Discovery of potent and selective PPARα/δ dual antagonists and initial biological studies.

We previously published on the design and synthesis of novel, potent and selective PPARα antagonists suitable for either i.p. or oral in vivo administration for the potential treatment of cancer. Described herein is SAR for a subsequent program, where we set out to identify selective and potent PPARα/δ dual antagonist molecules. Emerging literature indicates that both PPARα and PPARδ antagonism may be helpful in curbing the proliferation of certain types of cancer. This dual antagonism could also be used to study PPARs in other settings. After testing for selective and dual potency, off-target counter screening, metabolic stability, oral bioavailability and associated toxicity, compound 11, the first reported PPARα/δ dual antagonist was chosen for more advanced preclinical evaluation.

In vitro and in vivo pharmacology of NXT629, a novel and selective PPARα antagonist.

Peroxisome-proliferator activated receptors (PPAR) are members of the nuclear hormone receptor superfamily which regulate gene transcription. PPARα is a key regulator of lipid homeostasis and a negative regulator of inflammation. Under conditions of metabolic stress such as fasting or glucose deprivation, PPARα is upregulated in order to control gene expression necessary for processing alternate fuel sources (e.g. fatty acid oxidation) and thereby promote maintenance of cell viability. Clinically, PPARα expression is upregulated in diseased tissues such as melanoma, chronic lymphocytic leukemia, ovarian and prostate cancer. This may allow for cellular proliferation and metastasis. Importantly, genetic knockouts of PPARα have been shown to be protected against tumor growth in a variety of syngeneic tumors models. We hypothesized that a potent and selective PPARα antagonist could represent a novel cancer therapy. Early in our discovery research, we identified NXT629 (Bravo et al., 2014). Herein we describe the pharmacology of NXT629 and demonstrate that it is a potent and selective PPARα antagonist. We identify NXT629 as a valuable tool for use in in vivo assessment of PPARα due to its good systemic exposure following intraperitoneal injection. We explore the in vivo pharmacology of NXT629 and demonstrate that it is efficacious in pharmacodynamic models that are driven by PPARα. Finally, we probe the efficacy of NXT629 in disease models where PPARα knockouts have shown to be protected. We believe that PPARα antagonists will be beneficial in diseases such as ovarian cancer and melanoma where PPARα and fatty acid oxidation may be involved.

5-Lipoxygenase-activating protein (FLAP) inhibitors. Part 4: development of 3-[3-tert-butylsulfanyl-1-[4-(6-ethoxypyridin-3-yl)benzyl]-5-(5-methylpyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethylpropionic acid (AM803), a potent, oral, once daily FLAP inhibitor.

The potent 5-lipoxygenase-activating protein (FLAP) inhibitor 3-[3-tert-butylsulfanyl-1-[4-(6-ethoxypyridin-3-yl)benzyl]-5-(5-methylpyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethylpropionic acid 11cc is described (AM803, now GSK2190915). Building upon AM103 (1) (Hutchinson et al. J. Med Chem.2009, 52, 5803-5815; Stock et al. Bioorg. Med. Chem. Lett. 2010, 20, 213-217; Stock et al. Bioorg. Med. Chem. Lett.2010, 20, 4598-4601), SAR studies centering around the pyridine moiety led to the discovery of compounds that exhibit significantly increased potency in a human whole blood assay measuring LTB(4) inhibition with longer drug preincubation times (15 min vs 5 h). Further studies identified 11cc with a potency of 2.9 nM in FLAP binding, an IC(50) of 76 nM for inhibition of LTB(4) in human blood (5 h incubation) and excellent preclinical toxicology and pharmacokinetics in rat and dog. 11cc also demonstrated an extended pharmacodynamic effect in a rodent bronchoalveolar lavage (BAL) model. This compound has successfully completed phase 1 clinical studies in healthy volunteers and is currently undergoing phase 2 trials in asthmatic patients.

Pharmacology of AM803, a novel selective five-lipoxygenase-activating protein (FLAP) inhibitor in rodent models of acute inflammation.

