USOO8962619B2

(12) United States Patent (10) Patent No.: US 8,962,619 B2 Ashwell et al. (45) Date of Patent: *Feb. 24, 2015

(54) SUBSTITUTED (58) Field of Classification Search MIDAZOPYRIDINYL-AMNOPYRIDINE None COMPOUNDS See application file for complete search history. (71) Applicant: ArCule, Inc., Woburn, MA (US) (56) References Cited U.S. PATENT DOCUMENTS (72) Inventors: Mark A. Ashwell, Carlisle, MA (US); Chris Brassard, Somerville, MA (US); 4,522,811 A 6/1985 Eppstein et al. Anton Filikov, Rockville, MD (US); 33. R: ck 3. RileyShWell f et alal...... 514,303 ason Hill. Auburndale, MA (US); Steffi 8,609.688 B2 * 12/2013 Ashwell et al...... 514,303 Koerner, Medford, MA (US); 2002/0151549 A1 10/2002 Hayakawa et al. Jean-Marc Lapierre, Pelham, NH (US); 2006/0035921 A1 2/2006 Castelhano et al. Yanbin Liu, Acton, MA (US); Nivedita 2007. O142369 A1 6/2007 van Heek et al. 2007,0254.868 A1 11/2007 Lauffer et al. Namdev, Westford, MA (US); Robert 2009/0253734 A1 10/2009 Kelly, III et al. Nicewonger, Tyngsboro, MA (US); 2011/0059936 A1 3/2011 Lauffer et al. Rocio Palma, North Andover, MA (US); 2011/0172203 A1 7, 2011 Ashwell et al. Manish Tandon, Framingham, MA 2012/0329793 A1* 12/2012 Ashwell et al...... 514,234.2 (US); David Vensel, Brookline, MA (US); Akihisa Matsuda, Tokyo (JP); FOREIGN PATENT DOCUMENTS Shin Iimura, Tokyo (JP); Kenichi CN 1824665 A 8, 2006 Yoshida, Tokyo (JP); Takanori DE 3722.992 A1 1, 1989 Yamazaki, Tokyo (JP); Takahiro P 12: A. 3. E. Kitamura, Tokyo (JP); Takeshi JP 200034.4780 A 12/2000 Isoyama, Tokyo (JP) KR 20090007347 U 7, 2009 WO WO-96.19991 A1 T 1996 (73) Assignee: ArCule, Inc., Woburn, MA (US) WO WO-99.57 103 A1 11, 1999 WO WO-03.08.0125 A2 10/2003 (*) Notice: Subject to any disclaimer, the term of this WO WO-03080610 A1 10, 2003 WO WO-03O88967 A1 10, 2003 patent is extended or adjusted under 35 WO WO-20050117OO A1 2/2005 U.S.C. 154(b) by 0 days. WO WO-2007056155 A1 5, 2007 Thiis patenttent iis subjectbiect tot a terminalt inal didis WOwo WO-2007wo.-2008.007900 111904 A2A1 10/20071/2008 claimer. WO WO-2008108958 A2 9, 2008 (21) Appl. No.: 14/075,505 (Continued) OTHER PUBLICATIONS (22) Filed: Nov. 8, 2013 Bukowski et al. “Imidazo[4,5-b]pyridine Derivatives of Potential (65) Prior Publication Data Tuberculostatic Activity.” Archie der Pharmazie. 324.2(1991): 121 127. US 2014/0073635 A1 Mar. 13, 2014 CAS Registry No. 134330-74-8, retrieved Oct. 21, 2011. CAS Registry No. 135286-66-7, retrieved Oct. 21, 2011. CAS Registry No. 95 1258-58-5, retrieved Oct. 21, 2011. Related U.S. ApplicationO O Data CAS Registry No. 95.1258-60-9,95 1258-59-6, retrieved Oct. 21, 2011. (63) Continuation of application No. 13/529,459, filed on AS E. N. 2: ES t : 38: Jun. 21, 2012, now Pat. No. 8,609,688. CAS Registryegistry No. 95 1258-63-2,-oz-1, retrievedretrieve Oct.C. 21, 2011. (60) Provisional application No. 61/500,889, filed on Jun. (Continued) 24, 2011. (51) Int. Cl Primary Examiner — Anna Pagonakis A6 IK3I/437 (2006.01) Etc."gent or Firm — Cooley LLP. Ivor R. Elrifi: CO7D 47L/04 (2006.01) A 6LX3/5377 (2006.01) (57) ABSTRACT A 6LX3L/2197 (2006.01) - 0 A6 IK3I/506 (2006.01) The present invention relates to substituted imidazopyridinyl aminopyridine compounds and methods of synthesizing CO7D 49.8/08 (2006.01) these compounds. The present invention also relates to phar CO7D 451/0249/08 (2006.01) maceuticalical compositions containing Substitutedbstituted imid1m1 azopy ridinyl-aminopyridine compounds and methods of treating (52) U.S. Cl. cell proliferative disorders, such as cancer, by administering CPC ...... C07D 471/04 (2013.01); C07D498/08 these compounds and pharmaceutical compositions to Sub (2013.01); C07D 451/02 (2013.01); C07D jects in need thereof. 491/08 (2013.01) USPC ...... 514/234.2:544/127 6 Claims, No Drawings US 8,962,619 B2 Page 2

(56) References Cited CAS Registry No. 95 1258-69-8, retrieved Oct. 21, 2011. CAS Registry No. 95 1258-70-1, retrieved Oct. 21, 2011. CAS Registry No. 95 1258-71-2, retrieved Oct. 21, 2011. FOREIGN PATENT DOCUMENTS CAS Registry No. 95 1258-72-3, retrieved Oct. 21, 2011. WO WO-2009000413 A1 12/2008 CAS Registry No. 95 1258-73-4, retrieved Oct. 21, 2011. WO WO-2009051454 A2 4/2009 CAS Registry No. 95 1258-74-5, retrieved Oct. 21, 2011. WO WO-2011082270 A2 T 2011 CAS Registry No. 95 1258-79-0, retrieved Oct. 21, 2011. WO WO-2012.177852 A1 12/2012 CAS Registry No. 95 1259-09-9, retrieved Oct. 21, 2011. OTHER PUBLICATIONS CAS Registry No. 95.1259-10-2, retrieved Oct. 21, 2011. CAS Registry No. 95.1258-64-3, retrieved Oct. 21, 2011. CAS Registry No. 95.1259-11-3, retrieved Oct 21, 2011. CAS Registry No. 95.1258-65-4, retrieved Oct. 21, 2011. Tóth etal. "Nitrogen Bridgehead Compounds.”.J. Heterocyclic Chem CAS Registry No. 95.1258-66-5, retrieved Oct. 21, 2011. 28.2(1991);497-501. CAS Registry No. 95.1258-67-6, retrieved Oct. 21, 2011. CAS Registry No. 95.1258-68-7, retrieved Oct. 21, 2011. * cited by examiner US 8,962,619 B2 1. 2 SUBSTITUTED The AKT family regulates cellular survival and metabo MIDAZOPYRIDINYL-AMINOPYRONE lism by binding and regulating many downstream effectors, COMPOUNDS e.g. Nuclear Factor-KB, Bcl-2 family proteins and murine double minute 2 (MDM2). Akt1 is known to play a role in the cell cycle. Moreover, activated Akt1 may enable proliferation CROSS-REFERENCE TO RELATED and survival of cells that have sustained a potentially APPLICATIONS mutagenic impact and, therefore, may contribute to acquisi tion of mutations in other genes. Akt1 has also been impli This application is a continuation of U.S. Ser. No. 13/529, cated in angiogenesis and tumor development. Studies have 459, filed Jun. 21, 2012, which claims priority to, and the shown that deficiency of Akt1 enhanced pathological angio benefit of, U.S. Ser. No. 61/500,889, filed Jun. 24, 2011. The genesis and tumor growth associated with matrix abnormali contents of each of these applications are incorporated herein ties in skin and blood vessels. Since it can block apoptosis, by reference in their entirety. and thereby promote cell survival, Akt1 is a major factor in many types of cancer. BACKGROUND OF THE INVENTION Accordingly, new compounds and methods for modulating 15 AKT genes, proteins and treating proliferation disorders, Cancer is the second leading cause of death in the United including cancer, are needed. The present invention addresses States, exceeded only by heart disease. (Cancer Facts and these needs. Figures 2004, American Cancer Society, Inc.). Despite recent SUMMARY OF THE INVENTION advances in cancer diagnosis and treatment, Surgery and radiotherapy may be curative if a cancer is found early, but The present invention provides, in part, Substituted imida current drug therapies for metastatic disease are mostly pal Zopyridinyl-aminopyridine compounds of formula I or II, and liative and seldom offer a long-term cure. Even with new methods of preparing the compounds of formula I or II: chemotherapies entering the market, the need continues for new drugs effective in monotherapy or in combination with 25 (I) existing agents as first line therapy, and as second and third HN line therapies in treatment of resistant tumors. Cancer cells are by definition heterogeneous. For example, N EN within a single tissue or cell type, multiple mutational "mechanisms' may lead to the development of cancer. As 30 X CO)4. N () Such, heterogeneity frequently exists between cancer cells taken from tumors of the same tissue and same type that have O originated in different individuals. Frequently observed mutational "mechanisms' associated with some cancers may differ between one tissue type and another (e.g., frequently 35 observed mutational "mechanisms' leading to colon cancer HN may differ from frequently observed “mechanisms' leading to leukemias). It is therefore often difficult to predict whether a particular cancer will respond to a particular chemothera peutic agent (Cancer Medicine, 5' edition, Bast et al., B.C. 40 Decker Inc., Hamilton, Ontario). Components of cellular signal transduction pathways that (II) regulate the growth and differentiation of normal cells can, HN when dysregulated, lead to the development of cellular pro N N liferative disorders and cancer. Mutations in cellular signal 45 rN-y ing proteins may cause Such proteins to become expressed or activated at inappropriate levels or at inappropriate times elN/ \ / during the cell cycle, which in turn may lead to uncontrolled cellular growth or changes in cell-cell attachment properties. For example, dysregulation of receptor tyrosine kinases by 50 mutation, gene rearrangement, gene amplification, and over NRR' expression of both receptor and ligand has been implicated in the development and progression of human cancers. HN AKT protein family, which members are also called pro tein kinases B (PKB) plays an important role in mammalian 55 cellular signaling. In humans, there are three genes in the AKT family: Akt1, Akt2, and Akt3. These genes code for wherein: enzymes that are members of the serine/threonine-specific X is NRR, OR, SRs or protein kinase family. Akt1 is involved in cellular survival pathways, by inhibitingapoptotic processes. Akt1 is also able 60 to induce protein synthesis pathways, and is therefore a key :::::: signaling protein in the cellular pathways that lead to skeletal muscle hypertrophy, and general tissue growth. Akt2 is an important signaling molecule in the Insulin signaling path way and is required to induce glucose transport. The role of 65 Akt3 is less clear, though it appears to be predominantly expressed in brain. US 8,962,619 B2 4 wherein R, and R, for each occurrence, are independently H or unsubstituted methyl: R is NRR', C(O)NRR, NR'C(O)R, or NR'S(O), :::::: R7; R and R are each independently H or unsubstituted or Substituted C-C alkyl, or RandR', together with the nitro gen atom to which they are attached, form an unsubstituted or Substituted heterocycle comprising one 5- or 6-member ring and optionally 1 or 2 additional heteroatoms selected from N. 10 O and S: is linked to the imidazopyridinyl ring at the position indicated R, is unsubstituted or substituted C-C alkyl; by ***", R, is H or unsubstituted or substituted C-C alkyl: R, and Rs are each independently unsubstituted or substi R and Rare each H, or R and R', together with the tuted Co-Co aryl; nitrogen atom to which they are attached, form an unsubsti Ry is (CH),R or unsubstituted or Substituted Co-Co 15 tuted or Substituted heterocycle comprising one 6-member aryl; ring and optionally 1 or 2 additional heteroatoms selected R is H; or from N, O and S; Rx and R', together with the nitrogen atom to which they Ra is C(O)Rs or S(O).R.: are attached, form an unsubstituted or Substituted morpho R is unsubstituted or substituted C-C alkyl: line; Rs is unsubstituted or Substituted C-C alkyl or unsubsti m is 1, 2, 3 or 4: tuted or substituted C-C carbocycle; R is unsubstituted or Substituted heterocycle comprising Rs and Rs are each independently H, unsubstituted or one 6-member ring and 1 or 2 heteroatoms selected from N.O substituted C-C alkyl or C(O)NR'R'', provided that Rs and and S; Rs are not both H: 25 R and Ro' are each independently H or unsubstituted or R, is substituted C-C alkyl or unsubstituted C-C alkyl: substituted C-C alkyl: R is (CH), OH or C(O)R; R is H or OH: R" is H; or R is C(O)NRR'; and R and R', together with the nitrogen atom to which they R, and Rare each independently H or unsubstituted or are attached, form a ring selected from 30 substituted C-C alkyl. The present invention also provides pharmaceutical com

:::::: :::::: :::::: positions comprising one or more compounds of each of the formulae described herein and one or more pharmaceutically acceptable carriers. 35 The present invention also provides methods of treating a C D N C D cell proliferative disorder by administering to a subject in C N Rsu? O D Rs' S need thereof, a therapeutically effective amount of a com R3 R4 pound of each of the formulae described herein, or a pharma :::::: N ceutically acceptable salt, prodrug, metabolite, analog or :::::: :::::: 40 derivative thereof, in combination with a pharmaceutically N N acceptable carrier, such that the disorder is treated. The present invention also provides methods of treating s and s cancer by administering to a Subject in need thereof, a thera O R peutically effective amount of a compound of each of the 45 formulae described herein, or a pharmaceutically acceptable Raza salt, prodrug, metabolite, analog or derivative thereof, in combination with a pharmaceutically acceptable carrier, Such wherein the nitrogen atom indicated by “*” is the nitrogen that the cancer is treated. atom to which R and R' are attached; The present invention also provides methods of selectively 50 inducing cell death in precancerous or cancerous cells by o is 1, 2, 3 or 4: contacting a cell with an effective amount of a compound of R is each of the formulae described herein, or a pharmaceutically acceptable salt, prodrug, metabolite, analog or derivative thereof, in combination with a pharmaceutically acceptable 55 carrier, such that contacting the cell results in selective induc tion of cell death in the precancerous or cancer cells. -- rose Unless otherwise defined, all technical and scientific terms R.' used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention tert-butyl, unsubstituted or substituted C-C carbocycle or 60 belongs. In the specification, the singular forms also include unsubstituted or Substituted heterocycle comprising one the plural unless the context clearly dictates otherwise. 6-member ring and 1 or 2 heteroatoms selected from N, O and Although methods and materials similar or equivalent to S; those described herein can be used in the practice or testing of n is 0, 1, 2 or 3: the present invention, Suitable methods and materials are Ro is H, unsubstituted or substituted C-C alkyl or C(O) 65 described below. All publications, patent applications, pat R; ents, and other references mentioned herein are incorporated R is unsubstituted or Substituted C-C alkyl; by reference. The references cited herein are not admitted to US 8,962,619 B2 5 6 be prior art to the claimed invention. In the case of conflict, the R is H; or present specification, including definitions, will control. In R and R', together with the nitrogen atom to which they addition, the materials, methods, and examples are illustra are attached, form an unsubstituted or Substituted morpho tive only and are not intended to be limiting. line; Other features and advantages of the invention will be m is 1, 2, 3 or 4: apparent from the following detailed description and claims. R is unsubstituted or Substituted heterocycle comprising one 6-member ring and 1 or 2 heteroatoms selected from N.O and S; and DETAILED DESCRIPTION OF THE INVENTION R, is substituted C-C alkylor unsubstituted C-C alkyl. 10 For example, X is NRR. 1. Substituted Imidazopyridinyl-Aminopyridine For example, R is H and R is (CH), R. Compounds For example, m is 1 or 2 and R is heterocycle selected from pyrrolidinyl, imidazolidinyl, pyrazolidinyl, oxazolidi The present invention provides novel substituted imida 15 nyl, isoxazolidinyl, triazolidinyl, tetrahyrofuranyl, piperidi Zopyridinyl-aminopyridine compounds, synthetic methods nyl, piperazinyl, and morpholinyl. For example, m is 2 and for making the compounds, pharmaceutical compositions R is morpholinyl. containing them and various uses of the disclosed com For example, R is H and R is unsubstituted phenyl. pounds. For example, R is HandR is phenyl substituted with one The present invention provides the compounds of formula or more groups, each of which is independently selected from I: hydroxyl, halogen (e.g., fluorine, chlorine, bromine and iodine), cyano, nitro, unsubstituted or Substituted amino (e.g., amino, C-C alkylamino and di-C-C alkylamino), unsub (I) stituted or Substituted C-C alkyl (e.g., methyl, ethyl, n-pro HN 25 pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, Sl-N N s-pentyl, and n-hexyl), unsubstituted or Substituted C-C, \ alkoxy (e.g., methoxy, ethoxy, n-propyloxy, i-propyloxy, n-butoxy, and t-butoxy), and heterocycle comprising one 5 X N elN/ \ / or 6-member ring and 1-3 heteroatoms selected from N, O 30 and S (e.g., pyrrolidinyl, imidazolidinyl, pyrazolidinyl, oxazolidinyl, isoxazolidinyl, triazolidinyl, tetrahyrofuranyl. piperidinyl, piperazinyl, and morpholinyl). For example, R is H and R is phenyl substituted at the ortho-, meta- or para-position. 35 For example, R is H and R is phenyl substituted with morpholinyl at the ortho-position, morpholinyl at the meta Q. position or morpholinyl at the para-position. For example, Ry and R', together with the nitrogen atom wherein: to which they are attached, form an unsubstituted morpho 40 line. X is NRR, OR SRs or For example, Ry and R', together with the nitrogen atom to which they are attached, form a morpholine substituted with one or more groups, each of which is independently selected from hydroxyl, halogen (e.g., fluorine, chlorine, bro 45 mine and iodine), cyano, nitro, unsubstituted or Substituted amino (e.g., amino, C-C alkylamino and di-C-C alky lamino), unsubstituted or substituted C-C alkyl (e.g., C methyl, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl), and unsubstituted or 50 Substituted C-C alkoxy (e.g., methoxy, ethoxy, n-propy wherein loxy, i-propyloxy, n-butoxy, and t-butoxy). For example, X is OR. For example, R is unsubstituted phenyl. For example, R is phenyl substituted with one or more 55 groups, each of which is independently selected from hydroxyl, halogen (e.g., fluorine, chlorine, bromine and iodine), cyano, nitro, unsubstituted or Substituted amino (e.g., O amino, C-C alkylamino and di-C-C alkylamino), unsub stituted or Substituted C-C alkyl (e.g., methyl, ethyl, n-pro 60 pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, s-pentyl, and n-hexyl), unsubstituted or substituted C-C, is linked to the imidazopyridinyl ring at the position indicated alkoxy (e.g., methoxy, ethoxy, n-propyloxy, i-propyloxy, by &G 3". n-butoxy, and t-butoxy), and heterocycle comprising one 5 R and Rs are each independently unsubstituted or Substi or 6-member ring and 1-3 heteroatoms selected from N, O tuted Co-Co aryl; 65 and S (e.g., pyrrolidinyl, imidazolidinyl, pyrazolidinyl, Ry is (CH),R or unsubstituted or Substituted Co-Co oxazolidinyl, isoxazolidinyl, triazolidinyl, tetrahyrofuranyl. aryl; piperidinyl, piperazinyl, and morpholinyl). US 8,962,619 B2 7 For example, X is SRs. -continued :::::: For example, Rs is unsubstituted phenyl. N For example, Rs is phenyl substituted with one or more :::::: :::::: groups, each of which is independently selected from N N hydroxyl, halogen (e.g., fluorine, chlorine, bromine and s and s iodine), cyano, nitro, unsubstituted or Substituted amino (e.g., amino, C-C alkylamino and di-C-C alkylamino), unsub O R stituted or substituted C-C alkyl (e.g., methyl, ethyl, n-pro pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, Raza s-pentyl, and n-hexyl), unsubstituted or Substituted C-C, 10 alkoxy (e.g., methoxy, ethoxy, n-propyloxy, i-propyloxy, wherein the nitrogen atom indicated by “*” is the nitrogen n-butoxy, and t-butoxy), and heterocycle comprising one 5 atom to which R and Rare attached; or 6-member ring and 1-3 heteroatoms selected from N, O and S (e.g., pyrrolidinyl, imidazolidinyl, pyrazolidinyl, o is 1, 2, 3 or 4: oxazolidinyl, isoxazolidinyl, triazolidinyl, tetrahyrofuranyl. 15 R is piperidinyl, piperazinyl, and morpholinyl). For example, X is R --C--ORio, :::::: R

tert-butyl, unsubstituted or substituted C-C carbocycle or 25 unsubstituted or Substituted heterocycle comprising one 6-member ring and 1 or 2 heteroatoms selected from N, O and S; For example, R, is substituted straight-chain or branched n is 0, 1, 2 or 3: C-C alkyl, including but not limited to, n-propyl, i-propyl. Ro is H, unsubstituted or substituted C-C alkyl or C(O) n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and 30 R11; n-hexyl, each of which is substituted. For example, R, is R is unsubstituted or substituted C-C alkyl; unsubstituted Straight-chain or branched C-C alkyl, includ ing but not limited to, n-butyl, i-butyl, S-butyl, t-butyl, n-pen R, and R, for each occurrence, are independently H or tyl, s-pentyl, and n-hexyl. For example, R, is t-butyl or unsubstituted methyl: substituted n-propyl. For example, R, is 2-hydroxypropyl. 35 The present invention also provides the compounds of R and R are each independently H or unsubstituted or formula II: Substituted C-C alkyl, or RandR', together with the nitro gen atom to which they are attached, form an unsubstituted or (II) HN Substituted heterocycle comprising one 5- or 6-member ring 40 and optionally 1 or 2 additional heteroatoms selected from N. N N N O and S: N R, is unsubstituted or substituted C-C alkyl; N elN/ \ / R, is H or unsubstituted or substituted C-C alkyl: 45 R and Rare each H, or R and R', together with the nitrogen atom to which they are attached, form an unsubsti tuted or Substituted heterocycle comprising one 6-member NRR ring and optionally 1 or 2 additional heteroatoms selected from N, O and S; HN 50 Ra is C(O)Rs or S(O).R.: R is unsubstituted or substituted C-C alkyl: Rs is unsubstituted or Substituted C-C alkyl or unsubsti wherein: tuted or Substituted C-Cs carbocycle; R is (CH), OH or C(O)R; Rs and Rs are each independently H, unsubstituted or R" is H; or 55 substituted C-C alkyl or C(O)NRR", provided that Rs and R and R', together with the nitrogen atom to which they Rs are not both H; and are attached, form a ring selected from R and Ro' are each independently H or unsubstituted or Substituted C-C alkyl; and

:::::: :::::: :::::: 60 R is H or OH: N N :::::: N R is C(O)NRR'; and R, and Rare each independently H or unsubstituted or C D N C D substituted C-C alkyl. N Rsu? O D Rs' S 65 For example, R is Hand R is (CH), OH. R3 R4 For example, o is 2, 3 or 4. For example, o is 2. US 8,962,619 B2 9 10 For example, R is H and R is C(O)R. For example, R is C(O)NRR". For example, R is For example, R and Rare not both H. For example, Reis Hand R is unsubstituted or substituted, 5 straight chain C-C alkyl or branched C-C alkyl, including RC but not limited to, methyl, ethyl, n-propyl. i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl, each - to of which is optionally substituted with one or more groups RC independently selected from hydroxyl and unsubstituted or 10 Substituted C-C alkoxy (e.g., methoxy, ethoxy, propyloxy, For example, n is 0 and Rio is unsubstituted or substituted, i-propyloxy, butoxy, and t-butoxy). straight chain C-C alkyl or branched C-C alkyl, including For example, R is H and R is methyl, ethyl, propyl. but not limited to, methyl, ethyl, n-propyl. i-propyl. n-butyl, i-propyl or ethyl substituted with methoxy. i-butyl, S-butyl, t-butyl, n-penty1, S-pentyl, and n-hexyl, each 15 For example, R. and R are each independently unsubsti of which is optionally substituted. For example, Ro is tuted or substituted, straight chain C-C alkyl or branched methyl. C-C alkyl, including but not limited to, methyl, ethyl, n-pro For example, n is 1 and R, and Rare each methyl. pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, s-pentyl, and n-hexyl, each of which is optionally Substituted For example, Rio is H. with one or more groups independently selected from For example, Ro is unsubstituted or Substituted, straight hydroxyl and unsubstituted or substituted C-C alkoxy (e.g., chain C-C alkyl or branched C-C alkyl, including but not methoxy, ethoxy, propyloxy, propyloxy, butoxy, and t-bu limited to, methyl, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, toxy). S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl, each of which For example, RandR are each methyl. For example, R. is optionally Substituted. For example, Ro is methyl. 25 and R are each ethyl. For example, Ro is C(O)R and R is unsubstituted or Substituted, straight chain C-C alkyl or branched C-C, For example, RandR', together with the nitrogenatom to alkyl, including but not limited to, methyl, ethyl, n-propyl. which they are attached, form a heterocycle selected from i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, piperidine, piperazine and morpholine. For example, Re and and n-hexyl, each of which is optionally substituted. For 30 R", together with the nitrogen atom to which they are example, R is methyl. attached, form a morpholine. For example, R is tert-butyl. For example, R is NR'C(O)R. For example, R is unsubstituted or substituted C-C car For example, R, is unsubstituted or substituted, straight bocycle, including but not limited to, cyclopropyl, cyclobu 35 chain C-C alkyl or branched C-C alkyl (e.g., methyl, tyl, cyclopentyl, cyclohexyl, and cycloheptyl. For example, ethyl, n-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, R is cyclopropyl. s-pentyl, and n-hexyl), and R, is unsubstituted or substituted, For example, R is unsubstituted or substituted heterocycle straight chain C-C alkyl or branched C-C alkyl (e.g., selected from pyrrolidinyl, imidazolidinyl, pyrazolidinyl, 40 methyl, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, S-butyl, oxazolidinyl, isoxazolidinyl, triazolidinyl, tetrahyrofuranyl. piperidinyl, piperazinyl, morpholinyl, tetrahydropyranyl, and t-butyl, n-pentyl, S-pentyl, and n-hexyl, each of which is dioxanyl. For example, R is tetrahydropyranyl or 1,4-dioxa optionally Substituted with one or more groups independently nyl. selected from hydroxyl and unsubstituted or substituted For example, RandR', together with the nitrogenatom to 45 C-C alkoxy (e.g., methoxy, ethoxy, propyloxy, i-propyloxy, which they are attached, form butoxy, and t-butoxy)). For example, R, is methyl and R is methyl.

:::::: For example, R, is Hand R, is unsubstituted or substituted, 50 straight chain C-C alkyl or branched C-C alkyl (e.g., methyl, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl, each of which is optionally Substituted with one or more groups independently 55 selected from hydroxyl and unsubstituted or substituted C-C alkoxy (e.g., methoxy, ethoxy, propyloxy, i-propyloxy, For example, R is NRR". butoxy, and t-butoxy)). For example, R, is methyl or methyl For example, R and R2 are each H. substituted with methoxy. 60 For example, R2 and R', together with the nitrogenatom For example, R is NR,'S(O).R. to which they are attached, form an unsubstituted or substi For example, R, is H, and R, is unsubstituted or substi tuted heterocycle selected from pyrrolidinyl, imidazolidinyl, pyrazolidinyl, oxazolidinyl, isoxazolidinyl, triazolidinyl, tet tuted, straight chain C-C alkyl or branched C-C alkyl rahyrofuranyl, piperidinyl, piperazinyl, and morpholinyl. For 65 (e.g., methyl, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, S-bu example, R and R', together with the nitrogen atom to tyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl). For example, R, which they are attached, form a morpholinyl. is methyl. US 8,962,619 B2 11 12 For example, RandR', together with the nitrogenatom to For example, Reis Hand R is unsubstituted or substituted, which they are attached, form straight chain C-C alkyl or branched C-C alkyl, including but not limited to, methyl, ethyl, n-propyl. i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl, each of which is optionally substituted. For example, RandR', together with the nitrogenatom to which they are attached, form

O 10

For example, R is C(O)Rs. For example, Rs is unsubstituted or substituted, straight chain C-C alkyl or branched C-C alkyl, including but not 15 limited to, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl, each of which is optionally substituted. For example, Rs is ethyl, i-propyl or For example, RandR', together with the nitrogenatom to t-butyl. which they are attached, form For example, Rs is unsubstituted or Substituted C-Cs car bocycle, including but not limited to, cyclopropyl, cyclobu tyl, cyclopentyl, cyclohexyl, and cycloheptyl. For example, :::::: Rs is cyclopropyl. For example, R is S(O).R. 25 For example, R, is unsubstituted or substituted, straight chain C-C alkyl or branched C-C alkyl, including but not limited to, methyl, ethyl, n-propyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, S-pentyl, and n-hexyl, each of which 30 For example, RandR', together with the nitrogenatom to is optionally substituted. For example, R is methyl. which they are attached, form For example, RandR', together with the nitrogenatom to which they are attached, form :::::: 35

Rsul O Rs. 40

For example, Rs' and Rs are each independently unsubsti tuted or substituted, straight chain C-C alkyl or branched 45 For example, R is OH. C-C alkyl, including but not limited to, methyl, ethyl, n-pro For example, RandR', together with the nitrogenatom to pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, which they are attached, form s-pentyl, and n-hexyl, each of which is optionally Substituted with one or more hydroxyl. For example, Rs. Rs are each methyl or methyl substituted with hydroxyl. 50 ::::::

For example, Rs' is Hand Rs is unsubstituted or substituted, straight chain C-C alkyl or branched C-C alkyl, including 55 but not limited to, methyl, ethyl, n-propyl. i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-penty1, S-pentyl, and n-hexyl, each For example, only one of RandR is H. For example, R. of which is optionally substituted. For example, Rs' is Hand is H and R is methyl. Rs is methyl substituted with hydroxyl. For example, R, and Rare each independently unsubsti For example, Rs' is Hand Rs is C(O)NR'R''. 60 tuted or substituted, straight chain C-C alkyl or branched For example, R.," and R are each independently unsubsti C-C alkyl, including but not limited to, methyl, ethyl, n-pro tuted or Substituted, straight chain C-C alkyl or branched pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, C-C alkyl, including but not limited to, methyl, ethyl, n-pro s-pentyl, and n-hexyl, each of which is optionally Substituted. pyl, i-propyl. n-butyl, i-butyl, S-butyl, t-butyl, n-pentyl, 65 For example, R, and Rare each methyl. s-pentyl, and n-hexyl, each of which is optionally Substituted. Representative compounds of the present invention For example, Ro" and Ro are each methyl. include compounds listed in Table 1.

