Abstract A119: The BET-bromodomain inhibitor OTX015 shows synergism with several anticancer agents in preclinical models of diffuse large B-cell lymphoma (DLBCL).
Authors: Gaudio Eugenio , Kwee Ivo , Bernasconi Elena , Rinaldi Andrea , Testoni Monica , Boi Michela , Stathis Anastasios , Riveiro Eugenia , Noel Kay , Zucca Emanuele , Bertoni Francesco 
Published: August 2014

Abstract
Relapsed/refractory DLBCL is still the cause of death in a portion of patients, hence novel active therapies are needed. BET Bromodomain inhibitors have shown promising preclinical activity in hematological and solid tumors and are currently investigated in early clinical trials. We have shown preclinical single-agent activity of the BET Bromodomain OTX015 on mature lymphoid tumors, including DLBCL (AACR 2012; ICML 2013). Here, we report on OTX015 combinations with other anticancer compounds in DLBCL cell lines.
Methods: 3 germinal center B cell (GCB) DLBCL and 2 activated B cell (ABC) DLBCL cell lines were exposed to increasing doses of OTX015 alone or in combination with increasing doses of other drugs. The MTT assay was performed after 72 hours of exposure. Synergy was assessed by Chou-Talalay combination index (CI) with the Synergy R package: CI&lt;0.3, strong synergism; 0.3-0.9, synergism; 0.9-1.1, additive effect. Baseline gene expression profiles (GEP) were obtained in 38 lymphoma cell lines, including 22 DLBCL with Illumina HumanHT-12 v4 Expression BeadChip. GEP before and after OTX015 treatment were done in 3 DLBCL cell lines, too. The relationship between GEP and IC50 values was assessed by Pearson correlation. LIMMA was used for differential expression analysis, followed by Benjamini-Hochberg multiple test correction, and GSEA to test for enrichment of functionally-related genes.
Results: Strong synergism was observed with OTX015 combined with the mTOR inhibitor everolimus (median CI, 0.11; range 0.1-0.17) and with the BTK-inhibitor ibrutinib in ABC-cells (CI=0.04; 0.02-0.1). A synergistic effect was estimated for OTX015 in combinations with the class I and II HDAC-inhibitor vorinostat (CI=0.45; 0.31-0.56), the anti-CD20 moAb rituximab (CI=0.47; 0.37-0.54), the hypomethylating agent decitabine (CI=0.62; 0.56-0.66), and the immunomodulant lenalidomide (CI=0.66; 0.59-0.72). OTX015 combinations with the class I HDAC inhibitor romidepsin (CI=1.08; 1-1.22) and with the chemotherapy agents bendamustine (CI=0.92; 0.83-1.1) and doxorubicin (CI=0.83; 0.71-0.96) presented a moderate additive effect. A stronger synergism was observed in ABC than in GCB DLBCL cells for ibrutinib (P&lt;0.0001), lenalidomide (P=0.0001), and rituximab (P=0.007). Data mining of GEP obtained at baseline across 38 lymphoma cell lines with known OTX015 IC50s and GEP changes observed after OTX015 exposure indicated the relevance of genes involved in MYD88/JAK/STAT pathway and glucose metabolism as possible explanations of the observed synergism of OTX015 with targeted agents, such as ibrutinib and everolimus.
Conclusions: Several targeted agents appeared to synergize with OTX015: additional studies are warranted to clarify the potential clinical role of BET bromodomain inhibitors.
Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A119.
Citation Format: Eugenio Gaudio, Ivo Kwee, Elena Bernasconi, Andrea Rinaldi, Monica Testoni, Michela Boi, Anastasios Stathis, Eugenia Riveiro, Kay Noel, Emanuele Zucca, Francesco Bertoni. The BET-bromodomain inhibitor OTX015 shows synergism with several anticancer agents in preclinical models of diffuse large B-cell lymphoma (DLBCL). [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A119.