A Point Mutation at C151 of Keap1 of Mice Abrogates NRF2 Signaling, Cytoprotection In Vitro and Hepatoprotection In Vivo by Bardoxolone Methyl (CDDO-Me)

by Selleckchem | Jul 16 2023

Bardoxolone methyl (CDDO-Me) is an oleanane triterpenoid in late-stage clinical development for the treatment of patients with diabetic kidney disease. Preclinical studies in rodents demonstrate the efficacy of triterpenoids against carcinogenesis and other diseases including renal ischemia-reperfusion injury, hyperoxia-induced acute lung injury and immune hepatitis. Genetic disruption of NRF2 abrogates protection by triterpenoids, suggesting that induction of the NRF2 pathway may drive this protection. Herein, we examined the effect of a point mutation (C151S) in KEAP1, a repressor of NRF2 signaling, at cysteine 151 in mouse embryo fibroblasts and mouse liver. Induction of target gene transcripts and enzyme activity by CDDO-Me was lost in C151S mutant fibroblasts compared to wild-type. Protection against menadione toxicity was also nullified in the mutant fibroblasts. In mouse liver, CDDO-Me evoked the nuclear translocation of NRF2 followed by increased transcript and activity levels of a prototypic target gene Nqo1 in wild-type, but not C151S mutant mice. To test the role of KEAP1 Cys151 in governing the broader pharmacodynamic action of CDDO-Me, wild-type and C151S mutant mice were challenged with concanavalin A to induce immune hepatitis. Strong protection was seen in wild-type but not C151S mutant mice. RNA-seq analysis of mouse liver from wild-type, C151S mutant and Nrf2-knockout mice revealed a vigorous response of the Nrf2 transcriptome in wild-type, but in neither C151S mutant nor Nrf2-knockout mice. Activation of "off-target" pathways by CDDO were not observed. These data highlight the singular importance of the KEAP1 cysteine 151 sensor for activation of NRF2 signaling by CDDO-Me. Significance Statement Keap1 serves as a key sensor for induction of the cytoprotective signaling pathway driven by the transcription factor NRF2. Mutation of a single cysteine (C151) in KEAP1 abrogates the induction of NRF2 signaling and its downstream cytoprotective actions in vitro and in vivo by bardoxolone methyl (CDDO-Me), a drug in late-stage clinical development. Further, at these bioeffective concentrations/doses, activation of "off-target" pathways by CDDO-Me are not observed, highlighting the singular importance of NRF2 in its mode of action.

Comments:

The passage you provided discusses the role of bardoxolone methyl (CDDO-Me), an oleanane triterpenoid, in the treatment of diabetic kidney disease and its potential protective effects against various diseases. It focuses on the interaction between CDDO-Me and KEAP1, a repressor of NRF2 signaling, which plays a crucial role in the cytoprotective pathway mediated by the transcription factor NRF2.

The passage describes a point mutation (C151S) in the KEAP1 gene, specifically at cysteine 151, and its impact on the response to CDDO-Me in mouse embryo fibroblasts and mouse liver. In wild-type cells and mice, CDDO-Me induces the activation of NRF2 signaling, resulting in the increased expression of target genes and enzyme activity associated with the cytoprotective pathway. However, in cells and mice with the C151S mutation, the induction of NRF2 signaling by CDDO-Me is lost, suggesting that the cysteine 151 residue in KEAP1 is essential for this activation.

The passage further demonstrates that the C151S mutation affects the broader pharmacodynamic actions of CDDO-Me. In wild-type mice, CDDO-Me triggers the nuclear translocation of NRF2, leading to increased expression and activity of the Nqo1 gene, a representative target gene. In contrast, the C151S mutant mice do not exhibit this response. Moreover, when wild-type and C151S mutant mice are subjected to immune hepatitis induction, CDDO-Me provides significant protection only in the wild-type mice, highlighting the importance of KEAP1 cysteine 151 in the activation of NRF2 signaling and the subsequent cytoprotective effects of CDDO-Me.

The passage concludes that the KEAP1 cysteine 151 sensor is crucial for the induction of NRF2 signaling by CDDO-Me. It also emphasizes that the activation of "off-target" pathways by CDDO-Me was not observed, underscoring the singular importance of NRF2 in its mode of action.

Overall, this passage provides insights into the molecular mechanisms underlying the therapeutic potential of bardoxolone methyl (CDDO-Me) and highlights the significance of the KEAP1-NRF2 pathway in mediating its cytoprotective effects.