[Frontiers in Bioscience E3, 801-807, June 1, 2011]

Src-family tyrosine kinases as therapeutic targets in advanced cancer

Irwin H. Gelman

Department of Cancer Genetics, Roswell Park Cancer Institute, Elm and Carlton Streets, Buffalo, NY

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Critical role for SFK in cancer malignancy: preclinical models
4. Therapeutic SFK inhibitors
5. Acknowledgements
6. References

1. ABSTRACT

Src-family tyrosine kinases (SFK) play critical roles in mediating many cellular pathways such as proliferation, adhesion, survival, differentiation and cell motility. There is clear evidence that SFK activity is increased in many human cancers, either through gene amplification, transcriptional upregulation, posttranslational modification by activated upstream growth factor receptors, and even in rare cases, by mutations known to increase intrinsic tyrosine kinase activity in oncoviral forms of SFK. Many recent studies using animal models of human cancer seem to indicate that SFK may only be appropriate therapeutic targets in a subset of primary tumors because of the existence of multiple independent pathways that mediate oncogenic signaling. In contrast, SFK seem to be required for specific parameters of malignant progression, such as recurrence and/or metastasis- especially involving growth in the bone microenvironment. The resulting development of SFK antagonists, and their progression through clinical trials, has brought renewed focus on this tyrosine kinase family as critical mediators of the so-called lethal phenotype of cancer.

2. INTRODUCTION

Src is the prototypic member of a family of non-receptor tyrosine kinases consisting of two subgroups: the ubiquitously expressed Src, Yes and Fyn, versus the kinases expressed in specialized cell types such as hematopoietic (Blk, Fgr, Hck, Lck, Lyn and Yrk) or epithelial (Frk) cells (1,2). The original identification of Src as the oncogene of Rous sarcoma virus indicated that even a single point mutation or the truncation of a C-terminal Tyr-527 residue (Tyr-530 in human Src) in the viral Src allele was sufficient to induce constitutive tyrosine kinase activity when introduced into the so-called cellular-Src proto-oncogene allele (3). Analysis of these mutations subsequently identified several so-called Src-homology (SH) domains: the kinase domain (SH1), a phosphotyrosine-binding domain (SH2) and a PxxPxP-binding domain (SH3) (4). In addition to all the SFK encoding these three functional motifs, many other signaling proteins including so-called adaptor proteins encode one or more SH2 and/or SH3 modules that facilitate protein-protein interactions during signal transduction (5). Structural studies of SFK suggest that internal interactions between the SH2/3 domains with their cognate ligands normally hold the proteins in a "closed", kinase-inactive conformation, whereas the kinase-active conformation can be achieved by mutation at these sites, by interaction of these sites with other signaling proteins, or by specific posttranslational modifications such as autophosphorylation at Y416 (Y419 in human Src)(6,1).

Many studies in the last 20 years have demonstrated upregulated SFK in cancer cell lines and in primary tumors, both at the protein and tyrosine kinase activity level (reviewed in (7). Although "viral-Src-like" activating mutations have been identified in small subsets of colon and breast cancers (8,9), the vast majority of cancers involve upregulation of WT-Src. The current belief in the field, therefore, is that genetic mutations and epigenetic changes that accumulate during cancer progression result in overexpressed, activated growth factor or adhesion receptors that directly or indirectly activate SFK and their downstream oncogenic pathways. Indeed, most cancers exhibit increasing activity levels of one or more SFK with progression to more aggressive phenotypes (7). Taken together, these data suggest that SFK inhibitors would have their greatest clinical impact in preventing or treating advanced stages of cancer such as recurrence and metastasis. In the paragraphs below, I will review the most recent advances in our understanding of how SFK drive cancer malignancy, the flurry of SFK antagonist development and testing, and the future prospects for targeting SFK in advanced cancers.

3. CRITICAL ROLE FOR SFK IN CANCER MALIGNANCY: PRECLINICAL MODELS

There is a large corpus of data showing that SFK members are required for the proliferation, differentiation, cell motility or survival of many untransformed and cancer cell lines in culture (reviewed in (10,11). With the advent of genetic knockdown technologies, many groups have shown that the loss of specific SFKs has little effect on primary tumor growth in animal models but can prevent parameters of malignant, secondary growth. Below is an incomplete set of examples of such studies. Trevino et al. showed that shRNA-mediated Src knockdown had no effect on the incidence of primary orthotopic human pancreatic L3.6pl tumors in nude mice, yet this strongly suppressed the generation of liver metastases (12). Although Src knockdown in human FG pancreatic cancer cells partially reduced primary orthotopic tumor growth in nude mice, it severely suppressed the increased spontaneous metastatic potential induced by ectopic alpha(v)beta(3) integrin expression (13). Park et al. showed similar effects of Src knockdown on the generation of lymph node metastases by orthotopic PC3-LN4 tumors (14). Rucci et al. (15) and Myoui et al. (16) used kinase-dead versions of Src as a dominant-interfering allele to show that the loss of Src kinase activity suppressed experimental bone metastases formed after intracardiac injection of the human MDA-MB-231 breast cancer cell line. The knockdown of Lyn resulted in a small but significant inhibition of primary Ewing's sarcoma growth but this had even greater effects on the generation of metastatic lesions (17). Lastly, Guo et al. demonstrated that the loss of Src-mediated tyrosine phosphorylation of the androgen receptor (resulting from Src shRNA expression) in CWR22R1 human prostate cancer cells prevented tumor recurrence in castrated male mice (18).

