Synthesis,in vitro and in vivo biological evaluation of novel lappaconitine derivatives as potential anti-inflammatory agents

Lappaconitine (LA), a natural compound with a novel C18-diterpenoid alkaloid skeleton, displayed extensive biological profile. Recent research on LA is focused mainly on its anti-tumor and analgesic effects, and therefore we aimed to investigate its anti-inflammatory potential. A series of novel LA derivatives with various substituents on the 20-N position was designed and synthesized. In the initial screening of LA derivatives against NO production, all the target compounds, except compound E2, exhibited excellent inhibitory ability relative to that of LA. Particularly, compound A4 exhibited the most potent inhibition with IC₅₀ of 12.91 μmol/L. The elementary structure–activity relationships (SARs) of NO inhibitory activity indicated that replacement of the benzene ring with an electron donating group could improve the anti-inflammatory efficacy. Furthermore, compound A4 shows an anti-inflammatory mechanism by inhibiting NO, PGE2, and TNF-α generation via the suppression of NF-κB and MAPK signaling pathways. Notably, compound A4 could exert a significant therapeutic effect on LPS-induced acute lung injury (ALI) in vivo. Based on the above research, we further investigated the preliminary pharmacokinetic property of A4 in rats. Therefore, compound A4 could be a promising candidate for the development of anti-inflammatory agents in the future.     

Zingiber cassumunar blended patches for skin application: Formulation,physicochemical properties,and in vitro studies

Our work was to study the preparation, physicochemical characterization, and in vitro characteristic of Zingiber cassumunar blended patches. The Z. cassumunar blended patches incorporating Z. cassumunar Roxb. also known as Plai were prepared from chitosan and polyvinyl alcohol with glycerin as plasticizer. They were prepared by adding all ingredients in a beaker and homogeneously mixing them. Then, they were transferred into Petri-dish and dried in hot air oven. The hydrophilic nature of the Z. cassumunar blended patches was confirmed by the moisture uptake, swelling ratio, erosion, and porosity values. The FTIR, DSC, XRD, and SEM studies showed revealed blended patches with amorphous region that was homogeneously smooth and compact in both surface and cross section dimensions. They exhibited controlled the release behavior of (E)-4-(3′,4′-dimethoxyphenyl) but-3-en-l-ol (compound D) that is the main active compound in Z. cassumunar for anti-inflammation activity. However, in in vitro skin permeation study, the compound D was accumulated in newborn pig skin more than in the receptor medium. Thus, the blended patches showed the suitable entrapment and controlled release of compound D. Accordingly, we have demonstrated that such chitosan and polyvinyl alcohol formulated patches might be developed for medical use.     

Synthesis,in vitro evaluation and molecular docking studies of novel coumarin‐isatin derivatives as α‐glucosidase inhibitors

This study synthesized a series of novel coumarin‐isatin derivatives and evaluated them for α‐glucosidase inhibitory activity. The majority of the screened compounds exhibited excellent inhibition activities with IC₅₀ values of 2.56 ± 0.08–268.79 ± 3.04 μm , when compared to acarbose. Among the newly derivatives, compound 5p was found to be the most active compound in the library of coumarin‐isatin derivatives. Furthermore, enzyme kinetic studies showed that compound 5p is a non‐competitive inhibitor with a K_i of 2.14 μm . Molecular docking analysis revealed the existence of hydrophobic and hydrogen interactions between compound 5p and the active site of α‐glucosidase. Our results indicate that coumarin‐isatin derivatives as a new class of α‐glucosidase inhibitors.     

Click chemistry‐based synthesis and cytotoxic activity evaluation of 4α‐triazole acetate podophyllotoxin derivatives

A series of novel 4α‐triazole acetate podophyllotoxin derivatives were synthesized via click chemistry. In vitro cytotoxic activity evaluation showed that most of the derivatives exhibited potent inhibitory activities against the tested cancer cell lines with low nanomolar IC₅₀ values. Further studies demonstrated that compound 31 exhibited broad‐spectrum cytotoxic activities, effectively overcame drug‐resistance, and showed relatively weak cytotoxicity on non‐cancer cells. Preliminary mechanistic studies indicated that 31 might have action on microtubule, cause cell cycle arrest at G₂/M phase, and induce apoptosis in human PC‐3 cancer cells.     

