Abstract 1502: Microtubule stabilization alters tumor secretome and fibroblast activation

Abstract

Abstract Microtubules (MTs) represent one of the most effective targets in cancer chemotherapy. However, drugs that target MTs, like the taxanes, often fail in the metastatic setting. Therefore it is of utmost importance to fully understand the roles of MTs and MT-targeting agents in tumor biology. An important process for tumor development and metastasis involves the remodeling of the 3D microenvironment surrounding tumor cells. In order to successfully proliferate and metastasize, tumor cells need to dynamically respond to, manipulate, and remodel their surrounding microenvironment. We hypothesized that MTs, as a dynamic cytoskeletal component, regulates the cellular signaling process that controls tumor-mediated remodeling of the microenvironment. To test this hypothesis, we assessed the MT-dependence of metastatic MDA-MB-231 (231) breast cancer cell activation of HMF3S human mammary fibroblasts. We collected media conditioned by vehicle- or paclitaxel (PTX)-pretreated 231 cells and then applied this conditioned media to HMF3S fibroblasts. Application of 231-derived conditioned media activated HMF3S fibroblasts as evidenced by their enhanced directional 3D motility towards a serum gradient. However, application of conditioned media from PTX-pretreated 231 cells to HMF3S fibroblasts inhibited their 3D directional motility. Importantly, we demonstrated that this effect was due to PTX-induced MT stabilization as conditioned media from PTX-treated 231 cells resistant to PTX did not inhibit fibroblasts’ 3D motility. Together, these results suggest that the composition of the tumor secretome changes upon MT stabilization by PTX, which in turn affects the ability of tumor cells to communicate with fibroblasts. In order to identify the factors responsible for this MT-dependent cell-cell communication we performed a differential proteomic analysis of the conditioned media from vehicle- and PTX-pretreated 231 cells. Using splitless nanoflow chromatography coupled with quadrupole time-of-flight mass spec, we identified a total of 2124 proteins within these tumor secretomes. Using a 2-fold change threshold, we identified 78 under-represented and 31 over-represented proteins in the secretome of PTX- versus vehicle-pretreated cells. We focused our analysis on under-represented proteins based on the hypothesis that MT stabilization would inhibit their secretion. From this analysis we have identified proteins involved in cell motility, cell-cell communication, or ECM remodeling. TGFβ, CTGF, cMet, fibronectin, and lysyl oxidase 2 were key down-regulated proteins identified in the tumor secretome upon PTX-induced MT stabilization. Currently we are investigating the mechanisms by which stabilization of the MT cytoskeleton results in the down-regulation of these factors. These findings point to a novel role of interphase MTs in tumor biology and can possibly enhance our understanding of taxane clinical efficacy. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1502. doi:1538-7445.AM2012-1502