BCL2-Family Dysregulation in B-Cell Malignancies: From Gene Expression Regulation to a Targeted Therapy Biomarker

Abstract

BCL2-family proteins have a central role in the mitochondrial apoptosis machinery and their expression is known to be deregulated in many cancer types. Effort in the development of small molecules that selectively target anti-apoptotic members of this family i.e., Bcl-2, Bcl-xL, Mcl-1 recently opened novel therapeutic opportunities. Among these apoptosis-inducing agents, BH3-mimetics (i.e., venetoclax) led to promising preclinical and clinical activity in B cell malignancies. However, several mechanisms of intrinsic or acquired resistance have been described ex vivo therefore predictive markers of response as well as mechanism-based combinations have to be designed. In the present study, we analyzed the expression of the BCL2-family genes across 10 mature B cell malignancies through computational normalization of 21 publicly available Affimetrix datasets gathering 1,219 patient samples. To better understand the deregulation of anti- and pro-apoptotic members of the BCL2-family in hematological disorders, we first compared gene expression profiles of malignant B cells to their relative normal control (naïve B cell to plasma cells, n = 37). We further assessed BCL2-family expression according to tissue localization i.e., peripheral blood, bone marrow, and lymph node, molecular subgroups or disease status i.e., indolent to aggressive. Across all cancer types, we showed that anti-apoptotic genes are upregulated while pro-apoptotic genes are downregulated when compared to normal counterpart cells. Of interest, our analysis highlighted that, independently of the nature of malignant B cells, the pro-apoptotic BH3-only BCL2L11 and PMAIP1 are deeply repressed in tumor niches, suggesting a central role of the microenvironment in their regulation. In addition, we showed selective modulations across molecular subgroups and showed that the BCL2-family expression profile was related to tumor aggressiveness. Finally, by integrating recent data on venetoclax-monotherapy clinical activity with the expression of BCL2-family members involved in the venetoclax response, we determined that the ratio (BCL2+BCL2L11+BAX)/BCL2L1 was the strongest predictor of venetoclax response for mature B cell malignancies in vivo.

Keywords: B-cell malignancy; BCL2; cell death; data mining; lymphoma; microenvironment; predictive markers.

Figure 1

BCL2-family is strongly deregulated in the landscape of B-cell malignancies. (A) Heat-map of Bcl-2 gene expression profiles among B-cell malignancies. The color corresponds to the intensity of the median gene expression. Blue indicates lower and red higher transcript abundance. MCL, Mantle Cell Lymphoma; BL, Burkitt lymphoma; DLBCL, Diffuse Large B-cell Lymphoma; FL, Follicular Lymphoma; BPLL, B-cell Prolymphocytic Leukemia; CLL, Chronic lymphocytic leukemia; HCL, Hairy Cell Lymphoma; MALT, mucosa-associated lymphoid tissue lymphoma; SMZL, Splenic Marginal Zone Lymphoma; BMPC, Bone Marrow Plasma Cell, MM: multiple Myeloma. (B) Expression of BCL2, BCL2A1, and BCL2L11 in the different B-cell malignancies compared to their respective control. Wilcoxon-Mann-Whitney tests. ^*p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001. (C) Representation of the individual factor map of each sample for the PCA and according to the two first dimensions. Colored ellipses are drawn around the mean of the group (= barycenter), with the 95% confidence interval of the mean in the corresponding plan. BCL2 is coding for Bcl-2 protein, BCL2L1 for Bcl-xL, MCL1 for Mcl-1, BCL2L2 for Bcl-w, BCL2A1 for Bfl1, BIK for Bik, PMAIP1 for Noxa, BMF for Bmf, BID for Bid, BCL2L11 for Bim, BAX for Bax, and BAK1 for Bak.

Figure 2

BCL2-family is regulated by the tumor microenvironment. (A) Heat-map of Bcl-2 gene expression profiles for MCL, FL, CLL, and SMZL in function of their tissue localization. Wilcoxon-Mann-Whitney tests. ^*p < 0.05. (B) Comparison of BCL2L11, PMAIP1, and BCL2L1 gene expression according to their localization. LN, lymph nodes; PB, peripheral blood; BM, bone marrow. Wilcoxon-Mann-Whitney tests. ^**p < 0.01, ^****p < 0.0001. (C) Representation of the individual factor map for the PCA according to the two first dimensions and their respective correlation circle. Colored ellipses are drawn around the mean of the group (= barycenter), with the 95% confidence interval of the mean in the corresponding plan.

Figure 3

Molecular subtypes display differential expression of the BCL2-family. (A) left; Heat-map of BCL2-family expression profile comparing PB MCL according to SOX11 gene expression (right). MCL1 and BIK expression in the two molecular subgroups. (B) left; Heat-map of BCL2-family expression profile comparing the different subtypes of DLBCL: GCB (germinal center B cell, ABC (activated B-cell) and PMBL (Primary mediastinal B-cell lymphoma). right; Gene expression of BCL2L1, BCL2, BID, and BMF for the three subtypes of DLBCL. (C) Representation of the individual factor map for the PCA and according to the two first dimensions of multiple myeloma samples and their respective correlation circle. Colored ellipses are drawn around the mean of the group (= barycenter), with the 95% confidence interval of the mean in the corresponding plan. (D) BCL2L2, BCL2L11, and BMF expression in the different multiple myeloma subtypes. ^*p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001.

Figure 4

Intra-entities BCL2-family heterogeneity is related to aggressiveness. (A) Heat-map of BCL2-family expression profile comparing FL and MALT samples according to their indolent (ind) or aggressive (agr) status. (B) BCL2, BCL2L11, BID, BAX, and BAK1 expression in FL and MALT sample according to their aggressiveness. (C) Representation of the individual factor map for the PCA and according to the two first dimensions of lymphoma samples and their respective correlation circle. Colored ellipses are drawn around the mean of the group (= barycenter), with the 95% confidence interval of the mean in the corresponding plan. ^*p < 0.05, ^**p < 0.01, ^****p < 0.0001.

Figure 5

(BCL2+BCL2L11+BAX)/BCL2L1 ratio predicts response to Bcl2 specific BH3 mimetic. (A) Evaluation of the (BCL2+BCL2L11+BAX)/BCL2L1 ratio for the different B-cell malignancies associated to ORR and PFS of patients treated with venetoclax-monotherapy when available. (B,C) Evaluation of the (BCL2 + BCL2L11 + BAX)/BCL2L1 ratio for the different subtypes of (B) MM and (C) MCL and DLBCL. (D) Evaluation of the (BCL2 + BCL2L11 + BAX)/BCL2L1 ratio for the different subtypes of FL and MALT. (E) Evaluation of the (BCL2 + BCL2L11 + BAX)/BCL2L1 ratio for MCL, FL, CLL, SMZL according to their tissue localization (peripheral blood, PB, lymph nodes, LN, spleen, SPL). ^*p < 0.05, ^**p < 0.01, ^****p < 0.0001.