Cellular Characterization of a Novel Focal Adhesion Kinase Inhibitor
Focal adhesion kinase (FAK) is a member of a family of non-receptor protein-tyrosine kinases that regulates integrin and growth factor signaling pathways involved in cell migration, proliferation, and survival. FAK expression is increased in many cancers, including breast and prostate cancer. Here we describe perturbation of adhesion-mediated signaling with a FAK inhibitor, PF-573,228. In vitro, this compound inhibited purified recombinant catalytic fragment of FAK with an IC50 of 4 nm. In cultured cells, PF-573,228 inhibited FAK phosphorylation on Tyr397 with an IC50 of 30–100 nm. Treatment of cells with concentrations of PF-573,228 that significantly decreased FAK Tyr397 phosphorylation failed to inhibit cell growth or induce apoptosis. In contrast, treatment with PF-573,228 inhibited both chemotactic and haptotactic migration concomitant with the inhibition of focal adhesion turnover. These studies show that PF-573,228 serves as a useful tool to dissect the functions of FAK in integrin-dependent signaling pathways in normal and cancer cells and forms the basis for the generation of compounds amenable for preclinical and patient trials. Focal adhesion kinase (FAK) is a member of a family of non-receptor protein-tyrosine kinases that regulates integrin and growth factor signaling pathways involved in cell migration, proliferation, and survival. FAK expression is increased in many cancers, including breast and prostate cancer. Here we describe perturbation of adhesion-mediated signaling with a FAK inhibitor, PF-573,228. In vitro, this compound inhibited purified recombinant catalytic fragment of FAK with an IC50 of 4 nm. In cultured cells, PF-573,228 inhibited FAK phosphorylation on Tyr397 with an IC50 of 30–100 nm. Treatment of cells with concentrations of PF-573,228 that significantly decreased FAK Tyr397 phosphorylation failed to inhibit cell growth or induce apoptosis. In contrast, treatment with PF-573,228 inhibited both chemotactic and haptotactic migration concomitant with the inhibition of focal adhesion turnover. These studies show that PF-573,228 serves as a useful tool to dissect the functions of FAK in integrin-dependent signaling pathways in normal and cancer cells and forms the basis for the generation of compounds amenable for preclinical and patient trials. The ability of cells to respond appropriately to environmental cues is critical to maintaining cellular, tissue, and organism homeostasis. One such environmental cue is derived from cellular adhesion to the extracellular matrix. The loss of adhesion-dependent cellular regulation can lead to increased cellular proliferation, decreased cell death, changes in cellular differentiation status, and altered cellular migratory capacity, all of which are critical components of cell carcinogenesis and metastatic progression. The FAK 4The abbreviations used are: FAK, focal adhesion kinase; ELISA, enzyme-linked immunosorbant assay; FN, fibronectin; PBS, phosphate-buffered saline; TIRF, total interference reflection fluorescence; MDCK, Madin-Darby canine kidney. 4The abbreviations used are: FAK, focal adhesion kinase; ELISA, enzyme-linked immunosorbant assay; FN, fibronectin; PBS, phosphate-buffered saline; TIRF, total interference reflection fluorescence; MDCK, Madin-Darby canine kidney. family kinases (which include FAK and Pyk2) regulate cell adhesion, migration, and proliferation in a variety of cell types (for review see Refs. 1Parsons J.T. J. Cell Sci. 2003; 116: 1409-1416Crossref PubMed Scopus (1119) Google Scholar, 2Mitra S.K. Hanson D.A. Schlaepfer D.D. Nat. Rev. Mol. Cell Biol. 2005; 6: 56-68Crossref PubMed Scopus (1902) Google Scholar, 3Abbi S. Guan J.L. Histol. Histopathol. 