Discovery and molecular basis of subtype-selective cyclophilin inhibitors

Peterson, Alexander A.; Rangwala, Aziz M.; Thakur, Manish K.; Ward, Patrick S.; Hung, Christie; Outhwaite, Ian R.; Chan, Alix I.; Usanov, Dmitry L.; Mootha, Vamsi K.; Seeliger, Markus A.; Liu, David R.

doi:10.1038/s41589-022-01116-1

Discovery and molecular basis of subtype-selective cyclophilin inhibitors

Journal Article · 26 September 2022 · Nature Chemical Biology

DOI:https://doi.org/10.1038/s41589-022-01116-1· OSTI ID:2425461

; ; Thakur, Manish K.; ; ; ; ; ; Mootha, Vamsi K.; ;

Abstract

Although cyclophilins are attractive targets for probing biology and therapeutic intervention, no subtype-selective cyclophilin inhibitors have been described. We discovered novel cyclophilin inhibitors from the in vitro selection of a DNA-templated library of 256,000 drug-like macrocycles for cyclophilin D (CypD) affinity. Iterated macrocycle engineering guided by ten X-ray co-crystal structures yielded potent and selective inhibitors (half maximal inhibitory concentration (IC₅₀) = 10 nM) that bind the active site of CypD and also make novel interactions with non-conserved residues in the S2 pocket, an adjacent exo-site. The resulting macrocycles inhibit CypD activity with 21- to >10,000-fold selectivity over other cyclophilins and inhibit mitochondrial permeability transition pore opening in isolated mitochondria. We further exploited S2 pocket interactions to develop the first cyclophilin E (CypE)-selective inhibitor, which forms a reversible covalent bond with a CypE S2 pocket lysine, and exhibits 30- to >4,000-fold selectivity over other cyclophilins. These findings reveal a strategy to generate isoform-selective small-molecule cyclophilin modulators, advancing their suitability as targets for biological investigation and therapeutic development.

Research Organization:: Brookhaven National Laboratory (BNL), Upton, NY (United States). National Synchrotron Light Source II (NSLS-II)

Sponsoring Organization:: USDOE Office of Science (SC)

DOE Contract Number:: SC0012704

OSTI ID:: 2425461

Journal Information:: Nature Chemical Biology, Journal Name: Nature Chemical Biology Journal Issue: 11 Vol. 18; ISSN 1552-4450

Publisher:: Nature Publishing Group

Country of Publication:: United States

Language:: English

References (59)

