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Supplemental table 1: Semi-quantitative immunohistochemical (IHC) analysis of s.c. DLBCL cell line xenografts. from Targeting of BCL2 Family Proteins with ABT-199 and Homoharringtonine Reveals BCL2- and MCL1-Dependent Subgroups of Diffuse Large B-Cell Lymphoma

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posted on 2023-03-31, 18:25 authored by Magdalena Klanova, Ladislav Andera, Jan Brazina, Jan Svadlenka, Simona Benesova, Jan Soukup, Dana Prukova, Dana Vejmelkova, Radek Jaksa, Karel Helman, Petra Vockova, Lucie Lateckova, Jan Molinsky, Bokang Calvin Lenyeletse Maswabi, Mahmudul Alam, Roman Kodet, Robert Pytlik, Marek Trneny, Pavel Klener

To investigate whether the expression level of BCL2 proteins detected by western blot is comparable with the expression level assessed by IHC analysis, we analyzed protein expression (by IHC) of BCL2, MCL1 and BCL-XL from subcutaneous lymphoma xenografts obtained from mice xenotransplanted with selected DLBCL cell lines with known protein expression (by western blot).

Funding

Czech Ministry of Health Research

Czech Republic Grant Agency Research

Charles University Research Center Financial

Charles University Programs of Research Development

Charles University Grant Agency Research

Specific University Research Program

Charles University in Prague

History

ARTICLE ABSTRACT

Purpose: To investigate the roles of BCL2, MCL1, and BCL-XL in the survival of diffuse large B-cell lymphoma (DLBCL).Experimental designs: Immunohistochemical analysis of 105 primary DLBCL samples, and Western blot analysis of 18 DLBCL cell lines for the expression of BCL2, MCL1, and BCL-XL. Pharmacologic targeting of BCL2, MCL1, and BCL-XL with ABT-199, homoharringtonine (HHT), and ABT-737. Analysis of DLBCL clones with manipulated expressions of BCL2, MCL1, and BCL-XL. Immunoprecipitation of MCL1 complexes in selected DLBCL cell lines. Experimental therapy aimed at inhibition of BCL2 and MCL1 using ABT-199 and HHT, single agent, or in combination, in vitro and in vivo on primary cell-based murine xenograft models of DLBCL.Results: By the pharmacologic targeting of BCL2, MCL1, and BCL-XL, we demonstrated that DLBCL can be divided into BCL2-dependent and MCL1-dependent subgroups with a less pronounced role left for BCL-XL. Derived DLBCL clones with manipulated expressions of BCL2, MCL1, and BCL-XL, as well as the immunoprecipitation experiments, which analyzed MCL1 protein complexes, confirmed these findings at the molecular level. We demonstrated that concurrent inhibition of BCL2 and MCL1 with ABT-199 and HHT induced significant synthetic lethality in most BCL2-expressing DLBCL cell lines. The marked cytotoxic synergy between ABT-199 and HHT was also confirmed in vivo using primary cell-based murine xenograft models of DLBCL.Conclusions: As homoharringtonine is a clinically approved antileukemia drug, and ABT-199 is in advanced phases of diverse clinical trials, our data might have direct implications for novel concepts of early clinical trials in patients with aggressive DLBCL. Clin Cancer Res; 22(5); 1138–49. ©2015 AACR.

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