Metabolism of cancer chemotherapeutics by the human gut microbiome

PROJECT SUMMARY Rigorous data from our lab and others indicate that the gut microbiome may an underappreciated contributor to inter-individual variations in cancer drug efficacy and side effect profiles; however, we currently lack the mechanistic insights and data from preclinical mouse models necessary to inform ongoing studies in cancer patients. We selected fluoropyrimidines, including 5-fluorouracil (5-FU) and its prodrug capecitabine (CAP), as an initial test case due to their critical role in colorectal cancer (CRC) therapy, increasing oral administration, highly variable pharmacokinetics, and unexplained differences in efficacy and toxicity. We propose a series of in vitro and mouse studies to dissect the human gut bacterial species, genes, and enzymes responsible for the metabolism of 5-FU (Aim 1) and CAP (Aim 2), including their downstream consequences for drug pharmacokinetics (PK) and pharmacodynamics (PD). Our overarching hypothesis is that the oral bioavailability and therapeutic effects of fluoropyrimidines are influenced by pathways for drug metabolism encoded by diverse human gut bacterial species. In Aim 1, we will identify and characterize the primary gut bacterial taxon responsible for 5-FU inactivation through a combination of biochemical and cell-based assays coupled to studies in gnotobiotic and xenograft mouse models. Based on our Preliminary Results, we hypothesize that Anaerostipes is the primary gut bacterial genus responsible for inter-individual variations in the metabolism of 5-FU. In Aim 2, we seek to discover the bacterial enzymes responsible for the activation of CAP to 5-FU, motivated by the surprising finding that E. coli can activate CAP leading to reduced bacterial growth at high concentrations. We hypothesize that E. coli catalyzes a 3-step metabolic pathway that mirrors the mammalian conversion of CAP to 5-FU. Our results in Aim 1 will provide a valuable proof-of-principle for dissecting the conservation and redundancies in clinically relevant microbial biotransformations, helping to move beyond studies of model gut bacteria to identify the most translationally relevant species. Aim 2 is potentially paradigm-shifting in that it would provide definitive evidence for CAP bioactivation outside of hepatocytes and cancer cells, creating new opportunities to improve treatment outcomes and study the physiological role and broader impacts of this metabolic pathway. Taken together, this research plan emphasizes the conservation of the pathways for metabolism of therapeutics across domains of life, highlighting the need to distinguish the relative contributions of human and microbial cells to drug disposition, efficacy, and side effect profiles. Due to our focus on drugs used as current standard of care and naturally occurring bacterial species prevalent in the human gut microbiome, this preclinical research program has clear translational relevance and is highly synergistic with ongoing clinical studies of cancer patients conducted by our team and the broader microbiome field.