Identification of determinants of variable carbonyl reductase 1 (CBR1) expression
Kalabus, James Lee
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Human carbonyl reductases (CBRs) are ubiquitous, monomeric, low molecular weight, NADPH-dependent, cytosolic enzymes belonging to the short-chain dehydrogenase family. CBRs are responsible for the two-electron reduction of reactive aldehyde or ketone moieties into a hydroxyl group. To date, CBR1 is the best-characterized member of this family of enzymes. Among its most notable substrates are the anticancer anthracyclines, which are considered to be some of the most effective chemotherapeutics ever developed and have been successfully employed for more than 40 years in the treatment of an array of solid (sarcomas and carcinomas) and hematological (leukemias and lymphomas) tumors. Their utilization in the clinic, however, is limited by the risk of acute and/or chronic cardiotoxicity. Doxorubicin and daunorubicin, the two natural antineoplastic anthracyclines, are extensively reduced by carbonyl reductases into their corresponding C-13 alcohol metabolites, a metabolic pathway that has been firmly linked to the pathogenesis of anthracycline-induced cardiotoxicity in biochemical studies and murine models. Carbonyl reductase activity varies widely among individuals and may contribute to the unpredictable pharmacodynamics of CBR1 drug substrates. To better understand the apparently capricious nature of anthracycline-related cardiotoxicity, we identified three major determinants of CBR1 variability. First, we examined variability in CBR1 expression through the lens of a relevant disease state in Down syndrome. Individuals with Down syndrome have an increased risk of developing acute myeloid leukemia. Down syndrome cancer patients are most frequently treated with chemotherapy regimens heavily reliant on daunorubicin. Unfortunately, this population is at significantly greater risk of developing anthracycline-related cardiotoxicity. Because CBR1 is located in the Down syndrome critical region of chromosome 21, 21q22.12, we aimed to determine if CBR1 is subjected to a Down syndrome-induced gene dosage effect. We found that heart samples from donors with Down syndrome exhibited significantly increased CBR1 mRNA and CBR1 protein expression as well as CBR1 activity compared to heart samples from donors without Down syndrome. The second determinant of CBR1 expression we detected was exposure to benzo[ a ]pyrene, a prominent constituent of cigarette smoke. A recent microarray study reported that levels of CBR1 mRNA expression were elevated in the lungs of smokers with cancer versus the lungs of smokers without cancer. Therefore, we sought to pinpoint the chemical that might be responsible for inducing CBR1 expression. We discovered that benzo[ a ]pyrene exposure is capable of up-regulating expression of CBR1 mRNA and CBR1 protein in human lung tissue and in a lung cancer cell line, A549, via the aryl hydrocarbon receptor pathway. Finally, we characterized an epigenetic pathway responsible for post-transcriptional regulation of CBR1 . Our work revealed two microRNAs that are capable of binding to the 3'-untranslated region (3'-UTR) of CBR1 to diminish CBR1 protein synthesis. We extended these findings by defining the impact of a common single-nucleotide polymorphism in the 3'-UTR on microRNA interactions with CBR1 . Our work elucidates several factors contributing to the complex regulation of CBR1 expression.