Bortezomib-Resistant Mutant Proteasomes: Structural and Biochemical Evaluation with Carfilzomib and ONX 0914

• DOI: 10.1016/j.str.2014.11.019
• Publication Date: 2015-01-15T16:48:48Z
• Authors: Eva M. Huber; Wolfgang Heinemeyer; Michael Groll
• Keywords: Models, Molecular; Proteasome Endopeptidase Complex; 0303 health sciences; Molecular Structure; Protein Conformation; Crystallography, X-Ray; Boronic Acids; 3. Good health; Bortezomib; 03 medical and health sciences; X-Ray Diffraction; Structural Biology; Drug Resistance, Neoplasm; Mutagenesis; Catalytic Domain; Pyrazines; Yeasts; Mutation; Molecular Biology; Oligopeptides

Inhibition of the 20S proteasome by bortezomib (Velcade) constitutes a successfully applied therapy for blood cancer. However, emerging resistance restricts its medicinal use. For example, mutations in the proteolytically active β5-subunit of the proteasome, the main target of inhibitors, were reported to impair drug binding and thus to reduce therapeutic efficacy. Using yeast as a model system, we describe here a systematic evaluation of these mutations by cell growth analysis, proteasome inhibition assays, and X-ray crystallography. The 11 mutants examined display decreased proliferation rates, impaired proteolytic activity, and marked resistance to bortezomib as well as the α',β'-epoxyketone inhibitors carfilzomib (Kyprolis) and ONX 0914, while the second-generation compound carfilzomib was the least affected. In total, 49 proteasome X-ray structures, including structural data on proteasome-carfilzomib complexes, reveal three distinct molecular mechanisms that hamper both drug binding and natural substrate turnover to an extent that is still compatible with cell survival.