Indoloquinoxaline derivatives as promising multi-functional anti-Alzheimer agents.

To confront a disease like Alzheimer's disease having complex pathogenesis, development of multitarget-directed ligands has emerged as a promising drug discovery approach. In our endeavor towards the development of multitarget-directed ligands for Alzheimer's disease, a series of indoloquinoxaline derivatives were designed and synthesized. In vitro cholinesterase inhibition studies revealed that all the synthesized compounds exhibited moderate to good cholinesterase inhibitory activity. 6-(6-(Piperidin-1-yl)hexyl)-6H-indolo[2,3-b]quinoxaline 9f was identified as the most potent and selective BuChE inhibitor (IC50 = 0.96 µM, selectivity index = 0.17) that possessed 2 fold higher BuChE inhibitory activity compared to the commercially approved reference drug donepezil (IC50 = 1.87 µM). Moreover, compound 9f is also endowed with self-induced Aß1-42 aggregation inhibitory activity (51.24% inhibition at 50 µM concentration). Some of the compounds of the series also displayed moderate anti-oxidant activity. To perceive a putative binding mode of the compound 9f, molecular docking studies were carried out, and the results pointed out significant interactions of compound 9f with the enzymes in the binding sites of cholinesterases as well as Aß1-42. Additionally, compound 9f exhibited favorable in silico ADMET properties. Put together these findings project compound 9f as a potential multitarget-directed ligand in the direction of developing novel anti-AD drugs.Communicated by Ramaswamy H. Sarma.

Carbazole-based semicarbazones and hydrazones as multifunctional anti-Alzheimer agents.

With the aim to combat a multi-faceted neurodegenerative Alzheimer's disease (AD), a series of carbazole-based semicarbazide and hydrazide derivatives were designed, synthesized and assessed for their cholinesterase (ChE) inhibitory, antioxidant and biometal chelating activity. Among them, (E)-2-((9-ethyl-9H-carbazol-3-yl)methylene)-N-(pyridin-2-yl)hydrazinecarbothioamide (62) and (E)-2-((9-ethyl-9H-carbazol-3-yl)methylene)-N-(5-chloropyridin-2-yl)hydrazinecarbothioamide (63) emerged as the premier candidates with good ChE inhibitory activities (IC50 values of 1.37 µM and 1.18 µM for hAChE, IC50 values of 2.69 µM and 3.31 µM for EqBuChE, respectively). All the test compounds displayed excellent antioxidant activity (reduction percentage of DPPH values for compounds (62) and (63) were 85.67% and 84.49%, respectively at 100 µM concentration). Compounds (62) and (63) conferred specific copper ion chelating property in metal chelation study. Molecular docking studies of compounds (62) and (63) indicate strong interactions within the active sites of both the ChE enzymes. Besides that, these compounds also exhibited significant in silico drug-like pharmacokinetic properties. Thus, taken together, they can serve as a starting point in the designing of multifunctional ligands in pursuit of potential anti-AD agents that might further prevent the progression of ADs.Communicated by Ramaswamy H. Sarma.

Further Studies on Triazinoindoles as Potential Novel Multitarget-Directed Anti-Alzheimer's Agents.

The inadequate clinical efficacy of the present anti-Alzheimer's disease (AD) drugs and their low impact on the progression of Alzheimer's disease in patients have revised the research focus from single targets to multitarget-directed ligands. A novel series of substituted triazinoindole derivatives were obtained by introducing various substituents on the indole ring for the development of multitarget-directed ligands as anti-AD agents. The experimental data indicated that some of these compounds exhibited significant anti-AD properties. Among them, 8-(piperidin-1-yl)-N-(6-(pyrrolidin-1-yl)hexyl)-5H-[1,2,4]triazino[5,6-b]indol-3-amine (60), the most potent cholinesterase inhibitor (AChE, IC50 value of 0.32 µM; BuChE, IC50 value of 0.21 µM), was also found to possess significant self-mediated Aß1-42 aggregation inhibitory activity (54% at 25 µM concentration). Additionally, compound 60 showed strong antioxidant activity. In the PAMPA assay, compound 60 exhibited blood-brain barrier penetrating ability. An acute toxicity study in rats demonstrated no sign of toxicity at doses up to 2000 mg/kg. Furthermore, compound 60 significantly restored the cognitive deficits in the scopolamine-induced mice model and Aß1-42-induced rat model. In the in silico ADMET prediction studies, the compound satisfied all the parameters of CNS acting drugs. These results highlighted the potential of compound 60 to be a promising multitarget-directed ligand for the development of potential anti-AD drugs.

