Abstract C208: Targeting the actin cytoskeleton of neuroblastoma: Elucidating the mechanism by which a novel class of anti-cancer compounds induces tumor cell death.

Abstract

Abstract The actin cytoskeleton is an ideal chemotherapeutic target due to its role in numerous biological processes essential for tumor cell growth and survival. Targeting actin however has been problematic due to unacceptable levels of toxicity associated with impacting actin containing structures essential for normal cell function. We have developed a novel class of compounds which target tropomyosin, the second core component of an actin microfilament. By targeting the cancer associated tropomyosin, Tm5NM1 we are able to discriminate between the actin filament populations in normal and transformed cells. We have demonstrated that our first in class anti-Tm compound, TR100, impacts actin filament integrity leading to tumor cell death in vitro and in vivo in neuroblastoma and melanoma models. In this study we investigate the molecular mechanisms by which disruption of the actin cytoskeleton with the next generation anti-Tm compounds leads to tumor cell death. Preliminary studies have indicated that these compounds initiate cell death via the intrinsic mitochondrial apoptotic pathway. SH-EP neuroblastoma cells treated with TR200 showed a dose dependent increase in both caspase activation and annexin V staining as measured by Western Blotting and FACS analysis respectively. Significant changes in mitochondrial membrane integrity were also observed using TMRE staining in SH-EP cells treated with 5 µM TR200 for 24h. Interestingly, mitochondrial permeabilization was independent of the presence of proapoptotic factors Bax and Bak as Bax/Bak null MEFs still displayed sensitivity to TR200. To delineate the signaling pathways leading to apoptosis, a kinexus phospho-antibody array was conducted using SH-EP cells treated with 10 µM DTR200 for 8 and 24h. DTR200 treatment for 8h reduced the phospho-activity of key intermediates of the MEK/ERK pathway, in particular a significant decrease in the phosphorylation of the RSK1/2 family of kinases. Preliminary data would suggest that the compounds at 24h are mediating their effect through the downregulation of the MEK/ERK and p38/JNK cell survival pathways. We are now validating these findings to elucidate which signaling pathways mediate the apoptotic response to the anti-Tm compounds. Understanding the molecular mechanisms involved in anti-Tm mediated cell death will allow us to determine how these compounds can be used in combination with existing therapies to lower the apoptotic threshold of a tumor cell and ultimately improve the outcome of patients with neuroblastoma. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C208. Citation Format: Melissa Desouza, Margaret Nguyen, Peter W. Gunning, Justine R. Stehn. Targeting the actin cytoskeleton of neuroblastoma: Elucidating the mechanism by which a novel class of anti-cancer compounds induces tumor cell death. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C208.