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Activation of the glucagon-like peptide-1 (GLP-1) receptor stimulates insulin release, lowers plasma glucose levels, delays gastric emptying, increases satiety, suppresses food intake, and affords weight loss in humans. These beneficial attributes have made peptide-based agonists valuable tools for the treatment of type 2 diabetes mellitus and obesity. However, efficient, and consistent delivery of peptide agents generally requires subcutaneous injection, which can reduce patient utilization. Traditional orally absorbed small molecules for this target may offer improved patient compliance as well as the opportunity for co-formulation with other oral therapeutics. Herein, we describe an SAR investigation leading to small-molecule GLP-1 receptor agonists that represent a series that parallels the recently reported clinical candidate danuglipron. In the event, identification of a benzyloxypyrimidine lead, using a sensitized high-throughput GLP-1 agonist assay, was followed by optimization of the SAR using substituent modifications analogous to those discovered in the danuglipron series. A new series of 6-azaspiro[2.5]octane molecules was optimized into potent GLP-1 agonists. Information gleaned from cryogenic electron microscope structures was used to rationalize the SAR of the optimized compounds., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

The α7 nicotinic acetylcholine receptor is a calcium permeable, ligand-gated ion channel that modulates synaptic transmission in the hippocampus, thalamus, and cerebral cortex. Previously disclosed work described PNU-120596 that acts as a powerful positive allosteric modulator of the α7 nicotinic acetylcholine receptor. The initial structure-activity relationships around PNU-120596 were gleaned from screening a large thiazole library. Independent systematic examination of the aryl and heteroaryl groups resulted in compounds with enhanced potency and improved physico-chemical properties culminating in the identification of 16 (PHA-758454). In the presence of acetylcholine, 16 enhanced evoked currents in rat hippocampal neurons. In a rat model of impaired sensory gating, treatment with 16 led to a reversal of the gating deficit in a dose-dependent manner. These results demonstrate that aryl heteroaryl ureas, like compound 16, may be useful tools for continued exploration of the unique biology of the α7 nicotinic acetylcholine receptor., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Our previous work on the optimization of a new class of small molecule PCSK9 mRNA translation inhibitors focused on empirical optimization of the amide tail region of the lead PF-06446846 (1). This work resulted in compound 3 that showed an improved safety profile. We hypothesized that this improvement was related to diminished binding of 3 to non-translating ribosomes and an apparent improvement in transcript selectivity. Herein, we describe our efforts to further optimize this series of inhibitors through modulation of the heterocyclic head group and the amine fragment. Some of the effort was guided by an emerging cryo electron microscopy structure of the binding mode of 1 in the ribosome. These efforts led to the identification of 15 that was deemed suitable for evaluation in a humanized PCSK9 mouse model and a rat toxicology study. Compound 15 demonstrated a dose dependent reduction of plasma PCSK9 levels. The rat toxicological profile was not improved over that of 1, which precluded 15 from further consideration as a clinical candidate., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. All authors were employees of Pfizer when this work was completed., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Peptide agonists of the glucagon-like peptide-1 receptor (GLP-1R) have revolutionized diabetes therapy, but their use has been limited because they require injection. Herein, we describe the discovery of the orally bioavailable, small-molecule, GLP-1R agonist PF-06882961 (danuglipron). A sensitized high-throughput screen was used to identify 5-fluoropyrimidine-based GLP-1R agonists that were optimized to promote endogenous GLP-1R signaling with nanomolar potency. Incorporation of a carboxylic acid moiety provided considerable GLP-1R potency gains with improved off-target pharmacology and reduced metabolic clearance, ultimately resulting in the identification of danuglipron. Danuglipron increased insulin levels in primates but not rodents, which was explained by receptor mutagensis studies and a cryogenic electron microscope structure that revealed a binding pocket requiring a primate-specific tryptophan 33 residue. Oral administration of danuglipron to healthy humans produced dose-proportional increases in systemic exposure (NCT03309241). This opens an opportunity for oral small-molecule therapies that target the well-validated GLP-1R for metabolic health.

