Effect of S-Se Bioisosteric Exchange on Affinity and Intrinsic Efficacy of Novel N-acylhydrazone Derivatives at the Adenosine A2A Receptor.

In this work, we evaluated the conformational effect promoted by the isosteric exchange of sulfur by selenium in the heteroaromatic ring of new N-acylhydrazone (NAH) derivatives (3-8, 13, 14), analogues of the cardioactive compounds LASSBio-294 (1) and LASSBio-785 (2). NMR spectra analysis demonstrated a chemical shift variation of the iminic Csp2 of NAH S/Se-isosters, suggesting a stronger intramolecular chalcogen interaction for Se-derivatives. To investigate the pharmacological profile of these compounds at the adenosine A2A receptor (A2AR), we performed a previously validated functional binding assay. As expected for bioisosteres, the isosteric-S/Se replacement affected neither the affinity nor the intrinsic efficacy of our NAH derivatives (1-8). However, the N-methylated compounds (2, 6-8) presented a weak partial agonist profile at A2AR, contrary to the non-methylated counterparts (1, 3-5), which appeared as weak inverse agonists. Additionally, retroisosterism between aromatic rings of NAH on S/Se-isosters mimicked the effect of the N-methylation on intrinsic efficacy at A2AR, while meta-substitution in the phenyl ring of the acyl moiety did not. This study showed that the conformational effect of NAH-N-methylation and aromatic rings retroisosterism changed the intrinsic efficacy on A2AR, indicating the S/Se-chalcogen effect to drive the conformational behavior of this series of NAH.

Novel phosphatidylinositol 4-kinases III beta (PI4KIIIß) inhibitors discovered by virtual screening using free energy models.

Herein, the LASSBio Chemical Library is presented as a valuable source of compounds for screening to identify hits suitable for subsequent hit-to-lead optimization stages. A feature of the LASSBio Chemical Library worth highlighting is the fact that it is a smart library designed by medicinal chemists with pharmacological activity as the main priority. The great majority of the compounds part of this library have shown in vivo activity in animal models, which is an indication that they possess overall favorable bioavailability properties and, hence, adequate pharmacokinetic profiles. This, in turn, is supported by the fact that approximately 85% of the compounds are compliant with Lipinski's rule of five and ca. 95% are compliant with Veber's rules, two important guidelines for oral bioavailability. In this work it is presented a virtual screening methodology combining a pharmacophore-based model and an empirical Gibbs free energy-based model for the ligand-protein interaction to explore the LASSBio Chemical Library as a source of new hits for the inhibition of the phosphatidylinositol 4-kinase IIIß (PI4KIIIß) enzyme, which is related to the development of viral infections (including enteroviruses, SARS coronavirus, and hepatitis C virus), cancers and neurological diseases. The approach resulted in the identification of two hits, LASSBio-1799 (7) and LASSBio-1814 (10), which inhibited the target enzyme with IC50 values of 3.66 µM and IC50 and 6.09 µM, respectively. This study also enabled the determination of the structural requirements for interactions with the active site of PI4KIIIß, demonstrating the importance of both acceptor and donor hydrogen bonding groups for forming interactions with binding site residues Val598 and Lys549.

Recent Advances in Development of Polyphenols as Anticancer Agents.

Still now, for many forms of the disseminated cancers there is no curative therapy available. The discovery of novel active chemotherapeutic agents is largely essential to overcome this problem. Natural compounds polyphenols are mainly characterized by a huge structural variance; they can render them intrinsic dietary components due to their common occurrence in plants. Now-a-days, polyphenols (secondary metabolites) are characterized by a vast spectrum of physiological significance. From the past twenty years in the world of scientific research, polyphenols play an important role in a wide range of physiological processess. This review focuses on the development of polyphenols as antitumor agent in recent research studies.

LASSBio-1135: a dual TRPV1 antagonist and anti-TNF-alpha compound orally effective in models of inflammatory and neuropathic pain.