PRODRG : a tool for high-throughput crystallography of protein–ligand complexes Schüttelkopf, Alexander W.; van Aalten, Daan M. F. Acta Crystallographica Section D Biological Crystallography, Vol. 60, Issue 8 https://doi.org/10.1107/S0907444904011679	journal	July 2004
Selective Inhibition of the Mitochondrial Permeability Transition Pore Protects against Neurodegeneration in Experimental Multiple Sclerosis Warne, Justin; Pryce, Gareth; Hill, Julia M. Journal of Biological Chemistry, Vol. 291, Issue 9 https://doi.org/10.1074/jbc.M115.700385	journal	February 2016
Mitochondrial permeability transition pore is a potential drug target for neurodegeneration Rao, Valasani Koteswara; Carlson, Emily A.; Yan, Shirley Shidu Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease, Vol. 1842, Issue 8 https://doi.org/10.1016/j.bbadis.2013.09.003	journal	August 2014
GNX-4728, a novel small molecule drug inhibitor of mitochondrial permeability transition, is therapeutic in a mouse model of amyotrophic lateral sclerosis Martin, Lee J.; Fancelli, Daniele; Wong, Margaret Frontiers in Cellular Neuroscience, Vol. 8 https://doi.org/10.3389/fncel.2014.00433	journal	December 2014
Features and development of Coot Emsley, P.; Lohkamp, B.; Scott, W. G. Acta Crystallographica Section D Biological Crystallography, Vol. 66, Issue 4 https://doi.org/10.1107/S0907444910007493	journal	March 2010
Mitochondrial Permeability Uncouples Elevated Autophagy and Lifespan Extension Zhou, Ben; Kreuzer, Johannes; Kumsta, Caroline Cell, Vol. 177, Issue 2 https://doi.org/10.1016/j.cell.2019.02.013	journal	April 2019
Mitochondrial permeability transition pore: a promising target for the treatment of Parkinson’s disease Rasheed, Md Zeeshan; Tabassum, Heena; Parvez, Suhel Protoplasma, Vol. 254, Issue 1 https://doi.org/10.1007/s00709-015-0930-2	journal	January 2016
Evaluation of NV556, a Novel Cyclophilin Inhibitor, as a Potential Antifibrotic Compound for Liver Fibrosis Simón Serrano, Sonia; Grönberg, Alvar; Longato, Lisa Cells, Vol. 8, Issue 11 https://doi.org/10.3390/cells8111409	journal	November 2019
Guidelines on experimental methods to assess mitochondrial dysfunction in cellular models of neurodegenerative diseases Connolly, Niamh M. C.; Theurey, Pierre; Adam-Vizi, Vera Cell Death & Differentiation, Vol. 25, Issue 3 https://doi.org/10.1038/s41418-017-0020-4	journal	December 2017
Phaser crystallographic software McCoy, Airlie J.; Grosse-Kunstleve, Ralf W.; Adams, Paul D. Journal of Applied Crystallography, Vol. 40, Issue 4 https://doi.org/10.1107/S0021889807021206	journal	July 2007
Probing Mitochondrial Permeability Transition Pore Activity in Nucleated Cells and Platelets by High-Throughput Screening Assays Suggests Involvement of Protein Phosphatase 2B in Mitochondrial Dynamics Chen, Mao Xiang; Ward, Emma; Caivano, Matilde ASSAY and Drug Development Technologies, Vol. 16, Issue 8 https://doi.org/10.1089/adt.2018.872	journal	December 2018
Structural and Biochemical Characterization of the Human Cyclophilin Family of Peptidyl-Prolyl Isomerases Davis, Tara L.; Walker, John R.; Campagna-Slater, Valérie PLoS Biology, Vol. 8, Issue 7 https://doi.org/10.1371/journal.pbio.1000439	journal	July 2010
Inhibition of Mcl-1 through covalent modification of a noncatalytic lysine side chain Akçay, Gizem; Belmonte, Matthew A.; Aquila, Brian Nature Chemical Biology, Vol. 12, Issue 11 https://doi.org/10.1038/nchembio.2174	journal	September 2016
Regulation of the Expression of Cytoplasmic Polyadenylation Element Binding Proteins for the Treatment of Cancer Chen, Yun; Tsai, Ya-Hui; Tseng, Sheng-Hong Anticancer Research, Vol. 36, Issue 11 https://doi.org/10.21873/anticanres.11150	journal	November 2016
Second-generation DNA-templated macrocycle libraries for the discovery of bioactive small molecules Usanov, Dmitry L.; Chan, Alix I.; Maianti, Juan Pablo Nature Chemistry, Vol. 10, Issue 7 https://doi.org/10.1038/s41557-018-0033-8	journal	April 2018
