Discovery of (S)-1-(1-(4-Chloro-3-fluorophenyl)-2-hydroxyethyl)-4-(2-((1-methyl-1H-pyrazol-5-yl)amino)pyrimidin-4-yl)pyridin-2(1H)-one (GDC-0994), an Extracellular Signal-Regulated Kinase 1/2 (ERK1/2) Inhibitor in Early Clinical Development.

The extracellular signal-regulated kinases ERK1/2 represent an essential node within the RAS/RAF/MEK/ERK signaling cascade that is commonly activated by oncogenic mutations in BRAF or RAS or by upstream oncogenic signaling. While targeting upstream nodes with RAF and MEK inhibitors has proven effective clinically, resistance frequently develops through reactivation of the pathway. Simultaneous targeting of multiple nodes in the pathway, such as MEK and ERK, offers the prospect of enhanced efficacy as well as reduced potential for acquired resistance. Described herein is the discovery and characterization of GDC-0994 (22), an orally bioavailable small molecule inhibitor selective for ERK kinase activity.

Structure-Guided Rescaffolding of Selective Antagonists of BCL-XL.

Because of the promise of BCL-2 antagonists in combating chronic lymphocytic leukemia (CLL) and non-Hodgkin's lymphoma (NHL), interest in additional selective antagonists of antiapoptotic proteins has grown. Beginning with a series of selective, potent BCL-XL antagonists containing an undesirable hydrazone functionality, in silico design and X-ray crystallography were utilized to develop alternative scaffolds that retained the selectivity and potency of the starting compounds.

Discovery of 5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine inhibitors of Erk2.

The discovery and optimization of a series of tetrahydropyridopyrimidine based extracellular signal-regulated kinase (Erks) inhibitors discovered via HTS and structure based drug design is reported. The compounds demonstrate potent and selective inhibition of Erk2 and knockdown of phospho-RSK levels in HepG2 cells and tumor xenografts.

Discovery of 2-{3-[2-(1-isopropyl-3-methyl-1H-1,2-4-triazol-5-yl)-5,6-dihydrobenzo[f]imidazo[1,2-d][1,4]oxazepin-9-yl]-1H-pyrazol-1-yl}-2-methylpropanamide (GDC-0032): a ß-sparing phosphoinositide 3-kinase inhibitor with high unbound exposure and robust in vivo antitumor activity.

Dysfunctional signaling through the phosphoinositide 3-kinase (PI3K)/AKT/mTOR pathway leads to uncontrolled tumor proliferation. In the course of the discovery of novel benzoxepin PI3K inhibitors, we observed a strong dependency of in vivo antitumor activity on the free-drug exposure. By lowering the intrinsic clearance, we derived a set of imidazobenzoxazepin compounds that showed improved unbound drug exposure and effectively suppressed growth of tumors in a mouse xenograft model at low drug dose levels. One of these compounds, GDC-0032 (11l), was progressed to clinical trials and is currently under phase I evaluation as a potential treatment for human malignancies.

Discovery of thiazolobenzoxepin PI3-kinase inhibitors that spare the PI3-kinase ß isoform.

A series of suitable five-membered heterocyclic alternatives to thiophenes within a thienobenzoxepin class of PI3-kinase (PI3K) inhibitors was discovered. Specific thiazolobenzoxepin 8-substitution was identified that increased selectivity over PI3Kß. PI3Kß-sparing compound 27 (PI3Kß Ki,app/PI3Kα Ki,app=57) demonstrated dose-dependent knockdown of pAKT, pPRAS40 and pS6RP in vivo as well as differential effects in an in vitro proliferation cell line screen compared to pan PI3K inhibitor GDC-0941. A new structure-based hypothesis for reducing inhibition of the PI3K ß isoform while maintaining activity against α, δ and γ isoforms is presented.

Discovery of a series of imidazo[4,5-b]pyridines with dual activity at angiotensin II type 1 receptor and peroxisome proliferator-activated receptor-γ.

Mining of an in-house collection of angiotensin II type 1 receptor antagonists to identify compounds with activity at the peroxisome proliferator-activated receptor-γ (PPARγ) revealed a new series of imidazo[4,5-b]pyridines 2 possessing activity at these two receptors. Early availability of the crystal structure of the lead compound 2a bound to the ligand binding domain of human PPARγ confirmed the mode of interaction of this scaffold to the nuclear receptor and assisted in the optimization of PPARγ activity. Among the new compounds, (S)-3-(5-(2-(1H-tetrazol-5-yl)phenyl)-2,3-dihydro-1H-inden-1-yl)-2-ethyl-5-isobutyl-7-methyl-3H-imidazo[4,5-b]pyridine (2l) was identified as a potent angiotensin II type I receptor blocker (IC(50) = 1.6 nM) with partial PPARγ agonism (EC(50) = 212 nM, 31% max) and oral bioavailability in rat. The dual pharmacology of 2l was demonstrated in animal models of hypertension (SHR) and insulin resistance (ZDF rat). In the SHR, 2l was highly efficacious in lowering blood pressure, while robust lowering of glucose and triglycerides was observed in the male ZDF rat.