Structure‐based Design,Synthesis, and Biological Evaluation of Isatin Derivatives as Potential Glycosyltransferase Inhibitors

Peptidoglycan glycosyltransferase (PGT) has been shown to be an important pharmacological target for the inhibition of bacterial cell wall biosynthesis. Structure‐based virtual screening of about 3 000 000 commercially available compounds against the crystal structure of the glycosyltransferase (GT) domain of the Staphylococcus aureus penicillin‐binding protein 2 (S. aureus PBP2) resulted in identification of an isatin derivative, 2‐(3‐(2‐carbamimidoylhydrazono)‐2‐oxoindolin‐1‐yl)‐N‐(m‐tolyl)acetamide ( 4 ) as a novel potential GT inhibitor. A series of 4 derivatives were synthesized. Several compounds showed more active antimicrobial activity than the initial hit compound 4 , in particular 2‐(3‐(2‐carbamimidoylhydrazono)‐2‐oxoindolin‐1‐yl)‐N‐(3‐nitrophenyl)acetamide ( 4l ), against Gram‐positive Bacillus subtilis and S. aureus with MIC values of 24 and 48 μg/mL, respectively. Saturation transfer difference (STD) NMR study revealed that there is a binding contact between 4l and the GT domain of S. aureus PBP2. Competitive STD‐NMR further proved that 4l and moenomycin A bind to GT domain in a competitive manner. Molecular docking study suggests a potential binding pocket of 4l in the GT domain of S. aureus PBP2. Taken together, compound 4l would provide a new scaffold for further development of potent GT inhibitors.     

Tubulin Inhibitor Identification by Bioactive Conformation Alignment Pharmacophore‐Guided Virtual Screening

Microtubules are important cellular component that are critical for proper cellular function. Microtubules are synthesized by polymerization of αβ tubulin heterodimers called protofilaments. Microtubule dynamics facilitate proper cell division during mitosis. Disruption of microtubule dynamics by small‐molecule agents inhibits mitosis, resulting in apoptotic cell death and preventing cell cycle progression. To identify a novel small molecule that binds the αβ tubulin interface to affect microtubule dynamics, we developed a bioactive conformation alignment pharmacophore (BCAP) model to screen tubulin inhibitors from a huge database. The application of BCAP model generated based on the known αβ‐tubulin interface binders enabled us to identify several small‐molecules that cause apoptosis in human promyelocytic leukemia (HL‐60) cells. Virtual screening combined with an in vitro assay yielded 15 cytotoxic molecules. In particular, ethyl 2‐(4‐(5‐methyl‐3‐nitro‐1H‐pyrazol‐1‐yl)butanamido)‐4‐phenylthiophene‐3‐carboxylate ( H05 ) inhibited tubulin polymerization with an IC₅₀ of 17.6 μm concentration. The virtual screening results suggest that the application of an unbiased BCAP pharmacophore greatly eliminates unlikely compounds from a huge database and maximizes screening success. From the limited compounds tested in the tubulin polymerization inhibitor (TPI) assay, compound H05 was discovered as a tubulin inhibitor. This compound requires further structure activity optimization to identify additional potent inhibitors from the same class of molecules.     

Design and discovery of novel pyrazole-pyrrolopyrimidine derivatives as anti-glioma agents via promoting apoptosis,inhibiting cell cycle and EGFR-TK

Glioma is an aggressive type of brain malignancy responsible for significant morbidity and mortality. In the current scenario, epidermal growth factor receptor (EGFR) kinases targeted therapy showed significant benefits in glioma patients. Therefore, in the present study, we intend to investigate the anti-glioma potential of a novel class of pyrazole-pyrrolopyrimidine derivatives and their mechanism of action. The compounds will be synthesized in a straight-forward synthetic route in excellent yields and subsequently tested for EGFR kinase inhibition. The compounds showed a diverse range of inhibitory activity against EGFR (IC₅₀ = 3.4–873.2 nM). With an IC₅₀ of 1.5 nM, compound 4i was determined to be the most effective EGFR inhibitor, even superior to the standard erlotinib (IC₅₀ 2.3 nM). Among them, the three most potent compounds ( 4i , 4j , and 4k ) were further subjected to the anticancer activity against the panel of various cancer cell lines MCF-7 (breast cancer), A549 (lung cancer), U87 (glioblastoma cell)-EGFR-Wild Type, U87 (mutant glioblastoma cells) EGFR-mutant cell, MCF-12A (normal cells). The compound 4i showed the most potent activity against glioblastoma cells as compared to other cancer cells. The effect of compound 4i was also studied on the apoptosis of U87 cells, where it showed induction of apoptosis in a concentration-dependent manner. It also showed inhibition of the G2/M cell cycle phase of U87 cells. Our study demonstrated the development of novel pyrazole-pyrrolopyrimidine derivatives as a novel class of anti-glioma agents.     

