Studies on depigmenting activities of dihydroxyl benzamide derivatives containing adamantane moiety

Six diphenolic compounds containing adamantane moiety were synthesized and evaluated as potent inhibitors on tyrosinase activity and melanin formation in melan-a cells. The inhibitory activity of 4-adamantyl resorcinol 1 was similar to that of 4-n-butyl resorcinol in both assays. However, dihydroxyl benzamide derivatives 6a–e showed different inhibitory patterns. All derivatives significantly suppressed the cellular melanin formation without tyrosinase inhibitory activities. These behaviors indicated that the introduction of amide bond changes the binding mode of dihydroxyl groups to tyrosinase. Among derivatives, 6d (3,4-dihydroxyl compound) and 6e (2,3-dihydroxyl compound) showed stronger inhibitory activities (IC₅₀ = 1.25 μM and 0.73 μM, respectively) as compared to 4-n-butyl resorcinol (IC₅₀ = 21.64 μM) and hydroquinone (IC₅₀ = 3.97 μM). This study showed that the position of dihydroxyl substituent at aromatic ring is important for the intercellular inhibition of melanin formation, and also amide linkage and adamantane moiety enhance the inhibition.     

Discovery of novel α1-adrenoceptor ligands based on the antipsychotic sertindole suitable for labeling as PET ligands

The synthesis and in vitro preclinical profile of a series of 5-heteroaryl substituted analogs of the antipsychotic drug sertindole are presented. Compounds 1-(4-fluorophenyl)-3-(1-methylpiperidin-4-yl)-5-(pyrimidin-5-yl)-1H-indole (Lu AA27122, 3i) and 1-(4-fluorophenyl)-5-(1-methyl-1H-1,2,4-triazol-3-yl)-3-(1-methylpiperidin-4-yl)-1H-indole (3l) were identified as high affinity α_1A-adrenoceptor ligands with K_i values of 0.52 and 0.16 nM, respectively, and with a >100-fold selectivity versus dopamine D₂ receptors. Compound 3i showed almost equal affinity for α_1B- (K_i = 1.9 nM) and α_1D-adrenoceptors (K_i = 2.5 nM) as for α_1A, as well as moderate affinity for 5-HT_1B (K_i = 13 nM) and 5-HT₆ (K_i = 16 nM) receptors, whereas 3l showed >40-fold selectivity toward all other targets tested. Based on in vitro assays for assessment of permeability rates and extent, it is predicted that both compounds enter the brain of rats, non-human primates, as well as humans, and as such are good candidates to be carried forward for further evaluation as positron emission tomography (PET) ligands.     

Exploring the 2- and 5-positions of the pyrazolo[4,3-d]pyrimidin-7-amino scaffold to target human A1 and A2A adenosine receptors

A new series of 7-aminopyrazolo[4,3-d]pyrimidine derivatives (1–31) were synthesized to evaluate some structural modifications at the 2- and 5-positions aimed at shifting affinity towards the human (h) A_2A adenosine receptor (AR) or both hA_2A and hA₁ ARs. The most active compounds were those featured by a 2-furyl or 5-methylfuran-2-yl moiety at position 5, combined with a benzyl or a substituted-benzyl group at position 2. Several of these derivatives (22–31) displayed nanomolar affinity for the hA_2A AR (K_i = 3.62–57 nM) and slightly lower for the hA₁ ARs, thus showing different degrees (3–22 fold) of hA_2A versus hA₁ selectivity. In particular, the 2-(2-methoxybenzyl)-5-(5-methylfuran-2-yl) derivative 25 possessed the highest hA_2A and hA₁ AR affinities (K_i = 3.62 nM and 18 nM, respectively) and behaved as potent antagonist at both these receptors (cAMP assays). Its 2-(2-hydroxybenzyl) analog 26 also showed a high affinity for the hA_2A AR (K_i = 5.26 nM) and was 22-fold selective versus the hA₁ subtype. Molecular docking investigations performed at the hA_2A AR crystal structure and at a homology model of the hA₁ AR allowed us to represent the hypothetical binding mode of our derivatives and to rationalize the observed SARs.     

