The discovery of potent glycine transporter type-2 inhibitors: Design and synthesis of phenoxymethylbenzamide derivatives

We describe the discovery of phenoxymethylbenzamide derivatives as a novel class of glycine transporter type-2 (GlyT-2) inhibitors. We found hit compound 1 (human GlyT-2, IC₅₀ = 4040 nM) in our library and converted its 1-(1-(naphthalen-2-ylmethyl)piperidin-4-yl)pyrrolidin-3-yl group to an 1-(N,N-dimethylaminopropyl)piperidyl group and its tert-butyl group to a trifluoromethyl group to obtain N-(1-(3-(dimethylamino)propyl)piperidin-4-yl)-4-((4-(trifluoromethyl)phenoxy)methyl)benzamide (20). Compound 20 showed good inhibitory activity against human GlyT-2 (IC₅₀ = 15.3 nM) and exhibited anti-allodynia effects in a mouse neuropathic pain model.     

Design and synthesis of novel anti-Alzheimer’s agents: Acridine-chromenone and quinoline-chromenone hybrids

A novel series of acridine-chromenone and quinoline-chromenone hybrids were designed, synthesized, and evaluated as anti-Alzheimer’s agents. All synthesized compounds were evaluated as cholinesterases (ChEs) inhibitors and among them, 7-(4-(6-chloro-2,3-dihydro-1H-cyclopenta[b]quinolin-9-ylamino)phenoxy)-4-methyl-2H-chromen-2-one (8e) exhibited the most potent anti-acetylcholinesterase (AChE) inhibitory activity (IC₅₀ = 16.17 μM) comparing with rivastigmine (IC₅₀ = 11.07 μM) as the reference drug. Also, compound 8e was assessed for its β-secretase (BACE1) inhibitory and neuroprotective activities which demonstrated satisfactory results. It should be noted that both kinetic study on the inhibition of AChE and molecular modeling revealed that compound 8e interacted simultaneously with both the catalytic active site (CAS) and peripheral anionic site (PAS) of AChE.     

Synthesis and biological evaluation of salicylic acid and N-acetyl-2-carboxybenzenesulfonamide regioisomers possessing a N-difluoromethyl-1,2-dihydropyrid-2-one pharmacophore: Dual inhibitors of cyclooxygenases and 5-lipoxygenase with anti-inflammatory activity

A novel class of salicylic acid and N-acetyl-2-carboxybenzenesulfonamide regioisomers possessing a N-difluoromethyl-1,2-dihydropyrid-2-one pharmacophore attached to its C-4 or C-5 position was designed for evaluation as anti-inflammatory (AI) agents. Replacement of the 2,4-difluorophenyl ring in diflunisal by the N-difluoromethyl-1,2-dihydropyrid-2-one moiety provided compounds showing dual selective cyclooxygenase-2 (COX-2)/5-lipoxygenase (5-LOX) inhibitory activities. AI structure–activity studies showed that the C-4 (14a) and C-5 (14b) salicylate regioisomers were 1.4- and 1.6-fold more potent than aspirin, and the C-5 N-acetyl-2-carboxybenzenesulfonamide regioisomer (22b) was 1.3- and 2.8-fold more potent than ibuprofen and aspirin, respectively. In vivo ulcer index (UI) studies showed that the 4- and 5-(N-difluoromethyl-1,2-dihydropyrid-2-one-4-yl)salicylic acids (14a and 14b) were completely non-ulcerogenic since no gastric lesions were present (UI = 0) relative to aspirin (UI = 57) at an equivalent μmol/kg oral dose. The N-difluoromethyl-1,2-dihydropyridin-2-one moiety provides a novel 5-LOX pharmacophore for the design of cyclic hydroxamic mimetics for exploitation in the development of dual COX-2/5-LOX inhibitory AI drugs.     

