Asparagine synthetase promoter hypermethylation is required, but not sufficient, for sensitivity to Asparlas in patient-derived models of hepatocellular carcinoma.

Therapies that deplete plasma asparagine, such as Asparlas, are a key component of the standard-of-care for acute lymphoblastic leukemia (ALL). The basis of this therapy is that some cancer types lack the ability to produce endogenous asparagine via the enzyme asparagine synthetase (ASNS) and are thus entirely dependent on exogenous asparagine for survival. The methylation status of the ASNS gene promoter in tumor cells has been associated with asparaginase sensitivity clinically in T-ALL, and in solid tumors preclinically. We investigated whether ASNS promoter methylation (pASNSmet) can serve as a biomarker for Asparlas efficacy in solid tumors with a focus on hepatocellular carcinoma (HCC) based on a relatively high frequency of pASNSmet in this indication in the TCGA. Asparlas treatment of a panel of HCC cell lines demonstrated that the highest pASNSmet lines were the most sensitive to asparagine depletion in vitro and in vivo. However, when a panel of patient derived xenograft models representing a range of pASNSmet levels was tested we found that high pASNSmet levels were required but not necessarily sufficient to confer sensitivity to treatment with Asparlas. Collectively, these results are in support of ASNS promoter hypermethylation as a targetable phenotype but indicate that pASNSmet alone is not a viable biomarker of this phenotype, and that significant advancements in our understanding of the underlying biology may be required before asparaginase-depleting therapy can be successfully implemented clinically in HCC.