US 8,962,619 B2 35 36 TABLE 1-continued Cmpd Structure Chemical Name 45 HN N-(3-(3-(4-(1- aminocyclobutyl)phenyl)-2-(2- 21 N N aminopyridin-3-yl)-3H N imidazo[4,5-b]pyridin-5- yl)phenyl)-2-methoxy-2- su-MN \ / methylpropanamide

No NH O NH2

46 HN 3-(3-4-(1- aminocyclobutyl)phenyl-5-3- 21 N N 4-(methylsulfonyl)piperazin-1- N yl)phenyl-3H-imidazo[4,5- bipyridin-2-yl)pyridin-2-amine Null? \ / hydrochloride

N C D NH2 N o==o CH3

35 As used herein, “alkyl”, “C, C, C, C, Cs or C alkyl or rylamino and alkylarylamino), acylamino (including alkyl "C-C alkyl is intended to include C, C, C, C, Cs or C carbonylamino, arylcarbonylamino, carbamoyl and ureido), straight chain (linear) saturated aliphatic hydrocarbon groups amidino, imino, Sulfhydryl, alkylthio, arylthio, thiocarboxy and C, C, Cs or C. branched saturated aliphatic hydrocar late, Sulfates, alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfona bon groups. For example, C-C alkyl is intended to include 40 mido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alky C, C, C, C, Cs and C alkyl groups. Examples of alkyl laryl, or an aromatic or heteroaromatic moiety. Cycloalkyls include, moieties having from one to six carbon atoms, such can be further substituted, e.g., with the substituents as, but not limited to, methyl, ethyl, n-propyl, i-propyl. n-bu described above. An “alkylaryl or an “aralkyl moiety is an tyl, S-butyl, t-butyl, n-pentyl, S-pentyl or n-hexyl. alkyl Substituted with an aryl (e.g., phenylmethyl (benzyl)). In certain embodiments, a straight chain or branched alkyl 45 Unless the number of carbons is otherwise specified, has six or fewer carbon atoms (e.g., C-C for straight chain, “lower alkyl includes an alkyl group, as defined above, hav C-C for branched chain), and in another embodiment, a ing from one to six, or in another embodiment from one to straight chain or branched alkyl has four or fewer carbon four, carbonatoms in its backbone structure. “Lower alkenyl atOmS. and “lower alkynyl have chain lengths of, for example, two “Heteroalkyl groups are alkyl groups, as defined above, 50 to six or of two to four carbon atoms. that have an oxygen, nitrogen, Sulfur or phosphorous atom As used herein, “alkyl linker is intended to include C, C, replacing one or more hydrocarbon backbone carbon atoms. C, C, Cs or C. Straight chain (linear) saturated aliphatic As used herein, the term "cycloalkyl”, “C, C, Cs, C, C, hydrocarbon groups and C, C, Cs or C. branched saturated or Cs cycloalkyl” or "C-C cycloalkyl is intended to include aliphatic hydrocarbon groups. For example, C-C alkyl hydrocarbon rings having from three to eight carbon atoms in 55 linker is intended to include C, C, C, C, Cs and C alkyl their ring structure. In one embodiment, a cycloalkyl group linker groups. Examples of alkyl linker include, moieties has five or six carbons in the ring structure. having from one to six carbon atoms, such as, but not limited The term “substituted alkyl refers to alkyl moieties having to, methyl (—CH2—), ethyl (—CH2CH2—), n-propyl Substituents replacing one or more hydrogenatoms on one or (—CH2CHCH ), i-propyl ( CHCHCH ), n-butyl more carbons of the hydrocarbon backbone. Such substitu 60 (—CH2CH2CHCH ), s-butyl ( CHCHCHCH ), ents can include, for example, alkyl, alkenyl, alkynyl, halo i-butyl (—C(CH)2CH ), n-pentyl gen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxy (—CH2CH2CHCHCH ), s-pentyl carbonyloxy, aryloxycarbonyloxy, carboxylate, (-CHCHCHCHCH ) O n-hexyl alkylcarbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbo (—CH2CH2CH2CH2CH2CH2—). nyl, alkylaminocarbonyl, dialkylaminocarbonyl, alkylthio 65 Alkenyl' includes unsaturated aliphatic groups analogous carbonyl, alkoxyl, phosphate, phosphonato, phosphinato, in length and possible substitution to the alkyls described amino (including alkylamino, dialkylamino, arylamino, dia above, but that contain at least one double bond. For example, US 8,962,619 B2 37 38 the term “alkenyl' includes straight chain alkenyl groups "Aryl includes groups with aromaticity, including “con (e.g., ethenyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, jugated, or multicyclic, systems with at least one aromatic octenyl, nonenyl, decenyl), branched alkenyl groups, ring. Examples include phenyl, benzyl, etc. cycloalkenyl (e.g., alicyclic) groups (e.g., cyclopropenyl, “Heteroaryl groups are aryl groups, as defined above, cyclopentenyl, cyclohexenyl, cycloheptenyl, cyclooctenyl), having from one to four heteroatoms in the ring structure, and alkyl or alkenyl Substituted cycloalkenyl groups, and may also be referred to as “arylheterocycles' or "heteroaro cycloalkyl or cycloalkenyl Substituted alkenyl groups. In cer matics’. As used herein, the term "heteroaryl is intended to tain embodiments, a straight chain or branched alkenyl group include a stable 5-, 6-, or 7-membered monocyclic or 7-8- has six or fewer carbon atoms in its backbone (e.g., C-C for 9-, 10-, 11- or 12-membered bicyclic aromatic heterocyclic straight chain, C-C for branched chain). Likewise, 10 ring which consists of carbon atoms and one or more heteroa cycloalkenyl groups may have from five to eight carbon toms, e.g., 1 or 1-2 or 1-3 or 1-4 or 1-5 or 1-6 heteroatoms, atoms in their ring structure, and in one embodiment, independently selected from the group consisting of nitrogen, cycloalkenyl groups have five or six carbons in the ring struc oxygen and Sulfur. The nitrogen atom may be substituted or ture. The term "C-C, includes alkenyl groups containing unsubstituted (i.e., Nor NR wherein R is Hor other substitu two to six carbon atoms. The term "C-C includes alkenyl 15 ents, as defined). The nitrogen and Sulfur heteroatoms may groups containing three to six carbon atoms. optionally be oxidized (i.e., N->O and S(O), where p=1 or “Heteroalkenyl' includes alkenyl groups, as defined 2). It is to be noted that total number of S and O atoms in the herein, having an oxygen, nitrogen, Sulfur or phosphorous aromatic heterocycle is not more than 1. atom replacing one or more hydrocarbon backbone carbons. Examples of heteroaryl groups include pyrrole, furan, The term “substituted alkenyl refers to alkenyl moieties thiophene, thiazole, isothiazole, imidazole, triazole, tetra having Substituents replacing one or more hydrogenatoms on Zole, pyrazole, oxazole, isoxazole, pyridine, pyrazine, one or more hydrocarbon backbone carbonatoms. Such sub pyridazine, pyrimidine, and the like. stituents can include, for example, alkyl, alkenyl, alkynyl, Furthermore, the terms “aryl and "heteroaryl' include halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, multicyclic aryl and heteroaryl groups, e.g., tricyclic, bicy alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkyl 25 clic, e.g., naphthalene, benzoxazole, benzodioxazole, ben carbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, Zothiazole, benzoimidazole, benzothiophene, methylene alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbo dioxyphenyl, quinoline, isoquinoline, naphthridine, indole, nyl, alkoxyl, phosphate, phosphonato, phosphinato, amino benzofuran, purine, benzofuran, deaZapurine, indolizine. (including alkylamino, dialkylamino, arylamino, diary In the case of multicyclic aromatic rings, only one of the lamino and alkylarylamino), acylamino (including alkylcar 30 rings needs to be aromatic (e.g., 2,3-dihydroindole), although bonylamino, arylcarbonylamino, carbamoyl and ureido), all of the rings may be aromatic (e.g., quinoline). The second amidino, imino, sulfhydryl, alkylthio, arylthio, thiocarboxy ring can also be fused or bridged. late, Sulfates, alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfona The aryl or heteroaryl aromatic ring can be substituted at mido, nitro, trifluoromethyl, cyano, heterocyclyl alkylaryl, one or more ring positions with Such Substituents as described or an aromatic or heteroaromatic moiety. 35 above, for example, alkyl, alkenyl, akynyl, halogen, Alkynyl includes unsaturated aliphatic groups analogous hydroxyl, alkoxy, alkylcarbonyloxy, arylcarbonyloxy, in length and possible substitution to the alkyls described alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkyl above, but which contain at least one triple bond. For carbonyl, alkylaminocarbonyl, aralkylaminocarbonyl, alk example, "alkynyl' includes straight chain alkynyl groups enylaminocarbonyl, alkylcarbonyl, arylcarbonyl, aralkylcar (e.g., ethynyl, propynyl, butynyl, pentynyl, hexynyl, hepty 40 bonyl, alkenylcarbonyl, alkoxycarbonyl, aminocarbonyl, nyl, octynyl, nonynyl, decynyl), branched alkynyl groups, alkylthiocarbonyl, phosphate, phosphonato, phosphinato, and cycloalkyl or cycloalkenyl Substituted alkynyl groups. In amino (including alkylamino, dialkylamino, arylamino, dia certain embodiments, a straight chain or branched alkynyl rylamino and alkylarylamino), acylamino (including alkyl group has six or fewer carbon atoms in its backbone (e.g., carbonylamino, arylcarbonylamino, carbamoyl and ureido), C-C for straight chain, C-C for branched chain). The term 45 amidino, imino, Sulfhydryl, alkylthio, arylthio, thiocarboxy "C-C includes alkynyl groups containing two to six carbon late, Sulfates, alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfona atoms. The term "C-C, includes alkynyl groups containing mido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alky three to six carbon atoms. laryl, or an aromatic or heteroaromatic moiety. Aryl groups “Heteroalkynyl' includes alkynyl groups, as defined can also be fused or bridged with alicyclic or heterocyclic herein, having an oxygen, nitrogen, Sulfur or phosphorous 50 rings, which are not aromatic So as to form a multicyclic atom replacing one or more hydrocarbon backbone carbons. system (e.g., tetralin, methylenedioxyphenyl). The term “substituted alkynyl refers to alkynyl moieties As used herein, "carbocycle' or “carbocyclic ring is having Substituents replacing one or more hydrogenatoms on intended to include any stable monocyclic, bicyclic or tricy one or more hydrocarbon backbone carbonatoms. Such sub clic ring having the specified number of carbons, any of which stituents can include, for example, alkyl, alkenyl, alkynyl, 55 may be saturated, unsaturated, or aromatic. For example, a halogen, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, C-C carbocycle is intended to include a monocyclic, bicy alkoxycarbonyloxy, aryloxycarbonyloxy, carboxylate, alkyl clic or tricyclic ring having 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13 or carbonyl, arylcarbonyl, alkoxycarbonyl, aminocarbonyl, 14 carbon atoms. Examples of carbocycles include, but are alkylaminocarbonyl, dialkylaminocarbonyl, alkylthiocarbo not limited to, cyclopropyl, cyclobutyl, cyclobutenyl, cyclo nyl, alkoxyl, phosphate, phosphonato, phosphinato, amino 60 pentyl, cyclopentenyl, cyclohexyl, cycloheptenyl, cyclohep (including alkylamino, dialkylamino, arylamino, diary tyl, cycloheptenyl, adamantyl, cyclooctyl, cyclooctenyl, lamino and alkylarylamino), acylamino (including alkylcar cyclooctadienyl, fluorenyl, phenyl, naphthyl, indanyl, ada bonylamino, arylcarbonylamino, carbamoyl and ureido), am manty1 and tetrahydronaphthyl. Bridged rings are also idino, imino, Sulfhydryl, alkylthio, arylthio, thiocarboxylate, included in the definition of carbocycle, including, for Sulfates, alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfonamido, 65 example, 3.3.0 bicyclooctane, 4.3.0 bicyclononane, 4.4.0 nitro, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, bicyclodecane and 2.2.2 bicyclooctane. A bridged ring or an aromatic or heteroaromatic moiety. occurs when one or more carbonatoms link two non-adjacent US 8,962,619 B2 39 40 carbonatoms. In one embodiment, bridge rings are one or two When any variable (e.g., R) occurs more than one time in carbon atoms. It is noted that a bridge always converts a any constituent or formula for a compound, its definition at monocyclic ring into a tricyclic ring. When a ring is bridged, each occurrence is independent of its definition at every other the substituents recited for the ring may also be present on the occurrence. Thus, for example, if a group is shown to be bridge. Fused (e.g., naphthyl, tetrahydronaphthyl) and spiro Substituted with 0-2 R moieties, then the group may option rings are also included. ally be substituted with up to two R moieties and R at each As used herein, "heterocycle' includes any ring structure occurrence is selected independently from the definition of (Saturated or partially unsaturated) which contains at least R. Also, combinations of Substituents and/or variables are one ring heteroatom (e.g., N, O or S). Examples of hetero permissible, but only if such combinations result in stable 10 compounds. cycles include, but are not limited to, morpholine, pyrroli The term “hydroxy” or “hydroxyl includes groups with an dine, tetrahydrothiophene, piperidine, piperazine and tet - OH or - O. rahydrofuran. As used herein, “halo' or “halogen refers to fluoro, Examples of heterocyclic groups include, but are not lim chloro, bromo and iodo. The term “perhalogenated' gener ited to, acridinyl, azocinyl, benzimidazolyl, benzofuranyl. 15 ally refers to a moiety wherein all hydrogen atoms are benzothiofuranyl, benzothiophenyl, benzoxazolyl, benzox replaced by halogen atoms. azolinyl, benzthiazolyl, benztriazolyl, benztetrazolyl, ben The term “carbonyl or “carboxy” includes compounds Zisoxazolyl, benzisothiazolyl, benzimidazolinyl, carbazolyl, and moieties which contain a carbon connected with a double 4aH-carbazolyl, carbolinyl, chromanyl, chromenyl, cinnoli bond to an oxygen atom. Examples of moieties containing a nyl, decahydroquinolinyl, 2H,6H-1.5.2-dithiazinyl, dihydro carbonyl include, but are not limited to, aldehydes, ketones, furo2.3-btetrahydrofuran, furanyl, furazanyl, imidazolidi carboxylic acids, amides, esters, anhydrides, etc. nyl, imidazolinyl, imidazolyl, 1H-indazolyl, indolenyl, Acyl' includes moieties that contain the acyl radical ( C indolinyl, indolizinyl, indolyl, 3H-indolyl, isatinoyl, isoben (O)—) or a carbonyl group. “Substituted acyl includes acyl Zofuranyl, isochromanyl, isolindazolyl, isoindolinyl, isoin groups where one or more of the hydrogenatoms are replaced dolyl, isoquinolinyl, isothiazolyl, isoxazolyl, methylenediox 25 by, for example, alkyl groups, alkynyl groups, halogen, yphenyl, morpholinyl, naphthyridinyl, hydroxyl, alkylcarbonyloxy, arylcarbonyloxy, alkoxycarbo octahydroisoquinolinyl, oxadiazolyl, 1.2.3-oxadiazolyl, 1.2, nyloxy, aryloxycarbonyloxy, carboxylate, alkylcarbonyl, 4-oxadiazolyl, 1,2,5-oxadiazolyl, 1.3,4-oxadiazolyl, 1,2,4- arylcarbonyl, alkoxycarbonyl, aminocarbonyl, alkylami oxadiazols(4H)-one, oxazolidinyl, oxazolyl, oxindolyl pyri nocarbonyl, dialkylaminocarbonyl, alkylthiocarbonyl, midinyl, phenanthridinyl, phenanthrolinyl, phenazinyl, 30 alkoxyl, phosphate, phosphonato, phosphinato, amino (in phenothiazinyl, phenoxathinyl, phenoxazinyl, phthalazinyl, cluding alkylamino, dialkylamino, arylamino, diarylamino piperazinyl, piperidinyl, piperidonyl, 4-piperidonyl, pipero and alkylarylamino), acylamino (including alkylcarbony nyl, pteridinyl, purinyl, pyranyl, pyrazinyl, pyrazolidinyl, lamino, arylcarbonylamino, carbamoylandureido), amidino, pyrazolinyl, pyrazolyl, pyridazinyl, pyridooxazole, pyri imino, sulfhydryl, alkylthio, arylthio, thiocarboxylate, sul doimidazole, pyridothiazole, pyridinyl, pyridyl, pyrimidinyl, 35 fates, alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfonamido, nitro, pyrrolidinyl, pyrrolinyl, 2H-pyrrolyl pyrrolyl, quinazolinyl, trifluoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an quinolinyl, 4H-quinolizinyl, quinoxalinyl, quinuclidinyl, tet aromatic or heteroaromatic moiety. rahydrofuranyl, tetrahydroisoquinolinyl, tetrahydroquinoli “Aroyl includes moieties with an aryl or heteroaromatic nyl, tetrazolyl, 6H-1,2,5-thiadiazinyl, 1,2,3-thiadiazolyl, 1.2, moiety bound to a carbonyl group. Examples of aroyl groups 4-thiadiazolyl, 1,2,5-thiadiazolyl, 1,3,4-thiadiazolyl, 40 include phenylcarboxy, naphthyl carboxy, etc. thianthrenyl, thiazolyl, thienyl, thienothiazolyl, thienoox “Alkoxyalkyl”, “alkylaminoalkyl and “thioalkoxyalkyl azolyl, thienoimidazolyl, thiophenyl, triazinyl, 1.2.3-triaz include alkyl groups, as described above, wherein oxygen, olyl, 1,2,4-triazolyl, 1.2.5-triazolyl, 1,3,4-triazolyl and Xan nitrogen or Sulfur atoms replace one or more hydrocarbon thenyl. backbone carbon atoms. The term “substituted, as used herein, means that any one 45 The term “alkoxy' or “alkoxyl includes substituted and or more hydrogen atoms on the designated atom is replaced unsubstituted alkyl, alkenyl and alkynyl groups covalently with a selection from the indicated groups, provided that the linked to an oxygen atom. Examples of alkoxy groups or designated atoms normal Valency is not exceeded, and that alkoxyl radicals include, but are not limited to, methoxy, the substitution results in a stable compound. When a sub ethoxy, isopropyloxy, propoxy, butoxy and pentoxy groups. stituent is keto (i.e., =O), then 2 hydrogenatoms on the atom 50 Examples of Substituted alkoxy groups include halogenated are replaced. Keto Substituents are not present on aromatic alkoxy groups. The alkoxy groups can be substituted with moieties. Ring double bonds, as used herein, are double groups such as alkenyl, alkynyl, halogen, hydroxyl, alkylcar bonds that are formed between two adjacent ring atoms (e.g., bonyloxy, arylcarbonyloxy, alkoxycarbonyloxy, aryloxycar C—C, C=N or N=N). “Stable compound” and “stable bonyloxy, carboxylate, alkylcarbonyl, arylcarbonyl, alkoxy structure' are meant to indicate a compound that is suffi 55 carbonyl, aminocarbonyl, alkylaminocarbonyl, ciently robust to survive isolation to a useful degree of purity dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, phos from a reaction mixture, and formulation into an efficacious phate, phosphonato, phosphinato, amino (including alky therapeutic agent. lamino, dialkylamino, arylamino, diarylamino, and alkylary When a bond to a substituent is shown to cross a bond lamino), acylamino (including alkylcarbonylamino, connecting two atoms in a ring, then such Substituent may be 60 arylcarbonylamino, carbamoyl and ureido), amidino, imino, bonded to any atom in the ring. When a substituent is listed sulfhydryl, alkylthio, arylthio, thiocarboxylate, sulfates, without indicating the atom via which Such substituent is alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfonamido, nitro, trif bonded to the rest of the compound of a given formula, then luoromethyl, cyano, azido, heterocyclyl, alkylaryl, or an aro Such substituent may be bonded via any atom in Such formula. matic or heteroaromatic moieties. Examples of halogen Sub Combinations of substituents and/or variables are permis 65 stituted alkoxy groups include, but are not limited to, sible, but only if such combinations result in stable com fluoromethoxy, difluoromethoxy, trifluoromethoxy, chlo pounds. romethoxy, dichloromethoxy and trichloromethoxy. US 8,962,619 B2 41 42 The term “ether” or “alkoxy' includes compounds or moi bound to the carbon of a carbonyl or thiocarbonyl group. It eties which contain an oxygenbonded to two carbonatoms or also includes “arylaminocarboxy' groups that include aryl or heteroatoms. For example, the term includes “alkoxyalkyl, heteroaryl moieties bound to an amino group that is bound to which refers to an alkyl, alkenyl, or alkynyl group covalently the carbon of a carbonyl or thiocarbonyl group. The terms bonded to an oxygen atom which is covalently bonded to an “alkylaminocarboxy”, “alkenylaminocarboxy”, “alkyny alkyl group. laminocarboxy” and “arylaminocarboxy” include moieties The term “ester includes compounds or moieties which wherein alkyl, alkenyl, alkynyl and aryl moieties, respec contain a carbon or a heteroatom bound to an oxygen atom tively, are bound to a nitrogen atom which is in turn bound to which is bonded to the carbon of a carbonyl group. The term the carbon of a carbonyl group. Amides can be substituted 'ester” includes alkoxycarboxy groups such as methoxycar 10 with Substituents such as Straight chain alkyl, branched alkyl, bonyl, ethoxycarbonyl, propoxycarbonyl, butoxycarbonyl, cycloalkyl, aryl, heteroaryl or heterocycle. Substituents on pentoxycarbonyl, etc. amide groups may be further Substituted. The term “thioalkyl includes compounds or moieties Compounds of the present invention that contain nitrogens which contain an alkyl group connected with a Sulfur atom. can be converted to N-oxides by treatment with an oxidizing The thioalkyl groups can be substituted with groups such as 15 agent (e.g., 3-chloroperoxybenzoic acid (m-CPBA) and/or alkyl, alkenyl, alkynyl, halogen, hydroxyl, alkylcarbonyloxy, hydrogen peroxides) to afford other compounds of the present arylcarbonyloxy, alkoxycarbonyloxy, aryl oxycarbonyloxy, invention. Thus, all shown and claimed nitrogen-containing carboxylate, carboxyacid, alkylcarbonyl, arylcarbonyl, compounds are considered, when allowed by Valency and alkoxycarbonyl, aminocarbonyl, alkylaminocarbonyl, structure, to include both the compound as shown and its dialkylaminocarbonyl, alkylthiocarbonyl, alkoxyl, amino N-oxide derivative (which can be designated as N->O or (including alkylamino, dialkylamino, arylamino, diary N" O). Furthermore, in other instances, the nitrogens in lamino and alkylarylamino), acylamino (including alkylcar the compounds of the present invention can be converted to bonylamino, arylcarbonylamino, carbamoyl and ureido), N-hydroxy or N-alkoxy compounds. For example, N-hy amidino, imino, Sulfhydryl, alkylthio, arylthio, thiocarboxy droxy compounds can be prepared by oxidation of the parent late, Sulfates, alkylsulfinyl, Sulfonato, Sulfamoyl, Sulfona 25 amine by an oxidizing agent such as m-CPBA. All shown and mido, nitro, trifluoromethyl, cyano, azido, heterocyclyl, alky claimed nitrogen-containing compounds are also considered, laryl, or an aromatic or heteroaromatic moieties. when allowed by valency and structure, to cover both the The term “thiocarbonyl or “thiocarboxy' includes com compound as shown and its N-hydroxy (i.e., N—OH) and pounds and moieties which contain a carbon connected with N-alkoxy (i.e., N—OR, wherein R is substituted or unsubsti a double bond to a sulfur atom. 30 tuted C-C alkyl, C-C alkenyl, C-C alkynyl, 3-14-mem The term “thioether' includes moieties which contain a bered carbocycle or 3-14-membered heterocycle) derivatives. sulfur atom bonded to two carbon atoms or heteroatoms. In the present specification, the structural formula of the Examples of thioethers include, but are not limited to alkthio compound represents a certain isomer for convenience in alkyls, alkthioalkenyls and alkthioalkynyls. The term “alk Some cases, but the present invention includes all isomers, thioalkyls' include moieties with an alkyl, alkenyl or alkynyl 35 Such as geometrical isomers, optical isomers based on an group bonded to a sulfur atom which is bonded to an alkyl asymmetrical carbon, Stereoisomers, tautomers, and the like. group. Similarly, the term “alkthioalkenyls' refers to moi In addition, a crystal polymorphism may be present for the eties wherein an alkyl, alkenyl or alkynyl group is bonded to compounds represented by the formula. It is noted that any a Sulfur atom which is covalently bonded to an alkenyl group; crystal form, crystal form mixture, or anhydride or hydrate and alkthioalkynyls' refers to moieties wherein an alkyl, 40 thereof is included in the scope of the present invention. alkenyl or alkynyl group is bonded to a Sulfur atom which is Furthermore, so-called metabolite which is produced by deg covalently bonded to an alkynyl group. radation of the present compound in vivo is included in the As used herein, “amine' or "amino” includes moieties Scope of the present invention. where a nitrogen atom is covalently bonded to at least one “Isomerism” means compounds that have identical carbon or heteroatom. Alkylamino' includes groups of com 45 molecular formulae but differ in the sequence of bonding of pounds wherein nitrogen is bound to at least one alkyl group. their atoms or in the arrangement of their atoms in space. Examples of alkylamino groups include benzylamino, Isomers that differ in the arrangement of their atoms in space methylamino, ethylamino, phenethylamino, etc. are termed “stereoisomers'. Stereoisomers that are not mirror “Dialkylamino” includes groups wherein the nitrogenatom is images of one another are termed "diastereoisomers', and bound to at least two additional alkyl groups. Examples of 50 Stereoisomers that are non-Superimposable mirror images of dialkylamino groups include, but are not limited to, dimethy each other are termed “enantiomers' or sometimes optical lamino and diethylamino. "Arylamino” and “diarylamino” isomers. A mixture containing equal amounts of individual include groups wherein the nitrogenis bound to at least one or enantiomeric forms of opposite chirality is termed a “racemic two aryl groups, respectively. “Alkylarylamino', 'alkylami mixture'. noaryl' or “arylaminoalkyl refers to an amino group which 55 A carbonatom bonded to four nonidentical substituents is is bound to at least one alkyl group and at least one aryl group. termed a "chiral center'. Alkaminoalkyl refers to an alkyl, alkenyl, or alkynyl group “Chiral isomer means a compound with at least one chiral bound to a nitrogen atom which is also bound to an alkyl center. Compounds with more than one chiral center may group. Acylamino' includes groups wherein nitrogen is exist either as an individual diastereomer or as a mixture of bound to an acyl group. Examples of acylamino include, but 60 diastereomers, termed "diastereomeric mixture'. When one are not limited to, alkylcarbonylamino, arylcarbonylamino, chiral center is present, a stereoisomer may be characterized carbamoyl and ureido groups. by the absolute configuration (R or S) of that chiral center. The term "amide' or "aminocarboxy” includes compounds Absolute configuration refers to the arrangement in space of or moieties that contain a nitrogen atom that is bound to the the substituents attached to the chiral center. The substituents carbon of a carbonyl or a thiocarbonyl group. The term 65 attached to the chiral center under consideration are ranked in includes "alkaminocarboxy' groups that include alkyl, alk accordance with the Sequence Rule of Cahn, Ingold and Pre enyl or alkynyl groups bound to an amino group which is log. (Cahn et al., Angew. Chem. Inter. Edit. 1966, 5, 385; US 8,962,619 B2 43 44 errata 511; Cahn et al., Angew. Chem. 1966, 78, 413: Cahn hydrates include monohydrates, dihydrates, etc. Nonlimiting and Ingold, J. Chem. Soc. 1951 (London), 612; Cahn et al., examples of Solvates include ethanol Solvates, acetone sol Experientia 1956, 12, 81; Cahn, J. Chem. Educ. 1964, 41, Vates, etc. 116). “Solvate” means solvent addition forms that contain either “Geometric isomer means the diastereomers that owe Stoichiometric or non Stoichiometric amounts of solvent. their existence to hindered rotation about double bonds. Some compounds have a tendency to trap a fixed molar ratio These configurations are differentiated in their names by the of solvent molecules in the crystalline solid state, thus form prefixes cis and trans, or Z and E, which indicate that the ing a solvate. If the solvent is water the solvate formed is a groups are on the same or opposite side of the double bond in hydrate; and if the solvent is alcohol, the solvate formed is an the molecule according to the Cahn-Ingold-Prelog rules. 10 alcoholate. Hydrates are formed by the combination of one or Furthermore, the structures and other compounds dis more molecules of water with one molecule of the substance cussed in this invention include all atropic isomers thereof. in which the water retains its molecular state as H.O. Atropic isomers' are a type of stereoisomer in which the As used herein, the term “analog refers to a chemical atoms of two isomers are arranged differently in space. Atro 15 compound that is structurally similar to another but differs pic isomers owe their existence to a restricted rotation caused slightly in composition (as in the replacement of one atom by by hindrance of rotation of large groups about a central bond. an atom of a different element or in the presence of a particu Such atropic isomers typically exist as a mixture, however as lar functional group, or the replacement of one functional a result of recent advances in chromatography techniques; it group by another functional group). Thus, an analog is a has been possible to separate mixtures of two atropic isomers compound that is similar or comparable in function and in select cases. appearance, but not in structure or origin to the reference "Tautomer' is one of two or more structural isomers that compound. exist in equilibrium and is readily converted from one iso As defined herein, the term "derivative' refers to com meric form to another. This conversion results in the formal pounds that have a common core structure, and are substituted migration of a hydrogen atom accompanied by a Switch of 25 with various groups as described herein. For example, all of adjacent conjugated double bonds. Tautomers exist as a mix the compounds represented by formula I are imidazopyridi ture of a tautomeric set in solution. In solid form, usually one nyl-aminopyridine derivatives, and have formula I as a com tautomer predominates. In solutions where tautomerization is O CO. possible, a chemical equilibrium of the tautomers will be The term “bioisostere” refers to a compound resulting from reached. The exact ratio of the tautomers depends on several 30 the exchange of an atom or of a group of atoms with another, factors, including temperature, solvent and pH. The concept broadly similar, atom or group of atoms. The objective of a of tautomers that are interconvertable by tautomerizations is bioisosteric replacement is to create a new compound with called tautomerism. similar biological properties to the parent compound. The Of the various types of tautomerism that are possible, two 35 bioisosteric replacement may be physicochemically or topo are commonly observed. In keto-enol tautomerism a simul logically based. Examples of carboxylic acid bioisosteres taneous shift of electrons and a hydrogen atom occurs. Ring include, but are not limited to, acyl Sulfonimides, tetrazoles, chain tautomerism arises as a result of the aldehyde group Sulfonates and phosphonates. See, e.g., Patani and LaVoie, (—CHO) in a Sugar chain molecule reacting with one of the Chem. Rev. 96, 3147-3176, 1996. hydroxy groups (-OH) in the same molecule to give it a 40 The present invention is intended to include all isotopes of cyclic (ring-shaped) form as exhibited by glucose. atoms occurring in the present compounds. Isotopes include Common tautomeric pairs are: ketone-enol, amide-nitrile, those atoms having the same atomic number but different lactam-lactim, amide-imidic acid tautomerism in heterocy mass numbers. By way of general example and without limi clic rings (e.g., in nucleobases such as guanine, thymine and tation, isotopes of hydrogen include tritium and deuterium, cytosine), amine-enamine and enamine-enamine. 45 and isotopes of carbon include C-13 and C-14. It is to be understood that the compounds of the present invention may be depicted as different tautomers. It should 2. Synthesis of Substituted also be understood that when compounds have tautomeric Imidazopyridinyl-Aminopyridine Compounds forms, all tautomeric forms are intended to be included in the Scope of the present invention, and the naming of the com 50 The present invention provides methods for the synthesis pounds does not exclude any tautomer form. of the compounds of each of the formulae described herein. The term “crystal polymorphs”, “polymorphs' or “crystal The present invention also provides detailed methods for the forms' means crystal structures in which a compound (or a synthesis of various disclosed compounds of the present salt or solvate thereof) can crystallize in different crystal invention according to the following schemes as shown in the packing arrangements, all of which have the same elemental 55 examples. composition. Different crystal forms usually have different X-ray diffraction patterns, infrared spectral, melting points, Throughout the description, where compositions are density hardness, crystal shape, optical and electrical proper described as having, including, or comprising specific com ties, stability and solubility. Recrystallization solvent, rate of ponents, it is contemplated that compositions also consist crystallization, storage temperature, and other factors may 60 essentially of or consist of the recited components. Simi cause one crystal form to dominate. Crystal polymorphs of larly, where methods or processes are described as having, the compounds can be prepared by crystallization under dif including, or comprising specific process steps, the processes ferent conditions. also consistessentially of, or consist of the recited processing Additionally, the compounds of the present invention, for steps. Further, it should be understood that the order of steps example, the salts of the compounds, can exist in either 65 or order for performing certain actions is immaterial so long hydrated or unhydrated (the anhydrous) form or as solvates as the invention remains operable. Moreover, two or more with other solvent molecules. Nonlimiting examples of steps or actions can be conducted simultaneously. US 8,962,619 B2 45 The synthetic processes of the invention can tolerate a wide -continued variety of functional groups, therefore various substituted O starting materials can be used. The processes generally pro 2 vide the desired final compound at or near the end of the NH2 overall process, although it may be desirable in certain N NO N instances to further convert the compound to a pharmaceuti cally acceptable salt, ester or prodrug thereof. 2 2N RX N NH Compounds of the present invention can be prepared in a 4 variety of ways using commercially available starting mate 10 Na2SO4 (85%) rials, compounds known in the literature, or from readily DMSO, MeOH prepared intermediates, by employing standard synthetic 100° C. methods and procedures either known to those skilled in the Step 2 H art, or which will be apparent to the skilled artisan in light of Boc-N the teachings herein. Standard synthetic methods and proce 15 dures for the preparation of organic molecules and functional group transformations and manipulations can be obtained 3 from the relevant scientific literature or from standard text HN books in the field. Although not limited to any one or several N sources, classic texts such as Smith, M.B., March, J., March's N N Advanced Organic Chemistry. Reactions, Mechanisms, and 2 N \ / Structure, 5' edition, John Wiley & Sons: New York, 2001; RX N and Greene, T. W. Wuts, P. G. M., Protective Groups in Organic Synthesis, 3" edition, John Wiley & Sons: NewYork, 1999, incorporated by reference herein, are useful and recog 25 nized reference textbooks of organic synthesis known to those in the art. The following descriptions of synthetic meth Boc-N ods are designed to illustrate, but not to limit, general proce H dures for the preparation of compounds of the present inven 30 tion. 5 Compounds of the present invention can be conveniently NO N RARNNHori RoOH prepared by a variety of methods familiar to those skilled in or RSSH the art. The compounds each of the formulae described herein 2 Her may be prepared according to the following procedures from 35 C N NH Basic condition commercially available starting materials or starting materi OC.-120° C. als which can be prepared using literature procedures. These procedures show the preparation of representative com Step 1-1 pounds of this invention. 40 All the abbreviations used in this application are found in Boc-N “Protective Groups in Organic Synthesis” by John Wiley & H Sons, Inc, or the MERCKINDEX by MERCK& Co., Inc, or other chemistry books or chemicals catalogs by chemicals 3a Vendor Such as Aldrich, or according to usage know in the art. 45 NO General Procedure A N One general procedure for R2-amino-Substituted imida 2 Zopyridine formation is described below in Scheme 1. Rx N NH

