Mapping the Melatonin Receptor. 8. Selective MT2 Agonists Derived from 5,6-Dihydroindolo[2,1-a]isoquinolines and Related Systems.

A series of substituted indolo[2,1-a]isoquinolines and indolo[1,2-a]benzoxazines have been prepared, as melatonin analogues, to investigate the nature of the binding site of the melatonin receptor. Agonist and antagonist potency of all the analogues was measured using the [35S]GTPγS binding assay protocol. The binding affinity of the analogues were measured by competition binding studies against the human MT1 (hMT1) and MT2 (hMT2) receptors stably transfected in Chinese Hamster Ovarian (CHO) cells, using 2-[125 I]-iodomelatonin, as a ligand. N-Acetyl 2-(10-methoxy-5,6-dihydroindolo[2,1-a]isoquinolin-12-yl)propyl-1-amine (12 a) binds strongly to both the hMT1 and hMT2 receptors, and shows a preference for the hMT2, as does its propanamido counterpart 12 b. The introduction of two methyl groups into their side chain, analogues 15 a and 15 b, leads to antagonism, in the case of the former, and drastically diminishes its hMT1 binding; an analogous profile is seen for 15 b, which, however, is a partial agonist. Introduction of chlorine or methoxy groups into ring 4 gives compounds, that are weakly binding, with a preference for MT2. Substitution of oxygen for carbon at position 5 gives the indolo[1,2-c]benzoxazines 33, 36 a and b, that bind strongly to the human receptors, 33, 36 b being potent agonists at the melatonin receptors, but do not discriminate between hMT1 and hMT2.

Synthesis and evaluation of novel 2,4-disubstituted arylthiazoles against T. brucei.

The design, synthesis and pharmacological evaluation of the 4-substituted-2-[3-(adamant-1-yl)-4-fluorophenyl]thiazoles 1a-j, the 4-substituted-2-[4-(adamant-1-yl)phenyl]thiazoles 2a-h, the 2-substituted-4-[4-(adamant-1-yl)phenyl]thiazoles 3a-e, the N-substituted 2-phenylthiazol-4-ethylamides 4a, b and the N-substituted 4-phenylthiazol-2-ethylamides 4c, d is described. Compounds 1a and 2a exhibit trypanocidal activity in the range of IC50 = 0.42 µM and IC50 = 0.80 µM, respectively. Both of these derivatives bear a lipophilic end, which consists of a 4-(1-adamantyl) phenyl or a 3-(1-adamantyl)phenyl moiety, a 1,3-thiazole ring and a functional end, which comprises of an alkylamine and can be considered as promising candidates for the treatment of Trypanosoma brucei infections.

Development and Characterization of Eudragit®-Based Electrospun Nanofibrous Mats and Their Formulation into Nanofiber Tablets for the Modified Release of Furosemide.

Furosemide, a chloride channel blocker ordinarily used as a high-ceiling or loop diuretic, is practically insoluble in water and dilute acids. Due to its acidic nature, furosemide is mostly absorbed in the stomach and in the upper small intestine. Efforts have focused on the development of sustained release systems of furosemide in order to improve the effectiveness of the drug, which exhibits poor aqueous solubility and poor permeability. Recently, electrospun nanofibrous drug delivery systems have emerged as promising alternative solid-dosage forms due to their advantages of high porosity, high surface to volume ratio, and high drug-loading efficacy. Herein, a number of nanofibrous mats composed of different types of Eudragit® polymers in various concentrations and combinations loaded with furosemide were designed, successfully electrospun, and characterized using SEM, FTIR, DSC, and TGA analyses. The nanofibrous nonwovens were formulated in nanofiber tablets and the release profile of furosemide from them was evaluated at pH 1.2 and 6.8 and compared to that of physical mixture matrix tablets of analogous composition as well as to that of a commercial formulation. It was found that the release of furosemide was compatible with the gastroretentive and slower intestinal release requirements with a well-defined absorption window, while some nanofiber formulations could act as furosemide carriers in emergency situations where a relatively fast onset of its action is required, as in the case of critically ill post-traumatic patients.