Non-enzymatic dynamic kinetic resolution (DKR) of secondary alcohols by enantioselective acylation using an isothiourea-derived HyperBTM catalyst and racemization of slowly reacting alcohol by Bäckvall's ruthenium complex is reported. The DKR approach features high enantioselectivities (up to 99:1), employs easy-to-handle crystalline 4-nitrophenyl isobutyrate as the acylating reagent, and proceeds at room temperature and under an ambient atmosphere. The stereoinduction model featuring cation-π system interactions between the acylated HyperBTM catalyst and π electrons of an alcohol aryl subunit has been elaborated by DFT calculations.

Sickle cell disease (SCD) is a genetic disorder caused by a single point mutation (β6 Glu → Val) on the β-chain of adult hemoglobin (HbA) that results in sickled hemoglobin (HbS). In the deoxygenated state, polymerization of HbS leads to sickling of red blood cells (RBC). Several downstream consequences of polymerization and RBC sickling include vaso-occlusion, hemolytic anemia, and stroke. We report the design of a noncovalent modulator of HbS, clinical candidate PF-07059013 ( 23 ). The seminal hit molecule was discovered by virtual screening and confirmed through a series of biochemical and biophysical studies. After a significant optimization effort, we arrived at 23 , a compound that specifically binds to Hb with nanomolar affinity and displays strong partitioning into RBCs. In a 2-week multiple dose study using Townes SCD mice, 23 showed a 37.8% (±9.0%) reduction in sickling compared to vehicle treated mice. 23 (PF-07059013) has advanced to phase 1 clinical trials.

DNA-encoded library (DEL) technology has the potential to dramatically expedite hit identification in drug discovery owing to its ability to perform protein affinity selection with millions or billions of molecules in a few experiments. To expand the molecular diversity of DEL, it is critical to develop different types of DNA-encoded transformations that produce billions of molecules with distinct molecular scaffolds. Sequential functionalization of multiple C-H bonds provides a unique avenue for creating diversity and complexity from simple starting materials. However, the use of water as solvent, the presence of DNA, and the extremely low concentration of DNA-encoded coupling partners (0.001 M) have hampered the development of DNA-encoded C(sp 3 )-H activation reactions. Herein, we report the realization of palladium-catalyzed C(sp 3 )-H arylation of aliphatic carboxylic acids, amides and ketones with DNA-encoded aryl iodides in water. Notably, the present method enables the use of alternative sets of monofunctional building blocks, providing a linchpin to facilitate further setup for DELs. Furthermore, the C-H arylation chemistry enabled the on-DNA synthesis of structurally-diverse scaffolds containing enriched C(sp 3 ) character, chiral centers, cyclopropane, cyclobutane, and heterocycles., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Preparative reactions that occur efficiently under dilute, buffered, aqueous conditions in the presence of biomolecules find application in ligation, peptide synthesis, and polynucleotide synthesis and sequencing. However, the identification of functional groups or reagents that are mutually reactive with one another, but unreactive with biopolymers and water, is challenging. Shown here are cobalt catalysts that react with alkenes under dilute, aqueous, buffered conditions and promote efficient cycloisomerization and formal Friedel-Crafts reactions. The constraining conditions of bioorthogonal chemistry are beneficial for reaction efficiency as superior conversion at low catalyst concentration is obtained and competent rates in dilute conditions are maintained. Efficiency at high dilution in the presence of buffer and nucleobases suggests that these reaction conditions may find broad application., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

DNA encoded libraries (DEL) have shown promise as a valuable technology for democratizing the hit discovery process. Although DEL provides relatively inexpensive access to libraries of unprecedented size, their production has been hampered by the idiosyncratic needs of the encoding DNA tag relegating DEL compatible chemistry to dilute aqueous environments. Recently reversible adsorption to solid support (RASS) has been demonstrated as a promising method to expand DEL reactivity using standard organic synthesis protocols. Here we demonstrate a suite of on-DNA chemistries to incorporate medicinally relevant and C-S, C-P and N-S linkages into DELs, which are underrepresented in the canonical methods., (© 2020 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

