Ritonavir (RTV) is on the World Health Organization’s list of essential medicines for antiretroviral therapy, but can cause hepatotoxicity by unknown mechanisms. Multiple clinical studies found that hepatotoxicity occurred in 100% of participants who were pretreated with rifampicin or efavirenz followed by RTV-containing regimens. Both rifampicin and efavirenz are activators of the pregnane X receptor (PXR), a transcription factor with marked interspecies differences in ligand-dependent activation. Using PXR-humanized mouse models, we recapitulated the RTV hepatotoxicity observed in the clinic. PXR was found to modulate RTV hepatotoxicity through CYP3A4-dependent pathways involved in RTV bioactivation, oxidative stress, and endoplasmic reticulum stress. In summary, the current work demonstrated the essential roles of human PXR and CYP3A4 in RTV hepatotoxicity, which can be applied to guide the safe use of RTV-containing regimens in the clinic.

The transporter ABCG2 determines phototoxicity and hepatotoxicity in erythropoietic protoporphyria.

Although safety of drug candidates is carefully monitored in preclinical and clinical studies using a variety of approaches, drug toxicity may still occur in clinical practice. Therefore, novel approaches are needed to complement the current drug safety evaluation system. Metabolomics comprehensively analyzes the metabolites altered by drug exposure, which can therefore be used to profile drug metabolism, endobiotic metabolism, and drug-microbiota interactions. The information from metabolomic analysis can be used to determine the off-targets of a drug candidate and thus provide a mechanistic understanding of drug toxicity. We herein discuss the opportunities of metabolomics in drug safety evaluation.

Multidrug Resistance Proteins (MRPs) are transporters that play critical roles in cancer even though the physiological substrates of these enigmatic transporters are poorly elucidated. In Caenorhabditis elegans, MRP5/ABCC5 is an essential heme exporter because mrp-5 mutants are unviable due to their inability to export heme from the intestine to extraintestinal tissues. Heme supplementation restores viability of these mutants but fails to restore male reproductive deficits. Correspondingly, cell biological studies show that MRP5 regulates heme levels in the mammalian secretory pathway even though MRP5 knockout (KO) mice do not show reproductive phenotypes. The closest homolog of MRP5 is MRP9/ABCC12, which is absent in C. elegans, raising the possibility that MRP9 may genetically compensate for MRP5. Here, we show that MRP5 and MRP9 double KO (DKO) mice are viable but reveal significant male reproductive deficits. Although MRP9 is highly expressed in sperm, MRP9 KO mice show reproductive phenotypes only when MRP5 is absent. Both ABCC transporters localize to mitochondrial-associated membranes, dynamic scaffolds that associate the mitochondria and endoplasmic reticulum. Consequently, DKO mice reveal abnormal sperm mitochondria with reduced mitochondrial membrane potential and fertilization rates. Metabolomics show striking differences in metabolite profiles in the DKO testes, and RNA sequencing shows significant alterations in genes related to mitochondrial function and retinoic acid metabolism. Targeted functional metabolomics reveal lower retinoic acid levels in the DKO testes and higher levels of triglycerides in the mitochondria. These findings establish a model in which MRP5 and MRP9 play a concerted role in regulating male reproductive functions and mitochondrial sufficiency. Significance Multidrug Resistance Proteins (MRPs) are typically implicated in cancer biology. Here, we show that MRP9 and MRP5 localize to mitochondrial-associated membranes and play a concerted role in maintaining mitochondrial homeostasis and male reproductive fitness. Our work fills in significant gaps in our understanding of MRP9 and MRP5 with wider implications in male fertility. It is plausible that variants in these transporters are associated with male reproductive dysfunction.