracted with ethyl acetate. The organic layer was evaporated to give the product isopropyl purine.

Preparation of 2-diethanolamino-6-(4-methoxybenzylamino)-9-isopropylpurine, The purine (1.65g, 4.98 mmol) was dissolved in DMSO (12 mL) and diethanolamine (4 mL) and then heated at 140 0 C for 2-3 days and then at 160°C for 1 day. The solution was cooled and water saturated butanol was added (100 mL). The solution was then washed with water (3 x 50 mL), before being evaporated to give a brown oil. The residue was chromatographed over silica gel eluting with ethyl acetate, followed by 3% methanol in ethyl acetate to give the product. 'H-NMR(5 CDC13): 7.29(br s 1H), 7.25(d, 2H), 6.94(br s. 1H), 6.83(d. 2H), 5.43(br 4.63(br s. 2H), 4.53(m 1H), 3.86(t. 4H), 3.76(m, 7H), 1.47(d 6H).

Table 1 identifies compounds of this invention that were prepared according to the synthesis method set forth in this Example.

ITABLE 1 Cornpoun s Prepared By The Method of Example 1

R

1 X (4-methoxyphenyl)methylamino 3-cyanopropyl Cl (4-methoxyphenyl)methylamino 3-chloropropyl c I (4-methoxyphenyl)methylamino benz1 ClI (4-methoxyphenyl)rnethylamino (4-methylcarboxyphenyl)methyl Cl (4-methoxyphenyl)methylamino 2-(N-phtyhaloyl)ethyl Cl (4-rnethoxyphenyl)methylamino isopropyl Ethanolaniine (4-methoxyphenyl)methylaznino isopropyl Diethanolammne (4-methoxyphenyl)rnethylarnino 3-inethylbutyl clI (4-methoxyphenyl)methylamino 2-methylbutyl Cl (4-methoxyphenyl)methylamino cyclopentyl c I (4-methoxyphenyl)methylamino (3-nitrophenyl)methyl clI (4-methoxyphenyl)methylamino (4-nitrophenyl)methyl Cl (4-methoxyphenyi)methyiamino ethyl Cl (4-methoxyphenyl)methylainino propyl

I

(4-methoxyphenyl)rnethylamino (3-methylphenyl)methyl c 1 (4-methoxyphenyi)methylamino (4-methylphenyl)rnethyl c heptylamino H ClI N-benzyl-N-hydroxylamino H Cl propylamino H clI noradarnantylammo H ClI cyclobutylamino H clI 3-methoxypropylamino H Cl 2-methoxyethylamino H ClI cyclapentylaxnzno H ClI I -hydroxy-2-methyl-2- H ClI propylamiuo I-benzylpipenidinyl)-4-annno H ClI heptylamino Methyl ClI

R

1 X R, R,21 N-benzyi-N-hydroxyl mmo Methyl Cl propylaniino Methyl Cl noradamantylamuno Methyl Cl cyclobutylammno Methyl Cl 3-methoxypropylainino Methyl Cl 2-rnethoxyethylamino Methyl Cl cyclopentylamino Methyl Cl I -hydroxy-2-methyl-2- Methyl Cl propylaino I -benzylpipenidinyl)-4-ammno Methyl Cl Methyl Cl diinethoxyphenvl)methylarnino________________ (2-methoxyphenyl)methylamino H Cl (2-pryidinyl)methylamino H Cl (3,4-dimethoxyphenyl)ethylamino H Cl (3-pyridinyl)methylamino H Cl (4-pyridinyl)methylamrino H Cl 6-bydroxy- I-hexylammno H C I phenethylamino H ClI (2.benzothiazolyl)amino H Cl H Cl dirnethoxyphenyl)methyalrnino (2-metboxyphenyl)rnethylamino Methyl ClI (2-pyridinyl)methylaxmno Methyl Cl (3,4-ditnethoxyphenyl)ethylarniino Methyl clI WO 00/44750 RI R2R (4-methoxyphenyl)xuethylamino Methyl Cl (3-pyridinyl)methylarmno isopropyl H Cl l-hydroxy-6-hexylamino H ClI phenethylamino H clI (2-benzothiazolyl)qmino H clI (4-methoxyphenyl)methyiaihno H c I 3-phenyl- I -propylaino isopropyl 3-hydroxypyrrolidino (2-indanyl)amino H ClI (4-methoxyphenyl)ethylano H C I (4-nitrophenyl)methylamino H C I (2,6-difluorophenyi)methylamino H ClI 3-phenyl-lI-propylaxnino H Cl (2-indanyl)aino Methyl ClI (4-methoxyphenyl)ethylammio Methyl ClI (4-nitrophenyl)methylamino Methyl C I (2,6-difluorophenyl)methylamino Methyl c I cyclopropyimethylamino Methyl clI 4-(1,2-methylenedioxyphenyl) H Cl1 nmthylamuno H ClI aminosulfonylphenyl)methylammno (cyclohexanol)- I-methylamino H Cl (2-benzimidazoly)methylamino H Cl cyclohexylmnethylarnino H Cl1 (4-methoxyphenyl)methylamino H Cl R,-X 1, (4-methoxyphenyl)methylamino isopropyl (2-hydroxy- I hydroxyrnethyl)ethylanaino cyclopropyimethylarnino isopr opyl 3-amino-2-hydroxypropylamilo 4-(l 2-rnethylene- Methyl cl dioxyphenyl)rnethylaiio (4-axninosulfonyl- Methyl Cl phenyl)methylamino________________ (cyclohexanol)- 1 -methylamino Methyl c I (2-benziiniidazolyl)methylamfino Methyl c cyclohexyiniethylamino Methyl (3-pyridinyl)nmethylamnlfo Methyl

I

(4-pyridinyl)methylamino 2-methyipropyl 1 6-hydroxyhexylamino cyclopentyl ClI phenethylamino propyl c 1 (2-benzothiazolyl)axnino ethyl clI 3-phenyl- 1 -propylainino isopropyl Cl (2-indanyl)axnino 2-metbylpropyl Cl 2-(4-methoxyphenyl)ethylamino cyclopentyl Cl (4-nitropbenyl)methylaniino propyl Cl (2,6-difluorophenyl)methylaxnmo ethyl Cl (4-methoxyphenyl)methylanino isopropyl c I 3-phenyl- 1 -propylamino isopropyl 4-hydroxypiperidino (2-indanyl)amino H Cl 2-(4-metboxyphenyl)ethylarnino H Cl (4-nitrophenyl)methylarnimo H Cl (2,6-difluorophenyl)methylamno H Cl

[,R

(4-methoxyphenyl)methylainino H Cl (4-methoxyphenyl)methylamino isopropyl N-(2-cyanoethyl)-N-benzlano 3-phyenyl-l1-propylam~ino isopropyl 1 -(RS)-hydroxyrnethyl-3methylbutylamnino (2-indanyl)aniino isopropyl Cl 2-(4-Methoxyphenylethylaxnino isopropyl Cl (4-aitrophenvi)methylanaino isopropyl Cl (2,6-difluorophenyl)methylamino isopropyl Cl (4-methoxyphenyl)methylaznino isopropyl Cl (4-mcthoxyphenyl)mnethylamino isopropyl Pipenidino (4-methoxyphenyl)methylaniino isopropyl 3-hydroxypiperidino 3-phenyl- I -propylamnimo isopropyl I -(S)-hydroxymethyl-2-(4'imidazolyl)etbylanmino (2-indanyl)amino isopropyl diethanolainino (4-methyoxyphenyi)methylamino isopropyl diethanolamino (4-methyoxyphenyl)methylamino isopropyl 2-(S)-hydroxymetbylpyrrolidino (4-rnethyoxyphenyl)methylamino isopropyl diethnolanino (4-methyoxyphenyl)methylamino benzyl morpholino (4-methyoxyphenyl)methylamino 3-methylbutyl diethanolainino (4-rnethyoxyphenyl)methylaniino 2-metbylbutyl diethanolainino (4-methyoxyphenyl)methylamino cyclopentyl diethanolainino (4-methyoxyphenyl)methylamino (3-aitrophenyi)methylamino diethanolamino (4-methyoxyphenyl)methylamino (4-nitrophenyl)methylamino diethanolarnino (4-rnethyoxyphenyl)methylamino ethyl diethanolamuino (4-methyoxyphenyl)methylaznino propyl diethanol amino (4-methyoxyphenyl)methylaxnino (3-methylphenyl)methylamino diethanolamino ,heptylarnino, (4-methylphenyl)methylaxnino, diethanolamino R,-X N-benzyl-N-hydroxyamino Methyl diedhzolammo propylamino Methyl diethanolamino noradamantylamino Methyl diethanolanuno cyciobutylamino Methyl diethanolamino 3-methoxypropylamino Methyl diethanolamino 2-methoxyethylamino Methyl diethanolamino cyclopentylainino Methyl diethanolannno I -hydroxy-2-methyl-2- Methyl diethanolamino propylamino 4-(lI -benzylpiperidinyl)anino Methyl diethanolannno (4-metboxyphenyl)rnethylaxnino Methyl diethanolamino (4-methoxyphenyl)methylamino isopropyl diethanolammno (2,4-dimethoxy-phenyl) isopropyl 3-hydroxypyrrolidino methylanmno (2-methoxyphenyl)methylamino Methyl 2-(3'indolyl)etbylamino (2-pyridinyl)methylaniino Methyl diethanolamino Methyl diethanolamino ditnethoxyphenyl)ethylamino (3-pyridinyl)methylamino Methyl diethaolamino (4-pyridinyl)methylamino Methyl diethanolamino 6-hyclroxy- I -hexylamino Methyl dietbanolzmino phenethylamaino Methyl diethanolamino (2-benzotbiazolyl)arnino Methyl diethanolamino 3-phenyl- I -propylamino Methyl diethanolamino (2-indanyl)amino Methyl diethaolainino 2-(4-methoxyphenyl)ethylamino Methyl diethanolamino (4-nitrophenvl)rnethylaxnino Methyl diethanolamino (2,6-difluorophenyl)methylamino Methyl diedhaoLamino R,-X R,

R,

cyclopropylmethylamino Methyl diethanolamino 1,2-rnethylenedioxy- Methyl diethanolamino phenyl)methylamino (4-aminosulfonylphenyl)- Methyl diethanolarnino methylamino (cyclohexanol)- 1 -methylammuo Methyl diethaolamino (2-benzimidazolyl)methylaniino Methyl diethanolamino cyclohexylmethylimino Methyl diethanolammino (3-pyridyl)methylamino Methyl diethanolainino (4-pyridyl,)methylamino 2-methylpropyl diethanolamino 6-hydroxy- I -hexylanuno cyclopentyl diethanolamino 2-phenethylamino propyl diethanolanino (2-benzothiazolyl)amino ethyl diethanolammno 3-phenyl- I -propyiamino isopropyl diethanolamino (2-indanyl)arnino 2-methyipropyl diethanolamino 2-(4-methoxyphenyl)ethylainino cyclopentyl diethanolarnino (4-nitropbenyl)methylammno propyl diethanoiaino (2,6-difluorophenyl)methylanino ethyl diethaoLamino (4-methyoxyphenylmethylamiino isopropyl diethanolammno (4-methyoxyphenyl)rnethylamino isopropyl 1-hydroxyrnethylcyclopenrylamino (4-methyoxyphenyl)methylamino isopropyl 2-(R.S)-hydroxymethylpiperidino cyclopropylxnethylanaino isopropyl 2,3-dihydroxy- 1 -propylamino 1,2-methylenedioxyphenyl) isopropyl cl1 methylamino (4-aminosulfonylphenyl) isopropyl ClI methylammlo (cyclohexanol)- I -rnethylarnio isopropyl IC 1 R,-X R2 R3 (2-benzimidazolyl)amino isopropyl cl cyclohexyimethylaino isopropyl Cl 3-phenyl- I -propylamino isopropyl Cl cyclopropylmethylamino cyclopentyl Cl 1,2-metbylenedioxyphenyl) isopropyl diethanolammno methylarnio (4-methoxyphenyl)methylamino isopropyl diethanolammo (4-methoxyphenyl)metbylanmmo isopropyl diisopropylamino (4-methoxyphenyl)methylamiino isopropyl (trans-2-hydroxycyclohexyl)immno (4-methoxyphenyl)methylamino isopropyl 1-hydroxy-3- (4-methoxyphenyl)metbylanino isopropyl 5-(S).(2,2-dimethyl-4(S)phenyldioxalanyl)amino (4-methoxyphenyl)methylarnino isopropyl I -imidazolyl)propyianiino (4-methoxyphenyl)methylamino isopropyl 4-hydroxyl-4-phenylpiperidino (4-methoxyphenyl)methylamino isopropyl (2-benzylthio-1 hydroxymethyl)ethylamino (4-methoxyphenyl)methylamino isopropyl N-methyl-N-(2-hydroxy-2-(3,4dihydroxyphenyl)ethyl)amino (4-methoxyphenyl)methylamino isopropyl diallylanino (4-methoxyphenyl)methylaniio isopropyl Piperazmo (4-methoxyphenyl)methylaruno isopropyl (+/-)N-methyl-N-(2-hydroxy-2phenylethyl)aniina (4-methoxyphenyl)methylarnino isopropyl (anilinomethyl)pyrrolidino (4-mttioyphnyi~ethianino soprpyl(+/-)N-(2-propenyl)-N-2-(4- (4-mthoyphnyl~ethlamno ioprpylhydroxy-2-methylpentyl)amiino (4-methoxyphenyl)methylaniino isopropyl N-(2-hydroxyethyl)-N-(3hydroxypropyl)amino R2 R (4-methoxyphenyl)methylamino isopropyl Di-N- 1-(2-hydroxy-2isopropyl Di-N-2-(3-hydroxybutyl)amino Example 2 This example describes a method for preparing compounds of this invention according to the following general synthesis scheme:

R

1

X

c N Ethanol, Reflux CH K2CO3

DMF

RR3H

R

3 N N NMP, 140 o C C N R, R2 Preparation of {2-chloropurin-6-yl} [(4-chlorophenyl)methyl]amine: To a suspension of 15 g (0.0794 mol) of 2,6-dichloropurine in 250 mL of absolute ethanol was added 12.7 mL (0.0873 mol) of triethylamine and 10.62 mL (0.0873 mol) of 4chlorobenzylamine. The mixture was refluxed at 80 °C for 16h (the formation of creamy white precipitate was observed). The reaction mixture was cooled and the precipitated product was removed by filtration. The precipitate was washed with ethanol (350 mL) and dried in high vacuum for 24h. Product was characterized by 'H-NMR.