DNA Encoded Libraries have proven immensely powerful tools for lead identification. The ability to screen billions of compounds at once has spurred increasing interest in DEL development and utilization. Although DEL provides access to libraries of unprecedented size and diversity, the idiosyncratic and hydrophilic nature of the DNA tag severely limits the scope of applicable chemistries. It is known that biomacromolecules can be reversibly, noncovalently adsorbed and eluted from solid supports, and this phenomenon has been utilized to perform synthetic modification of biomolecules in a strategy we have described as reversible adsorption to solid support (RASS). Herein, we present the adaptation of RASS for a DEL setting, which allows reactions to be performed in organic solvents at near anhydrous conditions opening previously inaccessible chemical reactivities to DEL. The RASS approach enabled the rapid development of C(sp 2 )-C(sp 3 ) decarboxylative cross-couplings with broad substrate scope, an electrochemical amination (the first electrochemical synthetic transformation performed in a DEL context), and improved reductive amination conditions. The utility of these reactions was demonstrated through a DEL-rehearsal in which all newly developed chemistries were orchestrated to afford a compound rich in diverse skeletal linkages. We believe that RASS will offer expedient access to new DEL reactivities, expanded chemical space, and ultimately more drug-like libraries.

A series of N-(piperidin-3-yl)-N-(pyridin-2-yl)piperidine/piperazine-1-carboxamides were identified as small molecule PCSK9 mRNA translation inhibitors. Analogues from this new chemical series, such as 4d and 4g, exhibited improved PCSK9 potency, ADME properties, and in vitro safety profiles when compared to earlier lead structures., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Two chemotypes were examined in vitro with CYPs 3A4 and 2C19 by molecular docking, metabolic profiles, and intrinsic clearance deuterium isotope effects with specifically deuterated form to assess the potential for enhancement of pharmacokinetic parameters. The results show the complexity of deuteration as an approach for pharmacokinetic enhancement when CYP enzymes are involved in metabolic clearance. With CYP3A4 the rate limiting step was chemotype-dependent. With one chemotype no intrinsic clearance deuterium isotope effect was observed with any deuterated form, whereas with the other chemotype the rate limiting step was isotopically sensitive, and the magnitude of the intrinsic clearance isotope effect was dependent on the position(s) and extent of deuteration. Molecular docking and metabolic profiles aided in identifying sites for deuteration and predicted the possibility for metabolic switching. However, the potential for an isotope effect on the intrinsic clearance cannot be predicted and must be established by examining select deuterated versions of the chemotypes. The results show how in a deuteration strategy molecular docking, in-vitro metabolic profiles, and intrinsic clearance assessments with select deuterated versions of new chemical entities can be applied to determine the potential for pharmacokinetic enhancement in a discovery setting. They also help explain the substantial failures reported in the literature of deuterated versions of drugs to elicit a systemic enhancement on pharmacokinetic parameters., Competing Interests: All authors were paid employees of Pfizer Inc at the time this work was done. Pfizer Inc provided all funding for this research. Some of the authors are owners of PFE stocks or shares and are named inventors on Pfizer patents. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

The optimization of a new class of small molecule PCSK9 mRNA translation inhibitors is described. The potency, physicochemical properties, and off-target pharmacology associated with the hit compound (1) were improved by changes to two regions of the molecule. The last step in the synthesis of the congested amide center was enabled by three different routes. Subtle structural changes yielded significant changes in pharmacology and off-target margins. These efforts led to the identification of 7l and 7n with overall profiles suitable for in vivo evaluation. In a 14-day toxicology study, 7l demonstrated an improved safety profile vs lead 7f. We hypothesize that the improved safety profile is related to diminished binding of 7l to nontranslating ribosomes and an apparent improvement in transcript selectivity due to the lower strength of 7l stalling of off-target proteins.

