Factors influencing the induction of DT-diaphorase activity by 1,2-dithiole-3-thione in human tumor cell lines

Abstract

NAD(P)H:(quinone acceptor)oxidoreductase (DT-diaphorase) is a two-electron reducing enzyme that activates bioreductive antitumor agents and is induced by a wide variety of compounds including 1,2-dithiole-3-thione (D3T). We investigated factors influencing DT-diaphorase induction in fourteen human tumor cell lines. Four cell lines had basal DT-diaphorase activity that was increased by D3T treatment (group A), six cell lines had basal DT-diaphorase activity but the activity was not increased by D3T (group B), and four cell lines had low enzyme activity without, or with, D3T (group C). Two cell lines in group A and two cell lines in group B had a C to T polymorphism at base 609 in the NQO 1, DT-diaphorase gene, in one allele, while all four cell lines in group C were homozygous mutants. The base 609 mutant NQO 1 gene produces a protein with little enzyme activity. In group A, D3T increased NQO 1 mRNA and wild-type protein, and also increased mutant protein in the two heterozygous cell lines. In group B, the inducer slightly increased NQO 1 mRNA, did not increase the wild-type protein, but did increase the mutant protein in the two heterozygous cell lines. In group C, D3T increased NQO 1 mRNA as well as its mutant enzyme product. Transfection of the mutant NQO 1 gene into cells with two wild-type alleles did not alter DT-diaphorase activity. The results suggest that the lack of induction of DT-diaphorase activity is transcriptional in nature, that basal and induced expression of DT-diaphorase are regulated independently, and that mutant NQO 1 does not act as a dominant-negative to suppress DT-diaphorase activity.