Preparation of {2-chloro-9-(methylethyl)purin-6-yl} [4-chlorophenyl)methyl] amine: To a solution of 6 g (0.020 imol) of 2-chloro-6-(4-chlorophenyl)methylaminopurine in 41 mL of anhydrous DMF was added 5.64 g (0.041mol) anhydrous. potassium carbonate and 3.41 mL,(0.035 mol of 2-iodopropane) and stirred at room temperature for 16h. To the mixture 500 mL of water was added and stirred for Ih. The precipitate was filtered, washed with water (350 mL), and dried in vacuum oven at 50 'C for 16h. The product was obtained as an off white solid and characterized by 'H-NMR.

Preparation of {2-[(2-aminoethyl)amino-9-(methylethyl)purin-6-yl} chlorophenyl) methyll amine: To a solution of 3.36 g (0.01 mol) of (2-chloro-9-(methylethyl)purin-6-yl} [4chlorophenyl)methyl]amine in 13 mL of anhy. 1-methyl-2-pyrrolidinone was added 4.68 mL (0.70 mol) of 2-aminoethylamine and the mixture was heated at 140 oC for 24h. The compound was subjected to variable gradient chromatography on silica gel with dichloromethane/methanol mixtures and yielded 1-methyl-2-pyrrolidinone. The mixture was dissolved in dichloromethane and extracted with water (520 mL). The organic layer was dried over anhydrous. Sodium sulfate and evaporated to an off white solid. The product was characterized by 'H-NMR and purity checked by RP-HPLC (YMC C-18 column; 50X4.4 mm; S-5 120 A' 0.1% TFA-water/ 0.1% TFA-acetonitrile).

Table 2, below identifies compounds of this invention that were prepared according to the general synthesis method set forth in this Example. In Table 2, MS Mass Spectrum and MH+ mass of parent molecular ion plus one hydrogen atom.

TABLE 2 COMPOUNDS PREPARED BY METHOD OF EXAMPLE 2 R,X R, R, T MS(MB+) (4-MethvlphenvI)methylamino Isopropyl 2-Aminoethylamino 340 (2.4-Dichlorophenyl)methylamino Isopropyl Diethanolamino 439 (2,4-Dichlorophenyl)methylamino Isopropyl 2-Aminoethylamino 394 (3-Methylphenyi)methylamino Isopropyl Diethanolamino 385 (3-Methylphenyl)muthylamino Isopropyl 2-Aminoethylamino 340 (3-Methylphenyi)methylamino Isopropyl 2-(N2-Dimethylamino)-N1- 458 benzvlethylamino (4-Trifluoromethvlphenyl)methylamino Isopropyl 2-Aminoethylamino 394 (4-Trifluoromethylphenyl)methylamino Isopropyl 1-Hydroxymethyl-2- 437 methylpropylamino Isopropyl Diethanolamino 507 trifluoromethylphenyl)methylamino I -I Isopropyl 2-Aminoethylammo 462 trifluoromethylphenvl)methvlamino I I I RX RRms

M+)

Isopropyl I -Hydroxyrnethyl-2- 505 trifluorornethvlphenvl)rnethviamino methyl ropylaino' (3-Chlorophenyl)methylamina Isopropyl 1 -Hvdroxvmethvlethvlamino 375 (2-Trifluoromethyl-phenyl)methvianhino Isopropyl 1 -Hydroxymethylethylaxnino 409 (4-Chloro-3-trifluoromethyl- Isopropyl 1 -Hydroxymethylethyian-dno 443 phenyl)inethylammno (3-Chlorophenvl)rnethylamino Isopropyl 2-Hvdroxyethylamino 361 (2-Trifluromethvl-phenvlmethylamino Isopropyl 2-Hydroxyethylamino 395 (4-Chloro-3-trifluoromethyl- Isopropyl 2-Hydroxyethyiammno 429 phenyl)rnethvlamino (3-Chlorophenyl)mcthylamino Isopropyl (I,2S)-2-Hydroxy- 1- 451 methyl-2:phenylethylanino (2-Chlorophe vflmethylan-ino Isopropyl Diethanolaino 450 pI Isopropyl Diethanolamino 407 (1 -Naphthyl)methylamino Isopropyl Diethanolamino 421 (2-Chlorophenvl)methvlaniino Isopropyl 2-Armnoethylamino 360 I Isopropyl 2-Arninoethvlamilo 362 (I -Naphthyl)methvlarrmino Isopropyl 2-Aminoethylamino 376 (2-Chlorophenyl)methylarnino Isopropyl 1 -Hydroxyrnethyl-2- 403 methviethviamino Isopropyl 1 -Hydroxymethyl-2- 405 methylethvlanaino (1 -Naphthyl)methylaxnino Isopropyl 1 -Hydroxymethyl-2- 419 methvlethylainino (3-Methvlphenvl)methylaniino Isopropyl 2-Aminopropylamino 354 (2-Chlorophenvl)methvlaniino Isopropyl 2-Axninovropylamliflo 374 (3-Chlorophenyl)methylaxruno Isorpropyl 2-An-dnopropylarnino 374 (2,S.Difluorophenyl)methylaxn'no Isopropyl 2-Axninopropvlammno 376 (1 -Naphthyl)rnethvlamimo Isopropyl 2-Aniinopropylaxnino 390 Isopropyl Diethanolamino 473 t-ifluoromethvlphenvl)methylanaino (3-Chlorophenyl)methylamino Isopropyl 1-Hydroxymethyl-2- 403 methyipropylaiino (2-Trifluoromethylphenyl)methylamino Isopropyl 1 .Hydroxymethyl-2- 437 methyipropyLmnino (3-Methylphenyl)methylamino 1 Isopropyl 2-(2-Hydroxyethylaniino)- 384 (2-Chlorophenyl)methylamino Isopropyl 2-(2-Hydroxyethylamlino)- 404 ethvlamino (3-Chlorophenyl)methylamino j Isopropyl 2-(2-Hydroxyethylarnino)- f 404 Isopropyl 2-(2-Hydroxyethylanhino)- 406 (1 -Naphthyl)methylamino Isopropyl 2-(2-Hydroxyethylamino)- 420 _________ethvlamino 1 I]sopropyl 2-(2-Hydroxyethylammno)- 506 p~henvl)methylamino J________ethylanuno (4-Isopropylphenyl)methylamino Isopropyl 2-(2-Hydroxyethylamnio)- 412 ethylamuno (2-Trifluoroxnethylphenyl)methylan-Lino Isopropyl 2-(2-Hydroxyethylammno)- 438 ethylamino (4-Methylphenyl)methylanxno Isopropyl 2-(2-Hydroxyethylaxnino)- 384 ethvlanmmnoII R,X

MS(MHF+)

(4-Chloro-3- Isopropyl 2-(2-Hydroxyethylamino)- 472 trifuorometb 1 henvl')methylamino ethylamino Isopropyl 2-(2-Hydroxyethylarmino)- 472 triflucromethylphenyl)methvlamiino ________ethviainino (3 ,5-Dichiorophenyl)methylammno Isopropyl 2-(2-Hydroxyethylaxnino)- 438 ethylanuno (1 R)-(4-Methyiphenyl)ethyamilo Isopropyl 2-(2-Hydroxyethylainfo)- 398 ethvianimo (IR)-(2-Naphthyl)ethylamino isopropyl 2-(2-Hydroxyetylamfilo)- 434 ethvlamnio (2,4-Dichloro-6- Isopropyl 2-(2-Hydroxyethylamfilo)- 1 452 methviphenyl)methylamino (4-Trifluoromethypheny1)methylamiflo Isopropyl 2-(2.Hydroxyethylamino)- 438 (3-Metbviphenvl)rnethylamilo Isopropyl (2S)-pyrrolomethylaniino 380 (2-Chlorophenvl)methvlamino Isopropyl (2S-prOlomethvLnio 400 (3-Chlorophenyl)methylanio isopropyl (2S)-pyrrolomethylaniino henyl)methvianino Isopropyl (2S)-pyrro omethvlamino 402 (I -Naphthyl)methylanuno ISOpropyl (2S)-pyrrolomethylamlflo 416 (3 ,5-Bistuifluoromethyl- Isopropyl (2S)-pyrrolomethylamiflo 502 phenyl)methylamino (4-Isopropylphenvl)methvamiDo Isopropyl (2S)-pyrrolomethvlamino 408 (2-Trifluor-omethylphenv)methylaliino Isopropyl (2S)-pyrrolomethylanino 434 (4-Methylphenyl)methviamilo Isopropyl (2S)-pyrrolomethviamino 380 (4-Chloro-3- Isopropyl (2S)-pyrrolomethylamino 468 trifluoromethylphenvl)nethvanmino Isopropyl (2S)-pyrrolometbylanino 468 trifluoromethylphenyl)methviamino 43 Isopropyl (S-vrlmtvann 3 (I R -4-Methy1 henyP)ethylamino j Isopropyl -(2S)-pyffolomethylamino 394 (IR)-2-Naphthy1)ethylamino Isopropyl -(2S)-pyrrolomethylaino 430 (2,4-Dichloro-6- Isopropyl (2S)-pyffolomethyla O 448 methylphenyl)mnethylaniino (4-TrifluOTOMethylphenvl)methvlahino Isopropvl (2S)-pvrrolomethylamnu'o 434 (3-Methylphenyl)methylamino Isopropyl 2-Hydroxy- 355 methylethvlano (2-Chlorophenyl)methylaniino Isopropyl 2-Hydroxy-l1-(S)- 375 methylethylaino isopropyl 2-Hydroxy- 377 methylethylamino (1 -Naphtbyl)rnethylamino Isopropyl 2-Hydroxy- 391 methylethylamino Isopropyl 2-Hydroxy- 1 477 phenyl)methylamino methylethylamino (4-Isopropylphenyl)methyi n1 n Isopropyl 2-Hydroxy- 383 methylethylamino (4-Methylphenyl)rnethyinmino Isopropyl 2-Hydroxy- 355 methylethylamino Isopropyl 2-Hydroxy-l-(S)- 443 trifluoromethylphenyl)rethvlamino methylethylaniino (5-Fluoro-2- Isopropyl 2-Hydroxy-l-{S)- 427 tnflucromethylphenylhmethvlaniino methylethylamino I j Isopropyl 2-Hdroylami 409 R,X R R MS(ME+) I -(4-Methylphenyl)ethylamino Isopropyl 2-Hydroxy-l1-(S)- 369 1-(2-Naphthyl)etbylamino Isopropyl 2-Hydroxy-l-(S)- 405 methylethylarnino (2 ,4-Dichloropheny)nethyimmno Isopropyl 2-Hydroxy- 1 409 methylethviaxnino (2,4-DichLoro-6- Isopropyl 2-Hydroxy-l1-(5)- 423 methylphenvl)methylamino methylethylamino (4-Trifluoromethyiphenyl)methylamino Isopropyl 2-Hydroxy-l1-(5)- 409 rnethylethylarnino (3-Methviphenyl)methylamino Isopropyl 2-Hvdroxyethylaxnino34 (2-Chlorophenvl)methylanhno Isopropyl 2-Hydroxvethylaniino 361 Isopropyl 2-Hydr xvethylamino 363 (1 -Naphthvl)methylarmino Isopropyl 2-Hvdroxvethylamino 377 (3 ,5-Bistrifluoromethyl- Isopropyl 2-Hydroxyethylaxmino 463 phenyl)methviamino (4-Isoprop~vlpbenvi')methviaminmo Isopropyl 2-Hydroxyethviamino 369 (4-Methvlphenv1'~methvlan-ino Isopropyl 2-Hydroxvethvianmino 341 T Isopropyl 2-Hydroxyethylaxnino 429 triflucoromethvlphenflmethylamino..

(5-Fluoro.2- Isopropyl 2-Hydroxyethylaxnhno 413 uifluoromethylphenyi)methylamino (3 .5-Dichlorophenvl)rnethvl:mino T Isopropyl 2-Hvdroxvethylainino 395 (lR)-(4-Methvlphenv1)ethylamino Isopropyl 2-Hydroxvethylamino 355 (IR)-(2-Naphthyl)ethylamino Isopropyl 2-Hydroxvethvlaniino 391 (2,4-Dichlorophenyl~methvlamino Isopropyl 2-Hvdroxyethylaxnino (2,4-Dichloro-6- Isopropyl 2-Hydroxyethylamino40 methylphenvl)metbvlamino- (4-Trifluoromethylphenyl~methylaxnino Isopropyl 2-Hvdroxyethvlamino 395 (3-Methylphenyl)methylamino Isopropyl 1- 369 Hvdroxymethylgropylaniino (2-Cb~lorophenyl)methylammno Isopropyl 1- 389 Hvdroxymethylpropylaxnino (3-Cblorophenyl)methyammno Isopropyl 1-389 Hydroxymethylpropylamino Isopropyl 1- 391 Hvdroxvmethvlpropylamino (1 -Naphthyl)methylarnino Isopropyl 1- 405 ____________Hvdroxvinethvipropylamuno Isopropyl 1- 491 pbenyl)methylaxnino ________Hydroxymethylpropylamino (4-Isopropyiphenyl)rnethylamino Isopropyl 1- 397 Hvdroxymcthylpropylainino (2-Trifluoromethylphenyl)methylamino Isopropyl 1-423 ydroxymethylpropyLamino (4-Methylphenyl)methylamino Isopropyl 1- 1 369 Hydroxymethvipropylamino (4-Chloro-3 Isopropyl 1- 457 trfluoromethvlphenyl)methylanmino j _______Hydroxymetbvlpropylanxino Isopropyl 1- 457 trifhioromethylphenyl)rnethylamino ________Hvdroxvinethylpropylamino (S-Fluoro-2- 1- 441 trffluoromcthylphenyl)methvlarnino jHvdroxymethyipropylaminoI R,X J_ R, MS(M]H+) (3 ,5-Dichlorophenyl)methylamino Isopropyl 1- 423 _____________Hydroxyniethvipropvlamino (1 R)-(4-Methylphenyl)ethylamino Isopropyl 1 383 (1 R)-(2-Naphthyl)ethylamino Isopropyl I1- 419 Hydroxymethylpropylarino (2,4-Dichlorophenyl)rnethylamino Isopropyl 1- 423 1HydroxyrnetylpropylarnioI_______ (2,4-Dichloro-6- Isopropyl 1- 437 metbvlphenvl)methylamino Hydroxvtnethylpropylaxm (4-Trifluorometbylphenyl)methylamino Isopropyl 1- 423 1 Hydroxymethylpropylarnio (3 ,5-Dichloropbenyl)methylamino Isopropyl ]2-(2-Hydroxyethylamino)- 438 I ethylaino (3 ,5-Dichlorophenyl)metbvlanmmo Isopropyl I(2S)-pyrrolomethviamino 434 (4-Chlorophenyl)methylaino Isopropyl 2-Aminocyclohexyl i. o 414 (4-Chlorophenyi)methylarxmo Isop~rotpyl 3-Amiinocyclohexylaminn 414 (3-Fluoro-6-trifluoromethyl- Isopropyl Diethanolqmino phenyl)methvlamino Isopropyl 2-Aminoethylamino 428 Isopropyl 2-Aminopropylamino I 476 phenyl)methylaino____________ (2-Trifluoromethyl-pbenvl)metaylamino Isopropvl 2-Aminopropylamino 408 (4-methyiphenyl)methvlamino Isopropyl I 2-Axninopropylarnino j 354 (4-Chloro-3-trifluoromethyl- Isopropyl 2-Aminopropylamino 442 phenyl)rnethylaxnino_____________ Isopropyl 2-Aminopropylamino 442 p env mre FRl-I -(4-meth lphenyl)ethylamn Isopropyl 2-ainoethylamin2 354 1 -(2-Naphtbvl')ethylamnio Isopropyl 2-Aminoethylamino 390 Isopropyl I -Hydroxymethyl-2- 471 Phenvl)rnethylamino methylpropylarnino (3-fluoro-6-trifluorometbyl- Isopropyl 1 -Hydroxymethyl-2- 455 phenvl)methylamino methylpropylamino [Ri- I -(4-methylphenyl)ethvlamino Isopropvl 2-Aminopropylamino 368 [RI- I -(2-Naiphthvl)ethvlaxnino Isopropyl 2-Aminopropylamimo F 404 (4-Trifluoromethvl-phenyl)methvlainino J Isopropyl f 2-AmninopropyLamino 408 (3-Methylphenyl)methylaznino Isopropyl 2-Amiino-2- 368 (2-Chlorophcnyl)methylamino Isopropyl 2-Amnino-2- 388 methylpropylamino (3-Chiorophenyl)methylammo Isopropyl 2-Amino-2- 388 methylpropylamino Isopropyl 2-Amino-2- 390 methvlpropylamino (I -Naphthyl)methylamino Isopropyl 2-Amino-2- 404 methyipropylarnino (3 ,5-Bis-trffluoromethyl- T Isopropyl 2-Amino-2- 490 phenyl)methylamino tmethylpropylamino (2-Trifluorometbyl-phenyl)methylamino Isopropyl 2-Amino-2- 422 metlhylpropylarnino (4-Methylphenyl)rnethylaniino T Isopropyl 2-Amino-2- 368 T methylpropylamino R,X R, R, MS(MH+) (4-Chloro-3-trifluoromethyl- Isopropyl 2-Amino-2- 456 phenyl)methvlamino methylpropylamino Isopropyl 2-Amino-2- 456 phenyl)methylamino methylpropvlamino (3-Fluoro-6-triffluorometbyl- Isopropyl 2-Amino-2- 440 phenvi)methylamino methylpropylamino 1 -(4-Methylphenyl)ethylamino Isopropyl 2-Amino-2- 382 methylpropylamino 1 -(2-Naphtbyl)ethylamino Isopropyl 2-Amino-2- 418 methvlpropylamino (4-Trifluoromethyl-phenyl)methylamino Isopropyl 2-Anino-2- 422 1 1 methylpropylamino EXAMPLE 3 This Example describes a method for preparing compounds of this invention. The synthesis method disclosed in this Example is only slightly modified from that disclosed in Example 1.