Exploration of 4,4-disubstituted pyrrolidine-1,2-dicarboxamides as potent, orally active Factor Xa inhibitors with extended duration of action.

Aiming to improve upon previously disclosed Factor Xa inhibitors, a series of 4,4-disubstituted pyrrolidine-1,2-dicarboxamides were explored with the intent of increasing the projected human half-life versus 5 (projected human t(1/2)=6 h). A stereospecific route to compounds containing a 4-aryl-4-hydroxypyrrolidine scaffold was developed, resulting in several compounds that demonstrated an increase in the half-life as well as an increase in the in vitro potency compared to 5. Reported herein is the discovery of 26, containing a (2R,4S)-4-hydroxy-4-(2,4-difluorophenyl)-pyrrolidine scaffold, which is a selective, orally bioavailable, efficacious Factor Xa inhibitor that appears suitable for a once-daily dosing (projected human t(1/2)=23 h).

The discovery of (2R,4R)-N-(4-chlorophenyl)-N- (2-fluoro-4-(2-oxopyridin-1(2H)-yl)phenyl)-4-methoxypyrrolidine-1,2-dicarboxamide (PD 0348292), an orally efficacious factor Xa inhibitor.

Herein, we report the discovery of novel, proline-based factor Xa inhibitors containing a neutral P1 chlorophenyl pharmacophore. Through the additional incorporation of 1-(4-amino-3-fluoro-phenyl)-1H-pyridin-2-one 22, as a P4 pharmacophore, we discovered compound 7 (PD 0348292). This compound is a selective, orally bioavailable, efficacious FXa inhibitor that is currently in phase II clinical trials for the treatment and prevention of thrombotic disorders.

Structure-based drug design of pyrrolidine-1, 2-dicarboxamides as a novel series of orally bioavailable factor Xa inhibitors.

A novel series of pyrrolidine-1,2-dicarboxamides was discovered as factor Xa inhibitors using structure-based drug design. This series consisted of a neutral 4-chlorophenylurea P1, a biphenylsulfonamide P4 and a D-proline scaffold (1, IC(50) = 18 nM). Optimization of the initial hit resulted in an orally bioavailable, subnanomolar inhibitor of factor Xa (13, IC(50) = 0.38 nM), which was shown to be efficacious in a canine electrolytic model of thrombosis with minimal bleeding.

The discovery of fluoropyridine-based inhibitors of the factor VIIa/TF complex--Part 2.

The activated factor VII/tissue factor complex (FVIIa/TF) is known to play a key role in the formation of blood clots. Inhibition of this complex may lead to new antithrombotic drugs. A fluoropyridine-based series of FVIIa/TF inhibitors was discovered which utilized a diisopropylamino group for binding in the S2 and S3 binding pockets of the active site of the enzyme complex. In this series, an enhancement in binding affinity was observed by substitution at the 5-position of the hydroxybenzoic acid sidechain. An X-ray crystal structure indicates that amides at this position may increase inhibitor binding affinity through interactions with the S1'/S2' pocket.

The discovery of fluoropyridine-based inhibitors of the Factor VIIa/TF complex.

The activated Factor VII/tissue factor complex (FVIIa/TF) plays a key role in the formation of blood clots. Inhibition of this complex may lead to new antithrombotic drugs. An X-ray crystal structure of a fluoropyridine-based FVIIa/TF inhibitor bound in the active site of the enzyme complex suggested that incorporation of substitution at the 5-position of the hydroxybenzoic acid side chain could lead to the formation of more potent inhibitors through interactions with the S1'/S2' pocket.

Design, synthesis, and biological activity of potent and selective inhibitors of blood coagulation factor Xa.

Factor Xa (FXa) has materialized as a key enzyme for the intervention of the blood coagulation cascade and for the development of new antithrombotic agents. FXa is the lone enzyme responsible for the production of thrombin and therefore is an attractive target for the control of thrombus formation. We have designed and synthesized a unique series of quinoxalinone FXa inhibitors. This series resulted in 3-[4-[5-((2S,6R)-2,6-dimethylpiperidin-1-yl)pentyl]-3-oxo-3,4-dihydroquinoxolin-2-yl]benzamidine (35) with 0.83 nM activity against FXa and excellent selectivity over similar serine proteases. An X-ray crystal structure of compound 35 bound to trypsin along with molecular modeling has led to a predicted binding conformation of compound 35 in FXa. Compound 35 has also been proven to be efficacious in vivo in both the rabbit veno-venous shunt and dog electrolytic injury models. In addition, it was shown that compound 35 did not significantly increase bleeding times in a rabbit model except at the highest doses and plasma concentrations were elevated in a dose dependent manner following a bolus dose and continuous intravenous infusion.