Abstract 2120: Asparaginases-Treatment potential for the biomarker selected hepatocellular carcinoma (HCC) population
Authors: Tyagi Ethika , Pohorecka Magdalena , Duvivier Valérie , Studeny Aurelie , Valour Damien , Moulharat Natacha , Darville Hélène , Provost Nicolas , Hilaire Pierre Barbier Saint , Hoeben Eef , Simons Donald 
Published: March 2024

Abstract
Asparaginase is a key component of the therapeutic regimen used to treat patients with acute lymphoblastic leukemia (ALL). Calaspargase pegol (Asparlas) is composed of E. coli L-asparaginase and is PEGylated with approximately 5000 Da polyethylene glycol (PEG). Asparlas depletes asparagine (Asn) by deamination into aspartic acid. Asn synthetase (ASNS) is the key enzyme responsible for de novo Asn synthesis. In the absence of Asn in their direct environment, normal cells having adequate ASNS activity can survive with their own Asn supply while cancer cells lacking ASNS activity undergo apoptosis as they depend on extracellular sources of Asn. The ASNS gene has a typical CpG island in its promoter and its methylation status in tumor cells has been reported to be associated with asparaginase sensitivity clinically in T-ALL and in solid tumors preclinically. In this study we have investigated ASNS promoter hypermethylation as a biomarker for Asparlas activity in solid tumors and explored the feasibility of translating these results into the clinic. Analyses of the TCGA datasets indicated that a relatively high fraction of hepatocellular carcinoma (HCC) and gastric cancers are hypermethylated at the ASNS promoter. Testing Asparlas across a panel of HCC cell lines demonstrated that ASNS promoter-hypermethylated cell lines with lowest ASNS mRNA expression are the most responsive to Asparlas treatment in vitro. In vivo efficacy studies in mice bearing human ASNS promoter-hypermethylated xenograft models of HCC and gastric cancer showed that Asparlas treatment inhibited tumor growth in a dose-dependent manner and resulted in tumor rejections at the highest doses. Data from the HCC model was used to build a PK/PD efficacy model to project efficacious concentrations in patients. A panel of HCC PDX models with different levels of ASNS promoter methylation was evaluated to better understand the relationship between this biomarker and response to Asparlas. Treatment with Asparlas resulted in complete depletion of Asn in plasma and a partial depletion of Asn in the tumor for all models tested and strong anti-tumor activity against a model that had high methylation of the ASNS promoter. This activity was lost in models with lesser degrees of methylation indicating a strict threshold of promoter methylation for efficacy. These data confirm and extend previous findings that hypermethylation of the ASNS promoter has a synthetic lethal relationship with Asn-depleting therapies including Asparlas and highlight the need to carefully define the minimum threshold of hypermethylation to select HCC patients who could potentially benefit from Asparlas treatment.
Citation Format: Ethika Tyagi, Magdalena Pohorecka, Valérie Duvivier, Aurelie Studeny, Damien Valour, Natacha Moulharat, Hélène Darville, Nicolas Provost, Pierre Barbier Saint Hilaire, Eef Hoeben, Donald Simons. Asparaginases-Treatment potential for the biomarker selected hepatocellular carcinoma (HCC) population [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2120.