50 Scheme 1: Imidazopyridine formation and N, S, OSubstitution on pyridine

Boc-N 55 H

3b

Boc-N Rx =X or NO 60 2 -e- NRR THF Rx = RR'N - or RoO - or RsS C DIEA 1 -15°C.-25° C. 65 Step 1 Step 1. Synthesis of 3-nitro-N-phenylpyridin-2-amine (structure 3 as shown in Scheme 1). 2-chloro-3-nitropyridine US 8,962,619 B2 47 48 (structure 1 as shown in Scheme 1) was dissolved in THF (10 -continued mL/mmol) in a round bottom flask. Aniline (1.0 eq.) (struc ture 2 as shown in Scheme 1) and diisopropylethylamine (1.05 eq.) were added. The reaction mixture was heated to the appropriate temperature for 4 to 36 hours. After cooling to n-y room temperature the solvent was removed under reduced RX 4N pressure. The residue was dissolved in ethyl acetate (20 mL/mmol) and washed with water and brine (20 mL/mmol respectively). The organic phase was separated and dried over NaSO. After filtration the solvent was removed under 10 reduced pressure. The crude product (red to brown solid) was solidified with hexane/ethyl acetate and collected by filtra NH, HCI tion. The product was carried on to the next step without further purification. Step 1-1. Synthesis of N/O'/S'-alkyl/aryl-3-nitro-N'- 15 phenylpyridine-2,6-diamine (structure 3b as shown in Scheme 1). Intermediate (structure 3a as shown in Scheme 1) (1 eq.) was dissolved in dioxane (5 mL/mmol) in a round bottom flask. Alky/aryl amine/sulfur/alcohol (2 eq.) was added and diisopropylamine (2.5 eq.). The reaction mixture NRR was heated to 80°C. in an oil bath for 24 h. After cooling to Carbamate (structure 5 as shown in Scheme 2) (1 eq.) was room temperature the solvent was removed under reduced dissolved in methanol. HCl (20 eq., 4 M in dioxane) was pressure. The residue was dissolved in ethyl acetate (10 added and stirred at room temperature for 2 to 4 hours. Con mL/mmol) and washed with water and brine (5 mL/mmol 25 centration under reduced pressure and solidified with ethyl respectively). The organic phase was separated and dried over acetate collection by filtration gave the deprotected amine NaSO. After filtration the solvent was removed under (structure 6 as shown in Scheme 2) as hydrochloric acid salt reduced pressure. The crude product (structure3b as shown in as a powder, which was used for the next step without further Scheme 1) was carried on to the next step without further purification. purification. 30 General Procedure C Step 2. Synthesis of 3-(3-phenyl-3H-imidazo[4,5-b]pyri One general procedure for amide formation is described din-2-yl)pyridin-2-amine (structure 5 as shown in Scheme 1). below in Scheme 3. 3-Nitro-N-phenylpyridin-2-amine (structure 3 as shown in Scheme 1) was dissolved in dimethylsulfoxide (8 mL/mmol) and methanol (1.5 mL/mmol) in a round bottom flask. 2-ami 35 Scheme 3: Amido formation nonicotinaldehyde 4 (1.1 eq.) and NaSO (85%, 2.5 eq.) HN were added. The reaction mixture was heated at 100°C. for 15 =N to 36 hours. After cooling to room temperature the reaction O n v RphNH2, EDC1, as N N / DIEA mixture was diluted with dichloromethane (20 mL/mmol) -- and washed with water and brine. The organic phase was 40 HO N DMF separated and dried over NaSO. After filtration the solvent was removed under reduced pressure. The crude product was purified by Silica gel chromatography (dichloromethane/ methanol: 0-20% methanol over 60 min) to give a yellow to NHBoc brown solid. 45 General Procedure B 7 One general procedure for BOC group deprotection is HN described below in Scheme 2. O N N SN 50 2 N \ / Scheme 2: Deproctection of BOC-group RphHN N HN

Sl-N N 55

RX N elN/ \ / HCl, dioxane NHBoc Hip methanol 60 8 Rph is as described above

NHBoc To a mixture of amine (structure 7 as shown in Scheme 3) 65 (1 eq.) and aryl/alkyl amine (1.1 eq.) in dimethylformamide 5 (5 mL/mmol) was added 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (1.2 eq.) and diisopropylethylamine (3 eq.). US 8,962,619 B2 49 50 The reaction mixture was stirred for 15 hours at room tem was separated, washed with brine (5 mL/mmol) and dried perature then quenched with water (2 mL/mmol). The mix over NaSO. After filtration the solvent was removed under ture was diluted with ethyl acetate and the organic phase was reduced pressure. The crude product (structure 10 as shown in separated, washed with brine (5 mL/mmol) and dried over Scheme 4) was purified by silica gel chromatography NaSO. After filtration the solvent was removed under (0-100% ethyl acetate in hexanes). reduced pressure. The crude product (structure 8 as shown in General Procedure E Scheme 3) was purified by silica gel chromatography (hex One general procedure for saponification is described anes/ethyl acetate/methanol: 100/0/0 to 0/90/10). below in Scheme 5. General Procedure D 10 One general procedure for Sulfonamide formation is described below in Scheme 4. Scheme 5: Saponification HN Scheme 4: Sulfonamide formation 15 Sl-N N N NaOH, H2O HN % N \ / HeTHF, MeOH N N SN N

RSOCl, O O DIEA R co, \ / -e- N1 N DCE 25 H R N^ 11 H

30

HN 35 S-N N elN/N \ /

40

O OH N1 R 45 H

12 R = protecting group such as BOC

50 N Ethyl 2-(4-(2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b] pyridine-3-yl)phenyl) acetate (structure 11 as shown in R.'-N-SEO Scheme 5) (obtained by General Procedure A) was dissolved O R7 55 in tetrahydrofuran (2 mL/mmol) and methanol (0.7 10 mL/mmol) in around bottom flask. NaOHHO (5 eq.) in HO R = H or protecting group such as BOC (0.5 mL/mmol) was added. The reaction mixture was carried R7 and R7 is as described above out at room temperature for 18 hours. The organic solvent was removed under reduced pressure. The water phase was neu 60 tralized with HCl (5 eq) in HO (0.5 mL/mmol) and the solid was collected with filtration to yield product (structure 12 as shown in Scheme 5). The product 12 was carried on to the To a mixture of amine (structure 9 as shown in Scheme 4) next step without further purification. (1 eq.) and sulfonyl chloride (1.1 eq.) in dichloromethane (5 General Procedure F mL/mmol) was added diisopropylethylamine (3 eq.). The 65 reaction mixture was stirred for 15 hours at room temperature A general procedure for the Suzuki coupling reaction is then quenched with water (2 mL/mmol). The organic phase described below in Scheme 6 and Scheme 7. US 8,962,619 B2 52 -continued Scheme 6: Suzuki coupling 1 HN NO N Sl-NN N 2 C N NH Rt N elN/ \ A RB(OH)2 or pinacol ester, Pd(PPh3)4 His toluen?ethanolisat.NaHCO3 80 to 120 degree Cercius 10

Boc-N Boc-N H H 15 14 3a Rt = meta substituted phenyl

NO Organohalide (5a as shown in Scheme 7) (1 eq.), NaCO, N (1.2 eq.), AMPHOS (bis(di-tert-butyl (4-dimethylaminophe 2 nyl)phosphine)dichloropalladium(II)) (0.1 eq.) and aryl Rt N NH boronic acid (2 eq.) were dissolved in DMF/water. The reac tion mixture was heated in the microwave for 60 minto 160° 25 C. The solvent was removed under reduced pressure and the crude product was purified by Silica gel chromatography (hexane/3-20% methanol in ethyl acetate) to yield the product (14 as shown in Scheme 7). Boc-N General Procedure H 30 A general procedure for the boronic acid pinacol ester 13 formation is described below in Scheme 8. Rt = meta substituted phenyl