C1

RI

butariol, A

N>

4 2 NaH R2 N

N>

R

3

N

R2 6

R

3 A

N

N-methytpyrrolidinone Cr"JAN

N

R2 The following compound was prepared according to the method above.

Preparation of 2,6-dichloro-9-isopropylpurine To a solution of 0.67g of 2,6-dichloropurine in 5mL of dry DMF at room temperature was added 0. 16gms I eq.) of 50% sodium hydride/oil powder. Upon cessation of hydrogen evolution, a large excess (2 mL) of isopropyl iodide was added to the anionic solution. This reaction solution was stirred for three days at ambient temperature. The reaction was quenched with 30 mL of water and extracted with ethyl acetate (350 mL). The organic extracts were combined and back washed with 350 mL of water followed by 20 mL of brine. The ethyl acetate solution was dried over anhydrous magnesium sulfate and evaporated. The compound was subjected to variable gradient flash chromatography on silica gel with hexane/ethyl acetate mixtures and yielded the N-9 product and the N-7 isomer.

Preparation of 2-chloro-6-anilino-9-isopropylpurine 2,6-dichloro-9-isopropylpurine (0.019 g, 0.081 mmol) was dissolved in butanol ml) and aniline (0.044 ml, 0.244 mmol) was added. The reaction mixture was heated to 120 0 C for 10 hr, cooled, diluted with EtOAc and washed 3 times with water. The mixture was dried over MgSO 4 and concentrated to an off white solid.

Preparation of 2-diethanolamino-6-(4-phenylanilino)-9-isopropylpurine A solution of 67mgs of 2,6-dichloro-N-9-isopropylpurine and 100mgs of 4-phenylaniline in 1 mL of n-octanol was heated to 80°C for 24 hours. The n-octanol was removed in vacuo and then replaced with 1 mL of 40% diethanolamine in DMSO. The solution was heated at 130 0 C for 48 hours. The reaction was cooled to ambient temperature then diluted with 10 mL of water and subsequently extracted with ethyl acetate (3X30 mL). The organic extracts were combined and back washed with 3X20 mL of water followed by 10 mL of brine. The ethyl acetate solution was dried over anhydrous magnesium sulfate and filtered and the solvent was evaporated. The 65mgs of crude product was crystallized from Tiff-ether solution.

Table 3 below identifies compounds of this invention that were prepared according to the general synthesis method set forth in this Example.

TABLE 3 Compounds Prepared By The Method Of Example 3 R,X (8-guinolinvI)amino Isopropyl CI (6-guinolinyl~aniino Isopropyl C1 (3-gwinolinyl)amino ISOPrOPYl CI aihno Isopropyl CI I Isopropyl C1 4-butvianilino Isopropvl C1 (8-guinolinyl)amino Isopropyl Diethanolamino (6-guinolinyl)armno Isopropyl Diethanolammlo (3-quinolinyl)amino ISOPTOPVI Diethanoiamnino anilmo Isopropyl Diethanolamino Isopropyl Diethanolamino 4-butviaihno Isopropyl Diethanolaniino (6-ethoxy-2-benzothiazoivl)amino Isopropyl C1 4-morpholino-2-methylamino Isopropyl CI (4-anuinosulf'onyl-p henvflmethviamino Isopropyl CI 4-bromoanilino Isopropyl diethanolamino 3,4-dichloroanilino Isopropyl diethanolamino 2-(2-(l1 -methyl)pyrrolidinvI)ethyaxnino Isopropyl diethanolainino 3-bromoanilino Isopropyl CI 4-methoxvanilino Isopropyl diethanolamnio 4-iodoaniluao Isopropyl C1 3-iodoanilino Isopropyl CI 3-rnethoxyanilino Isol ro I C1 I -piperidinyl)ethylamino ISOPropyl diethanolamino I -pyrrofidinvI)ethvlamino Isopropyl diethanolamino (I -indanyl)amino Isopropyl diethanolarnino 2-(6-ethoxybenzothiazolylamino Isopropyl diethanolaxnino 4-morpholino-2-methylan-ino Isopropyl diethanolamino (4-aminosulfonyl-phenyl)methvlamino Isopropyl diethaolarmno 4-bromoanilino Isopropyl diethanolamino 3 .4-dichioroanilino Isopropyl diethanoiamino 2 -methyl)pyrrolidinvl~ethylamino Isopropyl diethanolammno 3-brmoanlinoIsopopyldiethanolamino 4-methvoxvanilino Isopropyl diethanolammno 4-iodoanilino Isopropyl diethanolaxnuno 3-iodoanilino Isopropyl diethaolamino 3-methoxvanifino Iorpldiethanolamino 1-piperidinyl)ethylamino Isopropyl diethanolammio 1-pyrrohiyl)ethylamj no Isopropyl diethanolamino (I -indanyl')aniino Isopropyl diethanolamino 3-iodonflino Isopropyl diethanolamino RAX

R,

3-phenoxyanilino Isopi-opyl diethanolarnino 4-iodoanilino Isopropyl diethanolamino 4-phenoxvanilino Isopropyl diethanolamino 3-phenoxvanilino Isopropyl diethanolamino 2-fluorenvlammno Isoipropyl diethanolarnino 1 -fluorenylamino Isopropyl diethanolamino 2-anthracenviarnino Isopropyl diethanolamino 1 -anthracenviamino Isopropyl diethanolano 2-(6-ethoxybenzotiazolyl)amino Isopropyl diethanolamino (1 -indanvl)ammno j Isopropyl diethanolamino 2-(6-ethoxybenzothiazoivl)axpino j Isopropyl diethanolano 4-inorpholino-2-metbviamino Isopropyl diethanolamino (4amnoulonl-hny~mthlaiio Isopropyl diethanolamino 4-bromoanilino Isopropyl diethanolaiiino 3,4-dichloroanilino Isopropvl diethanolaino I -methyl)pyrrolidinyl)ethylan-ino Isopropyl diethanolamnio 3-bromoanilino Isopropyl diethanolamino 4-methoxyanjiimo Isopropyl diethanolaniino 4-iodoanilino Isopropyl diethanolamino 3-iodoanilino j Isopropyl diethanolainino 3-methoxyanilino Isopropyl diethanolamino 241 -piperidinyl)ethyiainino Isopropyl diethanolarnino 241 -pyrrolidinyl)ethviarnino Isopropyl dietbanolaxnjno (1 -indanyl)amino Isopropyl diethanolamino 3-iodoanilino Isopropyl diethanolamino 3-pheoxvanilino Isopropyl diethanolamino 4-iodornilno Isopropyl diethanoiamino 4-phenoxyanilino Isoprcypyl diethanolainino 3-phenoxvanilino Isopropyl diethanolamino 2-fluorenvlamo -Isopropyl diethanolarnino I -fluorenviaiio Isopropyl diedmaolamino 2-anthracenylarnino Isopropyl diethanolamino 1 -anthracenylaxrno Isopropyl diethanolamino 2-(6-etboxvbenzothiazolyI amino Isopropyl diethanolamino (2-biphenyi)methylamino Isopropyl diethaolamino (4-biphenyl)methylaniino Isopropyl diethanolamino 2-naphthylmethyLimino Isopropyl diethanolamino 1 -naphthvimethvlamino Isopropyl diethanolainino (4-Chlorophenyl)methylamino Isopropvl Diethanolamino (4-Fluorophenyl)methylamino Isopropyl Diethanolamino (4-Methoxyphenyl methylainino Isopropyl 4 -Trifluoromethylphenyl)methyiarnino Isobutvi Diethanolamino (4-TrifluoromethylphenyI meth lamino Isopropyl Diethanoamno 4 -Chlorophenyl)rnethylamino Isopropyl (S)-2-An uno-3phenylpropylamino (4-Fluorophenyl)methvlaniino Isopropyl 2-Axninoethylaniino (4-Fluorophenyl)methylamino Isopropyl 1-Hydroxymnethyl-2methyl-propylaniino (4-Fluorophenyl)methyl ni o Isopropyl I -Hydroxymethyl-2rnethylmpropvlamino (4-Chlorophenyl)methylamino Isopropyl (D)-l1-Hydroxymethyl-2- I methyl-propylarnino RX RR (4-Cblorophenyl)niethylaqnino Isopropyl 1-Hydroxymethyl-2- (4-Cb~orophnyl~mthylamno Ispropy 2-Hyroy-2-henyl (4-Chlorophenyl)methylaniino Isopropyl -yrx2phnl hyroxethletbvlmin (4-Chlorophenyl)methylaxnino Isopropyl 2-Amino-N2-(2bvdroxyethyl)ethylamino (4.-Chlorophenyl)metliylaniino Isopropyl (S).2-Phenyl- 1-carboxamida- ___________ethylantino (4-Chlorophenyl)methylami o lsopropyl 2-Amino-N2-(2hydroxyethyl)-N I (hvdroxvethyl)ethvlamino (4-Sulfonarnidophenyl)methvlamino Isopropyl 2-Aminoetbylamino (4-Fluorophenyl)methylamino 2-Oxo-3-butvl Diethanolamino (4-Fiuoropheiivl)methylamino 2-Oxo-3-butvl 2-Anminoethylamino (4-Chlorophenvl)methylamino 2-Oxo-3-butvl Diethanolamino (4-Chlorophenvl)methvianiino 2-Qxo-3-butvl 2-Aniinoethylamino (4-Methylphenyl)methylamino Isopropyi 1 -Hydroxyrnetbyl-2-methylpylanaino (3-Methylphenyl)methylamino Isopropyl 1 -Hydroxyrnethyl-2-methyl- (4-Chlorophenyl)methyiamino Isopropyl 2-(N2-dimethylamino)-Nl- (4-Chlorophenyl)methylaniino Isopropyl 1 -Carboxamido-2-methyl- (4-Chlorophenyl~methylam~p Ipoyl2Aicylamino (4-Chlorophenyl)methylamino Isopropyl 2-AniinopropyLanino (4-Cblorophenvl)methvlaxmno Isopropy) (4-Cbilorophenyl)methylamino Isopropyl 2-Amino-2-methyl- (4-Chlorophenyl)methylamino Isopropyl 1- (Hydroxymethyl)propylaniin 0 (4-Chlorophenyl)methylamino Isopropyl (Hydroxymethyl)propylarniin 0 (4-Chlorophenyl)methylamino Isopropyl 1- (Hydroxymethvl)ethvlamino (4-Chiorophenyl)methylamino Isopropyl I (Hydroxymethvl)ethlaxn-no (4-Chlorophenyl)methylainino Isopropyl Hydroxypropylaniino (4-Chlorophenyl)methylamino Isopropyl Hydroxyprorviarino (4-Fluorophenyl)methviarnino Isopropyl 2-Amino-propylaxnino (3-Chloropbenyl)methvlamino Isopropyl Diethanolamino (3-Chlorophenyl)methylaxnino Isopropyl 2-Aminoethvlanhno (2-Trifluoromethylphenvl)methylamino isapro yl Diethaolamino (2-Trifluoromethvlphenyl)methyl 0in Isopropyl 2-Anainoethylamino (4-Chloro '-3-trifluoromethyiphenyl) Isopropyl Diethanolanaino methvlam' -niotvamino (4-Chloro-3-trifluoromethvlphenvl) Isopropyl 2Ainehlmn

