Metabolism of chlorambucil by rat liver microsomal glutathione S-transferase.
Clinical efficacy of alkylating anticancer drugs, such as chlorambucil (4-[p-[bis [2-chloroethyl] amino] phenyl]-butanoic acid; CHB), is often limited by the emergence of drug resistant tumor cells. Increased glutathione (gamma-glutamylcysteinylglycine; GSH) conjugation (inactivation) of alkylating anticancer drugs due to overexpression of cytosolic glutathione S-transferase (GST) is believed to be an important mechanism in tumor cell resistance to alkylating agents. However, the potential involvement of microsomal GST in the establishment of acquired drug resistance (ADR) to CHB remains uncertain. In our experiments, a combination of lipid chromatography/electrospray ionization mass spectrometry (LC/ESI/MS) was employed for structural characterization of the resulting conjugates between CHB and GSH. The spontaneous reaction of 1mM CHB with 5 mM GSH at 37 degrees C in aqueous phosphate buffer for 1 h gave primarily the monoglutathionyl derivative, 4-[p-[N-2-chloroethyl, N-2-S-glutathionylethyl] amino]phenyl]-butanoic acid (CHBSG) and the diglutathionyl derivative, 4-[p-[2-S-glutathionylethyl] amino]phenyl]-butanoic acid (CHBSG2) with small amounts of the hydroxy-derivative, 4-[p-[N-2-S-glutathionylethyl, N-2-hydroxyethyl] amino]phenyl]-butanoic acid (CHBSGOH), 4-[p-[bis[2-hydroxyethyl] amino]phenyl]-butanoic acid (CHBOH2), 4-[p-[N-2-chloroethyl, N-2-S-hydroxyethyl]amino]phenyl]-butanoic acid (CHBOH). We demonstrated that rat liver microsomal GST presented a strong catalytic effect on these reactions as determined by the increase of CHBSG2, CHBSGOH and CHBSG and the decrease of CHB. We showed that microsomal GST was activated by CHB in a concentration and time dependent manner. Microsomal GST which was stimulated approximately two-fold with CHB had a stronger catalytic effect. Thus, microsomal GST may play a potential role in the metabolism of CHB in biological membranes, and in the development of ADR.
Zhang
Jie
J
Department of Pharmacology and Toxicology, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, China.
Ye
Zhiwei
Z
Lou
Yijia
Y
D000818:Animals; D018906:Antineoplastic Agents, Alkylating / Q000378:metabolism / Q000494:pharmacology*; D002384:Catalysis / Q000187:drug effects; D002699:Chlorambucil / Q000378:metabolism / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D004305:Dose-Response Relationship, Drug; D019008:Drug Resistance, Neoplasm / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D005982:Glutathione Transferase / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects* / Q000201:enzymology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D021241:Spectrometry, Mass, Electrospray Ionization
D018906:Antineoplastic Agents, Alkylating; D004791:Enzyme Inhibitors; D002699:Chlorambucil; D005982:Glutathione Transferase; D005033:Ethylmaleimide