C-X-C chemokine receptor type 7 (CXCR7) is involved in cardiac and immune pathophysiology. We report the discovery of a novel 1,4-diazepine CXCR7 modulator, demonstrating for the first time the role of pharmacological CXCR7 intervention in cardiac repair. Structure-activity-relationship (SAR) studies demonstrated that a net reduction in lipophilicity (log D) and an incorporation of saturated ring systems yielded compounds with good CXCR7 potencies and improvements in oxidative metabolic stability in human-liver microsomes (HLM). Tethering an ethylene amide further improved the selectivity profile (e.g., for compound 18, CXCR7 K i = 13 nM, adrenergic α 1a K b > 10 000 nM, and adrenergic β 2 K b > 10 000 nM). The subcutaneous administration of 18 in mice led to a statistically significant increase in circulating concentrations of plasma stromal-cell-derived factor 1α (SDF-1α) of approximately 2-fold. Chronic dosing of compound 18 in a mouse model of isoproterenol-induced cardiac injury further resulted in a statistically significant reduction of cardiac fibrosis.

[This corrects the article DOI: 10.1371/journal.pbio.2001882.].

A novel series of morpholine-based nonsteroidal mineralocorticoid receptor antagonists is reported. Starting from a pyrrolidine HTS hit 9 that possessed modest potency but excellect selectivity versus related nuclear hormone receptors, a series of libraries led to identification of morpholine lead 10. After further optimization, cis disubstituted morpholine 22 was discovered, which showed a 45-fold boost in binding affinity and corresponding functional potency compared to 13. While 22 had high clearance in rat, it provided sufficient exposure at high doses to favorably assess in vivo efficacy (increased urinary Na + /K + ratio) and safety. In contrast to rat, the dog and human MetID and PK profiles of 22 were adequate, suggesting that it could be suitable as a potential clinical asset.

Targeting of the human ribosome is an unprecedented therapeutic modality with a genome-wide selectivity challenge. A liver-targeted drug candidate is described that inhibits ribosomal synthesis of PCSK9, a lipid regulator considered undruggable by small molecules. Key to the concept was the identification of pharmacologically active zwitterions designed to be retained in the liver. Oral delivery of the poorly permeable zwitterions was achieved by prodrugs susceptible to cleavage by carboxylesterase 1. The synthesis of select tetrazole prodrugs was crucial. A cell-free in vitro translation assay containing human cell lysate and purified target mRNA fused to a reporter was used to identify active zwitterions. In vivo PCSK9 lowering by oral dosing of the candidate prodrug and quantification of the drug fraction delivered to the liver utilizing an oral positron emission tomography 18 F-isotopologue validated our liver-targeting approach., (© 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Proprotein convertase subtilisin/kexin type 9 (PCSK9) plays a key role in regulating the levels of plasma low-density lipoprotein cholesterol (LDL-C). Here, we demonstrate that the compound PF-06446846 inhibits translation of PCSK9 by inducing the ribosome to stall around codon 34, mediated by the sequence of the nascent chain within the exit tunnel. We further show that PF-06446846 reduces plasma PCSK9 and total cholesterol levels in rats following oral dosing. Using ribosome profiling, we demonstrate that PF-06446846 is highly selective for the inhibition of PCSK9 translation. The mechanism of action employed by PF-06446846 reveals a previously unexpected tunability of the human ribosome that allows small molecules to specifically block translation of individual transcripts.

A new catalyst for the dynamic kinetic resolution of azole hemiaminals has been developed using late-stage structural modifications of the tert-leucinol-derived chiral subunit of DMAP species.

A versatile synthesis of pyridazine-based small molecule α-helix mimetics (A) is presented. Modular C-C, C-N, and C-O bond-forming reactions allow for the inclusion of a variety of aliphatic, basic, aromatic, and heteroaromatic side chain moieties. This robust synthesis is suitable for the preparation of small pyridazine-based libraries.

We report a modular three-component dynamic kinetic resolution (DKR) that affords enantiomerically enriched hemiaminal esters derived from azoles and aldehydes. The novel and scalable reaction can be used to synthesize valuable substituted azoles in a regioselective manner by capping (e.g., acylation) of the equilibrating azole-aldehyde adduct. With the use of a prolinol-derived DMAP catalyst as the chiral Lewis base, the products can be obtained in high chemical yield and with high enantiomeric excess. The DKR was performed on a multikilogram scale to produce a tetrazole prodrug fragment for a leading clinical candidate that posed formidable synthesis challenges.