Novel 2,6-diketopiperazine-derived acetohydroxamic acids as promising anti-Trypanosoma brucei agents.

Aim: Identification of new, effective and selective trypanocidal agents. Materials & methods: Twelve novel acetohydroxamic acid derivatives based on 2-alkyl-2-aryl-2,6-diketopiperazine scaffolds have been synthesized and evaluated in vitro for their growth inhibitory activity against bloodstream form Trypanosoma brucei. Results: All the analogs were remarkably potent inhibitors, with low micromolar to submicromolar activities. Structure-activity relationship studies demonstrated that the presence of an alkyl substituent at the N(4)-position of the 2,6-diketopiperazine ring portion was, in general, beneficial to trypanocidal activity in this series. Conclusion: The highest activity resulted from the introduction of a methyl, n-propyl or n-butyl substituent to the N(4)-position of the parent compound. Importantly, the most potent analogs were found to be highly selective against T. brucei with respect to mammalian cells.

Fluorine substituted methoxyphenylalkyl amides as potent melatonin receptor agonists.

A series of fluorine substituted methoxyphenylalkyl amides were prepared with different orientations of the fluorine and methoxy groups with respect to the alkylamide side chain and with alkyl sides of differing lengths (n = 1-3). ß-Dimethyl and α-methyl derivatives were also synthesised. The compounds were tested as melatonin agonists and antagonists using the pigment aggregation of Xenopus melanophores as the biological assay. A number of these compounds were potent melatonin agonists, the potency depending on the length of the alkyl chain, the orientation of the methoxy and fluorine substituents, the amide chain length and, for the ethyl side-chain analogues, the presence of ß-substituents.

Synthesis and Evaluation of Nifurtimox-Adamantane Adducts with Trypanocidal Activity.

The synthesis and pharmacological evaluation of C1-substituted adamantane hydrazones, their C2-substituted isomers, and C1-substituted adamantane furanoic carboxamides is described. These new adamantane derivatives exhibited an interesting pharmacological profile in terms of trypanocidal activity and selectivity. The most active adduct with the best selectivity in this study was found to be the phenylacetoxy hydrazone 1 b (2-[4-(tricyclo[3.3.1.13,7 ]dec-1-yl)phenyl]-N'-[(5-nitrofuran-2-yl)methylene]acetohydrazide; EC50 =11±0.9 nm, SITb =770).

Synthesis of diphenoxyadamantane alkylamines with pharmacological interest.

In this work, the synthesis and the pharmacological evaluation of diphenoxyadamantane alkylamines Ia-f and IIa-f is described. The new diphenoxy-substituted adamantanes share structural features present in trypanocidal and antitubercular agents. 1-Methylpiperazine derivative Ia is the most potent against T. brucei compound, whilst its hexylamine congener IIf exhibits a significant antimycobacterial activity.

Modified In Vitro Release of Melatonin Loaded in Nanofibrous Electrospun Mats Incorporated Into Monolayered and Three-Layered Tablets.

Modified release tablet formulations with melatonin (MLT) are clinically more useful in initiating and maintaining sleep in elderly insomniacs, compared with those designed for immediate release. Aiming at the modified release of MLT, monolayered and 3-layered tablets, incorporating nanofibrous mats composed of cellulose acetate and polyvinylpyrrolidone loaded with MLT, were prepared and studied. In vitro dissolution profiles of MLT in gastrointestinal-like fluids revealed tableting pressure/pH-dependence. The release of the hormone from physical mixture tablets was generally slower from the nanofibers-based tablets, thus exhibiting in the latter case properties that are necessary for the control of both the sleep-onset and the maintenance dysfunctions. The nature of the excipients (hydroxypropylmethylcellulose or lactose monohydrate) used in this study to produce 3-layered tablets was also found to affect the release of MLT, adjusting it to the endogenous hormone's chronobiotic profile. The release of MLT from formulation F(nf)2 (nanofiber mats incorporated into 3-layered tablets containing lactose monohydrate both in the upper and lower layers) was found to be in closer alignment with these effects than the other delivery systems.

