Selective BET Inhibitor Research Articles

Discovery of a series of dihydroquinoxalin-2(1H)-ones as selective BET inhibitors from a dual PLK1-BRD4 inhibitor

Recent years have seen much effort to discover new chemotypes of BRD4 inhibitors. Interestingly, some kinase inhibitors have been demonstrated to be potent bromodomain inhibitors, especially the PLK1 inhibitor BI-2536 and the JAK2 inhibitor TG101209, which can bind to BRD4 with IC50 values of 0.025 μM and 0.13 μM, respectively. Although the concept of dual inhibition is intriguing, selective BRD4 inhibitors are preferred as they may diminish off-target effects and provide more flexibility in anticancer drug combination therapy. Inspired by BI-2536, we designed and prepared a series of dihydroquinoxalin-2(1H)-one derivatives as selective bromodomain inhibitors. We found compound 54 had slightly higher activity than (+)-JQ1 in the fluorescence anisotropy assay and potent antiproliferative cellular activity in the MM.1S cell line. We have successfully solved the cocrystal structure of 52 in complex with BRD4-BD1, providing a solid structural basis for the binding mode of compounds of this series. Compound 54 exhibited high selectivity over most non-BET subfamily members and did not show bioactivity towards the PLK1 kinase at 10 or 1 μM. From in vivo studies, compound 54 demonstrated a good PK profile, and the results from in vivo pharmacological studies clearly showed the efficacy of 54 in the mouse MM.1S xenograft model.

Abstract 3780: Activity of the BET inhibitor INCB054329 in models of lymphoma

Abstract Inhibitors of the BET family of Bromodomain proteins have been shown to be growth inhibitory across a spectrum of tumor types due to their ability to regulate expression of key survival and cell fate determining genes such as c-myc. Among the various tumor histologies, hematologic malignancies are among the most sensitive cancers to BET inhibition. INCB054329 is a novel, non-benzodiazepine, selective BET inhibitor that is undergoing Phase 1 clinical trials and that has shown encouraging in vitro and in vivo preclinical activity in several models of hematologic malignancy. In the current study, the activity of INCB054329 was evaluated in models of B cell malignancy. INCB54329 effectively inhibited the in vitro growth of a panel of cell lines representing both Hodgkin and non-Hodgkin lymphoma. Treated cells arrested primarily in G1 with sensitive lines also exhibiting dose and time-dependent apoptosis. Within a panel of double-hit lymphoma cell lines, which have activating chromosomal rearrangements in both c-myc and bcl-2, INCB054329 potently inhibited cell growth and was more effective than antagonists of BTK, bcl-2, PIM and PI3Kδ. INCB054329 also showed in vivo efficacy in models of diffuse large B-cell lymphoma (DLBCL). As a single agent, oral administration of INCB054329 inhibited tumor growth in Pfeiffer (GBC) and WILL-2 (GCB, double-hit) subcutaneous xenograft models. The in vivo combination of bendamustine with INCB054329 enhanced anti-tumor efficacy compared with either agent alone in the Pfeiffer model, and the combination was well tolerated. A rational, targeted combination strategy was evaluated involving INCB054329 and a selective, orally active PI3Kδ inhibitor, INCB050465, which is currently in clinical trials in B cell malignancies. Combining INCB054329 with PI3Kδ inhibition markedly enhanced anti-tumor efficacy, increasing the incidence of partial tumor regressions in vivo. In this model, both INCB054329 and INCB050465 treatment led to a reduction in c-Myc protein levels, suggesting a convergence between modulation of BET transcriptional regulation and the PI3Kδ pathway. These data suggest that clinical investigation of INCB054329, both as monotherapy and in combination with standard of care or novel targeted therapies, in several classes of B cell lymphoma, including high risk double hit lymphoma, is warranted. Citation Format: Matthew Stubbs, Robert Collins, Alla Volgina, Mike Liu, Margaret Favata, Mark Rupar, Xiaomng Wen, Richard Sparks, Thomas Maduskuie, Maryanne Covington, Timothy Burn, Bruce Ruggeri, Andrew P. Combs, Wenqing Yao, Reid Huber, Gregory Hollis, Peggy Scherle, Phillip CC Liu. Activity of the BET inhibitor INCB054329 in models of lymphoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3780.

Abstract 4904: The BET inhibitor INCB054329 enhances the activity of checkpoint modulation in syngeneic tumor models

