Cytotoxicity of human recombinant arginase I (Co)-PEG5000 in the presence of supplemental L-citrulline is dependent on decreased argininosuccinate synthetase expression in human cells

Abstract

Human recombinant arginase I cobalt [HuArgI (Co)] coupled with polyethylene glycol 5000 [HuArgI (Co)-PEG5000] has shown potent in-vitro depletion of arginine from tissue culture medium. We now show that HuArgI (Co)-PEG5000 is toxic to almost all cancer cell lines and to some normal primary cells examined. In contrast, HuArgI (Co)-PEG5000 in combination with supplemental L-citrulline is selectively cytotoxic to a fraction of human cancer cell lines in tissue culture, including some melanomas, mesotheliomas, acute myeloid leukemias, hepatocellular carcinomas, pancreas adenocarcinomas, prostate adenocarcinomas, lung adenocarcinomas, osteosarcomas, and small cell lung carcinomas. Unfortunately, a subset of normal human tissues is also sensitive to HuArgI (Co)-PEG5000 with L-citrulline supplementation, including umbilical endothelial cells, bronchial epithelium, neurons, and renal epithelial cells. We further show that cell sensitivity is predicted by the level of cellular argininosuccinate synthetase protein expression measured by immunoblots. By comparing a 3-day and 7-day exposure to HuArgI (Co)-PEG5000 with supplemental L-citrulline, some tumor cells sensitive on short-term assay are resistant in the 7-day assay consistent with the induction of argininosuccinate synthetase expression. On the basis of these results, we hypothesize that HuArgI (Co)-PEG5000 in combination with L-citrulline supplementation may be an attractive therapeutic agent for some argininosuccinate synthetase-deficient tumors. These in-vitro findings stimulate further development of this molecule and may aid in the identification of tissue toxicities and better selection of patients who will potentially respond to this combination therapy.

l-Arginine Depletion Blunts Antitumor T-cell Responses by Inducing Myeloid-Derived Suppressor Cells

“…Previous studies established the therapeutic benefit of peg-Arg I in various pre-clinical malignancies (6-10). Moreover, peg-Arg I is undergoing evaluation in phase I/II trials for adults with liver carcinoma, with favorable safety profiles (34).…”

“…Metabolism of the non-essential amino acid L-Arginine (L-Arg) plays a central role in several biological systems including the modulation of immune responses and tumor growth (5). We and others described the therapeutic benefit of depleting L-Arg in hematological and solid tumors through a pegylated form of the human L-Arg-metabolizing enzyme arginase I (peg-Arg I) (6-10). Pegylation of arginase I increased its stability in vivo , without altering its activity (7).…”

Targeting Arginine-Dependent Cancers with Arginine-Degrading Enzymes: Opportunities and Challenges

“…Lastly, there is increasing interest in the development of novel types of human arginase, in particular a pegylated cobalt-modified (i.e., substituted for native manganese) enzyme (HuArg I [Co]-PEG5K) that has shown good efficacy in depleting arginine [93,94]. Animal toxicity testing indicates that citrulline supplementation is required since arginase, unlike ADI, produces ornithine which cannot be recycled readily to citrulline [95].…”

Nuclear Trapping through Inhibition of Exosomal Export by Indomethacin Increases Cytostatic Efficacy of Doxorubicin and Pixantrone

“…Cell viability was determined using the CellTiter 96 AQueous One Solution Reagent as previously published (18). EC 50 was defined as the concentration of drug causing a 50% inhibition of cell growth compared with untreated control and was determined using the curve-fitting function (sigmoidal dose response, variable slope) of GraphPad Prism version 4.03 for Windows (GraphPad Software, http://www.graphpad.com).…”