LASSBio-1135 is an imidazo[1,2-a]pyridine derivative with high efficacy in screening models of nociception and inflammation, presumed as a weak COX-2 inhibitor. In order to tease out its mechanism of action, we investigated others possible target for LASSBio-1135, such as TNF-α and TRPV1, to better characterize it as a multitarget compound useful in the treatment of chronic pain. TRPV1 modulation was assessed in TRPV1-expressing Xenopus oocytes against capsaicin and low pH-induced current. Modulation of TNF-α production was evaluated in culture of macrophages stimulated with LPS. In vivo efficacy of LASSBio-1135 was investigated in carrageenan and partial sciatic ligation-induced thermal hyperalgesia and mechanical allodynia. Corroborating its previous demonstration of efficacy in a model of capsaicin-induced hyperalgesia, LASSBio-1135 blocks capsaicin-elicited currents in a non-competitive way with an IC50 of 580 nM as well as low pH-induced current at 50 µM. As an additional action, LASSBio-1135 inhibited TNF-α release in these cells stimulated by LPS with an IC50 of 546 nM by reducing p38 MAPK phosphorilation. Oral administration of 100 µmol x Kg(-1) LASSBio-1135 markedly reduced thermal hyperalgesia induced by carrageenan, however at 10 µmol x Kg(-1) only a partial reduction was observed at the 4th h. Neutrophil recruitment and TNF-α production after carrageenan stimulus was also inhibited by the treatment with LASSBio-1135. Modulating TRPV1 and TNF-α production, two key therapeutic targets of neuropathic pain, 100 µmol x Kg(-1) LASSBio-1135 was orally efficacious in reversing thermal hyperalgesia and mechanical allodynia produced by partial sciatic ligation 7-11 days after surgery without provoking hyperthermia, a common side effect of TRPV1 antagonists. In conclusion LASSBio-1135, besides being a weak COX-2 inhibitor, is a non-competitive TRPV1 antagonist and a TNF-α inhibitor. As a multitarget compound, LASSBio-1135 is orally efficacious in a model of neuropathic pain without presenting hyperthermia.

New insights into pharmacological profile of LASSBio-579, a multi-target N-phenylpiperazine derivative active on animal models of schizophrenia.

Previous behavioral and receptor binding studies on N-phenylpiperazine derivatives by our group indicated that LASSBio-579, LASSBio-580 and LASSBio-581 could be potential antipsychotic lead compounds. The present study identified LASSBio-579 as the most promising among the three compounds, since it was the only one that inhibited apomorphine-induced climbing (5 mg/kg p.o.) and apomorphine-induced hypothermia (15 mg/kg p.o.). Furthermore, LASSBio-579 (0.5 mg/kg p.o.) was effective in the ketamine-induced hyperlocomotion test and prevented the prepulse inhibition deficits induced by apomorphine, DOI and ketamine with different potencies (1 mg/kg, 0.5 mg/kg and 5 mg/kg p.o., respectively). LASSBio-579 also induced a motor impairment, catalepsy and a mild sedative effect but only at doses 3-120 times higher than those with antipsychotic-like effects. In addition, LASSBio-579 (0.5 and 1 mg/kg p.o.) reversed the catalepsy induced by WAY 100,635, corroborating its action on both dopaminergic and serotonergic neurotransmission and pointing to the contribution of 5-HT(1A) receptor activation to its pharmacological profile. Moreover, co-administration of sub-effective doses of LASSBio-579 with sub-effective doses of clozapine or haloperidol prevented the apomorphine-induced climbing without induction of catalepsy. In summary, our results characterize LASSBio-579 as a multi-target ligand active in pharmacological animal models of schizophrenia, confirming that this compound could be included in development programs aiming at a new drug for treating schizophrenia.

Novel furfurylidene N-acylhydrazones derived from natural safrole: discovery of LASSBio-1215, a new potent antiplatelet prototype.

We describe herein the discovery of (E)-N-methyl-N'-((5-nitrofuran-2-yl)methylene)benzo[d]( 1 , 3 ) dioxole-5-carbohydrazide (9e), named LASSBio-1215, as a novel antiplatelet agent belonging to the N-methyl-N-acylhydrazone class, which exert their antiaggregating actions on human and rabbit platelets induced by different agonists, through cyclooxygenase-1 (COX-1) or thromboxane synthase inhibition. This compound was elected after screening of a series of functionalized furyl N-acylhydrazone derivatives, synthesized from natural safrole 10. In vitro assays showed that compound 9e presents platelet-aggregating activity in rabbit platelet-rich plasma (PRP) induced by arachidonic acid (IC(50) = 0.7 µM) and collagen (IC(50) = 4.5 µM). Moreover, LASSBio-1215 also inhibited almost completely the second wave of adenosine diphosphate-induced platelet aggregation in human PRP, and this effect was correlated with their ability to block the production of pro-aggregating autacoid thromboxane A(2).