Cyclic constraints are incorporated into an 11-residue analogue of the N-terminus of glucagon-like peptide-1 (GLP-1) to investigate effects of structure on agonist activity. Cyclization through linking side chains of residues 2 and 5 or 5 and 9 produced agonists at nM concentrations in a cAMP assay. 2D NMR and CD spectra revealed an N-terminal β-turn and a C-terminal helix that differentially influenced affinity and agonist potency. These structures can inform development of small molecule agonists of the GLP-1 receptor to treat type 2 diabetes.

A novel series of nonsteroidal mineralocorticoid receptor (MR) antagonists identified as part of our strategy to follow up on the clinical candidate PF-03882845 (2) is reported. Optimization departed from the previously described pyrazoline 3a and focused on improving the selectivity for MR versus the progesterone receptor (PR) as an approach to avoid potential sex-hormone-related adverse effects and improving biopharmaceutical properties. From this effort, (R)-14c was identified as a potent nonsteroidal MR antagonist (IC50 = 4.5 nM) with higher than 500-fold selectivity versus PR and other related nuclear hormone receptors, with improved solubility as compared to 2 and pharmacokinetic properties suitable for oral administration. (R)-14c was evaluated in vivo using the increase of urinary Na(+)/K(+) ratio in rat as a mechanism biomarker of MR antagonism. Treatment with (R)-14c by oral administration resulted in significant increases in urinary Na(+)/K(+) ratio and demonstrated this novel compound acts as an MR antagonist.

An enantioselective organocatalytic process for the one-step synthesis of poly-substituted 1,2,4-triazolines is reported. The heterocycle formation is believed to go through a step-wise mechanism of nucleophilic addition of an azlactone to an azodicarboxylate in the presence of an organic base catalyst, followed by a TMSCHN2 mediated heterocyclization. Both theoretical calculations and experimental evidence suggest the pre-organization of the transition state for the chirality determining step via a unique 7-membered intramolecular hydrogen bonding.

Hit-to-lead medicinal chemistry efforts are described starting from a screening hit 1, leading to a new class of aryl sulfonamide-based MR antagonist, exemplified by 17, that possesses favourable MR binding affinity, selectivity profile against closely related NHRs, physicochemical properties and metabolic stability., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Two regioselective and complementary hydroarylation reactions of an unsymmetrical cyclic olefin have been developed. The products can be transformed in one step into constrained γ-amino acids. Regioselective arylation of Vince lactam is controlled by the choice of phosphine ligand enantiomer and the substituent on the amide nitrogen atom. The method was extended to a general regiodivergent parallel kinetic resolution of the racemic lactam., (Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Optimization of a high-throughput screening hit led to the discovery of a new series of 5-phenoxy-1,3-dimethyl-1H-pyrazole-4-carboxamides as highly potent agonists of TGR5. This novel chemotype was rapidly developed through iterative combinatorial library synthesis. It was determined that in vitro agonist potency correlated with functional activity data from human peripheral blood monocytes., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Takeda G-protein-coupled receptor 5 (TGR5) represents an exciting biological target for the potential treatment of diabetes and metabolic syndrome. A new class of high-throughput screening (HTS)-derived tetrahydropyrido[4,3-d]pyrimidine amide TGR5 agonists is disclosed. We describe our effort to identify an orally available agonist suitable for assessment of systemic TGR5 agonism. This effort resulted in identification of 16, which had acceptable potency and pharmacokinetic properties to allow for in vivo assessment in dog. A key aspect of this work was the calibration of human and dog in vitro assay systems that could be linked with data from a human ex vivo peripheral blood monocyte assay that expresses receptor at endogenous levels. Potency from the human in vitro assay was also found to correlate with data from an ex vivo human whole blood assay. This calibration exercise provided confidence that 16 could be used to drive plasma exposures sufficient to test the effects of systemic activation of TGR5.

