BDA-366, a putative Bcl-2 BH4 domain antagonist, induces apoptosis independently of Bcl-2 in a variety of cancer cell models

Tamara Vervloessem,Anastassios Economou,Flore Sneyers,Spyridoula Karamanou,Justin Kale,David W Andrews,Rita M La Rovere,Supriya Chakraborty,Luca Laurenti,Elena Xerxa,Dimitar G Efremov,Geert Bultynck,Meike Vogler,Binu K Sasi

doi:10.1038/s41419-020-02944-6

Abstract

Several cancer cell types, including chronic lymphocytic leukemia (CLL) and diffuse large B-cell lymphoma (DLBCL) upregulate antiapoptotic Bcl-2 to cope with oncogenic stress. BH3 mimetics targeting Bcl-2’s hydrophobic cleft have been developed, including venetoclax as a promising anticancer precision medicine for treating CLL patients. Recently, BDA-366 was identified as a small molecule BH4-domain antagonist that could kill lung cancer and multiple myeloma cells. BDA-366 was proposed to switch Bcl-2 from an antiapoptotic into a proapoptotic protein, thereby activating Bax and inducing apoptosis. Here, we scrutinized the therapeutic potential and mechanism of action of BDA-366 in CLL and DLBCL. Although BDA-366 displayed selective toxicity against both cell types, the BDA-366-induced cell death did not correlate with Bcl-2-protein levels and also occurred in the absence of Bcl-2. Moreover, although BDA-366 provoked Bax activation, it did neither directly activate Bax nor switch Bcl-2 into a Bax-activating protein in in vitro Bax/liposome assays. Instead, in primary CLL cells and DLBCL cell lines, BDA-366 inhibited the activity of the PI3K/AKT pathway, resulted in Bcl-2 dephosphorylation and reduced Mcl-1-protein levels without affecting the levels of Bcl-2 or Bcl-xL. Hence, our work challenges the current view that BDA-366 is a BH4-domain antagonist of Bcl-2 that turns Bcl-2 into a pro-apoptotic protein. Rather, our results indicate that other mechanisms beyond switching Bcl-2 conformation underlie BDA-366’s cell-death properties that may implicate Mcl-1 downregulation and/or Bcl-2 dephosphorylation.

Highlights

An important survival strategy of many cancer cells is upregulation of antiapoptotic Bcl-2 proteins[1]
Primary chronic lymphocytic leukemia (CLL) and diffuse large B-cell lymphoma (DLBCL) cell lines are sensitive to BDA-366 To investigate whether BDA-366 can kill CLL cells with higher efficiency than normal cells, we analyzed the viability of CLL (n = 39) and normal peripheral blood mononuclear cells (PBMCs) (n = 6) incubated with BDA366 for 48 h (Fig. 1a)
To further examine this heterogeneity, we studied a collection of Bcl-2-dependent DLBCL cell lines

Summary

Introduction

An important survival strategy of many cancer cells is upregulation of antiapoptotic Bcl-2 proteins[1]. Cancer cells such as chronic lymphocytic leukemia (CLL) and diffuse large B-cell lymphoma (DLBCL) exploit the antiapoptotic effects of Bcl-2 to survive oncogenic stress, though such. Much effort has been undertaken in finding a therapeutic strategy to antagonize Bcl-2 with a focus on small molecules that occupy Bcl-2’s hydrophobic cleft[2,8]. BH3 mimetics, such as venetoclax, have been developed to induce apoptosis by targeting Bcl-2’s hydrophobic cleft without directly activating Bax[9]. Vervloessem et al Cell Death and Disease (2020)11:769 oncogenic stress, they are sensitive to Bcl-2-antagonizing therapeutics such as venetoclax[3,10]. A broad scale of on-target BH3 mimetic inhibitors of several antiapoptotic Bcl-2-family members have been developed[11]

Methods

Results

Conclusion