Molecular,Immunological, and Biological Characterization of Tityus serrulatus Venom Hyaluronidase: New Insights into Its Role in Envenomation

Background

Scorpionism is a public health problem in Brazil, and Tityus serrulatus (Ts) is primarily responsible for severe accidents. The main toxic components of Ts venom are low-molecular-weight neurotoxins; however, the venom also contains poorly characterized high-molecular-weight enzymes. Hyaluronidase is one such enzyme that has been poorly characterized.

Methods and principal findings

We examined clones from a cDNA library of the Ts venom gland and described two novel isoforms of hyaluronidase, TsHyal-1 and TsHyal-2. The isoforms are 83% identical, and alignment of their predicted amino acid sequences with other hyaluronidases showed conserved residues between evolutionarily distant organisms. We performed gel filtration followed by reversed-phase chromatography to purify native hyaluronidase from Ts venom. Purified native Ts hyaluronidase was used to produce anti-hyaluronidase serum in rabbits. As little as 0.94 µl of anti-hyaluronidase serum neutralized 1 LD₅₀ (13.2 µg) of Ts venom hyaluronidase activity in vitro. In vivo neutralization assays showed that 121.6 µl of anti-hyaluronidase serum inhibited mouse death 100%, whereas 60.8 µl and 15.2 µl of serum delayed mouse death. Inhibition of death was also achieved by using the hyaluronidase pharmacological inhibitor aristolochic acid. Addition of native Ts hyaluronidase (0.418 µg) to pre-neutralized Ts venom (13.2 µg venom+0.94 µl anti-hyaluronidase serum) reversed mouse survival. We used the SPOT method to map TsHyal-1 and TsHyal-2 epitopes. More peptides were recognized by anti-hyaluronidase serum in TsHyal-1 than in TsHyal-2. Epitopes common to both isoforms included active site residues.

Conclusions

Hyaluronidase inhibition and immunoneutralization reduced the toxic effects of Ts venom. Our results have implications in scorpionism therapy and challenge the notion that only neurotoxins are important to the envenoming process.     

Indole and benzimidazole derivatives as steroid 5α-reductase inhibitors in the rat prostate

A novel series of indole and benzimidazole derivatives were synthesized and evaluated for their inhibitory activity of rat prostatic 5α-reductase. Among these compounds, 4-{2-[1-(4,4′-dipropylbenzhydryl)indole-5-carboxamido]phenoxy}butyric acid (15) and its benzimidazole analogue 25 showed potent inhibitory activities for rat prostatic 5α-reductase (IC₅₀ values of 9.6 ± 1.0 and 13 ± 1.5nM, respectively), with the potency very close to that of finasteride. Compound 30, in which the moiety between the benzene ring and amide bond was replaced by quinolin-4-one ring, showed almost equipotent activity (IC₅₀ = 19 ± 6.2nM) with the correspondent amide derivative 13. This result was consistent with the previous observation that the coplanarity of this moiety might contribute to the potent inhibitory activity.     

The phytotoxicity of selected mycotoxins on mature,germinatingZea mays embryos

Mature maize (Zea mays) embryos were exposed to 5, 10 and 25 µg ml^–1 of deoxynivalenol (DON), zearalenone (ZEA), ochratoxin A (OA) and a mixture of zearalenone and deoxynivalenol (ZEA/DON) for 9 days. DON and the ZEA/DON combination were consistently more inhibitory of the measured parameters than either ZEA or OA. Based on the predicted additive values, it would appear that, in combination, ZEA and DON act synergistically to inhibit root and shoot growth. For ZEA alone, a concentration of 5 µg ml^–1 ZEA was generally inhibitory of root and shoot elongation and fresh mass accumulation, while at 10 and 25 µg ml^–1, this toxin had a stimulatory effect on these parameters. For OA, the measured effects on root and shoot growth at 5 and 25 µg ml^–1 were stimulatory, while at 10 µg ml^–1 OA, an inhibitory effect was observed. For all toxins, inhibitory/stimulatory effects were generally more marked for root parameters than for shoot elongation or mass.Abbreviations ADON acetyldeoxynivalenol - AFB₁ aflatoxin B₁ - DAS diacetyoxyscirpenol - DON deoxynivalenol - FB₁ fumonisin B₁ - FHB Fusaium head blight - MON moniliformin - NIV nivalenol - OA ochratoxin A - ZEA zearalenone     