The role of natural products in the discovery of new drug candidates for the treatment of neurodegenerative disorders II: Alzheimer's disease.

The present review is part II in a series (part I focuses on Parkinson's Disease) that addresses the value of natural product chemistry in the discovery of medicines for the treatment of neurodegenerative disorders. Data reviewed document that a host of products from plant species and derivatives have neuroprotectant effects in vitro and in vivo. In addition, besides neuroprotection, natural products also demonstrate biological effects that target biochemical pathways underlying associated symptoms of neurdegnerative disorders that include cognitive impairments, energy/fatigue, mood, and anxiety. This part of the review series focuses specifically upon Alzheimer's Disease (AD). AD is postulated to result from extracellular formation of amyloid plaques and intracellular deposits of neurofibrilary tangles in the hippocampus, cerebral cortex and other areas of the brain essential for cognitive function. Plaques are formed mostly from the deposition ß-amyloid (Aß), a peptide derived from the amyloid precursor protein (APP). Filamentous tangles are formed from paired helical filaments composed of neurofilament and hyperphosphorilated tau protein, a microtubule-associated protein. In addition, environmental factors can engender the production of cytokines that are closely related to the installation of an inflammatory process that contributes to neuronal death and the development and the progression of AD. In this review we focus on the recent main contribuitions of natural products chemistry to the discovery of new chemical entities usefull to the control and prevention of AD installation and progression. More than sixteen plant species, including Ginseng, Celastrus paniculatus, Centella asiatica, Curcuma longa, Ginkgo biloba, Huperzia serrata, Lycoris radiate, Galanthus nivalis, Magnolia officinalis, Polygala tenuifolia, Salvia lavandulaefolia, Salvia miltiorrhiza, Coptis chinensis, Crocus sativus, Evodia rutaecarpa, Sanguisorba officinalis, Veratrum grandiflorum and Picrorhiza kurvoa, are discussed as potential sources of active extracts. In addition, more than sixty secondary metabolites are under evaluation for their efficacy on controlling symptoms and to impede the development and progression of AD.

The role of natural products in the discovery of new drug candidates for the treatment of neurodegenerative disorders I: Parkinson's disease.

Neurodegenerative disorders such as Parkinson's disease (PD), Alzheimer's disease (AD), Huntington's disease (HD) and amyotrophic lateral sclerosis (ALS) are currently incurable pathologies with huge social and economic impacts closely related to the increasing of life expectancy in modern times. Although the clinical and neuropathological aspects of these debilitating disorders are distinct, they share a pattern of neurodegeneration in anatomically or functionally related regions. For each disease, presently available treatments only address symptoms and do not alter the course or progression of the underlying diseases. In this context, the search for new effective chemical entities, capable of acting on diverse biochemical targets, with new mechanisms of action and low toxicity are genuine challenges to research groups and the pharmaceutical industry. This medical need has led to the reemerging of modern natural products chemistry that has yielded sophisticated and complex new lead molecules for drug discovery and development. In this review we discuss some of the main contributions of the natural products chemistry that covers multiple and varied plant species. Advances in the discovery of active constituents of plants, herbs, and extracts prescribed by traditional medicine practices for the treatment of senile neurodegenerative disorders, especially for PD, in the period after the 2000s is reviewed. The most important contributions from the 1990s are also discussed. The review also focuses on the pharmacological mechanisms of action that might underlie the purported beneficial improvements in memory and cognition, neurovascular function, and in neuroprotection. It is concluded that natural product chemistry brings tremendous diversity and historical precedent to a huge area of unmet medical need.

Design of new dopamine D2 receptor ligands: biosynthesis and pharmacological evaluation of the hydroxylated metabolite of LASSBio-581.

