A reduced complexity cross between BALB/c substrains identifies Zhx2 as a candidate gene underlying oxycodone metabolite brain concentration and state‐dependent learning of opioid reward

Abstract

Understanding the pharmacokinetic profile of an opioid drug is vital to therapeutic success, and mutations in human PK genes can drastically alter therapeutic efficacy of opioids. We observed that at 30 min post‐oxycodone administration (1.25 mg/kg, i.p.) BALB/cJ mice showed a higher whole brain concentration of oxycodone, and female specific increase in noroxycodone, and oxymorphone compared to BALB/cByJ. This observation could explain the sex‐specific increase in oxycodone state‐dependent conditioned place preference in BALB/cJ female mice. To potentially link behavioral differences with PK differences, we conducted quantitative trait locus (QTL) mapping of whole brain oxycodone and metabolite concentrations in a reduced complexity cross (RCC). Because BALB/cJ and BALB/cByJ substrains differ by ~8,500 SNPs/indels, large genetic loci identified in an F2 cross are offset by a dramatic reduction in potentially causal variants. QTL mapping in 133 BALB/cJ x BALB/cByJ F2 mice (68F, 65M) revealed a single QTL on chromosome 15 associated with brain oxymorphone concentration that explained 29% of the phenotypic variance in females. Oxymorphone is a full agonist at the mu opioid receptor, with 8x the potency of oxycodone, and likely contributes to oxycodone addictive properties. Hippocampal and striatal cis‐eQTL analysis revealed genetically regulated expression of Zhx2, a transcriptional inhibitor known to harbor a private BALB/cJ retroviral insertion that dramatically reduces protein expression and leads to sex specific dysregulation of CYP450 genes within the liver. Whole brain mass spectroscopy proteomics in the parental strains corroborated these eQTL findings. We hypothesize that decreased Zhx2 expression leads to increased CYP450 expression, increased brain oxymorphone, and increased oxycodone‐induced behaviors. Interestingly, human GWAS of nicotine consumption identified a nominal association (10^‐7) with ZHX2, indicating that this transcriptional repressor could influence metabolism of multiple drugs of abuse.