Honey shows potent inhibitory activity against the bovine testes hyaluronidase

The purpose of this study was to investigate the anti-hyaluronidase activities of honeys from different botanical origins honeys in order to determine their anti-inflammatory properties. The total phenolic contents, total flavonoids and total tannin levels of six types of honey, chestnut, oak, heather, pine, buckwheat and mixed blossom, were determined. Concentration-related inhibition values were tested turbidimetrically on bovine testis hyaluronidase (BTHase) as IC₅₀ (mg/mL). All honeys exhibited various concentration-dependent degrees of inhibition against BTHase. Inhibition values varied significantly depending on honeys’ levels of phenolic contents, flavonoid and tannin. The honeys with the highest anti-hyaluronidase activity were oak, chestnut and heather. In conclusion, polyphenol-rich honeys have high anti-hyaluronidase activity, and these honeys have high protective and complementary potential against hyaluronidase-induced anti-inflammatory failures.     

New 3-O-substituted xanthone derivatives as promising acetylcholinesterase inhibitors

A new series of 3-O-substituted xanthone derivatives were synthesised and evaluated for their anti-cholinergic activities against acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). The results indicated that the xanthone derivatives possessed good AChE inhibitory activity with eleven of them (5, 8, 11, 17, 19, 21-23, 26-28) exhibited significant effects with the IC₅₀ values ranged 0.88 to 1.28 µM. The AChE enzyme kinetic study of 3-(4-phenylbutoxy)-9H-xanthen-9-one (23) and ethyl 2-((9-oxo-9H-xanthen-3-yl)oxy)acetate (28) showed a mixed inhibition mechanism. Molecular docking study showed that 23 binds to the active site of AChE and interacts via extensive π–π stacking with the indole and phenol side chains of Trp86 and Tyr337, besides the hydrogen bonding with the hydration site and π–π interaction with the phenol side chain of Y72. This study revealed that 3-O-alkoxyl substituted xanthone derivatives are potential lead structures, especially 23 and 28 which can be further developed into potent AChE inhibitors.     

In-vitro evaluation of antioxidant,anti-elastase,anti-collagenase,anti-hyaluronidase activities of safranal and determination of its sun protection factor in skin photoaging

Safranal, a monoterpene aldehyde, is present as one of the main volatile constituents of Crocus sativus Linn. (saffron flowers). This volatile constituent not only contributes to the aroma of saffron but has been reported to possess antidiabetic, antiulcer, antiasthamatic, anticonvulsant, antidepressant, cardioprotective, anticancer and UV protective properties. Most of these therapeutic actions are contributed by its potential to quench reactive oxygen species (ROS). Antioxidant properties of phytoconstituents are now being explored for developing photoprotective skin formulations. These bioactives have the potential to protect the epidermal and dermal layers of the skin which mainly comprises of elastin and collagen. When UV rays penetrate the dermal layers, there is an increased production of elastase, collagenase and hyaluronidase leading to degradation of collagen, elastin and hyaluronic acid respectively. These dermal components are responsible to provide strength, elasticity and moisture to the skin. Due to frequent exposure to sunlight, these conditions tend to augment leading to wrinkle formation and sagging of skin.Although antioxidant properties of safranal have been established on various cell lines but till date no studies have been reported regarding the dermal enzyme inhibition activities. In the current research work, a comprehensive in vitro evaluation of antioxidant, anti-elastase, anti-collagenase, anti-hyaluronidase activities of safranal along with determination of sun protection factor (SPF) was carried out. The in vitro antioxidant activity was carried out by diphenylpicrylhydrazyl (DPPH) method and its IC₅₀ value was found to be 22.7 μg/ml. The enzyme inhibition IC₅₀ values of safranal for anti elastase activity were found to be 43.6 μg/ml, 70 μg/ml for antihyaluronidase activity and 9.4 μg/ml for anticollagenase activity. Photoprotective activity of safranal was determined by UV absorbance method and SPF calculated by Mansur equation which was found to be 6.6.The significant inhibitory activity of safranal on matrix metalloproteinases (MMPs) responsible for aging and a higher SPF established that this bioorganic molecule is a strong photoprotective agent. Its established free radical scavenging capability along with above characteristics make it a valuable component to be incorporated into herbal antiaging formulations.     