The disposition of 3,3-difluoropyrrolidin-1-yl{(2S,4S)-4-[4-(pyrimidin-2-yl)piperazin-1-yl]pyrrolidin-2-yl}methanone (PF-00734200), a dipeptidyl peptidase IV inhibitor that progressed to phase 3 for the treatment of type 2 diabetes, was examined in rats, dogs, and humans after oral administration of a single dose of [(14)C]PF-00734200. Mean recoveries of administered radioactivity were 97.1, 92.2, and 87.2% in rats, dogs, and humans, respectively. The majority of radioactive dose was detected in the urine of dogs and humans and in the feces of rats. Absorption of PF-00734200 was rapid in all species, with maximal plasma concentrations of radioactivity achieved within 1 h after the dose. Circulating radioactivity was primarily composed of the parent drug (79.9, 80.2, and 94.4% in rat, dog, and human, respectively). The major route of metabolism was due to hydroxylation at the 5' position of the pyrimidine ring (M5) in all species. In vitro experiments with recombinant cytochrome P450 isoforms suggested that the formation of M5 was catalyzed both by CYP2D6 and CYP3A4. Molecular docking simulations showed that the 5' position of the pyrimidine moiety of PF-00734200 can access the heme iron-oxo of both CYP3A4 and CYP2D6 in an energetically favored orientation. Other metabolic pathways included amide hydrolysis (M2), N-dealkylation at the piperazine nitrogen (M3) and an unusual metabolite resulting from scission of the pyrimidine ring (M1). Phase II metabolic pathways included the following: carbamoyl glucuronidation (M9), glucosidation (M15) on the pyrrolidine nitrogen, and conjugation with creatinine to form an unusual metabolite/metabonate (M16). The data from these studies suggest that PF-00734200 is eliminated by both metabolism and renal clearance.

A new and useful procedure for the macrocyclization of linear peptides is described. The natural amino acid side chains of tyrosine (phenol), lysine (alkylamine), and histidine (imidazole) react in an intramolecular fashion with a pendent pyridine-N-oxide-carboxamide, which is selectively activated by the phosphonium salt, PyBroP. The reaction is mild, rapid, and efficient with a potentially large substrate scope. Multiple examples are provided with full characterization and analyses, including a novel aza-variant of the C-O-D ring system of vancomycin.

Background: Accumulating evidence supports the role of the mineralocorticoid receptor (MR) in the pathogenesis of diabetic nephropathy. These findings have generated renewed interest in novel MR antagonists with improved selectivity against other nuclear hormone receptors and a potentially reduced risk of hyperkalemia. Characterization of novel MR antagonists warrants establishing translatable biomarkers of activity at the MR receptor. We assessed the translatability of urinary sodium to potassium ratio (Na+/K+) and plasma aldosterone as biomarkers of MR antagonism using eplerenone (Inspra®), a commercially available MR antagonist. Further we utilized these biomarkers to demonstrate antagonism of MR by PF-03882845, a novel compound., Methods: The effect of eplerenone and PF-03882845 on urinary Na+/K+ and plasma aldosterone were characterized in Sprague-Dawley rats and spontaneously hypertensive rats (SHR). Additionally, the effect of eplerenone on these biomarkers was determined in healthy volunteers. Drug exposure-response data were modeled to evaluate the translatability of these biomarkers from rats to humans., Results: In Sprague-Dawley rats, eplerenone elicited a rapid effect on urinary Na+/K+ yielding an EC50 that was within 5-fold of the functional in vitro IC50. More importantly, the effect of eplerenone on urinary Na+/K+ in healthy volunteers yielded an EC50 that was within 2-fold of the EC50 generated in Sprague-Dawley rats. Similarly, the potency of PF-03882845 in elevating urinary Na+/K+ in Sprague-Dawley rats was within 3-fold of its in vitro functional potency. The effect of MR antagonism on urinary Na+/K+ was not sustained chronically; thus we studied the effect of the compounds on plasma aldosterone following chronic dosing in SHR. Modeling of drug exposure-response data for both eplerenone and PF-03882845 yielded EC50 values that were within 2-fold of that estimated from modeling of drug exposure with changes in urinary sodium and potassium excretion. Importantly, similar unbound concentrations of eplerenone in humans and SHR rats yielded the same magnitude of elevations in aldosterone, indicating a good translatability from rat to human., Conclusions: Urinary Na+/K+ and plasma aldosterone appear to be translatable biomarkers of MR antagonism following administration of single or multiple doses of compound, respectively., Trial Registration: For clinical study reference EE3-96-02-004, this study was completed in 1996 and falls out scope for disclosure requirements. Clinical study reference A6141115: http://clinicaltrials.gov, http://NIHclinicaltrails.gov; NCTID: NCT00990223.