Synthesis and pharmacological characterization of a new series of 5,7-disubstituted-[1,2,4]triazolo[1,5-a][1,3,5]triazine derivatives as adenosine receptor antagonists: A preliminary inspection of ligand–receptor recognition process

A new series of triazolotriazines variously substituted at the C5 and N7 (5–25) positions was synthesized and fully characterized at the four adenosine receptor (AR) subtypes. In particular, arylacetyl or arylcarbamoyl moieties were introduced at the N7 position, which enhanced affinity at the hA_2B and hA₃ ARs, respectively, when utilized on the pyrazolo-triazolopyrimidine nucleus as we reported in the past. In general, compounds with a free amino group at the 7 position (5, 6), showed good affinity at the rat (r) A_2A AR (range 18.3–96.5 nM), while the introduction of a phenylcarbamoyl moiety at the N7 position (12, 19, 24) slightly increased the affinity at the hA₃ AR (range 311–633 nM) with respect to the unsubstituted derivatives. The binding profiles of the synthesized analogues seemed to correlate with the substitutions at the C5 and N7 positions. At the hA_2B AR, derivative 5, which contained a free amino group at the 7 position, was the most potent (EC₅₀ 3.42 μM) and could represent a starting point for searching new non-xanthine hA_2B AR antagonists. Molecular models of the rA_2A and hA₃ ARs were constructed by homology to the recently reported crystallographic structure of the hA_2A AR. A preliminary receptor-driven structure–activity relationship (SAR) based on the analysis of antagonist docking has been provided.     

Inhibitory effect of novel tetrahydropyrimidine-2(1H)-thiones on melanogenesis

The series of imidazoldine-2-thiones 2 and tetrahydropyrimidine-2-thiones 3 were discovered as inhibitor of α-MSH-induced melanin production in melanoma B16 cells. The primary bioassay showed that 1-(4-ethylbenzyl)-tetrahydropyrimidine-2(1H)-thione 3e (>100% inhibition at 10 μM, IC₅₀ = 1.2 μM) and 1-(4-tert-butylbenzyl)-tetrahydropyrimidine-2(1H)-thione 3f (>100% inhibition at 10 μM, IC₅₀ = 0.76 μM) exhibited potent inhibitory effect against α-MSH-induced melanin production. Compounds 3 inhibit the biosynthesis of tyrosinase without affecting its catalytic activity in melanogenesis.     

Synthesis and biological evaluation of a series of benzoxazole/benzothiazole-containing 2,3-dihydrobenzo[b][1,4]dioxine derivatives as potential antidepressants

A series of benzoxazole/benzothiazole-2,3-dihydrobenzo[b][1,4]dioxine derivatives (5a–5d and 8a–8j) was synthesized. Compounds were evaluated for binding affinities at the 5-HT_1A and 5-HT_2A receptors. Antidepressant activities of the compounds were screened using the forced swimming test (FST) and the tail suspension test (TST). The results indicated that the compounds exhibited high affinities for the 5-HT_1A and 5-HT_2A receptors and showed a marked antidepressant-like activity. Compound 8g exhibited high affinities for the 5-HT_1A (K_i = 17 nM) and 5-HT_2A (K_i = 0.71 nM) receptors; it also produced a decrease of the immobility time and exhibited potent antidepressant-like effects in the FST and TST in mice.     

Antiviral and cytotoxic activities of aminoarylazo compounds and aryltriazene derivatives

Twelve aminoarylazocompounds (A–C) and 46 aryltriazene 7 derivatives (D–G) have been synthesized and evaluated in cell-based assays for cytotoxicity and antiviral activity against a panel of 10 RNA and DNA viruses.Eight aminoazocompounds and 27 aryltriazene derivatives exhibited antiviral activity, sometimes of high level, against one or more viruses. A marked activity against BVDV and YFV was prevailing among the former compounds, while the latter type of compounds affected mainly CVB-2 and RSV. None of the active compounds inhibited the multiplication of HIV-1, VSV and VV.Arranged in order of decreasing potency and selectivity versus the host cell lines, the best compounds are the following; BVDV: 1 > 7 > 8 > 4; YFV: 7 > 5; CVB-2: 25 > 56 > 18; RSV: 14 > 20 > 55 > 38 > 18 > 19; HSV-1: 2. For these compounds the EC₅₀ ranged from 1.6 μM (1) to 12 μM (18), and the S. I. from 19.4 (1) to 4.2 (2).Thus the aminoarylazo and aryltriazene substructures appear as interesting molecular component for developing antiviral agents against ss RNA viruses, particularly against RSV and BVDV, which are important human and veterinary pathogens.Finally, molecular modeling investigations indicated that compounds of structure A–C, active against BVDV, could work targeting the viral RNA-dependent RNA-polymerase (RdRp), having been observed a good agreement between the trends of the estimated IC₅₀ and the experimental EC₅₀ values.     

