Abstract B043: Role of IRS2-microtubule interactions in breast carcinoma cell survival

The biggest obstacle for the eradication of breast cancer is the metastatic spread of tumor cells beyond the primary cancer site. Insulin-like Growth Factor-1 receptor (IGF-1R) signaling has been implicated in mammary tumor progression from localized to metastatic disease. Signal transduction by the IGF-1R requires the Insulin Receptor Substrate (IRS) adaptor proteins IRS1 and IRS2. These scaffold proteins lack intrinsic kinase activity but organize signaling downstream of the receptor. Despite their homology, these proteins have distinct functions, with IRS1 regulating tumor cell proliferation and IRS2 regulating metabolism, survival and invasion. In vivo studies performed by our group demonstrated that Irs2-/- tumors are poorly metastatic, whereas Irs1-/- tumors that express elevated Irs2 are highly metastatic, when compared with wild type tumors. These studies suggest that IRS2 is a positive regulator of mammary tumor metastasis. Importantly, the mechanism(s) responsible for the divergent roles of these homologous proteins in tumor biology remains unknown. Recent studies from our group and others suggest that the subcellular localization of IRS1 and IRS2 may play a role in their distinct functions. Specifically, in human breast tumors, IRS1 is expressed either in the nucleus or cytoplasm and IRS2 is expressed either at the cell membrane or in the cytoplasm. Of note, patients with breast tumors that have IRS2 localized at the cell membrane have decreased overall survival. We hypothesized that trafficking of the IRS proteins to distinct subcellular compartments is essential for their differential functions in breast cancer and that the cytoskeleton would play an important role in this localization. Our preliminary studies using MDA-MB-231 and SUM-159 breast carcinoma cells revealed that IRS2-mediated signaling, but not IRS1-mediated signaling, requires an intact microtubule cytoskeleton. Treatment of breast carcinoma cells with microtubule disrupting agents inhibited IGF1 stimulated activation of PI3K/Akt signaling in cells that predominantly signal through IRS2. In contrast, cells that signal predominantly through IRS1 retained the ability to activate Akt after disruption of the microtubule cytoskeleton. Suppression of IRS2 expression abrogated the effect of microtubule disruption on Akt activation, whereas restoration of Irs2 in Irs2-/- mouse mammary tumor cells sensitized these cells to the effects of microtubule disrupting agents. Likewise, overexpression of IRS1 in MDA-MB-435 breast carcinoma cells that express predominantly IRS2 caused resistance to the effects of microtubule disruption on IGF1-stimulated Akt activation. Given the importance of Akt for tumor cell survival, we assessed the impact of IRS2 signaling on the survival of breast carcinoma cells after microtubule disruption. Cells that signal predominantly through IRS2 had diminished cell survival after forty-eight hours of treatment with Nocodazole and Vinblastine. Moreover, Irs2-/- cells are more resistant to these agents, and restoration of Irs2 expression resulted in a significant increase in cell death. Taken together our data suggest that IRS2 may interact with the microtubule cytoskeleton and that this interaction plays a role in the ability of IRS2 to activate Akt signaling and promote breast carcinoma cell survival. More importantly, our data implicates IRS2 as a biomarker of tumor sensitivity to microtubule disrupting agents. Understanding the mechanism by which IRS2 interacts with microtubules and how this influences its function may facilitate the development of new therapeutic approaches for the effective treatment of breast cancer metastasis. Citation Format: Jose R. Mercado-Matos, Jennifer Clark, Andrew J. Piper, Jenny Janusis, Leslie M. Shaw. Role of IRS2-microtubule interactions in breast carcinoma cell survival. [abstract]. In: Proceedings of the AACR Special Conference on Advances in Breast Cancer Research: Genetics, Biology, and Clinical Applications; Oct 3-6, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2013;11(10 Suppl):Abstract nr B043.