Novel carbazole-stilbene hybrids as multifunctional anti-Alzheimer agents.

Molecules capable of engaging with multiple targets associated with pathological condition of Alzheimer's disease have proved to be potential anti-Alzheimer's agents. In our goal to develop multitarget-directed ligands for the treatment of Alzheimer's disease, a novel series of carbazole-based stilbene derivatives were designed by the fusion of carbazole ring with stilbene scaffold. The designed compounds were synthesized and evaluated for their anti-AD activities including cholinesterase inhibition, Aß aggregation inhibition, antioxidant and metal chelation properties. Amongst them, (E)-1-(4-(2-(9-ethyl-9H-carbazol-3-yl)vinyl)phenyl)-3-(2-(pyrrolidin-1-yl)ethyl)thiourea (50) appeared to be the best candidate with good inhibitory activities against AChE (IC50 value of 2.64 µM) and BuChE (IC50 value of 1.29 µM), and significant inhibition of self-mediated Aß1-42 aggregation (51.29% at 25 µM concentration). The metal chelation study showed that compound (50) possessed specific copper ion chelating property. Additionally, compound (50) exhibited moderate antioxidant activity. To understand the binding mode of 50, molecular docking studies were performed, and the results indicated strong non-covalent interactions of 50 with the enzymes in the active sites of AChE, BuChE as well as of the Aß1-42 peptide. Additionally, it showed promising in silico ADMET properties. Putting together, these findings evidently showed compound (50) as a potential multitarget-directed ligand in the course of developing novel anti-AD drugs.

Novel Multitarget Directed Triazinoindole Derivatives as Anti-Alzheimer Agents.

The multifaceted nature of Alzheimer's disease (AD) demands treatment with multitarget-directed ligands (MTDLs) to confront the key pathological aberrations. A novel series of triazinoindole derivatives were designed and synthesized. In vitro studies revealed that all the compounds showed moderate to good anticholinesterase activity; the most active compound 23e showed an IC50 value of 0.56 ± 0.02 µM for AChE and an IC50 value of 1.17 ± 0.09 µM for BuChE. These derivatives are also endowed with potent antioxidant activity. To understand the plausible binding mode of the compound 23e, molecular docking studies and molecular dynamics simulation studies were performed, and the results indicated significant interactions of 23e within the active sites of AChE as well as BuChE. Compound 23e successfully diminished H2O2-induced oxidative stress in SH-SY5Y cells and displayed excellent neuroprotective activity against H2O2 as well as Aß-induced toxicity in SH-SY5Y cells in a concentration dependent manner. Furthermore, it did not show any significant toxicity in neuronal SH-SY5Y cells in the cytotoxicity assay. Compound 23e did not show any acute toxicity in rats at doses up to 2000 mg/kg, and it significantly reversed scopolamine-induced memory deficit in mice model. Additionally, compound 23e showed notable in silico ADMET properties. Taken collectively, these findings project compound 23e as a potential balanced MTDL in the evolution process of novel anti-AD drugs.

Breast cancer amelioration by Butea monosperma in-vitro and in-vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Butea monosperma belonging to family Fabaceae is used in the Indian traditional medicine (Ayurveda) for various ailments including abdominal tumors and possess anti-estrogenic activity. AIM OF THE STUDY: The present study is aimed at investigating the chemo-preventive potential of Butea monosperma in breast cancer and elucidating it's mechanism of action by assessing its effect on key processes like apoptosis, angiogenesis and metastasis. METHODS: Cytotoxic potential of methanol extract of Butea monosperma flower (MEBM) was tested in MCF-7 (estrogen receptor positive), MDA-MB-231 (triple negative) and MDA-MB-453 (HER2 positive) human breast cancer cells by MTT assay. Chemo-preventive potential was evaluated in-vivo in Methylnitrosourea (MNU) induced mammary cancer in nulliparous Sprague-Dawley rats. The mechanism for anticancer potential was screened by in-vitro studies involving Annexin V- FITC assay (apoptosis), Chick Chorioallantoic Membrane assay (angiogenesis) and Migration assay (metastasis). Statistical analysis was done by one way and two way ANOVA (for Growth Rate and feed consumption efficiency) followed by post hoc Bonferroni's test with P value < 0.05. RESULTS: It is observed that the exposure of MEBM, at various concentrations and time intervals to different cell lines, resulted in decreased cell proliferation. The IC50 value of MCF-7 cells was found significantly less than that of MDA-MB-231 and MDA-MB-453 cells, which indicated that the extract of said medicinal plant were more potent inhibitors of estrogen positive breast cancer cells than other types of breast cancer cells in vitro. Corroborative evidences were acquired in MNU actuated mammary carcinogenesis where MEBM constricted tumor parameters, decreased expression of estrogen and progesterone, nucleic acid content and increased latency period. MEBM also induced apoptosis, inhibited angiogenesis and metastasis in-vitro. CONCLUSION: Selective cytotoxic activity in MCF-7 estrogen positive breast cancer cells and inhibition of growth of mammary carcinoma in-vivo by methanol extract of Butea monosperma flowers (MEBM) suggests chemo-prevention through modulation of estrogen and progesterone receptor, apoptotic, anti-angiogenesis and anti-metastatic activity.

