An In Silico and an In Vitro Inhibition Analysis of Glycogen Phosphorylase by Flavonoids, Styrylchromones, and Pyrazoles

Glycogen phosphorylase (GP) is a key enzyme in the glycogenolysis pathway. GP inhibitors are currently under investigation as a new liver-targeted approach to managing type 2 diabetes mellitus (DM). The aim of the present study was to evaluate the inhibitory activity of a panel of 52 structurally related chromone derivatives; namely, flavonoids, 2-styrylchromones, 2-styrylchromone-related derivatives [2-(4-arylbuta-1,3-dien-1-yl)chromones], and 4- and 5-styrylpyrazoles against GP, using in silico and in vitro microanalysis screening systems. Several of the tested compounds showed a potent inhibitory effect. The structure–activity relationship study indicated that for 2-styrylchromones and 2-styrylchromone-related derivatives, the hydroxylations at the A and B rings, and in the flavonoid family, as well as the hydroxylation of the A ring, were determinants for the inhibitory activity. To support the in vitro experimental findings, molecular docking studies were performed, revealing clear hydrogen bonding patterns that favored the inhibitory effects of flavonoids, 2-styrylchromones, and 2-styrylchromone-related derivatives. Interestingly, the potency of the most active compounds increased almost four-fold when the concentration of glucose increased, presenting an IC50 < 10 µM. This effect may reduce the risk of hypoglycemia, a commonly reported side effect of antidiabetic agents. This work contributes with important considerations and provides a better understanding of potential scaffolds for the study of novel GP inhibitors.

Synthesis and preliminary biological evaluation of new phthalazinone derivatives with PARP-1 and cholinesterase inhibitory activities

The inhibition of Poly (ADP-ribose) polymerases-1 (PARP-1) has a potentially therapeutical value for AD. In order to search for a new agent based on multitarget-directed ligands (MTDLs) strategy, a series of 21 novel compounds incorporated the respective pharmacophores of two marketed drugs, namely 4-benzyl phthalazinone moiety of PARP-1 inhibitor Olaparib and N-benzylpiperidine moiety of AChE inhibitor Donepezil, into one molecule was synthesized. The inhibitory activities of all the synthesized compounds against the enzymes PARP-1, AChE and BChE were evaluated. Among them, 30 exhibited the most potent inhibitory effect on PARP-1 enzyme (8.18±2.81 nM) and moderate BChE inhibitory activity (1.63±0.52µM), while its AChE inhibitory activity (13.48±2.15µM) was weaker than Donepezil (0.04±0.01µM). Further molecular docking studies revealed that four hydrogen bonds were formed between 30 and PARP-1 which were similar with the interactions between Olaparib and PARP-1. 30 interacted with the critical residues His438 and Trp82 of huBChE through hydrogen bond and hydrophobic interaction which were necessary for huBChE inhibitory potency. Our research gave a clue to search for new agents based on PARP-1 and cholinesterase dual-inhibited activities to treat Alzheimer's disease.

Inhibitory Effect of Ascorbic Acid on in vitro Enzymatic Digestion of Raw and Cooked Starches

Ascorbic acid, also known as vitamin C, was previously reported to inhibit the activity of pancreatic α-amylase, the primary digestive enzyme for starch. A major implication of such inhibition is a slowed rate of starch digestion into glucose, which thereby reduces postprandial hyperglycemia. The aim of this study was to explore the inhibitory effects of ascorbic acid at various concentrations on the in vitro digestion of high amylose maize starch (HAMS) and potato starch (PS) in both raw and cooked conditions. Resistant starch (RS) content, defined as the starch that remained after 4 h of simulated in vitro enzymatic digestion, was measured for the starch samples. Upon the addition of ascorbic acid, the RS contents increased in both raw and cooked starches. Cooking significantly reduced the RS contents as compared to raw starches, and less increase in RS was observed with the addition of ascorbic acid. The inhibitory effect of ascorbic acid on the digestion of raw starches showed a dose-dependent trend until it reached the maximum extent of inhibition. At the concentrations of 12.5 and 18.75 mg/mL, ascorbic acid exhibited the most potent inhibitory effect on the in vitro starch digestion in raw and cooked conditions, respectively. Overall, our results strongly indicate that ascorbic acid may function as a glycemic modulatory agent beyond other important functions, and its effects persist upon cooking with certain concentrations applied.