35 Scheme 8: boronic acid pinacol ester formation. Organohalide (structure 3a as shown in Scheme 6) (1 eq.), Step 1 boronic acid or pinacol ester (2 eq.), and Pd(PPh) (tetrakis (triphenylphosphine)palladium(0)) (0.05 eq) were suspended HNRR Br r Pddba in a mixture of ethanol and toluene, 10 mL/mmol respec rac-BINAP, CsCO3 40 -e- tively. A solution of saturated NaHCO was added (1 toluene mL/mmol). The reaction mixture was degassed with nitrogen O.4M for 30 min. Subsequently it was heated to 100° C. overnight TL 64, 2938(2008). under nitrogen. After cooling down to room temperature the 15 solvent was removed in vacuo. The crude residue was purified Step 2 45 KOAc by silica gel chromatography (hexane/3-20% methanol in PdCldppf ethyl acetate) to give product (structure 13 as shown in Br NRR' pinacol diboron Scheme 6). -e- General Procedure G 50 16 Scheme 7: Suzuki coupling 2 HN RtB(OH)2 or pinacol R Sl-N o ester, AMPHOS, 55 B NRR' N Na2CO3 C N 2NN \ / DMF, Water Microwave oven 120 to 160° C. for 17 15 minto 60 min 60 KOAc PdCl2dppf pinacol diboron Boc-N -- H Step 1.1 65 5a 18 US 8,962,619 B2 53 54 -continued bird, mouse, rat, fowl, dog, cat, cow, horse, goat, camel, sheep or a pig. Preferably, the mammal is a human. As used herein, the term “cell proliferative disorder” refers to conditions in which unregulated or abnormal growth, or both, of cells can lead to the development of an unwanted condition or disease, which may or may not be cancerous. Exemplary cell proliferative disorders of the invention encompass a variety of conditions wherein cell division is R1, R1 and Rph is as described above deregulated. Exemplary cell proliferative disorder include, 10 but are not limited to, neoplasms, benign tumors, malignant tumors, pre-cancerous conditions, in situ tumors, encapsu Step 1: 1,3-dibromobenzene (1 eq.), Pd dba (tris(diben lated tumors, metastatic tumors, liquid tumors, Solid tumors, Zylideneacetone)dipalladium(0)) (5 mol%), rac-BINAP (7.5 immunological tumors, hematological tumors, cancers, car mol %), cesium carbonate (3 eq.) and amine (1 eq.) were cinomas, leukemias, lymphomas, sarcomas, and rapidly dissolved in toluene (2.5 mL/mmol). The reaction mixture 15 dividing cells. The term “rapidly dividing cell as used herein was stirred for 5 hr at 80°C. The mixture was filtered through is defined as any cell that divides at a rate that exceeds or is celite pad and the solvent was removed under vacuum. The greater than what is expected or observed among neighboring residue was purified by silica gel column chromatography or juxtaposed cells within the same tissue. A cell proliferative (hexane/ethyl acetate, 10:0 to 9:1) to yield product as a pale disorder includes a precancer or a precancerous condition. A brown oil (16 as shown in Scheme 8). Step 2: Amino-3- cell proliferative disorder includes cancer. Preferably, the bromobenzene (1 eq.), potassium acetate (3 eq.), PdCldppf methods provided herein are used to treat oralleviate a symp (1,1'-bis(diphenylphosphino) ferrocenedichloropalladium tom of cancer. The term "cancer includes solid tumors, as (II) complex with dichloromethane) (4 mol %), and pinacol well as, hematologic tumors and/or malignancies. A "precan diboron (1.5 eq.) were dissolved in dimethyl formamide cer cell' or “precancerous cell' is a cell manifesting a cell (DMF) (5 mL/mmol). The mixture was heated at 80° C. for 6 25 proliferative disorder that is a precancer or a precancerous h and diluted with ethyl acetate and mixed with water. The condition. A "cancer cell' or “cancerous cell' is a cell mani organic layer was separated, dried over anhydrous Na2SO4. festing a cell proliferative disorder that is a cancer. Any repro and evaporated under reduced pressure. The residue was puri ducible means of measurement may be used to identify can fied by silica gel column chromatography (hexane/ethyl cer cells or precancerous cells. Cancer cells or precancerous acetate, 10:0 to 0:10) to yield product as a pale brown solid 30 cells can be identified by histological typing or grading of a (structure 17 as shown in Scheme 8). Step 1.1: The compound tissue sample (e.g., a biopsy sample). Cancer cells or precan 19 was synthesized from 18 by a similar procedure to that cerous cells can be identified through the use of appropriate described above. molecular markers. Exemplary non-cancerous conditions or disorders include, 3. Methods of Treatment 35 but are not limited to, rheumatoid arthritis; inflammation; autoimmune disease; lymphoproliferative conditions; The present invention provides methods for the treatment acromegaly; rheumatoid spondylitis; osteoarthritis; gout, of a cell proliferative disorder in a subject in need thereof by other arthritic conditions; sepsis; septic shock; endotoxic administering to a Subject in need of such treatment, a thera shock, gram-negative sepsis: toxic shock syndrome; asthma, peutically effective amount of a compound of the present 40 adult respiratory distress syndrome; chronic obstructive pull invention, or a pharmaceutically acceptable salt, prodrug, monary disease; chronic pulmonary inflammation; inflam metabolite, polymorph or solvate thereof. The cell prolifera matory bowel disease; Crohn's disease; psoriasis: eczema: tive disorder can be cancer or a precancerous condition. The ulcerative colitis; pancreatic fibrosis; hepatic fibrosis: acute present invention further provides the use of a compound of and chronic renal disease; irritable bowel syndrome; pyresis; the present invention, or a pharmaceutically acceptable salt, 45 restenosis; cerebral malaria; stroke and ischemic injury; neu prodrug, metabolite, polymorph or solvate thereof, for the ral trauma; Alzheimer's disease; Huntington's disease; Par preparation of a medicament useful for the treatment of a cell kinson's disease; acute and chronic pain; allergic rhinitis; proliferative disorder. allergic conjunctivitis; chronic heart failure; acute coronary The present invention also provides methods of protecting syndrome; cachexia; malaria, leprosy; leishmaniasis; Lyme against a cell proliferative disorder in a subject in need thereof 50 disease; Reiter's syndrome; acute synovitis; muscle degen by administering a therapeutically effective amount of com eration, bursitis; tendonitis; tenosynovitis; herniated, rup pound of the present invention, or a pharmaceutically accept tures, or prolapsed intervertebral disk syndrome; osteopetro able salt, prodrug, metabolite, polymorph or Solvate thereof, sis; thrombosis; restenosis; silicosis; pulmonary sarcosis: to a subject in need of such treatment. The cell proliferative bone resorption diseases, such as osteoporosis; graft-Versus disorder can be cancer or a precancerous condition. The 55 host reaction; Multiple Sclerosis; lupus; fibromyalgia: AIDS present invention also provides the use of compound of the and other viral diseases such as Herpes Zoster, Herpes Sim present invention, or a pharmaceutically acceptable salt, pro plex I or II, influenza virus and cytomegalovirus; and diabetes drug, metabolite, polymorph or Solvate thereof, for the prepa mellitus. ration of a medicament useful for the prevention of a cell Exemplary cancers include, but are not limited to, adreno proliferative disorder. 60 cortical carcinoma, AIDS-related cancers, AIDS-relatedlym As used herein, a “subject in need thereof is a subject phoma, anal cancer, anorectal cancer, cancer of the anal canal, having a cell proliferative disorder, or a Subject having an appendix cancer, childhood cerebellar astrocytoma, child increased risk of developing a cell proliferative disorder rela hood cerebral astrocytoma, basal cell carcinoma, skin cancer tive to the population at large. A subject in need thereof can (non-melanoma), biliary cancer, extrahepatic bile duct can have a precancerous condition. Preferably, a Subject in need 65 cer, intrahepatic bile duct cancer, bladder cancer, urinary thereof has cancer. A “subject' includes a mammal. The bladder cancer, bone and joint cancer, osteosarcoma and mammal can be e.g., any mammal, e.g., a human, primate, malignant fibrous histiocytoma, brain cancer, brain tumor, US 8,962,619 B2 55 56 brain stem glioma, cerebellar astrocytoma, cerebral astrocy system can include hyperplasia, dysplasia, and metaplasia of toma/malignant glioma, ependymoma, medulloblastoma, cells of the hematologic system. Preferably, compositions of Supratentorial primitive neuroectodeimal tumors, visual the present invention may be used to treat a cancer selected pathway and hypothalamic glioma, breast cancer, bronchial from the group consisting of a hematologic cancer of the adenomas/carcinoids, carcinoid tumor, gastrointestinal, ner present invention or a hematologic cell proliferative disorder Vous system cancer, nervous system lymphoma, central ner of the present invention. A hematologic cancer of the present Vous system cancer, central nervous system lymphoma, cer invention can include multiple myeloma, lymphoma (includ Vical cancer, childhood cancers, chronic lymphocytic ing Hodgkin’s lymphoma, non-Hodgkin’s lymphoma, child leukemia, chronic myelogenous leukemia, chronic myelo hood lymphomas, and lymphomas of lymphocytic and cuta proliferative disorders, colon cancer, colorectal cancer, cuta 10 neous origin), leukemia (including childhood leukemia, neous T-cell lymphoma, lymphoid neoplasm, mycosis fun hairy-cell leukemia, acute lymphocytic leukemia, acute goides, Seziary Syndrome, endometrial cancer, esophageal myelocytic leukemia, chronic lymphocytic leukemia, chronic cancer, extracranial germ cell tumor, extragonadal germ cell myelocytic leukemia, chronic myelogenous leukemia, and tumor, extrahepatic bile duct cancer, eye cancer, intraocular mast cell leukemia), myeloid neoplasms and mast cell neo melanoma, retinoblastoma, gallbladder cancer, gastric (stom 15 plasms. ach) cancer, gastrointestinal carcinoid tumor, gastrointestinal A “cell proliferative disorder of the lung is a cell prolif stromal tumor (GIST), germ cell tumor, ovarian germ cell erative disorder involving cells of the lung. Cell proliferative tumor, gestational trophoblastic tumor glioma, head and neck disorders of the lung can include all forms of cell proliferative cancer, hepatocellular (liver) cancer, Hodgkin lymphoma, disorders affecting lung cells. Cell proliferative disorders of hypopharyngeal cancer, intraocular melanoma, ocular can the lung can include lung cancer, a precancer or precancerous cer, islet cell tumors (endocrine pancreas), Kaposi Sarcoma, condition of the lung, benign growths or lesions of the lung, kidney cancer, renal cancer, kidney cancer, laryngeal cancer, and malignant growths or lesions of the lung, and metastatic acute lymphoblastic leukemia, acute myeloid leukemia, lesions in tissue and organs in the body other than the lung. chronic lymphocytic leukemia, chronic myelogenous leuke Preferably, compositions of the present invention may be mia, hairy cell leukemia, lip and oral cavity cancer, liver 25 used to treat lung cancer or cell proliferative disorders of the cancer, lung cancer, non-small cell lung cancer, Small cell lung. Lung cancer can include all forms of cancer of the lung. lung cancer, AIDS-related lymphoma, non-Hodgkin lym Lung cancer can include malignant lung neoplasms, carci phoma, primary central nervous system lymphoma, Walden noma in situ, typical carcinoid tumors, and atypical carcinoid stram macroglobulinemia, medulloblastoma, melanoma, tumors. Lung cancer can include Small cell lung cancer intraocular (eye) melanoma, merkel cell carcinoma, mesothe 30 ('SCLC), non-small cell lung cancer (“NSCLC), squa lioma malignant, mesothelioma, metastatic squamous neck mous cell carcinoma, adenocarcinoma, Small cell carcinoma, cancer, mouth cancer, cancer of the tongue, multiple endo large cell carcinoma, adenosquamous cell carcinoma, and crine neoplasia syndrome, mycosis fungoides, myelodys mesothelioma. Lung cancer can include “scar carcinoma’. plastic syndromes, myelodysplastic/myeloproliferative dis bronchioalveolar carcinoma, giant cell carcinoma, spindle eases, chronic myelogenous leukemia, acute myeloid 35 cell carcinoma, and large cell neuroendocrine carcinoma. leukemia, multiple myeloma, chronic myeloproliferative dis Lung cancer can include lung neoplasms having histologic orders, nasopharyngeal cancer, neuroblastoma, oral cancer, and ultrastructual heterogeneity (e.g., mixed cell types). oral cavity cancer, oropharyngeal cancer, ovarian cancer, ova Cell proliferative disorders of the lung can include all rian epithelial cancer, ovarian low malignant potential tumor, forms of cell proliferative disorders affecting lung cells. Cell pancreatic cancer, islet cell pancreatic cancer, paranasal sinus 40 proliferative disorders of the lung can include lung cancer, and nasal cavity cancer, parathyroid cancer, penile cancer, precancerous conditions of the lung. Cell proliferative disor pharyngeal cancer, pheochromocytoma, pineoblastoma and ders of the lung can include hyperplasia, metaplasia, and Supratentorial primitive neuroectodermal tumors, pituitary dysplasia of the lung. Cell proliferative disorders of the lung tumor, plasma cell neoplasm/multiple myeloma, pleuropul can include asbestos-induced hyperplasia, squamous meta monary blastoma, prostate cancer, rectal cancer, renal pelvis 45 plasia, and benign reactive mesothelial metaplasia. Cell pro and ureter, transitional cell cancer, retinoblastoma, rhab liferative disorders of the lung can include replacement of domyosarcoma, Salivary gland cancer, ewing family of sar columnar epithelium with stratified squamous epithelium, coma tumors, Kaposi Sarcoma, soft tissue sarcoma, uterine and mucosal dysplasia. Individuals exposed to inhaled inju cancer, uterine sarcoma, skin cancer (non-melanoma), skin rious environmental agents such as cigarette Smoke and cancer (melanoma), merkel cell skin carcinoma, Small intes 50 asbestos may be at increased risk for developing cell prolif tine cancer, soft tissue sarcoma, squamous cell carcinoma, erative disorders of the lung. Prior lung diseases that may stomach (gastric) cancer, Supratentorial primitive neuroecto predispose individuals to development of cell proliferative dermal tumors, testicular cancer, throat cancer, thymoma, disorders of the lung can include chronic interstitial lung thymoma and thymic carcinoma, thyroid cancer, transitional disease, necrotizing pulmonary disease, Scleroderma, rheu cell cancer of the renal pelvis and ureter and other urinary 55 matoid disease, sarcoidosis, interstitial pneumonitis, tubercu organs, gestational trophoblastic tumor, urethral cancer, losis, repeated pneumonias, idiopathic pulmonary fibrosis, endometrial uterine cancer, uterine sarcoma, uterine corpus granulomata, asbestosis, fibrosing alveolitis, and Hodgkin’s cancer, vaginal cancer, Vulvar cancer, and Wilm's Tumor. disease. A “cell proliferative disorder of the hematologic system is A “cell proliferative disorder of the colon' is a cell prolif a cell proliferative disorder involving cells of the hematologic 60 erative disorder involving cells of the colon. Preferably, the system. A cell proliferative disorder of the hematologic sys cell proliferative disorder of the colon is colon cancer. Pref tem can include lymphoma, leukemia, myeloid neoplasms, erably, compositions of the present invention may be used to mast cell neoplasms, myelodysplasia, benign monoclonal treat colon cancer or cell proliferative disorders of the colon. gammopathy, lymphomatoid granulomatosis, lymphomatoid Colon cancer can include all forms of cancer of the colon. papulosis, polycythemia Vera, chronic myelocytic leukemia, 65 Colon cancer can include sporadic and hereditary colon can agnogenic myeloid metaplasia, and essential thromb cers. Colon cancer can include malignant colon neoplasms, ocythemia. A cell proliferative disorder of the hematologic carcinoma in situ, typical carcinoid tumors, and atypical car US 8,962,619 B2 57 58 cinoid tumors. Colon cancer can include adenocarcinoma, the skin, benign growths or lesions of the skin, melanoma, squamous cell carcinoma, and adenosquamous cell carci malignant melanoma and other malignant growths or lesions noma. Colon cancer can be associated with a hereditary Syn of the skin, and metastatic lesions in tissue and organs in the drome selected from the group consisting of hereditary non body other than the skin. Cell proliferative disorders of the polyposis colorectal cancer, familial adenomatous polyposis, skin can include hyperplasia, metaplasia, and dysplasia of the Gardner's syndrome, Peutz-Jeghers syndrome, Turcot's Syn skin. drome and juvenile polyposis. Colon cancer can be caused by A “cell proliferative disorder of the ovary” is a cell prolif a hereditary syndrome selected from the group consisting of erative disorder involving cells of the ovary. Cell proliferative hereditary nonpolyposis colorectal cancer, familial adenoma disorders of the ovary can include all forms of cell prolifera tous polyposis, Gardner's syndrome, Peutz-Jeghers Syn 10 tive disorders affecting cells of the ovary. Cell proliferative drome, Turcot's syndrome and juvenile polyposis. disorders of the ovary can include a precancer or precancer Cell proliferative disorders of the colon can include all ous condition of the ovary, benign growths or lesions of the forms of cell proliferative disorders affecting colon cells. Cell ovary, ovarian cancer, malignant growths or lesions of the proliferative disorders of the colon can include colon cancer, ovary, and metastatic lesions in tissue and organs in the body precancerous conditions of the colon, adenomatous polyps of 15 other than the ovary. Cell proliferative disorders of the skin the colon and metachronous lesions of the colon. A cell pro can include hyperplasia, metaplasia, and dysplasia of cells of liferative disorder of the colon can include adenoma. Cell the ovary. proliferative disorders of the colon can be characterized by A “cell proliferative disorder of the breast’ is a cell prolif hyperplasia, metaplasia, and dysplasia of the colon. Prior erative disorder involving cells of the breast. Cell prolifera colon diseases that may predispose individuals to develop tive disorders of the breast can include all forms of cell ment of cell proliferative disorders of the colon can include proliferative disorders affecting breast cells. Cell prolifera prior colon cancer. Current disease that may predispose indi tive disorders of the breast can include breast cancer, a pre viduals to development of cell proliferative disorders of the cancer or precancerous condition of the breast, benign colon can include Crohn's disease and ulcerative colitis. A growths or lesions of the breast, and malignant growths or cell proliferative disorder of the colon can be associated with 25 lesions of the breast, and metastatic lesions in tissue and a mutation in a gene selected from the group consisting of organs in the body other than the breast. Cell proliferative p53, ras, FAP and DCC. An individual can have an elevated disorders of the breast can include hyperplasia, metaplasia, risk of developing a cell proliferative disorder of the colon and dysplasia of the breast. due to the presence of a mutation in a gene selected from the A cell proliferative disorder of the breast can be a precan group consisting of p53, ras, FAP and DCC. 30 cerous condition of the breast. Compositions of the present A “cell proliferative disorder of the pancreas” is a cell invention may be used to treat a precancerous condition of the proliferative disorder involving cells of the pancreas. Cell breast. A precancerous condition of the breast can include proliferative disorders of the pancreas can include all forms of atypical hyperplasia of the breast, ductal carcinoma in situ cell proliferative disorders affecting pancreatic cells. Cell (DCIS), intraductal carcinoma, lobular carcinoma in situ proliferative disorders of the pancreas can include pancreas 35 (LCIS), lobular neoplasia, and stage 0 or grade 0 growth or cancer, a precancer or precancerous condition of the pan lesion of the breast (e.g., stage 0 or grade 0 breast cancer, or creas, hyperplasia of the pancreas, and dysaplasia of the pan carcinoma in situ). A precancerous condition of the breast can creas, benign growths or lesions of the pancreas, and malig be staged according to the TNM classification scheme as nant growths or lesions of the pancreas, and metastatic lesions accepted by the American Joint Committee on Cancer in tissue and organs in the body other than the pancreas. 40 (AJCC), where the primary tumor (T) has been assigned a Pancreatic cancer includes all forms of cancer of the pan stage of TO or T is; and where the regional lymph nodes (N) creas. Pancreatic cancer can include ductal adenocarcinoma, have been assigned a stage of N0; and where distant metasta adenosquamous carcinoma, pleomorphic giant cell carci sis (M) has been assigned a stage of M0. noma, mucinous adenocarcinoma, osteoclast-like giant cell The cell proliferative disorder of the breast can be breast carcinoma, mucinous cystadenocarcinoma, acinar carci 45 cancer. Preferably, compositions of the present invention may noma, unclassified large cell carcinoma, Small cell carci be used to treat breast cancer. Breast cancer includes all forms noma, pancreatoblastoma, papillary neoplasm, mucinous of cancer of the breast. Breast cancer can include primary cystadenoma, papillary cystic neoplasm, and serous cystad epithelial breast cancers. Breast cancer can include cancers in enoma. Pancreatic cancer can also include pancreatic neo which the breast is involved by other tumors such as lym plasms having histologic and ultrastructual heterogeneity 50 phoma, sarcoma or melanoma. Breast cancer can include (e.g., mixed cell types). carcinoma of the breast, ductal carcinoma of the breast, lobu A “cell proliferative disorder of the prostate' is a cell lar carcinoma of the breast, undifferentiated carcinoma of the proliferative disorder involving cells of the prostate. Cell breast, cystosarcoma phyllodes of the breast, angiosarcoma proliferative disorders of the prostate can include all forms of of the breast, and primary lymphoma of the breast. Breast cell proliferative disorders affecting prostate cells. Cell pro 55 cancer can include Stage I, II, IIIA, IIIB, IIIC and IV breast liferative disorders of the prostate can include prostate cancer, cancer. Ductal carcinoma of the breast can include invasive a precancer or precancerous condition of the prostate, benign carcinoma, invasive carcinoma in situ with predominant growths or lesions of the prostate, and malignant growths or intraductal component, inflammatory breast cancer, and a lesions of the prostate, and metastatic lesions in tissue and ductal carcinoma of the breast with a histologic type selected organs in the body other than the prostate. Cell proliferative 60 from the group consisting of comedo, mucinous (colloid), disorders of the prostate can include hyperplasia, metaplasia, medullary, medullary with lymphcytic infiltrate, papillary, and dysplasia of the prostate. Scirrhous, and tubular. Lobular carcinoma of the breast can A “cell proliferative disorder of the skin' is a cell prolif include invasive lobular carcinoma with predominant in situ erative disorder involving cells of the skin. Cell proliferative component, invasive lobular carcinoma, and infiltrating lobu disorders of the skin can include all forms of cell proliferative 65 lar carcinoma. Breast cancer can include Paget’s disease, disorders affecting skin cells. Cell proliferative disorders of Paget’s disease with intraductal carcinoma, and Paget’s dis the skin can include a precancer or precancerous condition of ease with invasive ductal carcinoma. Breast cancer can US 8,962,619 B2 59 60 include breast neoplasms having histologic and ultrastructual cancer relative to the population at large. A Subject with an heterogeneity (e.g., mixed cell types). increased risk of developing breast cancer relative to the Preferably, compound of the present invention, or a phar population at large is a female Subject with a family history or maceutically acceptable salt, prodrug, metabolite, polymorph personal history of breast cancer. A subject with an increased or solvate thereof, may be used to treat breast cancer. A breast 5 risk of developing breast cancer relative to the population at cancer that is to be treated can include familial breast cancer. large is a female Subject having a germ-line or spontaneous A breast cancer that is to be treated can include sporadic mutation in BRCA1 or BRCA2, or both. A subject with an breast cancer. A breast cancer that is to be treated can arise in increased risk of developing breast cancer relative to the a male Subject. A breast cancer that is to be treated can arise population at large is a female Subject with a family history of in a female Subject. A breast cancer that is to be treated can 10 breast cancer and a germ-line or spontaneous mutation in arise in a premenopausal female Subject or a postmenopausal BRCA1 or BRCA2, or both. A subject with an increased risk female Subject. A breast cancer that is to be treated can arise of developing breast cancer relative to the population at large in a Subject equal to or older than 30 years old, or a subject is a female who is greater than 30 years old, greater than 40 younger than 30 years old. A breast cancer that is to be treated years old, greater than 50 years old, greater than 60 years old, has arisen in a Subject equal to or older than 50 years old, or 15 greater than 70 years old, greater than 80 years old, or greater a subject younger than 50 years old. A breast cancer that is to than 90 years old. A subject with an increased risk of devel be treated can arise in a subject equal to or older than 70 years oping breast cancer relative to the population at large is a old, or a subject younger than 70 years old. Subject with atypical hyperplasia of the breast, ductal carci A breast cancer that is to be treated can be typed to identify noma in situ (DCIS), intraductal carcinoma, lobular carci a familial or spontaneous mutation in BRCA1, BRCA2, or noma in situ (LCIS), lobular neoplasia, or a stage 0 growth or p53. A breast cancer that is to be treated can be typed as lesion of the breast (e.g., stage 0 or grade 0 breast cancer, or having a HER2/neu gene amplification, as overexpressing carcinoma in situ). HER2/neu, or as having a low, intermediate or high level of A breast cancer that is to be treated can histologically HER2/neu expression. A breast cancer that is to be treated can graded according to the Scarff-Bloom-Richardson system, be typed for a marker selected from the group consisting of 25 wherein a breast tumor has been assigned a mitosis count estrogen receptor (ER), progesterone receptor (PR), human score of 1, 2, or 3; a nuclear pleiomorphism score of 1, 2, or epidermal growth factor receptor-2, Ki-67, CA15-3, CA 3; a tubule formation score of 1, 2, or 3; and a total Scarff 27-29, and c-Met. A breast cancer that is to be treated can be Bloom-Richardson score of between 3 and 9. A breast cancer typed as ER-unknown, ER-rich or ER-poor. A breast cancer that is to be treated can be assigned a tumor grade according that is to be treated can be typed as ER-negative or ER 30 to the International Consensus Panel on the Treatment of positive. ER-typing of a breast cancer may be performed by Breast Cancer selected from the group consisting of grade 1. any reproducible means. ER-typing of a breast cancer may be grade 1-2, grade 2, grade 2-3, or grade 3. performed as set forth in Onkologie 27: 175-179 (2004). A A cancer that is to be treated can be staged according to the breast cancer that is to be treated can be typed as PR-un American Joint Committee on Cancer (AJCC) TNM classi known, PR-rich or PR-poor. A breast cancer that is to be 35 fication system, where the tumor (T) has been assigned a treated can be typed as PR-negative or PR-positive. A breast stage of TX, T1, T1mic, T1a, T1b, T1c, T2, T3, T4, T4a, T4b, cancer that is to be treated can be typed as receptor positive or T4c, or T4d; and where the regional lymph nodes (N) have receptor negative. A breast cancer that is to be treated can be been assigned a stage of NX, NO, N1, N2, N2a, N2b, N3, N3a, typed as being associated with elevated blood levels of CA N3b, or N3c.; and where distant metastasis (M) can be 15-3, or CA 27-29, or both. 40 assigned a stage of MX, MO, or M1. A cancer that is to be A breast cancer that is to be treated can include a localized treated can be staged according to an American Joint Com tumor of the breast. A breast cancer that is to be treated can mittee on Cancer (AJCC) classification as Stage I, Stage IIA, include a tumor of the breast that is associated with a negative Stage IIB, Stage IIIA, Stage IIIB, Stage IIIC, or Stage IV. A sentinel lymph node (SLN) biopsy. A breast cancer that is to cancer that is to be treated can be assigned a grade according be treated can include a tumor of the breast that is associated 45 to an AJCC classification as Grade GX (e.g., grade cannot be with a positive sentinel lymph node (SLN) biopsy. A breast assessed), Grade 1, Grade 2, Grade 3 or Grade 4. A cancer that cancer that is to be treated can include a tumor of the breast is to be treated can be staged according to an AJCC pathologic that is associated with one or more positive axillary lymph classification (pN) of pNX, pN0, PNO (I-), PNO (I+), PNO nodes, where the axillary lymph nodes have been staged by (mol-), PNO (mol+), PN1, PN1(mi), PN1a, PN1b, PN1c, any applicable method. A breast cancer that is to be treated 50 pN2, pN2a, pN2b, pN3, pN3a, pN3b, or pN3c. can includea tumor of the breast that has been typed as having A cancer that is to be treated can include a tumor that has nodal negative status (e.g., node-negative) or nodal positive been determined to be less than or equal to about 2 centime status (e.g., node-positive). A breast cancer that is to be ters in diameter. A cancer that is to be treated can include a treated can include a tumor of the breast that has metastasized tumor that has been determined to be from about 2 to about 5 to other locations in the body. A breast cancer that is to be 55 centimeters in diameter. A cancer that is to be treated can treated can be classified as having metastasized to a location include a tumor that has been determined to be greater than or selected from the group consisting of bone, lung, liver, or equal to about 3 centimeters in diameter. A cancer that is to be brain. A breast cancer that is to be treated can be classified treated can include a tumor that has been determined to be according to a characteristic selected from the group consist greater than 5 centimeters in diameter. A cancer that is to be ing of metastatic, localized, regional, local-regional, locally 60 treated can be classified by microscopic appearance as well advanced, distant, multicentric, bilateral, ipsilateral, con differentiated, moderately differentiated, poorly differenti tralateral, newly diagnosed, recurrent, and inoperable. ated, or undifferentiated. A cancer that is to be treated can be A compound of the present invention, or a pharmaceuti classified by microscopic appearance with respect to mitosis cally acceptable salt, prodrug, metabolite, polymorph or Sol count (e.g., amount of cell division) or nuclear pleiomor vate thereof, may be used to treat or prevent a cell prolifera 65 phism (e.g., change in cells). A cancer that is to be treated can tive disorder of the breast, or to treat or prevent breast cancer, be classified by microscopic appearance as being associated in a Subject having an increased risk of developing breast with areas of necrosis (e.g., areas of dying or degenerating US 8,962,619 B2 61 62 cells). A cancer that is to be treated can be classified as having A compound of the present invention, or a pharmaceuti an abnormal karyotype, having an abnormal number of chro cally acceptable salt, prodrug, metabolite, polymorph or Sol mosomes, or having one or more chromosomes that are vate thereof, can also be used to prevent a disease, condition abnormal in appearance. A cancer that is to be treated can be or disorder. As used herein, “preventing or “prevent classified as being aneuploid, triploid, tetraploid, or as having describes reducing or eliminating the onset of the symptoms an altered ploidy. A cancer that is to be treated can be classi or complications of the disease, condition or disorder. fied as having a chromosomal translocation, or a deletion or As used herein, the term “alleviate' is meant to describe a duplication of an entire chromosome, or a region of deletion, process by which the severity of a sign or symptom of a duplication or amplification of a portion of a chromosome. disorder is decreased. Importantly, a sign or symptom can be A cancer that is to be treated can be evaluated by DNA 10 cytometry, flow cytometry, or image cytometry. A cancer that alleviated without being eliminated. In a preferred embodi is to be treated can be typed as having 10%, 20%, 30%, 40%, ment, the administration of pharmaceutical compositions of 50%. 60%, 70%, 80%, or 90% of cells in the synthesis stage the invention leads to the elimination of a sign or symptom, of cell division (e.g., in S phase of cell division). A cancer that however, elimination is not required. Effective dosages are is to be treated can be typed as having a low S-phase fraction 15 expected to decrease the severity of a sign or symptom. For or a high S-phase fraction. instance, a sign or symptom of a disorder Such as cancer, As used herein, a “normal cell' is a cell that cannot be which can occur in multiple locations, is alleviated if the classified as part of a “cell proliferative disorder. A normal severity of the cancer is decreased within at least one of cell lacks unregulated or abnormal growth, or both, that can multiple locations. lead to the development of an unwanted condition or disease. As used herein, the term “severity” is meant to describe the Preferably, a normal cell possesses normally functioning cell potential of cancer to transform from a precancerous, or cycle checkpoint control mechanisms. benign, state into a malignant state. Alternatively, or in addi As used herein, "contacting a cell” refers to a condition in tion, severity is meant to describe a cancer stage, for example, which a compound or other composition of matter is in direct according to the TNM system (accepted by the International contact with a cell, or is close enough to induce a desired 25 Union Against Cancer (UICC) and the American Joint Com biological effect in a cell. mittee on Cancer (AJCC)) or by other art-recognized meth As used herein, “candidate compound” refers to a com ods. Cancer stage refers to the extent or severity of the cancer, pound of the present invention, or a pharmaceutically accept based on factors such as the location of the primary tumor, able salt, prodrug, metabolite, polymorph or Solvate thereof, tumor size, number of tumors, and lymph node involvement that has been or will be tested in one or more invitro or in vivo 30 biological assays, in order to determine if that compound is (spread of cancer into lymph nodes). Alternatively, or in addi likely to elicit a desired biological or medical response in a tion, severity is meant to describe the tumor grade by art cell, tissue, system, animal or human that is being sought by recognized methods (see, National Cancer Institute). Tumor a researcher or clinician. A candidate compound is a com grade is a system used to classify cancer cells in terms of how pound of the present invention, or a pharmaceutically accept 35 abnormal they look under a microscope and how quickly the able salt, prodrug, metabolite, polymorph or Solvate thereof. tumor is likely to grow and spread. Many factors are consid The biological or medical response can be the treatment of ered when determining tumor grade, including the structure cancer. The biological or medical response can be treatment and growth pattern of the cells. The specific factors used to or prevention of a cell proliferative disorder. In vitro or in vivo determine tumor grade vary with each type of cancer. Severity biological assays can include, but are not limited to, enzy 40 also describes a histologic grade, also called differentiation, matic activity assays, electrophoretic mobility shift assays, which refers to how much the tumor cells resemble normal reporter gene assays, in vitro cell viability assays, and the cells of the same tissue type (see, National Cancer Institute). assays described herein. Furthermore, severity describes a nuclear grade, which refers As used herein, "monotherapy” refers to the administration to the size and shape of the nucleus in tumor cells and the ofa single active ortherapeutic compound to a subject in need 45 percentage of tumor cells that are dividing (see, National thereof. Preferably, monotherapy will involve administration Cancer Institute). of a therapeutically effective amount of an active compound. In another aspect of the invention, severity describes the For example, cancer monotherapy with one of the compound degree to which a tumor has secreted growth factors, of the present invention, or a pharmaceutically acceptable degraded the extracellular matrix, become vascularized, lost salt, prodrug, metabolite, analog or derivative thereof, to a 50 adhesion to juxtaposed tissues, or metastasized. Moreover, subject in need of treatment of cancer. Monotherapy may be severity describes the number of locations to which a primary contrasted with combination therapy, in which a combination tumor has metastasized. Finally, severity includes the diffi of multiple active compounds is administered, preferably culty of treating tumors of varying types and locations. For with each component of the combination present in a thera example, inoperable tumors, those cancers which have peutically effective amount. In one aspect, monotherapy with 55 greater access to multiple body systems (hematological and a compound of the present invention, or a pharmaceutically immunological tumors), and those which are the most resis acceptable salt, prodrug, metabolite, polymorph or Solvate tant to traditional treatments are considered most severe. In thereof, is more effective than combination therapy in induc these situations, prolonging the life expectancy of the Subject ing a desired biological effect. and/or reducing pain, decreasing the proportion of cancerous As used herein, “treating or “treat describes the manage 60 cells or restricting cells to one system, and improving cancer ment and care of a patient for the purpose of combating a stage/tumor grade/histological grade/nuclear grade are con disease, condition, or disorder and includes the administra sidered alleviating a sign or symptom of the cancer. tion of a compound of the present invention, or a pharmaceu As used herein the term "symptom' is defined as an indi tically acceptable salt, prodrug, metabolite, polymorph or cation of disease, illness, injury, or that something is not right Solvate thereof, to alleviate the symptoms or complications of 65 in the body. Symptoms are felt or noticed by the individual a disease, condition or disorder, or to eliminate the disease, experiencing the symptom, but may not easily be noticed by condition or disorder. others. Others are defined as non-health-care professionals. US 8,962,619 B2 63 64 As used herein the term “sign” is also defined as an indi bladder function (such as more frequent or less frequent uri cation that something is not right in the body. But signs are nation) could be related to bladder or prostate cancer. defined as things that can be seen by a doctor, nurse, or other Changes in skin condition or appearance of a new skin health care professional. condition could indicate cancer. Skin cancers may bleed and Cancer is a group of diseases that may cause almost any 5 look like Sores that do not heal. A long-lasting Sore in the sign or symptom. The signs and symptoms will depend on mouth could be an oral cancer, especially in patients who where the cancer is, the size of the cancer, and how much it Smoke, chew tobacco, or frequently drink alcohol. Sores on affects the nearby organs or structures. If a cancer spreads the penis or vagina may either besigns of infection oran early (metastasizes), then symptoms may appear in different parts CaCC. 10 Unusual bleeding or discharge could indicate cancer. of the body. Unusual bleeding can happen in either early or advanced As a cancer grows, it begins to push on nearby organs, cancer. Blood in the sputum (phlegm) may be a sign of lung blood vessels, and nerves. This pressure creates some of the cancer. Blood in the stool (or a dark or black stool) could be signs and symptoms of cancer. If the cancer is in a critical a sign of colon or rectal cancer. Cancer of the cervix or the area, such as certain parts of the brain, even the Smallest tumor 15 endometrium (lining of the uterus) can cause vaginal bleed can cause early symptoms. ing. Blood in the urine may be a sign of bladder or kidney But sometimes cancers start in places where it does not cancer. A bloody discharge from the nipple may be a sign of cause any symptoms until the cancer has grown quite large. breast cancer. Pancreas cancers, for example, do not usually grow large A thickening or lump in the breast or in other parts of the enough to be felt from the outside of the body. Some pancre- 20 body could indicate the presence of a cancer. Many cancers atic cancers do not cause symptoms until they begin to grow can be felt through the skin, mostly in the breast, testicle, around nearby nerves (this causes a backache). Others grow lymph nodes (glands), and the soft tissues of the body. A lump around the bile duct, which blocks the flow of bile and leads or thickening may be an early or late sign of cancer. Any lump to a yellowing of the skin known as jaundice. By the time a or thickening could be indicative of cancer, especially if the pancreatic cancer causes these signs or symptoms, it has 25 formation is new or has grown in size. usually reached an advanced Stage. Indigestion or trouble Swallowing could indicate cancer. A cancer may also cause symptoms such as fever, fatigue, While these symptoms commonly have other causes, indiges or weight loss. This may be because cancer cells use up much tion or Swallowing problems may be a sign of cancer of the of the body's energy Supply or release Substances that change esophagus, stomach, or pharynx (throat). the body's metabolism. Or the cancer may cause the immune 30 Recent changes in a wart or mole could be indicative of system to react in ways that produce these symptoms. cancer. Any wart, mole, or freckle that changes in color, size, Sometimes, cancer cells release substances into the blood or shape, or loses its definite borders indicates the potential stream that cause symptoms not usually thought to result development of cancer. For example, the skin lesion may be a from cancers. For example, Some cancers of the pancreas can melanoma. release substances which cause blood clots to develop in 35 A persistent cough or hoarseness could be indicative of veins of the legs. Some lung cancers make hormone-like cancer. A cough that does not go away may be a sign of lung substances that affect blood calcium levels, affecting nerves cancer. Hoarseness can be a sign of cancer of the larynx (voice and muscles and causing weakness and dizziness box) or thyroid. Cancer presents several general signs or symptoms that While the signs and symptoms listed above are the more occur when a variety of Subtypes of cancer cells are present. 40 common ones seen with cancer, there are many others that are Most people with cancer will lose weight at some time with less common and are not listed here. However, all art-recog their disease. An unexplained (unintentional) weight loss of nized signs and symptoms of cancer are contemplated and 10 pounds or more may be the first sign of cancer, particularly encompassed by the instant invention. cancers of the pancreas, stomach, esophagus, or lung. Treating cancer can result in a reduction in size of a tumor. Fever is very common with cancer, but is more often seen 45 A reduction in size of a tumor may also be referred to as in advanced disease. Almost all patients with cancer will have “tumor regression’. Preferably, after treatment, tumor size is fever at Some time, especially if the cancer or its treatment reduced by 5% or greater relative to its size prior to treatment; affects the immune system and makes it harder for the body to more preferably, tumor size is reduced by 10% or greater; fight infection. Less often, fever may be an early sign of more preferably, reduced by 20% or greater; more preferably, cancer. Such as with leukemia or lymphoma. 50 reduced by 30% or greater; more preferably, reduced by 40% Fatigue may be an important symptom as cancer or greater; even more preferably, reduced by 50% or greater; progresses. It may happen early, though, in cancers such as and most preferably, reduced by greater than 75% or greater. with leukemia, or if the cancer is causing an ongoing loss of Size of a tumor may be measured by any reproducible means blood, as in some colon or stomach cancers. of measurement. The size of a tumor may be measured as a Pain may be an early symptom with Some cancers such as 55 diameter of the tumor. bone cancers or testicular cancer. But most often pain is a Treating cancer can result in a reduction in tumor Volume. symptom of advanced disease. Preferably, after treatment, tumor volume is reduced by 5% or Along with cancers of the skin (see next section), some greater relative to its size prior to treatment; more preferably, internal cancers can cause skin signs that can be seen. These tumor volume is reduced by 10% or greater; more preferably, changes include the skin looking darker (hyperpigmenta- 60 reduced by 20% or greater; more preferably, reduced by 30% tion), yellow (jaundice), or red (erythema); itching; or exces or greater; more preferably, reduced by 40% or greater; even sive hair growth. more preferably, reduced by 50% or greater; and most pref Alternatively, or in addition, cancer Subtypes present spe erably, reduced by greater than 75% or greater. Tumor volume cific signs or symptoms. Changes in bowel habits or bladder may be measured by any reproducible means of measure function could indicate cancer. Long-term constipation, diar- 65 ment. rhea, or a change in the size of the stool may be a sign of colon Treating cancer results in a decrease in number of tumors. cancer. Pain with urination, blood in the urine, or a change in Preferably, after treatment, tumor number is reduced by 5% or US 8,962,619 B2 65 66 greater relative to number prior to treatment; more preferably, An increase in average Survival time of a population may be tumor number is reduced by 10% or greater; more preferably, measured, for example, by calculating for a population the reduced by 20% or greater; more preferably, reduced by 30% average length of Survival following initiation of treatment or greater; more preferably, reduced by 40% or greater; even with an active compound. An increase in average Survival more preferably, reduced by 50% or greater; and most pref 5 time of a population may also be measured, for example, by erably, reduced by greater than 75%. Number of tumors may calculating for a population the average length of Survival be measured by any reproducible means of measurement. The following completion of a first round of treatment with an number of tumors may be measured by counting tumors active compound. visible to the naked eye or at a specified magnification. Pref Treating cancer can result in a decrease in the mortality rate 10 of a population of treated Subjects in comparison to a popu erably, the specified magnification is 2x. 3x, 4x, 5x, 10x, or lation receiving carrier alone. Treating cancer can result in a 50X. decrease in the mortality rate of a population of treated sub Treating cancer can result in a decrease in number of meta jects in comparison to an untreated population. Treating can static lesions in other tissues or organs distant from the pri cer can result in a decrease in the mortality rate of a popula mary tumor site. Preferably, after treatment, the number of 15 tion of treated Subjects in comparison to a population metastatic lesions is reduced by 5% or greater relative to receiving monotherapy with a drug that is not a compound of number prior to treatment; more preferably, the number of the present invention, or a pharmaceutically acceptable salt, metastatic lesions is reduced by 10% or greater; more pref prodrug, metabolite, analog or derivative thereof. Preferably, erably, reduced by 20% or greater; more preferably, reduced the mortality rate is decreased by more than 2%: more pref by 30% or greater; more preferably, reduced by 40% or erably, by more than 5%; more preferably, by more than 10%: greater; even more preferably, reduced by 50% or greater; and and most preferably, by more than 25%. A decrease in the most preferably, reduced by greater than 75%. The number of mortality rate of a population of treated Subjects may be metastatic lesions may be measured by any reproducible measured by any reproducible means. A decrease in the mor means of measurement. The number of metastatic lesions tality rate of a population may be measured, for example, by may be measured by counting metastatic lesions visible to the 25 calculating for a population the average number of disease naked eye or at a specified magnification. Preferably, the related deaths per unit time following initiation of treatment specified magnification is 2x, 3x, 4x, 5x, 10x, or 50x. with an active compound. A decrease in the mortality rate of Treating cancer can result in an increase in average Survival a population may also be measured, for example, by calcu time of a population of treated Subjects in comparison to a lating for a population the average number of disease-related population receiving carrier alone. Preferably, the average 30 deaths per unit time following completion of a first round of survival time is increased by more than 30 days; more pref treatment with an active compound. erably, by more than 60 days; more preferably, by more than Treating cancer can result in a decrease in tumor growth 90 days; and most preferably, by more than 120 days. An rate. Preferably, after treatment, tumor growth rate is reduced increase in average Survival time of a population may be by at least 5% relative to number prior to treatment; more measured by any reproducible means. An increase in average 35 preferably, tumor growth rate is reduced by at least 10%: Survival time of a population may be measured, for example, more preferably, reduced by at least 20%; more preferably, by calculating for a population the average length of Survival reduced by at least 30%; more preferably, reduced by at least following initiation of treatment with an active compound. An 40%: more preferably, reduced by at least 50%; even more increase in average Survival time of a population may also be preferably, reduced by at least 50%; and most preferably, measured, for example, by calculating for a population the 40 reduced by at least 75%. Tumor growth rate may be measured average length of Survival following completion of a first by any reproducible means of measurement. Tumor growth round of treatment with an active compound. rate can be measured according to a change in tumor diameter Treating cancer can result in an increase in average Survival per unit time. time of a population of treated Subjects in comparison to a Treating cancer can result in a decrease in tumor regrowth. population of untreated subjects. Preferably, the average sur 45 Preferably, after treatment, tumor regrowth is less than 5%; vival time is increased by more than 30 days; more preferably, more preferably, tumor regrowth is less than 10%: more pref by more than 60 days; more preferably, by more than 90 days; erably, less than 20%; more preferably, less than 30%; more and most preferably, by more than 120 days. An increase in preferably, less than 40%; more preferably, less than 50%: average Survival time of a population may be measured by any even more preferably, less than 50%; and most preferably, reproducible means. An increase in average Survival time of a 50 less than 75%. Tumor regrowth may be measured by any population may be measured, for example, by calculating for reproducible means of measurement. Tumor regrowth is mea a population the average length of Survival following initia Sured, for example, by measuring an increase in the diameter tion of treatment with an active compound. An increase in of a tumor after a prior tumor shrinkage that followed treat average Survival time of a population may also be measured, ment. A decrease in tumor regrowth is indicated by failure of for example, by calculating for a population the average 55 tumors to reoccur after treatment has stopped. length of survival following completion of a first round of Treating or preventing a cell proliferative disorder can treatment with an active compound. result in a reduction in the rate of cellular proliferation. Pref Treating cancer can result in increase in average Survival erably, after treatment, the rate of cellular proliferation is time of a population of treated Subjects in comparison to a reduced by at least 5%: more preferably, by at least 10%: population receiving monotherapy with a drug that is not a 60 more preferably, by at least 20%; more preferably, by at least compound of the present invention, or a pharmaceutically 30%: more preferably, by at least 40%; more preferably, by at acceptable