Synthesis and structure–activity relationships of new carbonyl guanidine derivatives as novel dual 5-HT2B and 5-HT7 receptor antagonists

To identify potent dual 5-HT_2B and 5-HT₇ receptor antagonists, we synthesized a series of novel carbonyl guanidine derivatives and examined their structure–activity relationships. Among these compounds, N-(9-hydroxy-9H-fluorene-2-carbonyl)guanidine (10) had a good in vitro profile, that is, potent affinity for human 5-HT_2B and 5-HT₇ receptor subtypes (K_i = 1.8 nM and K_i = 17.6 nM, respectively) and high selectivity over 5-HT_2A, 5-HT_2C, α₁, D₂ and M₁ receptors. Compound 10 also showed a suppressing effect on 5-HT-induced dural protein extravasation in guinea pigs when orally administered.     

Synthesis of theophylline derivatives and study of their activity as antagonists at adenosine receptors

The synthesis of oligo(ethylene glycol)-alkene substituted theophyllines in positions 7 and/or 8 is described. The binding activity at adenosine receptors of selected derivatives was studied. Compound 2 showed high affinity for human A_2B receptor (K_i = 4.16 nM) with a selectivity K_iA2A/K_iA2B of 24.1, and a solubility in water of 1 mM. The alkenyl substituent in some of the theophylline derivatives allows for covalent attachment of them onto hydrogen-terminated silicon substrate surfaces via hydrosilylation. Alternatively, an azido group was incorporated to an oligo(ethylene glycol)theophylline derivative as an anchor for tethering the molecules on ethynyl presenting surfaces via click reaction.     

Synthesis and pharmacological characterization of novel xanthine carboxylate amides as A2A adenosine receptor ligands exhibiting bronchospasmolytic activity

The carboxylate amides of 8-phenyl-1,3-dimethylxanthine described herein represent a new series of selective ligands of the adenosine A_2A receptors exhibiting bronchospasmolytic activity. The effects of location of 8-phenyl substitutions on the adenosine receptor (AR) binding affinities of the newly synthesized xanthines have also been studied. The compounds displayed moderate to potent binding affinities toward various adenosine receptor subtypes when evaluated through radioligand binding studies. However, most of the compounds showed the maximum affinity for the A_2A subtype, some with high selectivity versus all other subtypes. Xanthine carboxylate amide 13b with a diethylaminoethylamino moiety at the para-position of the 8-phenylxanthine scaffold was identified as the most potent A_2A adenosine receptor ligand with K_i = 0.06 μM. Similarly potent and highly A_2A-selective are the isovanillin derivatives 16a and 16d. In addition, the newly synthesized xanthine derivatives showed good in vivo bronchospasmolytic activity when tested in guinea pigs.     

Synthesis of dammarane-type triterpene derivatives and their ability to inhibit HIV and HCV proteases

We synthesized dammarane-type triterpene derivatives and evaluated their ability to inhibit HIV-1 and HCV proteases to understand their structure–activity relationships. All of the mono- and di-succinyl derivatives (5a–5f) were powerful inhibitors of HIV-1 protease (IC₅₀ < 10 μM). However, only di-succinyl (5e) and 2,3-seco-2,3-dioic acid (3b) derivatives similarly inhibited HCV protease (IC₅₀ < 10 μM). A-nor dammarane-type triterpenes (4a and 4b, IC₅₀ 10.0 and 29.9 μM, respectively) inhibited HIV-1 protease moderately or strongly, but were inactive against HCV protease. All compounds that powerfully inhibited HIV-1 or HCV protease did not appreciably inhibit the general human proteases, renin and trypsin (IC₅₀ > 1000 μM). These findings indicated that the mono-succinyl dammarane type derivatives (5a–5d) selectively inhibited HIV-1 protease and that the di-succinyl (5e, 5f) as well as 2,3-seco-2,3-dioic acid (3b) derivatives preferably inhibited both viral proteases.     