LASSBio-581 is a N-phenylpiperazine derivative designed for the treatment of schizophrenia. In this study, four strains of filamentous fungi were screened for their capabilities to biotransform LASSBio-581. Cunninghamella echinulata ATCC 9244 was chosen to scale up the biosynthesis of the p-hydroxylated metabolite of LASSBio-581. The chemical structure of the metabolite was confirmed by NMR, LC-MS and X-ray crystallography. Binding studies performed on brain homogenate indicated that the p-hydroxylated metabolite can be considered more selective for dopamine receptors than LASSBio-581, and, therefore, can be used to design new selective dopamine inhibitors.

Design and synthesis of 3,4-methylenedioxy-6-nitrophenoxyacetylhydrazone derivatives obtained from natural safrole: new lead-agents with analgesic and antipyretic properties.

In this work, we reported the synthesis and evaluation of the analgesic, anti-inflammatory, and antipyretic properties of new 2-(6-nitro-benzo[1,3]dioxol-5-yloxy)-acetylhydrazone derivatives (3), designed exploring molecular hybridization and isosteric replacement approaches between nimesulide (1) and carbanalogue NAH series (2) developed at LASSBio. Target compounds were synthesized in very good yields exploiting abundant Brazilian natural product safrole (4) as starting material. The evaluation of the antinociceptive properties of this series led us to discover a new potent prototype of analgesic and antipyretic agent, that is, NAH derivative 3c, named LASSBio-891, which showed to be more potent than dipyrone used as standard.

Design, synthesis, and pharmacological evaluation of new neuroactive pyrazolo[3,4-b]pyrrolo[3,4-d]pyridine derivatives with in vivo hypnotic and analgesic profile.

The present study describes the synthesis and pharmacological profiles of four novel pyrazolo[3,4-b]pyrrolo[3,4-d]pyridine derivatives 2-5, which were structurally designed by using the sedative and analgesic drug zolpidem 1 as lead compound. The heterotricyclic system present in the target compounds 2-5 was constructed in good yields, exploiting a regioselective hetero Diels-Alder reaction of the key azabutadiene derivative 7 and functionalized N-phenylmaleimides 9-12. Additionally, we identified that 1-methyl-7-(4-nitrophenyl)-3-phenyl-3,6,7,8-tetrahydropyrazolo[3,4-b]pyrrolo[3,4-d]pyridine-6,8-dione derivative (LASSBio-873, 5) presented not only the most potent ability to promote sedation, which was similar to that induced by the standard benzodiazepine drug midazolam, but also potent central antinociceptive effect.

New selective acetylcholinesterase inhibitors designed from natural piperidine alkaloids.

Five new piperidine alkaloids were designed from natural (-)-3-O-acetyl-spectaline and (-)-spectaline that were obtained from the flowers of Senna spectabilis (sin. Cassia spectabilis, Leguminosae). Two semi-synthetic analogues (7 and 9) inhibited rat brain acetylcholinesterase, showing IC50 of 7.32 and 15.1 microM, and were 21 and 9.5 times less potent against rat brain butyrylcholinesterase, respectively. Compound 9 (1mg/kg, i.p.) was fully efficacious in reverting scopolamine-induced amnesia in mice. The two active compounds (7 and 9) did not show overt toxic effects at the doses tested in vivo.

Evaluating the prophylactic potential of the phtalimide derivative LASSBio 552 on allergen-evoked inflammation in rats.

A previous study showed that the novel tetrazolephtalimide derivative LASSBio 552 (2-4-[3-(1H-1,2,3,4-tetraazol-5-yl)propoxy]phenethyl-1,3-isoindolinedione) prevents LTD(4)-evoked tracheal contraction. This led us to examine the putative anti-inflammatory effect of LASSBio 552 in comparison with the leukotriene CysLT(1) receptor antagonist zafirlukast using a model of allergic pleurisy in rats. Treatment with either LASSBio 552 (24-96 micromol/kg, i.p.) or zafirlukast (9-72 micromol/kg, i.p.), 1 h before challenge, inhibited eosinophil and mononuclear cell influx into the pleural cavity 24 h post-challenge, but failed to alter the increased levels of eotaxin, plasma leakage, mast cell degranulation and neutrophil infiltration noted 6 h post-challenge. CD4(+) T cell recruitment 24 h post-challenge was also sensitive to LASSBio 552. This treatment failed to alter cysteinyl leukotriene production at 6 h, but clearly inhibited the phenomenon 24 h and 48 h post-challenge. In in vitro settings LASSBio 552 inhibited allergen-evoked cysteinyl leukotriene generation from isolated mast cells, while histamine release remained unchanged. It also slightly inhibited cysteinyl leukotriene production by eosinophils and mononuclear cells triggered by Ca(+2) ionophore A23187. A leukotriene CysLT(1) receptor transfected cell-based assay revealed that LASSBio 552 did not prevent LTD(4)-evoked Ca(+2) influx, indicating that it was not a leukotriene CysLT(1) receptor antagonist. These findings indicate that LASSBio 552 is able to inhibit eosinophil influx triggered by allergen chalenge in a mechanism at least partially associated with suppression of CD4(+) T cell influx and cysteinyl leukotriene production.

