Phosphodiesterase 1b (PDE1B) Regulates Spatial and Contextual Memory in Hippocampus.

Augmentation of cyclic nucleotide signaling through inhibition of phosphodiesterase (PDE) activity has long been understood to enhance memory. Efforts in this domain have focused predominantly on PDE4, a cAMP-specific phosphodiesterase implicated in consolidation. But less is known about the function of other PDEs expressed in neuroanatomical regions critical to memory. The PDE1 isoforms are the only PDEs to regulate neuronal cAMP and cGMP levels in a Ca2+/Calmodulin (CaM) dependent manner. Here, we show that knock-down of PDE1B in hippocampus of adult mice enhances contextual and spatial memory without effect on non-cognitive behaviors. Pharmacological augmentation of memory in rats was observed with a selective inhibitor of PDE1 dosed before and immediately after training, but not with drug dosed either 1 h after training or before recall. Our data clearly demonstrate a role for the PDE1B isoforms as negative regulators of memory, and they implicate PDE1 in an early phase of consolidation, but not retrieval. Inhibition of PDE1B is a promising therapeutic mechanism for treating memory impairment.

Solvent-Controlled, Site-Selective N-Alkylation Reactions of Azolo-Fused Ring Heterocycles at N1-, N2-, and N3-Positions, Including Pyrazolo[3,4- d]pyrimidines, Purines, [1,2,3]Triazolo[4,5]pyridines, and Related Deaza-Compounds.

Alkylation of 4-methoxy-1 H-pyrazolo[3,4- d]pyrimidine (1b) with iodomethane in THF using NaHMDS as base selectively provided N2-methyl product 4-methoxy-2-methyl-2 H-pyrazolo[3,4- d]pyrimidine (3b) in an 8/1 ratio over N1-methyl product (2b). Interestingly, conducting the reaction in DMSO reversed selectivity to provide a 4/1 ratio of N1/N2 methylated products. Crystal structures of product 3b with N1 and N7 coordinated to sodium indicated a potential role for the latter reinforcing the N2-selectivity. Limits of selectivity were tested with 26 heterocycles which revealed that N7 was a controlling element directing alkylations to favor N2 for pyrazolo- and N3 for imidazo- and triazolo-fused ring heterocycles when conducted in THF. Use of 1H-detected pulsed field gradient-stimulated echo (PFG-STE) NMR defined the molecular weights of ionic reactive complexes. This data and DFT charge distribution calculations suggest close ion pairs (CIPs) or tight ion pairs (TIPs) control alkylation selectivity in THF and solvent-separated ion pairs (SIPs) are the reactive species in DMSO.

Discovery of Selective Phosphodiesterase 1 Inhibitors with Memory Enhancing Properties.

A series of potent thienotriazolopyrimidinone-based PDE1 inhibitors was discovered. X-ray crystal structures of example compounds from this series in complex with the catalytic domain of PDE1B and PDE10A were determined, allowing optimization of PDE1B potency and PDE selectivity. Reduction of hERG affinity led to greater than a 3000-fold selectivity for PDE1B over hERG. 6-(4-Methoxybenzyl)-9-((tetrahydro-2H-pyran-4-yl)methyl)-8,9,10,11-tetrahydropyrido[4',3':4,5]thieno[3,2-e][1,2,4]triazolo[1,5-c]pyrimidin-5(6H)-one was identified as an orally bioavailable and brain penetrating PDE1B enzyme inhibitor with potent memory-enhancing effects in a rat model of object recognition memory.

Characterization of thien-2-yl 1S,2R-milnacipran analogues as potent norepinephrine/serotonin transporter inhibitors for the treatment of neuropathic pain.

Thien-2-yl 1S,2R-milnacipran analogues were synthesized and characterized as norepinephrine/serotonin transporter inhibitors. These compounds possessed higher potencies than 1S,2R-milnacipran (2R-1) while maintaining low molecular weight and moderate lipophilicity, which are the important features for the pharmacological and pharmacokinetic characteristics of milnacipran (1). Thus, compound 5c exhibited IC50 values of 2.3 and 32 nM, respectively, at NET and SERT, which were more than 10-fold better than those of 1 (NET IC50 = 77 nM, SERT IC50 = 420 nM). Moreover, 5c achieved the same efficacy as 1, but with much lower doses, in a rodent spinal nerve ligation pain model. In addition, 5c displayed desirable pharmacokinetic properties in several species, including high oral availability and significant brain penetration.

Studies on the structure-activity relationship of bicifadine analogs as monoamine transporter inhibitors.

Compounds with various activities and selectivities were discovered through structure-activity relationship studies of bicifadine analogs as monoamine transporter inhibitors. The norepinephrine-selective 2-thienyl compound S-6j was efficacious in a rodent pain model.

Identification of 1S,2R-milnacipran analogs as potent norepinephrine and serotonin transporter inhibitors.

A series of milnacipran analogs were synthesized and studied as monoamine transporter inhibitors, and several potent compounds with moderate lipophilicity were identified from the 1S,2R-isomers. Thus, 15l exhibited IC(50) values of 1.7nM at NET and 25nM at SERT, which were, respectively, 20- and 13-fold more potent than 1S,2R-milnacipran 1-II.

Studies on a series of milnacipran analogs containing a heteroaromatic group as potent norepinephrine and serotonin transporter inhibitors.

