Bv6, an antagonist of inhibitor of apoptosis proteins, sensitizes resistant mantle cell lymphoma cells to venetoclax

Introduction: Although Bruton’s tyrosine kinase inhibitors (BTKi) and CAR T-cell therapy have achieved remarkable efficacy in the treatment of mantle cell lymphoma (MCL), it remains incurable due to the inevitable development of therapeutic resistance. The BCL-2 inhibitor venetoclax is effective in many relapsed MCL patients, including those who progressed on BTKi, but subsequent relapse to venetoclax yields poor clinical outcomes. Hence, novel therapeutics and combination therapies are required to tackle the clinical challenges. Here, we investigated BV6, an antagonist of inhibitor of apoptosis proteins (IAPs), alone and in combination with venetoclax in multiple therapy-resistant MCL models. Methods: We performed single-cell RNA-seq to assess gene expression in ibrutinib-sensitive, ibrutinib-resistant, and dual ibrutinib-/CAR T-resistant MCL patients. Cell lines with acquired resistance to ibrutinib or venetoclax, as well as primary tumor cells from patients who relapsed after multiple therapies, were used as models of MCL resistance. CellTiter-Glo® luminescent assay was used to measure cell viability; annexin V/PI staining was used to measure the induction of apoptosis; and western blotting was used to assess IAP and apoptosis signaling protein expression across MCL cell lines after drug treatment. Results: The single-cell RNA-seq analysis demonstrated that IAP gene expression was not influenced by patient response to ibrutinib and CAR T-cell therapy, and western blotting showed that IAP proteins were broadly expressed across all MCL cell lines. To determine the on-target effects of BV6, we treated JeKo-1, JeKo-IBN-R, Mino, and Mino-VEN-R cells with BV6 and examined the expression of IAPs. BV6 treatment for 4h markedly reduced c-IAP1 and c-IAP2 expression, and moderately reduced XIAP expression. BV6 inhibited cell viability in both ibrutinib- and venetoclax-resistant cell lines, and BV6 treatment for 24 h dose-dependently induced apoptosis in MCL cells, including not only cell lines with acquired resistance but also patient primary cells, independent of their resistance to ibrutinib, venetoclax, and CAR T-cell therapy. We observed a striking and significant synergy in the cell viability assay when ibrutinib- or venetoclax-resistant cells were treated with BV6 combined with venetoclax (combination index < 1), and a dramatic enhancement of apoptosis (p < 0.001). The combination eradicated c-IAP1 and c-IAP2 expression, while XIAP was synergistically diminished. This was accompanied by strong caspase-3 activation and PARP cleavage. This combination therapy is currently under evaluation in venetoclax-resistant mouse model in vivo. Conclusions: Our findings indicate that a combination of IAP antagonists with apoptosis inducers can be exploited as a rational strategy to overcome therapeutic resistance in MCL. This study was supported by philanthropic funds from the Kimmel Research fund and various donor funds. Keywords: Aggressive B-cell non-Hodgkin lymphoma, Combination Therapies, Molecular Targeted Therapies No conflicts of interests pertinent to the abstract.