Relationship between activation of microsomal glutathione S-transferase and metabolism behavior of chlorambucil.
Chlorambucil (4-[p-[bis [2-chloroethyl] amino] phenyl]-butanoic acid; CMB) is a bifunctional alkylating agent that exhibits acquired drug resistance upon repeated dosing in human. This compound reacts with glutathione (gamma-glutamylcysteinylglycine, GSH) both non-enzymatically and enzymatically in the presence of glutathione S-transferases (GSTs) to produce several GSH conjugates. The formation of GSH conjugates of this class of alkylating agents may play a role in the development of acquired drug resistance. In this study, the effect of CMB on microsomal GST (mGST) activity and the catalytic effect of mGST on the formation of GSH conjugates of CMB were investigated. mGST was activated by CMB in a concentration- and time-dependent manner. NEM failed to reactivate the CMB pretreated mGST. This suggests that mGST could be activated by CMB, possibly via reacting with the single cysteine (Cys49) in mGST. The resulting conjugates of GSH and CMB were characterized by a combination of lipid chromatography/electrospray ionization mass spectrometry (LC/ESI/MS). mGST presented a strong catalytic effect on the reaction as determined by the increase of the diglutathionyl derivative, 4-[p-[bis[2-S-glutathionylethyl] amino]phenyl]-butanoic acid (CMBSG2), monoglutathionyl derivatives, 4-[p-[N-2-chloroethyl, N-2-S-glutathionylethyl] amino]phenyl]-butanoic acid (CMBSG) and 4-[p-[N-2-S-glutathionylethyl, N-2-hydroxyethyl] amino]phenyl]-butanoic acid (CMBSGOH) and the decrease of CMB. mGST which was stimulated approximately 2.5-fold with CMB had a stronger catalytic effect. These results suggest the potential role of the activation of mGST in the CMB inactivation and in the development of acquired drug resistance.
Zhang
Jie
J
Department of Pharmacology and Toxicology, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310031, PR China.
Lou
Yi-Jia
YJ
D000818:Animals; D002384:Catalysis / Q000187:drug effects; D002699:Chlorambucil / Q000737:chemistry / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D005978:Glutathione / Q000737:chemistry; D005982:Glutathione Transferase / Q000302:isolation & purification / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D013997:Time Factors
D002699:Chlorambucil; D005982:Glutathione Transferase; D005978:Glutathione