Abstract Inhibitors of the BET family of bromodomain proteins have been shown to be growth inhibitory across a spectrum of tumor types due to their ability to regulate the expression of key survival and cell fate determining genes such as c-myc. In addition to their role in cancer, studies using genetic knockdown and small molecule inhibitors have demonstrated that targeting BET proteins controls the expression of pro-inflammatory cytokine genes in macrophages and is therapeutic in models of acute inflammation. These data suggest that in addition to their tumor intrinsic effects, BET inhibitors may also regulate the cytokine milieu within the tumor microenvironment and have immunomodulatory activity in cancer. To study this aspect, we evaluated INCB054329, a novel and selective BET inhibitor currently in Phase 1 trials, alone and in combination either with epacadostat, a highly selective IDO1 inhibitor, or with PD-1/PD-L1 axis blockade in syngeneic tumor models using immunocompetent animals. When used alone, INCB054329 suppressed a panel of cytokines and chemokines in a whole blood assay, confirming that INCB054329 can antagonize a pro-inflammatory response. The potency of INCB054329 in reducing the levels of these inflammatory mediators in the whole blood assay was similar to that for inhibition of c-myc, suggesting that the effects were on-target. INCB054329 was capable of inhibiting the growth of multiple syngeneic tumor models in immunocompetent mice, whereas only modest tumor growth inhibition was observed in immunodeficient mice and a lack of activity was observed in vitro, supporting the immunomodulatory activity of the compound. Because maximal in vivo tumor growth inhibition required an intact immune system, we investigated the impact of INCB054329 on various immune cell subsets, both in vitro and in vivo. Of note, increases in effector T cell populations were observed and efforts are ongoing to further characterize the tumor infiltrating immune cells following INCB054329 treatment. The mechanistic complimentarity of this novel BET inhibitor-mediated immunomodulation was also evaluated in combination with other therapeutically relevant mechanisms, including IDO1 inhibition and PD-1 axis blockade. Enhanced efficacy was observed with all INCB054329-containing regimens. These data demonstrate for the first time that BET inhibition can suppress tumor growth through both tumor-intrinsic and immune modulatory mechanisms, and support the potential of epigenetic-based, immunotherapy combinations as a novel approach to cancer therapy. Citation Format: Holly K. Koblish, Michael Hansbury, Leslie Hall, Liang-Chuan Wang, Yue Zhang, Maryanne Covington, Timothy Burn, Mark Rupar, Christine Gardiner, Thomas Condamine, Kerri Lasky, Matthew C. Stubbs, Eddy Yue, Richard Sparks, Richard Sparks, Thomas Maduskuie, Andrew P. Combs, Gregory Hollis, Reid Huber, Phillip CC Liu, Peggy Scherle. The BET inhibitor INCB054329 enhances the activity of checkpoint modulation in syngeneic tumor models. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4904.

Disrupting Acetyl-Lysine Recognition: Progress in the Development of Bromodomain Inhibitors

Bromodomains, small protein modules that recognize acetylated lysine on histones, play a significant role in the epigenome, where they function as "readers" that ultimately determine the functional outcome of the post-translational modification. Because the initial discovery of selective BET inhibitors have helped define the role of that protein family in oncology and inflammation, BET bromodomains have continued to garner the most attention of any other bromodomain. More recently, non-BET bromodomain inhibitors that are potent and selective have been disclosed for ATAD2, CBP, BRD7/9, BRPF, BRPF/TRIM24, CECR2, SMARCA4, and BAZ2A/B. Such novel inhibitors can be used to probe the physiological function of these non-BET bromodomains and further understanding of their role in certain disease states. Here, we provide an update to the progress in identifying selective bromodomain inhibitors and their use as biological tools, as well as our perspective on the field.

Abstract 17127: RVX-208 a Selective Bromodomain and Extra-terminal BET Protein Inhibitor Acts on Several Pathways to Benefit Cardiovascular Risks

The epigenetic modifying compound RVX-208 selectively inhibits the second ligand binding domain in BET proteins to increase ApoA-I/HDL production. In recent trials of CVD patients given standard of care and RVX-208 (200mg/d) there was a 55% relative risk reduction of MACE vs. placebo. This reduction is greater than expected from modest increases in ApoA-I/HDL, thus suggesting benefits of RVX-208 beyond lipid effects. Whether added benefits exist was explored by microarray survey of differential gene expression in primary human hepatocytes (PHH) and whole blood (WB). In PHH, RVX-208 downregulated genes within the complement, fibrin clotting, cholesterol biosynthesis, fatty acid biosynthesis, diabetes and acute phase response (APR) pathways. These data guided more focused analyses on the known HDL proteome encoded by 89 genes of which 30 were affected by RVX-208 and many of these were members of the APR or complement pathways. In support of the microarray data, we used RT-PCR and found RVX-208 lowered mRNA encoding complement 3, 4a/4b and 5 by >50% in Huh-7 and HepG2 cells exposed to RVX-208. Consistent with the mRNA data, secretion of these complement proteins were less from the treated cells. Next the cells were exposed to cytokines in order to mimic an inflammatory state thus inducing complement expression. Treatment with RVX-208 blocked this effect by >50%. Furthermore, in Huh-7 cells, RVX-208 repressed both fibrin clotting factors II (prothrombin) and XI (thromboplastin antecedent) consistent with the PHH microarray data thus suggesting an anti-thrombotic effect for RVX-208. To further extend findings in vitro from hepatoma cells, plasma from patients in our clinical trials who received RVX-208 were assayed and found to have decreased levels of both APR and complement proteins. Lastly, microarray studies of WB from healthy donors treated ex-vivo with RVX-208 uncovered differential regulation of anti- and pro-atherogenic gene sets that were activated and suppressed, respectively by RVX-208. In summary, RVX-208, a selective BET inhibitor, affects several pathways involved in CVD including; reverse cholesterol transport, atherogenesis, complement, thrombosis and vascular inflammation that cumulatively may reduce MACE in high risk CVD patients.