Synthesis and antiprotozoal activity of 2,5-bis[amidinoaryl]thiazoles

Seven novel diamidino 2,5-bis(aryl)thiazoles (5a–g) were synthesized and evaluated against Trypanosoma brucei rhodensiense (T. b. r.) and Plasmodium falciparum (P. f.). The diamidines were obtained directly from the corresponding bis-nitriles (4a–g) by the action of lithium bis(trimethylsilyl)amide. The bis-nitriles 4a–f were synthesized in four steps starting with the Stille coupling of 2-tributyltinthiazole with the appropriate cyanoaryl halide. The bis-nitrile 5g was obtained by the palladium facilitated coupling of the mixed tin-silyl reagent 2-trimethylsilyl-5-trimethyltinthiazole with 2-bromo-5-cyanopyridine. The amidoxime potential prodrugs 6a–e, 6g were obtained by the reaction of hydroxylamine with the bis-nitriles. O-Methylation of the amidoximes gave the corresponding N-methoxyamidines 7a–c, 7e, 7g. The diamidines showed strong DNA binding affinity as reflected by ΔT_m measurements. Four of the diamidines 5a, 5b, 5d and 5e were highly active in vitro against P. f. giving IC₅₀ values between 1.1 and 2.5 nM. The same four diamidines showed IC₅₀ values between 4 and 6 nM against T. b. r. The selectivity indices ranged from 233 to 9175. One diamidine 5a produced one of four cures at an ip dose of 4 × 5 mg/kg in the STIB900 mouse model for acute African trypanosomiasis. The amidoxime and N-methoxyamidine of 5a were the only produgs to provide cures (1/4 cures) in the same mouse model on oral dosage at 4 × 25 mg/kg.     

N-Alkylated arylsulfonamides of (aryloxy)ethyl piperidines: 5-HT7 receptor selectivity versus multireceptor profile

The N-alkylation of the sulfonamide moiety, in a group of arylsulfonamide derivatives of (aryloxy)ethyl piperidines, may be considered as a strategy for the design of selective 5-HT₇ receptor ligands or multifunctional agents to extend a polypharmacological approach to the treatment of complex diseases. The study allowed for the identification of 31 (1-methyl-N-{1-[2-(2-(t-butyl)phenoxy)ethyl]piperidin-4-yl}-N-cyclopropylmethyl-1H-pyrazole-4-sulfonamide), a potent and selective 5-HT₇ receptor antagonist and 33 (1-methyl-N-{1-[2-(biphenyl-2-yloxy)ethyl]piperidin-4-yl}-N-cyclopropylmethyl-1H-pyrazole-4-sulfonamide), as multimodal 5-HT/dopamine receptor ligand, as 5-HT_2A/5-HT₇/D₂ receptor antagonists. Both selected compounds were evaluated in vivo in a forced swim test (FST) in mice and in a novel object recognition (NOR) task in rats, demonstrating distinct antidepressant-like and pro-cognitive properties (MED = 1.25 mg/kg and 1 mg/kg, ip, respectively). These findings warrant further studies to explore the therapeutic potential of N-alkylated arylsulfonamides for the treatment of CNS disorders.     

Discovery of substituted N′-(2-oxoindolin-3-ylidene)benzohydrazides as new apoptosis inducers using a cell- and caspase-based HTS assay

We report the discovery of a series of substituted N′-(2-oxoindolin-3-ylidene)benzohydrazides as inducers of apoptosis using our proprietary cell- and caspase-based ASAP HTS assay. Through SAR studies, N′-(4-bromo-5-methyl-2-oxoindolin-3-ylidene)-3,4,5-trimethoxybenzohydrazide (3g) was identified as a potent apoptosis inducer with an EC₅₀ value of 0.24 μM in human colorectal carcinoma HCT116 cells, more than a 40-fold increase in potency from the initial screening hit N′-(5-bromo-2-oxoindolin-3-ylidene)-3,4,5-trimethoxybenzohydrazide (2a). Compound 3g also was found to be highly active in a growth inhibition assay with a GI₅₀ value of 0.056 μM in HCT116 cells. A group of potentially more aqueous soluble analogs were prepared and found to be highly active. Among them, compound 4e incorporating a methyl piperazine moiety was found to have EC₅₀ values of 0.17, 0.088 and 0.14 μM in human colorectal carcinoma cells HCT116, hepatocellular carcinoma cancer SNU398 cells and human colon cancer RKO cells, respectively. Compounds 3g and 4e were found to function as inhibitors of tubulin polymerization.     