Synthesis,in vitro and in silico anti-proliferative activity of 4-aryl-4<Emphasis Type="Italic">H</Emphasis>-chromene derivatives

A new series of C4-N,N-dialkylaniline-substituted 4-aryl-4H-chromenes were synthesized, and their anti-proliferative properties were evaluated against human cancer cell lines, namely, laryngeal carcinoma (Hep2), lung adenocarcinoma (A549), and cervical cancer (HeLa). The best among them, the 4-aryl-4H-chromene with C4-1-phenylpiperidine substitution was selected for further structure activity relationship (SAR) studies. Among the derivatives, N,6-dimethyl-3-nitro-4-(4-(piperidine-1-yl)phenyl)-4H-chromene-2-amine 3k showed most potent cytotoxic activity against all three cancer cell lines. Toxicity studies revealed that the 4-aryl-4H-chromenes specifically target the cancer cell lines. Molecular docking studies of this compound revealed its efficient interaction with the active site of αβ-tubulin protein.     

Design,Synthesis and in vivo Evaluation of Novel C‐Aryl Glucosides as Potent Sodium‐Dependent Glucose Cotransporters Inhibitors for the Treatment of Diabetes

A series of novel C‐aryl glucosides with substituents at the 3′‐position or cyclization at 3′, 4′‐positions of the distal aryl ring were designed and synthesized, which might decrease the oxidative metabolism of dapagliflozin. Preliminary evaluation for hypoglycemic effect and the risk of hypoglycemia were carried out both in normal and in streptozotocin‐induced diabetic mice. Among the synthesized compounds, compound 19a exerted potency‐similarity with dapagliflozin and triggered the hypoglycemic effect in a dose‐dependent manner. Besides, compound 19a , even at the high dose of 10 mg/kg, revealed a low risk of hypoglycemia. In further studies, 19a exhibited sustained antihyperglycemic effect without particular side‐effects in 30‐day chronic diabetic mice studies. Moreover, histological changes in the pancreas of diabetic mice indicated 19a might protect pancreatic β‐cell from apoptosis by reducing the damage of glucotoxicity. All of these results demonstrated that compound 19a , with excellent in vivo pharmacological activity and safety profile, was considered to be a promising drug candidate for the treatment of diabetes mellitus.     

Synthesis,adenosine receptor binding and molecular modelling studies of novel thieno[2,3‐d]pyrimidine derivatives

A series of new molecules containing a thieno[2,3‐d]pyrimidine scaffold was synthesized and characterized by adopting an efficient synthetic scheme. The effect of a free or substituted amino group at 2‐position as well as an oxo‐group, imidazole or 1,2,4‐triazole ring at 4‐position of the scaffold on the affinity and selectivity towards adenosine receptors (ARs) was evaluated. Compounds 17–19 with a free amino group at 2‐position along with the presence of an imidazole/1,2,4‐triazole ring at 4‐position of the scaffold showed selective binding affinities for hA_2A AR, whereas carbamoylation of the amino group at 2‐position (in the presence of an oxo‐group at 4‐position of the scaffold) increased the affinity and selectivity of certain compounds ( 7–10 ) for hA₃ AR. Molecular dynamic simulation study of one of the most active compound 8 (K_i hA₁ > 30 μm , hA_2A = 0.65 μm , and hA₃ = 0.124 μm ) revealed the role of important amino acid residues for imparting good affinity towards hA₃ and hA_2A ARs. Molecular docking studies were carried out for other compounds using the crystal structure of hA_2A AR and a homology model of hA₃ AR to rationalize their structure–activity relationships. The molecular docking results were in agreement with the experimental binding affinity data of ARs.     

3‐Aminomethyl pyridine chalcone derivatives: Design,synthesis, DNA binding and cytotoxic studies

Herein we report design, synthesis, and anticancer activity of compounds 6a–h and 11a–j . Compounds 6a–f were designed based on 3‐aminomethyl pyridine attached to different acetamide derivatives and in compounds 6g–h it was attached to coumarin moiety. Coumarin containing compounds 6g–h showed very poor anticancer activity against both A549 (Lungs cancer cell line), and MCF‐7 (Breast cancer cell line) cell lines in MTT assay. Compounds 11a–j were designed as derivatives of 3‐aminomethyl pyridine and 4‐amino chalcones. A series of chalcone derivatives of 3‐aminomethyl pyridine 11a–j have been synthesized and screened for their in vitro anticancer activity and DNA binding affinity. Most of the compounds showed very good antimitotic activity against A549 cell line as compared to fluorouracil. Compounds 11g and 11i were selected for DNA‐binding studies as they showed excellent activity against cancer cell lines in MTT assay. CT‐DNA binding affinity of compounds 11g and 11i have been investigated by UV based DNA titration and fluorescence emission study against DNA‐EtBr complex. Interestingly, compound 11i has displayed excellent antiproliferative activity, with IC₅₀ 0.0067 ± 0.0002 μm , against MCF‐7 cell line. Compound 11i has been studied for its cytotoxicity using MTT, LDH, as well as EtBr/AO assay and was found to induce apoptosis in the cancerous cell line.     

