Optimization of Hydroxyethylamine Transition State Isosteres as Aspartic Protease Inhibitors by Exploiting Conformational Preferences.

NMR conformational analysis of a hydroxyethylamine peptide isostere developed as an aspartic protease inhibitor shows that it is a flexible architecture. Cyclization to form pyrrolidines, piperidines, or morpholines results in a preorganization of the whole system in solution. The resulting conformation is similar to the conformation of the inhibitor in the active site of BACE-1. This entropic gain results in increased affinity for the enzyme when compared with the acyclic system. For morpholines 27 and 29, the combination of steric and electronic factors is exploited to orient substituents toward S1, S1', and S2' pockets both in the solution and in the bound states. These highly preorganized molecules proved to be the most potent compounds of the series. Additionally, the morpholines, unlike the pyrrolidine and piperidine analogues, have been found to be brain penetrant BACE-1 inhibitors.

Totally Endoscopic Robotic Mitral Valve Surgery.

Mitral valve dysfunction can seriously impair patients' lives and may require valve repair or replacement. Surgery can be performed using techniques including sternotomy; right thoracotomy with or without robot assistance; and the totally endoscopic robotic technique, which requires percutaneous techniques, femoral cannulation, and endovascular aortic cross-clamping. The totally endoscopic robotic technique has been facilitated by minimally invasive surgical techniques, the evolution of endoscopic techniques, and the development of surgical robots. These advances have enhanced the view of the surgical field and provide better exposure for the repair or replacement of the mitral valve and subvalvular apparatus. This article describes the totally endoscopic robotic approach to mitral valve surgery as performed at Temple University Hospital, Philadelphia, Pennsylvania.

Synthesis, crystallization, and biological evaluation of an orally active prodrug of gemcitabine.

The design, synthesis, and biological characterization of an orally active prodrug (3) of gemcitabine are described. Additionally, the identification of a novel co-crystal solid form of the compound is presented. Valproate amide 3 is orally bioavailable and releases gemcitabine into the systemic circulation after passing through the intestinal mucosa. The compound has entered clinical trials and is being evaluated as a potential new anticancer agent.

Mitsunobu approach to the synthesis of optically active alpha,alpha-disubstituted amino acids.

Chiral tertiary alpha-hydroxy esters of known stereochemical configuration were transformed to alpha-azido esters by Mitsunobu reaction with HN3. Optimization of this reaction was shown to proceed at room temperature with high chemical yield using 1,1-(azodicarbonyl)dipiperidine (ADDP) and trimethylphosphine (PMe3). Complete inversion of configuration was observed at the alpha-carbon. Several alpha,alpha-disubstituted amino acids were synthesized in high overall chemical yield and optical purity.

Cyclopropanecarboxylic acid esters as potential prodrugs with enhanced hydrolytic stability.

Esters of cyclopropanecarboxylic acid demonstrate a substantial increase in stability under both acid- and base-catalyzed hydrolytic conditions. Comparison of the stability of valacyclovir 13 with the cyclopropane analogue 14 shows that at 40 degrees C and pH 6 the half-life of 14 is >300 h while the value for 13 is 69.7 h. CBS-QB3 calculations on isodesmic reactions for transfer of groups from an alkane to an ester show that a cyclopropyl group provides hyperconjugative stabilization.

Macrocyclic peptidomimetic inhibitors of beta-secretase (BACE): first X-ray structure of a macrocyclic peptidomimetic-BACE complex.

The synthesis of novel macrocyclic peptidomimetic inhibitors of the enzyme BACE1 is described. These macrocycles are derived from a hydroxyethylene core structure. Compound 7 was co-crystallized with BACE1 and the X-ray structure of the complex elucidated at 1.6 Angstrom resolution. This molecule inhibits the production of the Abeta peptide in HEK293 cells overexpressing APP751sw.

Preorganization of the hydroxyethylene dipeptide isostere: the preferred conformation in solution resembles the conformation bound to BACE.

Conformational analysis in solution of beta-secretase inhibitors 1 and 2 by NMR spectroscopy reveals that the hydroxyethylene isostere, an apparently flexible fragment widely used as a scissile bond replacement in aspartic protease inhibitors, exists in one predominant conformation in solution. This preferred conformation is similar to that adopted by the hydroxyethylene core of 1 in complex with beta-secretase and that adopted by hydroxyethylene cores of related compounds when bound to aspartic proteases, indicating that this structural unit is preorganized in solution.

