Halala Saed

Thioridazine inhibits autophagy and sensitizes glioblastoma cells to temozolomide: Thioridazine blocks autophagy in glioblastoma cells

Glioblastoma multiforme (GBM) has a poor prognosis with an overall survival of 14–15 months after surgery, radiation and chemotherapy using temozolomide (TMZ). A major problem is that the tumors acquire resistance to therapy. In an effort to improve the therapeutic efficacy of TMZ, we performed a genome‐wide RNA interference (RNAi) synthetic lethality screen to establish a functional gene signature for TMZ sensitivity in human GBM cells. We then queried the Connectivity Map database to search for drugs that would induce corresponding changes in gene expression. By this approach we identified several potential pharmacological sensitizers to TMZ, where the most potent drug was the established antipsychotic agent Thioridazine, which significantly improved TMZ sensitivity while not demonstrating any significant toxicity alone. Mechanistically, we show that the specific chemosensitizing effect of Thioridazine is mediated by impairing autophagy, thereby preventing adaptive metabolic alterations associated with TMZ resistance. Moreover, we demonstrate that Thioridazine inhibits late‐stage autophagy by impairing fusion between autophagosomes and lysosomes. Finally, Thioridazine in combination with TMZ significantly inhibits brain tumor growth in vivo, demonstrating the potential clinical benefits of compounds targeting the autophagy‐lysosome pathway. Our study emphasizes the feasibility of exploiting drug repurposing for the design of novel therapeutic strategies for GBM.

Abstract LB-75: Oct7 is expressed in human gliomas and correlates with malignancy grade

INTRODUCTION: The transcription factor Oct 7 (also called POU3f2/Brn2) is expressed during neurogenesis, and constitutes the CNS equivalent of Oct 4, a critical regulator of induced pluripotent stem cells. Moreover, data suggest its expression is regulated by hypoxia, and Oct 7 expression has been reported in malignant melanoma of the skin. Since both melanomas and CNS-malignancies arise in organs of neuroectodermal origin, we investigated whether human gliomas expressed Oct7. MATERIAL & METHODS: We performed immunohistochemistry of 150 grade II-IV gliomas from our tumor bank, and subsequently performed western blots of 20 glioma samples. In addition, we performed flow cytometry analysis of Oct 7 expression in 5 acutely dissociated tumors. Using lentiviral transfection we established Oct7 overexpression in a panel of constitutively negative glioma cell lines, as well as knock-down in an Oct7 positive glioma cell line, to investigate the effect of Oct 7 expression on proliferation, migration and differentiation. RESULTS: Immunohistochemistry showed that Oct7 was almost uniformly expressed in human gliomas, although at a varying degree. Microscopy revealed a predominantly nuclear staining pattern, but cytoplasmic immunopositivity for Oct7 could also be detected in some tumors. Both western blot and flow cytometry confirmed expression of Oct7 in human gliomas. With two independent observers we obtained a nuclear staining index for all tumors, with 61% positive nuclei in GBM specimens, which was significantly higher than for grade II (37%) and grade III (36%) tumors (p= 0.0001). Moreover, ongoing studies suggest that overexpression of Oct7 increases the proportion of cells in G2/M phase of the cell cycle, suggesting that this transcription factor has a role in regulating tumor cell proliferation, and hence possibly overall tumor aggressiveness. Ongoing studies aim at further elucidating the multiple roles of Oct7 in brain tumor progression. Citation Format: Mohummad Aminur Rahman, Lina Leiss, Halala S. Saed, Christiane H. Gjerde, Mohammad S. Lellahi, Ercan Mutlu, Per Oyvind Enger. Oct7 is expressed in human gliomas and correlates with malignancy grade. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr LB-75. doi:10.1158/1538-7445.AM2014-LB-75