Data from Discovery and Targeting of a Noncanonical Mechanism of Sarcoma Resistance to ADI-PEG20 Mediated by the Microenvironment

Abstract

<div>AbstractPurpose:<p>Many cancers lack argininosuccinate synthetase 1 (ASS1), the rate-limiting enzyme of arginine biosynthesis. This deficiency causes arginine auxotrophy, targetable by extracellular arginine-degrading enzymes such as ADI-PEG20. Long-term tumor resistance has thus far been attributed solely to ASS1 reexpression. This study examines the role of ASS1 silencing on tumor growth and initiation and identifies a noncanonical mechanism of resistance, aiming to improve clinical responses to ADI-PEG20.</p>Experimental Design:<p>Tumor initiation and growth rates were measured for a spontaneous <i>Ass1</i> knockout (KO) murine sarcoma model. Tumor cell lines were generated, and resistance to arginine deprivation therapy was studied <i>in vitro</i> and <i>in vivo</i>.</p>Results:<p>Conditional <i>Ass1</i> KO affected neither tumor initiation nor growth rates in a sarcoma model, contradicting the prevalent idea that ASS1 silencing confers a proliferative advantage. <i>Ass1</i> KO cells grew robustly through arginine starvation <i>in vivo</i>, while ADI-PEG20 remained completely lethal <i>in vitro</i>, evidence that pointed toward a novel mechanism of resistance mediated by the microenvironment. Coculture with <i>Ass1</i>-competent fibroblasts rescued growth through macropinocytosis of vesicles and/or cell fragments, followed by recycling of protein-bound arginine through autophagy/lysosomal degradation. Inhibition of either macropinocytosis or autophagy/lysosomal degradation abrogated this growth support effect <i>in vitro</i> and <i>in vivo</i>.</p>Conclusions:<p>Noncanonical, ASS1-independent tumor resistance to ADI-PEG20 is driven by the microenvironment. This mechanism can be targeted by either the macropinocytosis inhibitor imipramine or the autophagy inhibitor chloroquine. These safe, widely available drugs should be added to current clinical trials to overcome microenvironmental arginine support of tumors and improve patient outcomes.</p></div>