Effect of inactivation of argininosuccinate synthetase on sensitivity of lymphomas to caspase-dependent apoptosis following treatment with arginine deiminase.

Abstract

8093 Background: Arginine is a semi-essential amino acid, critical to the growth of human cancers. Downregulation of the enzyme argininosuccinate synthetase (ASS1), a key rate-limiting enzyme involved in arginine metabolism, results in an intrinsic dependence on extracellular arginine with arginine deprivation offering a potential treatment option for patients with arginine auxotrophic tumors. Several early phase clinical trials of the arginine lowering drug, pegylated arginine deiminase (ADI-PEG20), have demonstrated clinical efficacy in patients with solid tumors. Methods: We assessed ASS1 expression (ASS1 promoter methylation, ASS1 mRNA and protein levels) in normal and malignant lymphoid cell lines and tissues, and tested the effect of ADI-PEG20. Results: The ASS1 promoter is hypermethylated in lymphomas using the Illumina Infinium methylation platform and provided a discriminator between lymphoma (cell lines and primary tumor samples, n=20) and B cell controls (benign lymph nodes and lymphoblastoid cell lines, n=6). A good correlation was observed between ASS1 mRNA and protein levels in tumor (n=12) and normal control cell lines (n=2), with most malignant, but not normal, cell lines showing absence of ASS1 expression. Notably, ASS1 protein was absent in >99% of lymphoma cases (n=300), as assessed by immunohistochemical staining. The ASS1-negative t(14;18) follicular lymphoma cell line, Karpas-422, and the cutaneous T cell lymphoma cell line, MyLa, were sensitive to ADI-PEG 20 with activation of caspases 3 and 7, PARP cleavage and downregulation of mTOR signalling. In contrast, the normal lymphoblastoid cell lines, NcNC and HRC57 and the Ramos Burkitt's lymphoma cell line, expressed constitutive and inducible levels of ASS1 protein, and were resistant to the apoptotic effects of ADI-PEG20. Conclusions: ASS1 is epigenetically inactivated in human lymphomas resulting in a potential dependency on extracellular arginine for cell growth. ASS1-negative lymphomas are sensitive to arginine depletion suggesting that this approach is worth exploring as a potential novel therapeutic approach in the management of patients with lymphoma. Author Disclosure Employment or Leadership Position Consultant or Advisory Role Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Polaris Pharmaceuticals, Inc. Polaris Pharmaceuticals, Inc.