1,3-Dialkyl-8-N-substituted benzyloxycarbonylamino-9-deazaxanthines as potent adenosine receptor ligands: Design,synthesis, structure–affinity and structure–selectivity relationships

A number of 1,3-dialkyl-9-deazaxanthines (9-dAXs), bearing a variety of N-substituted benzyloxycarbonylamino substituents at position 8, were prepared and evaluated for their binding affinity to the recombinant human adenosine receptors (hARs), chiefly to the hA_2B and hA_2A AR subtypes. Several ligands endowed with excellent binding affinity to the hA_2B receptors, but low selectivity versus hA_2A and hA₁ were identified. Among these, 1,3-dimethyl-N-3′-thienyl carbamate 15 resulted as the most potent ligand at hA_2B (K_i = 0.8 nM), with a low selectivity versus hA_2A (hA_2A/hA_2B = 12.6) and hA₁ (hA₁/hA_2B = 12.5) and a higher selectivity versus hA₃ (hA₃/hA_2B = 454). When tested in functional assays in vitro, compound 15 exhibited high antagonist activities and efficacies versus both the A_2A and A_2B receptor subtypes, with pA₂ values close to the corresponding pK_is. A comparative analysis of structure–affinity and structure–selectivity relationships of the similar analogues 8-N-substituted benzyloxycarbonylamino- and 8-N-substituted phenoxyacetamido-9-dAXs suggested that their binding modes at the hA_2B and hA_2A ARs may strongly differ. Computational studies help to clarify this striking difference arising from a simple, albeit crucial, structural change, from CH₂OCON to OCH₂CON, in the para-position of the 8-phenyl ring.     

Synthesis and biological activity of 5-styryl and 5-phenethyl-substituted 2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indoles

Syntheses, biological evaluation, and structure–activity relationships for a series of novel 5-styryl and 5-phenethyl analogs of dimebolin are disclosed. The novel derivatives and dimebolin share a broad spectrum of activities against therapeutically relevant targets. Among all synthesized derivatives, 2,8-dimethyl-5-[(Z)-2-phenylvinyl]-2,3,4,5-tetrahydro-1H-pyrido[4,3-b]indole and its 5-phenethyl analog are the most potent blockers of 5-HT₇, 5-HT₆, 5-HT_2C, Adrenergic α₂ and H₁ receptors. The general affinity rank order towards the studied receptors was Z-3(2) > 4(2) ? 4(3) ? dimebolin, all of them having highest affinities to 5-HT₇ receptors.     

Synthesis and pharmacological evaluation of optically pure,novel carbonyl guanidine derivatives as dual 5-HT2B and 5-HT7 receptor antagonists

A series of 9-disubstituted N-(9H-fluorene-2-carbonyl)guanidine derivatives have been discovered as potent and orally active dual 5-HT_2B and 5-HT₇ receptor antagonists. Upon screening several compounds, N-(diaminomethylene)-4′,5′-dihydro-3′H-spiro[fluorene-9,2′-furan]-2-carboxamide (17) exhibited potent affinity for both 5-HT_2B (K_i = 5.1 nM) and 5-HT₇ (K_i = 1.7 nM) receptors with high selectivity over 5-HT_2A, 5-HT_2C, α₁, D₂ and M₁ receptors. Optical resolution of the intermediate carboxylic acid 16 via the formation of diastereomeric salts using chiral alkaloids gave the optically pure compounds (R)-17 and (S)-17. Both enantiomers suppressed 5-HT-induced dural protein extravasation in guinea pigs in a dose-dependent manner and the amount of leaked protein was suppressed to near normal levels when orally administrated at 10 mg/kg. (R)-17 and (S)-17 were therefore selected as candidates for human clinical trials.     