Acyl-CoA:diacylglycerol acyltransferase-1 (DGAT-1) catalyzes the final committed step in the biosynthesis of triglycerides. DGAT-1 knockout mice have been shown to be resistant to diet-induced obesity and have increased insulin sensitivity. Thus, inhibition of DGAT-1 may represent an attractive target for the treatment of obesity or type II diabetes. Herein, we report the discovery and characterization of a potent and selective DGAT-1 inhibitor PF-04620110 (3). Compound 3 inhibits DGAT-1 with an IC50 of 19 nM and shows high selectivity versus a broad panel of off-target pharmacologic end points. In vivo DGAT-1 inhibition has been demonstrated through reduction of plasma triglyceride levels in rodents at doses of ≥0.1 mg/kg following a lipid challenge. On the basis of this pharmacologic and pharmacokinetic profile, compound 3 has been advanced to human clinical studies.

A potent, small molecule inhibitor with a favorable pharmacokinetic profile to allow for sustained SCD inhibition in vivo was identified. Starting from a low MW acyl guanidine (5a), identified with a RapidFire High-Throughput Mass Spectrometry (RF-MS) assay, iterative library design was used to rapidly probe the amide and tail regions of the molecule. Singleton synthesis was used to probe core changes. Biological evaluation of a SCD inhibitor (5b) included in vitro potency at SCD-1 and in vivo modulation of the plasma desaturation index (DI) in rats on a low essential fatty acid (LEFA) diet. In addition to dose-dependent decrease in DI, effects on rodent ocular tissue were noted. Therefore, in rat, these SCD inhibitors only recapitulate a portion of phenotype exhibited by the SCD-1 knockout mouse., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

A 3-amino-4-substituted pyrrolidine series of dipeptidyl peptidase IV (DPP-4) inhibitors was rapidly developed into a candidate series by identification of a polar valerolactam replacement for the lipophilic 2,4,5-trifluorophenyl pharmacophore. The addition of a gem-difluoro substituent to the lactam improved overall DPP-4 inhibition and an efficient asymmetric route to 3,4-diaminopyrrolidines was developed. Advanced profiling of a subset of analogs identified 5o with an acceptable human DPP-4 inhibition profile based on a rat PK/PD model and a projected human dose that was suitable for clinical development., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

The design and profile of a series of zwitterionic calcium sensing receptor negative allosteric modulators is described. Evaluation of key analogues using a rat model demonstrate a robust response, significantly improved potency over ronacaleret and have the potential as an oral, anabolic treatment for osteoporosis.

The development of a series of novel 1,2,3,4-tetrahydroisoquinolin-1-ones as antagonists of G protein-coupled receptor 40 (GPR40) is described. The synthesis, in vitro inhibitory values for GPR40, in vitro microsomal clearance and rat in vivo clearance data are discussed. Initial hits displayed high rat in vivo clearances that were higher than liver blood flow. Optimization of rat in vivo clearance was achieved and led to the identification of 15i, whose rat oral pharmacokinetic data is reported.

A series of 4-substituted proline amides was synthesized and evaluated as inhibitors of dipeptidyl pepdidase IV for the treatment of type 2 diabetes. (3,3-Difluoro-pyrrolidin-1-yl)-[(2S,4S)-(4-(4-pyrimidin-2-yl-piperazin-1-yl)-pyrrolidin-2-yl]-methanone (5) emerged as a potent (IC(50) = 13 nM) and selective compound, with high oral bioavailability in preclinical species and low plasma protein binding. Compound 5, PF-00734200, was selected for development as a potential new treatment for type 2 diabetes.

A novel alpha7 nAChR agonist, N-[(3R,5R)-1-azabicyclo[3.2.1]oct-3-yl]furo[2,3-c]pyridine-5-carboxamide (3a, PHA-709829), has been identified for the potential treatment of cognitive deficits in schizophrenia. The compound shows potent and selective alpha7 in vitro activity, excellent brain penetration, good rat oral bioavailability and robust in vivo efficacy in a rat auditory sensory gating model.