Synthesis,structure–activity relationships,and biological profiles of a dihydrobenzoxathiin class of histamine H3 receptor inverse agonists

A series of novel dihydrobenzoxathiin derivatives was synthesized and evaluated as potent human histamine H₃ receptor inverse agonists. After systematic modification of lead 1a, the potent and selective histamine H₃ inverse agonist 1-(3-{4-[(2S,3S)-8-methoxy-3-methyl-4,4-dioxido-2,3-dihydro-1,4-benzoxathiin-2-yl]phenoxy}propyl)pyrrolidine (5k) was identified. Compound 5k showed good pharmacokinetic profiles and brain penetrability in laboratory animals. After 3 mg/kg oral administration of 5k, significant elevation of brain histamine levels was observed in rats where the brain H₃ receptor was fully occupied.     

Synthesis and biological study of 3-(phenylsulfonyl)thieno[2,3-e][1,2,3]triazolo[1,5-a]pyrimidines as potent and selective serotonin 5-HT6 receptor antagonists

A number of 3-(phenylsulfonyl)thieno[2,3-e][1,2,3]triazolo[1,5-a]pyrimidines were prepared and their 5-HT₆ receptor binding affinity and ability to inhibit the functional cellular responses to serotonin were evaluated. 3-[(3-Chlorophenyl)sulfonyl]-N-(tetrahydrofuran-2-ylmethyl)thieno[2,3-e][1,2,3]triazolo[1,5-a]pyrimidin-5-amine 2{5,26} appeared to be the most active in a functional assay (IC₅₀ = 29.0 nM) and 3-(phenylsulfonyl)-N-(2-thienylmethyl) thieno[2,3-e][1,2,3]triazolo[1,5-a]pyrimidin-5-amine 2{1,28} demonstrated the greatest affinity in a 5-HT₆ receptor radioligand binding assay (K_i = 1.7 nM). A screening of 5-HT_2A and 5-HT_2B receptor affinity revealed that 3-(phenylsulfonyl)thieno[2,3-e][1,2,3]triazolo[1,5-a]pyrimidines are highly selective 5-HT₆ receptor ligands.     

Synthesis and biological evaluation of 2-substituted-4-(3′,4′,5′-trimethoxyphenyl)-5-aryl thiazoles as anticancer agents

Antitumor agents that bind to tubulin and disrupt microtubule dynamics have attracted considerable attention in the last few years. To extend our knowledge of the thiazole ring as a suitable mimic for the cis-olefin present in combretastatin A-4, we fixed the 3,4,5-trimethoxyphenyl at the C4-position of the thiazole core. We found that the substituents at the C2- and C5-positions had a profound effect on antiproliferative activity. Comparing compounds with the same substituents at the C5-position of the thiazole ring, the moiety at the C2-position influenced antiproliferative activities, with the order of potency being NHCH₃ > Me ? N(CH₃)₂. The N-methylamino substituent significantly improved antiproliferative activity on MCF-7 cells with respect to C2-amino counterparts. Increasing steric bulk at the C2-position from N-methylamino to N,N-dimethylamino caused a 1–2 log decrease in activity. The 2-N-methylamino thiazole derivatives 3b, 3d and 3e were the most active compounds as antiproliferative agents, with IC₅₀ values from low micromolar to single digit nanomolar, and, in addition, they are also active on multidrug-resistant cell lines over-expressing P-glycoprotein. Antiproliferative activity was probably caused by the compounds binding to the colchicines site of tubulin polymerization and disrupting microtubule dynamics. Moreover, the most active compound 3e induced apoptosis through the activation of caspase-2, -3 and -8, but 3e did not cause mitochondrial depolarization.     

