Discovery and optimization of thieno[2,3-d]pyrimidines as B-Raf inhibitors.

The serine/threonine specific protein kinase B-Raf is part of the MAPK pathway and is an interesting oncology target. We have identified thieno[2,3-d]pyrimidines as a core scaffold of small molecule B-Raf inhibitors. The SAR of analogs in this series will be described.

Structure-Guided Optimization Provides a Series of TTK Protein Inhibitors with Potent Antitumor Activity.

TTK is an essential spindle assembly checkpoint enzyme in many organisms. It plays a central role in tumor cell proliferation and is aberrantly overexpressed in a wide range of tumor types. We recently reported on a series of potent and selective TTK inhibitors with strong antiproliferative activity in triple negative breast cancer (TNBC) cell lines (8: TTK IC50 = 3.0 nM; CAL-51 IC50 = 84.0 nM). Inspired by previously described potent tricyclic TTK inhibitor 6 (TTK IC50 = 0.9 nM), we embarked on a structure-enabled design and optimization campaign to identify an improved series with excellent potency, TTK selectivity, solubility, CYP inhibition profile, and in vivo efficacy in a TNBC xenograft model. These efforts culminated in the discovery of 25 (TTK IC50 = 3.0 nM; CAL-51 IC50 = 16.0 nM), which showed significant single-agent efficacy when dosed iv in a TNBC xenograft model without body weight loss.

Effect of intramolecular Paternò-Büchi reaction on the thermodynamics and kinetics of nearly degenerate [3,3]-sigmatropic shift in fluxional polycycles.

In reactions with weak dienophiles, cyclooctatetraene (COT) often yields 2:1 adducts possessing the fluxional bicyclo[5.1.0]octadiene moiety. They undergo fast, nearly degenerate Cope rearrangement with an activation barrier similar to that of the parent dihydrobullvalene. Irradiation to excite the carbonyl moiety induces an intramolecular Paterno-Buchi cyclization yielding endo-oxetanes and significantly changing the Cope-averaged NMR spectra. In this paper we examine the effect of skeletal distortion caused by intramolecular [2 + 2]-photoaddition on thermodynamics and the activation barrier of the [3,3]-sigmatropic tautomerism. Our finding is that such a distortion lifts the energetic degeneracy of the two valence tautomers, while not affecting the activation barrier.

Evolution of Cereblon-Mediated Protein Degradation as a Therapeutic Modality.

Many cellular processes and pathways are mediated by the regulation of protein-protein complexes. For example, E3 ubiquitin ligases recruit substrate proteins and transfer a ubiquitin tag to target those proteins for destruction by the proteasome. It has now been shown that this cellular process for protein destruction can be redirected by small molecules in both laboratory and clinical settings. This presents a new paradigm in drug discovery, enabling the rapid removal of target proteins linked to disease. In this Innovations review, we will describe the work done on cereblon as a case study on the different strategies available for targeted protein degradation.

Design and Optimization Leading to an Orally Active TTK Protein Kinase Inhibitor with Robust Single Agent Efficacy.

Triple negative breast cancer (TNBC) is an aggressive disease with high relapse rates and few treatment options. Outlined in previous publications, we identified a series of potent, dual TTK/CLK2 inhibitors with strong efficacy in TNBC xenograft models. Pharmacokinetic properties and kinome selectivity were optimized, resulting in the identification of a new series of potent, selective, and orally bioavailable TTK inhibitors. We describe here the structure-activity relationship of the 2,4-disubstituted-7 H-pyrrolo[2,3- d]pyrimidine series, leading to significant single agent efficacy in a TNBC xenograft model without body weight loss. The design effort evolving an iv-dosed TTK/CLK2 inhibitor to an orally bioavailable TTK inhibitor is described.

Synthetic Lethal Strategy Identifies a Potent and Selective TTK and CLK1/2 Inhibitor for Treatment of Triple-Negative Breast Cancer with a Compromised G1-S Checkpoint.