A novel set of azabicyclic aryl amides have been identified as potent and selective agonists of the alpha7 nAChR. A two-pronged approach was taken to improve the potential hERG liability of previously disclosed alpha7 nAChR agonist, PNU-282,987, while maintaining the compound's other desirable pharmacological properties. The first approach involved further exploration of the aryl carboxylic acid fragment of PNU-282,987, while the second approach focused on modification of the azabicyclic amine portion of PNU-282,987. The best compounds from each series are characterized by rapid brain penetration, good oral bioavailability in rat, and demonstrate in vivo efficacy in a rat P50 auditory sensory gating assay. At least one analog from each series (1h, 1o, 2a, 9a, and 18a) shows an improved hERG safety profile over PNU-282,987.

Several lines of evidence suggest a link between the alpha7 neuronal nicotinic acetylcholine receptor (nAChR) and brain disorders including schizophrenia, Alzheimer's disease, and traumatic brain injury. The present work describes a novel molecule, 1-(5-chloro-2,4-dimethoxy-phenyl)-3-(5-methyl-isoxazol-3-yl)-urea (PNU-120596), which acts as a powerful positive allosteric modulator of the alpha7 nAChR. Discovered in a high-throughput screen, PNU-120596 increased agonist-evoked calcium flux mediated by an engineered variant of the human alpha7 nAChR. Electrophysiology studies confirmed that PNU-120596 increased peak agonist-evoked currents mediated by wild-type receptors and also demonstrated a pronounced prolongation of the evoked response in the continued presence of agonist. In contrast, PNU-120596 produced no detectable change in currents mediated by alpha4beta2, alpha3beta4, and alpha9alpha10 nAChRs. PNU-120596 increased the channel mean open time of alpha7 nAChRs but had no effect on ion selectivity and relatively little, if any, effect on unitary conductance. When applied to acute hippocampal slices, PNU-120596 increased the frequency of ACh-evoked GABAergic postsynaptic currents measured in pyramidal neurons; this effect was suppressed by TTX, suggesting that PNU-120596 modulated the function of alpha7 nAChRs located on the somatodendritic membrane of hippocampal interneurons. Accordingly, PNU-120596 greatly enhanced the ACh-evoked inward currents in these interneurons. Systemic administration of PNU-120596 to rats improved the auditory gating deficit caused by amphetamine, a model proposed to reflect a circuit level disturbance associated with schizophrenia. Together, these results suggest that PNU-120596 represents a new class of molecule that enhances alpha7 nAChR function and thus has the potential to treat psychiatric and neurological disorders.

The evolution of the insecticidal pyrazoline moiety that was originally discovered in 1972 has led to the discovery of a new crop insecticide, indoxacarb, which is the first commercialized pyrazoline-type sodium-channel blocker. Both monocyclic and fused-tricyclic pyrazolines and pyridazines, as well as structurally related semicarbazones were examined prior to the discovery of analogous tricyclic oxadiazines which had similarly high activity as well as favorable environmental dissipation rates and low toxicity to non-target organisms. The eventual leading candidate, DPX-JW062, was originally obtained as a racemic molecule, but a chiral synthesis was developed which produces material that is 50% ee in the insecticidal (+)-S-enantiomer (DPX-MP062, indoxacarb).

Intermolecular alkylation of the aziridinyl oxazole 20 using PhSO(2)CH(2)CH(2)OTf is possible despite the presence of potentially nucleophilic aziridine nitrogen. The resulting oxazolium salt 22 reacts with BnNMe(3)(+)CN(-) to produce the azomethine ylide 24b via electrocyclic ring opening of an oxazoline 23b. Internal cycloaddition affords 26 in 66% yield. After saponification and base-induced cleavage of the N-phenylsulfonylethyl group, conventional cyclization provides access to 33. Deprotection and DDQ oxidation completes the synthesis of the aziridinomitosene derivative 9b. The starting cis-disubstituted aziridine ester 16 can be prepared by the aza-Darzens reaction of 15 with tert-butyl chloroacetate.