A model of proteasome inhibitor resistance driven by metabolic reprogramming

Authors

• SM Gaikwad
• A Michalowski
• AS Quintanilla
• A Zingone
• S Najera
• J Simmons
• N Watson
• M Lal
• E Buehler
• S Gorjifard
• TJ Peat
• BA Mock

Abstract

Introduction: Multiple myeloma (MM) is both a rare disease and the second-most common hematological malignancy. Despite treatment regimens with proteasome inhibitors (PI) and immunomodulatory drugs, MM patients relapse. PI-resistant MM cells were found to harbor high levels of MCL1, a pro-survival protein of the BCL2 family. When these cells were treated with a small molecule capable of targeting MCL1, the mitochondrial apoptotic pathway was induced via metabolic deregulation. Methods: We established a clinically relevant Oprozomib-resistant MM subline (KMS28BM-R) from parental KMS28BM-P. Microarray, exome sequencing and druggable-genome RNAi sensitizer screening approaches were employed to characterize molecular and genetic differences between parental and resistant cells. Host cell bioenergetics were assayed using a Seahorse XFe analyzer. Immunoblotting and Q-PCRs were used to validate protein and gene expression levels. Results: KMS28BM-R cells were pan-resistant to multiple PIs, with a 10-fold increase in IC50 for oprozomib compared to parental cells. No differences were seen between KMS28BM-P and -R cells with respect to expression of proteasomal subunits, and exome sequencing confirmed that all subunits were intact. As expected, knock-down of proteasomal subunits sensitized the drug- resistant cells to apoptosis. To identify genes involved in modulating PI resistance, gene expression profiles and high throughput druggable-genome RNAi sensitizer screen data were analyzed using a Foundry data integration platform. Mcl1 was one of the top hits upregulated in resistant cells, and RNAi against Mcl1 in the resistant cell line showed reduced viability. Drug resistant cells had higher bioenergetics, as measured by oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) indicative of higher glycolytic activity. Treatment with an MCl1 inhibitor reduced cell-viability in the resistant cells, with concomitant reductions in the gene expression of glycolytic enzymes LDHA and PKM. Ongoing: High expression of MCL1 has been confirmed in additional PI-resistant cell lines, and the in vivo efficacy of MCL1 inhibitors are being evaluated preclinically. Thus, a systematic screening approach has identified MCL1 as one of the molecular modulators of PI resistance and suggests metabolic reprogramming can be used to combat PI resistance in MM.

Scientific Focus Area: Cancer Biology

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