Abstract 2867: Inhibition of BET bromodomain proteins as a therapeutic approach in small cell lung cancer

Abstract BET (bromodomain and extra-terminal) family proteins are epigenetic readers that regulate expression of genes involved in cell growth and oncogenesis. Selective small molecule BET inhibitors, such as the GSK I-BETs (I-BET762, I-BET151), abrogate binding of BET proteins to acetylated histones and inhibit transcriptional activation of BET target genes. BET inhibitors attenuate proliferation and survival in a number of hematologic cancer models, partially through down-regulation of the critical oncogene, MYC. We and others have previously shown activity for BET inhibitors in solid tumor models such as neuroblastoma and prostate cancer, with concomitant down-regulation of MYC family gene expression. However, MYC or MYCN silencing only partially accounts for the activity of BET inhibitors in these models, suggesting that transcriptional regulation of multiple pathways promotes I-BET induced phenotypes. Here we describe the activity of the selective BET inhibitor, I-BET762, in small cell lung cancer (SCLC), a highly aggressive malignancy with few treatment options. We observe potent growth inhibition and apoptosis in a subset of cell lines and patient-derived SCLC models in vitro, as well as significant tumor growth inhibition in cell line and primary mouse xenografts. These anti-proliferative effects are likely mediated by inhibition of BRD2, BRD3, and BRD4, as siRNAs individually targeting all three proteins result in partial inhibition of SCLC cell line growth. Similar to our observations in neuroblastoma and prostate cancer, I-BET762 treatment in SCLC cell lines results in transcriptional changes affecting MYC and other pathways. We explore the contribution of these changes to the anti-proliferative effects observed in SCLC models, and identify potential combination strategies to enhance the activity of I-BET762. Taken together, our data highlight the potential of BET inhibitors as novel therapeutic agents to treat small cell lung cancer driven by various oncogenic pathways. All studies were conducted in accordance with the GSK Policy on the Care, Welfare and Treatment of Laboratory Animals and were reviewed by the Institutional Animal Care and Use Committee either at GSK or by the ethical review process at the institution where the work was performed. Citation Format: Anastasia Wyce, BaoChau Le, Yuchen Bai, David Soong, Xi-Ping Zhang, Jeanne J. Matteo, Susan Korenchuk, Michael F. Butticello, Ramona Plant, Maureen R. Bleam, Yan Degenhardt, Charles F. McHugh, Christopher Carpenter, Peter J. Tummino, Olena Barbash. Inhibition of BET bromodomain proteins as a therapeutic approach in small cell lung cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2867. doi:10.1158/1538-7445.AM2015-2867

A Chemical Genetic Approach Engineers Selective BET Inhibitors

Abstract Targeting only the first bromodomain of BRD4 is sufficient for displacement of BRD4 from chromatin.

EG-04 * DEVELOPMENT OF HIGHLY POTENT, SELECTIVE BET BROMODOMAIN INHIBITORS THAT ARE CNS PENETRANT AND EFFECTIVE IN RODENT MODELS OF BRAIN CANCER

BRD2, BRD3, and BRD4 are members of the BET family of bromodomain inhibitors that have received intense current interest as novel treatments for cancer. The BET proteins function as epigenetic modulators that recognize specifically marked regions of histones and thereby influence gene transcription. Numerous highly potent, highly selective BET inhibitors have emerged and some (e.g. IBET-762) have advanced to clinical trials. We aimed to develop BET inhibitors as drug for untreatable forms of brain cancer including GBM (glioblastoma multiforme) and metastatic brain cancer. Unfortunately, we found that they are poorly suited as drugs for any indications that require high CNS drug exposure because they have high susceptibility to efflux transporters and low passive cellular permeability. To improve CNS penetration, we employed a structure-based design approach that involved (1) disrupting key transporter pharmacophore points and (2) altering the physical properties to improve cellular permeability. In this manner we have identified EP11313 which has strong potency (BRD4 IC50 7 nM), low efflux (ER <2), high passive cellular permeability (Papp 13.3 × 10−6 cm/s), moderate oral bioavailability in mouse (59%) and high CNS exposure. EP11313 (after daily drug administration for 3 weeks) shows antiproliferative effects in TMZ-resistant cancer patient stem cell lines and dose-dependent tumor reduction in nu/nu mice that were centrally transplanted with luciferase-expressing GBM cell xenografts.

Patent Highlights

Patent Highlights

Doubling the BET

Researchers at GlaxoSmithKline and Rockefeller University have generated a selective BET inhibitor that has therapeutic activity in animal models of sepsis and endotoxic shock. GSK is now investigating the potential of BET inhibitors in a wide variety of therapeutic areas.