Analysis of arginine metabolism in acute myeloid leukaemia to provide insights into tumour immune escape strategies

Abstract

BackgroundAminoacid catabolism has emerged as an important immune escape mechanism in cancer. Arginase, the enzyme that breaks down arginine, is exploited by cancer cells to dampen tumour-specific immune responses. We have shown that arginase-2 is released in plasma of patients with acute myeloid leukaemia to suppress T-cell growth. Here we aimed to elucidate the adaptive mechanism through which cancer cells resist arginine depletion in the tumour microenvironment. MethodsWe used colorimetric assays to measure arginine concentrations and arginase activity in plasma of patients and mice with acute myeloid leukaemia. T-cell proliferation and activation were assessed with flow cytometry. Differentially expressed genes in THP1 leukaemic cells compared with T cells upon in-vitro arginine depletion were identified in a microarray (Illumina, San Diego, CA, USA) and confirmed by real-time PCR. RNA sequencing (RNA-Seq, Illumina) of human acute myeloid leukaemia cells versus healthy myeloid cells was done to verify results ex vivo. Functional effects were assessed by western blot and proliferation assays. FindingsArginase-2 was released in plasma of patients and mice with acute myeloid leukaemia to cause systemic arginine depletion. Arginine deprivation inhibited T-cell proliferation and function, effects abolished with arginase-specific inhibitors. In contrast to T cells, cancer cells continued to proliferate by converting citrulline to arginine. This dichotomy relies on the ability of cancer cells to upregulate ASS1, the enzyme that controls citrulline conversion to arginine. Knockdown of ASS1 abrogated the ability of cancer cells to proliferate in arginine-free medium. Knockdown of transcription factor ATF4 blocked ASS1 upregulation and citrulline recycling. RNA-sequencing analysis of acute myeloid leukaemia blasts showed ASS1 and ATF4 upregulation in patients compared with healthy controls. InterpretationThese results identify arginine metabolism as an important pathway for acute myeloid leukaemia to suppress T-cell proliferation. Furthermore, they prove that cancer cells, but not T cells, continue to proliferate in arginine-deprived environments by upregulating ASS1, a target of transcription factor ATF4. Our findings lead to potential therapeutic opportunities where arginase-2, ASS1, and ATF4 can be targeted as part of cancer immunotherapy. FundingWellcome Trust clinical research training fellowship (awarded to AVH).