Further bioactive piperidine alkaloids from the flowers and green fruits of Cassia spectabilis.

The flowers of Cassia spectabilis yielded three new piperidine alkaloids, (-)-3-O-acetylspectaline (1), (-)-7-hydroxyspectaline (2), and iso-6-spectaline (3), together with the known (-)-spectaline (4). The green fruits of this plant were also investigated, resulting in the isolation of 1 and 4. Their structures were elucidated using a combination of multidimensional NMR and MS techniques, and relative stereochemistries were established by NOESY correlations and analysis of coupling constants. The DNA-damaging activity of these compounds was evaluated using a mutant yeast, Saccharomyces cerevisiae, assay.

Antinociceptive profile of (-)-spectaline: a piperidine alkaloid from Cassia leptophylla.

The antinociceptive activity of (-)-spectaline (1), a piperidine alkaloid isolated from Cassia leptophylla Vog. (Leguminosae), was investigated. We have also studied the acute oral toxicity of 1 in mice and it did not show any signals of toxicity in doses lower than 400 micromol/kg. The antinociceptive effect of 1 was evaluated on chemical (acetic acid, formalin and capsaicin) and thermal (hot plate and tail flick) pain models in mice, using classical standard drugs. Dipyrone ID (50) = 14.68 micromol/kg (4.8 mg/kg), indomethacin ID (50) = 0.78 micromol/kg (0.28 mg/kg) and (-)-spectaline ID (50) = 48.49 micromol/kg (15.75 mg/kg), all produced a significant inhibition of acetic acid-induced abdominal writhing in mice. (-)-Spectaline was inactive in the hyperalgesic model of formalin and did not show any central analgesic activity (hot plate and tail flick models). In the capsaicin-induced neurogenic pain model, (-)-spectaline presented an important inhibitory effect with an ID (50) = 20.81 microg/paw and dipyrone ID (50) = 19.89 microg/paw. The ensemble of results permitted us to identify 1 as an antinociceptive compound. The mechanism underlying this antinociceptive effect of 1 remains unknown, but the results suggest that such an effect could be related to pathways associated to vanilloid receptor systems.

New isoxazole derivatives designed as nicotinic acetylcholine receptor ligand candidates.

In this work we report the synthesis and evaluation of the analgesic properties of new isosteric heterocyclic derivatives, presenting the isoxazole nucleus, designed as nicotinic acetylcholine receptor ligand candidates, analogues to alkaloid epibatidine. Compound 2-(3-methyl-5-isoxazolyl)pyridine (3) presented the best analgesic profile of this series in hot plate test, which was partially prevented by pretreatment with nicotinic receptor antagonist mecamylamine.

Design, synthesis and pharmacological evaluation of novel pyrazolo[3,4-b]thieno[2,3-d]pyridine acid derivatives: a new class of anti-inflammatory and anti-platelet agents.

A series of pyrazolo[3,4-b]thieno[2,3-d]pyridine alkanoic acid derivatives has been synthesized and evaluated as thromboxane synthetase inhibitors and leukotriene D(4) receptor antagonists. The glutaric acid derivative LASSBio341 (6) was shown to be active in arachidonic acid-induced platelet aggregation (IC(50)=0.14 microM) and inhibition of the contraction of guinea pig tracheal strip induced with LTD(4) (IC(50) = 43.7 microM), displaying still in vivo anti-inflammatory profile.