R,XR,,

(3 ,5-Dichlorophenyl)methvlamino ISOProPYI Diethanolainino (3 ,5-Dichlorophenyl)methylamino Isopropyl 2-Amino-ethvlaxnino (2-Trifluoromethyl-phenyl)methylamino Isopropyl 2-(N2-dimethylammno)-N 1benzyl-ethylamino (3-Chlorophenvl)rnethviamimo Isopropyl Diethanolamino (3-Chlorophenyvhmethylarmino ISOPTDPYI 2-Aminocthvlammio (2-Trifluoromethvlphenyl)methylamino Isopropyl Diethanoiamino (2-Trifluoromethvlphenyl)methylamino Isopropyl 2-Arninoethylanmino (4-Chloro-3- .Isopropyl Diethanolanino trifluoromethylphenv'~methviamino (4-Chloro-3- Isopropyl 2-Aminoethylamuno trifluoromethylphenyl)methvlamino (3 .5-Dichlorophenyl)methvlamino Isopropyl Diethanolamino (3 ,5-Dichlorophenvl)methylan-ino Isopropyl 2-Aminoethvlamino (2-Trifluoromethylphenyl)methylamino Isopropyl 2-(N2-Dimethylamino)-NlI benzvletbviamino (3-ChLorophenyl)methylamino Isopropyl 2-(N2-Diznethylamino)-N I1benzvlethviamino Isopropyi 2-(N2-Dimethylaamino)-N 1 benzvlethylamino (4-Ghloro-3- Isopropyl 2-(N2-Dimrethylaamino)-N I1- U-Mu~oromethvlphenvl)metbvlamino benzviethviamuno (4-Cblorophenyl)methylamino Isopropyl 2-Amino-2rethylpropylamino (4-Fluorophenyl)methylamino Isopropyl Tetrahydrofuranyl)methylam (4-Fluorophenyl)methylamino Isopropyl Teahydrofuranylmethylam (4-Methylphenvflrethylamino Isopropvl Diethanolanino (4-Fluorophenyl)methylan-ino Isopropyl 2-Hydroxy- 1 methvlethvlainino (4-Fluorophenyl)methylaniino Isopropyl (S)-2-Hydroxy-2methviethvlan'uno (4-Fluorophenyl)methylamin Isopropyl (R)-2-Hydroxy-2methylethlammno (4-Fluorophenyl)methylamino Isopropyl I- ___________HvdroxvmethylPTOPYlamn (4-Flurophenyi)methylamino Isopropyl 2-Amino-2- I methvipropvlamin 51 EXAMPLE 4 This Example describes a method for preparing compounds of this invention. The synthesis method disclosed in this Example is only slightly modified from that disclosed in Example 1.

Cl

CI

N N NaH N C"N 7 R 2 C1 N NC N N H 4 R2 bulanol RI R N N a. d, N-mothylpyrolidnone N N Ro N N upling CJ N N 6A R2 5A R2 The following compound was prepared according to the method above.

Preparation of 2,6-dichloro-9-isopropylpurine The 2,6-dichloropurine (5.00 g, 26.46 mmol) was suspended in 55 ml of dry DMF at room temperature and treated with sodium hydride, 60% dispersion (1.27 g, 31.75 mmol) added in portions. After stirring for 1 hr, 2-iodopropane (4.5 ml, 44.98 mmol) was added and the reaction stirred for 2 days. The reaction was poured into diethyl ether and washed once with saturated sodium bicarbonate solution and once with water. The mixture was dried over anhydrous sodium sulfate and concentrated in vacuo. The concentrate was chromatographed over silica gel eluting with 10% acetone in dichloromethane solution to give the desired N-9 52 alkylation product as a white solid.

Preparation of 2-chloro-6-(4-methylmercapto) anilino-9-isoproplypurine 2,6-Dichloro-9-isopropylpurine (0.15 g, 0.649 mmol) was dissolved in n-butanol (4 ml) and 4-(methylmercapato) aniline (0.089 ml, 0.714 mmol) and triethylamine (0.20 ml, 1.43 mmol) were added. The reaction mixture was heated at 80* overnight. The cooled reaction was diluted ethyl acetate and washed 1 x IM HCI, 1 x saturated sodium bicarbonate, and 1 x brine before being dried with anhydrous sodium sulfate and concentrated in vacuo. The residue was chromatographed over silica gel and eluting with 2% methanol in dichloromethane to give the desired product as a white solid.

Preparation of 2-diethanolamine-6-(4-methylmercapto) anilino-9-isopropylpurine (6A).

The purine (0.18 g, 539 mmol) was dissolved in N-methylyrrolidinone (3 ml) and diethanolamine (1 ml) and then heated at 120°C overnight. The cooled reaction was poured into diethyl ether and washed three times with water before drying over anhydrous sodium sulfate and concentrating in vacuo. The residue was chromatographed over silica gel eluting with 5% methanol in dichloromethane to give the desired product as an off-white solid. 'H- CDC1 3) 8.08(s,lH), 7.58(d, 2H), 7.47(s,1H), 7.18(d, 2H), 4.95(br s, 4.52(m, 1H), 3.94(m, 4H), 3.83(m,4H), 2.43(s, 3H), 1.47(d, 6H).

Preparation of 4-(2-thienyl) benzonitrile.

Some groups must first be synthesized before reacting with the 2,6-dichloro-9isopropylpurine. These groups can be synthesized through various coupling methods and other synthetic procedures known to those skilled in the art of organic synthesis.

To a pressure tube was added 4-bromobenzonitrile (0.20 g, 1.10 mmol), tetrakis(triphenylphosphine) palladium (0.127 g, 0.1 eq) and 2-thiopheneboronic acid 53 (0.211 g, 1.65 mmol). The reaction was flushed under vacuum and flushed with dry nitrogen three times. Following flushes, ethyleneglycol dimethyl ether (5.5 ml) and an aqueous solution of sodium carbonate (2.53 ml, IM) were added to the tube. The tube was then sealed and heated at 80 0 C overnight. The cooled reaction was the diluted with diethyl ether and washed twice with water before drying over sodium sulfate and concentrating in vacuo. The residue was chromatographed over silica gel eluting with 10% ethyl acetate in hexane to give the desired product as a white solid.

Preparation of 4-(2-thienyl) benzylamine.

The 4-(2-thienyl)benzonitrile (0.086 g, 0.464 mmol) was dissolved in dry tetrahydrofuran (1.6 ml) before lithium aluminum hydride (0.46 ml, 0.464 mmnol, 1 M in THF) was added dropwise. The reaction was allowed to stir at room temperature overnight.

TLC methanol in dichloromethane) still showed starting material remaining. Another I eq of LAH was added. After an additional hour, the reaction was quenched by the Fieser and Fieser method using wager (17.46.l1), aqueous sodium hydroxide solution (17.4641, soln.), and water (52.37 pl) added sequentially to the reaction. The reaction was then diluted with diethyl ether and water and extracted twice with diethyl ether before drying over sodium sulfate and concentrating in vacuo. The residue was carried on crude without any further purification.

Table 3 below identified compounds of this invention that were prepared according to the general synthesis method set forth in this Example.

Table 4 Compounds Prepared By The Method of Example 4 R 1XR2 R3 Ci Me ci ethanolaino Me ethanolaino cyclopropyilmethylaiio isopropyl

CI

cvclopropvLmethylaRninO isopropyl diethanolamino isopropyl Cl 3-phenoxvanilino isopropyl Cl 4-iodoanilino isopropyl

CI

3-amino uinolino isopropyl Cl isopropyl diethanolamino ciepoxymethyl

CI

4-methoxybenzviamino 2.3-dihydroxvpropvl diethanoLamino 4-phenylanilino isopropyl diethanolamino 4-phenylbenzvlamino isopropyl

CI

2-naphthalenyimethvlamino isopropyl Cl I -naphthalenvlmethviamino isopropyl Cl 2-phenylbenzvlamino isopropyl ci 3-guinolinylarruno isopropyl diethanolamino isopropyl diethanolamino 6-guinolinviamino isopropyl diethanolarnino 8-guinoiinylanino isopropyl diethanolanmino n-butylamino isopropyl Cl 4-(2-thiophen I~benziarnino isopropyl deithanolamino 4-(2-thiophenyl)benzylamino isopropyl Ci 3-thiomethoxyanilino isoprapyl Cl 4-thiomethoxyanilino isopropyl c 3-thiomethorxyanilino isopropyl diethanino 4-thiomethoxvanjlino isopropyl diethanoamino 4-(2-pvridinyl) benzyiamino isopropyl 3-methoxvbenzvlamino isprpyl j 3,4-diniethoxybenzvlamino isopropyl 3 ,4,5-trimethoxyenzylamino isopropyl c 3-methoxybenzvlanmno isopropyl diethanoiannno 3,4-diimethoxvbenzylarnino Isopropyl diethanolamino 3 ,4,5-trimethoxbenzvianiino Isopropvl diethanolamino 4-(3-thjiohenyI)benzvaiano Isopropyl Cl 4-(4-methoxphenvl) benzylamino Isopropyl

CI

4-(4-brornophenyl) benzylamino Isopropyl diethanolamimo metboxyphenvl) benzvlamino Isopropyi diethanolamino 4-(4-rnethoxyphenv) benzylamino Isopropyl diethanolaminio 4-(3-thiophenyl) benzylamino Isopropyl diethanolanino 4-(3-methylpheny) benzylamino Isopropyl Cl 4-(4-methylphenyl) benzylamio Isopropyl Cl 4-(4-trifluoromethylphenyl) Isopropyl Cl benzylanaino 3-(4 nitrilophenyl)anilino Isopropyl CI 3-(4-nitrilophenyl)anilino Isopropyl diethanolainino 4-C 2-pyridinyI)benzvan-dno Isopropyl Cl 4-(2-pryidinyI)benzvlajmno IsonropyI diethanolamino EXAMPLE This Example describes a method for preparing compounds of this invention. The synthesis method disclosed in this Example is only slightly modified from that disclosed in Example 1.

Cl Cl N N R 2 N N H N NaH H NN H 2 N HN 7

QI

R2 1 R 3

-CI

I pyndine

CH

2

CI

2 Cl N

N^

R3-, N N

N

H R2 The following compound was prepared according to the method above.

Preparation of 2 -amino-6-chloro-9-methylpurine The 2-amino-6-chloropurine (1.08 g, 6.4 mmol) was suspended in dry DMF (75 ml) and treated with sodium hydride, 60% dispersion (0.28 g, 7 mmol). The suspension was stirred for 15 min before iodomethane (0.44 ml, 7.06 rmol) was added and the resulting yellow solution stirred for 1 hr 45 min. The solid was filtered and the filtrate evaporated before addition of water for 10 min. The resulting solid was filtered and dried overnight to 56 give the product as a mixture of N-7 and N-9 alkylation products. The residual liquor was left overnight and more crystals were collected the next day and dried.

Preparation of 6-chloro-2-(2-methoxyacetylamino)-9-methylpurine The mixture of isomers from above was dissolved in dichloromethane and pyridine (2 eq) followed by treatment with methoxyacetyl chloride (4 eq). The reaction was stirred at room temperature until complete. The reaction was evaporated and filtered through a plug of silicia gel eluting with 2% methanol in dichloromethane followed by purification on a chomatotron using silica gel and eluting with 2% methanol in dichloromethane to isolate the desired product.

Table 5 identifies compounds of this invention that were prepared according to the synthesis method set forth in this Example.

Table Compounds Prepared By The Method of Example R1 R2 R3 Cl Me H Cl Me 2-methoxyacetylamino EXAMPLE 6 This Example describes a method for preparing compounds of this invention. The synthesis method disclosed in this Example is only slightly modified from that disclosed in Example 1.

Cl Ri RN Et3N

N

The 2,6-dichloropurine (5.0 g, 26.45 mmol) was suspended in n-butanol (50 ml) and excess n-butanol and washed precipitate with 100 ml 1M HC and 200 ml water. The solid RPreparation of 2-diethanolamino-6-(4-phenyl benzylamino) R The following compound was prepared according to the method above.

Preparation of 2-chloro-6-(4-phenyl benzylamino) purine The 2,6-dichloropurine (5.0 g, 26.45 mmol) was suspended in n-butanol (50 ml) and the 4-phenylbenzylamine (6.61 g, 29.1 mmol) and triethylamine (4.1 ml, 29.1 mmol) were added. The solution was heated at 120°C overnight then cooled. Filtered off product using excess n-butanol and washed precipitate with 100 ml 1M HCI and 200 ml water.. The solid was dried in vacuum over overnight at 70'C to give the desired product as a pale yellow solid Preparation of 2-diethanolamino-6-(4-phenyl benzylamino) purine 58 The 2-chloro-6-(4-phenyl benzylamino) punine (2.0 g, 5.96 mmol) was added together with diethanolarnine (11.4 ml, 119.2 rnmol) and N-methylpyrrolidinone (10 ml) and heated at 1 20'C overnight. The cooled reaction was poured into dichioromethane and washed twice with water. The organic layer was dried with anhydrous sodium sulfate and concentrated in vacuo to give the desired product as a pale green solid which was further dried in vacuum oven at 70'C for 2 days.

Preparation of 2-diethanolamino-6-(4-phen yl benzylamino)-9-methylpurine (11).

The 2-diethanolamino-6-(4-phenyl benzylamino) purine (0.050 g, 0.124 mmol) was dissolved in dry DMF and treated wit sodium hydride, 60% dispersion (5.5 mgs, 0. 13 6 rmol) for I hr. iodomethane (0.009 ml, 0.148 mmol) was added and the resultant solution stirred at room temperature overnight. Poured reaction into diethyl ether and washed twice with saturated sodium bicarbonate solution before drying over anhydrous sodium sulfate and concentrating in vacuo. The residue was chromatographed over silica gel eluting with methanol in dichloromethane to give the produce as a white solid.

1 H-NMvR(5, CDCI3): 7.55 7.41 (in, 4H) 7.35(m, 4H), 6.41 (br s, IH), 5.10(br s, 4.72 (br s, 2H), 3.86 (in, 4H), 3.74(m, 4H), 3.59(s, 3H).

Table 5 identified compounds of this invention that were prepared according to the synthesis method set forth in this Example.

Table 6 Compounds Prepared By The Method of Example 6 R2 113 4-phenylbenzylamino IMethyl diethanolamino 4-phenylbenzvlainino Cyclopentvl diethanolamio 4-phenvlberizvlamino Alvi diethanolamino 4-phenylbenzylamino Benzvl diethanolaniino 4-phenylbenzylamino 3-methylbutyl dietbanolamino 4-phenylbenzyiamino Isobutvi diethanolamino 4-phenvibenzykammio t-butvaceaxe diethanolamino I 4-phenylbenzyianuno IMethylacetate dietbaoaximo 4-phenyIbenzlammio Cyclobuty1 diethanolamino I 4-phenvibenzylamino Ethyl Idiethanolamino 4-phenvibenzvlamino Propyl diethanolarnino EXAMPLE 7 Composition of this invention were evaluated in the following assays.