Antrodia cinnamomea exerts an anti-hepatoma effect by targeting PI3K/AKT-mediated cell cycle progression in vitro and in vivo

Antrodia cinnamomea is extensively used as a traditional medicine to prevention and treatment of liver cancer. However, its comprehensive chemical fingerprint is uncertain, and the mechanisms, especially the potential therapeutic target for anti-hepatocellular carcinoma (HCC) are still unclear. Using UPLC?Q-TOF/MS, 139 chemical components were identified in A. cinnamomea dropping pills (ACDPs). Based on these chemical components, network pharmacology demonstrated that the targets of active components were significantly enriched in the pathways in cancer, which were closely related with cell proliferation regulation. Next, HCC data was downloaded from Gene Expression Omnibus database (GEO). The Cancer Genome Atlas (TCGA) and DisGeNET were analyzed by bioinformatics, and 79 biomarkers were obtained. Furtherly, nine targets of ACDP active components were revealed, and they were significantly enriched in PI3K/AKT and cell cycle signaling pathways. The affinity between these targets and their corresponding active ingredients was predicted by molecular docking. Finally, in vivo and in vitro experiments showed that ACDPs could reduce the activity of PI3K/AKT signaling pathway and downregulate the expression of cell cycle-related proteins, contributing to the decreased growth of liver cancer. Altogether, PI3K/AKT-cell cycle appears as the significant central node in anti-liver cancer of A. Cinnamomea.     

In vitro and in vivo characterization of erythrosin B and derivatives against Zika virus

Zika virus (ZIKV) causes significant human diseases without specific therapy. Previously we found erythrosin B, an FDA-approved food additive, inhibited viral NS2B?NS3 interactions, leading to inhibition of ZIKV infection in cell culture. In this study, we performed pharmacokinetic and in vivo studies to demonstrate the efficacy of erythrosin B against ZIKV in 3D mini-brain organoid and mouse models. Our results showed that erythrosin B is very effective in abolishing ZIKV replication in the 3D organoid model. Although pharmacokinetics studies indicated that erythrosin B had a low absorption profile, mice challenged by a lethal dose of ZIKV showed a significantly improved survival rate upon oral administration of erythrosin B, compared to vehicle control. Limited structure?activity relationship studies indicated that most analogs of erythrosin B with modifications on the xanthene ring led to loss or reduction of inhibitory activities towards viral NS2B?NS3 interactions, protease activity and antiviral efficacy. In contrast, introducing chlorine substitutions on the isobenzofuran ring led to slightly increased activities, suggesting that the isobenzofuran ring is well tolerated for modifications. Cytotoxicity studies indicated that all derivatives are nontoxic to human cells. Overall, our studies demonstrated erythrosin B is an effective antiviral against ZIKV both in vitro and in vivo.     

Design,synthesis, and biological evaluation of novel 4‐phenoxypyridine derivatives as potential antitumor agents

A series of novel 4‐phenoxypyridine derivatives containing the 4‐oxo‐1,4‐dihydropyridazine‐3‐carboxamide moiety were synthesized and evaluated for their in vitro cytotoxic activity against the A549 cancer cell line, and some compounds were further examined for their cytotoxic activity against the H460, BGC823, MKN45, and HT‐29 cancer cell lines. Most of the compounds exhibited moderate to significant cytotoxicity. The most promising compound 15b (with VEGFR2 inhibitory concentration [IC₅₀] value of 0.23 μM) showed remarkable cytotoxicity against A549, BGC‐823, MKN45, H460, and HT‐29 cells, with IC₅₀ values of 0.75, 1.68, 2.63, 5.08 and 7.22 μM, respectively. Their preliminary structure‐activity relationship studies indicate that electron‐withdrawing groups on the terminal phenyl rings are beneficial for improving the antitumor activity. Moreover, treatment of A549 cells with compound 15b resulted in cell cycle arrest in the G₀/G₁ phase in a dose‐dependent manner. Further apoptotic studies and acridine orange/ethidium bromide staining were also performed on A549 cells, which showed that compound 15b could induce apoptosis. Wound‐healing assay results indicated that compound 15b strongly inhibited A549 cell motility.     