Design and synthesis of tricyclic imidazo[4,5-b]pyridin-2-ones as corticotropin-releasing factor-1 antagonists.

The synthesis and SAR studies of tricyclic imidazo[4,5-b]pyridin-2-ones as human corticotropin-releasing factor receptor (CRF(1)) antagonists are discussed herein. Compound 16g was identified as a functional antagonist that inhibited CRF-stimulated cyclic adenosine monophosphate production and CRF-induced adrenocorticotrophic hormone release. Pharmacokinetics studies in rats showed that 16g was orally bioavailable, had good brain penetration, and had a moderate half-life. In our effort to identify CRF(1) antagonists with improved pharmacokinetics properties, 16g exhibited a favorably lower volume of distribution.

Potent, orally active corticotropin-releasing factor receptor-1 antagonists containing a tricyclic pyrrolopyridine or pyrazolopyridine core.

Two new classes of tricyclic-based corticotropin-releasing factor (CRF(1)) receptor-1 antagonists were designed by constraining known 1H-pyrrolo[2,3-b]pyridine and 1H-pyrazolo[3,4-b]pyridine ligands. Pyrrole- and pyrazole-based molecules 19g and 22a, respectively, were discovered that potently bind the recombinant CRF(1) receptor (K(i) = 3.5, 2.9 nM) and inhibit adrenocorticotropic hormone (ACTH) release from rat pituitary cell culture (IC(50) = 14, 6.8 nM). These compounds show good oral bioavailabity (F = 24%, 7.0%) and serum half-lives in rats (t(1/2) = 6.3, 12 h) and penetrate the rat brain ([brain]/[plasma] = 0.27, 0.52) but tend toward large volumes of distribution (V(D) = 38, 44 L kg(-1)) and rapid clearances (CL = 70, 43 mL min(-1) kg(-1)). When given orally, both the pyrazole and the pyrrole leads dose-dependently inhibit stress-induced ACTH release in vivo. ACTH reductions of 84-86% were observed for 30 mg kg(-1) doses.

Rational design, synthesis, and structure-activity relationships of aryltriazoles as novel corticotropin-releasing factor-1 receptor antagonists.

Following the discovery of the very high binding affinity of 4-anilinopyrimidines against corticotropin-releasing factor receptor-1 (CRF(1)) (e.g., 1, K(i) = 2 nM), a new series of triazoles bearing different groups has been synthesized and evaluated. The compounds were prepared by cyclizations of N-acyl-S-methylisothioureas with alkylhydrazines or by cyclizations with hydrazine followed by alkylation. While members of this series showed potent binding affinity against CRF(1) receptor, there were important differences between the different regio- (7 and 12) and stereoisomeric aryltriazoles where the R(1) or R(2) side chain in 7 has an asymmetric center. In terms of overall potency, aryltriazole analogues such as 7r bearing an N-(alpha-branched benzyl)-N-propylamino side chain were the most potent, followed by analogues such as 7a, with an N-bis(cyclopropyl)methyl-N-propylamino side chain, and analogues such as 7m, with an N-(alpha-branched aliphatic)-N-propylamino side chain. While the N-propyl group was crucial for high potency, we hypothesized that the terminal methyl mimicked the 5-methyl of pyrazolo[1,5-a]pyrimidines 3 and 4. Correlation of the low-energy conformers of compounds of type 3 and 7 generated by computational analyses was very good. The size and shape of the N-alkyl group dramatically changed the potency of the triazoles, which is in contrast to the SAR seen for bicyclic CRF(1) antagonists. In general, the S-enantiomer was much more potent than the corresponding R-isomer. Furthermore, to a limited extent in the aryltriazole series the substituent on the 5-phenyl ring changed the potency up to 9-fold. (S)-1-Methyl-3-[N-(4-fluorophenylpentyl)-N-propyl]amino-5-(2-methoxy-4-dichlorophenyl)-1H-[1,2,4]triazole [(S)-7r] showed very potent binding affinity (K(i) = 2.7 nM) to CRF(1) receptors with an IC(50) of 49 nM in a cAMP inhibition assay.

