Src family kinase oncogenic potential and pathways in prostate cancer as revealed by AZD0530 - PubMed (original) (raw)

Src family kinase oncogenic potential and pathways in prostate cancer as revealed by AZD0530

Y-M Chang et al. Oncogene. 2008.

Abstract

Prostate cancer is the most frequently diagnosed cancer in American men. We have previously demonstrated that Src mediates androgen-independent proliferation in prostate cancer. We sought to investigate the Src-mediated oncogenic pathways and tumor biology using AZD0530, a novel Src family kinase/Abl dual-kinase inhibitor that is entering phase II clinical trials. We show that while both Src and Abl are expressed in all prostate cancer cell lines, Src but not Abl is activated in the prostate. Furthermore, Src activation is inhibited by AZD0530 in a rapid and dose-dependent manner. We show that Src mediates cell proliferation in DU145 and PC3 cells at the G1 phase of cell cycle. Src inhibition resulted in decreased binding of beta-catenin to the promoters of G1 phase cell cycle regulators cyclin D1 and c-Myc. C-Myc may also be regulated at the protein level by extracellular signal-regulated kinase 1/2 and GSK3beta. Cell motility factors focal adhesion kinase, p130CAS and paxillin activation in DU145 and PC3 cells were also inhibited. Administration of AZD0530 in mice reduced orthotopic DU145 xenograft growth by 45%. We have further delineated the Src-mediated oncogenic growth and migration pathways in prostate cancer and established mechanistic rationale for Src inhibition as novel therapy in the treatment of prostate cancer.

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Figures

Figure 1

Figure 1

AZD0530 inhibits Src activation through inhibition of Y419 phosphorylation. (a) The chemical structure of AZD0530. (b) Commonly used cell lines were harvested and probed with Abl, p-Abl, phospho-Src Y419 and Src antibodies demonstrating relative increased ratio of activated-to-total Src in DU145 and PC3 cells. (c) Src autophosphorylation in DU145 and PC3 cells were inhibited in a dose-dependent manner by AZD0530 following 30-min treatment (left and right, respectively) or rapid manner by 1 μM of AZD0530 (middle).

Figure 2

Figure 2

AZD0530 inhibits cell proliferation at G0/G1-S transition. (a) Single administration of AZD0530 inhibited cell proliferation in a dose-dependent manner in 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide assay showing immortalized cells on average being more resistant than malignant cell lines. (b) AZD0530 inhibited DU145 and PC3 proliferation in a dose-dependent manner over time. (c) AZD0530 induced G1/S cell cycle arrest but not apoptosis in DU145 and PC3 cells. (d) AZD0530 did not induce apoptosis in DU145 and PC3 cells after 2 days, as shown by the lack of caspase 3 cleavage. Columns, mean; bars, standard error; *P<0.05 (_n_=3); **P<0.01 (_n_=3).

Figure 3

Figure 3

AZD0530 inhibits cell proliferation through β-catenin, ERK1/2 and GSK3β-mediated cyclin D1 and c-myc regulation. (a) DU145 and PC3 cells treated with AZD0530 shows dose-dependent decreases in levels of cyclin D1 and c-Myc. Corresponding rebound of phospho-Src and c-Myc is seen over time. (b) Cyclin D1 and c-Myc transcript levels decreased following 4 μM AZD0530 treatment. (c) AZD0530 treatment did not inhibit signal and transducer of transcription 3 activation after 30 min but downregulated β-catenin after 24 h (top). DU145 and PC3 treated with 4 μM of AZD0530 for 12 h shows decreased binding of β-catenin to cyclin D1 and c-Myc promoter regions (bottom). (d) AZD0530 inhibited ERK1/2 and GSK3β phosphorylation in DU145 and AKT and GSK3β phosphorylation in PC3 after 30 min.

Figure 4

Figure 4

AZD0530 inhibits cell migration through Src-mediated FAK activation. (a) DU145 (top) and PC3 (bottom) treated with AZD0530 shows dose-dependent decrease in cell migration. (b) Paxillin, p130CAS and p-FAK (Y576/577) phosphorylation were inhibited in DU145 and PC3, following AZD0530 treatment for 30 min. Columns, mean; bars, standard error; *P<0.05 (_n_=3); **P<0.01 (_n_=3).

Figure 5

Figure 5

AZD0530 inhibits tumor growth in vivo. (a) 25 mg/kg of AZD0530 was administered orally daily starting 2 days after orthotopic injection of 2 million DU145 cells. Mice were euthanized after 54 days. Established tumors were harvested and weighed. (b) Immunohistochemical analysis of tumor samples from (a) using specific phospho-Src Y419 antibody as described. Dots and triangles, tumor samples; columns, mean; bars, standard error; *P<0.05 (_n_=10).

References

    1. Albanese C, Johnson J, Watanabe G, Eklund N, Vu D, Arnold A, et al. Transforming p21ras mutants and c-Ets-2 activate the cyclin D1 promoter through distinguishable regions. J Biol Chem. 1995;270:23589–23597. - PubMed
    1. Bang YJ, Pirnia F, Fang WG, Kang WK, Sartor O, Whitesell L, et al. Terminal neuroendocrine differentiation of human prostate carcinoma cells in response to increased intracellular cyclic AMP. Proc Natl Acad Sci USA. 1994;91:5330–5334. - PMC - PubMed
    1. Barone MV, Courtneidge SA. Myc but not Fos rescue of PDGF signalling block caused by kinase-inactive Src. Nature. 1995;378:509–512. - PubMed
    1. Biscardi JS, Ishizawar RC, Silva CM, Parsons SJ. Tyrosine kinase signalling in breast cancer: epidermal growth factor receptor and c-Src interactions in breast cancer. Breast Cancer Res. 2000;2:203–210. - PMC - PubMed
    1. Bjelfman C, Meyerson G, Cartwright CA, Mellstrom K, Hammerling U, Pahlman S. Early activation of endogenous pp60src kinase activity during neuronal differentiation of cultured human neuroblastoma cells. Mol Cell Biol. 1990;10:361–370. - PMC - PubMed

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