Genetic and pharmacologic inhibition of Keap1 dampens the pro-fibrotic program in human hepatic stellate cells

Abstract

Abstract The increasing prevalence of chronic liver diseases (CLDs) is a global health concern. CLDs can cause fibrosis, and there is an urgent need for antifibrotic agents. The primary driver of fibrogenesis, transforming growth factor beta 1 (TGF-β1), activates hepatic stellate cells (HSCs), which are responsible for hepatofibrosis. Activation of the transcription factor NF-E2 p45-related factor 2 (Nrf2) can ameliorate hepatic fibrosis in mice; therefore, we hypothesized that activating/restoring Nrf2 activity could inhibit TGF-β1-mediated fibrogenesis in HSCs. Here, we show that TGF-β1 suppressed the expression of several Nrf2 targets in human HSCs. Genetic manipulation of Nrf2 and its principal repressor Kelch-like ECH-associated protein 1 (Keap1) revealed that Nrf2 knockdown (Nrf2-kd) increased expression of pro-fibrotic markers in unstimulated and TGF-β1-stimulated cells. In Keap1-kd cells (high Nrf2 activity), expression of pro-fibrotic markers was suppressed. To enhance Nrf2 activity, we used Keap1 inhibitors of two types, the electrophilic triterpenoid RTA-405, and the non-electrophilic Keap1-Nrf2 protein–protein interaction inhibitor PRL-295. High-resolution proteomics revealed that pre-treatment with either inhibitor dampens the TGF-β1-driven pro-fibrotic program. Considering the recent approval of the triterpenoid RTA-408 (omaveloxolone) for clinical use, our findings support the development/repurposing of such compounds for prevention and treatment of hepatofibrosis.