2002; 17: 1163-1171PubMed Google Scholar). Adhesion of cells to the extracellular matrix is mediated by heterodimeric transmembrane integrin receptors located within sites of close opposition to the underlying matrix called focal adhesions. Integrin engagement and clustering stimulates FAK phosphorylation on Tyr397, creating a high affinity binding site for Src and Src family kinases. The FAK·Src complex phosphorylates many components of the focal adhesion, resulting in changes in adhesion dynamics and the initiation of signaling cascades. In addition to FAK catalytic activity, FAK also functions as a scaffold to organize structural and signaling proteins within focal adhesions. The importance of FAK as a regulator of normal cellular function is underscored by the number of cancers reported to have alterations in FAK expression and/or activity, including colon, breast, thyroid, prostate, cervical, ovarian, head and neck, oral, liver, stomach, sarcoma, glioblastoma, and melanoma (4Gabarra-Niecko V. Schaller M.D. Dunty J.M. Cancer Metastasis Rev. 2003; 22: 359-374Crossref PubMed Scopus (296) Google Scholar, 5McLean G.W. Carragher N.O. Avizienyte E. Evans J. Brunton V.G. Frame M.C. Nat. Rev. Cancer. 2005; 5: 505-515Crossref PubMed Scopus (839) Google Scholar). Additionally, alterations in FAK expression and/or activity have been associated with tumorigenesis and increased metastatic potential (4Gabarra-Niecko V. Schaller M.D. Dunty J.M. Cancer Metastasis Rev. 2003; 22: 359-374Crossref PubMed Scopus (296) Google Scholar, 5McLean G.W. Carragher N.O. Avizienyte E. Evans J. Brunton V.G. Frame M.C. Nat. Rev. Cancer. 2005; 5: 505-515Crossref PubMed Scopus (839) Google Scholar). Currently, it is unclear how the catalytic and/or scaffolding function of FAK contributes to tumor progression. To date studies of FAK function have relied on the expression of dominant interfering mutants or elimination of FAK expression by genetic knock-out, antisense oligonucleotide expression, or small interfering RNA. Herein, we report the biochemical and cellular characterization of a novel small molecule inhibitor, PF-573,228 (here after referred to as PF-228), that targets FAK catalytic activity. The inhibitor interacts with FAK in the ATP-binding pocket and effectively blocks the catalytic activity of recombinant FAK protein or endogenous FAK expressed in a variety of normal and cancer cell lines. Treatment of cells with PF-228 blocked FAK phosphorylation on Tyr397 and concomitantly reduced the tyrosine phosphorylation of paxillin, a recognized downstream effector of FAK signaling. Drug treatment of normal and cancer cells resulted in decreased cell migration and inhibited adhesion turnover, biological activities previously ascribed to FAK. Interestingly, inhibition of FAK activity had little effect on normal or cancer cell growth or apoptosis in culture. PF-573,228 provides an appropriate tool to dissect the role of FAK in regulation of cell adhesion signaling and the regulation of adhesion dynamics. Chemical Synthesis—PF-573,228 was identified through a combination of high throughput screening, structure based drug design, and conventional medicinal chemistry approaches (38Luzzio M.J. Autry C. Berliner M. Coleman K. Cooper B. Emerson E. Griffor M. Hulford C. Jani J. Kath J. LaGreca S. Lin J. Lorenzen M. Matt E. Martinez-Alsina L. Patel N. Richter D. Schmitt E. Ung E. Vajdos F. Wessell M. Whalen P. Yao L. Roberts W.G. Proc. Annu. Meet. Am. Assoc. Cancer Res. 2007; 48: 1283Google Scholar). PF-573,228 was prepared according to the procedures described in a patent (35Kath J.C. Luzzio M.J. U. S. Patent Application WO 2004056786. 2004; Google Scholar). The structure and inhibitory activity are shown in Fig. 1. Recombinant Kinase Assay—Purified activated FAK kinase domain (amino acids 410–689) was reacted with 50 μm ATP, and 10 μg/well of a random peptide polymer of Glu and Tyr (molar ratio of 4:1), poly(Glu/Tyr) in kinase buffer (50 mm HEPES, pH 7.5, 125 mm NaCl, 48 mm MgCl2) for 15 min. Phosphorylation of poly(Glu/Tyr) was challenged with serially diluted compounds at ½-Log concentrations starting at a top concentration of 1 μm. Each concentration was run in triplicate. Phosphorylation of poly(Glu/Tyr) was detected with a general anti-phospho-tyrosine (PY20) antibody, followed by horseradish peroxidase-conjugated goat anti-mouse IgG antibody. The standard horseradish peroxidase substrate 3, 3′, 5, 5′-tetramethylbenzidine was added, and Optical Density readings at 450 nm were obtained following the addition of stop solution (2 m H2SO4). The IC50 values were determined using the Hill slope model. Broad kinase selectivity profiling was performed using the KinaseProfiler™ selectivity screening service available through Upstate Biotechnology, Inc. For more information please see: www.upstate.com/discovery/services/kp_overview.q. Cellular Kinase Assays—Using the GeneSwitch inducible system from Invitrogen, stable A431 epithelial carcinoma clones were generated to express either wild type V5-tagged FAK protein or mutant FAK Y397F V5-tagged protein under the inducible regulation of mifepristone (36Ung E.J. Whalen P.M. Roberts W.G. Cell Assay for Inhibitors of Focal Adhesion Kinase (FAK). 