CDK2 assays: Compositions of this invention were assayed to determine their CDK2 inhibitory activity. The assay system (total volume of 50 l1) contained 50 mM Tris-C1, pH 7.4, 10 mM MgC1 2 5 mM DTT, 1 gg of histone HI, 30 gM ATP (1 gCi of gamma 2

P

labeled ATP), 10 gg of BSA and 1 ng of purified CDK2. After incubation at 30 0 C for min, the reaction was terminated by the addition of 10 .1 of 10% TCA and the samples were blotted onto to nitrocellulose filters. These filters were washed extensively in 10% TCA and assayed for radioactivity. Blanks contained no enzyme.

To ascertain the potency of various compounds of this invention, the compounds were added to the above assay at concentrations ranging from 100 to 0.02 gg/ml. After incubation at 30 min., the assay tubes were processed as above. In all assays, various concentrations of olomoucine were added and were used as a standard positive control. The IC 50 (enzyme) listed in Table 7 is defined as the concentration required to inhibit CDK2 activity by 61 EXAMPLE 8 Cell Proliferation Assays: Early passage rat aortic smooth muscle cells (CV Therapeutics Cell repository) were seeded in 48 well dishes (Falcon, ml/well) at a density of 20,000 cells/ml of DME containing 5% heat inactivated bovine serum. The cells were incubated in a standard tissue culture incubator for 48 hr. The medium was aspirated and the wells were replenished with 0.2 ml of fresh medium. Compounds of this invention were added at concentrations ranging from 100 to 0.37 gg/ml. After 48 hr incubation, the medium was aspirated and the cultures were treated with 0.2 ml of saline 0.25 p of phenozine methosulfate solution containing MTS (Cell Titer 96® Aqueous Non-radioactive cell proliferation assay kit, Catalog G 5430, Promega, 2800 Woods Hollow Road, Madison, WI 53711-5399). The ICs 0 cells listed in Table 6 is defined as the concentration required to inhibit cell proliferation by Olomoucine at various concentrations was added and was used as a standard positive control.

TABLE 7 Bioactivity of Selected Representatives of this Invention R2 R3 ICo (g/mL) IC, (pg/mL) enzyme cells benzylamino Me Ethanolamino 7 4-methoxybenzylamino H Cl 60 NA 4-methoxybenzylamino Me Cl 6 4-methoxybenzylamino Me Ethanolamino 4 48 4-chlorobenzyloxy H Cl 60 NA 4-chlorobenzyloxy Me Cl 60 NA 4-chlorobenzyloxy trifluoromethyl Cl >60 NA 4-methoxybenzylamino isopropyl Cl 4 77 RI~ R2 R3 IC., (J9/mL) IC, p/ enzyme cells 4-methoxvbenzylaxnino isopropyl Ethanolaznino 4 43 4-methoxybenzylamino Me Diethanoiamino 4 48 4-methoxybenzyiamino 2-methylpropyl cl 60 ethanolamino Me Ethanolamino >60 4-methoxybenzylamino trifluoromethyl C 1 >60 4-methoxybenzylamina benzyl Cl >60 ethanolamino H Benzylamino >60 NA 4-methoxybenzyianiino isopropyl Diethanolamino 0.2 2.! 4-methoxybenzylainino perflUOTOisopropyl C 1 >45 NA 4-methoxybenzylamino perfluoroisopropyl Diethanolamno 40 NA 4-methoxybenzylamino ispropyl 3-pyrroline 1 12.5 4-methox ybenzyl amino hydroxyethyl Diethanolamino 0.5 62 4-methoxybenzylamino isopropyl Serinol 0.4 4-methoxybenzyiamino isopropyl I .3-diamino-2- 0.6 4-methoxybenzylamino 3-cyanopropyl CI >60 NA 4-methoxybenzylarnino 3-chioropTOPY1 CI >60 NA 4-methoxybenzyiamino benzyl Cl >60 NA 4-methoxybenzylamino Methyl 4- CI >60 NA carboxybenzyi 4-methoxybenzyiarnino Naphthaloylethyl cI >60 NA 4-chborobenzylamino Trifluoromethyl Jl 4-methoxybenzylamino isopropyl N-(2-cyaflopropyi)-

NA

N-(3pyridylmethyl)amino 4-methox ybenzyl amino isopropyl 2-(hydroxymethyl).I 3-rnethylbutan-2- NA amino 4-methoxybenzyiarnino isopropyl 3hydroxypipenidino NA cyclohexylmethylarnino isopropyl CI

NA

piperonylarnino isopropyl Diethanolamino 0.8 NA 4-methoxybenzylaniino isopropyl Diisopropanolamin 0.8 NA 0 R2 1 R3 (jig/ML) (pg/mL) enzyme cells aniiino isopropyl C1 I NA 4-rnethoxybenzyiamino isopropyl N-benzyl-N-2- INA hydroxyethylamino 4-phenylanilino isopropyl Diethanolamino 0.6 NA 4-phenylbenzyiamino isopropyl Diethanolamino 0.6 NA 4-phenyibenzylamina isopropyl 3-amino-l,2- 0.6 NA propanediol thiophenyl)benzyiaminoDehanamo0.

4-(4-methylphenyl) isopropyl Diethanolamino 0.6 NA benzylarnino 4-4-iopoylDitanlmio0.6

NA

trifluoromethylphenyl) iorplDehnlmn benzyalmino 4-thiomethoxyaniiino isopropyl C1 0.6 NA 3-(4-nitrilophenyl) isopropyl Diethanolarnino 0.5 NA anilino 3-thiomethoxyanilino isopropyl Diethanolamino 0.1 NA 4-thiomethoxyanilino isopropyl Diethanolarnino 0.07 NA 3-methoxybenzyianiino isopropyl CI 0.9 NA 4-(2-pyridi .nyl) isopropyl Diethanolamino 0.16 N benzylanino

N

3-methoxybenzylarnino isopropyl Diethanolamino 0.5 NA Isopropyl Diethanolamino 0.12 0.3 Cblorophenyl)methyla rnino Isopropyl Diethanolamino 0.15 2.2 Fluorophenyl)methyla miuno Isobutyl Diethanolamino 59 NA Trifluoromethylphenyl) methylamino Isopropyl Diethanolamino 0.56 NA Trifluoromethylphenyl) methviam-ino_______ Isopropyl (S)-2-Aniino-3- 1.07 NA Cbhlorcphenyl)methyla phenyipropylanimno midno Isopropyl 2- 0.17 1.4 Fluorophenyl)methyla Aminoethylam~ino Isopropyl (D--0.06 2.7 Fluorophenyl)methyla Hydroxymethyl-2wino methylprooviamino________ (4-Fluorophenyl) Isopropyl 0.19 NA methviamino Hydroxymethyl-2-_______ R-XR2 R3 ICso (ji g/ML) 1 ICso (11glML) 1 enzyme cells methylpropylamino Isopropyl I- 0.19 NA Cblorophenyl)methyla Hydroxymethyl.2mino methyl- Isopropyl 0.05 NA Chlorophenyl)methyla Hydroxymethyl.2- Mino methylpropylainino________ Isopropyl 2-Hydroxy-2- 0.08 Chlorophenyl)rnethyla phenyi-ethylamino ndno Isopropyl 2-Axnino-NI-(2- 0.07 0.2 Cblorophenyl)methyla hydrox~yethyl)ethyl m~ino ammno Isopropyl 2-Amino-N2-(2- 2.02 NA Chlorophenyl)methyla hydroxyethyl)ethyl Mino amino Isopropyl (S)-2-Phenyl-l- 1.07 NA Chlorophenyl)methyla carboxaniidorruno ethylamino Isopropyl 2-Amino-N2-(2- 0.43 NA Chlorophenyl)methyla hydroxyethyl)-N I mino (hydroxyethyl)ethy Isopropyl 2- 9 NA Sulfonarmdophenyl)me Aminoethylamino thvlamino 2-Oxo-3-butyl Diethanolamino 11 NA Fluorophenyl)methyla Mino 2-Oxo-3-butyl Diethanolamino 37 NA Chlorophenyl)methyla nino Isopropyl 2- 0.35 0.1 Chlorophenyl)methyla Am-inoethylamino ""no Isopropyl 3- 1.0 NA Chlorophenyi)methyla Aminopropylamino mnno Isopropyl 5- 31 NA Chlorophenyl)methyla Aminopentylainino "no0 Isopropyl 2-Amino-2-methyl- 0.05 0.1 Chlorophenyl)methyla ethylamino Isopropyl

NA

Chlorophenyl)methyla (Hydroxymethyl)pr ""DO ovviamino Isopropyl 1- 0.18 NA Chlorophenyflrnethyla (Hvdroxymnethyl)vr

R

1 R2 R 13 I (9g/niL) I (jig/mL) jenzymej cells rnino opylamino Isopropyl 0.26 NA Cblorophenyl)methyla (Hydroxymethyl)et mino hylaxnino Isopropyl I- 0.35 NA Chlorophenyl)methyla (Hydroxymethyl)et mino hvlam'no Isopropyl 0.38 NA Chlorophenyl)methyla Hydroxypropylami niino no Isopropyl 0.43 NA Chlorophenyl)methyla HydroxypropyLami inino no Isopropyl 2-Amnino- 0.48 NA Fluorophenyl)methyia propylarnino no.

Isopropyl 0.63 NA Fluorophenyl)methyla Tearahydrofuranyl) rnino methvlaniino Isopropyl 0.58 NA Fluorophenyl)methyla Tetrahydrofmanyl) rnino methvlamino Isopropyl 2-Hydroxy- 1 0.18 NA Fluorophenyl)methyla methylethylamino flfl Isopropyl (S)-2-Hydroxy-1- 0.22 NA Fluorophenyl)methyla methylethylarnino Isopropyl (R)-2-Hydroxy-2- 0.23 Fluorophe'ny1)methyla rnethylethylamino mino Isopropyl 1 0.11 2.4 Fluorophenyl)methyla Hydroxymethyipro mino pylaino The inhibition of cell proliferation properties of the compounds of this invention are demonstrated by their ability to inhibit cell proliferation in the range of about 0.05 pg/mi to 100 g.g/ml, preferably less than 0. 5 pLg/ml.

Similar assays were performed using the following cell lines; P388 mouse lymphoid neoplasm; L 12 10 mouse lymphcytic leukemia; Caco2 human colon adenocarcinoma; MCF7 human breast adenocarcinoina; *PupVSMC rat neonatal aortic smooth muscle cells; Ovcar human ovarian Carcinoma; Pancl human pancreatic adenocarcinomna; and HUVEC human umbilical cord endothelial cells. The inhibitory activity of several compositions of this invention against one or more of the cell lines are reported in Tables 8 and 9 below.

Table 8

IC,

0 (pg/ml) for Inhibition of Cell Proliferation R,X R, R, P L Caco MCF PancI OvC Pup HU 388 120 2 7 at VS VE MC C Isopropyl Diethanolamino 1.5 2.5 4.5 8.0 10.0 11.0 0.5 methoxyphenyl )-methylamino Isopropyl Diethanolamino 1.0 4.0 0.5 4.0 4.0 7.0 phenylphenyl)a mino Isopropyl Diethanolamino 1.0 1.0 3.5 1.0 1.3 phenylphenyl)methvlamino Isopropyl Morpholino >5 5.5 methoxyphenyl )-methylamino 3- isopropyl Diethanolamino 1.5 2.0 2.5 phenoxyphenyl '3benzyloxyphen Table 9 RX R, R, MRC-5 PupVSMC (4-methoxyphenyl)methylamino Isopropyl Diethanolamino 5 0.4 (4-Chlorophenyl)methylamino Isopropyl 2-Aminoethylamino 1 0.1 (4-Chlorophenyl)-methylamino Isopropyl 2-Amino-2- 1 .0.1 methylethylamino (4-Chlorophenyl)-methylamino Isopropyl 2-Amino-N1-(2- 1 0.3 hydroxyethyl)ethylamin 0 (4-Chlorophenyl)-methylamino Isopropyl Diethanolamino 3 0.3 Human Fibroblast PupVSMC Rat neonatal aortic smooth muscle cells EXAMPLE 8 Compounda of this invention was evaluated for effectiveness using the Murine Leukemia Model. The Murine Leukemia Model is a standard model used in the evaluation of antitumor agents. CDF1 mice were injected ip with L1210 cells (1xl0 cells/mouse).

Twenty-four hours later, these mice were treated with various doses (ip) of compound 3 of Example I in saline. The dosing regimen used in this study is outlined in Table 10, below.

Mice were dosed with compound 3 daily or on alternate days. Control mice received saline.

After 7 days, dosing was suspended and survival monitored.

Table Treatment N Median T/CxIOO survival time Days Saline control 7 10(9-13) 100 Compound 3 0.5 mg/kg bid 7 11(10-15) 110 mg/kg bid 7 13 (11-13) 130 2 mg/kg bid 7 12(10-14) 120 4 mg/kg days 7 13 (10-15) 130 1,3,5,7 8 mg/kg days 7 13(12-16) 130 1,3,5,7 The results indicate that rats administered compound 3 survived longer than the control rats.

68 EXAMPLE 9 This example measured the effect of an acute local delivery of compound 3 of Example 1 in reducing neointima formation following balloon angioplasty in the rat carotid artery model. In this example, the left common carotid arteries of adult male rats (n=10 per experimental group) were surgically injured using a Fogarty arterial embolectomy catheter.

Immediately after injury, the common carotid artery was bisected with a vascular clamp, thereby establishing an untreated and treated segment. A drug delivery catheter was then inserted into the distal half of the common carotid. After drug delivery, the catheter was removed and excess drug was washed out by removing the vascular clamp and re-establishing blood flow before closing the artery. The animals were allowed to recover for 14 days before harvesting the common carotid artery. The harvested tissue was sectioned and the neointimal area was digitized and measured with a computer planimetery system. For each animal, measurements were averaged for the untreated segment and 15 for the treated. aminoethyl)amino]-9-( methylethyl)purin-6-yl} [(4-chlorophenyl)methyl]amine was administered at a dose of 5 mg/mL reducing the neointimal area about 90% in comparison to the 6% reduction of saline alone.

The results of this Example are found in Figure 1. According to Figure 1, administering compound 3 of Example 1 to a damaged carotid artery reduced the neointimal area about 88% in comparison to the 6% reduction produced by the saline vehicle alone.

A/^

69 EXAMPLE IKB-ct Kinase Assays: Compositions of this invention were assayed to determine their IKB-a kinase inhibitory activity. The human umbilical vein endothelial cell line (HUVEC) used in these studies was purchased from Clonetics (San Diego, CA) and was maintained in endothelial cell growth medium supplemented with 2% fetal bovine serum, lOng/ml human recombinant epidermal growth factor, 1 upg/ml hydrocortisone, 50 .±g/ml gentamicin, 50 ng/ml amphotericin B and 12 .g/ml bovine brain extract at 37°C in a tissue culture incubator. All growth media and supplements were purchased from Clonetics (San Diego, CA). E. coli lipopolysaccharide (LPS) serotype 0111 :B4 was purchased from Sigma (Saint Louis, MI). All other chemicals were of reagent grade.