Biochemical and kinetic characterization of BACE1: investigation into the putative species-specificity for beta- and beta'-cleavage sites by human and murine BACE1.

Beta-amyloid peptides (Abeta) are produced by a sequential cleavage of amyloid precursor protein (APP) by beta- and gamma-secretases. The lack of Abeta production in beta-APP cleaving enzyme (BACE1)(-/-) mice suggests that BACE1 is the principal beta-secretase in mammalian neurons. Transfection of human APP and BACE1 into neurons derived from wild-type and BACE1(-/-) mice supports cleavage of APP at the canonical beta-secretase site. However, these studies also revealed an alternative BACE1 cleavage site in APP, designated as beta', resulting in Abeta peptides starting at Glu11. The apparent inability of human BACE1 to make this beta'-cleavage in murine APP, and vice versa, led to the hypothesis that this alternative cleavage was species-specific. In contrast, the results from human BACE1 transgenic mice demonstrated that the human BACE1 is able to cleave the endogenous murine APP at the beta'-cleavage site. To address this discrepancy, we designed fluorescent resonance energy transfer peptide substrates containing the beta- and beta'-cleavage sites within human and murine APP to compare: (i) the enzymatic efficiency; (ii) binding kinetics of a BACE1 active site inhibitor LY2039911; and (iii) the pharmacological profiles for human and murine recombinant BACE1. Both BACE1 orthologs were able to cleave APP at the beta- and beta'-sites, although with different efficiencies. Moreover, the inhibitory potency of LY2039911 toward recombinant human and native BACE1 from mouse or guinea pig was indistinguishable. In summary, we have demonstrated, for the first time, that recombinant BACE1 can recognize and cleave APP peptide substrates at the postulated beta'-cleavage site. It does not appear to be a significant species specificity to this cleavage.

Conversion of human-selective PPARalpha agonists to human/mouse dual agonists: a molecular modeling analysis.

To understand the species selectivity in a series of alpha-methyl-alpha-phenoxy carboxylic acid PPARalpha/gamma dual agonists (1-11), structure-based molecular modeling was carried out in the ligand binding pockets of both human and mouse PPARalpha. This study suggested that interaction of both 4-phenoxy and phenyloxazole substituents of these ligands with F272 and M279 in mouse PPARalpha leads to the species-specific divergence in ligand binding. Insights obtained in the molecular modeling studies of these key interactions resulted in the ability to convert a human-selective PPARalpha agonist to a human and mouse dual agonist within the same platform.

Design of 2,5-dimethyl-3-(6-dimethyl-4-methylpyridin-3-yl)-7-dipropylaminopyrazolo[1,5-a]pyrimidine (NBI 30775/R121919) and structure--activity relationships of a series of potent and orally active corticotropin-releasing factor receptor antagonists.

We have previously shown that 3-phenylpyrazolo[1,5-a]pyrimidines exemplified by 8 were potent antagonists of the human corticotropin-releasing factor-1 receptor. A series of 3-pyridylpyrazolo[1,5-a]pyrimidines 15, 25-30, 34, and 35 containing a weakly basic pyridine ring at the 3-position of the bicyclic nucleus was designed to reduce lipophilicity from the initial leads such as 7. Here, we showed that these 3-pyridyl compounds exhibited potent antagonists at the human CRF(1) receptor. Moreover, the hydrophilic and weakly basic pyridine moiety increased the water solubility of some analogues. Compound 26 h exhibited good binding affinity at the human CRF(1) receptor with a K(i) value of 3.5 nM. As a functional antagonist, it dose-dependently inhibited CRF-stimulated cAMP production in cells expressing the CRF(1) receptor (IC(50) = 50 nM), and CRF-stimulated ACTH release from cultured rat pituitary cells (IC(50) = 20 nM). 26 h had a log P value of 4.9 and water solubility of greater than 10 mg/mL. Pharmacokinetic studies in rats showed that 26 h was orally bioavailable and able to penetrate into the brain. 26 h has been demonstrated in vivo efficacy in animal behavioral models that measure anxiolytic activity. These results suggest that analogues from this series were potent CRF(1) receptor antagonists with proper physicochemical properties and good pharmacokinetic profiles. 26 h was developed into a clinical compound and exhibited efficacy in patients with major depression.