One mechanism by which Src seems to inhibit metastatic growth is by suppressing neovascularization at distal sites of tumor cell dissemination. c-Src activation is required to induce vascular endothelial growth factor expression and secretion (19), most probably due to its ability to downregulate HIF-1" expression (20), such that Src knockdown suppresses the metastatic potential of human colon cancer cells in mouse models (21). Our lab showed that the SSeCKS/Gravin/AKAP12 gene, an antagonist of Src-mediated oncogenic transformation (22) and podosome/invadosome formation (23), can selectively suppress neovascularization at lung metastatic sites by downregulating VEGF expression (24) through the disengagement of growth factor activation of Src from MAP kinase pathways (25). Several studies from the Cheresh lab argue for the requirement for Src signaling to facilitate activation and recruitment of vascular endothelial cells and other cellular effectors of the tumor microenvironment to peripheral sites of growing metastases (26,27).

Increased Src activation has been reported in cancer cell lines exhibiting resistance to chemotherapeutics (28-34), and thus, groups have focused on preventing resistance by combining SFK inhibitors with standard cytotoxic chemotherapies (30,34-36) or on treating resistant cells with SFK inhibitors (37).

Many cancers, such as breast and prostate cancer and multiple myeloma, metastasize to the bone. The crosstalk of secreted factors between tumor and specific bone cells results in a so-called "vicious cycle" that increases both bone destruction and tumor growth. Metastatic cells interfere with normal bone maintenance and remodeling programs involving a crosstalk between osteoblasts and osteoclasts via the RANKL-osteoprotegerin axis (38,39). Src activity is required for osteoclast activation (40-43) and its inhibition suppresses the formation and growth of bone metastases (16, 44-49). Moreover, in patients with androgen-independent prostate cancer, higher SFK activity levels correlated with increased levels of bone metastases (31). Higher SFK activity levels in primary breast cancer lesions also correlated with increased chances of disease relapse as bone metastases (50). Lastly, there is mounting evidence that insulin-like growth factor (IGF) and IGF-binding proteins facilitate bone development of prostate metastases (51), and thus, Src is thought to be an appropriate target in this context because of its requirement for IGF-1 receptor upregulation by androgens (52).

4. THERAPEUTIC SFK INHIBITORS

A host of small molecule SFK inhibitors are being tested in pre-clinical and clinical trials as monotherapies and in drug combination studies. These include: Dasatinib (BMS-354825; Spryceltrademark), Bosutinib (SKI-606), Saracatinib (AZD0530), PD180970 (53), SU6656 (54,55), KXO1 (KX2-391) (56,57), CGP76030 (58), AP23451 (59), AZM475271 (60-62). Others, such as INNO-406 (NS-187), XL-999 and XL-228, were developed as inhibitors of the Abl non-receptor tyrosine kinase, although they have shown some ability to inhibit SFK (63,64). Dasatinib is the only of these that is approved by the U.S. Food and Drug Administration, and this was for use in chronic myelogenous leukemia or Philadelphia-chromosome-positive acute lymphocytic leukemia. Several recent reviews describe the testing phase and tumor settings for these compounds (65-67). A review of the NCI Clinical Trials Website (http://www.clinicaltrials.gov/ct2/results?term=Src+inhibitors&pg=1), accessed in May 2010, indicates that Dasatinib, Bosutinib and Saracatinib are being tested at the Phase II and III levels in small and non-small cell lung carcinoma, breast, prostate, colorectal, liver, ovarian pancreatic cancer, sarcomas, head and neck squamous cell carcinoma and melanomas, with the remaining SFK inhibitors being tested in Phase II trials on limited tumor settings.