Structure–activity relationships of truncated adenosine derivatives as highly potent and selective human A3 adenosine receptor antagonists

On the basis of potent and selective binding affinity of truncated 4′-thioadenosine derivatives at the human A₃ adenosine receptor (AR), their bioisosteric 4′-oxo derivatives were designed and synthesized from commercially available 2,3-O-isopropylidene-d-erythrono lactone. The derivatives tested in AR binding assays were substituted at the C2 and N⁶ positions. All synthesized nucleosides exhibited potent and selective binding affinity at the human A₃ AR. They were less potent than the corresponding 4′-thio analogues, but showed still selective to other subtypes. The 2-Cl series generally were better than the 2-H series in view of binding affinity and selectivity. Among compounds tested, compound 5d (X = Cl, R = 3-bromobenzyl) showed the highest binding affinity (K_i = 13.0 ± 6.9 nM) at the hA₃ AR with high selectivity (at least 88-fold) in comparison to other AR subtypes. Like the corresponding truncated 4′-thio series, compound 5d antagonized the action of an agonist to inhibit forskolin-stimulated adenylate cyclase in hA₃ AR-expressing CHO cells. Although the 4′-oxo series were less potent than the 4′-thio series, this class of human A₃ AR antagonists is also regarded as another good template for the design of A₃ AR antagonists and for further drug development.     

Synthesis and biological investigation of new equatorial (β) stereoisomers of 3-aminotropane arylamides with atypical antipsychotic profile

A series of novel 3β-aminotropane derivatives containing a 2-naphthalene or a 2-quinoline moiety was synthesised and evaluated for their affinity for 5-HT_1A, 5-HT_2A and D₂ receptors. Their affinity for the receptors was in the nanomolar to micromolar range. p-Substitution (6c, 6f, 6i, 6l, 6o), as well as substitution with chlorine atoms (6g, 6h, 6i), led to a significant increase in binding affinity for D₂ receptors with compounds 6f (K_i = 0.6 nM), 6c and 6i (K_i = 0.4 nM), having the highest binding affinities. m-Substituted derivatives were the most promising ligands in terms of 5-HT_2A receptor binding affinity whereas 2-quinoline derivatives (10a, 10b) displayed the highest affinity for 5-HT_1AR and were the most selective ligands with K_i = 62.7 nM and K_i = 30.5 nM, respectively. Finally, the selected ligands 6b, 6d, 6e, 6g, 6h, 6k, 6n and 6o, with triple binding activity for the D₂, 5-HT_1A and 5-HT_2A receptors, were subjected to in vivo tests, such as those for induced hypothermia, climbing behaviour and the head twitch response, in order to determine their pharmacological profile. The tested ligands presented neither agonist nor antagonist properties for the 5-HT_1A receptors in the induced hypothermia and lower lip retraction (LLR) tests. All tested compounds displayed antagonistic activity against 5-HT_2A, with 6n and 6o being the most active. Four (6b, 6k, 6n and 6o) out of eight tested compounds could be classified as D₂ antagonists. Additionally, evaluation of metabolic stability was performed for selected ligands, and introduction of halogen atoms into the benzene ring of 6h, 6k, 6n and 6o improved their metabolic stability. The project resulted in the selection of the lead compounds 6n and 6o, which had antipsychotic profiles, combining dopamine D₂-receptor and 5-HT_2A antagonism and metabolic stability.     

Discovery of simplified N2-substituted pyrazolo[3,4-d]pyrimidine derivatives as novel adenosine receptor antagonists: Efficient synthetic approaches,biological evaluations and molecular docking studies