2002; (Patent Cooperation Treaty application number W004027018)Google Scholar). Stable clones were grown in Dulbecco’s modified Eagle’s medium, 10% fetal bovine serum, 750 μg/ml Zeocin, and 50 μg/ml Hygromycin. One day prior to running the FAK cell ELISA, A431·FAKwt cells were seeded at 1.2 × 106 cells/ml in growth medium in 96-well U-bottom plates. After 4–6 h at 37 °C, 5% CO2, FAK expression was induced with 0.1 nm miferpristone. Uninduced controls were included. Goat anti-mouse or anti-rabbit plates were subsequently coated with either anti-V5 or anti-FAK (1.0 μg/ml) or an irrelevant antibody control in Superblock Tris-buffered saline buffer. Anti-V5- or anti-FAK-coated plates were blocked in 3% bovine serum albumin, for 1 h at The cells were with ½-Log starting at a top concentration of 1 μm for at 37 °C, 5% from cells with concentrations of compound were prepared in buffer (50 mm pH mm NaCl, 1 mm 1 mm 1 mm and and to the or anti-FAK-coated plates to total induced or total FAK was used to FAK Tyr397, followed by antibody. peroxidase substrate was added, and plates were at 450 nm. The IC50 values were determined using the Hill slope model. For cells were with the concentrations of inhibitor for the of prior to in buffer (50 mm HEPES, m NaCl, mm and pH 1 mm mm and 1 mm mm and 10 mm For cells were in medium in the or of the concentrations of inhibitor and were to to plates coated with μg/ml for in the or of The cells were in and of cell was performed using of Cell and were performed by 1 × or of a in h prior to treatment with the concentrations of inhibitor for the cells were and were performed using a cell according to the or cells were for h h for with the concentrations of inhibitor prior to for h in medium as a The cell were in in the The the standard of of performed in Cell migration was using modified as described previously M.J. J.T. J. Biol. PubMed Scopus Google Scholar). was determined to the number of cells to the of the in For growth 10% fetal bovine serum was in the To the of the was coated with μg/ml in in the and was in the After the at 4 °C, the were and Dulbecco’s modified Eagle’s medium serum was to For inhibitor 1 × cells were with inhibitor for prior to and addition to the The inhibitor in both of for the of the migration The cells were to for h at 37 cells on the of the were with a The cells that had to the of the were in PBS, with at for with PBS, and with was by using a The the standard of of performed in to the Each was using of the for of the was at the 5% we to which from controls using was as on were with a and cells the were for h at 37 by PF-573,228 was at the of and was 1 h was performed using a with a interference and a The were with a and using the of cells were as the and the cell was by the of the cell by the total of the and Cell cells were and with concentrations of PF-573,228 for 1 h in the of For cells were in for serum and in the of of the FAK The cells were μg/ml) (for or (for interference for in the of The cells were for with with for and blocked in goat serum, bovine serum for min. The cells were with to or FAK were with or were using a with a a and the and Adhesion cells were with using the system on were with PF-573,228 and with a as described were at 37 using a with a × a and The were for min. was performed using The of adhesion was and the for adhesion was Adhesion was as the of the for the adhesion to the from and the for the adhesion to to from were of FAK Kinase in and in is a inhibitor of with for FAK family protein-tyrosine kinases PF-573,228 was of compounds for biochemical and cellular activities FAK and was for studies based on for FAK, activity kinases and for in a