Future directions- A major consideration in the development and testing of SFK-based therapeutics is the ability to identify valid biomarkers of drug efficacy in patients, and additionally, to develop appropriate patient-specific genomic signatures that predict or explain clinical response. One confounding factor in this regard has been the realization that compounds developed initially as Src- or SFK-specific inhibitors subsequently are shown to have a wider range of targets. For example, Dasatinib, originally envisioned as a Src/Abl inhibitor (68), actually targets at least 40 receptor and non-receptor tyrosine kinases including SFK and EGFR (69-71). Thus, given that many human cancer types exhibit activation of multiple, redundant oncogenic pathways (33), it will be difficult in pre-clinical models and in clinical trials to attribute whether anti-tumor efficacy is due to the loss of SFK activation or whether this is a bystander effect. Nonetheless, some groups have already identified potential SFK-related biomarkers for Dasatinib action such as the loss of a shared SFK-poY419 epitope or an epitope (poY845) for the autophosphorylated EGFR (71-73). This raises the issue as to whether a more promiscuous inhibitor might have clinical advantage because of its ability to target multiple oncogenic pathways. Clearly, this will require further study to resolve. However, the advantage for the more specific SFK inhibitors might lie in their lack of toxicity, based on their narrow range of targets. As an example, the widely reported cardiotoxicity induced by Dasatinib is believed to be attributed to its strong inhibition of c-Abl (74), and moreover, Dasatinib has strong immunosuppressive activity (75-77), possibly correlating with its ability to inhibit ZAP-70 (Gelman, I.H., unpublished data). In contrast, KXO1 is a poor inhibitor of Abl and has little effect on ZAP-70 (Gelman, I.H., unpublished data), correlating with no reported cardiotoxicity and little lymphocytopenia in Phase I trials (56). Nevertheless, the clinical side-effects of most SFK inhibitors are manageable, and some are even well tolerated based on Phase I data (56). With the renewed appreciation for SFK as important therapeutic targets in cancer malignancy, and with the growing development of drugs in this sector, there is confidence that SFK inhibitors will offer benefit for many cancer patients, especially those with recurrent and/or metastatic disease. In this regard, there will have to be increased future commitment to clinical studies that include parameters of advanced cancer as therapeutic endpoints rather than just shrinkage of primary tumors.

5. ACKNOWLEDGEMENTS

This work is supported by Department of Defense (W81XWH-08-1-0026, W81XWH-09-1-0465) and National Institutes of Health (CA094108, CA116430) grants to I.H.G.

6. REFERENCES

1. G. S. Martin: The hunting of the Src. Nat Rev Mol Cell Biol 2, 467-475 (2001)
doi:10.1038/35073094

2. M. C. Frame: Newest findings on the oldest oncogene; how activated src does it. J Cell Sci 117, 989-998 (2004)
doi:10.1242/jcs.01111

3. R. Jove, H. Hanafusa: Cell Transformation by the Viral src Oncogene. Ann Rev Cell Biol 3, 31-56 (1987)
doi:10.1146/annurev.cb.03.110187.000335
doi:10.1146/annurev.cellbio.3.1.31

4. T. Pawson, J. D. Scott: Signaling through scaffold, anchoring, and adaptor proteins. Science 278, 2075-2080 (1997)
doi:10.1126/science.278.5346.2075

5. J. R. Engen, T. E. Wales, J. M. Hochrein, M. A. Meyn, III, Ozkan S. Banu, I. Bahar, T. E. Smithgall: Structure and dynamic regulation of Src-family kinases. Cell Mol Life Sci 65, 3058-3073 (2008)
doi:10.1007/s00018-008-8122-2

6. C. B. Breitenlechner, N. A. Kairies, K. Honold, S. Scheiblich, H. Koll, E. Greiter, S. Koch, W. Schafer, R. Huber, R. A. Engh: Crystal structures of active SRC kinase domain complexes. J Mol Biol 353, 222-231 (2005)
doi:10.1016/j.jmb.2005.08.023

7. J. M. Summy, G. E. Gallick: Src family kinases in tumor progression and metastasis. Cancer Metastasis Rev 22, 337-358 (2003)
doi:10.1023/A:1023772912750

8. R. B. Irby, W. G. Mao, D. Coppola, J. Kang, J. M. Loubeau, W. Trudeau, R. Karl, D. J. Fujita, R. Jove, T. J. Yeatman: Activating SRC mutation in a subset of advanced human colon cancers. Nature Genet 21, 187-190 (1999)
doi:10.1038/5971

9. T. J. Yeatman: A renaissance for SRC. Nat Rev Cancer 4, 470-480 (2004)
doi:10.1038/nrc1366

10. P. L. Schwartzberg: The many faces of Src: multiple functions of a prototypical tyrosine kinase. Oncogene 17, 1463-1468 (1998)
doi:10.1038/sj.onc.1202176

11. S. M. Thomas, J. S. Brugge: Cellular functions regulated by Src family kinases. Annu Rev Cell Dev Biol 13, 513-609 (1997)
doi:10.1146/annurev.cellbio.13.1.513

