A discovery study of daunorubicin induced cardiotoxicity in a sample of acute myeloid leukemia patients prioritizes P450 oxidoreductase polymorphisms as a potential risk factor.

Anthracyclines are very effective chemotherapeutic agents; however, their use is hampered by the treatment-induced cardiotoxicity. Genetic variants that help define patient's sensitivity to anthracyclines will greatly improve the design of optimal chemotherapeutic regimens. However, identification of such variants is hampered by the lack of analytical approaches that address the complex, multi-genic character of anthracycline induced cardiotoxicity (AIC). Here, using a multi-SNP based approach, we examined 60 genes coding for proteins involved in drug metabolism and efflux and identified the P450 oxidoreductase (POR) gene to be most strongly associated with daunorubicin induced cardiotoxicity in a population of acute myeloid leukemia (AML) patients (FDR adjusted p-value of 0.15). In this sample of cancer patients, variation in the POR gene is estimated to account for some 11.6% of the variability in the drop of left ventricular ejection fraction (LVEF) after daunorubicin treatment, compared to the estimated 13.2% accounted for by the cumulative dose and ethnicity. In post-hoc analysis, this association was driven by 3 SNPs-the rs2868177, rs13240755, and rs4732513-through their linear interaction with cumulative daunorubicin dose. The unadjusted odds ratios (ORs) and confidence intervals (CIs) for rs2868177 and rs13240755 were estimated to be 1.89 (95% CI: 0.7435-4.819; p = 0.1756) and 3.18 (95% CI: 1.223-8.27; p = 0.01376), respectively. Although the contribution of POR variants is expected to be overestimated due to the multiple testing performed in this small pilot study, given that cumulative anthracycline dose is virtually the only factor used clinically to predict the risk of cardiotoxicity, the contribution that genetic analyses of POR can make to the assessment of this risk is worthy of follow up in future investigations.

Single-nucleotide polymorphisms in reductase genes are not associated with response to daunorubicin-based remission induction.

BACKGROUND: To improve the quality of care for patients with acute myeloid leukemia (AML), biomarkers predictive of response to the standard daunorubicin-based induction therapy are needed. Genetic variants affecting daunorubicin metabolism are attractive candidates for such biomarkers. METHODS: We have previously shown that 13 of the naturally occurring nonsynonymous single-nucleotide polymorphisms (SNP) in the reductase genes affect daunorubicin metabolism in vitro. Here, we test these SNPs individually and jointly for association with response to one cycle of daunorubicin-based chemotherapy in a sample of 189 patients with acute myelogenous leukemia. RESULTS: Of the 13 SNPs included in this study, only 5 passed quality control filters. No association was found between these 5 SNPs and response to one cycle of daunorubicin-based induction therapy in either individual or joint effect tests. CONCLUSIONS: Despite their showing in vitro effect on metabolic rate of daunorubicin, the nonsynonymous SNPs in the reductase genes on their own are not significant contributors to the observed variability in response to daunorubicin therapy and thus, as singularities, are not useful biomarkers of this outcome. IMPACT: The results of this investigation provide important information for studies on personalization of anthracycline-based therapies.

Single-nucleotide polymorphisms in aldo-keto and carbonyl reductase genes are not associated with acute cardiotoxicity after daunorubicin chemotherapy.

BACKGROUND: Evidence suggests that interpatient variability in anthracycline metabolic rate may contribute to the cardiotoxicity associated with anthracycline-based chemotherapy. Therefore, polymorphisms in the anthracycline metabolizing enzymes have been proposed as potential biomarkers of anthracycline-induced cardiotoxicity (AIC). METHODS: We have previously shown that 13 of the naturally occurring nonsynonymous single-nucleotide polymorphisms (nsSNP) in the aldo-keto reductases (AKR) and carbonyl reductases (CBR) reduce anthracycline metabolic rate in vitro. Here, we test these SNPs individually and jointly for association with daunorubicin-induced cardiotoxicity in patients with acute myeloid leukemia (AML). RESULTS: Five of the 13 nsSNPs exhibiting an in vitro effect on anthracycline metabolism were detected among the 185 patients with AML. No association was found between the SNPs and daunorubicin-induced cardiotoxicity in either individual or joint effect analyses. CONCLUSIONS: Despite the shown in vitro effect of nsSNPs in reductase genes on anthracycline metabolic rate, on their own these SNPs do not explain enough variability in cardiotoxicity to be useful markers of this adverse event. IMPACT: The results of this study provide important information for biomarker studies on side effects of anthracycline chemotherapy.

A functional assay for G-protein-coupled receptors using stably transformed insect tissue culture cell lines.

Insect cells are an underexplored resource for functional G-protein-coupled receptor (GPCR) assays, despite a strong record in biochemical (binding) assays. Here we describe the use of vectors capable of creating stably transformed insect cell lines to generate a cell-based functional GPCR assay. This assay employs the luminescent photoprotein aequorin and the promiscuous G-protein subunit Galpha16 and is broadly applicable to human GPCRs. We demonstrate that the assay can quantitate ligand concentration-activity relationships for seven different human GPCRs, can differentiate between partial and full agonists, and can determine rank order potencies for both agonists and antagonists that match those seen with other assay systems. Human Galpha16 improves signal strength but is not required for activity with some receptors. The coexpression of human and bovine betagamma subunits and/or phospholipase Cbeta makes no difference to agonist efficacy or potency. Two different receptors expressed in the same cell line respond to their specific agonists, and two different cell lines (Sf9 and High 5) are able to functionally detect the same expressed GPCR. Sf9 cells have the capability to produce fully functional human receptors, allied to a low background of endogenous receptors, and so are a valuable system for investigating orphan GPCRs and receptor dimerization.