Mouse liver and kidney carboxylesterase (M-LK) rapidly hydrolyzes antitumor prodrug irinotecan and the N-terminal three quarter sequence determines substrate selectivity.
Antitumor prodrug irinotecan is used for a variety of malignancies such as colorectal cancer. It is hydrolyzed to the metabolite, 7-ethyl-10-hydroxycamptothecin (SN-38), which exerts its antineoplastic effect. Several human and rodent carboxylesterases are shown to hydrolyze irinotecan, but the overall activity varies from enzyme to enzyme. This report describes a novel mouse liver and kidney carboxylesterase (M-LK) that is highly active toward this prodrug. Northern analyses demonstrated that M-LK was abundantly expressed in the liver and kidney and slightly in the intestine and lung. Lysates from M-LK transfected cells exhibited a markedly higher activity on irinotecan hydrolysis than lysates from the cells transfected with mouse triacylglycerol hydrolase (TGH) (6.9 versus 1.3 pmol/mg/min). Based on the immunostaining intensity with purified rat hydrolase A, M-LK had a specific activity of 173 pmol/mg/min, which ranked it as one of the most efficient esterases known to hydrolyze irinotecan. A chimeric carboxylesterase and its wild-type enzyme (e.g., M-LKn and M-LK), sharing three quarters of the entire sequence from the N-terminus, exhibited the same substrate preference toward irinotecan and two other substrates, suggesting that the N-terminal sequence determines substrate selectivity. M-LK transfected cells manifested more severe cytotoxicity than TGH transfected cells upon being exposed to irinotecan. Topoisomerase I inhibitors such as irinotecan represent a promising class of anticancer drugs. Identification of M-LK as an efficient carboxylesterase to activate irinotecan provides additional sequence information to locate residues involved in irinotecan hydrolysis and thus facilitates the design of new analogs.
Xie
Mingxing
M
Department of Biomedical Sciences, University of Rhode Island, Kingston, Rhode Island 02881, USA.
Yang
Dongfang
D
Wu
Micheal
M
Xue
Bob
B
Yan
Bingfang
B
D000595:Amino Acid Sequence / Q000502:physiology; D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000378:metabolism* / Q000493:pharmacokinetics; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000493:pharmacokinetics; D043182:Carboxylesterase; D002265:Carboxylic Ester Hydrolases / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis / Q000187:drug effects; D000077146:Irinotecan; D007668:Kidney / Q000201:enzymology*; D008049:Lipase / Q000378:metabolism / Q000493:pharmacokinetics; D008099:Liver / Q000201:enzymology*; D051379:Mice; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism* / Q000502:physiology; D011355:Prodrugs / Q000378:metabolism* / Q000493:pharmacokinetics; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity / Q000187:drug effects / Q000502:physiology
D000972:Antineoplastic Agents, Phytogenic; D010446:Peptide Fragments; D011355:Prodrugs; D000077146:Irinotecan; D002265:Carboxylic Ester Hydrolases; D043182:Carboxylesterase; D008049:Lipase; D002166:Camptothecin