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Published February 7, 2022 | Published + Supplemental Material
Journal Article Open

The p97 Inhibitor UPCDC-30245 Blocks Endo-Lysosomal Degradation

Abstract

The diverse modes of action of small molecule inhibitors provide versatile tools to investigate basic biology and develop therapeutics. However, it remains a challenging task to evaluate their exact mechanisms of action. We identified two classes of inhibitors for the p97 ATPase: ATP competitive and allosteric. We showed that the allosteric p97 inhibitor, UPCDC-30245, does not affect two well-known cellular functions of p97, endoplasmic-reticulum-associated protein degradation and the unfolded protein response pathway; instead, it strongly increases the lipidated form of microtubule-associated proteins 1A/1B light chain 3B (LC3-II), suggesting an alteration of autophagic pathways. To evaluate the molecular mechanism, we performed proteomic analysis of UPCDC-30245 treated cells. Our results revealed that UPCDC-30245 blocks endo-lysosomal degradation by inhibiting the formation of early endosome and reducing the acidity of the lysosome, an effect not observed with the potent p97 inhibitor CB-5083. This unique effect allows us to demonstrate UPCDC-30245 exhibits antiviral effects against coronavirus by blocking viral entry.

Additional Information

© 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). Received: 2 January 2022. Accepted: 27 January 2022. Published: 7 February 2022. We thank the Merkin Institute for Translational Research at Caltech for supporting the antiviral research. This work was supported in part with funds from the National Institute of Neurological Disorders and Stroke, R01NS100815 and R01NS102279. Author Contributions: F.W. wrote the manuscript. F.W. and S.L. performed all cellular assays and proteomics studies. K.-W.C. assisted with virus experiments. W.M.R. edited the manuscript and provided experimental suggestions. T.-F.C. supervised the project and designed the research. All authors have read and agreed to the published version of the manuscript. Data Availability Statement: All relevant data generated during this study are included in the article and the supplementary Information. The mass spectrometry raw data are deposited to the ProteomeXchance Consortium (https://www.ebi.ac.uk/pride/, accessed on 28, December, 2021) via the PRIDE repository with the dataset identifier PXD025822 and 10.6019/PXD025822". Additional raw data generated during the current study and relevant information are available from the corresponding authors upon request. The data are not publicly available due to the larger size and complexity. Supplementary Materials: The following are available online at www.mdpi.com/article/10.3390/ph15020204/s1, Figure S1: (A) Functional enrichment analysis on proteins affected by UPCDC-30245. (B) The log2 fold change of LDLR, APOB and NAGLU from previous and new data set, Figure S2: UPCDC-30245 rapidly reduced the LysoTracker staining in H1299 cells. Table S1: qPCR probes used in this study. Institutional Review Board Statement: Not applicable. Informed Consent Statement: Not applicable. The authors declare no conflict of interest.

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Published - pharmaceuticals-15-00204.pdf

Supplemental Material - pharmaceuticals-15-00204-s001.zip

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Additional details

Created:
October 9, 2023
Modified:
December 22, 2023