Therapeutic Assessment of Targeting ASNS Combined with <scp>l</scp> -Asparaginase Treatment in Solid Tumors and Investigation of Resistance Mechanisms

Abstract

Asparagine deprivation by l-asparaginase (L-ASNase) is an effective therapeutic strategy in acute lymphoblastic leukemia, with resistance occurring due to upregulation of ASNS, the only human enzyme synthetizing asparagine (<i>Annu. Rev. Biochem.</i> <b>2006</b>, <i>75</i> (1), 629-654). l-Asparaginase efficacy in solid tumors is limited by dose-related toxicities (<i>OncoTargets and Therapy</i> 2017, pp 1413-1422). Large-scale loss of function genetic <i>in vitro</i> screens identified ASNS as a cancer dependency in several solid malignancies (<i>Cell</i> <b>2017</b>, <i>170</i> (3), 564-576.e16. <i>Cell</i> <b>2017</b>, <i>170</i> (3), 577-592.e10). Here we evaluate the therapeutic potential of targeting ASNS in melanoma cells. While we confirm <i>in vitro</i> dependency on ASNS silencing, this is largely dispensable for <i>in vivo</i> tumor growth, even in the face of asparagine deprivation, prompting us to characterize such a resistance mechanism to devise novel therapeutic strategies. Using <i>ex vivo</i> quantitative proteome and transcriptome profiling, we characterize the compensatory mechanism elicited by ASNS knockout melanoma cells allowing their survival. Mechanistically, a genome-wide CRISPR screen revealed that such a resistance mechanism is elicited by a dual axis: GCN2-ATF4 aimed at restoring amino acid levels and MAPK-BCLXL to promote survival. Importantly, pharmacological inhibition of such nodes synergizes with l-asparaginase-mediated asparagine deprivation in ASNS deficient cells suggesting novel potential therapeutic combinations in melanoma.