Evaluation of the effect of Punica granatum juice and punicalagin on NFκB modulation in inflammatory bowel disease.

Punica granatum L. (Lythraceae) inhibits cancer cell proliferation and apoptosis through the modulation of cellular transcription factors and signaling proteins. No pharmacological work is reported on the effects of P. granatum juice on the cellular signaling pathways involved in initiation and progression of inflammation. The present investigation evaluates the effect of P. granatum juice (PJ) and purified punicalagin (PW) on nuclear factor kappa B (NFκB) and the signaling pathways leading to its expression in colon inflammation. Male Sprague-Dawley rats were divided into six groups: positive and negative control, vehicle (50 % ethanol), standard (5-ASA 100 mg/kg, p.o.), PJ (400 mg/kg, p.o.), PW (4 mg/kg, p.o.). Colitis was induced with 2,4-dinitrobenzene sulfonic acid and animals were euthanized on 18th day. Colon samples collected were subjected to various histological assessment (CMDI, DAI), and biochemical parameters (MPO, MDA, SOD, NO). Gene expression study was carried out using RT-PCR for cytokines (TNF-α, IL-1ß, IL-18 and NF-κß). Pretreatment with PJ and PW significantly (p < 0.05) lowered the disease extent and severity as indicated by reduction in CMDI (2 ± 0.31) and DAI (1.83(#) ± 0.22) when compared with DNBS-treated rats (3.83* ± 0.17). Gene expression studies showed decreased mRNA levels of TNF-α, IL-18, and IL-1ß in PJ and PW-treated groups. NF-κß mRNA levels were found to be reduced 84 and 64 % by PJ and PW, respectively. These results suggest that P. granatum juice is more biologically active over punicalagin alone and can be potentially used for the treatment of inflammatory bowel disease.

Validated HPTLC Method for Quantification of Luteolin and Apigenin in Premna mucronata Roxb., Verbenaceae.

A simple, rapid, and precise high-performance thin-layer chromatographic method was developed for quantitative estimation of luteolin and apigenin in Premna mucronata Roxb., family Verbenaceae. Separation was performed on silica gel 60 F254 HPTLC plates using toluene : ethyl acetate : formic acid (6 : 4 : 0.3) as mobile phase for elution of markers from extract. The determination was carried out in fluorescence mode using densitometric absorbance-reflection mode at 366 nm for both luteolin and apigenin. The methanolic extract of Premna mucronata was found to contain 10.2 mg/g % luteolin and 0.165 mg/g % of apigenin. The method was validated in terms of linearity, LOD and LOQ, accuracy, precision, and specificity. The calibration curve was found to be linear between 200 and 1000 ng/band for luteolin and 50 and 250 ng/band for apigenin. For luteolin and apigenin, the limit of detection was found to be 42.6 ng/band and 7.97 ng/band while the limit of quantitation was found to be 129.08 ng/band and 24.155 ng/band, respectively. This developed validated method is capable of quantifying and resolving luteolin and apigenin and can be applicable for routine analysis of extract and plant as a whole.

Validated HPTLC method for quantitative determination of gallic acid in stem bark of Myrica esculenta Buch.-Ham. ex D. Don, myricaceae.