A series of milnacipran analogs containing a heteroaromatic group were synthesized and studied as monoamine transporter inhibitors. Many compounds exhibited higher potency than milnacipran at NET and NET/SERT with no significant change in lipophilicity. For example, compound R-26f was about 10-fold more potent than milnacipran with IC(50) values of 8.7 and 26nM at NET and SERT, respectively.

Studies on the SAR and pharmacophore of milnacipran derivatives as monoamine transporter inhibitors.

Derivatives of milnacipran were synthesized and studied as monoamine transporter inhibitors. Potent analogs were discovered at NET (9k) and at both NET and SERT (9s and 9u). A pharmacophore model was established based on the conformational analysis of milnacipran in aqueous solution using NMR techniques and was consistent with the SAR results.

Synthesis and structure-activity relationships of spirohydantoin-derived small-molecule antagonists of the melanin-concentrating hormone receptor-1 (MCH-R1).

The design, synthesis, and SAR of a series of substituted spirohydantoins are described. Optimization of an in-house screening hit gave compounds that exhibited potent binding affinity and functional activity at MCH-R1.

Thienopyrimidinone bis-aminopyrrolidine ureas as potent melanin-concentrating hormone receptor-1 (MCH-R1) antagonists.

A series of thienopyrimidinone bis-aminopyrrolidine ureas were designed, synthesized, and evaluated for their ability to bind melanin-concentrating hormone receptor-1. These compounds exhibit potent binding affinity (K(i)=3 nM) and good in vitro metabolic stability.

Synthesis and structure-activity relationships of retro bis-aminopyrrolidine urea (rAPU) derived small-molecule antagonists of the melanin-concentrating hormone receptor-1 (MCH-R1). Part 1.

The design, synthesis, and SAR of a series of retro bis-aminopyrrolidine ureas are described. Compounds from this series exhibited potent binding affinity and functional activity at MCH-R1, and good oral bioavailability in rat.

A thienopyridazinone-based melanin-concentrating hormone receptor 1 antagonist with potent in vivo anorectic properties.

Melanin-concentrating hormone receptor antagonists containing thieno- and a benzopyridazinone cores were designed and tested as potential anorectic agents. These ligands showed high affinity for the receptor, potent functional activity in vitro, and good oral bioavailabilty in rats. The thiophene analogue exhibited low iv clearance, long half-life, and high brain penetration. In obese rats, the thienopyridazinone demonstrated a dose-dependent reduction in feeding and body weight with doses between 1 and 10 mg kg-1.

Substituted chromones and quinolones as potent melanin-concentrating hormone receptor 1 antagonists.

A series of substituted chromones were designed, synthesized, and evaluated for their ability to bind melanin-concentrating hormone receptor 1. Compounds with subnanomolar binding affinity and 66% oral bioavailability in rats were discovered.

Design and synthesis of tricyclic corticotropin-releasing factor-1 antagonists.

Antagonists of the corticotropin-releasing factor (CRF) neuropeptide should prove to be effective in treating stress and anxiety-related disorders. In an effort to identify antagonists with improved physicochemical properties, new tricyclic CRF(1) antagonists were designed, synthesized, and tested for biological activity. As a result of studies aimed at establishing a relationship between structure and CRF(1) binding affinity, NBI 35965 (12a) was identified as a high-affinity antagonist with a pK(i) value of 8.5. Compound 12a proved to be a functional CRF(1) antagonist with pIC(50) values of 7.1 and 6.9 in the in vitro CRF-stimulated cAMP accumulation and ACTH production assays, respectively, and 12a also reduced CRF or stress induced ACTH production in vivo.

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 and orally active non-peptide antagonists of the human melanocortin-4 receptor based on a series of trans-2-disubstituted cyclohexylpiperazines.

The melanocortin-4 receptor (MC4R) plays an important role in the regulation of energy homeostasis. Recent studies have shown that blockade of the MC4R reverses tumor-induced weight loss in mice. Herein, we describe the synthesis and identification of potent and selective non-peptide antagonists of the human MC4R from a series of 2-ethoxycarbonylcyclohexyl-piperazines. Compound 12i was found to possess low nanomolar affinity for the MC4R, and exhibit oral bioavailability in rats. More importantly, when administered orally to mice (10 mg/kg), it led to statistically significant increases in food intake over a 24-h period.

Synthesis and structure-activity relationships of biarylcarboxamide bis-aminopyrrolidine urea derived small-molecule antagonists of the melanin-concentrating hormone receptor-1 (MCH-R1).

A novel series of bis-aminopyrrolidine ureas containing either a 4-biphenylcarboxmide or 5-phenyl-2-thiophenecarboxamide group have been identified as potent and functional antagonists of the melanin-concentrating hormone receptor-1. Syntheses and SAR are described, which led to the discovery of compounds with high binding affinity (Ki = 1 nM) for the receptor. Preliminary in vitro metabolic stability data are also reported for key compounds.

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.

Bis(aminopyrrolidine)-derived ureas (APUs) as potent MCH1 receptor antagonists.

Ureas derived from two substituted 3-aminopyrrolidine subunits were prepared as constrained analogs of a linear lead compound and tested as antagonists of the MCH(1) receptor. The series was optimized for substitution and stereochemistry to generate a functional antagonist with a K(i) of 3.3 nM and IC(50) of 12 nM (GTPgammaS).