Apoptotic Blocks in Primary Non-Hodgkin B Cell Lymphomas Identified by BH3 Profiling

SIMPLE SUMMARY: The BCL2 protein is expressed in many non-Hodgkin lymphomas (NHLs) as well as associated leukemias, e.g., chronic lymphocytic leukemia (CLL). It functions as a cell survival protein that reduces that ability of a cell to undergo mitochondrial apoptosis. However, the BCL2 inhibitor venetoclax is mainly effective in CLL, despite the expression of its protein target in NHL. We hypothesized that other mechanisms are inhibiting apoptosis in NHL: defects in pro-apoptotic signaling and/or the expression of anti-apoptotic proteins other than BCL2. Our study makes use of a technique known as BH3 profiling, which is a functional assay that determines the apoptotic competency of cells on primary NHL samples. By determining how cells in NHL avoid apoptosis upon exposure to venetoclax, we can identify patients who may benefit from additional therapies and potentially improve the response of drugs currently undergoing clinical trials for NHL. ABSTRACT: To determine causes of apoptotic resistance, we analyzed 124 primary B cell NHL samples using BH3 profiling, a technique that measures the mitochondrial permeabilization upon exposure to synthetic BH3 peptides. Our cohort included samples from chronic lymphocytic leukemia (CLL), follicular lymphoma (FL), diffuse large B-cell lymphoma (DLBCL), high-grade B cell lymphoma with translocations in MYC and BCL2 (HGBL-DH), mantle cell lymphoma (MCL) and marginal zone lymphoma (MZL). While a large number of our samples displayed appropriate responses to apoptosis-inducing peptides, pro-apoptotic functional defects, implicating BAX, BAK, BIM or BID, were seen in 32.4% of high-grade NHLs (12/37) and in 3.4% of low-grade NHLs (3/87, p < 0.0001). The inhibition of single anti-apoptotic proteins induced apoptosis in only a few samples, however, the dual inhibition of BCL2 and MCL1 was effective in 83% of samples, indicating MCL1 was the most common cause of lack of response to the BCL2 inhibitor, venetoclax. We then profiled Toledo and OCI-Ly8 high-grade lymphoma cell lines to determine which drugs could reduce MCL1 expression and potentiate venetoclax responses. Doxorubicin and vincristine decreased levels of MCL1 and increased venetoclax-induced apoptosis (all p < 0.05). Overall, in primary NHLs expressing BCL2 that have no defects in pro-apoptotic signaling, a poor response to venetoclax is primarily due to the presence of MCL1, which may be overcome by combining venetoclax with doxorubicin and vincristine-based chemotherapy or with other anti-microtubule inhibitors.