Fabrication and Characterization of Electrospun Nanofibers for the Modified Release of the Chronobiotic Hormone Melatonin.

OBJECTIVE: Aiming at the modified release of melatonin (MLT), electrospun-MLT loaded nanofibers, filled into hard gelatin and DRcapsTM capsules, were used as formulants. METHODS: Cellulose acetate, polyvinylpyrrolidinone and hydroxypropylmethylcellusose (HPMC 2910) were used for the preparation of the fiber matrices through electrospinning. The in vitro modified release profile of MLT from the fabricated matrices in gastrointestinal-like fluids was studied. At pH 1.2, the formulations CA1, CA2, PV1, HP1, HP2 and the composite formulations CAPV1-CAPV5 in hard gelatin capsules exhibited fast MLT release. RESULTS: In general, the same trend was observed at pH 6.8, with the exception of CAPV1 and CAPV2. These two composite formulations delivered 52.08% and 75.25% MLT, respectively at a slower pace (6 h) when encapsulated in DRcapsTM capsules. In all other cases, the release of MLT from DRcapsTM capsules filled with the MLT-loaded nanofibers reached 100% at 6h. CONCLUSION: These findings suggest that the MLT-loaded nanofibrous mats developed in this study exhibit a promising profile for treating sleep dysfunctions.

Lipophilic conformationally constrained spiro carbocyclic 2,6-diketopiperazine-1-acetohydroxamic acid analogues as trypanocidal and leishmanicidal agents: An extended SAR study.

We have previously described a number of lipophilic conformationally constrained spiro carbocyclic 2,6-diketopiperazine (2,6-DKP)-1-acetohydroxamic acids as potent antitrypanosomal agents. In this report, we extend the SAR analysis in this class of compounds with respect to in vitro growth inhibition of Trypanosoma and Leishmania parasites. Introduction of bulky hydrophobic substituents at the vicinal position of the basic nitrogen atom in the spiro carbocyclic 2,6-DKP ring system can provide analogues which are potently active against bloodstream form Trypanosoma brucei and exhibit significant activities toward Trypanosoma cruzi epimastogotes and Leishmania infantum promastigotes and intracellular amastigotes. In particular, compounds possessing a benzyl or 4-chlorobenzyl substituent were found to be the most active growth inhibitors, with activities in the low nanomolar and low micromolar ranges for T. brucei and L. infantum, respectively. The benzyl-substituted (S)-enantiomer was the most potent derivative against T. brucei (IC50  = 6.8 nm), T. cruzi (IC50  = 0.21 µm), and L. infantum promastigotes (IC50  = 2.67 µm) and intracellular amastigotes (IC50  = 2.60 µm). Moreover, the (R)-chiral benzyl-substituted derivative and its racemic counterpart displayed significant activities against L. donovani. Importantly, the active compounds show high selectivity in comparison with two mammalian cell lines.

Synthesis of Adamantane Aminoethers with Antitubercular Potential.

