155514-79-7Relevant academic research and scientific papers
Disulfiram generates a stable N,N-diethylcarbamoyl adduct on Cys-125 of rat hemoglobin β-chains in vivo
Erve, John C. L.,Jensen, Ole N.,Valentine, Holly S.,Amarnath, Venkataraman,Valentine, William M.
, p. 237 - 244 (2000)
Disulfiram (DSF) is a drug used in aversion therapy to treat alcoholics and acts by inhibiting mitochondrial low-K(m) aldehyde dehydrogenase. Investigations into the mechanisms for in vivo inactivation suggest that the DSF metabolite S-methyl-N,N-diethylthiocarbamate sulfoxide reacts irreversibly with an active site Cys. This work aimed to determine if DSF generates monothiocarbamate adducts on cysteine residues in vivo by examining hemoglobin. Sprague-Dawley rats were treated with DSF po for 2, 4, and 6 weeks. Rats have four different globin β-chains, of which three (β-1-3) contain two cysteine residues each. MALDI-TOF MS analysis of two new globin species from DSF-treated rats collected by HPLC revealed increments of 99 Da above the mass of the unmodified chains (β-2 and β-3). In a separate experiment, the globin mixture was digested for 2 h with Glu-C and reanalyzed by MALDI-TOF MS. Results showed a peptide at m/z 2716.3 having a mass 99 Da higher than a known Cys-containing peptide. Subsequently, the Glu-C digest was analyzed using Q-TOF tandem MS, enabling observation of the +4 charge state of the peptide with m/z 2716.3. This peptide was fragmented to produce y-sequence ions that located the modification to Cys-125 (present on both β- 2 and β-3). Cys-125 is the most reactive of two cysteine residues on these β-chains. To confirm the structure of the modification, globin was hydrolyzed with 6 N HCl at 110 °C for 18 h. The adduct survived these conditions so that S-(N,N-diethylcarbamoyl)cysteine was detected in the hydrolysates of treated rats on the basis of comparison with the tandem MS spectrum of a standard. These results extend the findings of others obtained using glutathione conjugates and demonstrate the ability of DSF to covalently modify Cys residues of proteins in a manner consistent with the production of S-methyl-N,N-diethylthiocarbamate sulfoxide, or sulfone, intermediates.
Glutathione carbamoylation with S-methyl N,N-diethylthiolcarbamate sulfoxide and sulfone: Mitochondrial low K(m) aldehyde dehydrogenase inhibition and implications for its alcohol-deterrent action
Ningaraj, Nagendra S.,Schloss, John V.,Williams, Todd D.,Faiman, Morris D.
, p. 749 - 756 (1998)
S-Methyl N,N-diethylthiolcarbamate sulfoxide (DETC-MeSO) and sulfone (DETC-MeSO2) both inhibit rat liver low K(m) aldehyde dehydrogenase (ALDH2) in vitro and in vivo (Nagendra et al., Biochem Pharmacol 47: 1465-1467, 1994). DETC-MeSO has been shown to be a metabolite of disulfiram, but DETC- MeSO2 has not. Studies were carried out to further investigate the inhibition of ALDH2 by DETC-MeSO and DETC-MeSO2. In an in vitro system containing hydrogen peroxide and horseradish peroxidase, the rate of DETC- MeSO oxidation corresponded to the rate of DETC-MeSO2 formation. Carbamoylation of GSH by both DETC-MeSO and DETC-MeSO2 was observed in a rat liver S9 fraction. Carbamoylation of GSH was not observed in the presence of N-methylmaleimide. In in vitro studies, DETC-MeSO and DETC-MeSO2 were equipotent ALDH2 inhibitors when solubilized mitochondria were used, but DETC-MeSO was approximately four times more potent than DETC-MeSO2 in intact mitochondria. In studies with rats, the dose (i,p. or oral) required to inhibit 50% ALDH2 (ED50) was 3.5 mg/kg for DETC-MeSO and approximately 35 mg/kg for DETC-MeSO2, approximately a 10-fold difference. Furthermore, maximum ALDH2 inhibition occurred 1 hr after DETC-MeSO administration, whereas maximal ALDH2 inhibition occurred 8 hr after DETC-MeSO2 dosing. DETC-MeSO is, therefore, not only a more potent ALDH2 inhibitor than DETC- MeSO2 in vivo, but also in vitro when intact mitochondria are utilized. The in vitro results thus support the in vivo findings. Since oxidation of DETC- MeSO can occur both enzymatically and non-enzymatically, it is possible that DETC-MeSO2 is formed in vivo. DETC-MeSO2, however, is not as effective as DETC-MeSO in inhibiting ALDH2, probably because it has difficulty penetrating the mitochondrial membrane. Thus, even if DETC-MeSO2 is formed in vivo from DETC-MeSO, it is the metabolite DETC-MeSO that is most likely responsible for the inhibition of ALDH2 after disulfiram administration.
S-Methyl N,N-Diethylthiocarbamate Sulfone, a Potential Metabolite of Disulfiram and Potent Inhibitor of Low Km Mitochondrial Aldehyde Dehydrogenase
Mays, Dennis C.,Nelson, Albert N.,Fauq, Abdul H.,Shriver, Zachary H.,Veverka, Karen A.,et al.
, p. 693 - 700 (2007/10/03)
Disulfiram inhibits hepatic aldehyde dehydrogenase (ALDH) causing an accumulation of acetaldehyde after ethanol ingestion. It is thought that disulfiram is too short-lived in vivo to directly inhibit ALDH, but instead is biotransformed to reactive metabolites that inhibit the enzyme. S-Methyl N,N-diethylthiocarbamate (MeDTC) sulfoxide has been identified in the blood of animals given disulfiram and is a potent inhibitor of ALDH (Hart and Faiman, Biochem Pharmacol 46: 2285-2290, 1993). MeDTC sulfone is a logical metabolite of MeDTC sulfoxide. Therefore, we investigated the effects of MeDTC sulfone on the activity of rat hepatic low Km mitochondrial ALDH, the major enzyme in the metabolism of acetaldehyde. MeDTC sulfone inhibited the low Km mitochondrial ALDH in vitro with an IC50 of 0.42 +/- 0.04 μM (mean +/- SD, N = 5) compared with disulfiram, which had an IC50 of 7.5 +/- 1.2 μM under the same conditions. The inhibition of ALDH by MeDTC sulfone was time dependent. The decline in ALDH activity followed pseudo first-order kinetics with an apparent half-life of 2.1 min at 0.6 μM MeDTC sulfone. Inhibition of ALDH by MeDTC sulfone was apparently irreversible; dilution of the inhibited enzyme did not restore lost activity. The substrate (acetaldehyde, 80 μM) and cofactor (NAD, 0.5 mM) together completely protected ALDH from inhibition by MeDTC sulfone; substrate alone partially protected the enzyme. Addition of either thiol-containing compound glutathione (GSH) or dithiothreitol (DTT) to MeDTC sulfone before incubation with the enzyme increased the IC50 of MeDTC sulfone by 7- to 14-fold. Neither GSH nor DTT could restore lost ALDH activity after exposure of the enzyme to MeDTC sulfone. Results of these studies indicate that MeDTC sulfone, a potential metabolite of disulfiram, is a potent, irreversible inhibitor of low Km mitochondrial ALDH.