Design and synthesis of the novel oleanolic acid-cinnamic acid ester derivatives and glycyrrhetinic acid-cinnamic acid ester derivatives with cytotoxic properties

Oleanolic acid (OA) and glycyrrhetinic acid (GA) are natural products with anticancer effects. Cinnamic acid (CA) and its derivatives also exhibited certain anticancer activity. In order to improve the anticancer activity of OA and GA, we designed and synthesized a series of novel OA-CA ester derivatives and GA-CA ester derivatives by using molecular hybridization approach. The 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay was used to assess their in vitro cytotoxicity on three cell lines (HeLa (cervical cancer), MCF-7 (breast cancer) and L-O2 (a normal hepatic cell)). Among the evaluated compounds, 3o presented the strongest selective cytotoxicity on HeLa cells (IC₅₀ = 1.35 μM) and showed no inhibitory activity against MCF-7 cells (IC₅₀ > 100 μM) and L-O2 cells (IC₅₀ > 100 μM), and 3e presented the strongest selective inhibition of the MCF-7 cells (IC₅₀ = 1.79 μM). What’s more, compound 2d also showed very strong selective inhibitory activity against HeLa cells (IC₅₀ = 1.55 μM). The further research using Hoechst 33342, AO/EB dual-staining, flow cytometric analysis and DCFH-DA fluorescent dye staining assay presented that 2d and 3o could induce HeLa cells apoptosis and autophagy.     

Quinazolin-4(3H)-one based potential multiple tyrosine kinase inhibitors with excellent cytotoxicity

A series of quinazolin-4(3H)-one derivatives were synthesised and evaluated for their cytotoxicity against human Caucasian breast adenocarcinoma (MCF-7) and human ovarian carcinoma (A2780) cell lines. Cytotoxicity of the most tested compounds was 2- to 30-fold more than the positive control lapatinib (IC₅₀ of 2j = 3.79 ± 0.96; 3j = 0.20 ± 0.02; and lapatinib = 5.9 ± 0.74) against MCF7 cell lines except two compounds (IC₅₀ of 2 b = 15.72 ± 0.07 and 2e = 14.88 ± 0.99). On the other hand, cytotoxicity was 4 − 87 folds (IC₅₀ of 3a = 3.00 ± 1.20; 3 g = 0.14 ± 0.03) more the positive control lapatinib (IC₅₀ = 12.11 ± 1.03) against A2780 cell lines except compound 2e (IC₅₀ = 16.43 ± 1.80). Among the synthesised quinazolin-4(3H)-one derivatives, potent cytotoxic 2f-j and 3f-j were investigated for molecular mechanism of action. Inhibitory activities of the compounds were tested against multiple tyrosine protein kinases (CDK2, HER2, EGFR and VEGFR2) enzymes. As expected, all the quinazolin-4(3H)-one derivatives were showed comparable inhibitory activity against those kinases tested, especially, compound 2i and 3i showed potent inhibitory activity against CDK2, HER2, EGFR tyrosine kinases. Therefore, molecular docking analysis for quinazolin-4(3H)-one derivatives 2i and 3i were performed, and it was revealed that compounds 2i and 3i act as ATP non-competitive type-II inhibitor against CDK2 kinase enzymes and ATP competitive type-I inhibitor against EGFR kinase enzymes. However, in case of HER2, compounds 2i act as ATP non-competitive type-II inhibitor and 3i act as ATP competitive type-I inhibitor. Docking results of known inhibitors were compared with synthesised compounds and found synthesised 2i and 3i are superior than the known inhibitors in case of interactions. In addition, in silico drug likeness properties of quinazolin-4(3H)-one derivatives showed better predicted ADME values than lapatinib.     

Vascular Endothelium-Dependent and Independent Actions of Oleanolic Acid and Its Synthetic Oleanane Derivatives as Possible Mechanisms for Hypotensive Effects

Purpose

Plant-derived oleanolic acid (OA) and its related synthetic derivatives (Br-OA and Me-OA) possess antihypertensive effects in experimental animals. The present study investigated possible underlying mechanisms in rat isolated single ventricular myocytes and in vascular smooth muscles superfused at 37°C.