12. J. G. Trevino, J. M. Summy, D. P. Lesslie, N. U. Parikh, D. S. Hong, F. Y. Lee, N. J. Donato, J. L. Abbruzzese, C. H. Baker, G. E. Gallick: Inhibition of SRC expression and activity inhibits tumor progression and metastasis of human pancreatic adenocarcinoma cells in an orthotopic nude mouse model. Am J Pathol 168, 962-972 (2006)
doi:10.2353/ajpath.2006.050570

13. J. S. Desgrosellier, L. A. Barnes, D. J. Shields, M. Huang, S. K. Lau, N. Prevost, D. Tarin, S. J. Shattil, D. A. Cheresh: An integrin alpha(v)beta(3)-c-Src oncogenic unit promotes anchorage-independence and tumor progression. Nat Med 15, 1163-1169 (2009)
doi:10.1038/nm.2009

14. S. I. Park, J. Zhang, K. A. Phillips, J. C. Araujo, A. M. Najjar, A. Y. Volgin, J. G. Gelovani, S. J. Kim, Z. Wang, G. E. Gallick: Targeting SRC family kinases inhibits growth and lymph node metastases of prostate cancer in an orthotopic nude mouse model. Cancer Res 68, 3323-3333 (2008)
doi:10.1158/0008-5472.CAN-07-2997

15. N. Rucci, I. Recchia, A. Angelucci, M. Alamanou, Fattore A. Del, D. Fortunati, M. Susa, D. Fabbro, M. Bologna, A. Teti: Inhibition of protein kinase c-Src reduces the incidence of breast cancer metastases and increases survival in mice: implications for therapy. J Pharmacol Exp Ther 318, 161-172 (2006)
doi:10.1124/jpet.106.102004

16. A. Myoui, R. Nishimura, P. J. Williams, T. Hiraga, D. Tamura, T. Michigami, G. R. Mundy, T. Yoneda: C-SRC tyrosine kinase activity is associated with tumor colonization in bone and lung in an animal model of human breast cancer metastasis. Cancer Res 63, 5028-5033 (2003)

17. H. Guan, Z. Zhou, G. E. Gallick, S. F. Jia, J. Morales, A. K. Sood, S. J. Corey, E. S. Kleinerman: Targeting Lyn inhibits tumor growth and metastasis in Ewing's sarcoma. Mol Cancer Ther 7, 1807-1816 (2008)
doi:10.1158/1535-7163.MCT-08-0058

18. Z. Guo, B. Dai, T. Jiang, K. Xu, Y. Xie, O. Kim, I. Nesheiwat, X. Kong, J. Melamed, V. D. Handratta, V. C. Njar, A. M. Brodie, L. R. Yu, T. D. Veenstra, H. Chen, Y. Qiu: Regulation of androgen receptor activity by tyrosine phosphorylation. Cancer Cell 10, 309-319 (2006)
doi:10.1016/j.ccr.2006.08.021

19. D. Mukhopadhyay, L. Tsiokas, X. M. Zhou, D. Foster, J. S. Brugge, V. P. Sukhatme: Hypoxic induction of human vascular endothelial growth factor expression through c-Src activation. Nature 375, 577-581 (1995)
doi:10.1038/375577a0

20. B. H. Jiang, F. Agani, A. Passaniti, G. L. Semenza: V-SRC induces expression of hypoxia-inducible factor 1 (HIF- 1) and transcription of genes encoding vascular endothelial growth factor and enolase 1: Involvement of HIF-1 in tumor progression. Cancer Res 57, 5328-5335 (1997)

21. L. M. Ellis, C. A. Staley, W. Liu, R. Y. Fleming, N. U. Parikh, C. D. Bucana, G. E. Gallick: Down-regulation of vascular endothelial growth factor in a human colon carcinoma cell line transfected with an antisense expression vector specific for c-src. J Biol Chem 273, 1052-1057 (1998)
doi:10.1074/jbc.273.2.1052

22. X. Lin, I. H. Gelman: Re-expression of the major protein kinase C substrate, SSeCKS, suppresses v-src-induced morphological transformation and tumorigenesis. Cancer Res 57, 2304-2312 (1997)

23. I. H. Gelman, L. Gao: The SSeCKS/Gravin/AKAP12 Metastasis Suppressor Inhibits Podosome Formation Via RhoA- and Cdc42-Dependent Pathways. Mol Cancer Res 4, 151-158 (2006)
doi:10.1158/1541-7786.MCR-05-0252

24. B. Su, Q. Zheng, M. M. Vaughan, Y. Bu, I. H. Gelman: SSeCKS metastasis-suppressing activity in MatLyLu prostate cancer cells correlates with VEGF inhibition. Cancer Res 66, 5599-5607 (2006)
doi:10.1158/0008-5472.CAN-05-4123

25. B. Su, Y. Bu, D. Engelberg, I. H. Gelman: SSeCKS/Gravin/AKAP12 inhibits cancer cell invasiveness and chemotaxis by suppressing a PKC-RAF/MEK/ERK pathway. J Biol Chem 285, 4578-4586 (2010)
doi:10.1074/jbc.M109.073494

