Characterization of inhibitors of specific carboxylesterases: development of carboxylesterase inhibitors for translational application.
Carboxylesterases, expressed at high levels in human liver and intestine, are thought to detoxify xenobiotics. The anticancer prodrug 7-ethyl-10-[4-1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11) is also metabolized by carboxylesterases to produce the active drug 7-ethyl-10-hydroxycamptothecin. Activation of CPT-11 by human intestinal carboxylesterase (hiCE) in the human intestine may contribute to delayed onset diarrhea, a dose-limiting side effect of this drug. The goal of this study was to develop small molecule inhibitors selective for hiCE to circumvent or treat the toxic side effects of CPT-11. A secondary goal was to develop molecules that specifically inhibit activation of CPT-11 by a rabbit liver carboxylesterase (rCE). rCE is the most efficient CPT-11-activating enzyme thus far identified, and this enzyme is being developed for viral-directed enzyme prodrug therapy applications. Based on in vitro assays with partially purified hiCE and rCE proteins and on growth inhibition assays using U373MG human glioma cells transfected to express hiCE or rCE (U373pIREShiCE or U373pIRESrCE), we identified specific inhibitors of each enzyme. Lead compounds are derivatives of nitrophenol having 4-(furan-2-carbonyl)-piperazine-1-carboxylic acid or 4-[(4-chlorophenyl)-phenylmethyl]-piperazine-1-carboxylic acid substitutions in the p position. Kinetic analysis of each compound for hiCE compared with rCE showed that the Ki values of the most selective of these inhibitors differed by 6- to 10-fold. In growth inhibition assays, nontoxic, low micromolar concentrations of these inhibitors increased the EC50 of CPT-11 for U373pIREShiCE or U373pIRESrCE cells by 13- to >1,500-fold. The four compounds characterized in this study will serve as lead compounds for a series of inhibitors to be constructed using a combinatorial approach.
Department of Molecular Pharmacology, St. Jude Children's Research Hospital, 332 North Lauderdale, Memphis, TN 3810, USA.
Kyoung Jin P
KJ
Yoon
Janice L
JL
Hyatt
Christopher L
CL
Morton
Richard E
RE
Lee
Philip M
PM
Potter
Mary K
MK
Danks
D000818:Animals; D002166:Camptothecin / Q000031:analogs & derivatives* / Q000494:pharmacology*; D043182:Carboxylesterase / Q000737:chemistry; D002265:Carboxylic Ester Hydrolases / Q000037:antagonists & inhibitors* / Q000737:chemistry; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism; D000077146:Irinotecan; D007700:Kinetics; D008099:Liver / Q000201:enzymology; D008956:Models, Chemical; D009596:Nitrophenols / Q000737:chemistry; D014176:Protein Biosynthesis; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry; D014162:Transfection
D004791:Enzyme Inhibitors; D009596:Nitrophenols; D011994:Recombinant Proteins; D000077146:Irinotecan; C028098:10-hydroxycamptothecin; D002265:Carboxylic Ester Hydrolases; D043182:Carboxylesterase; D002166:Camptothecin