Synthesis and structure–activity relationship of 3β-(4-alkylthio, -methylsulfinyl,and -methylsulfonylphenyl)tropane and 3β-(4-alkylthiophenyl)nortropane derivatives for monoamine transporters

Early studies led to the identification of 3β-(4-methoxyphenyl)tropane-2β-carboxylic acid methyl ester (5) with high affinity at the DAT (IC₅₀ = 6.5 nM) and 5-HTT (K_i = 4.3 nM), while having much less affinity at the NET (K_i = 1110 nM). In the present study, we replaced the 4′-methoxy group of the 3β-phenyl ring with a bioisosteric 4′-methylthio group to give 7a. We also synthesized a number of 3β-(4-alkylthiophenyl)tropanes 7b–e, 3β-(4-methylsulfinylphenyl) and 3β-(4-methylsulfonylphenyl)tropane analogues 7f–h as well as the 3β-(4-alkylthiophenyl)nortropane derivatives 8–11 to further characterize the structure–activity relationship of this type of compound for binding at monoamine transporters. With exception of the 4′-methylsulfonyl analogue 7h, all the tested compounds possessed high binding affinities at the 5-HTT. The K_i values ranged from 0.19 nM to 49 nM. The 3β-(4-methylthiophenyl)tropane 7a and its N-(3-fluoropropyl) analogue 9a and N-allyl analogue 10a are the most selective compounds for the 5-HTT over the NET (NET/5-HTT = 314–364) in the series. However, none of the compounds showed selectivity similar to 5 for both the DAT and 5-HTT relative to the NET. This study provided useful SAR information for rational design of potent and selective monoamine transporter inhibitors.     

(R)-3-Amino-1-((3aS,7aS)-octahydro-1H-indol-1-yl)-4-(2,4,5-trifluorophenyl)butan-1-one derivatives as potent inhibitors of dipeptidyl peptidase-4: Design,synthesis, biological evaluation,and molecular modeling

A series of (R)-3-amino-1-((3aS,7aS)-octahydro-1H-indol-1-yl)-4-(2,4,5-trifluorophenyl)butan-1-one derivatives was designed, synthesized, and evaluated as novel inhibitors of dipeptidyl peptidase-4 (DPP-4) for the treatment of type 2 diabetes. Most of the synthesized compounds demonstrated good inhibition activities against DPP-4. Among these, compounds 3e, 4c, 4l, and 4n exhibited prominent inhibition activities against DPP-4, with IC₅₀s of 0.07, 0.07, 0.14, and 0.17 μM, respectively. The possible binding modes of compounds 3e and 4n with dipeptidyl peptidase-4 were also explored by molecular docking simulation. These potent DPP-4 inhibitors were optimized for the absorption, distribution, metabolism, and excretion (ADME) properties, and compound 4n displayed an attractive pharmacokinetic profile (F = 96.3%, t_1/2 = 10.5 h).     

Microbial alcohol dehydrogenase screening for enantiopure lactone synthesis: Down-stream process from microtiter plate to bench bioreactor