Preparation of cell Lvsate: Monolayers (75 cm 2 of HUVEC cells were treated with LPS (100 ng/ml) for 5 minutes after which the cell media was rapidly removed and the monolayer washed three times with ice cold PBS. The cell layer was scraped into 10 ml PBS and the cells pelleted by centrifugation (3000 rpm, 5 min, Cell lysate was prepared by incubating the cell pellet in 0.2 ml lysis buffer (20mM HEPES, pH7.3, 50mM NaCI, MgCl,, ImM EDTA, ImM EGTA, ImM sodium orthovanadate, 10mM p-glycerophospate, ImM phenylmethylsulfonylfuoride, ImM dithiothreitol, 0.5% Nonidet P-40 for 15 minutes at 37 0 C for frequent vortexing. Cell debris was removed from the sample by microcentrifugation (10,000xg, 15 minutes, 4°C) and the supernatant was "precleared" by the addition of 100 ml of a suspension of sepharose 4B in lysis buffer and mixing gently for 1 hour at 4°C. The speharose 4B beads were removed by microcentrifugation and the supernatant aliquotted and stored at Solid Phase IKB-a kinase assay: 1 4.g of GST- IicB-a, corresponding to full length IKB-a of human origin, (Santa Cruz Biotechnology,) was incubated with 20 4l of a 50% slurry of glutathione S sepharose 4B (Pharmacia) in reaction buffer (20mM HEPES, pH7.3, MgCl 2 15mM P-glycerophosphate, 0.5mM sodium orthovanadate, 0.5mM EGTA) for minutes at room temperature. The GST- IrB-bead complex was the washed three times with ml of reaction buffer by resuspension and microcentrifugation. 10g of HUVEC cell lysate protein in 100p.l of reaction buffer was then added to the GST- IKB-bead complex and the mixture incubated with gentle mixing at 4°C for 1 hour. The bead complex was then washed three times with reaction buffer containing'0.2 M NaCl and once with reaction buffer alone. Finally the bead complex was resuspended in 20±1 of reaction buffer containing 54Ci [y-"P]ATP (>5000 ci/mmol, New England Nuclear Corp. Boston, MA) and incubated at room temperature for 15 minutes. The reaction was terminated by the addition of 1 0(l of SDS- PAGE sample buffer and boiled for 3 minutes before separation by SDS-PAGE (10-20% gradient Readygel, BioRad). Following electrophoresis the gel was fixed (50% methanol 10% acetic acid) for 15 minutes, washed three times for 5 minutes each with distilled H,0 and treated with 5% glycerol for 15 minutes before drying down and exposing to film for autoradiography (X-OMAT XAR-5 Kodak).

In gel kinase assay: IKB-a isozymes were assayed for activity using a modification of previously published methods (11, 19, 20). Briefly duplicate samples of the IKcB-glutathione sepharose 4B bead complex were prepared as described above and were separated by electrophoresis through a 12% SDS-PAGE gel which had been polymerised in the presence of .g/ml GST- IKB-a. Following electrophoresis the gel was washed gently twice for minutes each with 50mM Tris-HCI pH8.0, 5mM B-mercaptoethanol; 20% isopropanol to remove SDS. Proteins were then denatured within the gel by incubation for 45 minutes in 100ml 50mM Tris-HCI pH8.0; 5mM P-mercaptoethanol; 0.04% Tween 40. The gel was then cut in half to separate the duplicate samples, one half was incubated in 10 ml reaction buffer alone and the other in 10 ml reaction buffer containing 10g/ml of 2-diethanolamino-6(4phenyl anilino)-9-isopropyl purine (compound 6 of Example 2) for 1 hour at room temperature which 10)Ci[y- 3 2 P]ATP was added and the incubations continued for a further hour at room temperature. The gels were then subjected to multiple 15 minute washes of 100ml each 5% trichloroacetic acid containing 1% sodium pyrophosphate until 1 ml of wash solution gave close to background radioactivity. The gels were then dried down and exposed to file for autoradiograhy.

Preparation of 2-diethanolamino-6-(4-phenvbenzvlamino)-9-isopropyv purine Epoxy activated Sepharose 6B Affinity Matrix. Freeze dried epoxy activated Sepharose 6B (Pharmacia LKB, Piscataway, NJ) was chosen for the coupling reaction due to its ability to form an ether bond between an hydroxyl-containing ligand and the epoxide group on the sepharose. The gel was swollen according to the manufacturer's instructions, (100mg) of compound 6 of Example 2 was dissolved in Iml coupling solution (1.2:1 v/v dimethylformamide 0.1N NaOH) and mixed with 0.5ml of swollen gel at pH 10-11 for 72 hours at room temperature with gentle agitation. Excess reactive groups were blocked with lM ethanolamine for 4 hours at and the gel slurry was poured into 1 ml syringe column. The resin was activated with three alternating cycles of twenty column volumes each of oH 4.0 (0.1M acetate, 0.5M NaC1) and pH 8.0 (0.1M Tris-HC1, 0.5M NaCi) buffers followed by twenty column volumes of reaction buffer (20mM HEPES, pH7.3, 10mM MgCl2, 15mM p-glycerophophate, 0.5mM sodium orthovanadate, 0.5mM EGTA). The column was stored at 4°C in reaction buffer containing 72 sodium azide and regenerated prior to each use with alternating cycles of low and high pH as described above.

Activated HUVEC cell lysate (500jg protein in Iml reaction buffer) was passed over the CVT-1545 sepharose matrix sequentially five times and the flow through was saved (unbound material). The matrix was then washed three times with lml of reaction buffer (wash 1-3) then three times each with reaction buffer containing 0.5M NaCI (eluate 1-3).

Aliquots (20tl from Iml) of each sample were assayed for their ability to phosphorylate at GST- IcB-sepharose bead complex and analyzed by SDS-PAGE as described above.

Assay of affinity enriched IKcB-(x kinase. The bulked 0.5 M NaCl eluates from the affinity matrix were used as the source of enzyme for development of an IicB-a kinase filter assay.

Each reaction contained affinity enriched IcB-a kinase (lpig protein), 10ng GST IcB-a kinase and 0.5pCi[y-" P]ATP (>5000 Ci/mmol, New England Nuclear Corp, Boston, MA) in reaction buffer. The reaction was incubated for 15 minutes at room temperature and was terminated by the addition of 24l 0.5M EDTA. Reaction mixtures were blotted onto phosphocellulose disks (Gibco BRL Life Technologies, Gaithersburg, MD) and the filters washed three times with 0.15M phosphoric acid with gentle shaking for 15 minutes (up to ten filters were washed with 300 ml of 0.15M phosphoric acid.) Following a third wash the filters were air dried, added to scintillation fluid and assayed by liquid scintillation spectrometry.

Electrophoretic Mobility Shift Assay: Nuclear extracts were prepared using a high-salt buffer extraction procedure. 10 pmol of double stranded NF-xB consenses oligonucleotide AGTTGAGGGGACTTTCCCAGGC-3') )Promega) was 5' end labeled with 5[iCi [y- 3 2

P]ATP

(>5000 Ci/mmol, New England Nuclear Corp, Boston, MA) by incubaton with T4 polynucleotide kinase for 1 hr at 37 0 C. Unincorporated nucleotides were removed by pasing the reaction mixture over I ml Sephadex G-5-spin column. Binding assays were performed at room temperature for 1 hr and consisted of 1 04g nuclear extract protein, 14~g salmon sperm DNA, and Sx 10' cpin of 2 P labeled consensus of oligonucleotide in the presence and absence of fifty fold unlabeled oligonucleotide. DNA-protein complexes were resolved by 8% non denaturing polyacrylamide gel electrophoresis, the gels were dried onto filter paper and visualized by autoradiography.

Table 11 Ac'ti'viv of Selectedl R nrespntntivpc nf thi TnVPi*nn En ~v m p R 1XR2 R3 IC5O(4.nM 4-phenylbenzylamino Isopropyl Diethanolamino 1.1 4-phenylbenzylaznino Isopropyl Diethylamino >2.4 4-phenylbenzylamino Isopropyl Ethanolamino 4-bromoanilino Isopropyl Diethanolanuino 14 4-(3-methoxphenyl) Isopropyl Diethanolamino benzylamino__________ 4-(4-methoxphenyl) Isopropyl Diethanolanuno 11 3-(4-nitrilophenyl) Isopropyl Diethanolanuno 2.2 anilino 4-thiomethoxyanilirio Isopropyl Diethanolamino 12.4 4-(2-pyridinyl) Isopropyl Diethanolaniino [benzylamnino

Claims (43)

1. A 2,6,9-trisubstituted purine composition of matter having the following formula: N N N N N R3R R 3 NR wherein R, is halogen or wherein X NH, 0, S, S(0 2 is alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl, and alkynyl, each having one to 20.carbon atoms, which alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl, and alkynyl, are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, CF, heteroaryl, heterocyclyl, R' 2 SR 20 S(O)R 21 S0 2 R 21 SO 2 N RRY S0 2 NR 0 C0R 2 1 ONR 20 CONR 0 R 23 SO-,NR 2 0 00 2 2 NR 0 23 NR 0 0 2 ,NR 20 COR, NR 20 CNR 0 N(R 20 )C(N 20 )NHR 3 NR 20 S0 2 R 1 OR 20 0C0NR 2 R 23 OCONR 0 SO 2R 21 0C0NR 0 R 2 3 CN, C0 2 R 20 CONR 20 R 2 1, CONR 20 SO,R 2 and C0R 20 R 2 is a hydrogen or hydrocarbon selected from the group ailkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and ailcynyl, each having one to 20 carbon atoms, which ailkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R" 2 SRZ 0 S(O)R 1 S0 2 R 2 1 SO,NR'OR", S 0 2 NR 20 00R 2 1 S ONR 20 C0NR 0 R 2 3 S0 2 NR 20 00 2 R, NR 2 0 R 23 NR 20 C0R 2 1 NR 20 GOR 21 NROCONRlIR 23 N(R 2 0 )C(NR 20 )NHR', NR 2 0 S 2 R 2 OR 20 0C0NR 2 R 23 000NR 20 S0 2 R 1 OCONR 2 0 R23, ON, CO,R 20 C0NR 20 R 2 3 CONRO 0 2 R 2 and C0R 0 R 3 is a halogen, hydroxyl, thia, alkoxy, alkylthio, alkyl, -NR, 4 R, or a component having the formula: R where m=1-3, o=1,3, y=carbonyl,'.NR, 4 hydroxyl, thiol, alkoxy, ailkyitbiol; R, and R, are each independently hydrogen, OR 2 NR 2 0 R. 3 or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R" 2 SR 20 S(O)R 21 S0 2 R 21 S0 2 NR 2 0 R 23 S0 2 NR 2 C0R 2 1 SO 2 NR 20 00NR" 2 S O.,NR 20 0 2 R 2 1 NR 2 31 NR 20 0R 21 NR 20 00 2 R 21 NR 20 00NR 2 R 23 N(R 20 )C(NR 20 )NHjR 2 3, NRa 2 0 SO,R 21 OR 20 OCONR 0 RY,' 0C0NR 2 S0 2 R 1 OCONR 2 D 2 ',CN C0 2 R 2 0 C0NR 0 R 2 C0NR 0 S0 2 R 2 and C0R 20 with the proviso that when Y is carbonyl, Y and together may be a single oxygen atom, and R 5 together may be a single oxygen atom, and may together be a single oxygen atom, and wherein when R 3 is 2- hydroxyethylamino and R, is methyl, is not amino, 3-methyl-2-butenylamino, benzylamino, or m-hydroxybenzylamino, when R 3 is not 2-hydroxyethylamino, when is isopropyl, R 1 is not benzylamino, m-hydroxybenzylamino, or 3-methylbutylamino, when R 3 is 2-hydroxyethylamino and is 2-hydroxyethyl, R, X is not benzylamino and when R 3 is selected from the group consisting of 2-methyl-2-hydroxypropylamino, and 2- dimethyl aminoethyl amino, and when R, is methyl, then R 1 is not benzylamino; R" 0 is a member selected from the group consisting of H, C,, 1 alkyl, C 21 5 alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with I to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl or heteroaryl anmide, CN, O-CI- 6 alkyl, CF 3 is aryl, and heteroaryl; R" 1 is a member selected from the group consisting of alkyl, C 2 1 alkenyl, 1, alkynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkcynyl, aryl, heterocyclyl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from the group of halo, heterocyclyl, aryl, heteroaryl, CF 3 CN, OR 20 SR 20 N(R 20 2 S(O)R2I, S0 2 R1 2 SO 2 N(P. 20 2 SO,NR 20 1C0R 2 2 SON 2 O00 2 R 22 SO 2 NR 20 1CON(R 0 2 N(R 20 2 NR 2 0 COR 1 2 NR' 0 C0,R 2 2 NR 20 CON(R 20 2 NR 20 C(NR 2 0 )NHR 23 COP. 20 C0 2 R 20 CON(R 20 2 C0NR 2 0 S0 2 R 2 NR 20 SOR 2 2 SO.2NR 20 CO2R22, OR 20 0C0N 2 0 S0 2 R 22 OC(O)R? 0 C(O)OCH,OC(O)R 20 and OCON(R 0 2 and each optional heteroaryl, aryl, and heterocyclyl substituent is optionally substituted with halo, ailkyl, CF 3 amino, mono- or di- alkylamino, alkyl or aryl or heteroaryl amnide, NCOR. 2 NR 20 S0 2 R 2 C0R 0 C0 2 R' 0 CON(R) 2 NR' 0 CON(R 20 2 OC(O)R 0 OC(O)N(R 20 2 SR 20 S(O)R2, SO,R 22 SO 2 N(R 0 CN, or OR 20 R 22 is a member selected from the group consisting of ailkyl, C2- 5 alkenyl, C 2 1 alkynyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, alkynyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl or heteroaryl an-lde, CN, alkyl, CF 3 aryl, and heteroaryl; and R 23 is R 2 or H.

