A Genetic Signature of Spina Bifida Risk from Pathway-Informed Comprehensive Gene-Variant Analysis

Despite compelling epidemiological evidence that folic acid supplements reduce the frequency of neural tube defects (NTDs) in newborns, common variant association studies with folate metabolism genes have failed to explain the majority of NTD risk. The contribution of rare alleles as well as genetic interactions within the folate pathway have not been extensively studied in the context of NTDs. Thus, we sequenced the exons in 31 folate-related genes in a 480-member NTD case-control population to identify the full spectrum of allelic variation and determine whether rare alleles or obvious genetic interactions within this pathway affect NTD risk. We constructed a pathway model, predetermined independent of the data, which grouped genes into coherent sets reflecting the distinct metabolic compartments in the folate/one-carbon pathway (purine synthesis, pyrimidine synthesis, and homocysteine recycling to methionine). By integrating multiple variants based on these groupings, we uncovered two provocative, complex genetic risk signatures. Interestingly, these signatures differed by race/ethnicity: a Hispanic risk profile pointed to alterations in purine biosynthesis, whereas that in non-Hispanic whites implicated homocysteine metabolism. In contrast, parallel analyses that focused on individual alleles, or individual genes, as the units by which to assign risk revealed no compelling associations. These results suggest that the ability to layer pathway relationships onto clinical variant data can be uniquely informative for identifying genetic risk as well as for generating mechanistic hypotheses. Furthermore, the identification of ethnic-specific risk signatures for spina bifida resonated with epidemiological data suggesting that the underlying pathogenesis may differ between Hispanic and non-Hispanic groups

Pharmacokinetics and bioavailability of the isoflavone biochanin A in rats

Biochanin A(BCA) is a dietary isoflavone present in legumes, most notably red clover, and in many herbal dietary supplements. BCA has been reported to have chemopreventive properties and is metabolized to the isoflavone genistein (GEN), BCA conjugates, and GEN conjugates. The metabolites may contribute to the chemopreventive effects of BCA. The absorption, metabolism, and disposition of BCA have not been determined in rats. Our objective was to evaluate the pharmacokinetics and metabolism of BCA in rats. Male Sprague-Dawley rats were administered BCA by intravenous injection (1 and 5 mg/kg), by intraperitoneal injection (5 and 50 mg/kg), and orally (5 and 50 mg/kg). Plasma and bile samples were enzymatically hydrolyzed in vitro to determine conjugate concentrations for BCA and GEN. Equilibrium dialysis was used to determine protein binding. The BCA and GEN concentrations in plasma, urine, and bile were determined by liquid chromatography-tandem mass spectrometry (LC/MS/MS). The pharmacokinetic parameters of BCA were analyzed by noncompartmental analysis. Significant levels of BCA conjugates and GEN conjugates were detected in plasma and bile. Both BCA and GEN were found to have a high clearance and a large apparent volume of distribution; the bioavailability of both was poor (<4%). Reentry peaks were evident after oral administration of both BCA and GEN, suggesting enterohepatic cycling. The free fraction of BCA in rat plasma was 1.5%. A2-compartment model that included both linear and nonlinear clearance terms and enterohepatic recirculation best described the plasma data. This represents the first evaluation of the dose-dependent pharmacokinetics and metabolism of BCA in rats