Synthesis and Biological Activity Evaluation of Novel 5-Methyl-7-phenyl-3H-thiazolo[4,5-b]pyridin-2-ones

A series of 5-methyl-7-phenyl-3H-thiazolo[4,5-b]pyridin-2-ones has been designed, synthesized, and characterized by spectral data. Target compounds were screened for their antimicrobial activity against some pathogenic bacteria and fungi, and most of them showed moderate activity, especially compound 3g, which displayed the potent inhibitory effect against Pseudomonas aeruginosa and Escherichia coli with MIC value of 0.21 μM. The active thiazolopyridine derivatives 3c, 3f, and 3g were screened for their cytotoxicity effects on HaCat, Balb/c 3T3 cells using MTT assay, which revealed promising results. In silico assessment for compounds 3c, 3f, and 3g also revealed suitable drug-like parameters and ADME properties. The binding interactions of the most active compound 3g were performed through molecular docking against MurD and DNA gyrase, with binding energies and an inhibitory constant compared to the reference drug ciprofloxacin. The tested thiazolo[4,5-b]pyridines constitute an exciting background for the further development of new synthetic antimicrobial agents.

Chromane Derivatives from Underground Parts of Iris tenuifolia and Their In Vitro Antimicrobial, Cytotoxicity and Antiproliferative Evaluation

Phytochemical investigation of the ethanol extract of underground parts of Iris tenuifolia Pall. afforded five new compounds; an unusual macrolide termed moniristenulide (1), 5-methoxy-6,7-methylenedioxy-4-O-2′-cycloflavan (2), 5,7,2′,3′-tetrahydroxyflavanone (3), 5-hydroxy-6,7-dimethoxyisoflavone-2′-O-β-d-glucopyranoside (9), 5,2′,3′-dihydroxy-6,7-dimethoxyisoflavone (10), along with seven known compounds (4–8, 11–12). The structures of all purified compounds were established by analysis of 1D and 2D NMR spectroscopy and HR-ESI-MS. The antimicrobial activity of the compounds 1–3, 5, 9, and 10 was investigated using the agar diffusion method against fungi, Gram-positive and Gram-negative bacteria. In consequence, new compound 3 was found to possess the highest antibacterial activity against Enterococcus faecalis VRE and Mycobacterium vaccae. Cell proliferation and cytotoxicity tests were also applied on all isolated compounds and plant crude extract in vitro with the result of potent inhibitory effect against leukemia cells. In particular, the newly discovered isoflavone 10 was active against both of the leukemia cells K-562 and THP-1 while 4–6 of the flavanone type compounds were active against only THP-1.

Evaluation of Hedgehog Pathway Inhibitors as a Therapeutic Option for Uterine Leiomyosarcoma Using the Xenograft Model

Uterine leiomyosarcoma (LMS) contributes to a significant proportion of uterine cancer deaths. It is a rare and high-risk gynecological cancer. LMS is challenging to the treatment due to the resistance of several therapies. The activation of the Hedgehog (HH) pathway has been reported in several types of female cancers. Uterine LMS presents an upregulation of the crucial HH signaling pathway members such as SMO and GLI1. Although targeting the HH pathway exhibited a potent inhibitory effect on the phenotype of uterine LMS in vitro, the effect of the HH inhibitors on LMS growth in vivo has not been identified. The present study aimed to assess the effect of Hedgehog pathway inhibitors (SMO-LDE225 and GLI-Gant61) as a therapeutic option in the xenograft model of uterine LMS. The results demonstrated that LDE225 treatment did not show any inhibitory effect on LMS tumor growth; however, treatment with GLI inhibitor (Gant61) induced a remarkable tumor regression with a significant decrease in Ki67 expression, compared to control (p < 0.01). Moreover, administration of Gant61 decreased the expression of GLI1, GLI target genes BMP4 and c-MYC (p < 0.05), indicating that the HH pathway is implicated in the LMS experimental model. In conclusion, our studies demonstrate for the first time that GLI inhibitor (Gant61), but not SMO inhibitor (LDE225), shows a potent inhibitory effect on LMS tumor growth and concomitantly suppresses the expression of GLI1- and GLI-targeted genes using the xenograft model of uterine LMS.

Kechuangping Mixture Inhibits LPS‑induced Macrophage Immune Response Through Suppressing the TLR4/MyD88/NF-κB Signal Pathway

Background: Kechuangping mixture (KCPM) based on Maxingshigan decoction has a significant clinical effect in the treatment of pediatric pneumonia, especially for asthmatic pneumonia. This study explored the effects of KCPM on lipopolysaccharide (LPS)-induced macrophage immune response and the molecular mechanism involved. Methods: CCK-8 assays was used to detect the effect of KCPM on cell viability. The RAW 264.7 macrophages were divided into four groups: blank, LPS, KCPM, LPS+KCPM. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to examine the secretion of inflammatory factors and the expression of TLR4/MyD88/NF-κB signal pathways with or without KCPM stimulation. Results: KCPM inhibited the differentiation of RAW264.7 macrophages induced by LPS. The secretion of pro-inflammatory cytokines IL-6 and TNF-α induced by LPS was decreased by pre-treatment with KCPM. While, treatment with KCPM advanced, the anti-inflammatory cytokines IL-10 and TGF-β increased significantly. Additionally, KCPM exhibited a potent inhibitory effect on the expression of iNOS and a promotive effect on the expression of Arg-1. Moreover, KCPM clearly suppressed the transcription level of TLR4, MyD88 and NF-κB signal pathways.Conclusion: KCPM promoted the macrophages polarization toward to M2 phenotype, as a result, limited the macrophage immune response. TLR4/MyD88/NF-κB signal pathways may play an important role in this process.