fragment of FAK, the inhibitory of PF-228 were by the phosphorylation of poly(Glu/Tyr) by recombinant FAK PF-228 inhibited catalytic activity in this with an IC50 of 4 nm and In a PF-228 inhibited with an IC50 of 1 μm PF-228 was a available of recombinant Biotechnology, M. P. J. PubMed Scopus Google at 1 μm In this 1 μm PF-228 the IC50 for inhibited the kinases and and and the inhibitory effect of PF-228 on recombinant and kinase activity was The IC50 values for PF-228 on all kinases were the IC50 for FAK selectivity of of of in a were used to the activity of A431 epithelial carcinoma cells FAK under the inducible regulation of mifepristone were with concentrations of and the phosphorylation of FAK Tyr397 was in a PF-228 inhibited FAK phosphorylation in A431 cells with IC50 of nm To the ability of PF-228 to endogenous FAK activity, the phosphorylation of FAK Tyr397 was in cultured cells cell Treatment of cells with concentrations of PF-228 for inhibition of Tyr397 phosphorylation of nm. inhibition was with PF-228 concentrations of μm PF-228 also blocked FAK Tyr397 phosphorylation in and and cells with IC50 of nm FAK Tyr397 phosphorylation decreased within 15 of treatment with 1 μm PF-228 and was by a in the phosphorylation of a downstream FAK to and is activated within focal J.T. J. Cell Sci. 2003; 116: 1409-1416Crossref PubMed Scopus (1119) Google Scholar, 2Mitra S.K. Hanson D.A. Schlaepfer D.D. Nat. Rev. Mol. Cell Biol. 2005; 6: 56-68Crossref PubMed Scopus (1902) Google Scholar). the of FAK was in cells 1 h after treatment with concentrations of the FAK in the of the was a in FAK phosphorylation within focal in the underlying the cell of FAK on Cell and signaling have been reported to regulate cell growth and apoptosis Schaller M.D. 2004; PubMed Scopus Google Scholar, Guan J.L. Histol. Histopathol. Google Scholar). increased FAK expression been with increased in Refs. V. Schaller M.D. Dunty J.M. Cancer Metastasis Rev. 2003; 22: 359-374Crossref PubMed Scopus (296) Google and 5McLean G.W. Carragher N.O. Avizienyte E. Evans J. Brunton V.G. Frame M.C. Nat. Rev. Cancer. 2005; 5: 505-515Crossref PubMed Scopus (839) Google the growth inhibitory of PF-228 on cells, a prostate with increased FAK expression J.T. PubMed Scopus Google as as a cell were or cells were with PF-228 day for after which the total number of cells was determined and with the controls Treatment of cells with 1 μm PF-228 failed to inhibit cell this concentration of PF-228 was to inhibit FAK phosphorylation by of and cells was significantly inhibited at the concentration of 10 μm PF-228 To the inhibition of growth at concentrations was to of D. S. N. K. N. S. J. M. PubMed Scopus Google were with concentrations of of growth was using 10 μm PF-228 that treatment of cells at high concentrations of drug inhibit cell growth in a of PF-228 on cell of of in a To inhibition of FAK catalytic activity induced or cells were with the concentrations of PF-228 for and apoptosis was using an which the of in previously reported of cells for h induced apoptosis J. Cell Biol. PubMed Scopus Google Scholar). cells for h also induced to a the apoptosis induced in cells apoptosis in Treatment of or cells with PF-228 failed to induce at concentrations shown to significantly FAK Tyr397 of FAK on Cell and adhesion to extracellular matrix components such as stimulates FAK activity and cell J.T. J. Cell Sci. 2003; 116: 1409-1416Crossref PubMed Scopus (1119) Google Scholar, 2Mitra S.K. Hanson D.A. Schlaepfer D.D. Nat. Rev. Mol. Cell Biol. 2005; 6: 56-68Crossref PubMed Scopus (1902) Google Scholar). To the ability of PF-228 to adhesion-dependent FAK cells were in medium concentrations of PF-228 or control for 1 h and on for in the or of inhibitor Treatment of cells with μm PF-228 reduced FAK Tyr397 phosphorylation by In treatment of cells with PF-228 also reduced the tyrosine phosphorylation of on in a To the of PF-228 on the of cell cells were in the of the inhibitor and on μg/ml for in the of a inhibition of FAK phosphorylation and a loss of in focal with μm PF-228 role for FAK catalytic activity in cell migration was in using 10% serum or as cells were with the concentrations of PF-228 for and to 10% serum or for h in the of