Historically, phenotypic-based drug discovery has yielded a high percentage of novel drugs while uncovering new tumor biology. CC-671 was discovered using a phenotypic screen for compounds that preferentially induced apoptosis in triple-negative breast cancer cell lines while sparing luminal breast cancer cell lines. Detailed in vitro kinase profiling shows CC-671 potently and selectively inhibits two kinases-TTK and CLK2. Cellular mechanism of action studies demonstrate that CC-671 potently inhibits the phosphorylation of KNL1 and SRp75, direct TTK and CLK2 substrates, respectively. Furthermore, CC-671 causes mitotic acceleration and modification of pre-mRNA splicing leading to apoptosis, consistent with cellular TTK and CLK inhibition. Correlative analysis of genomic and potency data against a large panel of breast cancer cell lines identifies breast cancer cells with a dysfunctional G1-S checkpoint as more sensitive to CC-671, suggesting synthetic lethality between G1-S checkpoint and TTK/CLK2 inhibition. Furthermore, significant in vivo CC-671 efficacy was demonstrated in two cell line-derived and one patient tumor-derived xenograft models of triple-negative breast cancer (TNBC) following weekly dosing. These findings are the first to demonstrate the unique inhibitory combination activity of a dual TTK/CLK2 inhibitor that preferably kills TNBC cells and shows synthetic lethality with a compromised G1-S checkpoint in breast cancer cell lines. On the basis of these data, CC-671 was moved forward for clinical development as a potent and selective TTK/CLK2 inhibitor in a subset of patients with TNBC. Mol Cancer Ther; 17(8); 1727-38. ©2018 AACR.

The Discovery of a Dual TTK Protein Kinase/CDC2-Like Kinase (CLK2) Inhibitor for the Treatment of Triple Negative Breast Cancer Initiated from a Phenotypic Screen.

Triple negative breast cancer (TNBC) remains a serious unmet medical need with discouragingly high relapse rates. We report here the synthesis and structure-activity relationship (SAR) of a novel series of 2,4,5-trisubstituted-7H-pyrrolo[2,3-d]pyrimidines with potent activity against TNBC tumor cell lines. These compounds were discovered from a TNBC phenotypic screen and possess a unique dual inhibition profile targeting TTK (mitotic exit) and CLK2 (mRNA splicing). Design and optimization, driven with a TNBC tumor cell assay, identified potent and selective compounds with favorable in vitro and in vivo activity profiles and good iv PK properties. This cell-based driven SAR produced compounds with strong single agent in vivo efficacy in multiple TNBC xenograft models without significant body weight loss. These data supported the nomination of CC-671 into IND-enabling studies as a single agent TNBC therapy.

Discovery of mammalian target of rapamycin (mTOR) kinase inhibitor CC-223.

We report here the synthesis and structure-activity relationship (SAR) of a novel series of mammalian target of rapamycin (mTOR) kinase inhibitors. A series of 4,6- or 1,7-disubstituted-3,4-dihydropyrazino[2,3-b]pyrazine-2(1H)-ones were optimized for in vivo efficacy. These efforts resulted in the identification of compounds with excellent mTOR kinase inhibitory potency, with exquisite kinase selectivity over the related lipid kinase PI3K. The improved PK properties of this series allowed for exploration of in vivo efficacy and ultimately the selection of CC-223 for clinical development.

Optimization of a Series of Triazole Containing Mammalian Target of Rapamycin (mTOR) Kinase Inhibitors and the Discovery of CC-115.

We report here the synthesis and structure-activity relationship (SAR) of a novel series of triazole containing mammalian target of rapamycin (mTOR) kinase inhibitors. SAR studies examining the potency, selectivity, and PK parameters for a series of triazole containing 4,6- or 1,7-disubstituted-3,4-dihydropyrazino[2,3-b]pyrazine-2(1H)-ones resulted in the identification of triazole containing mTOR kinase inhibitors with improved PK properties. Potent compounds from this series were found to block both mTORC1(pS6) and mTORC2(pAktS473) signaling in PC-3 cancer cells, in vitro and in vivo. When assessed in efficacy models, analogs exhibited dose-dependent efficacy in tumor xenograft models. This work resulted in the selection of CC-115 for clinical development.

Factor VIIa inhibitors: a prodrug strategy to improve oral bioavailability.

We have developed a series of potent and selective factor VIIa inhibitors based on the 2-[5-(5-carbamimidoyl-1H-benzoimidazol-2-yl)-6-hydroxy-biphenyl-3-yl]-succinic acid scaffold. These amidine-containing compounds have low oral bioavailability. Herein, we describe our efforts to improve the oral bioavailability of the parent amidine via a prodrug strategy where the amidine basicity and polarity were reduced with either an alkoxy-amidine or a carbamate prodrug.

Potent 4-amino-5-azaindole factor VIIa inhibitors.

The 4-amino-5-azaindole as an amidino-benzimidazole replacement is described. A series of potent and selective analogs were discovered and showed desirable ex vivo efficacy as measured by PT.

Efforts toward oral bioavailability in factor VIIa inhibitors.

Efforts toward developing orally bioavailable factor VIIa inhibitors starting from parenteral lead compound 1 are described. SAR resulted in improved physicochemical properties, leading to enhanced oral absorption in rat.

Novel 5-azaindole factor VIIa inhibitors.

The discovery and development of 5-azaindole factor VIIa inhibitors will be described.