One-pot conversion with whole cells of bacteria was performed for biooxidation of meso monocyclic (3a–b) and bicyclic diols (3c–e) into corresponding chiral lactones of bicyclo[4.3.0]nonane structure (2a–b) as well as exo- and endo-bridged lactones with the structure of [2.2.1] (3c–d) and [2.2.2] (3e). Micrococcus sp. DSM 30771 was selected as biocatalyst with significant alcohol dehydrogenase activity. Among tested strains, microbial oxidation of meso diols 3a–e catalyzed by Micrococcus sp. afforded enantiomerically pure ((+)-(2S,3R)-2c (ee = 99%), (+)-(2S,3R)-2e (ee = 99%)) or enriched ((+)-(1S,5R)-2a (ee = 90%), (−)-(1S,5R)-2b (ee = 86%), (+)-(2S,3R)-2d (ee = 80%)) lactone moieties. Comparative study with respect to microbial cultivation as well as biooxidation was undertaken to verify agreement of secondary metabolite biosynthesis in different scales: from MTP (4 mL), across shake flask (100 mL) till bioreactor (4 L). The results from biotransformations showed quite similar dependence in oxidation of all substrates 3a–e in MTP and flasks as well, thereby confirmed the validity and reasonable approach of using MTP for preliminary studies.     

Synthesis of 5- and 6-substituted 2-(4-dimethylaminophenyl)-1,3-benzoxazoles and their in vitro and in vivo evaluation as imaging agents for amyloid plaque

A series of novel 5- and 6-substituted 2-(4-dimethylaminophenyl)-1,3-benzoxazoles was synthesized and their potential as imaging probes for Alzheimer’s Disease (AD)-related amyloid plaque was evaluated in vitro and in vivo. In vitro binding affinities for Aβ1–40 peptide of several of these compounds were in the low-nanomolar range . The lowest K_i of 9.3 nM was found for N-(2-(4-(dimethylamino)phenyl)-1,3-benzoxazol-5-yl)-4-iodobenzamide (1e). Its ¹²³I-radiolabeled form ([¹²³I]1e) was subsequently prepared by iododestannylation of the corresponding tributylstannyl precursor and evaluated in vivo in a baboon model using SPECT imaging. Contrary to our expectations, 1e did not cross the blood–brain barrier (BBB) to any significant extent.     

1,5-Benzodiazepine inhibitors of HCV NS5B polymerase

Optimization through parallel synthesis of a novel series of hepatitis C virus (HCV) NS5B polymerase inhibitors led to the identification of (R)-11-(4-benzyloxy-2-fluorophenyl)-6-hydroxy-3,3-dimethyl-10-(6-methylpyridine-2-carbonyl)-2,3,4,5,10,11-hexahydro-dibenzo[b,e][1,4]diazepin-1-one 11zc and (R)-11-(4-benzyloxy-2-fluorophenyl)-6-hydroxy-3,3-dimethyl-10-(2,5-dimethyloxazol-4-carbonyl)-2,3,4,5,10,11-hexahydro-dibenzo[b,e][1,4]diazepin-1-one 11zk as potent (replicon EC₅₀ = 400 nM and 270 nM, respectively) and selective (CC₅₀ > 20 μM) inhibitors of HCV replication. These data warrant further lead-optimization efforts.     

Synthesis and biological evaluation of (phenylpiperazinyl-propyl)arylsulfonamides as selective 5-HT2A receptor antagonists

A novel series of 5-HT_2A ligands that contain a (phenylpiperazinyl-propyl)arylsulfonamides skeleton was synthesized. Thirty-seven N-(cycloalkylmethyl)-4-methoxy-N-(3-(4-arylpiperazin-1-yl)propyl)-arylsulfonamide and N-(4-(4-arylpiperazin-1-yl)butan-2-yl)-arylsulfonamide compounds were obtained. The binding of these compounds to the 5-HT_2A, 5-HT_2C, and 5-HT₇ receptors was evaluated. Most of the compounds showed IC₅₀ values of less than 100 nM and exhibited high selectivity for the 5-HT_2A receptor. Among the synthesized compounds, 16a and 16d showed good affinity at 5-HT_2A (IC₅₀ = 0.7 nM and 0.5 nM) and good selectivity over 5-HT_2C (50–100 times) and 5-HT₇ (1500–3000 times).     

