Abstract
Human carbonyl reductase 1 (CBR1) metabolizes endogenous and xenobiotic substrates such as the fever mediator, prostaglandin E2 (PGE2), and the anticancer anthracycline drug, daunorubicin. We screened 33 CBR1 full-length cDNA samples from white and black liver donors and performed database analyses to identify genetic determinants of CBR1 activity. We pinpointed a single nucleotide polymorphism on CBR1 (CBR1 V88I) that encodes for a valine-to-isoleucine substitution for further characterization. We detected the CBR1 V88I polymorphism in DNA samples from individuals with African ancestry (p = 0.986, q = 0.014). Kinetic studies revealed that the CBR1 V88 and CBR1 I88 isoforms have different maximal velocities for daunorubicin (Vmax CBR1 V88, 181 ± 13 versus Vmax CBR1 I88, 121 ± 12 nmol/min · mg, p < 0.05) and PGE2 (Vmax CBR1 V88, 53 ± 7 versus Vmax CBR1 I88, 35 ± 4 nmol/min · mg, p < 0.01). Concomitantly, CBR1 V88 produced higher levels of the cardiotoxic metabolite daunorubicinol compared with CBR1 I88 (1.7-fold, p < 0.0001). Inhibition studies demonstrated that CBR1 V88 and CBR1 I88 are distinctively inhibited by the flavonoid, rutin (IC50 CBR1 V88, 54.0 ± 0.4 μM versus IC50 CBR1 I88, 15.0 ± 0.1 μM, p < 0.001). Furthermore, isothermal titration calorimetry analyses together with molecular modeling studies showed that CBR1 V88I results in CBR1 isoforms with different binding affinities for the cofactor NADPH (Kd CBR1 V88, 6.3 ± 0.6 μM versus Kd CBR1 I88, 3.8 ± 0.5 μM). These studies characterize the first functional genetic determinant of CBR1 activity toward relevant physiological and pharmacological substrates.
Footnotes
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This work was supported by National Institutes of Health/National Institute of General Medical Sciences Grant R01GM73646 to J.G.B., and by National Institutes of Health/National Institute of General Medical Sciences Grant U01GM61393 and National Institutes of Health/National Institute of General Medical Sciences Pharmacogenetics Research Network and Database Grant U01GM61374 (http://pharmgkb.org). V.G.-C. is a Merck fellow.
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doi:10.1124/dmd.107.014779.
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ABBREVIATIONS: CBR, carbonyl reductase; SNP, single nucleotide polymorphism; PCR, polymerase chain reaction; PGE2, prostaglandin E2; HPLC, high-performance liquid chromatography.
- Received January 10, 2007.
- Accepted March 6, 2007.
- The American Society for Pharmacology and Experimental Therapeutics
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