George D. Hartman

Inhibition of farnesyltransferase induces regression of mammary and salivary carcinomas in ras transgenic mice.

For Ras oncoproteins to transform mammalian cells, they must be post-translationally modified with a farnesyl group in a reaction catalysed by the enzyme farnesyl-protein transferase (FPTase). Inhibitors of FPTase have therefore been proposed as anti-cancer agents. We show that L-744,832, which mimics the CaaX motif to which the farnesyl group is added, is a potent and selective inhibitor of FPTase. In MMTV-v-Ha-ras mice bearing palpable tumours, daily administration of L-744,832 caused tumour regression. Following cessation of treatment, tumours reappeared, the majority of which regressed upon retreatment. No systemic toxicity was found upon necropsy of L-744,832-treated mice. This first demonstration of anti-FPTase-mediated tumour regression suggests that FPTase inhibitors may be safe and effective anti-tumour agents in some cancers.

HMG CoA-reductase inhibitors

SummaryLovastatin and simvastatin are the 2 best-known members of the class of hypolipidaemic agents known as HMG CoA reductase inhibitors. Clinical experience with lovastatin includes over 5000 patients, 700 of whom have been treated for 2 years or more, and experience with simvastatin includes over 3500 patients, of whom 350 have been treated for 18 months or more. Lovastatin has been marketed in the United States for over 6 months. Both agents show substantial clinical efficacy, with reductions in total cholesterol of over 30% and in LDL-cholesterol of 40% in clinical studies. Modest increases in HDL-cholesterol levels of about 10% are also reported. Clinical tolerability of both agents has been good, with fewer than 3% of patients withdrawn from treatment because of clinical adverse experiences. Ophthalmological examinations in over 1100 patients treated with one or the other agent have revealed no evidence of significant short term (up to 2 years) cataractogenic potential. One to 2% of patients have elevations of serum transaminases to greater than 3 times the upper limit of normal. These episodes are asymptomatic and reversible when therapy is discontinued. Minor elevations of creatine kinase levels are reported in about 5% of patients. Myopathy, associated in some cases with myoglobinuria, and in 2 cases with transient renal failure, has been rarely reported with lovastatin, especially in patients concomitantly treated with cyclosporin, gemfibrozil or niacin. Lovastatin and simvastatin are both effective and well-tolerated agents for lowering elevated levels of serum cholesterol. As wider use confirms their safety profile, they will gain increasing importance in the therapeutic approach to hypercholesterolaemia and its consequences.

Discovery of the dual orexin receptor antagonist [(7R)-4-(5-chloro-1,3-benzoxazol-2-yl)-7-methyl-1,4-diazepan-1-yl][5-methyl-2-(2H-1,2,3-triazol-2-yl)phenyl]methanone (MK-4305) for the treatment of insomnia.

Despite increased understanding of the biological basis for sleep control in the brain, few novel mechanisms for the treatment of insomnia have been identified in recent years. One notable exception is inhibition of the excitatory neuropeptides orexins A and B by design of orexin receptor antagonists. Herein, we describe how efforts to understand the origin of poor oral pharmacokinetics in a leading HTS-derived diazepane orexin receptor antagonist led to the identification of compound 10 with a 7-methyl substitution on the diazepane core. Though 10 displayed good potency, improved pharmacokinetics, and excellent in vivo efficacy, it formed reactive metabolites in microsomal incubations. A mechanistic hypothesis coupled with an in vitro assay to assess bioactivation led to replacement of the fluoroquinazoline ring of 10 with a chlorobenzoxazole to provide 3 (MK-4305), a potent dual orexin receptor antagonist that is currently being tested in phase III clinical trials for the treatment of primary insomnia.

Design, Synthesis, and Evaluation of a Novel 4-Aminomethyl-4-fluoropiperidine as a T-Type Ca2+ Channel Antagonist

The novel T-type antagonist ( S)- 5 has been prepared and evaluated in in vitro and in vivo assays for T-type calcium ion channel activity. Structural modification of the piperidine leads 1 and 2 afforded the fluorinated piperidine ( S)- 5, a potent and selective antagonist that displayed in vivo CNS efficacy without adverse cardiovascular effects.

Kinesin spindle protein (KSP) inhibitors. 9. Discovery of (2S)-4-(2,5-difluorophenyl)-n-[(3R,4S)-3-fluoro-1-methylpiperidin-4-yl]-2-(hydroxymethyl)-N-methyl-2-phenyl-2,5-dihydro-1H-pyrrole-1-carboxamide (MK-0731) for the treatment of taxane-refractory cancer.

Inhibition of kinesin spindle protein (KSP) is a novel mechanism for treatment of cancer with the potential to overcome limitations associated with currently employed cytotoxic agents. Herein, we describe a C2-hydroxymethyl dihydropyrrole KSP inhibitor ( 11) that circumvents hERG channel binding and poor in vivo potency, issues that limited earlier compounds from our program. However, introduction of the C2-hydroxymethyl group caused 11 to be a substrate for cellular efflux by P-glycoprotein (Pgp). Utilizing knowledge garnered from previous KSP inhibitors, we found that beta-fluorination modulated the p K a of the piperidine nitrogen and reduced Pgp efflux, but the resulting compound ( 14) generated a toxic metabolite in vivo. Incorporation of fluorine in a strategic, metabolically benign position by synthesis of an N-methyl-3-fluoro-4-(aminomethyl)piperidine urea led to compound 30 that has an optimal in vitro and metabolic profile. Compound 30 (MK-0731) was recently studied in a phase I clinical trial in patients with taxane-refractory solid tumors.