Methods

Cell shortening was assessed at 1 Hz using a video-based edge-detection system and the L-type Ca²⁺ current (I_CaL) was measured using the whole-cell patch-clamp technique in single ventricular myocytes. Isometric tension was measured using force transducer in isolated aortic rings and in mesenteric arteries. Vascular effects were measured in endothelium-intact and denuded vessels in the presence of various enzyme or channel inhibitors.

Results

OA and its derivatives increased cell shortening in cardiomyocytes isolated from normotensive rats but had no effect in those isolated from hypertensive animals. These triterpenes also caused relaxation in aortic rings and in mesenteric arteries pre-contracted with either phenylephrine or KCl-enriched solution. The relaxation was only partially inhibited by endothelium denudation, and also partly inhibited by the cyclooxygenase (COX) inhibitor indomethacin, with no additional inhibitory effect of the NO synthase inhibitor, N-ω-Nitro-L-arginine. A combination of both ATP-dependent channel inhibition by glibenclaminde and voltage-dependent K⁺ channel inhibition by 4-aminopyridine was necessary to fully inhibit the relaxation.

Conclusion

These data indicate that the effects of OA and its derivatives are mediated via both endothelium-dependent and independent mechanisms suggesting the involvement of COX in the endothelium-dependent effects and of vascular muscle K⁺ channels in the endothelium-independent effects. Finally, our results support the view that the antihypertensive action of OA and its derivatives is due to a decrease of vascular resistance with no negative inotropic effect on the heart.     

Design and biological evaluation of substituted 5,7-dihydro-6H-indolo[2,3-c]quinolin-6-one as novel selective Haspin inhibitors

A library of substituted indolo[2,3-c]quinolone-6-ones was developed as simplified Lamellarin isosters. Synthesis was achieved from indole after a four-step pathway sequence involving iodination, a Suzuki-Miyaura cross-coupling reaction, and a reduction/lactamization sequence. The inhibitory activity of the 22 novel derivatives was assessed on Haspin kinase. Two of them possessed an IC₅₀ of 1 and 2 nM with selectivity towards a panel of 10 other kinases including the parent kinases DYRK1A and CLK1. The most selective compound exerted additionally a very interesting cell effect on the osteosarcoma U-2 OS cell line.     

Synthesis,potential antitumor activity,cell cycle analysis,and multitarget mechanisms of novel hydrazones incorporating a 4-methylsulfonylbenzene scaffold: a molecular docking study

Hydrazone is a bioactive pharmacophore that can be used to design antitumor agents. We synthesised a series of hydrazones (compounds 4–24) incorporating a 4-methylsulfonylbenzene scaffold and analysed their potential antitumor activity. Compounds 6, 9, 16, and 20 had the most antitumor activity with a positive cytotoxic effect (PCE) of 52/59, 27/59, 59/59, and 59/59, respectively, while compounds 5, 10, 14, 15, 18, and 19 had a moderate antitumor activity with a PCE of 11/59–14/59. Compound 20 was the most active and had a mean 50% cell growth inhibition (GI₅₀) of 0.26 µM. Compounds 9 and 20 showed the highest inhibitory activity against COX-2, with a half-maximal inhibitory concentration (IC₅₀) of 2.97 and 6.94 μM, respectively. Compounds 16 and 20 significantly inhibited EGFR (IC₅₀ = 0.2 and 0.19 μM, respectively) and HER2 (IC₅₀ = 0.13 and 0.07 μM, respectively). Molecular docking studies of derivatives 9, 16, and 20 into the binding sites of COX-2, EGFR, and HER2 were carried out to explore the interaction mode and the structural requirements for antitumor activity.     

A Novel Synthesized Sulfonamido-Based Gallate—JEZ-C as Potential Therapeutic Agents for Osteoarthritis