26. B. P. Eliceiri, R. Paul, P. L. Schwartzberg, J. D. Hood, J. Leng, D. A. Cheresh: Selective requirement for Src kinases during VEGF-induced angiogenesis and vascular permeability. Mol Cell 4, 915-924 (1999)
doi:10.1016/S1097-2765(00)80221-X

27. S. Weis, J. Cui, L. Barnes, D. Cheresh: Endothelial barrier disruption by VEGF-mediated Src activity potentiates tumor cell extravasation and metastasis. J Cell Biol 167, 223-229 (2004)
doi:10.1083/jcb.200408130

28. M. S. Duxbury, H. Ito, M. J. Zinner, S. W. Ashley, E. E. Whang: Inhibition of SRC tyrosine kinase impairs inherent and acquired gemcitabine resistance in human pancreatic adenocarcinoma cells. Clin Cancer Res 10, 2307-2318 (2004)
doi:10.1158/1078-0432.CCR-1183-3

29. M. S. Duxbury, H. Ito, M. J. Zinner, S. W. Ashley, E. E. Whang: siRNA directed against c-Src enhances pancreatic adenocarcinoma cell gemcitabine chemosensitivity. J Am Coll Surg 198, 953-959 (2004)
doi:10.1016/j.jamcollsurg.2004.01.037

30. J. A. George, T. Chen, C. C. Taylor: SRC tyrosine kinase and multidrug resistance protein-1 inhibitions act independently but cooperatively to restore paclitaxel sensitivity to paclitaxel-resistant ovarian cancer cells. Cancer Res 65, 10381-10388 (2005)
doi:10.1158/0008-5472.CAN-05-1822

31. O. Tatarov, T. J. Mitchell, M. Seywright, H. Y. Leung, V. G. Brunton, J. Edwards: SRC family kinase activity is up-regulated in hormone-refractory prostate cancer. Clin Cancer Res 15, 3540-3549 (2009)
doi:10.1158/1078-0432.CCR-08-1857

32. V. S. Hawthorne, W. C. Huang, C. L. Neal, L. M. Tseng, M. C. Hung, D. Yu: ErbB2-mediated Src and signal transducer and activator of transcription 3 activation leads to transcriptional up-regulation of p21Cip1 and chemoresistance in breast cancer cells. Mol Cancer Res 7, 592-600 (2009)
doi:10.1158/1541-7786.MCR-08-0316

33. P. Laurent-Puig, A. Lievre, H. Blons: Mutations and response to epidermal growth factor receptor inhibitors. Clin Cancer Res 15, 1133-1139 (2009)
doi:10.1158/1078-0432.CCR-08-0905

34. I. Ischenko, P. Camaj, H. Seeliger, A. Kleespies, M. Guba, E. N. De Toni, B. Schwarz, C. Graeb, M. E. Eichhorn, K. W. Jauch, C. J. Bruns: Inhibition of Src tyrosine kinase reverts chemoresistance toward 5-fluorouracil in human pancreatic carcinoma cells: an involvement of epidermal growth factor receptor signaling. Oncogene 27, 7212-7222 (2008)
doi:10.1038/onc.2008.326

35. S. Kopetz, D. P. Lesslie, N. A. Dallas, S. I. Park, M. Johnson, N. U. Parikh, M. P. Kim, J. L. Abbruzzese, L. M. Ellis, J. Chandra, G. E. Gallick: Synergistic activity of the SRC family kinase inhibitor dasatinib and oxaliplatin in colon carcinoma cells is mediated by oxidative stress. Cancer Res 69, 3842-3849 (2009)
doi:10.1158/0008-5472.CAN-08-2246

36. Y. Ueda, T. Igishi, K. Hashimoto, H. Suyama, K. Araki, T. Sumikawa, K. Takeda, H. Nakazaki, K. Matsunami, M. Kodani, Y. Shigeoka, S. Matsumoto, E. Shimizu: Synergistic cell growth inhibition by the combination of amrubicin and Akt-suppressing tyrosine kinase inhibitors in small cell lung cancer cells: implication of c-Src and its inhibitor. Int J Oncol 34, 689-696 (2009)

37. T. Yoshida, I. Okamoto, W. Okamoto, E. Hatashita, Y. Yamada, K. Kuwata, K. Nishio, M. Fukuoka, P. A. Janne, K. Nakagawa: Effects of Src inhibitors on cell growth and epidermal growth factor receptor and MET signaling in gefitinib-resistant non-small cell lung cancer cells with acquired MET amplification. Cancer Sci 101, 167-172 (2010)
doi:10.1111/j.1349-7006.2009.01368.x

38. R. K. Murthy, P. K. Morrow, R. L. Theriault: Bone biology and the role of the RANK ligand pathway. Oncology (Williston Park) 23, 9-15 (2009)

