Search

Proteins of the bromodomain and extra-terminal (BET) domain family are epigenetic readers that bind acetylated histones through their bromodomains to regulate gene transcription. Dual-bromodomain BET inhibitors (DbBi) that bind with similar affinities to the first (BD1) and second (BD2) bromodomains of BRD2, BRD3, BRD4 and BRDt have displayed modest clinical activity in monotherapy cancer trials. A reduced number of thrombocytes in the blood (thrombocytopenia) as well as symptoms of gastrointestinal toxicity are dose-limiting adverse events for some types of DbBi1–5. Given that similar haematological and gastrointestinal defects were observed after genetic silencing of Brd4 in mice6, the platelet and gastrointestinal toxicities may represent on-target activities associated with BET inhibition. The two individual bromodomains in BET family proteins may have distinct functions7–9 and different cellular phenotypes after pharmacological inhibition of one or both bromodomains have been reported10,11, suggesting that selectively targeting one of the bromodomains may result in a different efficacy and tolerability profile compared with DbBi. Available compounds that are selective to individual domains lack sufficient potency and the pharmacokinetics properties that are required for in vivo efficacy and tolerability assessment10–13. Here we carried out a medicinal chemistry campaign that led to the discovery of ABBV-744, a highly potent and selective inhibitor of the BD2 domain of BET family proteins with drug-like properties. In contrast to the broad range of cell growth inhibition induced by DbBi, the antiproliferative activity of ABBV-744 was largely, but not exclusively, restricted to cell lines of acute myeloid leukaemia and prostate cancer that expressed the full-length androgen receptor (AR). ABBV-744 retained robust activity in prostate cancer xenografts, and showed fewer platelet and gastrointestinal toxicities than the DbBi ABBV-07514. Analyses of RNA expression and chromatin immunoprecipitation followed by sequencing revealed that ABBV-744 displaced BRD4 from AR-containing super-enhancers and inhibited AR-dependent transcription, with less impact on global transcription compared with ABBV-075. These results underscore the potential value of selectively targeting the BD2 domain of BET family proteins for cancer therapy. ABBV-744, a selective inhibitor of the BD2 domains of BET family proteins, is effective against prostate cancer in mouse xenograft models, with lower toxicities than the dual-bromodomain BET inhibitor ABBV-075.

Dual bromodomain BET inhibitors that bind with similar affinities to the first and second bromodomains across BRD2, BRD3, BRD4, and BRDT have displayed modest activity as monotherapy in clinical trials. Thrombocytopenia, closely followed by symptoms characteristic of gastrointestinal toxicity, have presented as dose-limiting adverse events that may have prevented escalation to higher dose levels required for more robust efficacy. ABBV-744 is a highly selective inhibitor for the second bromodomain of the four BET family proteins. In contrast to the broad antiproliferative activities observed with dual bromodomain BET inhibitors, ABBV-744 displayed significant antiproliferative activities largely although not exclusively in cancer cell lines derived from acute myeloid leukemia and androgen receptor positive prostate cancer. Studies in acute myeloid leukemia xenograft models demonstrated antitumor efficacy for ABBV-744 that was comparable with the pan-BET inhibitor ABBV-075 but with an improved therapeutic index. Enhanced antitumor efficacy was also observed with the combination of ABBV-744 and the BCL-2 inhibitor, venetoclax compared with monotherapies of either agent alone. These results collectively support the clinical evaluation of ABBV-744 in AML (Clinical Trials.gov identifier: NCT03360006).

The TMPRSS2–ERG gene fusion and subsequent overexpression of the ERG transcription factor occurs in ∼50% of prostate tumors, making it the most common abnormality of the prostate cancer genome. While ERG has been shown to drive tumor progression and cancer-related phenotypes, as a transcription factor it is difficult to target therapeutically. Using a genetic screen, we identified the toll-like receptor 4 (TLR4) signaling pathway as important for ERG function in prostate cells. Our data confirm previous reports that ERG can transcriptionally activate TLR4 gene expression; however, using a constitutively active ERG mutant, we demonstrate that the critical function of TLR4 signaling is upstream, promoting ERG phosphorylation at serine 96 and ERG transcriptional activation. The TLR4 inhibitor, TAK-242, attenuated ERG-mediated migration, clonogenic survival, target gene activation and tumor growth. Together these data indicate a mechanistic basis for inhibition of TLR4 signaling as a treatment for ERG-positive prostate cancer., Graphical Abstract Graphical AbstractToll-like receptor 4 signaling activates ERG function in prostate cancer and provides a therapeutic target.