Cyclophilin D is a component of mitochondrial permeability transition and mediates neuronal cell death after focal cerebral ischemia Schinzel, Anna C.; Takeuchi, Osamu; Huang, Zhihong Proceedings of the National Academy of Sciences, Vol. 102, Issue 34 https://doi.org/10.1073/pnas.0505294102	journal	August 2005
Discovery of novel Cyclophilin D inhibitors starting from three dimensional fragments with millimolar potencies Grädler, Ulrich; Schwarz, Daniel; Blaesse, Michael Bioorganic & Medicinal Chemistry Letters, Vol. 29, Issue 23 https://doi.org/10.1016/j.bmcl.2019.126717	journal	December 2019
Crystal structure of human cyclophilin D in complex with its inhibitor, cyclosporin A at 0.96‐Å resolution Kajitani, Kenji; Fujihashi, Masahiro; Kobayashi, Yukiko Proteins: Structure, Function, and Bioinformatics, Vol. 70, Issue 4 https://doi.org/10.1002/prot.21855	journal	December 2007
Molecular nature and regulation of the mitochondrial permeability transition pore(s), drug target(s) in cardioprotection Carraro, Michela; Carrer, Andrea; Urbani, Andrea Journal of Molecular and Cellular Cardiology, Vol. 144 https://doi.org/10.1016/j.yjmcc.2020.05.014	journal	July 2020
Cyclophilin D-dependent mitochondrial permeability transition regulates some necrotic but not apoptotic cell death Nakagawa, Takashi; Shimizu, Shigeomi; Watanabe, Tetsuya Nature, Vol. 434, Issue 7033 https://doi.org/10.1038/nature03317	journal	March 2005
F‐ATP synthase and the permeability transition pore: fewer doubts, more certainties Carraro, Michela; Checchetto, Vanessa; Szabó, Ildikó FEBS Letters, Vol. 593, Issue 13 https://doi.org/10.1002/1873-3468.13485	journal	June 2019
Mitochondria-Localized Self-Reporting Small-Molecule-Decorated Theranostic Agents for Cancer-Organelle Transporting and Imaging Zhou, Lihua; Wu, Yayun; Luo, Yuan ACS Applied Bio Materials, Vol. 2, Issue 11 https://doi.org/10.1021/acsabm.9b00811	journal	October 2019
Loss of cyclophilin D reveals a critical role for mitochondrial permeability transition in cell death Baines, Christopher P.; Kaiser, Robert A.; Purcell, Nicole H. Nature, Vol. 434, Issue 7033 https://doi.org/10.1038/nature03434	journal	March 2005
Use of a Real-Time Fluorescence Monitoring System for High-Throughput Screening for Prolyl Isomerase Inhibitors Mori, Tadashi; Itami, Seima; Yanagi, Tomotaka SLAS Discovery, Vol. 14, Issue 4 https://doi.org/10.1177/1087057109333979	journal	April 2009
A fluorescence polarization-based assay for peptidyl prolyl cis/trans isomerase cyclophilin A Liu, Yaya; Jiang, Jianjun; Richardson, Paul L. Analytical Biochemistry, Vol. 356, Issue 1 https://doi.org/10.1016/j.ab.2006.04.040	journal	September 2006
Mitochondrial Permeability Transition: New Findings and Persisting Uncertainties Izzo, Valentina; Bravo-San Pedro, José Manuel; Sica, Valentina Trends in Cell Biology, Vol. 26, Issue 9 https://doi.org/10.1016/j.tcb.2016.04.006	journal	September 2016
Persistence of the permeability transition pore in human mitochondria devoid of an assembled ATP synthase Carroll, Joe; He, Jiuya; Ding, Shujing Proceedings of the National Academy of Sciences, Vol. 116, Issue 26 https://doi.org/10.1073/pnas.1904005116	journal	June 2019
Novel cyclophilin D inhibitors derived from quinoxaline exhibit highly inhibitory activity against rat mitochondrial swelling and Ca2+ uptake/release Guo, Hong-xia; Wang, Feng; Yu, Kun-qian Acta Pharmacologica Sinica, Vol. 26, Issue 10 https://doi.org/10.1111/j.1745-7254.2005.00189.x	journal	October 2005
A 20/20 view of ANT function in mitochondrial biology and necrotic cell death Bround, Michael J.; Bers, Donald M.; Molkentin, Jeffery D. Journal of Molecular and Cellular Cardiology, Vol. 144 https://doi.org/10.1016/j.yjmcc.2020.05.012	journal	July 2020
Mitochondrial Membrane Permeabilization in Cell Death Kroemer, Guido; Galluzzi, Lorenzo; Brenner, Catherine Physiological Reviews, Vol. 87, Issue 1 https://doi.org/10.1152/physrev.00013.2006	journal	January 2007