A simple, rapid, and precise HPTLC method was developed for quantitative estimation of gallic acid in stem bark of Myrica esculenta, family Myricaceae. Separation was performed on silica gel 60F254 HPTLC plates using toluene-ethyl acetate-formic acid-methanol (3 + 3 + 0.6 + 0.4, v/v/v/v) mobile phase for separation of the extracted components. The determination was carried out in the UV densitometric absorbance-reflection mode at 280 nm. The amount of gallic acid in free and combined form in the stem bark powder was found to be 0.276 and 0.541%, respectively, on a dry weight basis. The method was validated in terms of linearity, accuracy, precision, and specificity according to International Conference on Harmonization guidelines. Gallic acid response was found to be linear over a broad concentration range of 0.4-2.0 microg/band. LOD and LOQ were 0.103 and 0.312 microg/spot, respectively. The developed method is capable of quantifying amounts of gallic acid in stem bark powder of M. esculenta.

Is cytochrome modulation the new frontier for decreasing the risk of cataract?

AIM: The present study was designed to study the effect of cytochrome P450 (CYP) modulators on the occurrence of cataract using male Sprague-Dawley rats weighing 40:50 gm. MATERIALS AND METHODS: Macroscopical examination of the lens isolated from rats pretreated with diltiazem (30 mg/kg; once daily; PO) showed delayed occurrence of cataract while pioglitazone (3.8 mg/kg; once daily; PO) pretreatment demonstrated an early cataract. RESULTS AND CONCLUSION: A delayed occurrence of cataract with diltiazem (CYP inhibitor) and an early onset of cataract with pioglitazone (CYP inducer) indicate that a cytochrome P450 mediated pathway may affect the initiation of cataract but not the maturation pattern.

Evaluation of bronchodilator and anti-anaphylactic activity of Myrica sapida.

BACKGROUND: Asthma is a chronic inflammatory disorder of the airways. The available treatment options have major limitations owing to low efficacy, associated adverse events and compliance issues. Therefore, the health burden of bronchial asthma is increasing globally at an alarming rate, providing a strong impetus for the development of new therapeutics. Myrica sapida is known traditionally in Ayurveda to possess anti-asthmatic activity. Hence, the present investigation was undertaken to evaluate the bronchodilator and anti-anaphylactic activity of the stem bark of Myrica sapida. METHODS: Experimental models studied were acetylcholine induced bronchospasm in guinea pigs, egg albumin induced anaphylaxis in guinea pigs, in vitro studies on tracheal strip of egg albumin sensitized guinea pigs. RESULTS: Treatment with ethanolic extract of M. sapida, 75 mg/kg, orally resulted in significant protection against acetylcholine aerosol induced bronchospasm and allergen induced anaphylaxis in guinea pigs. Ethanolic extract of M. sapida (75 mg/kg, p.o.) prevented the potentiation of responses and also produced a decrease in pD2 value of histamine and acetylcholine in guinea pig tracheal strip. CONCLUSION: These results suggest that M. sapida possesses bronchodilator activity, has potent inhibitory effect on immediate hyper-sensitivity reactions and decreases bronchial hyper-responsiveness.

Novel hepatocyte growth factor (HGF) binding domains on fibronectin and vitronectin coordinate a distinct and amplified Met-integrin induced signalling pathway in endothelial cells.

BACKGROUND: The growth of new blood vessels in adult life requires the initiation of endothelial cell migration and proliferation from pre-existing vessels in addition to the recruitment and differentiation of circulating endothelial progenitor cells. Signals emanating from growth factors and the extracellular matrix are important in regulating these processes. RESULTS: Here we report that fibronectin (FN) and vitronectin (VN) modulate the responses of endothelial cells to HGF (Scatter Factor), an important pro-angiogenic mediator. Novel binding sites for HGF were identified on both FN and VN that generate molecular complexes with enhanced biological activity and these were identified in the supernatants of degranulated platelet suspensions implicating their release and formation in vivo. In the absence of co-stimulation with an ECM glycoprotein, HGF could not promote endothelial cell migration but retained the capacity to induce a proliferative response utilising the Map kinase pathway. Through promoting Met-Integrin association, HGF-FN and HGF-VN complexes coordinated and enhanced endothelial cell migration through activation of the PI-3 kinase pathway involving a Ras-dependent mechanism whereas a Ras-independent and attenuated migratory response was promoted by co-stimulation of cells with HGF and a non-binding partner ECM glycoprotein such as collagen-1. CONCLUSIONS: These studies identify a novel mechanism and pathway of HGF signalling in endothelial cells involving cooperation between Met and integrins in a Ras dependent manner. These findings have implications for the regulation of neovascularization in both health and disease.