BACKGROUND: Intrigued by the fact that aminoadamantane derivatives, bearing the active 1,2-ethylenediamine moiety, are promising antitubercular agents, we report herein the synthesis and the antitubercular evaluation of N,N'-substituded-4,4'-[adamantane-2,2-diyl]bis(phe-noxyalkylamines) 1a-g, N,N'-substituded-4,4'-[adamantane-1,3-diyl]bis(phenoxyalkylamines) 2a-f, N,N'- substituded-[4-(1-adamantyl)phenoxy]alkylamines 3a-d and N,N'-substituded-[4-(2-adamantyl)- phenoxy]alkylamines 4a,b. METHOD: A substituted diarylmethane moiety was introduced on the adamantane skeleton of the new derivatives. The synthesis of the above compounds involved the nucleophilic attack of the corresponding phenoxide, to the appropriate aminoalkylchloride hydrochloride under heating. RESULTS: The double substituted adamantane derivatives with an aminoether side chain exhibit significant activity against Mycobacterium tuberculosis. CONCLUSION: The length and the nature of the amino end of the side chain influence the antitubercular activity. The double phenolic substitution of the adamantane scaffold and the aminoether side chain with a three-methylene spacer between the phenoxy group and the nitrogen atom present the better results. (analogues 1f,g and 2e,f). These findings merit further investigation aiming at the design of more potent adamantane antituberculars, bearing a number of different substituents on the diarylmethane pharmacophore, which will also be translocated to other posititions on the adamantane ring.

Novel 1-(2-aryl-2-adamantyl)piperazine derivatives with antiproliferative activity.

Novel 1-(2-aryl-2-adamantyl)piperazine derivatives have been synthesized and evaluated in vitro for their antitumor properties against HeLa cervical carcinoma, MDA MB 231 breast cancer, MIA PaCa2 pancreatic cancer, and NCI H1975 non-small cell lung cancer. The parent piperazine 6 was found to exhibit a reasonable activity toward the HeLa and MDA MB 231 tumor cell lines (IC50= 9.2 and 8.4 µΜ, respectively). Concurrent benzene ring C4-fluorination and piperidine acetylation of the piperazino NH of compound 6 resulted in the most active compound 13 of the series in both of the above cell lines (IC50=8.4 and 6.8 µΜ, respectively). Noticeably, compounds 6 and 13 exhibited a significantly low cytotoxicity level over the normal human cells HUVEC (Human Umbilical Vein Endothelial Cells) and NHDF (Normal Human Dermal Fibroblasts).

Benzocyclobutane, benzocycloheptane and heptene derivatives as melatonin agonists and antagonists.

Two series of analogues were designed, synthesised and evaluated as potential human melatonin type 1 and 2 receptor (hMT1 and hMT2 ) ligands. Their biological effects were assessed by a well-established, specific model of melatonin action, the pigment response of Xenopus laevis melanophores. Compounds containing a benzocyclobutane scaffold and a methoxy group in the "melatonin" orientation were found to be potent agonists, with one of the analogues exhibiting activity comparable to melatonin. In contrast, analogues with a methoxy group in non-melatonin positions or with multiple methoxy groups showed either weaker agonist activity or were antagonists. Benzocycloheptene derivatives with one methoxy group are found to be weak agonists, whereas those with two methoxy groups were found to be antagonists, as were all of the benzocycloheptane derivatives evaluated. The most active compounds were assessed in a human receptor radio ligand binding assay but showed little discrimination between MT1 and MT2 . These results again show that the indole nitrogen of melatonin is not a necessary component for analogue activity and also illustrate that replacement of the indole ring with a 4-membered carbocycle can provide highly active compounds when the methoxy group is in the melatonin position.

An E/Z conformational behaviour study on the trypanocidal action of lipophilic spiro carbocyclic 2,6-diketopiperazine-1-acetohydroxamic acids.

An explanation for the vast difference observed in the trypanocidal activity between the new secondary (N-methylated) hydroxamic acids 5 and 6, and their primary (nonmethylated) congeners 1a and 2, based on their E/Z conformational behaviour in DMSO, is presented.

New adamantane phenylalkylamines with σ-receptor binding affinity and anticancer activity, associated with putative antagonism of neuropathic pain.

The synthesis of the adamantane phenylalkylamines 2a-d, 3a-c, and 4a-e is described. These compounds exhibited significant antiproliferative activity, in vitro, against eight cancer cell lines tested. The σ(1), σ(2), and sodium channel binding affinities of compounds 2a, 3a, 4a, and 4c-e were investigated. The most interesting analogue, 4a, exhibited significant in vivo anticancer profile on pancreas, prostate, leukemia, and ovarian cancer cell line xenografts together with apoptosis and caspase-3 activation. Inhibition of the cancer cells cycle at the sub-G1 level was also obtained with 4a. Finally, encouraging results were observed with 4a in vivo on mice, suggesting putative antimetastatic and analgesic activities of this compound.