Metastatic colonization involves paracrine/juxtacrine interactions with the microenvironment inducing an adaptive response through transcriptional regulation. However, the identities of transcription factors (TFs) induced by the metastatic microenvironment in ovarian cancer (OC) and their mechanism of action is poorly understood. Using an organotypic 3D culture model recapitulating the early events of metastasis, we identified ETS1 as the most upregulated member of the ETS family of TFs in metastasizing OC cells as they interacted with the microenvironment. ETS1 was regulated by p44/42 MAP kinase signaling activated in the OC cells interacting with mesothelial cells at the metastatic site. Human OC tumors had increased expression of ETS1, which predicted poor prognosis. ETS1 regulated OC metastasis both in vitro and in mouse xenografts. A combination of ChIP-seq and RNA-seq analysis and functional rescue experiments revealed FAK as the key transcriptional target and downstream effector of ETS1. Taken together, our results indicate that ETS1 is an essential transcription factor induced in OC cells by the microenvironment, which promotes metastatic colonization though the transcriptional upregulation of its target FAK.

Summary: More than 50% of prostate tumors have a chromosomal rearrangement resulting in aberrant expression of an oncogenic ETS family transcription factor. However, mechanisms that differentiate the function of oncogenic ETS factors expressed in prostate tumors from non-oncogenic ETS factors expressed in normal prostate are unknown. Here, we find that four oncogenic ETS (ERG, ETV1, ETV4, and ETV5), and no other ETS, interact with the Ewing’s sarcoma breakpoint protein, EWS. This EWS interaction was necessary and sufficient for oncogenic ETS functions including gene activation, cell migration, clonogenic survival, and transformation. Significantly, the EWS interacting region of ERG has no homology with that of ETV1, ETV4, and ETV5. Therefore, this finding may explain how divergent ETS factors have a common oncogenic function. Strikingly, EWS is fused to various ETS factors by the chromosome translocations that cause Ewing’s sarcoma. Therefore, these findings link oncogenic ETS function in both prostate cancer and Ewing’s sarcoma. : A subset of ETS transcription factors is oncogenic in prostate. Kedage et al. show that oncogenic ETS, but not other ETS, interact with EWS, and this interaction is necessary for oncogenic functions. Because EWS is fused to ETS factors in Ewing’s sarcoma, this finding links the mechanisms of these diseases. Keywords: prostate cancer, ETS, EWS, Ewing’s sarcoma, ERG

Metastatic colonization of ovarian cancer involves productive paracrine/juxtacrine interactions with the microenvironment. The resulting induction of an adaptive response in the cancer cells enables them to establish themselves in the new microenvironment and take advantage of the new factors available. A key feature of this adaptation is induced changes in gene expression through transcriptional regulation as a result of microenvironmental cues. However, the identities of transcription factors induced by the metastatic microenvironment in ovarian cancer and their mechanism of action are poorly understood. Using an organotypic 3D culture model recapitulating the early events of metastasis, we identified ETS1, a member of the ETS family of TFs, as an essential driver of metastatic colonization. Increased ETS1 expression was induced in metastasizing ovarian cancer cells interacting with the mesothelial cells covering the surface of the omentum. The mechanism of upregulation was through the activation of p44/42 MAP kinase signaling in the cancer cells induced by TGFbeta from the microenvironment. We also found an increased ETS1 expression in human ovarian cancer samples as compared to normal fallopian tubes using a tissue microarray. Moreover, higher expression of ETS1 was a predictor of poor prognosis in ovarian cancer patients. Knocking down ETS1 decreased migration, proliferation, and colony formation as well as invasion through and colonization of the organotypic 3D culture. Overexpression of ETS1 had the opposite effect. CRISPR/Cas9-mediated knockout of ETS1 resulted in decreased tumor burden in mouse xenografts. A combination of ChIP-seq and RNA-seq analysis revealed that ETS1 promoted an EMT phenotype and FAK was identified as a novel transcriptional target. Inhibition of FAK functionally mimicked the effects of ETS1 inhibition in the ovarian cancer cells. Moreover, functional rescue experiments established FAK as a downstream effector of ETS1 during ovarian cancer metastasis. Taken together, our results indicate that ETS1 is an essential transcription factor induced in ovarian cancer cells by the microenvironment, which promotes metastatic colonization. This is the first report establishing FAK as a transcriptional target and functional effector of ETS1 in establishing metastatic tumors. Citation Format: Sunil Tomar, Joshua P. Plotnik, James Haley, Joshua Scantland, Zahir Sheikh, Robert Emerson, Dean Lenz, Peter C. Hollenhorst, Anirban K. Mitra. Induction of a novel ETS1/FAK pathway in metastasizing ovarian cancer cells by the omental microenvironment primes them for metastatic colonization. [abstract]. In: Proceedings of the AACR Conference: Addressing Critical Questions in Ovarian Cancer Research and Treatment; Oct 1-4, 2017; Pittsburgh, PA. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(15_Suppl):Abstract nr A34.