Discovery of 1,4-Substituted Piperidines as Potent and Selective Inhibitors of T-Type Calcium Channels

The discovery of a novel series of potent and selective T-type calcium channel antagonists is reported. Initial optimization of high-throughput screening leads afforded a 1,4-substituted piperidine amide 6 with good potency and limited selectivity over hERG and L-type channels and other off-target activities. Further SAR on reducing the basicity of the piperidine and introducing polarity led to the discovery of 3-axial fluoropiperidine 30 with a significantly improved selectivity profile. Compound 30 showed good oral bioavailability and brain penetration across species. In a rat genetic model of absence epilepsy, compound 30 demonstrated a robust reduction in the number and duration of seizures at 33 nM plasma concentration, with no cardiovascular effects at up to 5.6 microM. Compound 30 also showed good efficacy in rodent models of essential tremor and Parkinsons disease. Compound 30 thus demonstrates a wide margin between CNS and peripheral effects and is a useful tool for probing the effects of T-type calcium channel inhibition.

Discovery of [(2R,5R)-5-{[(5-fluoropyridin-2-yl)oxy]methyl}-2-methylpiperidin-1-yl][5-methyl-2-(pyrimidin-2-yl)phenyl]methanone (MK-6096): a dual orexin receptor antagonist with potent sleep-promoting properties.

Insomnia is a common disorder that can be comorbid with other physical and psychological illnesses. Traditional management of insomnia relies on general central nervous system (CNS) suppression using GABA modulators. Many of these agents fail to meet patient needs with respect to sleep onset, maintenance, and next‐day residual effects and have issues related to tolerance, memory disturbances, and balance. Orexin neuropeptides are central regulators of wakefulness, and orexin antagonism has been identified as a novel mechanism for treating insomnia with clinical proof of concept. Herein we describe the discovery of a series of α‐methylpiperidine carboxamide dual orexin 1 and orexin 2 receptor (OX1R/OX2R) antagonists (DORAs). The design of these molecules was inspired by earlier work from this laboratory in understanding preferred conformational properties for potent orexin receptor binding. Minimization of 1,3‐allylic strain interactions was used as a design principle to synthesize 2,5‐disubstituted piperidine carboxamides with axially oriented substituents including DORA 28. DORA 28 (MK‐6096) has exceptional in vivo activity in preclinical sleep models, and has advanced into phase II clinical trials for the treatment of insomnia.

Farnesyltransferase inhibitors versus Ras inhibitors

Over the past few years, the idea that farnesyl-protein transferase (FPTase) inhibitors might be effective antiproliferative/antitumor agents has been realized in studies of cultured cells and in rodent models of cancer. Most of the studies with FPTase inhibitors have focused on inhibiting the growth of ras-transformed cells in vitro or the growth of ras-dependent tumors in mice. More recently, it has been recognized that the antiproliferative effect of FPTase inhibitors may extend beyond ras-driven tumors. It now seems likely that the ability of FPTase inhibitors to reverse the malignant phenotype results, at least in part, from inhibiting the farnesylation of proteins other than Ras.

Dual In Vivo Quantification of Integrin-targeted and Protease-activated Agents in Cancer Using Fluorescence Molecular Tomography (FMT)

PurposeIntegrins, especially αvβ3 and αvβ5, are upregulated in tumor cells and activated endothelial cells and as such, serve as cancer biomarkers. We developed a novel near-infrared-labeled optical agent for the in vivo detection and quantification of αvβ3/αvβ5.ProceduresA small peptidomimetic αvβ3 antagonist was synthesized, coupled to a near-infrared fluorescent (NIRF) dye, and tested for binding specificity using integrin-overexpressing cells, inhibition of vitronectin-mediated cell attachment, binding to tumor and endothelial cells in vitro, and competition studies. Pharmacokinetics, biodistribution, specificity of tumor targeting, and the effect of an antiangiogenic treatment were assessed in vivo.ResultsThe integrin NIRF agent showed strong selectivity towards αvβ3/αvβ5in vitro and predominant tumor distribution in vivo, allowing noninvasive and real-time quantification of integrin signal in tumors. Antiangiogenic treatment significantly inhibited integrin signal in vivo but had no effect on a cathepsin-cleavable NIR agent. Simultaneous imaging revealed different patterns of distribution reflecting the underlying differences in integrin and cathepsin biology during tumor progression.ConclusionsNIRF-labeled integrin antagonists allow noninvasive molecular fluorescent imaging and quantification of tumors in vivo, improving and providing more refined approaches for cancer detection and treatment monitoring.

Discovery of a potent, CNS-penetrant orexin receptor antagonist based on an n,n-disubstituted-1,4-diazepane scaffold that promotes sleep in rats.

Silent Night: Antagonism of the orexin (or hypocretin) system has recently been identified as a novel mechanism for the treatment of insomnia. Herein, we describe discovery of a dual (OX1R/OX2R) orexin receptor antagonist featuring a 1,4‐diazepane central constraint that blocks orexin signaling in vivo. In telemetry‐implanted rats, oral administration of this antagonist produced a decrease in wakefulness, while increasing REM and non‐REM sleep.