Pgp and MRP Activities Using Calcein-AM Are Prognostic Factors in Adult Acute Myeloid Leukemia Patients Legrand, Ollivier; Simonin, Ghislaine; Perrot, Jean-Yves Blood, Vol. 91, Issue 12 https://doi.org/10.1182/blood.V91.12.4480	journal	June 1998
Multidomain Peptidyl Prolyl cis/trans Isomerases Schiene-Fischer, Cordelia Biochimica et Biophysica Acta (BBA) - General Subjects, Vol. 1850, Issue 10 https://doi.org/10.1016/j.bbagen.2014.11.012	journal	October 2015
Crystal engineering yields crystals of cyclophilin D diffracting to 1.7 Å resolution Schlatter, Daniel; Thoma, Ralf; Küng, Erich Acta Crystallographica Section D Biological Crystallography, Vol. 61, Issue 5 https://doi.org/10.1107/S0907444905003070	journal	April 2005
Novel quinazoline-urea analogues as modulators for Aβ-induced mitochondrial dysfunction: Design, synthesis, and molecular docking study Elkamhawy, Ahmed; Lee, Jiyoun; Park, Beoung-Geon European Journal of Medicinal Chemistry, Vol. 84 https://doi.org/10.1016/j.ejmech.2014.07.027	journal	September 2014
Prolyl cis/trans isomerase signalling pathways in cancer Theuerkorn, Martin; Fischer, Gunter; Schiene-Fischer, Cordelia Current Opinion in Pharmacology, Vol. 11, Issue 4 https://doi.org/10.1016/j.coph.2011.03.007	journal	August 2011
Investigating the Mitochondrial Permeability Transition Pore in Disease Phenotypes and Drug Screening Bhosale, Gauri; Duchen, Michael R. Current Protocols in Pharmacology, Vol. 85, Issue 1 https://doi.org/10.1002/cpph.59	journal	May 2019
Roles of Prolyl Isomerases in RNA-Mediated Gene Expression Thapar, Roopa Biomolecules, Vol. 5, Issue 2 https://doi.org/10.3390/biom5020974	journal	May 2015
Fragment-based discovery of a new family of non-peptidic small-molecule cyclophilin inhibitors with potent antiviral activities Ahmed-Belkacem, Abdelhakim; Colliandre, Lionel; Ahnou, Nazim Nature Communications, Vol. 7, Issue 1 https://doi.org/10.1038/ncomms12777	journal	September 2016
Cyclophilins as Modulators of Viral Replication Frausto, Stephen; Lee, Emily; Tang, Hengli Viruses, Vol. 5, Issue 7 https://doi.org/10.3390/v5071684	journal	July 2013
Cyclophilin D as a potential target for antioxidants in neurodegeneration: the X-ALD case López-Erauskin, Jone; Ferrer, Isidre; Galea, Elena bchm, Vol. 394, Issue 5 https://doi.org/10.1515/hsz-2012-0323	journal	March 2013
Discovery of Potent Cyclophilin Inhibitors Based on the Structural Simplification of Sanglifehrin A Steadman, Victoria A.; Pettit, Simon B.; Poullennec, Karine G. Journal of Medicinal Chemistry, Vol. 60, Issue 3 https://doi.org/10.1021/acs.jmedchem.6b01329	journal	January 2017
Sanglifehrin A Acts as a Potent Inhibitor of the Mitochondrial Permeability Transition and Reperfusion Injury of the Heart by Binding to Cyclophilin-D at a Different Site from Cyclosporin A Clarke, Samantha J.; McStay, Gavin P.; Halestrap, Andrew P. Journal of Biological Chemistry, Vol. 277, Issue 38 https://doi.org/10.1074/jbc.M202191200	journal	September 2002
Ca2+ ionophores are not suitable for inducing mPTP opening in murine isolated adult cardiac myocytes Panel, Mathieu; Ghaleh, Bijan; Morin, Didier Scientific Reports, Vol. 7, Issue 1 https://doi.org/10.1038/s41598-017-04618-4	journal	June 2017
Discovery of non-peptidic small molecule inhibitors of cyclophilin D as neuroprotective agents in Aβ-induced mitochondrial dysfunction Park, Insun; Londhe, Ashwini M.; Lim, Ji Woong Journal of Computer-Aided Molecular Design, Vol. 31, Issue 10 https://doi.org/10.1007/s10822-017-0067-9	journal	September 2017
Isoform-Specific Inhibition of Cyclophilins Daum, Sebastian; Schumann, Michael; Mathea, Sebastian Biochemistry, Vol. 48, Issue 26 https://doi.org/10.1021/bi9007287	journal	June 2009
Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix Liebschner, Dorothee; Afonine, Pavel V.; Baker, Matthew L. Acta Crystallographica Section D Structural Biology, Vol. 75, Issue 10 https://doi.org/10.1107/S2059798319011471	journal	October 2019
Implementation and evaluation of a docking-rescoring method using molecular footprint comparisons Balius, Trent E.; Mukherjee, Sudipto; Rizzo, Robert C. Journal of Computational Chemistry, Vol. 32, Issue 10 https://doi.org/10.1002/jcc.21814	journal	May 2011