Novel boronated chlorin e6-based photosensitizers: Synthesis,binding to albumin and antitumour efficacy

Chlorins, a class of plant porphyrins, are perspective as photosensitizing agents due to light absorption in the long wavelength spectral region and deeper photodamage of tissues. Aiming at optimization of antitumour properties of chlorins, we synthesized a series of boronated derivatives of chlorin e₆ and their complexes containing Zn(II), Pd(II) or Sn(IV). The compounds were synthesized by alkylation of amino or hydroxy derivatives of chlorin e₆ with 1-trifluoromethanesulfonylmethyl-o-carborane. Chlorin e₆ 13(1)-N-{2-[N-(o-carboran-1-yl)methyl]aminoethyl}amide-15(2), 17(3)-dimethyl ester (compound 5) formed complexes with serum albumin, a major porphyrin carrier. The binding constant of these complexes was ～4 times bigger than the respective value for the complexes of albumin with boron-free aminochlorin e₆. Compound 5 potently sensitized rat fibroblasts to illumination with monochromatic red light: >98% of cells were necrotic by 24 h post-illumination with 1 μM of 5. This compound demonstrated high efficacy in photodynamic therapy of rat M-1 sarcoma. After PDT with 25 mg/kg of 5 the residual tumours were significantly smaller than in animals subjected to PDT with equal concentration of boron-free aminochlorin e₆. No signs of general toxicity were detectable after PDT with 5. Thus, boronation can enhance the potency of chlorins in PDT, in particular, due to an increased binding to albumin. Our data expand the therapeutic applicability of boronated chlorins beyond boron neutron capture therapy; these agents emerge as dual efficacy photoradiosensitizers.     

Synthesis,characterization and in vitro biological evaluation of some novel 1,3,5-triazine–Schiff base conjugates as potential antimycobacterial agents

A series of some novel 1,3,5-triazine–Schiff base conjugates (1–32) have been synthesized and evaluated for their in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Rv using Alamar Blue assay and the activity expressed as the minimum inhibitory concentration (MIC) in μg/mL. Compounds 4 (4-Methoxy-6-methyl-N-(3,4,5-trimethoxybenzylidene)-1,3,5-triazin-2-amine), 11 (4-Methoxy-6-methyl-N-(2-hydroxy-3-bromo-5-chloro-benzylidene)-1,3,5-triazin-2-amine) and 24 (4-Methoxy-6-methyl-N-(1-(2,5-dihydroxyphenyl)ethylidene)-1,3,5-triazin-2-amine) exhibited a significant activity at 3.125, 6.25 and 6.25 μg/mL, respectively, when compared with the antitubercular drugs such as ethambutol (3.125 μg/mL), pyrazinamide (6.25 μg/mL) and streptomycin (6.25 μg/mL) and it could be a potential starting point to develop new lead compounds in the fight against Mycobacterium tuberculosis H37Rv.     

Synthesis and biological evaluation of novel propargyl amines as potential fluorine-18 labeled radioligands for detection of MAO-B activity

