S-METHYL-N,N-DIETHYLTHIOCARBAMATE SULFOXIDE AND S-METHYL-N,N-DIETHYLTHIOCARBAMATE SULFONE, 2 CANDIDATES FOR THE ACTIVE METABOLITE OF DISULFIRAM

The mechanism of action of disulfiram involves inhibition of hepatic a ldehyde dehydrogenase (ALDH). Although disulfiram inhibits ALDH in vit ro, it is believed that the drug is too short-lived in vivo to inhibit the enzyme directly. The ultimate inhibitor is thought to be a metabo lite of disulfiram. In this study, we examined the effects of S-methyl -N,N-diethylthiocarbamate (MeDTC) sulfoxide and S-methyl-N,N-diethylth iocarbamate sulfone (confirmed and proposed metabolites of disulfiram, respectively) on rat liver mitochondrial low K-m ALDH. MeDTC sulfoxid e and MeDTC sulfone, in 10-min incubations with detergent-solubilized mitochondria, inhibited ALDH activity with an IC50 (mean +/- SD) of 0. 93 +/- 0.04 and 0.53 +/- 0.11 mu M respectively, compared with 7.4 +/- 1.0 mu M for the parent drug disulfiram. Inhibition by MeDTC sulfone and MeDTC sulfoxide, both at 0.6 mu M, was time-dependent, following a pparent pseudo-first-order kinetics with a t(1/2) of inactivation of 3 .5 and 8.8 min, respectively. Dilution of ALDH inhibited by either sul foxide or sulfone did not restore activity, an indication of irreversi ble inhibition. Addition of glutathione (50 to 1000 mu M) to ALDH befo re the inhibitors did not alter the inhibition by MeDTC sulfoxide. In contrast, the inhibition by MeDTC sulfone was decreased >10-fold (IC50 = 6.3 mu M) by 50 mu M of glutathione and almost completely abolished by 500 mu M of glutathione. The cofactor NAD, in a concentration-depe ndent manner, protected ALDH from inhibition by MeDTC sulfoxide and Me DTC sulfone. In incubations with intact mitochondria, the potency of t he two compounds was reversed (IC50 of 9.2 +/- 3.6 and 0.95 +/- 0.30 m u M for the MeDTC sulfone and sulfoxide, respectively). Our results su ggest that MeDTC sulfone is highly reactive with normal cellular const ituents (e.g., glutathione), which may protect ALDH from inhibition, u nless this inhibitor is formed very near the target enzyme. In contras t, MeDTC sulfoxide is a better candidate for the ultimate active metab olite of disulfiram, because it is more likely to be sufficiently stab le to diffuse from a distant site of formation, such as the endoplasmi c reticulum, penetrate the mitochondria, and react with ALDH located i n the mitochondrial matrix.