Gallic acid (GA) and its derivatives are anti-inflammatory agents reported to have an effect on osteoarthritis (OA). However, GA has much weaker anti-oxidant effects and inferior bioactivity compared with its derivatives. We modified GA with the introduction of sulfonamide to synthesize a novel compound named JEZ-C and analyzed its anti-arthritis and chondro-protective effects. Comparison of JEZ-C with its sources i.e. GA and Sulfamethoxazole (SMZ) was also performed. Results showed that JEZ-C could effectively inhibit the IL-1-mediated induction of MMP-1 and MMP-13 and could induce the expression of TIMP-1, which demonstrated its ability to reduce the progression of OA. JEZ-C can also exert chondro-protective effects by promoting cell proliferation and maintaining the phenotype of articular chondrocytes, as evidenced by improved cell growth, enhanced synthesis of cartilage specific markers such as aggrecan, collagen II and Sox9. Meanwhile, expression of the collagen I gene was effectively downregulated, revealing the inhibition of chondrocytes dedifferentiation by JEZ-C. Hypertrophy that may lead to chondrocyte ossification was also undetectable in JEZ-C groups. The recommended dose of JEZ-C ranges from 6.25×10^-7 μg/ml to 6.25×10^-5 μg/ml, among which the most profound response was observed with 6.25×10^-6 μg/ml. In contrast, its source products of GA and SMZ have a weak effect not only in the inhibition of OA but also in the bioactivity of chondrocytes, which indicated the significance of this modification. This study revealed JEZ-C as a promising novel agent in the treatment of chondral and osteochondral lesions.     

Synthesis and bioactivities evaluation of oleanolic acid oxime ester derivatives as α-glucosidase and α-amylase inhibitors

Different oleanolic acid (OA) oxime ester derivatives (3a-3t) were designed and synthesised to develop inhibitors against α-glucosidase and α-amylase. All the synthesised OA derivatives were evaluated against α-glucosidase and α-amylase in vitro. Among them, compound 3a showed the highest α-glucosidase inhibition with an IC₅₀ of 0.35 µM, which was ∼1900 times stronger than that of acarbose, meanwhile compound 3f exhibited the highest α-amylase inhibitory with an IC₅₀ of 3.80 µM that was ∼26 times higher than that of acarbose. The inhibition kinetic studies showed that the inhibitory mechanism of compounds 3a and 3f were reversible and mixed types towards α-glucosidase and α-amylase, respectively. Molecular docking studies analysed the interaction between compound and two enzymes, respectively. Furthermore, cytotoxicity evaluation assay demonstrated a high level of safety profile of compounds 3a and 3f against 3T3-L1 and HepG2 cells.

Highlights

1. Oleanolic acid oxime ester derivatives (3a–3t) were synthesised and screened against α-glucosidase and α-amylase.
2. Compound 3a showed the highest α-glucosidase inhibitory with IC50 of 0.35 µM.
3. Compound 3f presented the highest α-amylase inhibitory with IC50 of 3.80 µM.
4. Kinetic studies and in silico studies analysed the binding between compounds and α-glucosidase or α-amylase.

     

Novel chalcone/aryl carboximidamide hybrids as potent anti-inflammatory via inhibition of prostaglandin E2 and inducible NO synthase activities: design,synthesis, molecular docking studies and ADMET prediction

Two series of chalcone/aryl carboximidamide hybrids 4a–f and 6a–f were synthesised and evaluated for their inhibitory activity against iNOS and PGE2. The most potent derivatives were further checked for their in vivo anti-inflammatory activity utilising carrageenan-induced rat paw oedema model. Compounds 4c, 4d, 6c and 6d were proved to be the most effective inhibitors of PGE2, LPS-induced NO production, iNOS activity. Moreover, 4c, 4d, 6c and 6d showed significant oedema inhibition ranging from 62.21% to 78.51%, compared to indomethacin (56.27 ± 2.14%) and celecoxib (12.32%). Additionally, 4c, 6a and 6e displayed good COX2 inhibitory activity while 4c, 6a and 6c exhibited the highest 5LOX inhibitory activity. Compounds 4c, 4d, 6c and 6d fit nicely into the pocket of iNOS protein (PDB ID: 1r35) via the important amino acid residues. Prediction of physicochemical parameters exhibited that 4c, 4d, 6c and 6d had acceptable physicochemical parameters and drug-likeness. The results indicated that chalcone/aryl carboximidamides 4c, 4d, 6c and 6d, in particular 4d and 6d, could be used as promising lead candidates as potent anti-inflammatory agents.     