salt, prodrug, metabolite, analog or derivative least 50%; even more preferably, by at least 50%; and most thereof. Preferably, the average survival time is increased by preferably, by at least 75%. The rate of cellular proliferation more than 30 days; more preferably, by more than 60 days; may be measured by any reproducible means of measure more preferably, by more than 90 days; and most preferably, 65 ment. The rate of cellular proliferation is measured, for by more than 120 days. An increase in average Survival time example, by measuring the number of dividing cells in a of a population may be measured by any reproducible means. tissue sample per unit time. US 8,962,619 B2 67 68 Treating or preventing a cell proliferative disorder can example, cell death would be said to occur selectively in result in a reduction in the proportion of proliferating cells. cancer cells if it occurred greater than twice as frequently in Preferably, after treatment, the proportion of proliferating cancer cells as compared to normal cells. cells is reduced by at least 5%: more preferably, by at least A compound of the present invention, or a pharmaceuti 10%; more preferably, by at least 20%; more preferably, by at cally acceptable salt, prodrug, metabolite, polymorph or Sol least 30%; more preferably, by at least 40%; more preferably, vate thereof, can modulate the activity of a molecular target by at least 50%; even more preferably, by at least 50%; and (e.g., a target kinase). Modulating refers to stimulating or most preferably, by at least 75%. The proportion of prolifer inhibiting an activity of a molecular target. Preferably, a com ating cells may be measured by any reproducible means of pound of the present invention, or a pharmaceutically accept measurement. Preferably, the proportion of proliferating cells 10 able salt, prodrug, metabolite, polymorph or Solvate thereof, is measured, for example, by quantifying the number of divid modulates the activity of a molecular target if it stimulates or ing cells relative to the number of nondividing cells in a tissue inhibits the activity of the molecular target by at least 2-fold sample. The proportion of proliferating cells can be equiva relative to the activity of the molecular target under the same lent to the mitotic index. conditions but lacking only the presence of said compound. Treating or preventing a cell proliferative disorder can 15 More preferably, a compound of the present invention, or a result in a decrease in size of an area or Zone of cellular pharmaceutically acceptable salt, prodrug, metabolite, poly proliferation. Preferably, after treatment, size of an area or morph or solvate thereof, modulates the activity of a molecu Zone of cellular proliferation is reduced by at least 5% relative lar target if it stimulates or inhibits the activity of the molecu to its size prior to treatment; more preferably, reduced by at lar target by at least 5-fold, at least 10-fold, at least 20-fold, at least 10%: more preferably, reduced by at least 20%; more least 50-fold, at least 100-fold relative to the activity of the preferably, reduced by at least 30%; more preferably, reduced molecular target under the same conditions but lacking only by at least 40%; more preferably, reduced by at least 50%: the presence of said compound. The activity of a molecular even more preferably, reduced by at least 50%; and most target may be measured by any reproducible means. The preferably, reduced by at least 75%. Size of an area or Zone of activity of a molecular target may be measured in vitro or in cellular proliferation may be measured by any reproducible 25 vivo. For example, the activity of a molecular target may be means of measurement. The size of an area or Zone of cellular measured in vitro by an enzymatic activity assay or a DNA proliferation may be measured as a diameter or width of an binding assay, or the activity of a molecular target may be area or Zone of cellular proliferation. measured in vivo by assaying for expression of a reporter Treating or preventing a cell proliferative disorder can gene. result in a decrease in the number or proportion of cells 30 A compound of the present invention, or a pharmaceuti having an abnormal appearance or morphology. Preferably, cally acceptable salt, prodrug, metabolite, polymorph or Sol after treatment, the number of cells having an abnormal mor vate thereof, does not significantly modulate the activity of a phology is reduced by at least 5% relative to its size prior to molecular target if the addition of the compound does not treatment; more preferably, reduced by at least 10%: more stimulate or inhibit the activity of the molecular target by preferably, reduced by at least 20%; more preferably, reduced 35 greater than 10% relative to the activity of the molecular by at least 30%; more preferably, reduced by at least 40%; target under the same conditions but lacking only the pres more preferably, reduced by at least 50%; even more prefer ence of said compound. ably, reduced by at least 50%; and most preferably, reduced As used herein, the term “isozyme selective” means pref by at least 75%. An abnormal cellular appearance or morphol erential inhibition or stimulation of a first isoform of an ogy may be measured by any reproducible means of measure 40 enzyme in comparison to a second isoform of an enzyme ment. An abnormal cellular morphology can be measured by (e.g., preferential inhibition or stimulation of a kinase microscopy, e.g., using an inverted tissue culture microscope. isozyme alpha in comparison to a kinase isozyme beta). Pref An abnormal cellular morphology can take the form of erably, a compound of the present invention, or a pharmaceu nuclear pleiomorphism. tically acceptable salt, prodrug, metabolite, polymorph or As used herein, the term “selectively’ means tending to 45 solvate thereof, demonstrates a minimum of a four fold dif occurat a higher frequency in one population than in another ferential, preferably a ten fold differential, more preferably a population. The compared populations can be cell popula fifty fold differential, in the dosage required to achieve a tions. Preferably, a compound of the present invention, or a biological effect. Preferably, a compound of the present pharmaceutically acceptable salt, prodrug, metabolite, poly invention, or a pharmaceutically acceptable salt, prodrug, morph or Solvate thereof, acts selectively on a cancer or 50 metabolite, polymorph or solvate thereof, demonstrates this precancerous cell but not on a normal cell. Preferably, a differential across the range of inhibition, and the differential compound of the present invention, or a pharmaceutically is exemplified at the ICs, i.e., a 50% inhibition, for a molecu acceptable salt, prodrug, metabolite, polymorph or Solvate lar target of interest. thereof, acts selectively to modulate one molecular target Administering a compound of the present invention, or a (e.g., a target kinase) but does not significantly modulate 55 pharmaceutically acceptable salt, prodrug, metabolite, poly another molecular target (e.g., a non-target kinase). The morph or solvate thereof, to a cell or a subject in need thereof invention also provides a method for selectively inhibiting the can result in modulation (i.e., stimulation or inhibition) of an activity of an enzyme. Such as a kinase. Preferably, an event activity of a kinase of interest. occurs selectively in population A relative to population B if The present invention provides methods to assess biologi it occurs greater than two times more frequently in population 60 cal activity of a compound of the present invention, or a Aas compared to population B. An event occurs selectively if pharmaceutically acceptable salt, prodrug, metabolite, poly it occurs greater than five times more frequently in population morph or Solvate thereof. In one method, an assay based on A. An event occurs selectively if it occurs greater than ten enzymatic activity can be utilized. In one specific enzymatic times more frequently in population A; more preferably, activity assay, the enzymatic activity is from a kinase. As used greater than fifty times; even more preferably, greater than 65 herein, “kinase' refers to a large class of enzymes which 100 times; and most preferably, greater than 1000 times more catalyze the transfer of the Y-phosphate from ATP to the frequently in population A as compared to population B. For hydroxyl group on the side chain of Ser/Thr or Tyr in proteins US 8,962,619 B2 69 70 and peptides and are intimately involved in the control of or more members of the cell cycle checkpoint pathway. Pref various important cell functions, perhaps most notably: signal erably, a cell cycle checkpoint pathway is a biochemical transduction, differentiation, and proliferation. There are esti signaling pathway. mated to be about 2,000 distinct protein kinases in the human As used herein, “cell cycle checkpoint regulator” refers to body, and although each of these phosphorylates particular a composition of matter that can function, at least in part, in protein/peptide Substrates, they all bind the same second Sub modulation of a cell cycle checkpoint. A cell cycle checkpoint strate ATP in a highly conserved pocket. About 50% of the regulator may have stimulatory or inhibitory effects, or both, known oncogene products are protein tyrosine kinases on one or more functions comprising a cell cycle checkpoint. (PTKs), and their kinase activity has been shown to lead to A cell cycle checkpoint regulator can be a protein or not a cell transformation. Preferably, the kinase assayed is a 10 protein. tyrosine kinase. Treating cancer or a cell proliferative disorder can result in A change in enzymatic activity caused by a compound of cell death, and preferably, cell death results in a decrease of at the present invention, or a pharmaceutically acceptable salt, least 10% in number of cells in a population. More preferably, prodrug, metabolite, polymorph or Solvate thereof, can be cell death means a decrease of at least 20%; more preferably, measured in the disclosed assays. The change in enzymatic 15 a decrease of at least 30%; more preferably, a decrease of at activity can be characterized by the change in the extent of least 40%; more preferably, a decrease of at least 50%; most phosphorylation of certain Substrates. As used herein, phos preferably, a decrease of at least 75%. Number of cells in a phorylation” refers to the addition of phosphate groups to a population may be measured by any reproducible means. A Substrate, including proteins and organic molecules; and, number of cells in a population can be measured by fluores plays an important role in regulating the biological activities cence activated cell sorting (FACS), immunofluorescence of proteins. Preferably, the phosphorylation assayed and mea microscopy and light microscopy. Methods of measuring cell Sured involves the addition of phosphate groups to tyrosine death are as shown in Li et al., Proc Natl Acad Sci USA. residues. The Substrate can be a peptide or protein. 100(5): 2674-8, 2003. In an aspect, cell death occurs by In some assays, immunological reagents, e.g., antibodies apoptosis. and antigens, are employed. Fluorescence can be utilized in 25 Preferably, an effective amount of a compound of the the measurement of enzymatic activity in Some assays. As present invention, or a pharmaceutically acceptable salt, pro used herein, “fluorescence” refers to a process through which drug, metabolite, polymorph or Solvate thereof, is not signifi a molecule emits a photon as a result of absorbing an incom cantly cytotoxic to normal cells. A therapeutically effective ing photon of higher energy by the same molecule. Specific amount of a compound is not significantly cytotoxic to nor methods for assessing the biological activity of the disclosed 30 mal cells if administration of the compound in a therapeuti compounds are described in the examples. cally effective amount does not induce cell death in greater As used herein, an activity of c-Met refers to any biological than 10% of normal cells. Atherapeutically effective amount function or activity that is carried out by c-Met. For example, of a compound does not significantly affect the viability of a function of c-Met includes phosphorylation of downstream normal cells if administration of the compound in a therapeu target proteins. Other functions of c-Met include autophos 35 tically effective amount does not induce cell death in greater phorylation, binding of adaptor proteins such as Gab-1, Grb than 10% of normal cells. In an aspect, cell death occurs by 2, Shc, SHP2 and c-Cbl, and activation of signal transducers apoptosis. such as Ras, Src, PI3K, PLC-Y, STATs, ERK1 and 2 and FAK. Contacting a cell with a compound of the present invention, Administering a compound of the present invention, or a or a pharmaceutically acceptable salt, prodrug, metabolite, pharmaceutically acceptable salt, prodrug, metabolite, poly 40 polymorph or Solvate thereof, can induce or activate cell morph or solvate thereof, to a cellor a subject in need thereof death selectively in cancer cells. Administering to a Subject in results in modulation (i.e., stimulation or inhibition) of an need thereof a compound of the present invention, or a phar activity of an intracellular target (e.g., Substrate). Several maceutically acceptable salt, prodrug, metabolite, polymorph intracellular targets can be modulated with the compounds of or solvate thereof, can induce or activate cell death selectively the present invention, including, but not limited to, adaptor 45 in cancer cells. Contacting a cell with a compound of the proteins such as Gab-1, Grb-2, Shc, SHP2 and c-Cbl, and present invention, or a pharmaceutically acceptable salt, pro signal transducers such as Ras, Src, PI3K, PLC-Y, STATs, drug, metabolite, polymorph or Solvate thereof, can induce ERK1 and 2 and FAK. cell death selectively in one or more cells affected by a cell Activating refers to placing a composition of matter (e.g., proliferative disorder. Preferably, administering to a subject protein or nucleic acid) in a state Suitable for carrying out a 50 in need thereof a compound of the present invention, or a desired biological function. A composition of matter capable pharmaceutically acceptable salt, prodrug, metabolite, poly of being activated also has an unactivated State. An activated morph or solvate thereof, induces cell death selectively in one composition of matter may have an inhibitory or stimulatory or more cells affected by a cell proliferative disorder. biological function, or both. The present invention relates to a method of treating or Elevation refers to an increase in a desired biological activ 55 preventing cancer by administering a compound of the ity of a composition of matter (e.g., a protein or a nucleic present invention, or a pharmaceutically acceptable salt, pro acid). Elevation may occur through an increase in concentra drug, metabolite, polymorph or Solvate thereof, to a Subject in tion of a composition of matter. need thereof, where administration of the compound of the As used herein, “a cell cycle checkpoint pathway' refers to present invention, or a pharmaceutically acceptable salt, pro a biochemical pathway that is involved in modulation of a cell 60 drug, metabolite, polymorph or Solvate thereof, results in one cycle checkpoint. A cell cycle checkpoint pathway may have or more of the following: accumulation of cells in G1 and/or stimulatory or inhibitory effects, or both, on one or more S phase of the cell cycle, cytotoxicity via cell death in cancer functions comprising a cell cycle checkpoint. A cell cycle cells without a significant amount of cell death in normal checkpoint pathway is comprised of at least two composi cells, antitumor activity in animals with atherapeutic index of tions of matter, preferably proteins, both of which contribute 65 at least 2, and activation of a cell cycle checkpoint. As used to modulation of a cell cycle checkpoint. A cell cycle check herein, “therapeutic index' is the maximum tolerated dose point pathway may be activated through an activation of one divided by the efficacious dose. US 8,962,619 B2 71 72 One skilled in the art may refer to general reference texts A compound of the present invention, or a pharmaceuti for detailed descriptions of known techniques discussed cally acceptable salt, prodrug, metabolite, analog or deriva herein or equivalent techniques. These texts include Ausubel tive thereof, may be administered in combination with a sec et al., Current Protocols in Molecular Biology, John Wiley ond chemotherapeutic agent. The second chemotherapeutic and Sons, Inc. (2005); Sambrook et al., Molecular Cloning, A agent (also referred to as an anti-neoplastic agent or anti Laboratory Manual (3" edition), Cold Spring Harbor Press, proliferative agent) can be an alkylating agent; an antibiotic; Cold Spring Harbor, N.Y. (2000); Coligan et al., Current an anti-metabolite; a detoxifying agent; an interferon; a poly Protocols in Immunology, John Wiley & Sons, N.Y.; Enna et clonal or monoclonal antibody; an EGFR inhibitor; a HER2 al., Current Protocols in Pharmacology, John Wiley & Sons, inhibitor; a histone deacetylase inhibitor; a hormone; a N.Y.; Fingletal. The Pharmacological Basis of Therapeutics 10 mitotic inhibitor; an MTOR inhibitor; a multi-kinase inhibi tor; a serine/threonine kinase inhibitor, a tyrosine kinase (1975), Remington's Pharmaceutical Sciences, Mack Pub inhibitors; a VEGF/VEGFR inhibitor; a taxane or taxane lishing Co., Easton, Pa., 18" edition (1990). These texts can, derivative, an aromatase inhibitor, an anthracycline, a micro of course, also be referred to in making or using an aspect of tubule targeting drug, atopoisomerase poison drug, an inhibi the invention 15 tor of a molecular target or enzyme (e.g., a kinase inhibitor), As used herein, “combination therapy’ or “co-therapy' a cytidine analogue drug or any chemotherapeutic, anti-neo includes the administration of a compound of the present plastic or anti-proliferative agent listed in www.cancer.org/ invention, or a pharmaceutically acceptable salt, prodrug, docroot/cdg/cdg 0.asp. metabolite, polymorph or Solvate thereof, and at least a sec Exemplary alkylating agents include, but are not limited to, ond agent as part of a specific treatment regimen intended to cyclophosphamide (Cytoxan; Neosar); chlorambucil (Leuke provide the beneficial effect from the co-action of these thera ran); melphalan (Alkeran); carmustine (BiCNU); busulfan peutic agents. The beneficial effect of the combination (Busulfex); lomustine (CeeNU); dacarbazine (DTIC-Dome); includes, but is not limited to, pharmacokinetic or pharmaco oxaliplatin (Eloxatin); carmustine (Gliadel); Ifosfamide dynamic co-action resulting from the combination of thera (Ifex); mechlorethamine (Mustargen); busulfan (Myleran); peutic agents. Administration of these therapeutic agents in 25 carboplatin (Paraplatin); cisplatin (CDDP: Platinol); temozo combination typically is carried out over a defined time lomide (Temodar); thiotepa (Thioplex); bendamustine (Tre period (usually minutes, hours, days or weeks depending anda); or Streptozocin (Zanosar). upon the combination selected). “Combination therapy” may Exemplary antibiotics include, but are not limited to, doxo be, but generally is not, intended to encompass the adminis rubicin (Adriamycin); doxorubicin liposomal (Doxil); mitox tration of two or more of these therapeutic agents as part of 30 antrone (Novantrone); bleomycin (Blenoxane); daunorubicin separate monotherapy regimens that incidentally and arbi (Cerubidine); daunorubicin liposomal (DaunoXome); dacti trarily result in the combinations of the present invention. nomycin (Cosmegen); epirubicin (Ellence); idarubicin (Ida “Combination therapy” is intended to embrace administra mycin); plicamycin (Mithracin); mitomycin (Mutamycin); tion of these therapeutic agents in a sequential manner, pentostatin (Nipent); or valrubicin (Valstar). wherein each therapeutic agent is administered at a different 35 Exemplary anti-metabolites include, but are not limited to, time, as well as administration of these therapeutic agents, or fluorouracil (Adrucil); capecitabine (Xeloda): hydroxyurea at least two of the therapeutic agents, in a Substantially simul (Hydrea); mercaptopurine (Purinethol); pemetrexed (Al taneous manner. Substantially simultaneous administration imta); fludarabine (Fludara); nelarabine (Arranon); cladrib can be accomplished, for example, by administering to the ine (Cladribine Novaplus); clofarabine (Clolar); cytarabine Subject a single capsule having a fixed ratio of each therapeu 40 (Cytosar-U); decitabine (Dacogen); cytarabine liposomal tic agent or in multiple, single capsules for each of the thera (DepoCyt); hydroxyurea (Droxia); pralatrexate (Folotyn); peutic agents. Sequential or Substantially simultaneous floxuridine (FUDR); gemcitabine (Gemzar); cladribine administration of each therapeutic agent can be effected by (Leustatin); fludarabine (Oforta); methotrexate (MTX; Rheu any appropriate route including, but not limited to, oral matrex); methotrexate (Trexall); thioguanine (Tabloid); TS-1 routes, intravenous routes, intramuscular routes, and direct 45 or cytarabine (Tarabine PFS). absorption through mucous membrane tissues. The therapeu Exemplary detoxifying agents include, but are not limited tic agents can be administered by the same route or by differ to, amifostine (Ethyol) or mesna (Mesnex). ent routes. For example, a first therapeutic agent of the com Exemplary interferons include, but are not limited to, inter bination selected may be administered by intravenous feron alfa-2b (Intron A) or interferon alfa-2a (Roferon-A). injection while the other therapeutic agents of the combina 50 Exemplary polyclonal or monoclonal antibodies include, tion may be administered orally. Alternatively, for example, but are not limited to, trastuzumab (Herceptin); ofatumumab all therapeutic agents may be administered orally or all thera (Arzerra); bevacizumab (Avastin); rituximab (Rituxan); peutic agents may be administered by intravenous injection. cetuximab (Erbitux); panitumumab (Vectibix); tositumomab/ The sequence in which the therapeutic agents are adminis iodine''' to situmomab (Bexxar); alemtuzumab (Campath); tered is not narrowly critical. 55 ibritumomab (Zevalin: In-111; Y-90 Zevalin); gemtuzumab “Combination therapy' also embraces the administration (Mylotarg); eculizumab (Soliris) ordenosumab. of the therapeutic agents as described above in further com Exemplary EGFR inhibitors include, but are not limited to, bination with other biologically active ingredients and non gefitinib (Iressa); lapatinib (Tykerb); cetuximab (Erbitux); drug therapies (e.g., Surgery or radiation treatment). Where erlotinib (Tarceva); panitumumab (Vectibix); PKI-166; can the combination therapy further comprises a non-drug treat 60 ertinib (CI-1033); matuzumab (Emd7200) or EKB-569. ment, the non-drug treatment may be conducted at any Suit Exemplary HER2 inhibitors include, but are not limited to, able time so long as a beneficial effect from the co-action of trastuzumab (Herceptin); lapatinib (Tykerb) or AC-480. the combination of the therapeutic agents and non-drug treat Histone Deacetylase Inhibitors include, but are not limited ment is achieved. For example, in appropriate cases, the ben to, Vorinostat (Zollinza). eficial effect is still achieved when the non-drug treatment is 65 Exemplary hormones include, but are not limited to, temporally removed from the administration of the therapeu tamoxifen (Soltamox; Nolvadex); raloxifene (Evista); mege tic agents, perhaps by days or even weeks. strol (Megace); leuprolide (Lupron; Lupron Depot; Eligard; US 8,962,619 B2 73 74 Viadur); fulvestrant (Faslodex); letrozole (Ferrara); triptore factor). In another aspect, a compound of the present inven lin (Trelstar LA; Trelstar Depot); exemestane (Aromasin): tion, or a pharmaceutically acceptable salt, prodrug, metabo goserelin (Zoladex); bicalutamide (CaSodex); anastroZole lite, analog or derivative thereof, may be administered in (Arimidex); fluoxymesterone (Androxy: Halotestin): combination with radiation therapy. Radiation therapy can medroxyprogesterone (Provera; Depo-Provera); estramus also be administered in combination with a compound of the tine (Emcyt); flutamide (Eulexin); toremifene (Fareston); present invention and another chemotherapeutic agent degarelix (Firmagon); nilutamide (Nilandron); abarelix described herein as part of a multiple agent therapy. In yet (Plenaxis); or testolactone (Teslac). another aspect, a compound of the present invention, or a Exemplary mitotic inhibitors include, but are not limited pharmaceutically acceptable salt, prodrug, metabolite, ana to, paclitaxel (Taxol; Onxol; Abraxane); docetaxel (Taxo 10 log orderivative thereof, may be administered in combination tere); Vincristine (Oncovin; Vincasar PFS); vinblastine (Vel with standard chemotherapy combinations such as, but not ban): etoposide (Toposar; Etopophos; VePesid); teniposide restricted to, CMF (cyclophosphamide, methotrexate and (Vumon); ixabepilone (Ixempra); nocodazole; epothilone; 5-fluorouracil), CAF (cyclophosphamide, adriamycin and vinorelbine (Navelbine); camptothecin (CPT); irinotecan 5-fluorouracil), AC (adriamycin and cyclophosphamide), (Camptosar); topotecan (Hycamtin); amsacrine or lamellarin 15 FEC (5-fluorouracil, epirubicin, and cyclophosphamide), D (LAM-D). ACT or ATC (adriamycin, cyclophosphamide, and pacli Exemplary MTOR inhibitors include, but are not limited taxel), rituximab, Xeloda (capecitabine), Cisplatin (CDDP), to, everolimus (Afinitor) or temsirolimus (Torisel); rapam Carboplatin, TS-1 (tegafur, gimestat and otastat potassium at une, ridaforolimus; or AP23573. a molar ratio of 1:0.4:1), Camptothecin-11 (CPT-11, Irinote Exemplary multi-kinase inhibitors include, but are not lim can or CamptosarTM) or CMFP (cyclophosphamide, methotr ited to, Sorafenib (Nexavar); Sunitinib (Sutent); BIBW 2992: exate, 5-fluorouracil and prednisone). E7080; Zdé474; PKC-412; motesanib; or AP24534. In preferred embodiments, a compound of the present Exemplary serine/threonine kinase inhibitors include, but invention, or a pharmaceutically acceptable salt, prodrug, are not limited to, ruboxistaurin; eril/easudil hydrochloride: metabolite, polymorph or Solvate thereof, may be adminis flavopiridol; seliciclib (CYC202; Roscovitrine); SNS-032 25 tered with an inhibitor of an enzyme. Such as a receptor or (BMS-387032); Pkc412; bryostatin: KAI-9803; SF1126; non-receptor kinase. Receptor and non-receptor kinases of VX-680: Azd 1152: Arry-142886 (AZD-6244); SCIO-469; the invention are, for example, tyrosine kinases or serine/ GW681323; CC-401; CEP-1347 or PD 332991. threonine kinases. Kinase inhibitors of the invention are small Exemplary tyrosine kinase inhibitors include, but are not molecules, polynucleic acids, polypeptides, or antibodies. limited to, erlotinib (Tarceva); gefitinib (Iressa); imatinib 30 Exemplary kinase inhibitors include, but are not limited to, (Gleevec); Sorafenib (Nexavar); Sunitinib (Sutent); trastu Bevacizumab (targets VEGF), BIBW 2.992 (targets EGFR Zumab (Herceptin); bevacizumab (Avastin); rituximab (Rit and Erb2), Cetuximab/Erbitux (targets Erb1), Imatinib/ uxan); lapatinib (Tykerb); cetuximab (Erbitux); panitu Gleevic (targets Bcr-Abl), Trastuzumab (targets Erb2), Gefi mumab (Vectibix); everolimus (Afinitor); alemtuzumab tinib/Iressa (targets EGFR), Ranibizumab (targets VEGF), (Campath); gemtuzumab (Mylotarg); temsirolimus (Torisel); 35 Pegaptainib (targets VEGF), Erlotinib/Tarceva (targets Erb 1), pazopanib (Votrient); dasatinib (Sprycel); nilotinib (Tasi Nilotinib (targets Bcr-Abl), Lapatinib (targets Erb1 and Erb2/ gna); vatalanib (Ptk787; ZK222584); CEP-701: SU5614; Her2), GW-572016/lapatinib ditosylate (targets HER2/ MLN518; XL999; VX-322: Azd0530; BMS-354825; SKI Erb2), Panitumumab/Vectibix (targets EGFR), Vandetinib 606 CP-690; AG-490; WHI-P154: WHI-P131; AC-220; or (targets RET/VEGFR), E7080 (multiple targets including AMG888. 40 RET and VEGFR), Herceptin (targets HER2/Erb2), PKI-166 Exemplary VEGF/VEGFR inhibitors include, but are not (targets EGFR), Canertinib/CI-1033 (targets EGFR), Suni limited to, bevacizumab (Avastin): Sorafenib (Nexavar); Suni tinib/SU-11464/Sutent (targets EGFR and FLT3), Matu tinib (Sutent); ranibizumab: pegaptanib; or vandetinib. Zumab/Emd7200 (targets EGFR), EKB-569 (targets EGFR), Exemplary microtubule targeting drugs include, but are not Zdé474 (targets EGFR and VEGFR), PKC-412 (targets limited to, paclitaxel, docetaxel, Vincristin, vinblastin, 45 VEGR and FLT3), Vatalanib/Ptk787/ZK222584 (targets nocodazole, epothilones and navelbine. VEGR), CEP-701 (targets FLT3), SU5614 (targets FLT3), Exemplary topoisomerase poison drugs include, but are MLN518 (targets FLT3), XL999 (targets FLT3), VX-322 not limited to, teniposide, etoposide, adriamycin, camptoth (targets FLT3), AZd0530 (targets SRC), BMS-354825 (tar ecin, daunorubicin, dactinomycin, mitoxantrone, amsacrine, gets SRC), SKI-606 (targets SRC), CP-690 (targets JAK), epirubicin and idarubicin. 50 AG-490 (targets JAK), WHI-P154 (targets JAK), WHI-P131 Exemplary taxanes or taxane derivatives include, but are (targets JAK), Sorafenib/Nexavar (targets RAF kinase, not limited to, paclitaxel and docetaxol. VEGFR-1, VEGFR-2, VEGFR-3, PDGFR-13, KIT, FLT-3, Exemplary general chemotherapeutic, anti-neoplastic, and RET), Dasatinib/Sprycel (BCR/ABL and Src), AC-220 anti-proliferative agents include, but are not limited to, altre (targets Flt3), AC-480 (targets all HER proteins, “panHER”), tamine (Hexylen); isotretinoin (Accutane; Amnesteem; Clar 55 Motesanib diphosphate (targets VEGF1-3, PDGFR, and avis; Sotret); tretinoin (Vesanoid); azacitidine (Vidaza); bort c-kit), Denosumab (targets RANKL, inhibits SRC), eZomib (Velcade) asparaginase (Elspar); levamisole AMG888 (targets HER3), and AP24534 (multiple targets (Ergamisol); mitotane (LySodren); procarbazine (Matulane); including Flt3). pegaspargase (Oncaspar); denileukin diftitoX (Ontak); por Exemplary serine/threonine kinase inhibitors include, but fimer (Photofrin); aldesleukin (Proleukin); lenalidomide 60 are not limited to, Rapamune (targets mTOR/FRAP1), (Revlimid); bexarotene (Targretin); thalidomide (Thalomid); Deforolimus (targets mTOR), Certican/Everolimus (targets temsirolimus (Torisel); arsenic trioxide (Trisenox); vertepor mTOR/FRAP1), AP23573 (targets mTOR/FRAP1), Eril/Fa fin (Visudyne); mimosine (Leucenol); (1 M tegafur 0.4 M Sudil hydrochloride (targets RHO), Flavopiridol (targets 5-chloro-2,4-dihydroxypyrimidine—1 M potassium CDK), Seliciclib/CYC202/Roscovitrine (targets CDK), oXonate) or lovastatin. 65 SNS-032/BMS-387032 (targets CDK), Ruboxistaurin (tar In another aspect, the second chemotherapeutic agent can gets PKC). Pkc412 (targets PKC), Bryostatin (targets PKC). be a cytokine Such as G-CSF (granulocyte colony stimulating KAI-9803 (targets PKC), SF1126 (targets PI3K), VX-680 US 8,962,619 B2 75 76 (targets Aurora kinase), AZd 1152 (targets Aurora kinase), can be adjusted with acids or bases, such as hydrochloric acid Arry-142886/AZD-6244 (targets MAP/MEK), SCID-469 or Sodium hydroxide. The parenteral preparation can be (targets MAP/MEK), GW681323 (targets MAP/MEK), enclosed in ampoules, disposable Syringes or multiple dose CC-401 (targets JNK), CEP-1347 (targets JNK), and PD vials made of glass or plastic. 332991 (targets CDK). A compound or pharmaceutical composition of the inven tion can be administered to a subject in many of the well 4. Pharmaceutical Compositions known methods currently used for chemotherapeutic treat ment. For example, for treatment of cancers, a compound of The present invention also provides pharmaceutical com the invention may be injected directly into tumors, injected into the blood stream or body cavities or taken orally or positions comprising a compound of each of the formulae 10 applied through the skin with patches. The dose chosen described herein in combination with at least one pharmaceu should be sufficient to constitute effective treatment but not as tically acceptable excipient or carrier. high as to cause unacceptable side effects. The state of the A "pharmaceutical composition' is a formulation contain disease condition (e.g., cancer, precancer, and the like) and ing the compounds of the present invention in a form Suitable the health of the patient should preferably be closely moni for administration to a Subject. In one embodiment, the phar 15 tored during and for a reasonable period after treatment. maceutical composition is in bulk or in unit dosage form. The The term “therapeutically effective amount’, as used unit dosage form is any of a variety of forms, including, for herein, refers to an amount of a pharmaceutical agent to treat, example, a capsule, an IV bag, a tablet, a single pump on an ameliorate, or prevent an identified disease or condition, or to aerosol inhaler or a vial. The quantity of active ingredient exhibit a detectable therapeutic or inhibitory effect. The effect (e.g., a formulation of the disclosed compound or salt, can be detected by any assay method known in the art. The hydrate, Solvate or isomer thereof) in a unit dose of compo precise effective amount for a subject will depend upon the sition is an effective amount and is varied according to the Subject's body weight, size, and health; the nature and extent particular treatment involved. One skilled in the art will of the condition; and the therapeutic or combination of thera appreciate that it is sometimes necessary to make routine peutics selected for administration. Therapeutically effective variations to the dosage depending on the age and condition 25 amounts for a given situation can be determined by routine of the patient. The dosage will also depend on the route of experimentation that is within the skill and judgment of the administration. A variety of routes are contemplated, includ clinician. In a preferred aspect, the disease or condition to be ing oral, pulmonary, rectal, parenteral, transdermal, Subcuta treated is cancer. In another aspect, the disease or condition to neous, intravenous, intramuscular, intraperitoneal, inhala be treated is a cell proliferative disorder. tional, buccal, Sublingual, intrapleural, intrathecal, intranasal, 30 For any compound, the therapeutically effective amount and the like. Dosage forms for the topical or transdermal can be estimated initially either in cell culture assays, e.g., of administration of a compound of this invention include pow neoplastic cells, or in animal models, usually rats, mice, rab ders, sprays, ointments, pastes, creams, lotions, gels, solu bits, dogs, or pigs. The animal model may also be used to tions, patches and inhalants. In one embodiment, the active determine the appropriate concentration range and route of compound is mixed under sterile conditions with a pharma 35 administration. Such information can then be used to deter ceutically acceptable carrier, and with any preservatives, mine useful doses and routes for administration in humans. buffers or propellants that are required. Therapeutic/prophylactic efficacy and toxicity may be deter As used herein, the phrase “pharmaceutically acceptable' mined by standard pharmaceutical procedures in cell cultures refers to those compounds, materials, compositions, carriers, or experimental animals, e.g., EDs (the dose therapeutically and/or dosage forms which are, within the scope of Sound 40 effective in 50% of the population) and LDs (the dose lethal medical judgment, Suitable for use in contact with the tissues to 50% of the population). The dose ratio between toxic and of human beings and animals without excessive toxicity, irri therapeutic effects is the therapeutic index, and it can be tation, allergic response, or other problem or complication, expressed as the ratio, LDso/EDso. Pharmaceutical composi commensurate with a reasonable benefit/risk ratio. tions that exhibit large therapeutic indices are preferred. The "Pharmaceutically acceptable excipient’ means an excipi 45 dosage may vary within this range depending upon the dosage ent that is useful in preparing a pharmaceutical composition form employed, sensitivity of the patient, and the route of that is generally safe, non-toxic and neither biologically nor administration. otherwise undesirable, and includes excipient that is accept Dosage and administration are adjusted to provide Suffi able for veterinary use as well as human pharmaceutical use. cient levels of the active agent(s) or to maintain the desired A "pharmaceutically acceptable excipient’ as used in the 50 effect. Factors which may be taken into account include the specification and claims includes both one and more than one severity of the disease state, general health of the Subject, age, Such excipient. weight, and gender of the Subject, diet, time and frequency of A pharmaceutical composition of the invention is formu administration, drug combination(s), reaction sensitivities, lated to be compatible with its intended route of administra and tolerance/response to therapy. Long-acting pharmaceuti tion. Examples of routes of administration include parenteral, 55 cal compositions may be administered every 3 to 4 days, e.g., intravenous, intradermal. Subcutaneous, oral (e.g., inha every week, or once every two weeks depending on half-life lation), transdermal (topical), and transmucosal administra and clearance rate of the particular formulation. tion. Solutions or Suspensions used for parenteral, intrader The pharmaceutical compositions containing active com mal, or Subcutaneous application can include the following pounds of the present invention may be manufactured in a components: a sterile diluent Such as water for injection, 60 manner that is generally known, e.g., by means of conven saline solution, fixed oils, polyethylene glycols, glycerine, tional mixing, dissolving, granulating, dragee-making, levi propylene glycol or other synthetic solvents; antibacterial gating, emulsifying, encapsulating, entrapping, or lyophiliz agents such as benzyl alcohol or methyl parabens; antioxi ing processes. Pharmaceutical compositions may be dants such as ascorbic acid or sodium bisulfite; chelating formulated in a conventional manner using one or more phar agents such as ethylenediaminetetraacetic acid; buffers such 65 maceutically acceptable carriers comprising excipients and/ as acetates, citrates or phosphates, and agents for the adjust or auxiliaries that facilitate processing of the active com ment oftonicity such as sodium chloride or dextrose. The pH pounds into preparations that can be used pharmaceutically. US 8,962,619 B2 77 78 Of course, the appropriate formulation is dependent upon the Systemic administration can also be by transmucosal or route of administration chosen. transdermal means. For transmucosal or transdermal admin Pharmaceutical compositions suitable for injectable use istration, penetrants appropriate to the barrier to be permeated include sterile aqueous solutions (where water soluble) or are used in the formulation. Such penetrants are generally dispersions and sterile powders for the extemporaneous known in the art, and include, for example, for transmucosal preparation of sterile injectable solutions or dispersion. For administration, detergents, bile salts, and fusidic acid deriva intravenous administration, Suitable carriers include physi tives. Transmucosal administration can be accomplished ological saline, bacteriostatic water, Cremophor ELTM through the use of nasal sprays or Suppositories. For trans (BASF, Parsippany, N.J.) orphosphate buffered saline (PBS). dermal administration, the active compounds are formulated In all cases, the composition must be sterile and should be 10 into ointments, salves, gels, or creams as generally known in fluid to the extent that easy syringeability exists. It must be the art. stable under the conditions of manufacture and storage and The active compounds can be prepared with pharmaceuti must be preserved against the contaminating action of micro cally acceptable carriers that will protect the compound organisms such as bacteria and fungi. The carrier can be a against rapid elimination from the body, Such as a controlled Solvent or dispersion medium containing, for example, water, 15 release formulation, including implants and microencapsu ethanol, polyol (for example, glycerol, propylene glycol, and lated delivery systems. Biodegradable, biocompatible poly liquid polyethylene glycol, and the like), and Suitable mix mers can be used, such as ethylene vinyl acetate, polyanhy tures thereof. The proper fluidity can be maintained, for drides, polyglycolic acid, collagen, polyorthoesters, and example, by the use of a coating Such as lecithin, by the polylactic acid. Methods for preparation of such formulations maintenance of the required particle size in the case of dis will be apparent to those skilled in the art. The materials can persion and by the use of surfactants. Prevention of the action also be obtained commercially from Alza Corporation and of microorganisms can be achieved by various antibacterial Nova Pharmaceuticals, Inc. Liposomal Suspensions (includ and antifungal agents, for example, parabens, chlorobutanol, ing liposomes targeted to infected cells with monoclonal phenol, ascorbic acid, thimerosal, and the like. In many cases, 25 antibodies to viral antigens) can also be used as pharmaceu it will be preferable to include isotonic agents, for example, tically acceptable carriers. These can be prepared according Sugars, polyalcohols such as manitol, Sorbitol, Sodium chlo to methods known to those skilled in the art, for example, as ride in the composition. Prolonged absorption of the inject described in U.S. Pat. No. 4,522,811. able compositions can be brought about by including in the It is especially advantageous to formulate oral or parenteral composition an agent which delays absorption, for example, 30 compositions in dosage unit form for ease of administration aluminum monostearate and gelatin. and uniformity of dosage. Dosage unit form as used herein Sterile injectable solutions can be prepared by incorporat refers to physically discrete units suited as unitary dosages for ing the active compound in the required amount in an appro the Subject to be treated; each unit containing a predetermined priate solvent with one or a combination of ingredients enu quantity of active compound calculated to produce the merated above, as required, followed by filtered sterilization. 35 desired therapeutic effect in association with the required Generally, dispersions are prepared by incorporating the pharmaceutical carrier. The specification for the dosage unit active compound into a sterile vehicle that contains a basic forms of the invention are dictated by and directly dependent dispersion medium and the required other ingredients from on the unique characteristics of the active compound and the those enumerated above. In the case of sterile powders for the particular therapeutic effect to be achieved. preparation of sterile injectable solutions, methods of prepa 40 In therapeutic applications, the dosages of the pharmaceu ration are vacuum drying and freeze-drying that yields a tical compositions used in accordance with the invention vary powder of the active ingredient plus any additional desired depending on the agent, the age, weight, and clinical condi ingredient from a previously sterile-filtered solution thereof. tion of the recipient patient, and the experience and judgment Oral compositions generally include an inert diluent or an of the clinician or practitioner administering the therapy, edible pharmaceutically acceptable carrier. They can be 45 among other factors affecting the selected dosage. Generally, enclosed in gelatin capsules or compressed into tablets. For the dose should be sufficient to result in slowing, and prefer the purpose of oral therapeutic administration, the active ably regressing, the growth of the tumors and also preferably compound can be incorporated with excipients and used in causing complete regression of the cancer. Dosages can range the form of tablets, troches, or capsules. Oral compositions from about 0.01 mg/kg per day to about 5000 mg/kg per day. can also be prepared using a fluid carrier for use as a mouth 50 In preferred aspects, dosages can range from about 1 mg/kg wash, wherein the compound in the fluid carrier is applied per day to about 1000 mg/kg per day. In an aspect, the dose orally and Swished and expectorated or swallowed. Pharma will be in the range of about 0.1 mg/day to about 50 g/day; ceutically compatible binding agents, and/or adjuvant mate about 0.1 mg/day to about 25 g/day; about 0.1 mg/day to rials can be included as part of the composition. The tablets, about 10 g/day; about 0.1 mg to about 3 g/day; or about 0.1 pills, capsules, troches and the like can contain any of the 55 mg to about 1 g/day, in single, divided, or continuous doses following ingredients, or compounds of a similar nature: a (which dose may be adjusted for the patients weight in kg, binder Such as microcrystalline cellulose, gum tragacanth or body surface area in m, and age in years). An effective gelatin; an excipient Such as starch or lactose, a disintegrating amount of a pharmaceutical agent is that which provides an agent such as alginic acid, Primogel, or corn starch; a lubri objectively identifiable improvement as noted by the clinician cant such as magnesium Stearate or Sterotes; a glidant Such as 60 or other qualified observer. For example, regression of a colloidal silicon dioxide; a Sweetening agent Such as Sucrose tumor in a patient may be measured with reference to the or saccharin; or a flavoring agent Such as peppermint, methyl diameter of a tumor. Decrease in the diameter of a tumor salicylate, or orange flavoring. indicates regression. Regression is also indicated by failure of For administration by inhalation, the compounds are deliv tumors to reoccur after treatment has stopped. As used herein, ered in the form of an aerosol spray from pressured container 65 the term “dosage effective manner” refers to amount of an or dispenser, which contains a Suitable propellant, e.g., a gas active compound to produce the desired biological effect in a Such as carbon dioxide, or a nebulizer. subject or cell. US 8,962,619 B2 79 80 The pharmaceutical compositions can be included in a “Prodrugs are intended to include any covalently bonded container, pack, or dispenser together with instructions for carriers that release an active parent drug of the present inven administration. tion in vivo when Such prodrug is administered to a Subject. The compounds of the present invention are capable of Prodrugs in the present invention are prepared by modifying further forming salts. All of these forms are also contemplated 5 functional groups present in the compound in Such a way that within the scope of the claimed invention. the modifications are cleaved, either in routine manipulation As used herein, “pharmaceutically acceptable salts' refer or in vivo, to the parent compound. Prodrugs include com to derivatives of the compounds of the present invention pounds of the present invention wherein a hydroxy, amino, wherein the parent compound is modified by making acid or Sulfhydryl, carboxy or carbonyl group is bonded to any group base salts thereof. Examples of pharmaceutically acceptable 10 salts include, but are not limited to, mineral or organic acid that may be cleaved in vivo to form a free hydroxyl, free salts of basic residues such as amines, alkali or organic salts of amino, free Sulfhydryl, free carboxy or free carbonyl group, acidic residues such as carboxylic acids, and the like. The respectively. pharmaceutically acceptable salts include the conventional Examples of prodrugs include, but are not limited to, esters non-toxic salts or the quaternary ammonium salts of the par 15 (e.g., acetate, dialkylaminoacetates, formates, phosphates, ent compound formed, for example, from non-toxic inorganic Sulfates and benzoate derivatives) and carbamates (e.g., N.N- or organic acids. For example, such conventional non-toxic dimethylaminocarbonyl) of hydroxy functional groups, salts include, but are not limited to, those derived from inor esters (e.g., ethyl esters, morpholinoethanol esters) of car ganic and organic acids selected from 2-acetoxybenzoic, boxyl functional groups, N-acyl derivatives (e.g., N-acetyl) 2-hydroxyethane Sulfonic, acetic, ascorbic, benzene Sulfonic, N-Mannich bases, Schiff bases and enaminones of amino benzoic, bicarbonic, carbonic, citric, edetic, ethane disul functional groups, oximes, acetals, ketals and enol esters of fonic, 1.2-ethane Sulfonic, fumaric, glucoheptonic, gluconic, ketone and aldehyde functional groups in compounds of the glutamic, glycolic, glycolyarsanilic, hexylresorcinic, hydra invention, and the like, See Bundegaard, H., Design of Pro bamic, hydrobromic, hydrochloric, hydroiodic, hydroxyma drugs, p 1-92, Elesevier, N.Y. Oxford (1985). leic, hydroxynaphthoic, isethionic, lactic, lactobionic, lauryl 25 Sulfonic, maleic, malic, mandelic, methane Sulfonic, nap The compounds, or pharmaceutically acceptable salts, sylic, nitric, oxalic, pamoic, pantothenic, phenylacetic, phos esters or prodrugs thereof, are administered orally, nasally, phoric, polygalacturonic, propionic, salicyclic, Stearic, Sub transdermally, pulmonary, inhalationally, buccally, Sublin acetic, succinic, Sulfamic, Sulfanilic, Sulfuric, tannic, tartaric, gually, intraperintoneally, Subcutaneously, intramuscularly, toluene Sulfonic, and the commonly occurring amine acids, 30 intravenously, rectally, intrapleurally, intrathecally and e.g., glycine, alanine, phenylalanine, arginine, etc. parenterally. In one embodiment, the compound is adminis Other examples of pharmaceutically acceptable salts tered orally. One skilled in the art will recognize the advan include hexanoic acid, cyclopentane propionic acid, pyruvic tages of certain routes of administration. acid, malonic acid, 3-(4-hydroxybenzoyl)benzoic acid, cin The dosage regimen utilizing the compounds is selected in namic acid, 4-chlorobenzenesulfonic acid, 2-naphthalene 35 Sulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, accordance with a variety of factors including type, species, 4-methylbicyclo-2.2.2-oct-2-ene-1-carboxylic acid, 3-phe age, weight, sex and medical condition of the patient; the nylpropionic acid, trimethylacetic acid, tertiary butylacetic severity of the condition to be treated; the route of adminis acid, muconic acid, and the like. The present invention also tration; the renal and hepatic function of the patient; and the encompasses salts formed when an acidic proton present in 40 particular compound or salt thereof employed. An ordinarily the parent compound either is replaced by a metalion, e.g., an skilled physician or veterinarian can readily determine and alkali metalion, an alkaline earth ion, or an aluminum ion; or prescribe the effective amount of the drug required to prevent, coordinates with an organic base Such as ethanolamine, counter or arrest the progress of the condition. diethanolamine, triethanolamine, tromethamine, N-methyl Techniques for formulation and administration of the dis glucamine, and the like. 45 closed compounds of the invention can be found in Reming It should be understood that all references to pharmaceu ton: the Science and Practice of Pharmacy, 19" edition, tically acceptable salts include solvent addition forms (sol Mack Publishing Co., Easton, Pa. (1995). In an embodiment, Vates) or crystal forms (polymorphs) as defined herein, of the the compounds described herein, and the pharmaceutically same salt. acceptable salts thereof, are used in pharmaceutical prepara The compounds of the present invention can also be pre 50 pared as esters, for example, pharmaceutically acceptable tions in combination with a pharmaceutically acceptable car esters. For example, a carboxylic acid function group in a rier or diluent. Suitable pharmaceutically acceptable carriers compound can be converted to its corresponding ester, e.g., a include inert solid fillers or diluents and sterile aqueous or methyl, ethyl or other ester. Also, an alcohol group in a organic solutions. The compounds will be present in Such compound can be converted to its corresponding ester, e.g., an 55 pharmaceutical compositions in amounts sufficient to pro acetate, propionate or other ester. vide the desired dosage amount in the range described herein. The compounds of the present invention can also be pre All percentages and ratios used herein, unless otherwise pared as prodrugs, for example, pharmaceutically acceptable indicated, are by weight. Other features and advantages of the prodrugs. The terms “pro-drug and “prodrug” are used inter present invention are apparent from the different examples. changeably herein and refer to any compound which releases 60 The provided examples illustrate different components and an active parent drug in vivo. Since prodrugs are known to methodology useful in practicing the present invention. The enhance numerous desirable qualities of pharmaceuticals examples do not limit the claimed invention. Based on the (e.g., Solubility, bioavailability, manufacturing, etc.), the present disclosure the skilled artisan can identify and employ compounds of the present invention can be delivered in pro other components and methodology useful for practicing the drug form. Thus, the present invention is intended to cover 65 prodrugs of the presently claimed compounds, methods of present invention. delivering the same and compositions containing the same.