inhibitor Treatment of cells with μm PF-228 concentration that FAK phosphorylation by had effect on random migration of cells at this concentration of both and migration were significantly Treatment of cells with 10 μm PF-228 blocked random migration and also blocked serum and migration of PF-228 on cell control standard significantly from The control standard significantly from in a of cell migration by PF-228 was also in performed with cells Treatment of with 1 μm PF-228 significantly reduced the of of cells the the from the Cell migration the and of focal a as focal adhesion K. J.T. Nat. Cell Biol. 2004; 6: PubMed Scopus Google Scholar). The of PF-228 treatment on focal adhesion were using a in cells Focal in cells at the of the were by The of the was by the of cells adhesion and and the adhesion was In contrast, all of the in the cells with PF-228 a in the of and significantly the in the cells adhesion of and for 1 and 10 μm Treatment with PF-228 inhibit the of Interestingly, the in the cells with PF-228 a of turnover, a of the of the cell and In this we the biochemical and cellular of a novel and small molecule inhibitor of FAK, The inhibitor effectively blocked recombinant FAK catalytic activity and FAK Tyr397 phosphorylation in cultured Treatment of cultured cells with concentrations of the inhibitor that significantly inhibited FAK of Tyr397 failed to inhibit growth or apoptosis. treatment of cells with PF-228 resulted in inhibition of serum or migration and decreased focal adhesion turnover. increased FAK expression and activity are of many cancers and the that increased cell migration tumor cells with increased metastatic FAK have potential for the treatment of cancer. PF-228 inhibited recombinant FAK kinase activity with an IC50 of 4 kinase In cultured cells, PF-228 inhibited FAK phosphorylation to of FAK phosphorylation in A431 cells to FAK an IC50 of nm with nm in cells by of inhibitory activity of PF-228 in cancer cell using a of IC50 of nm. it is that the to in and of FAK phosphorylation in cells generated to the it is that the in the cell The A431 cells FAK at significantly The of is to the and activity of FAK, which it more to inhibition by this is the that FAK to more to FAK inhibition normal cells, with in studies FAK functions have been in the adhesion-dependent signaling that regulate cell death, and migration J.T. J. Cell Sci. 2003; 116: 1409-1416Crossref PubMed Scopus (1119) Google Scholar, 2Mitra S.K. Hanson D.A. Schlaepfer D.D. Nat. Rev. Mol. Cell Biol. 2005; 6: 56-68Crossref PubMed Scopus (1902) Google Scholar, V. Schaller M.D. Dunty J.M. Cancer Metastasis Rev. 2003; 22: 359-374Crossref PubMed Scopus (296) Google Scholar, 5McLean G.W. Carragher N.O. Avizienyte E. Evans J. Brunton V.G. Frame M.C. Nat. Rev. Cancer. 2005; 5: 505-515Crossref PubMed Scopus (839) Google Scholar). The FAK kinase activity in the regulation of cell migration, concentrations of PF-228 that inhibit FAK activity cell growth or induce apoptosis in In contrast, studies have FAK in the regulation of both K. E. J. Cell Biol. PubMed Scopus Google Scholar, D.A. W.G. J. Biol. PubMed Scopus Google Scholar, G.W. B. L. F. K. D. P. Frame M.C. 2004; PubMed Scopus Google Scholar, E. S.K. W.G. Mol. Cell Biol. 2004; PubMed Scopus Google Scholar, D. Schlaepfer D.D. P. S. J. Cell Biol. PubMed Scopus Google Scholar). of the domain of FAK, which FAK apoptosis in cell D. Schlaepfer D.D. P. S. J. Cell Biol. PubMed Scopus Google Scholar, M.C. Res. 2002; PubMed Scopus Google Scholar). loss of FAK expression in a of cancer tumor which is by increased cell G.W. B. L. F. K. D. P. Frame M.C. 2004; PubMed Scopus Google Scholar). and a role for FAK in apoptosis on the loss of FAK or the of dominant interfering mutants of FAK and the scaffolding function of FAK in the regulation of apoptosis. of FAK kinase activity with PF-228 was to induce apoptosis in in cell types to the in which cells are grown and the pathways that are activated to or to the that the scaffolding function of FAK kinase activity is for the regulation of apoptosis. signaling have been shown to effect cellular proliferation by the from the to the role for FAK in this is of wild type FAK been shown to growth of a variety of (4Gabarra-Niecko V. Schaller M.D. Dunty J.M. Cancer Metastasis Rev. 2003; 22: 359-374Crossref PubMed Scopus (296) Google Scholar, 5McLean G.W. Carragher N.O. Avizienyte E. Evans J. Brunton V.G. Frame M.C. Nat. Rev. Cancer. 