The aim of this project was to synthesize and evaluate three novel fluorine-18 labeled derivatives of propargyl amine as potential PET radioligands to visualize monoamine oxidase B (MAO-B) activity.The three fluorinated derivatives of propargyl amine ((S)-1-fluoro-N,4-dimethyl-N-(prop-2-ynyl)-pent-4-en-2-amine (5), (S)-N-(1-fluoro-3-(furan-2-yl)propan-2-yl)-N-methylprop-2-yn-1-amine (10) and (S)-1-fluoro-N,4-dimethyl-N-(prop-2-ynyl)pentan-2-amine (15)) were synthesized in multi-step organic syntheses. IC₅₀ values for inhibition were determined for compounds 5, 10 and 15 in order to determine their specificity for binding to MAO-B. Compound 5 inhibited MAO-B with an IC₅₀ of 664 ± 48.08 nM. No further investigation was carried out with this compound. Compound 10 inhibited MAO-B with an IC₅₀ of 208.5 ± 13.44 nM and compound 15 featured an IC₅₀ of 131.5 ± 0.71 nM for its MAO-B inhibitory activity. None of the compounds inhibited MAO-A activity (IC₅₀ > 2 μM).The fluorine-18 labeled analogues of the two higher binding affinity compounds (10 and 15) (S)-N-(1-[¹⁸F]fluoro-3-(furan-2-yl)propan-2-yl)-N-methylprop-2-yn-1-amine (16) and (S)-1-[¹⁸F]fluoro-N,4-dimethyl-N-(prop-2-ynyl)pentan-2-amine (18) were both prepared from the corresponding precursors 9A, 9B and 14A, 14B by a one-step fluorine-18 nucleophilic substitution reaction. Autoradiography experiments on human postmortem brain tissue sections were performed with 16 and 18. Only compound 18 demonstrated a high selectivity for MAO-B over MAO-A and was, therefore, chosen for further examination by PET in a cynomolgus monkey.The initial uptake of 18 in the monkey brain was 250% SUV at 4 min post injection. The highest uptake of radioactivity was observed in the striatum and thalamus, regions with high MAO-B activity, whereas lower levels of radioactivity were detected in the cortex and cerebellum. The percentage of unchanged radioligand 18 was 30% in plasma at 90 min post injection.In conclusion, compound 18 is a selective inhibitor of MAO-B in vitro and demonstrated a MAO-B specific binding pattern in vivo by PET in monkey. It can, therefore, be considered as a candidate for further investigation in human by PET.     

Inhibitors of hepatitis C virus polymerase: Synthesis and characterization of novel 2-oxy-6-fluoro-N-((S)-1-hydroxy-3-phenylpropan-2-yl)-benzamides

SAR exploration from an initial hit, (S)-N-(2-cyclohexenylethyl)-2-fluoro-6-(2-(1-hydroxy-3-phenylpropan-2-ylamino)-2-oxoethoxy)benzamide (1), identified using our proprietary automated ligand identification system (ALIS),¹ has led to a novel series of selective hepatitis C virus (HCV) NS5B polymerase inhibitors with improved in vitro potency as exemplified by (S)-2-fluoro-6-(2-(1-hydroxy-3-phenylpropan-2-ylamino)-2-oxoethoxy)-N-isopentyl-N-methylbenzamidecarboxamide (41) (IC₅₀ = 0.5 μM). The crystal structure of an analogue (44) was solved and provided rationalization of the SAR of this series, which binds in a distinct manner in the palm domain of NS5B, consistent with biochemical analysis using enzyme mutant variants. These data warrant further lead optimization efforts on this novel series of non-nucleoside inhibitors targeting the HCV polymerase.     

Synthesis and pharmacological evaluation of novel N-aryl-3,4-dihydro-1′H-spiro[chromene-2,4′-piperidine]-1′-carboxamides as TRPM8 antagonists

A novel series of N-aryl-3,4-dihydro-1′H-spiro[chromene-2,4′-piperidine]-1′-carboxamides was identified as transient receptor potential melastatin 8 (TRPM8) channel blockers through analogue-based rational design, synthesis and screening. Details of the synthesis, effect of aryl groups and their substituents on in-vitro potency were studied. The effects of selected functional groups on the 4-position of the chromene ring were also studied, which showed interesting results. The 4-hydroxy derivatives showed excellent potency and selectivity. Optical resolution and screening of alcohols revealed that (R)-(–)-isomers were in general more potent than the corresponding (S)-(+)-isomers. The isomer (R)-(–)-10e (IC₅₀: 8.9 nM) showed a good pharmacokinetic profile upon oral dosing at 10 mg/kg in Sprague–Dawley (SD) rats. The compound (R)-(–)-10e also showed excellent efficacy in relevant rodent models of neuropathic pain.