39. P. D'Amelio, G. Isaia, G. C. Isaia: The osteoprotegerin/RANK/RANKL system: a bone key to vascular disease. J Endocrinol Invest 32, 6-9 (2009)

40. P. Soriano, C. Montgomery, R. Geske, A. Bradley: Targeted disruption of the c-src proto-oncogene leads to osteopetrosis in mice. Cell 64, 693-702 (1991)
doi:10.1016/0092-8674(91)90499-O

41. C. Lowe, T. Yoneda, B. F. Boyce, H. Chen, G. R. Mundy, P. Soriano: Osteopetrosis in Src-deficient mice is due to an autonomous defect of osteoclasts. Proc Natl Acad Sci U S A 90, 4485-4489 (1993)
doi:10.1073/pnas.90.10.4485

42. P. L. Schwartzberg, L. P. Xing, O. Hoffmann, C. A. Lowell, L. Garrett, B. F. Boyce, H. E. Varmus: Rescue of osteoclast function by transgenic expression of kinase- deficient Src in src-/- mutant mice. Genes Dev 11, 2835-2844 (1997)
doi:10.1101/gad.11.21.2835

43. A. Yano, S. Tsutsumi, S. Soga, M. J. Lee, J. Trepel, H. Osada, L. Neckers: Inhibition of Hsp90 activates osteoclast c-Src signaling and promotes growth of prostate carcinoma cells in bone. Proc Natl Acad Sci U S A 105, 15541-15546 (2008)
doi:10.1073/pnas.0805354105

44. B. F. Boyce, L. Xing, W. Shakespeare, Y. Wang, D. Dalgarno, J. Iuliucci, T. Sawyer: Regulation of bone remodeling and emerging breakthrough drugs for osteoporosis and osteolytic bone metastases. Kidney Int Suppl S2-S5 (2003)
doi:10.1046/j.1523-1755.63.s85.2.x

45. J. Edwards: Src kinase inhibitors: an emerging therapeutic treatment option for prostate cancer. Expert Opin Investig Drugs 19, 605-614 (2010)
doi:10.1517/13543781003789388

46. R. S. Finn: Targeting Src in breast cancer. Ann Oncol 19, 1379-1386 (2008)
doi:10.1093/annonc/mdn291

47. N. Rucci, M. Susa, A. Teti: Inhibition of protein kinase c-Src as a therapeutic approach for cancer and bone metastases. Anticancer Agents Med Chem 8, 342-349 (2008)
doi:10.2174/187152008783961905

48. F. Saad: Src as a therapeutic target in men with prostate cancer and bone metastases. BJU Int 103, 434-440 (2009)
doi:10.1111/j.1464-410X.2008.08249.x

49. F. Saad, A. Lipton: SRC kinase inhibition: Targeting bone metastases and tumor growth in prostate and breast cancer. Cancer Treat Rev 36, 177-184 (2009)
doi:10.1016/j.ctrv.2009.11.005

50. X. H. Zhang, Q. Wang, W. Gerald, C. A. Hudis, L. Norton, M. Smid, J. A. Foekens, J. Massague: Latent bone metastasis in breast cancer tied to Src-dependent survival signals. Cancer Cell 16, 67-78 (2009)
doi:10.1016/j.ccr.2009.05.017

51. C. Gennigens, C. Menetrier-Caux, J. P. Droz: Insulin-Like Growth Factor (IGF) family and prostate cancer. Crit Rev Oncol Hematol 58, 124-145 (2006)
doi:10.1016/j.critrevonc.2005.10.003

52. G. Pandini, R. Mineo, F. Frasca, C. T. Roberts, Jr., M. Marcelli, R. Vigneri, A. Belfiore: Androgens up-regulate the insulin-like growth factor-I receptor in prostate cancer cells. Cancer Res 65, 1849-1857 (2005)
doi:10.1158/0008-5472.CAN-04-1837

53. R. Nimmanapalli, E. O'Bryan, M. Huang, P. Bali, P. K. Burnette, T. Loughran, J. Tepperberg, R. Jove, K. Bhalla: Molecular characterization and sensitivity of STI-571 (imatinib mesylate, Gleevec)-resistant, Bcr-Abl-positive, human acute leukemia cells to SRC kinase inhibitor PD180970 and 17-allylamino-17-demethoxygeldanamycin. Cancer Res 62, 5761-5769 (2002)

54. R. A. Blake, M. A. Broome, X. Liu, J. Wu, M. Gishizky, L. Sun, S. A. Courtneidge: SU6656, a selective src family kinase inhibitor, used to probe growth factor signaling. Mol Cell Biol 20, 9018-9027 (2000)
doi:10.1128/MCB.20.23.9018-9027.2000