Design,Synthesis, Biological Screening,and Molecular Docking Studies of Piperazine-Derived Constrained Inhibitors of DPP-IV for the Treatment of Type 2 Diabetes

Novel piperazine-derived conformationally constrained compounds were designed, synthesized, and evaluated for in vitro Dipeptidyl peptidase-IV (DPP-IV) inhibitory activities. From a library of compounds synthesized, 1-(2-(4-(7-Chloro-4-quinolyl)piperazin-1-yl)acetyl)pyrrolidine ( 2g ) was identified as a potential DPP-IV inhibitor exhibiting better inhibitory activity than P32/98, reference inhibitor. The in vivo studies carried out in STZ and db/db mice models indicated that the compound 2g showed moderate antihyperglycemic activity as compared to the marketed drug Sitagliptin. A two-week repeated dose study in db/db mice revealed that compound 2g significantly declined blood glucose levels with no evidence of hypoglycemia risk. Furthermore, it showed improvement in insulin resistance reversal and antidyslipidemic properties. Molecular docking studies established good binding affinity of compound 2g at the DPP-IV active site and are in favor of the observed biological data. These data collectively suggest that compound 2g is a good lead molecule for further optimization studies.     

Neuroprotective effects of a novel poly (ADP‐ribose) polymerase‐1 inhibitor,JPI‐289, in hypoxic rat cortical neurons

Excessive activation of poly (ADP‐ribose) polymerase‐1 (PARP‐1) is known to develop neuronal apoptosis, necrosis and inflammation after ischaemic brain injury. Therefore, PARP‐1 inhibition after ischaemic stroke has been attempted in successful animal studies. The purpose of present work was to develop a novel water soluble PARP‐1 inhibitor (JPI‐289) and explore its neuroprotective effect on ischaemic injury in an in vitro model. The half‐life of JPI‐289 after intravenous or oral administration in rats was relatively long (1.4‐1.5 hours) with 65.6% bioavailability. The inhibitor strongly inhibited PARP‐1 activity (IC₅₀=18.5 nmol/L) and cellular PAR formation (IC₅₀=10.7 nmol/L) in the nanomolar range. In rat cortical neuronal cells, JPI‐289 did not affect cell viability up to 1 mmol/L as assayed by Trypan blue staining (TBS) and lactate dehydrogenase (LDH) assay. Treatment of JPI‐289 for 2 hours after 2 hours of oxygen glucose deprived (OGD) rat cortical neuron attenuated PARP activity and restored ATP and NAD+ levels. Apoptosis‐associated molecules such as apoptosis inducing factor (AIF), cytochrome C and cleaved caspase‐3 were reduced after JPI‐289 treatment in the OGD model. The present findings suggest that the novel PARP‐1 inhibitor, JPI‐289, is a potential neuroprotective agent which could be useful as a treatment for acute ischaemic stroke.     

Synthesis and biological evaluation of novel benzo[c]acridine‐diones as potential anticancer agents and tubulin polymerization inhibitors

A new series of novel benzo[c]acridine‐diones possessing pharmacophoric elements of antitubulins with central dihydropyridine bridge were designed and synthesized as potential anticancer agents and tubulin polymerization inhibitors. The cytotoxic activity of the synthesized compounds was evaluated against eight cancer cell lines including MCF‐7, A2780, HeLa, HepG2, DU145, A549, PC3, and LNCAP cancer cells and normal cells human umbilical vein endothelial cell (HUVEC) through 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5‐diphenyl tetrazolium bromide (MTT) assay, wherein β‐lapachone and combretastatin A‐4 were used as positive controls. Some of our compounds ( 4c and 4g ) showed significant cytotoxic activity on cancer cells with IC₅₀ values in the range of 5.23–24.32 μM. None of the synthesized compounds showed significant cytotoxicity on normal HUVEC cells. Among all investigated derivatives, compound 4g showed promising greater antiproliferative activity against all tested cancer cells with the highest sensitivity observed for the PC3 cell line. Results from the flow cytometry analysis of PC3 and MCF‐7 cancer cells treated with 4g showed an induced cell‐cycle arrest at G2/M, and therefore induced apoptosis which occurred at low concentration of test compound, whereas annexin V‐FITC/propidium iodide staining assay in the aforementioned cancer cell lines treated with 4g showed that 4g can cause necrosis in PC3 and MCF‐7 cancer cells at higher concentration. Compound 4g proved to be an inhibitor of tubulin polymerization in a mode similar to that of colchicine and in a dose‐dependent manner. Molecular docking studies of 4g into the colchicine‐binding site of tubulin exhibited a possible mode of interaction between this compound and tubulin.