The isoquinoline PRL-295 increases the thermostability of Keap1 and disrupts its interaction with Nrf2

TY - JOUR

T1 - The isoquinoline PRL-295 increases the thermostability of Keap1 and disrupts its interaction with Nrf2

AU - Dayalan Naidu, Sharadha

AU - Suzuki, Takafumi

AU - Dikovskaya, Dina

AU - Knatko, Elena V.

AU - Higgins, Maureen

AU - Sato, Miu

AU - Novak, Miroslav

AU - Villegas, José A.

AU - Moore, Terry W.

AU - Yamamoto, Masayuki

AU - Dinkova-Kostova, Albena

N1 - Funding: We thank Phillip Lazzara for synthesizing PRL-295, the Biotechnology and Biological Sciences Research Council (BB/L01923X/1 and BB/T508111/1), Tenovus Scotland (T17/14), MEXT/JSPS KAKENHI (19H05649, 19K07340 and 17KK0183), Cancer Research UK (C20953/A18644), and Reata Pharmaceuticals for financial support.

PY - 2022/1/21

Y1 - 2022/1/21

N2 - Transcription factor Nrf2 and its negative regulator Keap1 orchestrate a cytoprotective response against oxidative, metabolic, and inflammatory stress. Keap1 is a drug target, with several small molecules in drug development. Here, we show that the isoquinoline PRL-295 increased Keap1 thermostability in lysates from cells expressing fluorescently-tagged Keap1. The thermostability of endogenous Keap1 also increased in intact cells and murine liver following PRL-295 treatment. Fluorescence Lifetime Imaging - Förster Resonance Energy Transfer (FLIM-FRET) experiments in cells co-expressing sfGFP-Nrf2 and Keap1-mCherry further showed that PRL-295 prolonged the donor fluorescence lifetime, indicating disruption of the Keap1-Nrf2 protein complex. Orally-administered PRL-295 to mice activated the Nrf2-transcriptional target NAD(P)H:quinone oxidoreductase 1 (NQO1) in liver and decreased the levels of plasma alanine aminotransferase and aspartate aminotransferase upon acetaminophen-induced hepatic injury. Thus, PRL-295 engages the Keap1 protein target in cells and in vivo, disrupting its interaction with Nrf2, leading to activation of Nrf2-dependent transcription and hepatocellular protection.

AB - Transcription factor Nrf2 and its negative regulator Keap1 orchestrate a cytoprotective response against oxidative, metabolic, and inflammatory stress. Keap1 is a drug target, with several small molecules in drug development. Here, we show that the isoquinoline PRL-295 increased Keap1 thermostability in lysates from cells expressing fluorescently-tagged Keap1. The thermostability of endogenous Keap1 also increased in intact cells and murine liver following PRL-295 treatment. Fluorescence Lifetime Imaging - Förster Resonance Energy Transfer (FLIM-FRET) experiments in cells co-expressing sfGFP-Nrf2 and Keap1-mCherry further showed that PRL-295 prolonged the donor fluorescence lifetime, indicating disruption of the Keap1-Nrf2 protein complex. Orally-administered PRL-295 to mice activated the Nrf2-transcriptional target NAD(P)H:quinone oxidoreductase 1 (NQO1) in liver and decreased the levels of plasma alanine aminotransferase and aspartate aminotransferase upon acetaminophen-induced hepatic injury. Thus, PRL-295 engages the Keap1 protein target in cells and in vivo, disrupting its interaction with Nrf2, leading to activation of Nrf2-dependent transcription and hepatocellular protection.

KW - Biochemistry

KW - Biological sciences

KW - Molecular interaction

UR - https://www.scopus.com/pages/publications/85122539558

U2 - 10.1016/j.isci.2021.103703

DO - 10.1016/j.isci.2021.103703

M3 - Article

C2 - 35036882

AN - SCOPUS:85122539558

SN - 2589-0042

VL - 25

JO - iScience

JF - iScience

IS - 1

M1 - 103703

ER -