Synthesis of benzoylpyrimidines as antagonists of the corticotropin-releasing factor-1 receptor.

A series of benzoylpyrimidines derived from the anilinepyrimidine CRF(1) antagonists were synthesized. Several synthetic routes were developed to explore the SAR of this series of compounds. Compounds such as 8d (K(i) = 15 nM) exhibited high binding affinities at the human CRF(1) receptor.

Design and synthesis of 3-(2-pyridyl)pyrazolo[1,5-a]pyrimidines as potent CRF1 receptor antagonists.

A series of 3-(2-pyridyl)pyrazolo[1,5-a]pyrimidines was designed and synthesized as antagonists for the corticotrophin-releasing factor-1 (CRF(1)) receptor. Several compounds such as 20c (K(i)=10 nM) exhibited good binding affinities at the CRF(1) receptor. In addition, 20c had adequate solubility in water.

Optimization of 3-phenylpyrazolo[1,5-a]pyrimidines as potent corticotropin-releasing factor-1 antagonists with adequate lipophilicity and water solubility.

In our efforts to identify potent CRF(1) antagonists with proper physicochemical properties, a series of 3-phenylpyrazolo[1,5-a]pyrimidines bearing polar groups, such as amino, hydroxyl, methoxy, sulfoxide, were designed and synthesized. Several positions of the core structure were identified, where a polar group was tolerated with slight reduction in receptor binding. NBI 30545 (18n) was found to have good binding affinity and potent antagonistic activity at the human CRF(1) receptor. Moreover, this compound had proper lipophilicity (log D = 2.78) and good solubility in water (>10mg/mL), and exhibited good plasma and brain exposure when given orally.

Design and synthesis of alpha-aryloxy-alpha-methylhydrocinnamic acids: a novel class of dual peroxisome proliferator-activated receptor alpha/gamma agonists.

The design and synthesis of the dual peroxisome proliferator activated receptor (PPAR) alpha/gamma agonist (S)-2-methyl-3-[4-[2-(5-methyl-2-thiophen-2-yl-oxazol-4-yl)ethoxy]phenyl]-2-phenoxypropionic acid (2) for the treatment of type 2 diabetes and associated dyslipidemia are described. 2 possesses a potent dual hPPAR alpha/gamma agonist profile (IC(50) = 28 and 10 nM; EC(50) = 9 and 4 nM, respectively, for hPPARalpha and hPPARgamma). In preclinical models, 2 substantially improves insulin sensitivity and potently reverses diabetic hyperglycemia while significantly improving overall lipid homeostasis.

Design, synthesis, and SAR of 2-dialkylamino-4-arylpyrimidines as potent and selective corticotropin-releasing factor(1) (CRF(1)) receptor antagonists.

A series of 2-dialkylamino-4-phenylpyrimidines (7) was designed and synthesized as CRF(1) antagonists. SAR studies of this series resulted in the discovery of potent and selective antagonists 7b and 7n bearing a 4-(2,4,6-trisubstituted-phenyl) ring and a bulky 2-(N-bis(cyclopropane)methyl-N-propyl)amino group.

Design and synthesis of statine-containing BACE inhibitors.

Utilizing structure-based techniques and solid-phase synthesis, statine-based tetrapeptide BACE inhibitors were designed and synthesized using a heptapeptide BACE transition-state mimetic, 1, as the starting point. Structure-activity relationship studies at the P(3), P(2), and P(2)' positions as well as the N-terminal capping group on scaffold 5 led to the discovery of potent inhibitors 27, 32, and 34 (IC(50) <100 nM). In addition, computational analysis and the X-ray structure of BACE-inhibitor 38 are discussed.

Synthesis of 3-phenylpyrazolo[4,3-b]pyridines via a convenient synthesis of 4-amino-3-arylpyrazoles and SAR of corticotropin-releasing factor receptor type-1 antagonists.

3-Phenylpyrazolo[4,3-b]pyridines were synthesized via a cyclization of 4-amino-3-phenylpyrazoles 11-13 with ethyl acetoacetate. These compounds were found to be potent CRF(1) antagonists. The 2-alkylpyrazolo[4,3-b]pyridines were more polar but less active than the corresponding 1-alkyl-isomers.