Bioengineered Human Arginase I with Enhanced Activity and Stability Controls Hepatocellular and Pancreatic Carcinoma Xenografts

Abstract

Hepatocellular carcinoma (HCC) and pancreatic carcinoma (PC) cells often have inherent urea cycle defects rendering them auxotrophic for the amino acid l-arginine (l-arg). Most HCC and PC require extracellular sources of l-arg and undergo cell cycle arrest and apoptosis when l-arg is restricted. Systemic, enzyme-mediated depletion of l-arg has been investigated in mouse models and human trials. Non-human enzymes elicit neutralizing antibodies, whereas human arginases display poor pharmacological properties in serum. Co(2+) substitution of the Mn(2+) metal cofactor in human arginase I (Co-hArgI) was shown to confer more than 10-fold higher catalytic activity (k(cat)/K(m)) and 5-fold greater stability. We hypothesized that the Co-hArgI enzyme would decrease tumor burden by systemic elimination of l-arg in a murine model. Co-hArgI was conjugated to 5-kDa PEG (Co-hArgI-PEG) to enhance circulation persistence. It was used as monotherapy for HCC and PC in vitro and in vivo murine xenografts. The mechanism of cell death was also investigated. Weekly treatment of 8 mg/kg Co-hArgI-PEG effectively controlled human HepG2 (HCC) and Panc-1 (PC) tumor xenografts (P = .001 and P = .03, respectively). Both cell lines underwent apoptosis in vitro with significant increased expression of activated caspase-3 (P < .001). Furthermore, there was evidence of autophagy in vitro and in vivo. We have demonstrated that Co-hArgI-PEG is effective at controlling two types of l-arg-dependent carcinomas. Being a nonessential amino acid, arginine deprivation therapy through Co-hArgI-PEG holds promise as a new therapy in the treatment of HCC and PC.