A surface plasmon resonance-based assay for small molecule inhibitors of human cyclophilin A Wear, Martin A.; Patterson, Alan; Malone, Kirk Analytical Biochemistry, Vol. 345, Issue 2 https://doi.org/10.1016/j.ab.2005.06.037	journal	October 2005
A computationally designed binding mode flip leads to a novel class of potent tri-vector cyclophilin inhibitors De Simone, Alessio; Georgiou, Charis; Ioannidis, Harris Chemical Science, Vol. 10, Issue 2 https://doi.org/10.1039/C8SC03831G	journal	January 2019
Peptidyl-Proline Isomerases (PPIases): Targets for Natural Products and Natural Product-Inspired Compounds: Miniperspective Dunyak, Bryan M.; Gestwicki, Jason E. Journal of Medicinal Chemistry, Vol. 59, Issue 21 https://doi.org/10.1021/acs.jmedchem.6b00411	journal	July 2016
Mitochondrial permeability transition increases reactive oxygen species production and induces DNA fragmentation in human spermatozoa Treulen, F.; Uribe, P.; Boguen, R. Human Reproduction, Vol. 30, Issue 4 https://doi.org/10.1093/humrep/dev015	journal	February 2015
Mitochondrial Proton Leak Regulated by Cyclophilin D Elevates Insulin Secretion in Islets at Nonstimulatory Glucose Levels Taddeo, Evan P.; Alsabeeh, Nour; Baghdasarian, Siyouneh Diabetes, Vol. 69, Issue 2 https://doi.org/10.2337/db19-0379	journal	November 2019
Molecular identity of the mitochondrial permeability transition pore and its role in ischemia-reperfusion injury Morciano, Giampaolo; Giorgi, Carlotta; Bonora, Massimo Journal of Molecular and Cellular Cardiology, Vol. 78 https://doi.org/10.1016/j.yjmcc.2014.08.015	journal	January 2015
Identification of a Small Molecule Cyclophilin D Inhibitor for Rescuing Aβ-Mediated Mitochondrial Dysfunction Valasani, Koteswara Rao; Sun, Qinru; Fang, Du ACS Medicinal Chemistry Letters, Vol. 7, Issue 3 https://doi.org/10.1021/acsmedchemlett.5b00451	journal	January 2016
UCSF Chimera?A visualization system for exploratory research and analysis Pettersen, Eric F.; Goddard, Thomas D.; Huang, Conrad C. Journal of Computational Chemistry, Vol. 25, Issue 13 https://doi.org/10.1002/jcc.20084	journal	January 2004
Mitochondrial Permeability Transition: A Molecular Lesion with Multiple Drug Targets Briston, Thomas; Selwood, David L.; Szabadkai, Gyorgy Trends in Pharmacological Sciences, Vol. 40, Issue 1 https://doi.org/10.1016/j.tips.2018.11.004	journal	January 2019
Mitochondria and aging: A role for the mitochondrial transition pore? Panel, Mathieu; Ghaleh, Bijan; Morin, Didier Aging Cell, Vol. 17, Issue 4 https://doi.org/10.1111/acel.12793	journal	June 2018
Mitochondrial permeability transition pore induces mitochondria injury in Huntington disease Quintanilla, Rodrigo A.; Jin, Youngnam N.; von Bernhardi, Rommy Molecular Neurodegeneration, Vol. 8, Issue 1 https://doi.org/10.1186/1750-1326-8-45	journal	January 2013
Cyclosporin A Increases Mitochondrial Buffering of Calcium: An Additional Mechanism in Delaying Mitochondrial Permeability Transition Pore Opening Mishra, Jyotsna; Davani, Ariea J.; Natarajan, Gayathri K. Cells, Vol. 8, Issue 9 https://doi.org/10.3390/cells8091052	journal	September 2019

Similar Records

New cyclophilin D inhibitor rescues mitochondrial and cognitive function in Alzheimer’s disease

Journal Article · 2023 · Brain · OSTI ID:2582312

The cyclophilin D/Drp1 axis regulates mitochondrial fission contributing to oxidative stress-induced mitochondrial dysfunctions in SH-SY5Y cells

Journal Article · 2017 · Biochemical and Biophysical Research Communications · OSTI ID:22696850

Cyclophilin 1 (Cyp1) mutation ameliorates oxidative stress-induced defects in a Drosophila DJ-1 null mutant

Journal Article · 2018 · Biochemical and Biophysical Research Communications · OSTI ID:23134107

Related Subjects

Biochemistry & Molecular Biology

Discovery and molecular basis of subtype-selective cyclophilin inhibitors

Citation Formats

References (59)

Similar Records

Related Subjects