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

Sharadha Dayalan Naidu; Takafumi Suzuki; Dina Dikovskaya; Elena V. Knatko; Maureen Higgins; Miu Sato; Miroslav Novak; José A. Villegas; Terry W. Moore; Masayuki Yamamoto; Albena Dinkova-Kostova

doi:10.1016/j.isci.2021.103703

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

Sharadha Dayalan Naidu, Takafumi Suzuki, Dina Dikovskaya, Elena V. Knatko, Maureen Higgins, Miu Sato, Miroslav Novak, José A. Villegas, Terry W. Moore, Masayuki Yamamoto, Albena Dinkova-Kostova (Lead / Corresponding author)

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Abstract

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.

Original language	English
Article number	103703
Number of pages	19
Journal	iScience
Volume	25
Issue number	1
Early online date	27 Dec 2021
DOIs	https://doi.org/10.1016/j.isci.2021.103703
Publication status	Published - 21 Jan 2022

Keywords

Biochemistry
Biological sciences
Molecular interaction

ASJC Scopus subject areas

General

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10.1016/j.isci.2021.103703Licence: CC BY

Final Published VersionFinal published version, 3.61 MBLicence: CC BY

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@article{81423a65f88b415cb2b734490f6713dc,

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

abstract = "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 - F{\"o}rster 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.",

keywords = "Biochemistry, Biological sciences, Molecular interaction",

author = "{Dayalan Naidu}, Sharadha and Takafumi Suzuki and Dina Dikovskaya and Knatko, {Elena V.} and Maureen Higgins and Miu Sato and Miroslav Novak and Villegas, {Jos{\'e} A.} and Moore, {Terry W.} and Masayuki Yamamoto and Albena Dinkova-Kostova",

note = "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.",

year = "2022",

month = jan,

day = "21",

doi = "10.1016/j.isci.2021.103703",

language = "English",

volume = "25",

journal = "iScience",

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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

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DO - 10.1016/j.isci.2021.103703

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AN - SCOPUS:85122539558

SN - 2589-0042

VL - 25

JO - iScience

JF - iScience

IS - 1

M1 - 103703

ER -

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

Abstract

Keywords

ASJC Scopus subject areas

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The Role of the Keap1/Nrf2 Pathway in Tumour Metabolic Adaptation (Joint with University of Cambridge and University College London)

The Spatiotemporal Regulation of the Keap1/Nrf2 Pathway (Joint with University College London)

Cite this

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

Abstract

Keywords

ASJC Scopus subject areas

Access to Document

Other files and links

Fingerprint

Projects

The Role of the Keap1/Nrf2 Pathway in Tumour Metabolic Adaptation (Joint with University of Cambridge and University College London)

The Spatiotemporal Regulation of the Keap1/Nrf2 Pathway (Joint with University College London)

Cite this