Effects of Pb<Superscript>2+</Superscript> ions on Na<Superscript>+</Superscript> transport in the isolated skin of the toad <Emphasis Type="Italic">Pleurodema thaul</Emphasis>

The effects induced by lead ions on the short-circuit current (SCC) and on the potential difference (V) of the toad Pleurodema thaul skin were investigated. Pb²⁺ applied to the outer (mucosal) surface increased SCC and V and when applied to the inner (serosal) surface decreased both parameters. The stimulatory effect, but not the inhibitory action, was reversible after washout of the metal ion. The amiloride test showed that the increase was due principally to stimulation of the driving potential for Na⁺ (V-E_Na+) and that inhibition was accompanied by a reduction in the V-E_Na+ and also by a significant decrease in skin resistance indicating possible disruption of membrane and/or cell integrity. The effect of noradrenaline was increased by outer and decreased by inner administration of Pb²⁺. The results suggest that mucosal Pb²⁺ activates toad skin ion transport by stimulating the V-E_Na+ and that serosal Pb²⁺, with easier access to membrane and cellular constituents, inactivates this mechanism, revealing greater toxicity when applied to the inner surface of the skin. Abbreviations: SCC – short-circuit current; V – potential difference; V-E_Na+– driving potential for Na⁺; ENaC – epithelial sodium channel; R_Na+– active sodium resistance; RS – passive or shunt resistance; G_Na– active sodium conductance; GS – passive or shunt conductance; Gmax – total conductance; EC₅₀– half-maximal excitatory concentration; IC₅₀– half maximal inhibitory concentration; NA – noradrenaline.     

Discovery of new 3-methylquinoxalines as potential anti-cancer agents and apoptosis inducers targeting VEGFR-2: design,synthesis, and in silico studies

There is an urgent need to design new anticancer agents that can prevent cancer cell proliferation even with minimal side effects. Accordingly, two new series of 3-methylquinoxalin-2(1H)-one and 3-methylquinoxaline-2-thiol derivatives were designed to act as VEGFR-2 inhibitors. The designed derivatives were synthesised and evaluated in vitro as cytotoxic agents against two human cancer cell lines namely, HepG-2 and MCF-7. Also, the synthesised derivatives were assessed for their VEGFR-2inhibitory effect. The most promising member 11e were further investigated to reach a valuable insight about its apoptotic effect through cell cycle and apoptosis analyses. Moreover, deep investigations were carried out for compound 11e using western-plot analyses to detect its effect against some apoptotic and apoptotic parameters including caspase-9, caspase-3, BAX, and Bcl-2. Many in silico investigations including docking, ADMET, toxicity studies were performed to predict binding affinity, pharmacokinetic, drug likeness, and toxicity of the synthesised compounds. The results revealed that compounds 11e, 11g, 12e, 12g, and 12k exhibited promising cytotoxic activities (IC₅₀ range is 2.1 − 9.8 µM), comparing to sorafenib (IC₅₀ = 3.4 and 2.2 µM against MCF-7 and HepG2, respectively). Moreover, 11b, 11f, 11g, 12e, 12f, 12g, and 12k showed the highest VEGFR-2 inhibitory activities (IC₅₀ range is 2.9 − 5.4 µM), comparing to sorafenib (IC₅₀ = 3.07 nM). Additionally, compound 11e had good potential to arrest the HepG2 cell growth at G2/M phase and to induce apoptosis by 49.14% compared to the control cells (9.71%). As well, such compound showed a significant increase in the level of caspase-3 (2.34-fold), caspase-9 (2.34-fold), and BAX (3.14-fold), and a significant decrease in Bcl-2 level (3.13-fold). For in silico studies, the synthesised compounds showed binding mode similar to that of the reference compound (sorafenib).     

4-Methoxycinnamaldehyde inhibited human respiratory syncytial virus in a human larynx carcinoma cell line

4-Methoxycinnamaldehyde, an active constituent of Agastache rugosa, was examined for its cytoprotective activity against RSV by XTT method in human larynx carcinoma cell line. 4-Methoxycinnamaldehyde could effectively inhibit cytopathic effect of RSV (p<0.0001) with an estimated IC₅₀ of 0.055 μg/ml and a selectivity index (SI) of 898.2. 4-Methoxycinnamaldehyde (0.03 μg/ml) could inhibit viral entrance by interfering viral attachment (IC₅₀ of 0.06 μg/ml; p<0.0001) and internalization (IC₅₀ of 0.01 μg/ml; p<0.0001). 4-Methoxycinnamaldehyde significantly increased the basal production of IFN (p=0.0015), but not the virus-induced IFN production. Therefore, its cytoprotective activity against RSV was not mediated by interferon. In conclusion, 4-methoxycinnamaldehyde might be helpful to manage the disease induced by RSV infection.     

