The Relationship of Arginine Deprivation, Argininosuccinate Synthetase and Cell Death in Melanoma

Abstract

It has been shown that melanoma cells do not express argininosuccinate synthetase (ASS) and therefore are unable to synthesize arginine from citrulline. Depleting arginine using pegylated arginine deiminase (ADI-PEG20) results in cell death in melanoma but not normal cells. This concept was translated into clinical trial and responses were seen. However, induction of ASS expression does occur which results in resistance to ADI-PEG20. We have used 4 melanoma cell lines to study factors which may govern ASS expression. Although these 4 melanoma cell lines do not express ASS protein or mRNA as detected by both immunoblot and northernblot analysis, ASS protein can be induced after these cells are grown in the presence of ADI-PEG20, but again repressed after replenishing arginine in the media. The levels of induction are different and one cell line could not be induced. Interestingly, a melanoma cell line with the highest level of induction could also be made resistant to ADI-PEG20. This resistant line possesses high levels of ASS mRNA and protein expression which cannot be repressed with arginine. Our study indicates that ASS expression in melanoma cells is complex and governed by biochemical parameters which are different among melanoma cells.

Arginine Deprivation as a Targeted Therapy for Cancer

“…In our laboratory, we have studied the growth inhibitory effect of ADI-PEG20 in 4 melanoma cell lines, (A-375, SK-Mel2, A-2058 and Mel-1220 all ASS (−) one normal human fibroblast (BJ-1) and one ASS (+) NSCLC line. The growth inhibitory effect ranged from 0.05-0.09 μg/ml after 3 days exposure in 4 melanoma cell lines tested (see Table 1) and greater than 1 in BJ-1 and NSCLCS cell lines [18]. Interestingly, at 72 hr.…”

“…Furthermore, this enzyme is active in physiologic pH and has a high affinity for arginine. In fact, ADI has been reported to have antitumor activity both in vitro and in vivo in melanoma and hepatocellular carcinoma cell lines and in other cell lines [15][16][17][18][31][32][33][34]. However, one of the drawbacks of ADI is that it is not produced by humans and, consequently, it is highly antigenic.…”

“…We and others have shown that degrading arginine in the culture media using ADI or exposure of melanoma cell lines to arginine free media with citrulline and NH4Cl supplement can inhibit growth as well as have cytotoxic effects in melanoma cell lines [15,18]. The IC50 after 7 days of exposure ranged from 0.01-0.3 μg/ml among 16 melanoma cell lines tested [15].…”

“…Thus, depletion of this amino acid by asparaginase therapy is generally well tolerated since normal human cells do not require asparagine. Thus, eliminating a certain Several laboratories have reported that certain tumor cell lines such as melanoma, hepatocellular carcinoma, renal cell carcinoma and some mesotheliomas do not express ASS, and hence become auxotrophic for arginine [9,11,[15][16][17][18]. Consequently, arginine degrading enzyme has been shown by several laboratories to have antitumor effect in these tumors [9,13,[19][20][21][22][23][24][25].…”

“…To further prove that the lack of ASS expression in melanoma cell lines is a key to render them susceptible to cell death upon arginine deprivation, we and others have transfected melanoma cells with ASS. These transfectants are resistant to arginine deprivation [15,18].…”