Chemical modification by polyethylene glycol of the anti-tumor enzyme arginine deiminase from Mycoplasma arginini.
Amino acid-degrading enzymes are known to inhibit the growth of tumor cells in culture by depleting amino acids in the medium. Here we demonstrate that arginine deiminase (EC 3.5.3.6) from Mycoplasma arginini had stronger growth-inhibitory activity against all 4 kinds of tumor cell lines tested than L-asparaginase and arginase, which are well-known anti-tumor enzymes. Next, chemical modification of the arginine deiminase molecule with polyethylene glycol was shown to enhance its potency as an anti-tumor enzyme. The percentage of modified amino groups per molecule was estimated to be 51% of the total amino groups, and the average molecular weight was estimated to be about 400,000 by gel-filtration HPLC. The enzymic activity of the modified enzyme was 25.5 units/mg protein, which was equivalent to 57% of that of the native enzyme. The modified enzyme strongly inhibited growth of a mouse hepatoma cell line, MH134, at a concentration of more than 10 ng/ml, showing almost the same dose-response curve as the native enzyme. When a bolus of 5 units of the modified enzyme was intravenously injected into male BDF1 mice, L-arginine in the blood completely disappeared within 5 min, and remained undetectable for more than 8 days. On the other hand, in the case of bolus injection of the same number of units of native enzyme, the plasma L-arginine level recovered up to 66% of the control level at 8 days. These results suggest that this modified enzyme has a longer plasma clearance time and may be more effective as a new anti-tumor agent than the native enzyme.
Takaku
H
H
Pharmaceuticals and Biotechnology Laboratory, Nikko Kyodo Co., Ltd., Saitama.
Misawa
S
S
Hayashi
H
H
Miyazaki
K
K
D000818:Animals; D001119:Arginase / Q000494:pharmacology; D001120:Arginine / Q000097:blood; D001215:Asparaginase / Q000494:pharmacology; D002956:Citrulline / Q000097:blood; D006131:Growth Inhibitors / Q000737:chemistry; D006867:Hydrolases / Q000737:chemistry*; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D009174:Mycoplasma / Q000201:enzymology; D011092:Polyethylene Glycols / Q000737:chemistry*; D014407:Tumor Cells, Cultured
D006131:Growth Inhibitors; D002956:Citrulline; D011092:Polyethylene Glycols; D001120:Arginine; D006867:Hydrolases; D001215:Asparaginase; D001119:Arginase; C019723:arginine deiminase