Development of 3-methyl/3-(morpholinomethyl)benzofuran derivatives as novel antitumor agents towards non-small cell lung cancer cells

As one of the most lethal malignancies, lung cancer is considered to account for approximately one-fifth of all malignant tumours-related deaths worldwide. This study reports the synthesis and in vitro biological assessment of two sets of 3-methylbenzofurans (4a–d, 6a–c, 8a–c and 11) and 3-(morpholinomethyl)benzofurans (15a–c, 16a–b, 17a–b and 18) as potential anticancer agents towards non-small cell lung carcinoma A549 and NCI-H23 cell lines, with VEGFR-2 inhibitory activity. The target benzofuran-based derivatives efficiently inhibited the growth of both A549 and NCI-H23 cell lines with IC₅₀ spanning in ranges 1.48–47.02 and 0.49–68.9 µM, respectively. The three most active benzofurans (4b, 15a and 16a) were further investigated for their effects on the cell cycle progression and apoptosis in A549 (for 4b) and NCI-H23 (for 15a and 16a) cell lines. Furthermore, benzofurans 4b, 15a and 16a displayed good VEGFR-2 inhibitory activity with IC₅₀ equal 77.97, 132.5 and 45.4 nM, respectively.     

Ligand-based design,synthesis, computational insights,and in vitro studies of novel N-(5-Nitrothiazol-2-yl)-carboxamido derivatives as potent inhibitors of SARS-CoV-2 main protease

The global outbreak of the COVID-19 pandemic provokes scientists to make a prompt development of new effective therapeutic interventions for the battle against SARS-CoV-2. A new series of N-(5-nitrothiazol-2-yl)-carboxamido derivatives were designed and synthesised based on the structural optimisation principle of the SARS-CoV Mpro co-crystallized WR1 inhibitor. Notably, compound 3b achieved the most promising anti-SARS-CoV-2 activity with an IC₅₀ value of 174.7 µg/mL. On the other hand, compounds 3a, 3b, and 3c showed very promising SARS-CoV-2 Mpro inhibitory effects with IC₅₀ values of 4.67, 5.12, and 11.90 µg/mL, respectively. Compound 3b docking score was very promising (−6.94 kcal/mol) and its binding mode was nearly similar to that of WR1. Besides, the molecular dynamics (MD) simulations of compound 3b showed its great stability inside the binding pocket until around 40 ns. Finally, a very promising SAR was concluded to help to design more powerful SARS-CoV-2 Mpro inhibitors shortly.     

Indole carboxamides inhibit bovine testes hyaluronidase at pH 7.0 and indole acetamides activate the enzyme at pH 3.5 by different mechanisms

Hyaluronidases are enzymes controlling many crucial physiological processes. Imbalanced enzymatic activity is connected with severe diseases. Because there is limited availability of drugs modulating hyaluronidase activity, the search for hyaluronidase interacting compounds is getting more and more important. A series of fifteen indole carboxamides and acetamides were synthesized and tested on inhibition of bovine testes hyaluronidase. In vitro assays were performed using stains-all at pH 7 and the Morgan-Elson reaction at pH 3.5. At neutral pH, the most active inhibitory compound was N-(Pyridin-4yl)-[5-bromo-1-(4-fluorobenzyl)indole-3-yl]carboxamide (20) with an IC₅₀ value of 46 μM. Surprisingly, inhibition of all compounds was completely abolished by a decrease in pH. At pH 3.5 the activity of the enzyme was increased up to 134% by compound N-(4,6-Dimethylpyridin-2yl)-(1-ethylindole-3-yl)acetamide (24) at a concentration of 100 μM. The known activating effect of bovine serum albumine (BSA) on hyaluronidase activity was verified in the assay and compared to the effect of compound 24. Structure-activity relationships are discussed and a model is proposed, which explains the increase in activity at pH 3.5 by bonding of the protonated form of N-(4,6-Dimethylpyridin-2yl)-(1-ethylindole-3-yl)acetamide (24) to hyaluronic acid. The bonding results in an elongated form of the substrate with easier enzymatic access.