OTX015, a novel BET-BRD inhibitor, is active in non-small-cell lung cancer (NSCLC) cell lines harboring different oncogenic mutations

RIVEIRO, MARÍA EUGENIA; ASTORGUES-XERRI, LUCILE; IJAZ, NAZISH; BREKRADDA, MOHAMED; VAZQUEZ, RAMIRO; FRAPOLLI, ROBERTA; RINALDI, ANDREA; KWEE, IVO; CVITKOVIC, ESTEBAN; RAYMOND, ERIC

Barcelona

Congreso; 26th meeting 'Molecular Targets and Cancer Therapeutics'; 2014

AACR-NCI-EORTC

Background: The human BET bromodomain protein family is a druggabletarget through inhibitors that show potent antiproliferative effects in severaltumors including NSCLC. We report preclinical findings of the BETBRD inhibitor OTX015 in NSCLC cell lines harboring different oncogenicmutations (KRAS, MYC amplifications, fusion protein EML4−ALK) as asingle agent or combined with targeted therapies.Material and Methods: OTX015 growth inhibition 50% (GI50) valueswere determined with the MTT assay after 72 h exposure in 5 establishedNSCLC cell lines (H2228, H3122, A549, HOP62, HOP92). Protein levelswere analyzed by Western Blot with commercial antibodies; RT-PCR wasdone with Fast SYBR Green Master Mix on a StepOnePlus Real-TimePCR System at baseline, 4 and 24 h post-treatment. For cell cycle analysiscells were stained with propidium iodide and DNA content analyzed with aFACScan flow cytometer. OTX015 was combined with the targeted agentseverolimus and crizotinib and the combination index (CI) determined by theChou?Talalay method.Results: OTX015 had antiproliferative effects in 4 of 5 NSCLC cell lines(two harboring the fusion protein EML4−ALK+) with GI50 values from 110to 940 nM. In A549 cells, concurrent mutations in KRAS and LKB1 genesabrogated OTX015 effects, as described for JQ1, an OTX015 analog, whileHOP92 and HOP62 are LKB1 wild-type. Percent cells in the S phase werereduced by 40%, 42% and 52% in HOP92, H2228 and H3122, respectivelyafter 24 h exposure at 500 nM. OTX015 combined with the ALK+-inhibitorcrizotinib had additive effects (CI = 0.98) after 48 h concomitant exposurein H2228 cells and additive/synergistic effects with everolimus in HOP62,HOP92, A549 and H2228. OTX015-sensitive and -resistant lines hadsimilar basal mRNA and protein expression of BRD2/3/4, HEXIM1, BCL-2and P21. CMYC and NMYC were overexpressed in HOP92 and H3122,respectively. Following 24 h OTX015 (500 nM), CMYC protein and mRNAwere downregulated in HOP92 cells and NMYC downregulated in H3122and HOP62 cells, while HEXIM mRNA was upregulated in OTX015-sensitive cell lines.Conclusion: NSCLC cell lines, including those harboring the EML4-ALKfusion gene or KRAS mutation, are sensitive to OTX015 BET inhibition,downregulating MYC family members and upregulating HEXIM, suggestingthe therapeutic potential of OTX015 in NSCLC. In vivo studies of singleagent OTX015 and combined with everolimus in HOP92 xenografts will bepresented.