Conformational analysis of two novel cytotoxic C2-substituted pyrrolo[2,3-f]quinolines in aqueous media, organic solvents, membrane bilayers and at the putative active site.

We have performed: (i) conformational analysis of two novel cytotoxic C2-substituted pyrrolo[2,3-f]quinolines 5e and 5g in deuterated dimethylsulfoxide (DMSO-d(6)) utilizing NOE results from NMR spectroscopy; (ii) molecular dynamics (MD) calculations in water, DMSO and dimyristoyl phosphatidylcholine bilayers and (iii) molecular docking and MD calculations on DNA nucleotide sequences. The obtained results for the two similar in structure molecules showed differences in: (i) their conformational properties in silico and in media that reasonably simulate the biological environment; (ii) the way they are incorporated into the lipid bilayers and therefore their diffusion ability and (iii) molecular docking capacity as it is depicted from their different binding scores.

Synthesis, σ1, σ2-receptors binding affinity and antiproliferative action of new C1-substituted adamantanes.

The synthesis of N-{4-[a-(1-adamantyl)benzyl]phenyl}piperazines 2a-e is described. The in vitro antiproliferative activity of most compounds against main cancer cell lines is significant. The σ(1), σ(2)-receptors and sodium channels binding affinity of compounds 2 were investigated. One of the most active analogs, 2a, had an interesting in vivo anticancer profile against the BxPC-3 and Mia-Paca-2 pancreas cancer cell lines with caspase-3 activation, which was associated with an anagelsic activity against the neuropathic pain.

New adamantane derivatives with sigma affinity and antiproliferative activity.

The synthesis of 4-(1-adamantyl)-4,4-diarylbutylamines 1, 5-(1-adamantyl)-5,5-diarylpentylamines 2 and 6-(1-adamantyl)-6,6-diarylhexylamines 3 is described and the σ1, σ2-receptors and sodium channels binding affinity of compounds 1 were investigated. The in vitro activity of compounds 1, 2 and 3 against main cancer cell lines is significant. One of the most active analogs, 1a, had an interesting in vivo anticancer profile against the ovarian cancer cell line IGROV-1, which was associated with an anagelsic activity against the neuropathic pain induced by the main anticancer drugs.

Design and synthesis of Trypanosoma brucei active 1-alkyloxy and 1-benzyloxyadamantano 2-guanylhydrazones.

Treating African trypanosomiasis: The synthesis and biological evaluation of novel 1-alkyloxy and 1-benzyloxyadamantano 2-guanylhydrazones, their 1-dioxa congeners and two 1-benzyladamantano 2-guanylhydrazones is reported. Preliminary structure-activity relationship data were elucidated and lead compounds suitable for further optimization were discovered.

Design and synthesis of bioactive 1,2-annulated adamantane derivatives.

Adamantanopyrrolidines 8, 9 and 10, adamantanopyrrolidines 16 and 18, adamantanoxazolone 20, adamantanopyrazolone 23, adamantanopyrazolothione 24 and adamantanocyclopentanamine 32 were synthesized and tested for anti-influenza A virus and trypanocidal activity. The stereoelectronic requirements for optimal antiviral and trypanocidal potency were investigated. Pyrrolidine 16 proved to be the most active of the compounds tested against influenza A virus, being 4-fold more active than amantadine, equipotent to rimantadine and 19-fold more potent than ribavirin. Oxazolone 20 showed significant trypanocidal activity against bloodstream forms of the African trypanosome, Trypanosoma brucei, being approximately 3 times more potent than rimantadine and almost 50-fold more active than amantadine.