SPECTROPHOTOMETRIC METHOD DEVELOPMENT AND VALIDATION FOR SIMULTANEOUS ESTIMATION OF ABACAVIR AND LAMIVUDINE IN COMBINED DOSAGE FORM

Antiretroviral therapy [ART] has been developed remarkably within last three decades, since the first NRTI (nucleoside reverse transcriptase inhibitor) developed. For the mostly patients the challenge of complete and sustain suppression of viral has been clarify, since the triple therapy arrived. Convenient pill burden and tolerability are the main limiting factor for enhancing the long term benefit of ART. ABACAVIR (ABA) and LAMIVUDINE (LAM) are synthetic nucleotide analog (SNA) that showing a synergistic and potent inhibitory effect on human immunodeficiency virus-I (HIV-I), causative agent of AIDS. ABA and LAM belongs to nucleoside reverse transcriptase inhibitor (NRTI) class. As per new therapeutic strategy these two antiretroviral (ARV) drugs are required for the treatment of AIDS. When ARV regimens changed due to lack to virologic response ABA should be avoided. ABA is getting converted in to the active metabolite (carbovir triphosphate) which is an deoxyguanosine-5’ triphosphate analog by intracellular enzymes. LAM gets converted in its active 5’-triphosphate metabolite through phosphorylation. Chemically, ABA is (1S, cis)-4-[2-amino-6-(cyclopropyl amino)-9H-purin-9-yl]-2-cyclopentene-1-methanol sulphate and LAM is (2R, cis)-4-amino-1-(2-hydroxymethyl-1, 3-oxathiolan-5-yl)-(1H)-pyrimidin-2-one. Fig. 1 and 2 show ABA and LAM’s structure respectively.

Enzymatic Production of 3-OH Phlorizin, a Possible Bioactive Polyphenol from Apples, by Bacillus megaterium CYP102A1 via Regioselective Hydroxylation

Phlorizin is the most abundant glucoside of phloretin from the apple tree and its products. Phlorizin and its aglycone phloretin are currently considered health-beneficial polyphenols from apples useful in treating hyperglycemia and obesity. Recently, we showed that phloretin could be regioselectively hydroxylated to make 3-OH phloretin by Bacillus megaterium CYP102A1 and human P450 enzymes. The 3-OH phloretin has a potent inhibitory effect on differentiating 3T3-L1 preadipocytes into adipocytes and lipid accumulation. The glucoside of 3-OH phloretin would be a promising agent with increased bioavailability and water solubility compared with its aglycone. However, procedures to make 3-OH phlorizin, a glucoside of 3-OH phloretin, using chemical methods, are not currently available. Here, a biocatalytic strategy for the efficient synthesis of a possibly valuable hydroxylated product, 3-OH phlorizin, was developed via CYP102A1-catalyzed regioselective hydroxylation. The production of 3-OH phlorizin by CYP102A1 was confirmed by HPLC and LC–MS spectroscopy in addition to enzymatic removal of its glucose moiety for comparison to 3-OH phloretin. Taken together, in this study, we found a panel of mutants from B. megaterium CYP102A1 could catalyze regioselective hydroxylation of phlorizin to produce 3-OH phlorizin, a catechol product.

In-Silico Study of Some Natural Plant Phyto-compounds for the Identification of Novel Potent Cholinesterase Inhibitors Against Alzheimer Disease

The main aim of this study is to identify inhibitory binding potent of the available commercially alkaloids, against the crystal structure of acetylcholinesterase (AChE) protein by in silico studies. The inhibitory data of the compounds should be compared with the internal ligand as well as standard AChE inhibitor Aricept (which is used for the treatment of all stages of Alzheimer&rsquo;s disease). AutoDock 4.0 is used for the docking study, conformational orientation site analysis, and, with the help of docking, we have calculated parameters like binding energy and inhibition constant. Docking's study showed that Glabridin, Isorosmanol, Quercetin, Honokiol, Eckol, Sargaquinoic acid, and Ginsedosides revealed strong binding affinity with the enzyme. Moreover, The ADMET profiling and physicochemical properties of the selected compounds are evaluated using the Molinspiration and Data warrior software. By showing a strong binding affinity value, positive bioactivity score, and good pharmacokinetic properties, the top compound was determined. After evaluation with all parameters, the compound Glabridin and Ginsedosides show the most potent inhibitory effect towards the acetylcholinesterase, so this compound could be used as a novel is required to treat Alzheimer's disease.