We evaluated the in vivo pharmacological properties of AM803 3-[3-tert-butylsulfanyl-1-[4-(6-ethoxy-pyridin-3-yl)-benzyl]-5-(5-methyl-pyridin-2-ylmethoxy)-1H-indol-2-yl]-2,2-dimethyl-propionic acid, a selective five-lipoxygenase-activating protein (FLAP) inhibitor, using rat and mouse models of acute inflammation. Oral administration of AM803 (1 mg/kg) resulted in sustained inhibition of ex vivo ionophore-challenged whole blood LTB4 biosynthesis with >90% inhibition for up to 12 h and an EC50 of approximately 7 nM. When rat lungs were challenged in vivo with calcium-ionophore, AM803 inhibited LTB4 and cysteinyl leukotriene (CysLT) production with ED50s of 0.12 mg/kg and 0.37 mg/kg, respectively. The inhibition measured 16 h following a single oral dose of 3 mg/kg was 86% and 41% for LTB4 and CysLTs, respectively. In an acute inflammation setting, AM803 dose-dependently reduced LTB4, CysLTs, plasma protein extravasation and neutrophil influx induced by peritoneal zymosan injection. Finally, AM803 increased survival time in mice exposed to a lethal intravenous injection of platelet activating factor (PAF). The magnitude of effect was similar to that of an inhibitor of five-lipoxygenase (5-LO) and LTA4 hydrolase but superior to a leukotriene CysLT1 receptor antagonist. In summary, AM803 is a novel, potent and selective FLAP inhibitor that has excellent pharmacodynamic properties in vivo and is effective in animal models of acute inflammation and in a model of lethal shock.

Pharmacological blockade of the DP2 receptor inhibits cigarette smoke-induced inflammation, mucus cell metaplasia, and epithelial hyperplasia in the mouse lung.

Prostaglandin D(2) (PGD(2)) is one of a family of biologically active lipids derived from arachidonic acid via the action of COX-1 and COX-2. PGD(2) is released from mast cells and binds primarily to two G protein-coupled receptors, namely DP1 and DP2, the latter also known as chemoattractant receptor-homologous molecule expressed on Th2 cells. DP2 is predominantly expressed on eosinophils, Th2 cells, and basophils, but it is also expressed to a lesser extent on monocytes, mast cells, and epithelial cells. Interaction of PGD(2) and its active metabolites with DP2 results in cellular chemotaxis, degranulation, up-regulation of adhesion molecules, and cytokine production. Chronic obstructive pulmonary disease (COPD) is a chronic progressive inflammatory disease characterized by elevated lung neutrophils, macrophages, and CD8+ T lymphocytes and mucus hypersecretion. Cigarette smoke contributes to the etiology of COPD and was used here as a provoking agent in a murine model of COPD. In an acute model, {2'-[(cyclopropanecarbonyl-ethyl-amino)-methyl]-6-methoxy-4'-trifluoro-methyl-biphenyl-3-yl}-acetic acid, sodium salt (AM156) and (5-{2-[(benzoyloxycarbonyl-ethyl-amino)-methyl]-4-trifluoromethyl-phenyl}-pyridin-3-yl)-acetic acid, sodium salt) (AM206), potent DP2 receptor antagonists, dose-dependently inhibited influx of neutrophils and lymphocytes to smoke-exposed airways. In a subchronic model, AM156 and AM206 inhibited neutrophil and lymphocyte trafficking to the airways. Furthermore, AM156 and AM206 treatment inhibited mucus cell metaplasia and prevented the thickening of the airway epithelial layer induced by cigarette smoke. These data suggest that DP2 receptor antagonism may represent a novel therapy for COPD or other conditions characterized by neutrophil influx, mucus hypersecretion, and airway remodeling.

Novel tricyclic antagonists of the prostaglandin D2 receptor DP2 with efficacy in a murine model of allergic rhinitis.

The synthesis of a series of tricyclic antagonists for the prostaglandin D(2) receptor DP2 (CRTH2) is disclosed. The activities of the compounds were evaluated in a human DP2 binding assay and a human whole blood eosinophil shape change assay. Potential metabolic liabilities of the compounds were addressed through in vitro CYP studies. The lead compound was demonstrated to have efficacy in a mouse model of allergic rhinitis following oral dosing.