title: Naturally occurring variants of human aldo-keto reductases with reduced in vitro metabolism of daunorubicin and doxorubicin.
abstract: Doxorubicin (DOX) and daunorubicin (DAUN) are effective anticancer drugs; however, considerable interpatient variability exists in their pharmacokinetics. This may be caused by altered metabolism by nonsynonymous single-nucleotide polymorphisms (ns-SNPs) in genes encoding aldo-keto reductases (AKRs) and carbonyl reductases. This study examined the effect of 27 ns-SNPs, in eight human genes, on the in vitro metabolism of both drugs to their major metabolites, doxorubicinol and daunorubicinol. Kinetic assays measured metabolite levels by high-performance liquid chromatography separation with fluorescence detection using purified, histidine-tagged, human wild-type, and variant enzymes. Maximal rate of activity (V(max)), substrate affinity (K(m)), turnover rate (k(cat)), and catalytic efficiency (k(cat)/K(m)) were determined. With DAUN as substrate, variants for three genes exhibited significant differences in these parameters compared with their wild-type counterparts: the A106T, R170C, and P180S variants significantly reduced metabolism compared with the AKR1C3 wild-type (V(max), 23-47% decrease; k(cat), 22-47%; k(cat)/K(m), 38-44%); the L311V variant of AKR1C4 significantly decreased V(max) (47% lower) and k(cat) and k(cat)/K(m) (both 43% lower); and the A142T variant of AKR7A2 significantly affected all kinetic parameters (V(max) and k(cat), 61% decrease; K(m), 156% increase; k(cat)/K(m), 85% decrease). With DOX, the R170C and P180S variants of AKR1C3 showed significantly reduced V(max) (41-44% decrease), k(cat) (39-45%), and k(cat)/K(m) (52-69%), whereas the A142T variant significantly altered all kinetic parameters for AKR7A2 (V(max), 41% decrease; k(cat), 44% decrease; K(m), 47% increase; k(cat)/K(m), 60% decrease). These findings suggest that ns-SNPs in human AKR1C3, AKR1C4, and AKR7A2 significantly decrease the in vitro metabolism of DOX and DAUN.
authors.affiliation: Division of Pharmaceutics and Biopharmaceutics, Faculty of Pharmaceutical Sciences, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Onkar S
authors.initials: OS
authors.lastname: Bains
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Grigliatti
authors.forename: Ronald E
authors.initials: RE
authors.lastname: Reid
authors.forename: K Wayne
authors.initials: KW
authors.lastname: Riggs
mesh_terms: D015089:20-Hydroxysteroid Dehydrogenases / Q000235:genetics / Q000378:metabolism; D015096:3-Hydroxysteroid Dehydrogenases / Q000235:genetics / Q000378:metabolism; D000429:Alcohol Oxidoreductases / Q000235:genetics* / Q000378:metabolism*; D000449:Aldehyde Reductase / Q000235:genetics / Q000378:metabolism; D000074425:Aldo-Keto Reductase Family 1 Member C3; D000074408:Aldo-Keto Reductases; D055162:Biocatalysis; D003630:Daunorubicin / Q000378:metabolism*; D004317:Doxorubicin / Q000378:metabolism*; D005787:Gene Frequency; D005985:Glyceraldehyde / Q000378:metabolism; D006801:Humans; D006911:Hydroxyprostaglandin Dehydrogenases / Q000235:genetics / Q000378:metabolism; D006913:Hydroxysteroid Dehydrogenases / Q000235:genetics / Q000378:metabolism; D007189:Indans / Q000378:metabolism; D007700:Kinetics; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000235:genetics / Q000378:metabolism; D010088:Oxidoreductases / Q000235:genetics / Q000378:metabolism; D010616:Phenanthrenes / Q000378:metabolism; D020641:Polymorphism, Single Nucleotide / Q000502:physiology*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D024483:Vitamin K 3 / Q000378:metabolism; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)
chemical_list: D015089:20-Hydroxysteroid Dehydrogenases; D015096:3-Hydroxysteroid Dehydrogenases; D000429:Alcohol Oxidoreductases; D000449:Aldehyde Reductase; D000074425:Aldo-Keto Reductase Family 1 Member C3; D000074408:Aldo-Keto Reductases; D003630:Daunorubicin; D004317:Doxorubicin; D005985:Glyceraldehyde; D006911:Hydroxyprostaglandin Dehydrogenases; D006913:Hydroxysteroid Dehydrogenases; D007189:Indans; D024101:Mitochondrial Proteins; D016660:NAD(P)H Dehydrogenase (Quinone); D010088:Oxidoreductases; D010616:Phenanthrenes; D011994:Recombinant Proteins; D024483:Vitamin K 3; C005399:9,10-phenanthrenequinone; C016515:1-indanol; C115140:3 alpha-beta, 20 beta-hydroxysteroid dehydrogenase; C120471:AKR1B10 protein, human; C082765:aflatoxin B1 aldehyde reductase; C484670:AKR1B1 protein, human; C435496:AKR1C2 protein, human; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific); C487341:AKR1C3 protein, human; C048338:trans-1,2-dihydrobenzene-1,2-diol dehydrogenase; C533692:CBR4 protein, human