2. A 2,6,9-trisubstituted purine composition of claim I wherein: is a alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkcynyl, each having one to 20 carbon atoms, which alkcyl, heterocyclyl, aryl, heteroaryl, arailcyl, heteroarylalkyl, alkenyl,and alkynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, CF 3 aryl, heteroaryl, heterocyclyl, R 2 1, SR 20 S(O)R 21 SoR 2 SO.,NR 20 R23, NR 20 R 2 3 NR 20 C0R 21 NR 20 COR 21 NR 2 0CONR&R2n, NR 20 SO,R 2 OR 20 CN, C0 2 R 20 C0NR" 2 and C0R 0 R, is a hydrogen or hydrocarbon selected from the group substituted alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and ailkynyl, each having one to 20 carbon atoms, which alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalcyl, alkenyl,and alkynyl,are optionally substituted with from I to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R" 2 SR 20 S(O)R 21 S0 2 R 2 1 SO 2 NR 2 &R 23 NR 2 &R 23 NR 20 C0R 21 NR 20 C0 2 R 21 NR 20 CONlR 2 R3 NR-0R R 0 N C0 2 R 0 C0N RR, and C0R 0 78 R. and R, are each independently hydrogen, OR 20 NR 20 R 2 3 or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroar-ylalkyl, atkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and allcynyl,are optionally substituted with from I to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R 2 2 SR 2 1, S(O)R 21 SO,R 21 S0 2 NR OR 2 3 NRI'R 2 1 NR 20 00R 2 Nk 2 0 CO 2 R 21 NR 20 C0NO 2 1, NR 20 SO, 21 OR 0 ,OOR 20 CONR 20 R23, and CopR 20 R 20 is a member selected from the group consisting of H, 0 1 .,allcyl, alkenyl, 02- Sheterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with I to 3 substituents independently selected from halo, ailkyl, mono- or dialkylamino, alkyl or aryl or heteroaryl amide, ON, alkyl, OF 3 aryl, and heteroaryl; R 2 is a member selected from the group consisting of ailcyl, C2- alkenyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, aryl, heterocyclyl, and heteroaryl are' 1s optionally substituted with 1 to 3 substituents independently selected from the group of halo, heterocyclyl, aryl, lieteroaryl, OF 3 ON, OR 20 SR 20 N(R 2 0 2 S(O)R 22 SOR 22 SO 2 NR 0 S0 2 N 2 0 0R 22 SO'NR 0 OO 2 22 SO.,NR 20 00( 20 R) 2 NR 20 C0R 2 2 ,NR00R, NR. 20 0N(R 20 NR 20 C(NR2 0 )NI~e 3 OR 20 C0 2 R 20 OON(R 20 2 00NR 20 S0 2 R 2 NR 20 SORR2, SONR 20 C0 2 RW 2 OR 2 1, 000NR 20 S0 2 R 22 OO(O)R 20 O(O)OOH,OO(O)R 20 and OOON(R 20 2 and each optional heteroaryl, aryl, and heterocyclyl -substituent is optionally substituted with halo, alkyl, OF 3 amino, mono- or di- alkylamino, alkyl or aryl or heteroaryl amide, N00R 22 NR 20 SO 2 R 2 2 OR 20 OO,R 20 CON(R 2 2 NR 20 00N( 2 0 2 OO(O)R 20 OO(O)N(R 0 2 SR2', S(O)R 2 2 SO2R 2 SO 2 N(R 2 0 2 ON, or OR 20 and R' 2 is a member selected from the group consisting of alkyl, C 2 8 allcenyl, heterocyclyl, aryl, and heteroaryl, which alkyl, alkenyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or diallcylamino, alkyl or aryl or heteroaryl amide, ON, 0-C 1-6 alkyl, OF 3 aryl, and heteroaryl.

3. A 2,6,9-trisubstituted purine composition of claim 1 wherein: R11 is alkcyl, heterocyclyl, aryl, heteroaryl, arailcyl, heteroarylalkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, OF 3 aryl, heteroaryl, heterocyclyl, R- 2 SR 20 S(0)R2', S0 2 S O,NR 0 R 2 NR 20 R 23 NR 20 00R 2 1 NR 20 C 2 R 21 NR 20 SO,R 21 OR 20 CN, C0 2 R 20 C0NR 20 R 2 and C0R 0 R, is a hydrogen or hydrocarbon selected from the group including alkyl, heterocyclyl, and aryl, each having one to 10 carbon atoms, which alkyl, heterocyclyl, aryl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R" 2 SR 20 S(O)R 2 SO 2 R 21 S0 2 NR 20 R 3 NR 2 0R 23 NR 20 C0R 21 NR 2 uC0,R 1 NR 20 SO,R 2 OR 20 CN, O,R 2 CONR 2 and 00R 0 R 4 and R 5 are each independently hydrogen, OR 2 0 NR 2 ,R 23 or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, arailkyl, alkenyi,and alkynyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R' 2 SR 20 S(O)R 21 SO,R 21 SO 2 NR 20 R 2 1, WROW' 2 NR 20 C0R 2 NR 0 C0 2 R 2 1 NR. 2 SOR 2 OR 20 ON, C0 2 R 20 C0NR 2 0 R 23 and C0R 0 R 0 is a member selected from the group consisting of H, C,- 8 alkyl, aryl, and heteroaryl, which ailkyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, ailkyl. or aryl or heteroaryl. amide, ON, 0-01. alkyl, OF 3 R 2 1 is a member selected from the group consisting of 01.8 alkyl, aryl, and heteroaryl, which alkyl, aryl, and heteroaryl are optionally substituted with 1 to 2 substituents independently selected from the group of halo, OF, ON, OR" 0 SR. 2 N(R 20 S(0)R 2 2 SO R 22 SON(R 20 2 NR 20 CO.R 22 NR 20 00N(R 20 C0R 20 CO,R' 0 CON(R' 0 NR 20 SO2R 22 OR 20 and R 2 1 is a member selected from the group consisting of alkyl, aryl, and heteroaryl, which alkyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamnino, alkyl or aryl or heteroaryl amide, ON, 0-C I* alkyl, OF 3 aryl, and heteroaryl.

4. A 2,6,9-tri substituted purine composition of claim I wherein: is a alkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which ailkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl, alkenyl,and allcynyl,are optionally with from I to 2 substituents independently selected from the group consisting of halo, OF3,, aryl, S(O)R2', SO2R SONR' 0 R 23 NR 2 0 R 23 NR 20 00R 1 N'R' 0 C0 2 R NR' 0 S02R 1 OR" 0 ON, CO,R2 0 and R2 is a hydrogen or hydrocarbon selected from the group alkyl, heterocyciyl, and aryl, each having one to 10 carbon atoms which alkyl, heterocyclyl, aryl, are optionally substituted with from I to 2 substituents independently selected from the group consisting of halo, aryl, heteroaryl, heterocyclyl, R' 2 SR" 0 S(O)R 1 SO2R 1 S0 2 NR 20 R 2 3 NR 2 &R 23 NR' 0 C0R 1 NR 2 0 CO 2 R 21 NR 20 S0 2 R 21 OR" 0 CN, C0 2 R 0 C0NR 0 R 2 1, and C0R 0 R, and P- 5 are each independently hydrogen, or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkryl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, alkenyl,and allcynyl,are optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, aryl, R 22 SR 2 S(O)R 1 S0 2 R 1 S0 2 NR OR 3 NR OR 23 NR 20 C0R 21 NR 20 C0 2 R 21 NR 20 S0 1 OR 2 0 CN, C0 2 R 20 and 00NR 20 R 3 R 2 0 is a member selected from the group consisting of H, C 1 .,alkyl, aryl, and heteroaryl, which alkyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents; independently selected from halo, alkyl, mono- or dialkylamino, ailkyl or aryl or heteroaryl an-ide, CN, O-C,- 6 alkyl, OF 3 R 21 is a member selected from the group consisting of 01-, alkyl, aryl, and heteroaryl, which alkyl, aryl, and h~eteroaryl are optionally substituted with 1 to 2 substituents independently selected from the group of halo, CF,, ON, OR 20 SR 20 N(R 20 2 S(O)R 2 SO,R 2 SO 3 N(R 20 2 NR 20 COR 22 NRI 0 CON(R 2 0 COR 20 CO2R 2 1, CON(R' 0 2 NR 20 SO,R 1, OR 20 and R 22 is a member selected from the group consisting of 01.8 alkyl, aryl, and heteroaryl, which alkryl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl or heteroaryl amide, ON, 0-C, alkyl, CF 3 aryl, and heteroaryl. The 2,6,9-trisubstituted punine composition of claim 1 wherein X=NH.

6. The 2,6,9-trisubstituted purine composition of claim 1 wherein R 3 is a component having the formula: where m=1-3, n=1-3, o=1,3, y=carbonyl, -NRRs, hydroxyl, thiol, alkoxy, ailkyithiol with the provisos that when Y is carbonyl, Y and together may be a single oxygen atom, and may together be a single oxygen atom, and may together be a single oxygen atom; and R, and R, are each independently hydrogen, or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroa-yl, aralkyl, alkenyl,and alkynyl,are optionally substituted with from I to 2 substituients independently selected from the group consisting of halo, aryl, R" 2 SR 20 S(O)R 1 S0 2 R 2 1 S0 2 NR 20 R 2 1, NR 20 R 2 1, NR 20 C0R 2 2 NR 20 C0 2 R 21 NR' 0 SO,R 21 OR 20 CN, CO,R 20 and C0NR 20 R 1 3

7. The 2,6,9-trisubstituted purine composition of claim 3 wherein is selected from the group consisting of aralkyl and heteroarylalcyl.

8. The 2,6,9-trisubstituted purine composition of claim 7 wherein is selected. from the group consisting of aralkyl, unsubstituted pyridylalkyl and substituted pyridylalkyl and whereiih R. is selected from the group consisting of lower alkyl, substituted lower ailkyl, and alkyl cycloalkyl.

9. A 2,6,9-trisubstituted purine composition of claim 5 wherein: is an aryl, heteroaryl, heterocyclyl, aralkyl, heteroarylalkyl, each having one to carbon atoms, which aryl, heteroaryl, heterocyclyl, aralkyl, heteroarylalkyl, are optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, CF 3 aryl, R 2 SR 2 0 S(O)R 21 SO,R 21 SONR2 R 2 3 NR2°R 23 NR 2 0 COR 21 NR 20 COR 21 NR 20 SOR 2 1 OR 20 CN, COR 2 0 and CONRR R 2 is a hydrogen or hydrocarbon selected from the group substituted lower alkyl, substituted alkyl, cycloalkyl, substituted cycloalkyl each having one to 10 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, R 2 2 SR 2 0 S(O)R" 2 S0 2 R 21 NR 20 R 23 OR 2 0 and CN; R 4 and R 5 are each independently hydrogen, or a hydrocarbon selected from the group including alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, alkenyl,and alkynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, alkenyl,and alkynyl,are optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, aryl, R 22 SR 20 NR 20 R 23 NR 20 COR 21 NR 20 COR 21 NR 2 0 SOR 2 1, OR 2 0 CN, CO,R 20 and CONR 2 R"; R 2 0 is a member selected from the group consisting of H, C,. 8 alkyl, which alkyl is optionally substituted with 1 to 2 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or CN, alkyl, CF,; R 2 1 is a member selected from the group consisting of C,. 8 alkyl, which alkyl is optionally substituted with 1 to 2 substituents independently selected from the group of halo, CF 3 CN, OR 2 0 SR 20 N(R 2 0 2 and 84 R 22 is a member selected from the group consisting of C. 3 ,alkyl, aryl, heteroaryl which alkyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl, CN, O-C, alkyl, CF 3

10. The 2,6,9-trisubstituted purine composition of claim 3 wherein is selected from the group consisting of aryl, heterocyclyl, heteroaryl, substituted heteroaryl, and substituted aryl.

11. The 2,6,9-trisubstituted purine composition of claim 3 wherein is selected from the group consisting of aryl, unsubstituted pyridyl, substituted pyridyl, and substituted aryl, and R, is selected from the group consisting of alkyl, substituted alkyl.

12. The 2,6,9-trisubstituted purine composition of claim 2 wherein R, is -NR, 4 R wherein R, and R 5 are each selected from the group consisting of hydrogen, alkyl, heterocyclyl, acyl, aryl, heteroaryl, aralkyl, heteroaralkyl, alkyl alkenyl, alkyl alkynyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroarylalkyl,are optionally substituted with from 1 to 3 substituents independently selected from the group consisting of halo, aryl, heteroaryl,heterocyclyl, SR 2 0 S(O)R 2 1 SO,R 21 SO 2 NR 2 0 NR 2 R 2 3 NR 20 COR 21 NR 2 0 CO 2 R 21 NR 2 0 CONR 2 R"3, NR 20 SO,R 2 OR 20 CN, CO2R 2 0 CONR 2 °R 2 3 and COR 2 o.

13. A 2,6,9-trisubstituted purine composition of claim 12 wherein: is an aryl, substituted aryl, each having 6 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, CF,, aryl, R 2 2 NR 2 R 2 3 NR 2 0 COR 21 OR 20 CN; R 2 is a hydrogen or hydrocarbon selected from the group substituted lower alkyl, cycloalkyl, substituted cycloalkyl each having one to 6 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, R 2 2 NR 2 R 2 3 OR 2 0 R 4 and R 5 are each independently hydrogen, or a hydrocarbon selected from the group including alkyl, and heterocyclyl wherein each hydrocarbon has from 1 to 12 carbon atoms, which alkyl, and heterocyclyl are optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, SR 2 0 OR 2 0 ,NR 2 0 CN, COR 20 and CONR2 0 R 2 3 R 20 is a member selected from the group consisting of H, C,.salkyl; R 21 is a member selected from the group consisting of C,.3 alkyl, which alkyl is optionally substituted with 1 to 2 substituents independently selected from the group of halo, CF 3 CN, OR 2 0 SR 2 0 N(R 2 0 and R" is a member selected from the group consisting of C 1 3 alkyl, aryl, heteroaryl which alkyl, aryl, and heteroaryl are optionally substituted with 1 to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl, CN, O-C,.6 alkyl, CF 3

14. A 2,6,9-trisubstituted purine composition of claim 12 wherein: is an aryl, substituted aryl, each having 6 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, CF 3 OR 2 0 CN; R, is isopropyl; R 4 and R, are each independently hydrogen, or a hydrocarbon selected from the group including alkyl, and heterocyclyl wherein each hydrocarbon has from 1 to 12 carbon atoms, 86 which alkyl, and heterocyclyl are optionally substituted with from I substituent independently selected from the group consisting of R 2 2 OR 2 0 NR 20 R 2 3 R 2 0 is a member selected from the group consisting of H, C. 2 ,alkyl; R 2 is a member selected from the group consisting of C,.3 alkyl; R" is a member selected from the group consisting of C. 3 alkyl, aryl, which alkyl, aryl, are optionally substituted with 1 substituent independently selected from halo, alkyl, mono- or dialkylamino, CN, CF 3 and R 23 is R 21 or H. A 2,6,9-trisubstituted purine composition of claim 12 wherein: is an aralkyl, substituted aralkyl, each having 6-8 carbon atoms wherein substitution includes optional substitution with from 1 to 2 substituents independently selected from the group consisting of halo, CF 3 aryl, R 2 2 NR 2 R 23 NR 20 COR 2 OR 2 0 CN; R, is a hydrogen or hydrocarbon selected from the group substituted alkyl, cycloalkyl, substituted cycloalkyl each having one to 6 carbon atoms wherein substitution includes optional substitution with from I substituent independently selected from the group consisting of halo, R 22 NR 2 0 OR 2 0 and R s are each independently hydrogen, or a hydrocarbon selected from the group including alkyl and heterocyclyl wherein each hydrocarbon has from 1 to 12 carbon atoms, which alkyl and heterocyclyl are optionally substituted with from I to 2 substituents independently selected from the group consisting of halo, R 22 SR 2 0 OR 2 0 ,NR 2 oR 2 CN, COR 2 0 and CONR 2 R'; R 2 0 is a member selected from the group consisting of H, C,.,alkyl; R 2 is a member selected from the group consisting of C,.3 alkyl, which alkyl is optionally substituted with 1 to 2 substituents independently selected from the group of halo, CF,, CN, OR 2 0 SR 2 0 N(R 2 0 and R 2 2 is a member selected from the group consisting of C 1 3 alkyl, aryl, heteroaryl which alkyl, aryl, and heteroaryl are optionally substituted with I to 3 substituents independently selected from halo, alkyl, mono- or dialkylamino, alkyl or aryl, CN, O-Ci., alkyl, CF,.

