This is a proposal to study the mechanism of action of disulfiram. Disulfiram is currently the only drug which is approved in the United States for aversion therapy in the treatment of alcoholism. Although it is known that this drug causes an inhibition of the enzyme aldehyde dehydrogenase, and thus interferes with the metabolism of ethanol, the precise mechanism is unknown. The purpose of this study is to clearly define the manner in which disulfiram inhibits aldehyde dehydrogenase in vivo. Since it appears that activation of metabolites of disulfiram is involved in the inhibition, these studies will focus on such activating pathways, specifically methylation and oxidation. Studies will be performed in microsomal systems, animals, and humans. Active metabolites of disulfiram will be identified by mass spectrometry. In addition, we will address the question of the variation in the response to disulfiram which is seen when it is used clinically. A possible explanation for this finding is that genetic differences in the metabolism of disulfiram may lead to different degrees of aldehyde dehydrogenase inhibition. We will test, in animals as well as in humans whether genetic polymorphisms in methylation are responsible for the observed variations. The studies described in this proposal will provide a better understanding of the mechanism of action of disulfiram, the basis for clinical variation in its use, and may result in a rational strategy for developing new compounds for the treatment of alcoholism.

Agency
National Institute of Health (NIH)
Institute
National Institute on Alcohol Abuse and Alcoholism (NIAAA)
Type
Research Project (R01)
Project #
5R01AA009543-02
Application #
3113599
Study Section
Special Emphasis Panel (SRCA (51))
Project Start
1992-09-30
Project End
1995-08-31
Budget Start
1993-09-01
Budget End
1994-08-31
Support Year
2
Fiscal Year
1993
Total Cost
Indirect Cost
Name
Mayo Clinic, Rochester
Department
Type
DUNS #
City
Rochester
State
MN
Country
United States
Zip Code
55905
Shen, M L; Johnson, K L; Mays, D C et al. (2001) Determination of in vivo adducts of disulfiram with mitochondrial aldehyde dehydrogenase. Biochem Pharmacol 61:537-45
Shen, M L; Benson, L M; Johnson, K L et al. (2001) Effect of enzyme inhibitors on protein quaternary structure determined by on-line size exclusion chromatography-microelectrospray ionization mass spectrometry. J Am Soc Mass Spectrom 12:97-104
Lipsky, J J; Shen, M L; Naylor, S (2001) In vivo inhibition of aldehyde dehydrogenase by disulfiram. Chem Biol Interact 130-132:93-102
Pike, M G; Mays, D C; Macomber, D W et al. (2001) Metabolism of a disulfiram metabolite, S-methyl N,N-diethyldithiocarbamate, by flavin monooxygenase in human renal microsomes. Drug Metab Dispos 29:127-32
Shen, M L; Lipsky, J J; Naylor, S (2000) Role of disulfiram in the in vitro inhibition of rat liver mitochondrial aldehyde dehydrogenase. Biochem Pharmacol 60:947-53
Shen, M L; Johnson, K L; Mays, D C et al. (2000) Identification of the protein-drug adduct formed between aldehyde dehydrogenase and S-methyl-N,N-diethylthiocarbamoyl sulfoxide by on-line proteolytic digestion high performance liquid chromatography electrospray ionization mass spectrometry. Rapid Commun Mass Spectrom 14:918-23
Kathmann, E C; Naylor, S; Lipsky, J J (2000) Rat liver constitutive and phenobarbital-inducible cytosolic aldehyde dehydrogenases are highly homologous proteins that function as distinct isozymes. Biochemistry 39:11170-6
Pike, M G; Martin, Y N; Mays, D C et al. (1999) Roles of FMO and CYP450 in the metabolism in human liver microsomes of S-methyl-N,N-diethyldithiocarbamate, a disulfiram metabolite. Alcohol Clin Exp Res 23:1173-9
Kathmann, E C; Lipsky, J J (1999) Cloning and expression of a cDNA encoding a constitutively expressed rat liver cytosolic aldehyde dehydrogenase. Adv Exp Med Biol 463:237-41
Mays, D C; Tomlinson, A J; Johnson, K L et al. (1999) Inhibition of human mitochondrial aldehyde dehydrogenase by metabolites of disulfiram and structural characterization of the enzyme adduct by HPLC-tandem mass spectrometry. Adv Exp Med Biol 463:61-70

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