Pharmaceutical-Grade Rigosertib Is a Microtubule-Destabilizing Agent

Marco Jost; Yuwen Chen; Luke A Gilbert; Max A Horlbeck; Lenno Krenning; Grégory Menchon; Ankit Rai; Min Y Cho; Jacob J Stern; Andrea E Prota; Martin Kampmann; Anna Akhmanova; Michel O Steinmetz; Marvin E Tanenbaum; Jonathan S Weissman

doi:10.1016/j.molcel.2020.06.008

Pharmaceutical-Grade Rigosertib Is a Microtubule-Destabilizing Agent

Mol Cell. 2020 Jul 2;79(1):191-198.e3. doi: 10.1016/j.molcel.2020.06.008.

Authors

Affiliations

¹ Department of Cellular & Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, San Francisco, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA.
² Department of Cellular & Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, San Francisco, San Francisco, CA 94158, USA.
³ Department of Cellular & Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, San Francisco, San Francisco, CA 94158, USA; Helen Diller Family Comprehensive Cancer Center, Department of Urology, University of California, San Francisco, San Francisco, CA 94158, USA.
⁴ Hubrecht Institute - KNAW and University Medical Center Utrecht, 3584CT Utrecht, the Netherlands.
⁵ Laboratory of Biomolecular Research, Division of Biology and Chemistry, Paul Scherrer Institut, 5232 Villigen PSI, Switzerland.
⁶ Cell Biology, Department of Biology, Faculty of Science, Utrecht University, 3548CH Utrecht, the Netherlands.
⁷ Department of Cellular & Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, San Francisco, San Francisco, CA 94158, USA; Institute for Neurodegenerative Diseases and Department of Biochemistry & Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA; Chan-Zuckerberg Biohub, San Francisco, CA 94158, USA.
⁸ Laboratory of Biomolecular Research, Division of Biology and Chemistry, Paul Scherrer Institut, 5232 Villigen PSI, Switzerland; Biozentrum, University of Basel, 4056 Basel, Switzerland.
⁹ Hubrecht Institute - KNAW and University Medical Center Utrecht, 3584CT Utrecht, the Netherlands. Electronic address: m.tanenbaum@hubrecht.eu.
¹⁰ Department of Cellular & Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, San Francisco, San Francisco, CA 94158, USA. Electronic address: jonathan.weissman@ucsf.edu.

Abstract

We recently used CRISPRi/a-based chemical-genetic screens and cell biological, biochemical, and structural assays to determine that rigosertib, an anti-cancer agent in phase III clinical trials, kills cancer cells by destabilizing microtubules. Reddy and co-workers (Baker et al., 2020, this issue of Molecular Cell) suggest that a contaminating degradation product in commercial formulations of rigosertib is responsible for the microtubule-destabilizing activity. Here, we demonstrate that cells treated with pharmaceutical-grade rigosertib (>99.9% purity) or commercially obtained rigosertib have qualitatively indistinguishable phenotypes across multiple assays. The two formulations have indistinguishable chemical-genetic interactions with genes that modulate microtubule stability, both destabilize microtubules in cells and in vitro, and expression of a rationally designed tubulin mutant with a mutation in the rigosertib binding site (L240F TUBB) allows cells to proliferate in the presence of either formulation. Importantly, the specificity of the L240F TUBB mutant for microtubule-destabilizing agents has been confirmed independently. Thus, rigosertib kills cancer cells by destabilizing microtubules, in agreement with our original findings.

Keywords: CRISPRa; CRISPRi; chemical genetics; drug mechanism of action; drug target identification; microtubules; rigosertib.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Antineoplastic Agents / pharmacology*
Cell Proliferation*
Cells, Cultured
Crystallography, X-Ray
Drug Contamination
Glycine / analogs & derivatives*
Glycine / pharmacology
Humans
Microtubules / drug effects*
Mutation
Neoplasms / drug therapy
Neoplasms / metabolism
Neoplasms / pathology*
Pharmaceutical Preparations / chemistry
Pharmaceutical Preparations / metabolism*
Protein Conformation
Sulfones / pharmacology*
Tubulin / chemistry
Tubulin / genetics
Tubulin / metabolism*

Substances

Antineoplastic Agents
Pharmaceutical Preparations
Sulfones
Tubulin
ON 01910
Glycine

Abstract

Publication types

MeSH terms

Substances

Grants and funding