US 8,962,619 B2 111 112 -continued HN 21 NN N Null? N / O NYO

Ya O 10 HN O N H NHHCI

15 'H-NMR (400 MHz, CDC1) 8: 7.92-7.87 (im, 1H), 7.68 7.66 (m, 1H), 7.57-7.53 (m. 1H), 7.35 (t, 1H, J=7.6 Hz), 7.14 (brs, 1H), 4.10-4.03 (m, 2H), 3.46 (ddd, 2H, J=11.5, 11.5.2.8 To a solution of tert-butyl (1-4-2-(2-aminopyridin-3-yl)- Hz), 2.51-2.42 (m. 1H), 1.97-1.80 (m, 4H), 1.34 (s, 12H): 5-3-(1,4-dioxan-2-ylcarbonyl)aminolphenyl-3H-imi LCMS: 332 M+H. dazo[4,5-b]pyridin-3-yl)phenyl)cyclobutyl)carbamate (38 Step 2: Synthesis of tert-butyl (1-4-2-(2-aminopy mg) in dichloromethane (5 mL) was added trifluoroacetic ridin-3-yl)-5-3-(tetrahydro-2H-pyran-4-ylcarbonyl) acid (1 mL) dropwise at room temperature. After being stirred aminolphenyl-3H-imidazo[4,5-b]pyridin-3-yl) at the same temperature for 1 h, the reaction mixture was phenyl)cyclobutyl) carbamate concentrated under reduced pressure. Dichloromethane (5 mL) and MP-carbonate (Argonaout technologies 2.91 mmol/ 25 g, 400 mg) were added. The mixture was stayed for 30 min. with occasional shaking then filtered. The filtrate was con centrated under reduced pressure. To a solution of the residue in dichloromethane (5 mL) and methanol (0.4 mL) was added 30 hydrogen chloride in ethyl acetate (4M, 0.072 mL) at room Os-O temperature. After being stirred for 10 min., the mixture was HN concentrated under reduced pressure. The resulting Solid was suspended with ethyl acetate and filtered to give the product O)-() N (24 mg). 35 C N O 'H-NMR (400 MHz, DMSO-d) 8:9.91 (s, 1H), 8.90-8.74 (m,3H), 8.42-8.33 (m, 2H), 8.17-7.90 (m,3H), 7.80-7.64 (m, KPO4, dioxane, H2O 7H), 7.41 (t, 1H, J=8.0 Hz), 6.83-6.76 (m, 1H), 4.26 (dd. 1H, 80 deg., 18 h, 51% J=9.4, 3.0 Hz), 3.99-3.87 (im, 2H), 3.78-3.69 (m, 2H), 3.61 3.53 (m, 2H), 2.72-2.55 (m, 4H), 2.29-2.15 (m, 1H), 1.93 40 1.80 (m, 1H); LCMS: 562 M+H. Example 21

Synthesis of N-(3-3-4-(1-aminocyclobutyl)phe 45 nyl-2-(2-aminopyridin-3-yl)-3H-imdazo[4,5-b]pyri din-5-yl)phenyl)tetrahydro-2H-pyran-4-carboxamide hydrochloride of ( ) Step 1: Synthesis of N-3-(4.4.5.5-tetramethyl-1,3,2- 50 dioxaborolan-2-yl)phenyltetrahydro-2H-pyran-4- HN O R. x. carboxamide

To a solution of 3-(4.4.5.5-tetramethyl-1,3,2-dioxaboro O lan-2-yl)aniline (220 mg) and triethylamine (0.182 mL) in 55 N,N-dimethylacetamide (1.5 mL) chilled to 0°C. was added To a suspension of tert-butyl (1-4-2-(2-aminopyridin-3- tetrahydro-2H-pyran-4-carbonyl chloride (prepared from tet yl)-5-chloro-3H-imidazo[4,5-b]pyridin-3-yl) rahydro-2H-pyran-4-carboxylic acid (156 mg) and oxalyl phenyl)cyclobutyl)carbamate (50mg) and N-3-(4.4.5.5-tet chloride (0.15 mL) in dichloromethane (2.5 mL)) dropwise. 60 ramethyl-1,3,2-dioxaborolan-2-yl)phenyltetrahydro-2H After being stirred at room temperature for 1 h, the reaction pyran-4-carboxamide (67 mg) and potassium phosphate (65 mixture was diluted with ethyl acetate and washed with 1 M mg) and PdCl2(dppf)CH2Cl (8 mg) in dioxane (2 mL) and citric acid aqueous Solution (3 times) then brine. The organics HO(0.2 mL) was degassed for 5 minutes and then heated to were dried over anhydrous sodium Sulfate and concentrated 80° C. for 18 h. The reaction mixture was concentrated under under reduced pressure. Purification by column chromatog 65 reduced pressure. Purification by column chromatography (2 raphy (10-100% ethyl acetate in hexanes) gave the product times: 0-10% methanol in chloroform then 0-10% methanol (267 mg, 81%). in ethyl acetate) gave the product (34 mg, 51%).