55. J. Bain, H. McLauchlan, M. Elliott, P. Cohen: The specificities of protein kinase inhibitors: an update. Biochem J 371, 199-204 (2003)
doi:10.1042/BJ20021535

56. A. A. Adjei, R. B. Cohen, G. S. Gordon, D. Hangauer, L. Dyster, G. Fetterly, S. Barrientes, D. S. Hong, A. Naing: Results of a phase I trial of KX2-391, a novel non-ATP competitive substrate-pocket directed SRC inhibitor, in patients with advanced malignancies. J Clin Oncol 27(15s), abstr. 3511 (2009)

57. G. M. Lau, G. M. Lau, G. L. Yu, I. H. Gelman, A. Gutowski, D. Hangauer, J. W. Fang: Expression of Src and FAK in Hepatocellular Carcinoma and the Effect of Src Inhibitors on Hepatocellular Carcinoma In vitro. Dig Dis Sci 54, 1465-1474 (2008)
doi:10.1007/s10620-008-0519-0

58. I. Recchia, N. Rucci, C. Festuccia, M. Bologna, A. R. MacKay, S. Migliaccio, M. Longo, M. Susa, D. Fabbro, A. Teti: Pyrrolopyrimidine c-Src inhibitors reduce growth, adhesion, motility and invasion of prostate cancer cells in vitro. Eur J Cancer 39, 1927-1935 (2003)
doi:10.1016/S0959-8049(03)00394-0

59. W. C. Shakespeare, Y. Wang, R. Bohacek, T. Keenan, R. Sundaramoorthi, C. Metcalf, III, A. Dilauro, S. Roeloffzen, S. Liu, J. Saltmarsh, G. Paramanathan, D. Dalgarno, S. Narula, S. Pradeepan, M. R. van Schravendijk, J. Keats, M. Ram, S. Liou, S. Adams, S. Wardwell, J. Bogus, J. Iuliucci, M. Weigele, L. Xing, B. Boyce, T. K. Sawyer: SAR of carbon-linked, 2-substituted purines: synthesis and characterization of AP23451 as a novel bone-targeted inhibitor of Src tyrosine kinase with in vivo anti-resorptive activity. Chem Biol Drug Des 71, 97-105 (2008)
doi:10.1111/j.1747-0285.2007.00615.x

60. I. Ischenko, M. Guba, M. Yezhelyev, A. Papyan, G. Schmid, T. Green, M. Fennell, K. W. Jauch, C. J. Bruns: Effect of Src kinase inhibition on metastasis and tumor angiogenesis in human pancreatic cancer. Angiogenesis 10, 167-182 (2007)
doi:10.1007/s10456-007-9071-3

61. M. V. Yezhelyev, G. Koehl, M. Guba, T. Brabletz, K. W. Jauch, A. Ryan, A. Barge, T. Green, M. Fennell, C. J. Bruns: Inhibition of SRC tyrosine kinase as treatment for human pancreatic cancer growing orthotopically in nude mice. Clin Cancer Res 10, 8028-8036 (2004)
doi:10.1158/1078-0432.CCR-04-0621

62. I. Ischenko, H. Seeliger, P. Camaj, A. Kleespies, M. Guba, M. E. Eichhorn, K. W. Jauch, C. J. Bruns: Src Tyrosine Kinase Inhibition Suppresses Lymphangiogenesis in vitro and in vivo. Curr Cancer Drug Targets Apr. 6 (Epub ahead of print) (2010)

63. S. Kimura, H. Naito, H. Segawa, J. Kuroda, T. Yuasa, K. Sato, A. Yokota, Y. Kamitsuji, E. Kawata, E. Ashihara, Y. Nakaya, H. Naruoka, T. Wakayama, K. Nasu, T. Asaki, T. Niwa, K. Hirabayashi, T. Maekawa: NS-187, a potent and selective dual Bcr-Abl/Lyn tyrosine kinase inhibitor, is a novel agent for imatinib-resistant leukemia. Blood 106, 3948-3954 (2005)
doi:10.1182/blood-2005-06-2209

64. T. Niwa, T. Asaki, S. Kimura: NS-187 (INNO-406), a Bcr-Abl/Lyn dual tyrosine kinase inhibitor. Anal Chem Insights 2, 93-106 (2007)

65. D. L. Wheeler, M. Iida, E. F. Dunn: The role of Src in solid tumors. Oncologist 14, 667-678 (2009)
doi:10.1634/theoncologist.2009-0009

66. F. Saad, A. Lipton: SRC kinase inhibition: targeting bone metastases and tumor growth in prostate and breast cancer. Cancer Treat Rev 36, 177-184 (2010)
doi:10.1016/j.ctrv.2009.11.005

67. L. C. Kim, L. Song, E. B. Haura: Src kinases as therapeutic targets for cancer. Nat Rev Clin Oncol 6, 587-595 (2009)
doi:10.1038/nrclinonc.2009.129

