Abstract

Abstract The enzyme-drug L-asparaginase (L-ASP) has been used for four decades to treat acute lymphoblastic leukemia. However, its unique mechanism of action is still poorly understood, and its clinical efficacy has proven unpredictable. Those problems have prompted a continuing search for biomarkers that predict L-ASP response. We previously found that the expression of asparagine synthetase (ASNS) is strongly correlated with L-ASP activity in leukemia cell lines of the NCI-60 cell line panel. Unexpectedly, however, the correlation was also apparent in ovarian cancer lines, suggesting that L-ASP might be effective against a low-ASNS subset of ovarian cancers if salient characteristics of the cell lines reflect clinical ovarian tumors. Using siRNA to down-regulate ASNS by 5-fold produced a proportional ∼5-fold potentiation of L-ASP activity in some ovarian cancer lines (OVCAR-3 and OVCAR-4). Surprisingly, ASNS siRNAs yielded a striking >500-fold potentiation in other ovarian lines (OVCAR-8 and OVCAR-8/ADR). Taken together, those findings suggest that the L-ASP/ASNS correlation is causal but also support the hypothesis that L-ASP activity is modulated by additional molecules or pathways. To address that hypothesis and to further elucidate the L-ASP mechanism of action, we have initiated genome-wide siRNA library screening. Preliminary screens in the OVCAR-8 cell line using a 418-gene apoptosis library, a custom 392-gene metabolic library, and the MTS assay as an endpoint yielded Z′ factors >0.90, indicating a robust screen. RNAi-mediated down-regulation of apoptosis genes had little or no effect on L-ASP's anticancer activity. That was surprising because L-ASP has been reported to cause apoptosis through glutamine depletion. Knock-down of ATF4, a transcription factor involved in cell metabolism, did markedly potentiate L-ASP activity (>500-fold). Furthermore, ATF4 knock-down markedly enhanced L-ASP anticancer activity toward the highly resistant cell line, OVCAR-4. These findings suggest that ATF4 is an important mediator of L-ASP resistance and provide rationale for investigation of ATF4 as a predictive biomarker of L-ASP effect. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1683.

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