US 8,962,619 B2 117 118 -continued phase was dried and evaporated. The residue was purified with silica gel column chromatography (hexane/ethyl acetate 10:0 to 1:9) to give the titled compound (2.2g, 100%). OH 'H-NMR (CDC1,) 8: 7.08 (1H, td, J=8.2, 3.1 Hz), 7.03 (1H, Br N s), 6.93 (1H, d, J=8.0 Hz), 6.84 (1H, d, J=8.0 Hz), 3.72 (2H, d, J=10.3 Hz), 3.08 (3H, d, J=2.9 Hz), 2.96 (3H, d, J=2.3 Hz), 2.76 (2H, t, J=12.3 Hz), 2.64 (1H, dd, J=14.9, 3.8 Hz), 1.94 (2H, q, J=12.4 Hz), 1.81 (2H, d, J=13.2 Hz); LC/MS: 311 MW = 284.15 10 M+H. EXMS = 283.02 MF = C12H14BrNO2 Step 4: Synthesis of N,N-dimethyl-1-3-(4.4.5.5- tetramethyl-1,3,2-dioxaborolan-2-yl)phenylpiperi To a solution of ethyl 1-(3-bromophenyl)piperidine-4-car dine-4-carboxamide boxylate (5.6 g., 18 mmol) in THF/MeOH (360 mL, 1:1), was 15 added a solution of NaOH (1M, 90 mL) at rt. The mixture was heated at 50° C. for 30 min and organic solvent was removed under reduced pressure. The water solution was mixed with O ice and 1M HCl (90 mL) and extracted with dichloromethane/ KOAc(3 eq) methanol. The combined organic phase was dried over unhy N1 PdCl2dppf(4 mol%) pinacol diborane(1.5 eq) drous Na2SO4 and evaporated. The residue was solidified He with dichloromethane/hexane to give the titled compound Br N DMF 80° C. 6 (4.4g, 85%) as a white solid. O.2M 'H-NMR (DMSO-D) 8: 12.23 (1H, s), 7.13 (1H, t, J=8.0 25 Hz), 7.06 (1H, t, J=2.3 Hz), 6.93 (1H, dd, J=8.3, 2.0 Hz), 6.88 MW = 31122 (1H, dd, J=7.2, 1.4 Hz), 3.65 (2H, dt, J=12.6, 3.4 Hz), 3.33 EXMS = 310.07 (1H, s), 2.79 (2H, td, J=12.2, 2.3 Hz), 1.87 (2H, dd, J=13.2, MF = C14H19BrN2O 2.9 Hz), 1.60 (2H, ddd, J-24.6, 11.5, 4.0 Hz). 30 Step 3: Synthesis of 1-(3-bromophenyl)-N,N-dim ethylpiperidine-4-carboxamide

35 O 1) thionyl chloride, DCM OH 2) dimethylamine HCl, TEA, DCM O Her Br N 40 O N1 o-B N MW = 284.15 EXMS = 283.02 45 MF = C12H14BrNO2 O MW = 358.28 EXMS = 358.24 N1 MF = C2OH31BN2O3 50 Br N

To a solution of 1-(3-bromophenyl)-N,N-dimethylpiperi dine-4-carboxamide (2.2g, 7.3 mmol) were added bis(pina MW = 31122 55 EXMS = 310.07 colato) diboron (2.8 g., 11 mmol), PdCl2(dppf) (237 mg, 0.29 MF = C14H19BN2O mmol), and KOAc (2.2g, 22 mmol). The mixture was heated at 80°C. for 6 hand diluted with water and ethyl acetate then separated. The organic phase was dried and evaporated. The To a solution of 1-(3-bromophenyl)piperidine-4-carboxy residue was purified with silica gel column chromatography lic acid (2g, 7.0 mmol) in dioxane (70 mL), was added thionyl 60 (hexane/ethyl acetate 10:0 to 0:10) to give the titled com chloride (2.6 mL, 35 mmol) at 0°C. The mixture was kept at rt for 1 hand evaporated. The residue was co-evaporated with pound (1.94 g, 75%). dichloromethane and dissolved in dichloromethane (5 mL). H-NMR (CDC1) 8: 7.39 (1H, d, J=2.3 Hz), 7.30 (1H, d, To this mixture, were added the mixture of dimethylamine J=6.9 Hz), 7.27 (1H, t, J–74 Hz), 7.06-7.04 (1H, m), 3.78 HCl salt (1.7g, 21 mmol), triethylamine (4.9 mL, 35 mmol) 65 (2H, d, J=12.0 Hz), 3.09 (3H, s), 2.97 (3H, s), 2.74 (2H, td, and dichloromethane (20 mL) at 0°C. The mixture was stirred J=12.2, 2.7 Hz), 2.61 (1H, tt, J=11.5, 3.4 Hz), 1.97 (2H, ddd, at rt for 1 h and diluted with water and separated. The organic J=24.8, 12.5, 3.9 Hz), 1.83 (2H, d, J=13.2 Hz), 1.34 (12H, s).

US 8,962,619 B2 121 To a solution of tert-butyl (1-4-2-(2-aminopyridin-3-yl)- -continued 5-chloro-3H-imidazo[4,5-b]pyridin-3-yl) O Pd(PPh3)4 phenyl)cyclobutyl)carbamate (400 mg. 0.81 mmol) in DMF toluenEtOH (8.1 mL)/water (0.45 mL), were added N,N-dimethyl-1-3- N SatNaHCO3 Hs (4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenylpiperi O -B N dine-4-carboxamide. (584 mg, 1.6 mmol), AMPHOS (58 mg, 0.08 mmol) and NaCO (95 mg. 0.9 mmol). The mixture was heated at 160° C. for 1 h in microwave oven. The mixture was 10 MW = 358.28 evaporated under reduced pressure and the residue was puri EXMS = 358.24 fied with silica gel column chromatography (hexane/ethyl acetate/methanol, 10:0:0 to 0:10:1) to give tert-butyl (1-4- 2-(2-aminopyridin-3-yl)-5-3-4-(dimethylcarbamoyl)pip 15 eridin-1-yl)phenyl-3H-imidazo[4,5-b]pyridin-3-yl) phenyl)cyclobutyl)carbamate. This compound was deprotected with methanol (15 mL) and 4M HCl in ethyl acetate (15 mL) for 18 hat rt. The mixture was solidified with diethyl ether and the solid was purified with HPLC and HCl was added to the fraction and evaporated then solidified with O diethyl ether to give the titled compound (281 mg, 50%) as a NO N N N 2 white solid. 25 N 2 N NH 'H-NMR (DMSO-D) 8: 8.94 (3H, s), 8.52-8.39 (1H, m), 8.43 (1H, d. J=8.6Hz), 8.18 (1H, dd, J=6.3, 1.7 Hz), 8.13 (1H, 30 d, J=8.6 Hz), 7.94 (1H, dd, J=7.4, 1.7 Hz), 7.77 (2H, d, J=8.0 Hz), 7.70 (2H, d, J=8.6 Hz), 6.93 (1H, dd, J=74, 6.3 Hz), 3.74-3.64 (2H, m), 3.09 (4H, s), 2.85 (3H, s), 2.69-2.58 (4H, Boc m), 2.28-2.19 (1H, m), 1.95-1.81 (3H, m); LCMS: 587 35 N M+H.

40 To a solution of tert-butyl (1-4-(6-chloro-3-nitropyridin Alternative Synthesis Method Step 1: Synthesis of 2-yl)aminolphenyl)cyclobutyl)carbamate (4.1 g, 9.8 mmol) tert-butyl (1-4-(6-3-4-(dimethylcarbamoyl)pip 45 in toluene/EtOH/saturated NaHCO3 (200/200/20 mL), were eridin-1-yl)phenyl)-3-nitropyridin-2-yl)aminol added N,N-dimethyl-1-3-(4.4.5.5-tetramethyl-1,3,2-diox phenyl)cyclobutyl)carbamate aborolan-2-yl)phenylpiperidine-4-carboxamide. (4.2 g, 12 mmol), Pd(PPh4) (563 mg, 0.49 mmol). The mixture was 50 heated at 100° C. for 18 h. The mixture was filtered through NO celite, separated and evaporated. The residue was purified with silica gel column chromatography (hexane/ethyl acetate/methanol, 85:15:1.5) to give the titled compound as a C N NH 55 pale red solid (5.4g, 91%)

'H-NMR (CDC1) 8: 10.31 (1H, s), 8.56 (1H, d, J=9.2 Hz), 60 7.77 (3H, d, J=8.0 Hz), 7.67 (1H, dd, J=12.0, 8.0 Hz), 7.55 Boc NN H (1H, t, J=7.4 Hz), 7.47 (5H, d, J=6.9 Hz), 7.36 (1H, t, J=8.0 MW = 418.87 Hz), 7.30 (1H, d, J=9.2 Hz), 7.08 (1H, d, J=8.0 Hz), 5.21 (1H, EXMS = 418.14 s), 3.85 (2H, d, J–12.6 Hz), 3.11 (3H, s), 2.98 (3H, s), 2.86 65 MF = C20H23CN4O4 (2H, t, J=11.2 Hz), 2.75-2.66 (1H, m), 2.58-2.51 (3H, m), 2.14-1.95 (3H, m), 1.90-1.79 (3H, m), 1.4.6-1.20 (9H, brm).

US 8,962,619 B2 133 134 Step 2: Coupling A mixture of methyl (1-(3-bromophenyl)piperidin-4-yl) carbamate (127 mg, 0.406 mmol), bis(pinacolato) diboron (1-4-2-(2-aminopyridin-3-yl)-5-3-(cis-2,6-dimethyl (124 mg. 0.487 mmol), Pd(dppf)Cl. DCM (17 mg, 0.0203 morpholin-4-yl)phenyl-3H-imidazo[4,5-b]pyridin-3-yl) mmol) and potassium acetate (120 mg, 1.22 mmol) in diox phenyl)cyclobutyl)carbamate. A mixture of tert-butyl (1-4- ane (4 mL) was heated at 80°C. for 17.5 hours under nitrogen. 2-(2-aminopyridin-3-yl)-5-chloro-3H-imidazo[4,5-b] After cooling to room temperature, the mixture was diluted pyridin-3-ylphenyl)cyclobutyl)carbamate (101 mg, 0.205 with EtOAc and filtered through a Celite pad. The combined mmol), tert-butyl {1-3-(4.4.5.5-tetramethyl-1,3,2-diox filtrate and washings were concentrated. The residue was aborolan-2-yl)phenylpiperidin-4-yl)carbamate (99 mg, purified by silica gel column chromatography (hexane/ 0.246 mmol), Bis(di-tert-butyl (4-dimethylaminophenyl) AcOEt=75:25->50:50) to afford desired product (102 mg, 10 70%) as pale yellow solid. phosphine)dichloropalladium(II) (15 mg, 0.0205 mmol), and 500 MHz H-NMR (CDC1) 8: 7.51-748 (m. 1H), 7.37 2M NaCO, aq. (0.113 mL, 0.226 mmol) in DMF (3.5 mL) 7.35 (m. 1H), 7.27-7.24 (m, 1H), 7.06-7.02 (m, 1H), 5.86 was treated with microwave (160° C. for 1 hour). The mixture 5.80 (m, 1H), 3.69-3.63 (m, 5H), 2.89-2.82 (m, 2H), 2.05 was diluted with AcOEt, then washed with water(x3), brine, 1.99 (m, 2H), 1.65-1.56 (m, 2H), 1.34 (s, 12H); LCMS: 361 dried over NaSO, then filtrated. The filtrate was concen M+H. trated and the residue was purified by preparative thin-layer 15 chromatography (AcOEt only x2), and further purified by Synthesis of methyl (1-(3-3-4-(1-aminocyclobutyl) preparative thin-layer chromatography (CH.Cl/AcOEt=5: phenyl-2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b] 2x2) to afford desired product (66 mg, 51%) as pale yellow pyridin-5-yl)phenyl)piperidin-4-yl)carbamate trihy oil. drochloride Step 3: de-Boc N Starting material (66 mg, 0.105 mmol) in MeOH (1 mL) was added 4N HC1-dioxane (3 mL) and stirred at rit for 16 Sl-N N hours. The mixture was concentrated to afford desired prod N uct (59 mg, 83%) as yellow solid. 500MHz 'H-NMR (DMSO-d) 8:8.85-8.81 (m, 2H), 8.34 C N elNY \ A (d. J=8.6 Hz, 1H), 8.15-8.12 (m, 1H), 8.11-8.09 (m, 1H), 8.07 (d. J=8.6 Hz, 1H), 7.83-7.78 (m, 1H), 7.75 (d. J=8.6 Hz, 2H), 7.68 (d. J=8.6 Hz, 2H), 7.52-746 (m. 1H), 7.35-7.32 (m, 1H), 7.09-7.03 (m. 1H), 6.86-6.80 (m, 1H), 3.85-3.81 (m, 4H), 3.50-3.45 (m. 1H), 3.27-3.21 (m, 2H), 2.92-2.83 (m, 2H), 2.68-2.58 (m, 4H), 2.28-2.18 (m, 2H), 2.03-1.97 (m, 2H), 1.89-1.82 (m, 2H), 1.73-1.64 (m, 2H); LCMS: 531 M+H. Q Example 32 O Synthesis of methyl (1-(3-3-4-(1-aminocyclobutyl) d phenyl-2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b] Yo pyridin-5-yl)phenyl)piperidin-4-yl)carbamate trihy 40 drochloride Step 1: Synthesis of methyl {1-3-(4.4.5.5-tetram ethyl-1,3,2-dioxaborolan-2-yl)phenylpiperidin-4- O coupling de-Boc yl)carbamate 45 NN O 50 N Sl-NN N

N elNY \ / C -e- c 55 N

60 C C N y y N O N 65

US 8,962,619 B2 137 138 500MHz H-NMR (CDC1)8: 7.09(1H, t, J=8.3 Hz), 7.04 A mixture of N-1-(3-bromophenyl)piperidin-4-yl)-N-me (1H, t, J=1.7 Hz), 6.95-6.93 (1H, m), 6.83 (1H, dd, J=8.3, 1.7 thylacetamide (68 mg 0.217 mmol), bis(pinacolato) diboron Hz), 5.37-5.35 (1H, m), 3.99-3.91 (1H, m), 3.64-3.60 (2H, (66 mg, 0.260 mmol), Pd(dppf)Cl. DCM (9 mg, 0.0109 m), 2.88 (2H, td, J=12.6, 2.3 Hz), 2.06-2.01 (2H, m), 1.99 mmol) and potassium acetate (64 mg., 0.651 mmol) in diox (3H, s), 1.56-148 (2H, m); LCMS: 297, 299 M+H. ane (3 mL) was heated at 80°C. for 13 hours under nitrogen. After cooling to room temperature, the mixture was diluted Step 2: Synthesis of N-1-(3-bromophenyl)piperidin with EtOAc and filtered through a Celite pad. The combined 4-yl)-N-methylacetamide filtrate and washings were concentrated. The residue was purified by silica gel column chromatography (hexane/ Br 10 AcOEt=35:65->0:100) to afford desired product (61 mg, 78%) as white solid. 500 MHZ H-NMR (CDC1) 8: 741-7.38 (m, 1H), 7.35 7.28 (m, 2H), 7.06-7.03 (m, 1H), 4.68-4.61 (m, 1H), 3.84 3.76 (m, 2H), 2.88 (s. 2H), 2.85 (s, 1H), 2.83-2.76 (m, 2H), C. C.N 15 2.16 (s, 1H), 2.11 (s. 2H), 2.02-1.92 (m. 1H), 1.83-1.76 (m, 2H), 1.72-1.69 (m, 1H), 1.34-1.34 (m, 12H); LCMS: 359 M+H. Synthesis of N-1-(3-3-4-(1-aminocyclobutyl)phe CHN C nyl-2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b] 1 1. N pyridin-5-yl)phenyl)piperidin-4-yl)-N-methylaceta O O mide trihydrochloride FW:297.2 FW: 311.2 Exact Mass: 296.05 Exact Mass: 310.07 Formula: C3H1BrNO Formula: C14H10 BrNO 25 A solution of N-1-(3-bromophenyl)-4-piperidyllaceta HN mide (11.6 g. 39.2 mmol) in THF (392 mL) was added to Sl-N N sodium hydride (6.84 g, 55% in mineral oil, 157 mmol) N portionwise at 0°C. and stirred at room temperature for 10 30 elN/ \ / minutes under nitrogen. The reaction mixture was added to C N iodomethane (7.32 mL, 118 mmol) at 0° C. and stirred at room temperature for 6 hours. The mixture was slowly quenched with water at 0°C. and extracted with EtOAc. The -- organic layer was washed with brine, dried over NaSO and 35 filtered through a Celite pad. The combined filtrate and wash ings were concentrated. The residue was purified by silica gel NH column chromatography (hexane/AcOEt=80:20->0:100) to afford desired product (12.6 g., quant.) as brown oil. > 500 MHz H-NMR (CDC1) 8: 7.13-7.08 (1H, m), 7.06 40 7.04 (1H, m), 6.99-6.93 (1H, m), 6.86-6.82 (1H, m), 4.70 4.63 (1H, m), 3.79-3.65 (2H, m), 2.90-2.78 (5H, m), 2.17 (1H, As s), 2.11 (2H, s), 2.00-1.92 (0.5H, m), 1.82-1.73 (2H, m), 1.71-1.67 (1.5H, m); LCMS: 311, 313 M+H. 45 Step 3: Synthesis of N-methyl-N-(1-3-(4.4.5.5-tet ramethyl-1,3,2-dioxaborolan-2-yl)phenylpiperidin 4-yl)acetamide 50

O O Br B d Yo C NO 55 -e- N N coupling de-Boc 60

ON C O N ry ry 65 1. Y US 8,962,619 B2 139 140 -continued 4H), 2.26-2.18 (m, 2H), 2.10 (s, 1H), 2.02 (s. 2H), 1.90-1.82 HN (m. 2H), 1.82-1.80 (m. 1H), 1.67-1.60 (m, 2H); LCMS: 587 N SN M+H. rN-y 5 el? \ / Example 35 10 N Synthesis of 3-3-4-(1-aminocyclobutyl)phenyl-5- 3-(4-morpholin-4-ylpiperidin-1-yl)phenyl-3H-imi dazo[4,5-b]pyridin-2-yl)pyridin-2-amine tetrahydro chloride 15 -N- Step 1: Synthesis of 4-1-3-(4.4.5.5-tetramethyl-1,3, O 2-dioxaborolan-2-yl)phenylpiperidin-4- y1}morpholine

Step 4: Coupling 25 Br (1-4-2-(2-aminopyridin-3-yl)-5-3-(cis-2,6-dimethyl morpholin-4-yl)phenyl-3H-imidazo[4,5-b]pyridin-3-yl) phenyl)cyclobutyl)carbamate. A mixture of tert-butyl (1-4- 30 C.N c.N 2-(2-aminopyridin-3-yl)-5-chloro-3H-imidazo[4,5-b] pyridin-3-ylphenyl)cyclobutyl)carbamate (56 mg, 0.113 mmol), N-methyl-N-(1-3-(4.4.5.5-tetramethyl-1,3,2-diox aborolan-2-yl)phenylpiperidin-4-yl)acetamide (61 ng, C C 0.170 mmol), Bis(di-tert-butyl (4-dimethylaminophenyl) 35 phosphine)dichloropalladium(II) (8 mg, 0.0113 mmol), and C O) C O) 2M NaCO aq. (0.062 mL, 0.124 mmol) in DMF (2.5 mL) was treated with microwave (160° C. for 1 hour). The mixture was diluted with AcOEt, then washed with water(x3), brine, 40 dried over NaSO, then filtrated. The filtrate was concen trated and the residue was purified by preparative thin-layer A mixture of 4-1-(3-bromophenyl)piperidin-4-ylmor chromatography (AcOEt/MeOH-20:1), and further purified pholine (326 mg, 1.01 mmol), bis(pinacolato)diboron (306 by preparative thin-layer chromatography (CH2Cl2/ 45 mg, 1.21 mmol), Pd(dppf)Cl. DCM (41 mg 0.0505 mmol) MeOH-20:1 x2) to afford desired product (14 mg, 18%) as and potassium acetate (297 mg, 3.03 mmol) in dioxane (3 yellow solid. mL) was heated at 80° C. for 18 hours under nitrogen. After cooling to room temperature, the mixture was diluted with 50 EtOAc and filtered through a Celite pad. The combined fil trate and washings were concentrated. The residue was puri Step 5: de-Boc fied by silica gel column chromatography (CH2Cl2/ MeOH=100:0-95:5) to afford desired product (444 mg. Starting material (14 mg. 0.0204 mmol) in MeCH (1 mL) 55 quant) as dark brown oil. was added 4N HC1-dioxane (3 mL) and stirred at rit for 14 hours. The mixture was concentrated to afford desired prod uct (19 mg, quant) as pale yellow solid. 60 500 MHz H-NMR (CDC1) 8: 7.38 (d. J=2.3 Hz, 1H), 500 MHZ H-NMR (DMSO-d) 8: 8.86-8.82 (m, 2H), 7.30-7.24 (m, 2H), 7.05-7.03 (m, 1H), 3.78-3.70 (m, 4H), 8.41-8.37 (m, 1H), 8.36-8.23 (m, 2H), 8.27 (dd, J=10.0 Hz 2.72 (td, J=12.0 Hz and 2.3 Hz, 2H), 2.59 (t, J=4.0 Hz, 4H), and 5.0 Hz, 1H), 8.15 (d. J=6.3 Hz, 1H), 7.89-7.87 (n, 1H), 2.35-2.29 (m, 1H), 1.95-1.93 (m, 4H), 1.66 (ddd, J-24.1 Hz 7.75 (dd, J=8.6 Hz and 2.9 Hz, 2H), 7.69 (d. J=8.6 Hz, 2H), 65 7.51-7.36 (m, 2H), 6.88 (t, J=6.9 HZ, 1H), 3.74-3.64 (m, 2H), and 12.0 Hz and 4.0 Hz, 2H), 1.33 (s, 12H); LCMS: 373 3.50-345 (m, 3H), 2.85 (s. 2H), 2.70 (s, 1H), 2.68-2.57 (m, M+H. US 8,962,619 B2 141 142 Synthesis of 3-3-4-(1-aminocyclobutyl)phenyl-5- treated with microwave (160° C. for 2 hours). The mixture 3-(4-morpholin-4-ylpiperidin-1-yl)phenyl-3H-imi was diluted with AcOEt, then washed with water(x3), brine, dazo[4,5-b]pyridin-2-yl)pyridin-2-amine tetrahydro dried over NaSO, then filtrated. The filtrate was concen chloride trated and the residue was purified by preparative thin-layer chromatography (CH.Cl/MeOH=10:1), and further purified by NH silica gel column chromatography (AcOEt only) to afford desired product (26 mg, 36%) as yellow solid. N Sl-N SN Step 3: de-Boc Cy-K) 10 Starting material (26 mg 0.0371 mmol) in MeOH (1 mL) C N was added 4N HC1-dioxane (3 mL) and stirred at rit for 67 hours. The mixture was concentrated to afford desired prod 15 uct (25 mg, 90%) as yellow solid. 500MHz H-NMR (DMSO-d) 8:8.89-8.82 (m, 2H), 8.35 (d. J=8.6 Hz, 1H), 8.14 (dd, J=6.9 Hz and 1.7 Hz, 1H), 8.09 (d. J=8.6 Hz, 1H), 7.85-7.81 (m, 1H), 7.76 (d. J=8.6 Hz, 2H), 7.68 (d. J=8.6 Hz, 2H), 7.51-7.49 (m, 1H), 7.33 (t, J=8.6 Hz, Q 1H), 7.21-7.12 (m. 1H), 7.10-7.06 (m. 1H), 6.88-6.83 (m, O 1H), 4.01-3.94 (m, 3H), 3.89-3.84 (m, 2H), 3.41-3.33 (m, 2H), 3.14-3.06 (m, 2H), 2.82-2.73 (m, 2H), 2.66-2.60 (m, 3H), 2.26-2.19 (m, 2H), 1.88-1.81 (m, 2H), 1.28-1.13 (m, lYo X. 3H), 0.87-0.77 (m, 2H); LCMS: 601 M+H. 25

N coupling de-Boc Example 36 30 Synthesis of 8-(3-3-4-(1-aminocyclobutyl)phenyl 2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b]pyridin C 5-yl)phenyl)-8-azabicyclo[3.2.1]octan-3-ol trihydro chloride O 35 N Step 1: Synthesis of 8-3-(4.4.5.5-tetramethyl-1,3,2- N EN dioxaborolan-2-yl)phenyl-8-azabicyclo[3.2.1]octan N-y 3-ol 40 N elN/ \ /

45 O

50 C. C. CO

55 Step 2: Coupling as as (1-4-2-(2-aminopyridin-3-yl)-5-3-(cis-2,6-dimethyl morpholin-4-yl)phenyl-3H-imidazo[4,5-b]pyridin-3-yl) phenyl)cyclobutyl)carbamate. A mixture of tert-butyl (1-4- 60 A mixture of 8-(3-bromophenyl)-8-azabicyclo[3.2.1]oc 2-(2-aminopyridin-3-yl)-5-chloro-3H-imidazo[4,5-b] tan-3-ol (58 mg, 0.206 mmol), bis(pinacolato)diboron (63 pyridin-3-ylphenyl)cyclobutyl)carbamate (50 mg, 0.102 mg, 0.247 mmol), Pd(dppf)Cl. DCM (8 mg, 0.0103 mmol) mmol), 4-1-3-(4.4.5.5-tetramethyl-1,3,2-dioxaborolan-2- and potassium acetate (61 mg, 0.618 mmol) in dioxane (3 yl)phenylpiperidin-4-yl)morpholine (114 mg. 0.306 mmol), mL) was heated at 80°C. for 14.5 hours under nitrogen. After Bis(di-tert-butyl (4-dimethylaminophenyl)phosphine) 65 cooling to room temperature, the mixture was diluted with dichloropalladium(II) (14 mg. 0.0204 mmol), and 2M EtOAct, washed with water, brine, dried over NaSO, then NaCO aq. (0.056 mL, 0.112 mmol) in DMF (2.5 mL) was filtrated. The filtrate was concentrated and the residue was US 8,962,619 B2 143 144 purified by silica gel column chromatography (hexane/ microwave (160° C. for 1 hour). The mixture was diluted with AcOEt=85:15->75:25) to afford desired product (37 mg, AcOEt, then washed with water(x3), brine, dried over 55%) as colorless oil. NaSO, then filtrated. The filtrate was concentrated and the residue was purified by preparative thin-layer chromatogra Synthesis of 8-(3-3-4-(1-aminocyclobutyl)phenyl phy (CHC1/MeOH=20:1x2) to afford desired product (8 2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b]pyridin mg, 11%) as yellow solid. 5-yl)phenyl)-8-azabicyclo[3.2.1]octan-3-ol trihydro chloride Step 3: de-Boc 10 Starting material (8 mg 0.0122 mmol) in MeOH (0.5 mL) N was added 4N HC1-dioxane (2 mL) and stirred at rit for 18 hours. The mixture was concentrated to afford desired prod N EN uct (10 mg, quant) as yellow solid. Cy-K) 15 500 MHZ H-NMR (DMSO-d) 8: 8.92-8.87 (2H, m), C N elN/ \ / 8.53-8.43 (m, 1H),8.38-8.32 (m, 1H), 8.17 (d. J=7.4 Hz, 1H), 8.08-8.02 (m, 1H), 7.92 (d. J=7.4 Hz, 1H), 7.76 (d. J=9.2 Hz, -- 2H), 7.69 (d. J=9.2 Hz, 2H), 7.62-7.54 (m, 1H), 7.37-7.22 (m, 2H), 6.91 (t, J=7.4 Hz, 1H), 4.29-4.25 (m, 1H), 3.72-3.65 (m, 4H), 3.51-3.45 (m, 4H), 2.63 (t, J=8.0 Hz, 3H), 2.36-2.31 (m, 2H), 2.25-2.19 (m, 1H), 1.95-1.90 (m, 2H), 1.88-1.80 (m, N 1H), 1.63-1.54 (m, 1H); LCMS: 558 M+H.

