Biomarker and Ex Vivo Sensitivity Analysis of Venetoclax (ABT-199/GDC-0199) in Combination with Bortezomib and Dexamethasone in Relapsed/Refractory Multiple Myeloma

Introduction: Multiple myeloma (MM) tumor cell survival is largely dependent on a complex network of interactions between pro-survival (BCL-2, BCL-XL, MCL-1) and pro-death (BIM, NOXA, BID, BAD, BAK, BAX) BCL-2 family members. Venetoclax (VEN) is a potent, selective, orally bioavailable small-molecule BCL-2 inhibitor. Bortezomib (BTZ) is a proteasome inhibitor that can impede MCL-1 activity by increasing the MCL-1 antagonist NOXA. The addition of VEN has been shown to enhance the activity of BTZ in MM cell line and xenograft models. Chromosomal abnormalities (immunoglobulin heavy chain translocations, amplification, deletions) can also influence sensitivity of MM cells to treatment. Results presented herein describe retrospective biomarker analysis in a subset of dose-escalation patients (pts) from an ongoing phase 1b study of VEN in combination with BTZ and dexamethasone in relapsed/refractory (R/R) MM.

Methods: Exploratory objectives included evaluation of chromosomal abnormalities, BCL-2 family member gene expression levels, and ex vivo sensitivity of pt MM tumor cells to VEN and/or BTZ. Interphase FISH analysis was performed on CD138-positively selected bone marrow mononuclear cells (BMMC) using probes for t(11;14), t(4;14), deletion 17p (17p–), deletion 13q (13q–), and amplification of chromosomes 5, 9, or 15 (denoted as hyperdiploid [HY]). Absolute quantitation of BCL-2 family members (BCL-2, BCL-XL, MCL-1, BCL-2A1, BIM, BID, BAD, BAX, and NOXA) was performed on CD138-selected BMMCs collected at baseline using TaqMan probe-based droplet digital polymerase chain reaction. Ex vivo sensitivity of BMMCs collected at baseline to increasing doses of VEN and/or BTZ was determined by flow cytometry using a combined analysis for the loss of CD138 surface expression and the alteration of cellular morphology (lower FCS). Correlations between biomarkers and median best percentage change in M-protein were examined by Spearman test. Correlations between biomarkers and median time on study (mToS) were examined by log-rank and Wilcoxon tests for binary biomarkers, and by risk ratio from a Cox proportional hazard model for continuous biomarkers. No correction for multiple testing was performed.

Results: Forty-one pts were enrolled as of 06/15/2015. The biomarker study subgroups were similar to the overall pt population with respect to demographics. Of the 37/41 (90%) pts assessed by FISH, 5 were t(11;14), 3 were t(4;14), 20 were 13q–, 10 were 17p–, and 15 were HY. In pts with HY, median best percentage change in M-protein was –79.4% compared to +101.65% in non-HY, while mToS was 309 days compared to 29 days in non-HY (p <0.0001), consistent with literature regarding a positive prognostic effect in MM. No difference in efficacy was observed in other FISH subtypes. BCL-2 family gene expression levels were determined in 25/41 (61%) pts. BCL-2, but not BCL-XL or MCL-1 gene expression, correlated with M-protein response (Spearman –0.5116, p=0.0149) and with a trend for increased mToS (HR=0.42 [95% CI 0.13–1.0]). Responders (≥ partial response, n=13; assessed by International Myeloma Working Group criteria 2011) had higher BCL-2 levels than subjects with stable disease/progressive disease (n=12; p=0.015). Pts with high BCL-XL or high MCL-1 were found in both the responder and nonresponder populations. Of the 14 pt samples collected for ex vivo sensitivity, 8 had sufficient CD138-positive cells for VEN ± BTZ dose curve analysis. Four of 8 (50%) were sensitive to VEN alone (median lethal dose [LD50] ≤300 nM) while 4/8 (50%) were sensitive to BTZ alone (LD50 ≤5 nM). Seven of 8 (88%) were sensitive to the combination (LD50 ≤5 nM BTZ/300 nM VEN).

Conclusions: VEN in combination with BTZ and dexamethasone is active in R/R MM. CD138-enriched BMMCs were obtained in sufficient numbers to enable downstream FISH analysis, BCL-2 family gene expression profiling, and ex vivo sensitivity evaluation. The exploratory biomarker analysis showed that higher BCL-2 gene expression levels at baseline significantly correlated with clinical responses to therapy, with a trend toward increased mToS. Additionally, MM cells treated ex vivo demonstrated greater sensitivity to the combination of VEN/BTZ than to the 2 single agents alone. Further investigation of this dataset continues, including analysis of on-treatment tumor samples in a subset of pts.