Activating KRAS , NRAS , and BRAF mutants enhance proteasome capacity and reduce endoplasmic reticulum stress in multiple myeloma.

KRAS , NRAS , and BRAF mutations which activate p44/42 mitogen-activated protein kinase (MAPK) signaling are found in half of myeloma patients and contribute to proteasome inhibitor (PI) resistance, but the underlying mechanisms are not fully understood. We established myeloma cell lines expressing wild-type (WT), constitutively active (CA) (G12V/G13D/Q61H), or dominant-negative (DN) (S17N)- KRAS and - NRAS , or BRAF -V600E. Cells expressing CA mutants showed increased proteasome maturation protein (POMP) and nuclear factor (erythroid-derived 2)-like 2 (NRF2) expression. This correlated with an increase in catalytically active proteasome subunit β (PSMB)-8, PSMB9, and PSMB10, which occurred in an ETS transcription factor-dependent manner. Proteasome chymotrypsin-like, trypsin-like, and caspase-like activities were increased, and this enhanced capacity reduced PI sensitivity, while DN- KRAS and DN- NRAS did the opposite. Pharmacologic RAF or MAPK kinase (MEK) inhibitors decreased proteasome activity, and sensitized myeloma cells to PIs. CA- KRAS , CA- NRAS , and CA- BRAF down-regulated expression of endoplasmic reticulum (ER) stress proteins, and reduced unfolded protein response activation, while DN mutations increased both. Finally, a bortezomib (BTZ)/MEK inhibitor combination showed enhanced activity in vivo specifically in CA- NRAS models. Taken together, the data support the hypothesis that activating MAPK pathway mutations enhance PI resistance by increasing proteasome capacity, and provide a rationale for targeting such patients with PI/RAF or PI/MEK inhibitor combinations. Moreover, they argue these mutations promote myeloma survival by reducing cellular stress, thereby distancing plasma cells from the apoptotic threshold, potentially explaining their high frequency in myeloma.
Competing Interests: Competing interest statement: The authors declare a competing interest. H.C.L. has provided consultancy services to Amgen, Inc., Celgene, a wholly owned subsidiary of Bristol-Myers Squibb, GlaxoSmithKline, Janssen Pharmaceutical, Sanofi-Aventis, and Takeda Pharmaceutical and has received research funding from Amgen, Inc., Celgene, a wholly owned subsidiary of Bristol-Myers Squibb, Daiichi Sankyo, GlaxoSmithKline, Janssen Pharmaceutical, and Takeda Pharmaceuticals. L.D. and N.C. are employees of Takeda Pharmaceuticals U.S.A., Inc. R.Z.O. declares laboratory research funding from BioTheryX and clinical research funding from CARsgen Therapeutics, Celgene, Exelixis, Janssen Biotech, Sanofi-Aventis, Takeda Pharmaceuticals North America, Inc. Also, R.Z.O. has served on advisory boards for Amgen, Inc., Bristol-Myers Squibb, Celgene, EcoR1 Capital LLC, Forma Therapeutics, Genzyme, GSK Biologicals, Ionis Pharmaceuticals, Inc., Janssen Biotech, Juno Therapeutics, Kite Pharma, Legend Biotech USA, Molecular Partners, Regeneron Pharmaceuticals, Inc., Sanofi-Aventis, Servier, and Takeda Pharmaceuticals North America, Inc. and as a consultant for STATinMED Research. Finally, R.Z.O. is a Founder of Asylia Therapeutics, Inc., with associated patents and an equity interest, although this technology does not bear on the current paper. The remaining authors have no conflicts of interest to declare.