Metabolite mimicry identifies butyrate analogs with select protective functions in the intestinal mucosa

Microbial-derived short-chain fatty acids regulate a variety of pathways in the healthy colonic mucosa. In particular, butyrate serves as the primary energy source for colonocytes and regulates gene transcription by stabilizing the transcription factor hypoxia-inducible-factors (HIF) and functioning as a histone deacetylase (HDAC) inhibitor. A limitation of butyrate as a therapeutic is its rapid metabolism in differentiated colonocytes. Furthermore, intestinal stem cells (ISCs) respond differently to butyrate, preferentially using glucose for energy procurement. To address these limitations, we explored metabolite-mimicry to discover compounds with potent or selective biological responses within the butyrate pathway(s). We discovered an analog, 3-chlorobutyrate (3-Cl BA), that significantly enhances epithelial barrier formation and wound healing in vitro. Mechanistically, we revealed that 3-Cl BA is a potent HDAC inhibitor. Furthermore, unlike butyrate, 3-Cl BA does not stabilize HIF and it is not used as metabolic fuel. In vivo studies in a DSS-colitis model revealed that contrary to butyrate, 3-Cl BA is protective. Studies in stem-like colonoids demonstrated that only butyrate inhibits ISC proliferation and differentiation. Furthermore, it was recently reported that HIF stabilization inhibits ISCs activity. Given the fact that butyrate but not 3-Cl BA stabilizes HIF, we surmised that 3-Cl BA would circumvent these detrimental functional consequences. We demonstrate here that pharmacologic HIF stabilization inhibits colonoid differentiation and that genetic loss of HIF significantly promotes ISC differentiation. This study reveals a promising butyrate analog protective in colitis and demonstrates the advantages of metabolite-mimicry to dissect selective biological functions from major metabolites in the gut.

Butyrate is a well-studied microbial short-chain fatty acid that regulates a number of mucosal pathways and is paramount in maintaining intestinal integrity. In health, it is a major source of energy for colonocytes and regulates gene transcription. The role of butyrate in disease is still controversial and not well understood. When butyrate is not metabolized or well-utilized (e.g. disease), it accumulates in intestinal stem cells leading to reduced cell proliferation and differentiation, thereby hampering intestinal barrier recovery. In this study, we describe a butyrate analog that enhances epithelial barrier formation and wound healing. Furthermore, as opposed to native butyrate, this butyrate analog is protective in a colitis mouse model and does not exhibit detrimental influences on intestinal stem cells.