2005; 5: 505-515Crossref PubMed Scopus (839) Google which in was to an in cell through to Guan J.L. J. Cell Biol. PubMed Scopus Google Scholar). of the dominant domain of FAK growth (4Gabarra-Niecko V. Schaller M.D. Dunty J.M. Cancer Metastasis Rev. 2003; 22: 359-374Crossref PubMed Scopus (296) Google Scholar, 5McLean G.W. Carragher N.O. Avizienyte E. Evans J. Brunton V.G. Frame M.C. Nat. Rev. Cancer. 2005; 5: 505-515Crossref PubMed Scopus (839) Google and dominant interfering mutants decreased to Guan J.L. J. Cell Biol. PubMed Scopus Google Scholar). cells derived from FAK in cell growth D. S. N. K. N. S. J. M. PubMed Scopus Google Scholar). treatment of and cells with high concentrations of PF-228 for blocked cell growth this concentration and of PF-228 treatment also blocked growth of FAK concentrations of PF-228 inhibited many kinases the growth inhibitory reported at the high concentration of PF-228 inhibition of kinases in addition to FAK. The regulation of cell migration is a of FAK function on cellular activity. FAK knock-out, and expression of dominant interfering mutants all cell migration D. S. N. K. N. S. J. M. PubMed Scopus Google Scholar, Mol. Biol. PubMed Scopus Google Scholar, J. Res. 2004; Google Scholar, D.A. J.C. Schlaepfer D.D. Cancer Res. Google Scholar, N. M. J.T. J. Cell Sci. 2005; PubMed Scopus Google Scholar). migration is in FAK cells wild type FAK FAK, the mutant of FAK or the domain of FAK, both the scaffolding function and kinase activity of FAK in the regulation of cell migration Schlaepfer D.D. J. Cell Sci. PubMed Google Scholar, S.K. Mol. Cell Biol. PubMed Scopus Google Scholar). The that PF-228 inhibited cell migration in a a role for FAK kinase activity in the regulation of cell inhibition of FAK kinase activity with PF-228 decreased focal adhesion turnover, a of cell migration K. J.T. Nat. Cell Biol. 2004; 6: PubMed Scopus Google Scholar). the in the cells failed to a the of the is with the of in cells M.C. J. Cell Biol. 2002; PubMed Scopus Google Scholar, P. Proc. Sci. U. S. 2004; PubMed Scopus Google Scholar). it is that inhibition of FAK from the substrate to to the in the lead to in the generation and/or of at sites of adhesion, which cell migration and tumor M. J. Cell Biol. PubMed Scopus Google Scholar, N. K. D. Proc. Sci. U. S. PubMed Scopus Google Scholar). and reported the of a FAK inhibitor from on cell function L. S. D. D.D. Mol. 2007; Scholar). to the reported an inhibition of cell also induced apoptosis and inhibited growth of cells in culture. These to cell type or more of the is a with activity and FAK and activity in kinase with an IC50 of nm E. K. D. S. J. S. J. Google Scholar). PF-228 in kinase and selectivity to and many kinases for of FAK inhibition and resulting cellular The of on kinases was reported L. S. D. D.D. Mol. 2007; Scholar). PF-228 and compounds useful to the role of adhesion signaling in regulation of cellular proliferation, and migration and to have in the and/or treatment of cancer. small with FAK inhibitory activity have been are either and or have been in cellular K. D. V. J. J. N. N. PubMed Scopus Google Scholar, K. D. V. J. N. PubMed Scopus Google Scholar). The of and in to the role of FAK inhibition in the regulation of tumorigenesis and metastatic progression. The importance of FAK kinase activity in the regulation of cell migration as potential FAK activity in of to a concomitant in tumor or FAK W.G. Ung E. Whalen P. Cooper B. Hulford C. Autry C. Richter D. Emerson E. Lin J. Kath J. Coleman K. Yao L. Martinez-Alsina L. Lorenzen M. Berliner M. Luzzio M. Patel N. Schmitt E. LaGreca S. Proc. Annu. Meet. Am. Assoc. Cancer Res. 2007; 48: Scholar). a FAK inhibitor for the treatment of cancer W.G. Ung E. Whalen P. Cooper B. Hulford C. Autry C. Richter D. Emerson E. Lin J. Kath J. Coleman K. Yao L. Martinez-Alsina L. Lorenzen M. Berliner M. Luzzio M. Patel N. Schmitt E. LaGreca S. Proc. Annu. Meet. Am. Assoc. Cancer Res. 2007; 48: Scholar). for with the growth and for of the with