16. A 2,6,9-trisubstituted purine composition of claim 12 wherein: is -CH,-phenyl wherein the phenyl ring is optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, CF 3 R 2 OR 2 0 CN; R, is isopropyl; R, and R 5 are each independently hydrogen, or a hydrocarbon selected from the group including alkyl, and heterocyclyl wherein each hydrocarbon has from 1 to 12 carbon atoms, which alkyl, and heterocyclyl are optionally substituted with from 1 to 2 substituents independently selected from the group consisting of R 2 2 OR 2 0 NR 2 0Rn; R 20 is a member selected from the group consisting of H, C,. 2 alkyl; R 2 is a member selected from the group consisting of C,.3 alkyl; R 22 is a member selected from the group consisting of C,.3alkyl, aryl, which alkyl, aryl, are optionally substituted with 1 substituent independently selected from halo, alkcyl, mono- or dialkylamino, CN, CF3; and R" is R 2 1 or H.

17. The 2,6,9-trisubstituted purine composition of claim 12 wherein is selected from the group consisting of aralkyl, substituted pyridylalkyl, and unsubstituted pyridylalkyl; R 2 is selected from the group consisting of alkyl, which alkyl is optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, R", NR 2 R 2 3 OR 20 R, is a substituted alkyl having from 2 to 6 carbon atoms optionally substituted with from 1 to 3 substituents independently selected from the group consisting of R 2 2 OR 2 0 NR2R 2 3 R, is selected from the group consisting of hydrogen, alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl,are optionally substituted with from I to 2 substituents independently selected from the group consisting of halo, aryl, R 2 2 SR 2 0 NR 2 R NR 2 COR, NR 20 COR 2 1 NR 20 SO 2 R 2 1 OR 20 CN, C0 2 R 20 and CONR 2 R 2 3 R 2 is a member selected from the group consisting of H, C .2 alkyl; R 21 is a member selected from the group consisting of C,.3 alkyl; R 22 is a member selected from the group consisting of C,. 3 alkyl, aryl, which alkyl, aryl, are optionally substituted with 1 substituent independently selected from halo, alkyl, mono- or dialkylamino, CN, CF 3 and R 3 is R 2 1 or H.

18. The 2,6,9-trisubstituted purine composition of claim 12 wherein is selected from the group consisting of aryl, substituted aryl, pyridyl, and substituted pyridyl; R, is selected from the group consisting of alkyl, which alkyl is optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, R 22 NRR" OR 2 0 89 R 4 is a substituted alkyl having from 2 to 6 carbon atoms optionally substituted with from 1 to 3 substituents independently selected from the group consisting of OR 2 0 NR 2 R 23 R 5 is selected from the group consisting of hydrogen, alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl, each having one to 20 carbon atoms, which alkyl, acyl, heterocyclyl, aryl, heteroaryl, aralkyl,are optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, aryl, R 2 2 SR 2 0 NR 2 23 NR 20 COR 21 NR 20 COzR 2 NR 20 SOR 2 1 OR 2 0 CN, CO,R 0 and CONR"R 2 3 R 2 is a member selected from the group consisting of H, C,. 2 alkyl; R 21 is a member selected from the group consisting of C,.3 alkyl; R 22 is a member selected from the group consisting of C1.3 alkyl, aryl, which alkyl, aryl, are optionally substituted with 1 substituent independently selected from halo, alkyl, mono- or dialkylamino, CN, CF,; and R2 is R 2 or H.

19. The 2,6,9-trisubstituted purine composition of claim 12 wherein is selected from the group consisting of aralkyl, pyridylalkyl, and substituted pyridylalkyl; R, is selected from the group consisting of alkyl, which alkyl is optionally substituted with from 1 to 2 substituents independently selected from the group consisting of halo, R 2 2 and OR 20 R 4 and R, are each a substituted alkyl having from 2 to 6 carbon atoms optionally substituted with from 1 substituent independently selected from the group consisting of R 22 NR 2 1R 3 and OR 2 0 R 2 0 is a member selected from the group consisting of H, C,. 2 alkyl; R 2 is a member selected from the group consisting of Ci. 3 alkyl; R 2 2 is a member selected from the group consisting of C 1 3 alkyl; and R" is R 21 or H. The 2,6,9-trisubstituted purine composition of claim 12 wherein Ri' is CH, aryl or CH, substituted aryl, R 2 is lower alkyl or substituted lower alkyl, and R, and R, are each -CH 2 CHOH -CHR'CHOH, or -CH 2 CHR'OH wherein R' is hydrogen or alkyl having from 1 to 6 carbon atoms.

21. The 2,6,9-trisubstituted purine composition of claim 12 wherein is CH 2 Aryl or CH,-substituted aryl, R, is lower alkyl, and R 4 H, and R, is -CHCH.NH 2 CHR'CHNH 2 -CHCHR'NH, wherein R' is hydrogen or alkyl having from 1 to 6 carbon atoms.

22. The 2,6,9-trisubstituted purine composition of claim 21 wherein R 2 is isopropyl.

23. The 2,6,9-trisubstituted purine composition of claim 12 wherein is CH 2 Aryl or CH,-substituted aryl, R, is lower alkyl, and R 4 -CH 2 CH 2 OH R 5 is CHCH 2 NH 2 or CHR'CHNH 2 or CH 2 CHR'NH 2 wherein R' is hydrogen or alkyl having from 1 to 6 carbon atoms.

24. The 2,6,9-trisubstituted purine composition of claim 23 wherein R, is isopropyl.

25. The 2,6,9-trisubstituted purine composition of claim 20 wherein R 2 is isopropyl.

26. The 2,6,9-trisubstituted purine composition of claim 12 wherein is selected from the group consisting of aryl, substituted aryl, pyridyl, and substituted pyridyl, R 2 is 91 selected from the group consisting of lower alkyl, substituted lower alkyl, and alkyl cycloalkyl, and R 4 and R are each a substituted lower alkyl having from 2 to 6 carbon atoms.

27. The 2,6,9-trisubstituted purine composition of claim 12 wherein is aryl or substituted aryl, R 2 is lower alkyl, or substituted lower alkyl, and R 4 and R s are each CHCHOH, -CHR'CH 2 OH, or -CHCHR'OH wherein R' is hydrogen or alkyl having from I to 6 carbon atoms.

28. The 2,6,9-trisubstituted purine composition of claim 27 wherein R 2 is isopropyl.

29. The 2,6,9-trisubstituted purine composition of claim 12 wherein is benzyl substituted with a halogen, alkoxy, phenyl, pyridyl or nitro group, R, is isopropyl, and R, and R are each -CHCH 2 OH. The 2,6,9-trisubstuted purine composition of claim 12 wherein is benzyl substituted with a halogen, alkoxy, phenyl, pyridyl or nitro group, R, is isopropyl, R, H, and R s CH 2 CHNH,.

31. The 2,6,9-trisubstuted purine composition of claim 12 wherein is benzyl substituted with a halogen, alkoxy, alkyl, CF 3 phenyl, pyridyl or nitro group, R 2 is isopropyl, R, H, and R 5 CHCHR'NH, wherein R' is hydrogen or alkyl having from 1 to 6 carbon atoms.

32. The 2,6,9-trisubstuted purine composition of claim 12 wherein is benzyl substituted with a halogen, alkoxy, C,. 3 alkyl, CF, phenyl, pyridyl or nitro group, R, is isopropyl, R, H, and Rs CHCR'R'NH2 wherein R' is hydrogen or alkyl having from 1 to 6 carbon atoms. 92

33. The 2,6,9-trisubstuted purine composition of claim 1 selected from the group consisting of 2- {(2-hydroxyethyl)[9-(methylethyl)-6-( (trifluoromethyl)phenyl]niethyl} amino)purin-2 -yl] amino)} ethan- 1 -ol, {((2S)oxolan-2- yl)methyl](6- [(4-fluorophenyl)methyllaxnino -9-(methylethyl)purin-2-yl)amine, [((2R)oxolan-2-yI)methyl](6- fluoropheniyl)methyl] amino -9-(methylethyl)purin-2- yl)amnine, (2-amninoethyl)(6- f [3,5-dichlorophenyl)methyllamino -9-(methylethyl)purin-2- yl)amine, (2-amninoethyl)[6-( {[4-chl'oro-3-(trifluoromethyl)phenyl]methyl} amino)-9- (methylethyl)purin-2-yl] amine, f [(4-chlorophenyl)methyl] amino)} -9 -(methylethyl)purin- 2-yl)amino]-3-methylbutanamide, (2-amino-2-methylpropyl)(6- chlorophenyl)methyl] amino I -9-(methylethyl)purin-2-yl)amine, .3 -(2-[bis(2- hydroxyethyl) amino]j-6- {[4-chlorophenyl)methyl]anino~purin-9-yl)butan-2-one, [(4-chlorophenyl)methyll amino)} -9-(methylethyl)pu-in-2-yl)amino]-3-methylbutan- I1- ol, {2-[(2-aminoethyl)amino]-9-(methylethyl)purin-6- yI} amino)methyl]benzenesulfonamide, 2-[(2-hydroxyethyl)(6-{([(4- methoxyphenyl)methylljamino)} -9-(methylethyl)purin-2-yl)amino]ethan- 1 -ol, 2(2 hydroxyethyl) 19-( methylethyl)-6-[(4-phenylphenyl)amino]purin-2-yl} amnino)ethan- I -ol, {2-[(2-amino-2-propyl)amino]-9-(methylethyl)purin-6-ylI [(4-hlorophenyl)methyl] amine, {2-[(2-aminoethyl)amino] methylethyl)purin-6-yl }[(4-chlorophenyl)methyl]wmine, {2-[(2-aminopropyl)amino]-9-( methylethyl)purin-6-yl} [(4-chlorophenyl)inethyl~amr-ine and -aminoethyl)(6- [(4-chlorophenyl)methyl] am-ino }-9-(methylethyl)purin-2- yl)axnino]ethan-lI-ol.

34. The 2,6,9-trisubstituted purine composition of claim 12 wherein is phenyl substituted with a halogen, alkoxy, phenyl, pryidyl or itro group, R, is isopropyl, and R, and are each -CH 2 CH.,OH. The 2,6,9-trisubstituted purine composition of claim 12 wherein Rl' is biphenylmethyl, R, is isopropyl, and and R, are each -CH 2 CH.,OH.

36. The 2,6,9-tisubstituted purine composition of claim 12 wherein is selected from the group consisting of3-methylthiophenyl, 4-methyithiophenyl, 4-phenylbenzyl, 4- methoxybenzyl, 4-biphenyl, 3-methoxybenzyl, 4-(2-thienyl)benzyl, -4 methyl)phenylbenzyl, 4-(4-trifluoromethyl)phenylbenzyl, 4-(4-nitrilo)phenylbenzyl, 4-(2- pyridinyl)benzyl, piperonyl, 3-methoxbenzyl, 4-chlorobenzyl, and 4-nitrobenzyl, R 2 is isopropyl, and and R, are both GH,CH.,OH.

37. The 2,6,9-trisubstit-uted purine composition of claim 36 wherein is selected from the group of compounds consisting of 4-methoxybenzyl, 4-phenylbenzyl, 4- methoxybenzyl, 4-biphenyl, 3-methoxybenzyl, 4-(2-thienyl)benzyl, -4 methyl)phenylbenzyl, 4-(4-trifluoromethyl)phenylbenzyl, 4-(4-nitrilo)phenylbenzyl, 4-(2- pyridinyl)benzyl, piperonyl, 3-thiomethoxphenyl, 4-thiomethoxyphenyl and 4-bromophenyl.

38. A cationic salt of the composition of claim 1.

39. An acid addition salt of the composition of claim 1. A method for inhibiting cell proliferation in mammals comprising administering a therapeutically effective amount of the composition of claim 1 to the mammal.

41. The method of claim 40 wherein the therapeutically effective amount ranges from about 0.00 1 to about 100 mg/kg weight of the mammal.

42. The method of claim 40 wherein the composition is administered to a mammal suffering from a cell proliferation disorder selected from the group consisting of rheumatoid 94 arthritis, lupus, type I diabetes, multiple sclerosis, cancer, restenosis following ballon angioplasty or atherectomy, restenosis following vascular modifying surgical procedures, host graft disease, and gout.

43. The method of claim 42 wherein the cell proliferation disorder is restenosis.

44. The method of claim 42 wherein the cell proliferation is disorder cancer. The method of claim 42 wherein the cell proliferation disorder is polycystic kidney disease.

46. The method of claim 42 wherein the mammal is a human.

47. A pharmaceutical composition of matter comprising the composition of claim 1 and one or more pharmaceutical excipients.

48. An antifungal agent useful for treating fungal infections in humans, and animals comprising the composition of claim 1.

49. The method of claim 42 wherein the cell proliferation disorder is selected from the group consisting of lymphoyd neoplasm, cancer of the colon, breast cancer, ovarian cancer, pancreatic cancer, and cancers derived from endothelial cells. A 2,6,9-trisubstituted purine composition substantially as hereinbefore described with reference to the examples.

51. A method for inhibiting cell proliferation in mammals substantially as hereinbefore described with reference to the examples. 16 January 2004 By Freehills Carter Smith Beadle Registered Patent Attorneys for the Applicant CV Therapeutics, Inc.