25 O Example 37 dYo X. Synthesis of N-1-(3-3-4-(1-aminocyclobutyl)phe 30 coupling de-Boc nyl-2-(2-aminopyridin-3-yl)-3H-imidazo[4,5-b] N pyridin-5-yl)phenyl)piperidin-4-yl)-2-methoxyaceta mide trihydrochloride

35 Step 1: Synthesis of 2-methoxy-N-(1-3-(4.4.5.5- tetramethyl-1,3,2-dioxaborolan-2-yl)phenylpiperi O din-4-yl)acetamide N

40 Sl-NN N

N elN/ \ / O

Br 45 B O

N

N 50

O

55 Step 2: Coupling (1-4-2-(2-aminopyridin-3-yl)-5-3-(cis-2,6-dimethyl morpholin-4-yl)phenyl-3H-imidazo[4,5-b]pyridin-3-yl) A mixture of N-1-(3-bromophenyl)piperidin-4-yl)-2- phenyl)cyclobutyl)carbamate. A mixture of tert-butyl (1-4- 60 methoxyacetamide (66 mg, 0.202 mmol), bis(pinacolato)di 2-(2-aminopyridin-3-yl)-5-chloro-3H-imidazo[4,5-b] boron (61 mg 0.242 mmol), Pd(dppf)Cl. DCM (8 mg, pyridin-3-ylphenyl)cyclobutyl)carbamate (55 mg, 0.112 0.01.01 mmol) and potassium acetate (59 mg, 0.606 mmol) in mmol), 8-3-(4.4.5,5-tetramethyl-1,3,2-dioxaborolan-2-yl) dioxane (3 mL) was heated at 80° C. for 15.5 hours under phenyl-8-azabicyclo[3.2.1]octan-3-ol (37 mg, 0.112 mmol), nitrogen. After cooling to room temperature, the mixture was Bis(di-tert-butyl (4-dimethylaminophenyl)phosphine) 65 diluted with EtOAct, washed with water, brine, dried over dichloropalladium(II) (8 mg, 0.01 12 mmol), and 2MNaCO, NaSO, then filtrated. The filtrate was concentrated and the aq. (0.062 mL, 0.123 mmol) in DMF (3 mL) was treated with residue was purified by silica gel column chromatography

US 8,962,619 B2 152

Br HN Sl-N N N

C N elN/ \ / O -- a y -- MVS O O X-d N H MW = 290.1768 EM = 288.9772 MF = C10H12BrNO2S MW = 490.9846 EM = 490.1884

O

O O BNo. N M B-B He M M O O

MW = 253.9386 25 EM = 254.1861 MF = C12H24B2O4

O 30 B NO MW = 337.2421 EM = 337.1519 MF = C16H24BNO4S

35

OS MV O O 40 MW = 337.2421 co, (T)a y EM = 337.1519 N X-d 45 R. Step 2: 4-3-(4,4,5,5-Tetramethyl-1,3,2-dioxaboro lan-2-yl)phenylthiomorpholine 1,1-dioxide 50 A mixture of 4-(3-bromophenyl)thiomorpholine 1,1-diox ide (281 mg 0.968 mmol), bis(pinacolato)diboron (270 mg. Step 3: tert-Butyl (1-(4-2-(2-aminopyridin-3-yl)-5- 1.06 mmol), Pd(dppf)Cl. DCM (158 mg, 0.194 mmol) and 3-(1,1-dioxidothiomorpholin-4-yl)phenyl-3H-imi potassium acetate (294 mg, 2.90 mmol) in dioxane (5 mL) 55 dazo[4,5-b]pyridin-3-yl)phenyl)cyclobutylcarbam was heated at 80°C. for 11 hours under nitrogen. After cool ate ing to room temperature, the mixture was diluted with EtOAc A mixture of tert-butyl (1-4-2-(2-aminopyridin-3-yl)-5- and filtered through a Celite pad. The combined filtrate and chloro-3H-imidazo[4,5-b]pyridin-3-yl)phenyl)cyclobutyl) washings were concentrated. The residue was purified by 60 carbamate (50.0 mg, 0.101 mmol), 4-3-(4.4.5.5-tetramethyl silica gel column chromatography (EtOAc/hexane=90: 1,3,2-dioxaborolan-2-yl)phenylthiomorpholine 1,1-dioxide 10->85:15->80:20->17:25) to afford desired product (247 (51.5 mg 0.153 mmol), Pd(dppf)Cl. DCM (8.32 mg, 0.0102 mmol), and2MNaOHaq. (0.150 mL, 0.310) in DME (2 mL) mg, 75.7%) as white solid. was heated at 80°C. for 10 hours under nitrogen. After cool 400MHz H-NMR (CDC1) 8: 7.39 (d. J=7.2 Hz, 1H), 7.36 65 ing to room temperature, the mixture was diluted with DCM (d. J=2.3 Hz, 1H), 7.31 (t, J=7.7 Hz, 1H), 7.06-6.99 (m. 1H), and filtered through a Celite pad. The combined filtrate and 3.93-3.79 (m, 4H), 3.18-3.05 (m, 4H), 1.34 (s, 12H). washings were concentrated. The residue was purified by

US 8,962,619 B2 165 166 3.81-3.52 (m, 6H), 2.97-2.84 (m, 2H), 2.78-2.64 (m, 2H), -continued 2.40-2.24 (m. 1H), 2.14-1.97 (m, 1H); LCMS M+H: 548. Example 42 Synthesis of 3-3-4-(1-Aminocyclobutyl)phenyl-5- 3-(8-oxa-3-azabicyclo[3.2.1]oct-3-yl)phenyl-3H imidazo[4,5-b]pyridin-2-yl)pyridin-2-amine trihy drochloride 10

O Br 15 MW = 315.2149 EM = 315.2006 MF = C18H26BNO3

N Br O Step 2: 3-3-(4,4,5,5-Tetramethyl-1,3,2-dioxaboro lan-2-yl)phenyl-8-oxa-3-azabicyclo[3.2.1]octane -- -e- O N A mixture of 3-(3-bromophenyl)-8-oxa-3-azabicyclo H 3.2.1]octane (361 mg, 1.35 mmol), bis(pinacolato) diboron Br (376 mg, 1.48 mmol), Pd(dppf)Cl. DCM (220 mg, 0.269 25 MW = 235.9040 MW = 113.1576 MW = 268.1497 mmol) and potassium acetate (409 mg, 4.04 mmol) in diox EM = 233.868O EM = 113.0841 EM = 267.0259 ane (10 mL) was heated at 80°C. for 14 hours under nitrogen. MF = C6H4Br2 MF = C6H11NO MF = C12H14BrNO After cooling to room temperature, the mixture was diluted with EtOAc and filtered through a Celite pad. The combined filtrate and washings were concentrated. The residue was purified by silica gel column chromatography (EtOAC/hex 30 ane=95:5->90:10->80:20) to afford desired product (375 mg, 88.3%) as orange solid. 400 MHZ H-NMR (CDC1) 8: 7.30-7.23 (m, 3H), 6.94 Step 1: 6.87 (im, 1H), 4.53-4.42 (m, 2H), 3.39 (d. J=11.3 Hz, 2H), 3-(3-Bromophenyl)-8-oxa-3-azabicyclo[3.2.1]octane 3.01 (dd, J=11.3 Hz and 2.3 Hz, 2H), 2.00- 1.89 (m, 4H), 1.33 35 (s, 12H); LCMS M+H: 316. A mixture of 1,3-dibromobenzene (242 LL, 2.00 mmol), 8-Oxa-3-azabicyclo[3.2.1]octane (226 mg, 2.00 mmol), Pd HN (dba), (45.8 mg, 0.0500 mmol), rac-BINAP (96.3 mg, 0.150 N N mmol) and NaOtBu (231 mg, 2.40 mmol) in toluene was 40 heated at 80°C. for 13 hours under nitrogen. After cooling to Cy-K) C N / room temperature, the mixture was diluted with DCM and elN/ \ filtered through a Celite pad. The combined filtrate and wash ings were concentrated. The residue was purified by silica gel 45 column chromatography (EtOAc/hexane=9:1) to afford desired product (361 mg, 67.3%) as colorless oil. - 400 MHZ H-NMR (CDC1) 8: 7.09 (t, J=8.2 Hz, 1H), 6.95-6.90 (m, 2H), 6.74-6.68 (m, 1H), 4.53-4.43 (m, 2H), 50 MW'R) = 490.9846 3.31-3.26 (m, 2H), 3.01 (dd, J=11.7 Hz and 2.5 Hz, 2H), 2.03-1.86 (m, 4H); LCMS M+H: 268. EM = 490.1884

Br 55

O O N M N -- p- R -e- O O 60

O

MW = 268.1497 MW = 253.9386 EM = 267.0259 EM = 254.1861 65 MW = 315.2149 EM = 315.2006 MF = C18H26BNO3

US 8,962,619 B2 169 170 -continued O N O

- CroBr H MW = 248.3208 MW = 2359040 MW = 158.1983 EM = 248.1525 EM=233.8680 EM = 158.1055 MF = C14H2ON2O2

Step 1: 4-Benzyl-N,N-dimethylmorpholine-2-carboxamide 15 Oxalyl chloride (372 uL, 4.27 mmol) was added to a mix ture of 4-benzylmorpholine-2-carboxilic acid hydrochloride (1.00 g, 3.88 mmol) and N,N-dimethylformamide (15.0 LL. 0.194 mmol) in DCE (20 mL). The mixture was heated at 65° C. for 3 hours. After cooling to 0°C., MeNH (2M solution in THF, 9.70 mL, 19.4 mmol) was added. The mixture was O N stirred at room temperature for 4 hours. The mixture was concentrated and the residue was diluted with water and 25 MW = 313.1903 extracted with EtOAc (3x). The combined organic layers EM = 312.0473 were dried over anhydrous NaSO and concentrated to MF = C13H17BrN2O2 afford desired product (803 mg, 83.4%) as brown oil. 400 MHZ H-NMR (CDC1) 8: 7.40-7.21 (m, 5H), 4.28 (dd, J=10.3 Hz and 2.5 Hz, 1H), 3.97-3.90 (m. 1H), 3.71 (td, 30 J=11.3 Hz and 2.4 Hz, 1H), 3.58 (d. J=13.3 Hz, 1H), 3.54 (d. Step 3: 4-(3-Bromophenyl)-N,N-dimethylmorpho J=13.3 Hz, 1H), 3.05 (s.3H), 2.95-2.86 (m, 1H), 2.92 (s.3H), line-2-carboxamide 2.72-2.65 (m, 1H), 2.44-2.35 (m, 1H), 2.27 (td, J=11.4 Hz and 3.2 Hz, 1H); LCMS M+H: 249. A mixture of 1,3-dibromobenzene (387 uL, 3.20 mmol), 35 N,N-dimethylmorpholine-2-carboxamide (506 mg, 3.20 mmol), Pd(dba) (73.3 mg, 0.0800 mmol), rac-BINAP (154 mg, 0.240 mmol) and NaOtBu (369 mg, 3.84 mmol) in tolu O N O NH ene (8 mL) was heated at 80°C. for 5 hours under nitrogen. After cooling to room temperature, the mixture was diluted -e- 40 with DCM and filtered through a Celite pad. The combined O O filtrate and washings were concentrated. The residue was purified by silica gel column chromatography (EtOAC/hex N- - ane=95:5->90:10->80:20->67:33->50:50) to afford desired MW = 248.3208 MW = 158.1983 product (424 mg. 42.3%) as orange oil. EM = 248.1525 EM = 158.1055 45 400 MHZ H-NMR (CDC1) 8: 7.12 (t, J=8.0 Hz, 1H), MF = C7H14N2O2 7.08-7.06 (m. 1H), 7.02-6.98 (m, 1H), 6.88-6.84 (m, 1H), 4.33 (dd, J=10.1 Hz and 2.7 Hz, 1H), 4.07 (ddd, J=11.4 Hz, 3.2 Hz and 1.8 Hz, 1H), 3.80 (td, J=11.3 Hz and 2.9 HZ, 1H), Step 2: N,N-Dimethylmorpholine-2-carboxamide 3.63-3.56 (m. 1H), 3.45-3.38 (m, 1H), 3.13 (s.3H), 3.12 (dd. 50 J=12.4 Hz, and 10.1 Hz, 1H), 2.99 (s.3H), 2.95 (td, J=11.9Hz A mixture of 4-benzyl-N,N-dimethylmorpholine-2-car and 3.5 Hz, 1H); LCMS M+H: 313. boxamide (803 mg, 3.23 mmol), ammonium formate (2.15g. 32.3 mmol) and 10% Pd/C (400 mg) in EtOH (20 mL) was Br heated at 50° C. for 1 hour. After cooling to room temperature, the catalyst was filtered off, washed with EtOH. The com 55 bined filtrate and washings were concentrated, and the resi due was dissolved in EtOH (20 mL). Ammonium formate (2.15g, 32.3 mmol) and 10% Pd/C (400 mg) were added to N R / the mixture. The mixture was heated at 50° C. for 1 hour. After -- B-B -- cooling to room temperature, the catalyst was filtered off 60 washed with EtOH. The combined filtrate and washings were C O N N o? Y concentrated to afford desired product (506 mg, 98.9%) as colorless oil. O 400 MHz H-NMR (CDC1,) 8:4.44-4.37 (m. 1H), 3.93 (td, MW = 313.1903 MW = 253.9386 J=7.6 Hz and 3.8 Hz, 1H), 3.85-3.77 (m, 1H), 3.49 (brs, 1H), 65 EM = 312.0473 EM = 254.1861 3.19-3.14 (m, 2H), 3.09 (s.3H), 3.04-2.99 (m, 2H), 2.96 (s, MF = C13H17BrN2O2 MF = C12H24B2O4 3H); LCMS M+H: 159.

US 8,962,619 B2 177 178 Step 1: 1-(3-Bromophenyl)pyrrolidine-3-carboxylic -continued acid O O N M 1M NaOH ad. (2 mL) was added to a solution of methyl B-B -3- 1-(3-bromophenyl)pyrrolidine-3-carboxylate in THF-MeOH M N (1 mL-1 mL). After stirring the mixture at room temperature O O for 13 hours, the mixture was concentrated. The residue was MW = 253.9386 diluted with 1M NaOH aq. and washed with ether. The aque EM = 254.1861 ous layer was acidified with 1M HCl aq. and extracted with MF = C12H24B2O4 DCM (3x). The combined organic layer was dried over anhy drous NaSO and concentrated to afford desired product 10 (293 mg, 83.9%) as yellow solid. 500 MHz H-NMR (CDC1) 8: 705 (t, J=7.9 Hz, 1H), 6.82-6.77 (m, 1H), 6.67 (t, J=2.0 Hz, 1H), 6.45 (dd, J=7.9, 2.0 HZ, 1H), 3.58-3.47 (m, 2H), 3.43-3.35 (m. 1H), 3.34-3.18 (m, 2H), 2.37-2.22 (m, 2H); LCMS: 272 M+H. 15

Br Br

O

B NO N He- N

OH N 25 O O N N O O N MW = 270.1225 MW = 297.1909 N EM = 269.0051 EM = 296.0524 MF = C11H12BrNO2 MF = C13H17BrN2O 30 O Step 2: 1-(3-Bromophenyl)-N,N-dimethylpyrroli MW = 344.2562 dine-3-carboxamide EM = 344.2271 Oxalyl chloride (104 uL. 1.19 mmol) was added to a mix ture of 1-(3-bromophenyl)pyrrolidine-3-carboxylic acid (293 35 mg, 1.08 mmol) and N,N-dimethylformamide (8.4LL, 0.108 mmol) in DCE (5 mL). After heating the mixture at 65° C. for 2 hours, the mixture was coevaporated with toluene. The Step 3: N,N-Dimethyl-1-3-(4,4,5,5-tetramethyl-1,3, residue was diluted with THF (5 mL) and Me NH (2M/THF, 2-dioxaborolan-2-yl)phenylpyrrolidine-3-carboxam 2.71 mL, 5.42 mmol) was added to the mixture at 0°C. After 40 ide stirring the mixture for 16 hours at room temperature, the mixture was concentrated and the residue was diluted with 400 MHZ H-NMR (CDC1) 8: 7.24 (t, J=7.8 Hz, 1H), water and extracted with EtOAc (3x). The combined organic 7.17-7.11 (m. 1H), 7.04-6.98 (m, 1H), 6.72-6.63 (m, 1H), layers were dried over anhydrous NaSO and concentrated. 3.60 (t, J=8.7 Hz, 1H), 3.53-3.45 (m, 2H), 3.44-3.32 (m, 2H), The residue was purified by silica gel column chromatogra 45 phy (EtOAc/hexane=9:1->4:1->2:1) to afford desired prod 3.12 (s.3H), 2.99 (s.3H), 2.39-2.28 (m. 1H), 2.24-2.14 (m, uct (168 mg, 48.0%) as orange oil. 1H), 1.33 (s, 12H); LCMS: 345 M+H. 400 MHz H-NMR (CDC1,) 8: 'H-NMR (CDC1) 8: 7.05 (t, J=7.9 Hz, 1H), 6.80-6.75 (m. 1H), 6.68 (t, J=2.3 Hz, 1H), 6.46 (dd, J=7.9, 2.3 Hz, 1H), 3.56-3.28 (m, 5H), 3.12 (s.3H), 50 2.99 (s.3H), 2.38-2.16 (m, 2H); LCMS: 297 M+H. HN N N N Br

55 C N elNY \ /

-- O

60 X-d N H

O MW = 297.1909 MW = 490.9846 EM = 296.0524 65 EM = 490.1884 MF = C13H17BrN2O

US 8,962,619 B2 181 182 1H), 3.60-3.28 (m, 5H), 3.10 (s.3H), 2.86 (s.3H), 2.72-2.54 'H-NMR (400 MHz, DMSO-d) 8: 9.01-8.84 (m, 3H), (m, 4H), 2.29-2.16 (m. 2H), 2.14-2.02 (m. 1H), 1.92-1.78 (m, 8.54-8.32 (m, 2H), 8.16 (dd. 1H, J=6.2, 1.6 Hz), 8.09 (d. 1H, 1H); LCMS: 573 M+H). J=8.7 Hz), 7.90 (dd. 1H, J=7.6, 1.6 Hz), 7.77 (d. 2H, J=8.7 Hz), 7.72-7.64 (m, 3H), 7.54 (d. 1H, J=8.0 Hz), 7.35 (t, 1H, Example 46 J=8.0 Hz), 7.07 (dd. 1H, J=8.0, 2.3 Hz), 6.90 (dd. 1H, J=7.6, Synthesis of 3-(3-4-(1-aminocyclobutyl)phenyl-5- 6.2 Hz), 3.36-3.26 (m, 8H), 2.95 (s.3H), 2.70-2.57 (m, 4H), {3-4-(methylsulfonyl)piperazin-1-yl)phenyl)-3H 2.30-2.16 (m. 1H), 1.92-1.79 (m, 1H). LCMS: 595 M+H. imidazo 4,5-b]pyridin-2-yl)pyridin-2-amine hydro chloride Example 47 10 Step 1: Synthesis of 1-(methylsulfonyl)-4-3-(4.4.5, Inactive Akt Alpha Screen Assay 5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenylpip erazine AKT1 activity was assayed using the GSK3-derived bioti nylated peptide substrate, crosstide (biotin-GRPRTSS FAEG), and AlphaScreenTM (Amplified Luminescent Prox 15 imity Homogeneous Assay) technology. AKT1 activation was achieved by the addition of the activating kinases PDK1 and MAPKAPK2, lipid vesicles, and ATP. The extent of Sl peptide phosphorylation was determined using a phospho O O AKT Substrate antibody and acceptor beads conjugated to NY Protein A and donor beads conjugated to streptavidin that bind to the biotin on the peptide. Excitation of the donor beads converted ambient oxygen to excited singlet oxygen which, when in close proximity to acceptor beads, reacted with acceptor beads resulting in signal amplification. 25 Test inhibitors and controls (S)-1-(5-(3-methyl-1H-inda Zol-5-yl)pyridin-3-yl)oxy)-3-phenylpropan-2-amine, 1-(1- n O (4-(7-phenyl-1H-imidazo[4,5-gquinoxalin-6-yl)benzyl)pi N-s/ peridin-4-yl)-1H-benzodimidazol-2(3H)-one, and 8-(4-(1- / N aminocyclobutyl)phenyl)-9-phenyl-1,2,4-triazolo 3,4-f1. 30 6naphthyridin-3(2H)-one) were prepared in 10% DMSO at 10-fold the desired final concentration, and added to each well of a reaction plate (Corning 96-well half-area solid white 1-(Methylsulfonyl)-4-3-(4,4,5,5-tetramethyl-1,3,2-diox nonbinding Surface plate) in a Volume of 2.5 ul. Full-length aborolan-2-yl)phenylpiperazine was synthesized by the pro inactive AKT1 was diluted in assay buffer (50 mM Tris, pH cedure H (Step 1 and 2). 35 8.0, 0.02 mg/mL BSA, 10 mM MgCl, 1 mM EGTA, 10% 'H-NMR (400 MHz, CDC1) 8: 7.40-7.36 (2H, m), 7.30 glycerol, 0.2 mM NaVO, 1 mM DTT, 0.1 mM f-glycero (1H, t, J=7.8 Hz), 7.04 (1H, ddd, J=7.8, 2.8, 1.4 Hz), 3.41 phosphate, and 0.2 mM NaF) and added to each well in a 3.35 (4H, m), 3.34-3.28 (4H, m), 2.83 (3H, s), 1.34 (12H, s). volume of 17.5 uL for a final concentration in the 25 L LCMS: 367 M+H. reaction of 8 nM (Akt1). After a 20 minute pre-incubation at room temperature, the kinase reaction was initiated by the Step 2: Synthesis of 3-(3-4-(1-aminocyclobutyl) 40 addition of 5 uM of an activation mixture diluted in assay phenyl-5-3-4-(methylsulfonyl)piperazin-1-yl) buffer containing biotinylated crosstide, PDK1, MAP phenyl-3H-imidazo[4,5-b]pyridin-2-yl)pyridin-2- KAPK2, DOPS/DOPC, PtdIns(3,4,5)P3, and ATP for final amine hydrochloride concentrations of 60 nM biotinylated crosstide, 0.1 nM PDK1, 0.7 nM MK2, 5.5 uM DOPS, 5.5 M DOPC, 0.5uM 3-(3-4-(1-Aminocyclobutyl)phenyl-5-3-4-(methylsul 45 PtdIns(3,4,5)P3, and 50 uM ATP. The plates were incubated fonyl)piperazin-1-yl)phenyl-3H-imidazo[4,5-b]pyridin-2- for 30 minutes at room temperature, and then stopped in the yl)pyridin-2-amine hydrochloride was synthesized by the dark by the addition of 10 uL stop/detection mixture prepared procedures of G and B. in assay buffer containing EDTA, AlphaScreenTM Streptavi din Donor and Protein A Acceptor beads, and phospho-AKT 50 substrate antibody for final concentrations of 10 mM EDTA, HN 500 ng/well of both AlphaScreenTM Streptavidin Donor beads and Protein A Acceptor beads, and phospho-AKT Substrate Sl-N N antibody at a final dilution of 1:350. Assay plates were incu N bated for 90 minutes at room temperature in the dark, and the luN \ / 55 plates were read on a PerkinElmer Envision Multilabel plate reader (excitation wavelength:640 nm, emission wavelength: 570 nm). Reaction: 2.5uL 10x Akt inhibitor in 10% DMSO N 17.5 uL inactive Akt or buffer for blank 60 20 minute pre-incubation at room temperature 5 uL Reaction Mix (5xATP 5x substrate, 5xPDK1, C D NHHCI 5xMK2, and 5x lipid vesicle mixture) 30 minute incubation at room temperature OESEO 10 uL Detection Buffer 65 90 minute incubation at room temperature Detection (excitation: 640 nm, emission: 570 nm) Envision Instrument Settings: US 8,962,619 B2 183 184 Instrument: PerkinElmer Envision treatment with SuperSignal(R) ELISA for 15 minutes, the Plate: 96 well detection was performed using a luminometer. Data were Program Name: normalized to untreated controls and analyzed with Microsoft Excitation: Ex Top Excel. Mirror: General Dual Slot 2 Excitation Filter: CFP430 EX. Slot 2 Example 51 Emission Filter Emission 579 Em slot 2 2nd Emission Filter: None Measurement Height (mm): 3.8 ATP Ouantitation Assay Excitation light (%): 1 Detector gain: 1 10 Cell viability analysis. Cell viability was determined by the 2nd detector gain: 0 ATP quantitation assay. Briefly, cells were plated in a 96-well H. Flashes: 10 plate at 2,000-10,000 cells per well, cultured for 24 hours in H. Flashed/AD: 1 complete growth medium, and then treated with various Reference Signal: 383722 drugs or combination of drugs for 72 hours. CellTiter-Glo(R) AD gain: 4 15 reagent was added and incubated at room temperature for 10 Reference Excitation (%): 100 minutes, followed by assessment of cell viability using a luminometer. Data were normalized to untreated controls and Example 48 analyzed with Microsoft Excel. Table 2 shows the physical and biological properties of MTS Assay representative compounds of the present invention.

Cell proliferation analysis. Cell survival was determined TABLE 2 by the MTS assay. Briefly, cells were plated in a 96-well plate at 2,000-10,000 cells per well, cultured for 24 hours in com Thermal Cell Shift Elisa Akt1 biochemical assay ICso plete growth medium, and then treated with various drugs and 25 drug combinations for 72 hours. MTS was added and incu Cmpd Kd (nM) (nM) GIso (IM) AlphaScreen crosstide (IM) bated for 4 hour, followed by assessment of cell viability 1 N.A. 235 N.T. using the microplate reader at 570 nm. Data were normalized 3 7 29 444 O.OO309 4 0.4 17 26S O.OO17 to untreated controls and analyzed with Microsoft Excel. 5 0.7 O.OO446 30 6 1.4 O.OO163 Example 49 7 7.1 12 836 O.OO245 8 1.2 8 1348 O.OO15 Thermal Shift Assay 9 O.2 2.1 O.OO2S1 10 O.2 3.0 O.OO415 11 O.2 1.7 O.OO336 Binding analysis. Protein-ligand binding was identified 35 12 O.2 O.OO144 with the thermal shift assay which is based on the ligand 13 1.4 9 335 O.OO215 induced stabilization of the protein tertiary structure. The 14 1.1 11 296 O.OO184 thermal stability of the ligand-Akt1 complex was assessed by 15 1.1 4.0 Subjecting the complex to a set temperature gradient and by 16 1.7 O.OO342 comparison of the meltion temperature of the Akt1-ligand 17 3.1 O.OO436 40 18 O.8 O.OO179 complex with the melting temperature of the protein alone. 19 N.A. 60 1423 O.OO377 Protein unfolding was monitored by the fluorescence readout 2O 57.3 414 N.T. O.OO872 of an environmentally sensitive fluorescent dye, 1-anilinon 21 109.5 60S aphthalene-8-sulfonic acid (ANS). Protein-ligand mixture 22 83.1 containing 15 M compound, 200 ng/mL full length inactive Akt1, 200 uM ANS in the binding buffer (25 mM Tris-HCl 45 (pH. 7.5), 100 mMNaCl, 10% Glycerol, and 5 mMDTT) was prepared in a PCR plate. The PCR plate was placed into a The invention claimed is: RT-PCR instrument and a temperature gradient was per formed, increasing from 27 to 80°C., at 1° C./30 sec heat rate with a dwell time of 20 sec. The change in ANS fluorescence 50 1. A compound of formula I: was measured using the filter set of 475 nm/525 nm excita tion/emission. Ligand affinity parameter (Kd) at 25°C. was calculated using constant ligand binding enthalpy 4H--15 (I) kcal/mol. HN N N Example 50 55 Cell ELISA Assay X CO)-().4. N Phospho-Akt detection analysis. Phospho-Akt (S473) was detected using cell ELISA. Briefly, cells were plated in a 60 96-well plate at 7,000-10,000 cells per well, cultured for 2-3 days in complete growth medium, and then treated with vari ous drugs or combinations of drugs for 1 hour. The cells were HN fixed with ice-cold methanol, and washed with TBS supple mented with 1% H.O. Phospho-Akt (S473) was probed with 65 corresponding rabbit antibody as a primary antibody, and horseradish peroxidase conjugated goatanti-rabbit IgG. After or a pharmaceutically acceptable salt or ester thereof, US 8,962,619 B2 185 186 wherein: -continued X is NRR' or HN 21 N N | Y s slS^N \ /

C(O)NHR, 10 N-> O wherein ÖH. NH2

15

C(O)NHR,

is linked to the imidazopyridinyl ring of formula I at the position indicated by “*”: 25 Ry is Co-Co aryl Substituted with unsubstituted or Substi tuted heterocycle comprising one 5- or 6-member ring and 1-3 heteroatoms selected from N, O and S; ~ HN R is H; or 30 Ry. and R', together with the nitrogen atom to which they are attached, form an unsubstituted or substituted mor pholine; and R, is substituted C-C alkylor unsubstituted C-C alkyl. 2. The compound of claim 1, selected from the group 35 consisting of:

NH2 40 N N HN N N 21 N s r sCO)-(). N su 45 su? \ A

NH2 50 HN O HN -N NH2 N N HN 21 N s 55 N EN su? \ / 21 N s HN SaN / \ A

1sH O OH NH2

65 US 8,962,619 B2 187 188 -continued heterocycle selected from pyrrolidinyl, imidazolidinyl, pyra N Zolidinyl, oxazolidinyl, isoxazolidinyl, triazolidinyl, tetrahy N N rofuranyl, piperidinyl, piperazinyl, and morpholinyl. 21 N s 4. The compound of claim 3, wherein the heterocycle is suSM \ / morpholinyl. r HN N 5. The compound of claim 1, wherein X is NRR'; and R. and R, together with the nitrogen atom to which they are lu attached, form an unsubstituted morpholine or a morpholine substituted with a substituent selected from the group con 10 sisting of hydroxyl, halogen, cyano, nitro, unsubstituted or Substituted amino, unsubstituted or substituted C-C alkyl, and unsubstituted or Substituted C-C alkoxy. 6. A pharmaceutical composition comprising a therapeuti 15 cally effective amount of a compound of claim 1, or a salt or or a pharmaceutically acceptable salt or ester thereof. ester thereof, and a pharmaceutically acceptable carrier or 3. The compound of claim 1, wherein X is NRR'; and Ry excipient. is C-C aryl substituted with unsubstituted or substituted