Abstract 1171: LIMK1 functions as a modulator of expression and targeting of CXCR4 in prostate cancer cells through a negative feedback loop

Abstract

Abstract Increased activation of the GPCR CXCR4/CXCL12 signaling axis is commonly noted in metastatic prostate cancer and shows an association with poor prognosis. CXCR4, and its ligand, CXCL12/SDF-1a, are recognized as powerful mediators for tumor microenvironment remodeling and chemoprotection. LIM kinase 1 (LIMK1), a serine/threonine kinase, is an actin and microtubule modulatory protein that is also overexpressed in a variety of cancers including prostate cancer. Earlier, we showed that LIMK1 is involved in cell invasion and inhibition of LIMK1 expression reverted the invasive phenotype of PC3 prostate cancer cells. Activation of CXCR4/CXCL12 axis promotes activation of LIMK1 through RhoGTPAse pathway and promotes docetaxel resistance. Here we show that LIMK1 expression and functions have a positive correlation with CXCR4 expression and subcellular localization. We used metastatic prostate cancer cell lines PC3 and M12 and the non-tumorigenic prostate cell lines BPH and P69 to monitor expression and membrane localization of CXCR4 using western blots, flow cytometry and immunofluorescence analysis. Our analysis showed that cells with higher expression of LIMK1 (PC3) overexpress CXCR4, and that cell lines with low levels of LIMK1 (BPH and P69) express relatively lower amounts of CXCR4. Flow cytometric analysis showed a stronger surface staining of CXCR4 in the LIMK1 overexpressing PC3 cells compared to BPH cells. Next, we studied the effect of shRNA-mediated inhibition of LIMK1 expression on CXCR4 expression in PC3 or M12 cells. Western blot and immunofluorescence analysis showed noticeable reduction in CXCR4 levels, and its surface localization, as noted by flow cytometry, in LIMK1 shRNA transfected PC3 and M12 cells. To assess if the kinase activity of LIMK1 is essential for CXCR4 expression and surface localization, we treated these cells with LIMK1 specific kinase inhibitor BMS-5 and determined expression of CXCR4. Western blots and immunofluorescence analysis showed reduced expression of CXCR4 in treated cells compared to vehicle treated cells. Our observation indicates an indirect association between LIMK1 and CXCR4 expression and shows for the first time that expression of LIMK1 is required for the maintenance of CXCR4 expression. Our study demonstrates a novel positive feed back regulatory mechanism between LIMK1 and CXCR4 and suggest a possible combination treatment option through inhibition of LIMK1 expression and/or its kinase activity for improving chemosensitivity of prostate cancer cells with overactive CXCR4/CXCL12 axis. Citation Format: Richard Ottman, Lisa Ritchey, Ryan Herchan, Alexia Bossan, Ratna Chakrabarti. LIMK1 functions as a modulator of expression and targeting of CXCR4 in prostate cancer cells through a negative feedback loop. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 1171.