68. N. P. Shah: Dasatinib. Drugs Today (Barc) 43, 5-12 (2007)
doi:10.1358/dot.2007.43.1.1050791

69. J. Du, P. Bernasconi, K. R. Clauser, D. R. Mani, S. P. Finn, R. Beroukhim, M. Burns, B. Julian, X. P. Peng, H. Hieronymus, R. L. Maglathlin, T. A. Lewis, L. M. Liau, P. Nghiemphu, I. K. Mellinghoff, D. N. Louis, M. Loda, S. A. Carr, A. L. Kung, T. R. Golub: Bead-based profiling of tyrosine kinase phosphorylation identifies SRC as a potential target for glioblastoma therapy. Nat Biotechnol 27, 77-83 (2009)
doi:10.1038/nbt.1513

70. S. Moulder, K. Yan, F. Huang, K. R. Hess, C. Liedtke, F. Lin, C. Hatzis, G. N. Hortobagyi, W. F. Symmans, L. Pusztai: Development of Candidate Genomic Markers to Select Breast Cancer Patients for Dasatinib Therapy. Mol Cancer Ther Apr. 27 (Epub ahead of print) (2010)

71. J. Li, U. Rix, B. Fang, Y. Bai, A. Edwards, J. Colinge, K. L. Bennett, J. Gao, L. Song, S. Eschrich, G. Superti-Furga, J. Koomen, E. B. Haura: A chemical and phosphoproteomic characterization of dasatinib action in lung cancer. Nat Chem Biol 6, 291-299 (2010)
doi:10.1038/nchembio.332

72. F. R. Luo, Y. C. Barrett, Z. Yang, A. Camuso, K. McGlinchey, M. L. Wen, R. Smykla, K. Fager, R. Wild, H. Palme, S. Galbraith, A. Blackwood-Chirchir, F. Y. Lee: Identification and validation of phospho-SRC, a novel and potential pharmacodynamic biomarker for dasatinib (SPRYCELtrade mark), a multi-targeted kinase inhibitor. Cancer Chemother Pharmacol 62, 1065-1074 (2008)
doi:10.1007/s00280-008-0699-5

73. A. Serrels, I. R. MacPherson, T. R. Evans, F. Y. Lee, E. A. Clark, O. J. Sansom, G. H. Ashton, M. C. Frame, V. G. Brunton: Identification of potential biomarkers for measuring inhibition of Src kinase activity in colon cancer cells following treatment with dasatinib. Mol Cancer Ther 5, 3014-3022 (2006)
doi:10.1158/1535-7163.MCT-06-0382

74. G. S. Orphanos, G. N. Ioannidis, A. G. Ardavanis: Cardiotoxicity induced by tyrosine kinase inhibitors. Acta Oncol 48, 964-970 (2009) doi:10.1080/02841860903229124

75. R. Weichsel, C. Dix, L. Wooldridge, M. Clement, A. Fenton-May, A. K. Sewell, J. Zezula, E. Greiner, E. Gostick, D. A. Price, H. Einsele, R. Seggewiss: Profound inhibition of antigen-specific T-cell effector functions by dasatinib. Clin Cancer Res 14, 2484-2491 (2008)
doi:10.1158/1078-0432.CCR-07-4393

76. S. Blake, T. P. Hughes, G. Mayrhofer, A. B. Lyons: The Src/ABL kinase inhibitor dasatinib (BMS-354825) inhibits function of normal human T-lymphocytes in vitro. Clin Immunol 127, 330-339 (2008)
doi:10.1016/j.clim.2008.02.006

77. S. J. Blake, Lyons A. Bruce, C. K. Fraser, J. D. Hayball, T. P. Hughes: Dasatinib suppresses in vitro natural killer cell cytotoxicity. Blood 111, 4415-4416 (2008)
doi:10.1182/blood-2008-02-138701

Key Words: Src-family kinases, Src, Yes, Fyn, Blk, Fgr, Hck, Lck, Lyn, Yrk, Frk, cancer recurrence, metastasis, tyrosine kinase, chemotherapy resistance, osteoclasts, RANKL-osteoprotegerin, SSeCKS, Gravin, AKAP12, VEGF, tumor microenvironment, IGF-1, Dasatinib, BMS-354825, Sprycel, Bosutinib, SKI-606, Saracatinib, AZD0530, PD180970, SU6656, KXO1, KX2-391, CGP76030, AP23451, AZM475271, INNO-406, NS-187, XL-999, XL-228, Review

Send correspondence to: Irwin H. Gelman, Department of Cancer Genetics, Roswell Park Cancer Institute, Elm and Carlton Streets, Buffalo, NY 14263 U.S.A., Tel: 716-845-7681, Fax: 716-845-1698, E-mail:Irwin.Gelman@roswellpark.org