In the present study, a molecular simplification approach was employed to design novel bicyclic pyrazolo[3,4-d]pyrimidine (PP) derivatives from tricyclic pyrazolo[4,3-e]-1,2,4-triazolo-[1,5-c]pyrimidines (PTP) as promising human A₃ adenosine receptor (hA₃AR) antagonists. All the target compounds were synthesized using novel and efficient synthetic schemes and the structure–activity relationship studies of these PPs were explored through the synthesis of a series of PTP analogues with various substituents. Substituents with different lipophilicity and steric hindrance (e.g., alkyl and aryl–alkyl) functions were introduced at N² position of the pyrazole ring, while acyl groups with different electronic properties were introduced at C⁶ position of the bicyclic nucleus to probe both electronic and positional effects. Most of the synthesized derivatives of the PP series presented good affinity at the hA₃AR, as indicated by the low micromolar range of K_i values and among them, compound 63 with N² neopentyl substituents showed most potent hA₃AR affinity with K_i value of 0.9 μM and high selectivity (hA₁AR/hA₃AR = >111 & hA_2AAR/hA₃AR = >111) towards other adenosine receptor subtypes. Interestingly, small isopropyl groups at N² position displayed high affinity at another receptor subtype (hA_2AAR, e.g., compound 55, with K_i hA_2AAR = 0.8 μM), while they were less favorable at the hA₃AR. Molecular docking analysis was also performed to predict the possible binding mode of target compounds inside the hA₃AR and hA_2AAR. Overall, PP derivatives represent promising starting points for new AR antagonists.     

Synthesis and pharmacological evaluation of new arylpiperazines N-{4-[4-(aryl) piperazine-1-yl]-phenyl}-amine derivatives: Putative role of 5-HT1A receptors

In an attempt to design novel 5-HT_1A agonists/partial agonists, based on an arylpiperazine nucleus, a series of N-{4-[4-(aryl)piperazine-1-yl]-phenyl}-amine derivatives were synthesized and biologically tested. The anxiolytic effect of the compounds was investigated employing the Elevated plus Maze (EPM) task. On the basis of in vivo functional test, compound 1c (3 mg/kg) and 4c (3 mg/kg) induced significant increments in open arm entries and time on EPM as compared to Buspirone. The anxiolytic effects of compounds 1c and 4c were effectively antagonized by WAY-100635, a 5-HT_1A receptor antagonist (0.5 mg/kg). Furthermore, we have also evaluated the concentration of 5-HT in the brain tissue using HPLC with fluorescent detection. Our result showed that serotonin levels were significantly decreased by ～38% (p < 0.001) and ～32% (p < 0.001) after acute administration of compounds 1c and 4c, respectively. These findings suggest that the anxiolytic like activity of these new arylpiperazines is mediated via 5-HT_1A receptors in the brain.     

Natural ortho-dihydroxyisoflavone derivatives from aged Korean fermented soybean paste as potent tyrosinase and melanin formation inhibitors

Natural o-dihydroxyisoflavone (ODI) derivatives with variable hydroxyl substituent at the aromatic ring of isoflavone and three known isoflavones were isolated from five-year-old Korean fermented soybean paste (Doenjang) and evaluated as potent inhibitors on tyrosinase activity and melanin formation in melan-a cells comparing with other known isoflavones, 7,8,4′-trihydroxyisoflavone (1) and 7,3′,4′-trihydroxyisoflavone (2) inhibited tyrosinase by 50% at a concentration of 11.21 ± 0.8 μM and 5.23 ± 0.6 μM (IC₅₀), respectively, whereas, 6,7,4′-trihydroxyisoflavone (3), daidzein (4), glycitein (5) and genistein (6) showed very low inhibition activity. Furthermore, those compounds significantly suppressed the cellular melanin formation by 50% at a concentration of 12.23 ± 0.7 μM (1), 7.83 ± 0.7 μM (2), and 57.83 ± 0.5(6) and show more activity than arbutin. But, compounds 3, 4, and 5 showed lower inhibition activity. This study shows that the position of hydroxyl substituent at the aromatic ring of isoflavone plays an important role in the intracellular regulation of melanin formation in cell-based assay system.     

Design and synthesis of 4-aryl-4-oxobutanoic acid amides as calpain inhibitors

The involvement of μ-calpain in neurological disorders, such as stroke and Alzheimer’s disease has attracted considerable interest in the use of calpain inhibitors as therapeutic agents. 4-Aryl-4-oxobutanoic acid amide derivatives 4 were designed as acyclic variants of μ-calpain inhibitory chromone and quinolinone derivatives. Of the compounds synthesized, 4c-2, which possesses a 2-methoxymethoxy group at the phenyl ring and a primary amide at the warhead region most potently inhibited μ-calpain (IC₅₀ = 0.34 μM). Our findings suggest that the 4-aryl-4-oxobutanoic acid amide derivatives should be considered as a new family of μ-calpain inhibitors.