CCL2 is a potent regulator of prostate cancer cell migration and proliferation.

Tumor cells in the bone interact with the microenvironment to promote tumor cell survival and proliferation, resulting in a lethal phenotype for patients with advanced prostate cancer. Monocyte chemoattractant protein 1 (CCL2) is a member of the CC chemokine family and is known to promote monocyte chemotaxis to sites of inflammation. Here we have shown that human bone marrow endothelial (HBME) cells secrete significantly higher levels of CCL2 compared to human aortic endothelial cells and human dermal microvascular endothelial cells. Furthermore, we demonstrate that CCL2 is a potent chemoattractant of prostate cancer epithelial cells, and that stimulation of PC-3 and VCaP cells resulted in a dose-dependent activation of PI3 kinase/Akt signaling pathway. Activation of the PI3 kinase/Akt pathway was found to be vital to the proliferative effects of CCL2 stimulation of both PC-3 and VCaP cells. Additionally, CCL2 stimulated the phosphorylation of p70-S6 kinase (a downstream target of Akt) and induced actin rearrangement, resulting in a dynamic morphologic change indicative of microspike formation. These data suggest that bone marrow endothelial cells are a major source of CCL2, and that an elevated secretion of CCL2 recruits prostate cancer epithelial cells to the bone microenvironment and regulates their proliferation rate.

Development of the VCaP androgen-independent model of prostate cancer.

Prostate epithelial cell growth is dependent on the presence of androgens, and transition of prostate cancer to an androgen-independent phenotype results in a highly aggressive, currently incurable cancer. We have developed a new preclinical model of androgen-independent prostate cancer derived from the VCaP prostate cancer epithelial cell line. VCaP cells were subcutaneously implanted and serially passaged in castrated male severe combined immunodeficient mice. Androgen independence was confirmed by WST-1 (a tetrazolium salt) cell proliferation assay in the absence or presence of dihydrotesterone (1-100 nM). VCaP androgen-sensitive cells responded dose dependently to dihydrotesterone, whereas VCaP androgen-independent cells did not alter their proliferation in response to dihydrotesterone. Gene expression of androgen receptor, B-cell lymphoma-2, prostate cancer antigen 3, prostate acid phosphatase, 6 transmembrane epithelial antigen of the prostate, and survivin was determined by polymerase chain reaction amplification. B-cell lymphoma-2 expression was up regulated in the VCaP androgen-independent lines compared to the VCaP androgen-sensitive, suggesting a possible mechanism of androgen independence. Furthermore, tumor-associated angiogenesis was assessed by immunofluorescence confocal microscopy of CD31. VCaP androgen-independent tumors showed enhanced angiogenesis compared to VCaP androgen-sensitive tumors. These results illustrate the development of a novel model of prostate cancer androgen independence and provide a new system to study angiogenesis and the transition to androgen independence.

Differential expression analysis of MIM (MTSS1) splice variants and a functional role of MIM in prostate cancer cell biology.

We previously identified MIM-A (missing in metastasis, MTSS1) by differential display techniques as missing in invasive, metastatic bladder cancer cell lines and suggested that MIM-A is a novel putative metastasis suppressor gene. Characterization of the MIM gene revealed a WH2 (Wiskott-Aldrich syndrome protein homology 2) domain in the C-terminus that is known to bind actin monomers and regulate organization of the actin cytoskeleton. Here, we further describe two alternatively splice variants of MIM-A, called MIM(12del) and MIM-B, which share > 50% amino acid sequence homology with MIM-A in the C-terminal domain. We show that expression of all three transcripts is down-regulated in prostate cancer cell lines and tumor samples from patients. In addition, we generated stably-transfected PC-3 cells overexpressing MIM-A to evaluate the importance of MIM-A in prostate cancer biology. The initial experiments show that expression of MIM decreased the number of actin filaments and was associated with a decrease in the G:F actin ratio. Overexpression of MIM-A had no effect on PC-3 cell adhesion to extracellular matrices, as well as no effect on PC-3 motility. Further, overexpression of MIM-A reduced the rate of PC-3 cell proliferation. These results support the hypothesis that MIM-A is an actin-binding protein and implicate a role of MIM-A in the regulation of cellular proliferation. These data suggest that the reduction of MIM-A gene expression in prostate cancer and other cancers may contribute to tumor growth and development, as well as metastasis.