Cyclic lipopeptides as membrane fusion inhibitors against SARS-CoV-2: New tricks for old dogs.

With the resurgence of the coronavirus pandemic, the repositioning of FDA-approved drugs against coronovirus and finding alternative strategies for antiviral therapy are both important. We previously identified the viral lipid envelope as a potential target for the prevention and treatment of SARS-CoV-2 infection with plant alkaloids (Shekunov et al., 2021). Here, we investigated the effects of eleven cyclic lipopeptides (CLPs), including well-known antifungal and antibacterial compounds, on the liposome fusion triggered by calcium, polyethylene glycol 8000, and a fragment of SARS-CoV-2 fusion peptide (816-827) by calcein release assays. Differential scanning microcalorimetry of the gel-to-liquid-crystalline and lamellar-to-inverted hexagonal phase transitions and confocal fluorescence microscopy demonstrated the relation of the fusion inhibitory effects of CLPs to alterations in lipid packing, membrane curvature stress and domain organization. The antiviral effects of CLPs were evaluated in an in vitro Vero-based cell model, and aculeacin A, anidulafugin, iturin A, and mycosubtilin attenuated the cytopathogenicity of SARS-CoV-2 without specific toxicity.

Plant Alkaloids Inhibit Membrane Fusion Mediated by Calcium and Fragments of MERS-CoV and SARS-CoV/SARS-CoV-2 Fusion Peptides.

To rationalize the antiviral actions of plant alkaloids, the ability of 20 compounds to inhibit calcium-mediated fusion of lipid vesicles composed of phosphatidylglycerol and cholesterol was investigated using the calcein release assay and dynamic light scattering. Piperine, tabersonine, hordenine, lupinine, quinine, and 3-isobutyl-1-methylxanthine demonstrated the most potent effects (inhibition index greater than 50%). The introduction of phosphatidylcholine into the phosphatidylglycerol/cholesterol mixture led to significant changes in quinine, hordenine, and 3-isobutyl-1-methylxanthine efficiency. Comparison of the fusion inhibitory ability of the tested alkaloids, and the results of the measurements of alkaloid-induced alterations in the physical properties of model membranes indicated a potent relationship between a decrease in the cooperativity of the phase transition of lipids and the ability of alkaloids to prevent calcium-mediated vesicle fusion. In order to use this knowledge to combat the novel coronavirus pandemic, the ability of the most effective compounds to suppress membrane fusion induced by fragments of MERS-CoV and SARS-CoV/SARS-CoV-2 fusion peptides was studied using the calcein release assay and confocal fluorescence microscopy. Piperine was shown to inhibit vesicle fusion mediated by both coronavirus peptides. Moreover, piperine was shown to significantly reduce the titer of SARS-CoV2 progeny in vitro in Vero cells when used in non-toxic concentrations.

Antiviral activity of amides and carboxamides of quinolizidine alkaloid (-)-cytisine against human influenza virus A (H1N1) and parainfluenza virus type 3.

Novel derivatives of quinolizidine alkaloid (-)-cytisine were synthesised. ADME properties, cytotoxicity against HEK293 cells and activity against viruses of influenza A/California/07/09(H1N1)pdm09 virus (IAV) and human parainfluenza virus type 3 (HPIV3) were evaluated. It was shown, that 9-carboxamides of methylcytisine (with phenyl and allyl urea's fragments) are most active compounds against IAV probably due to predicted in silico peculiarity of their interactions with the 4R7B active site of IAV neuraminidase. Indexes of selectivity (SI) calculated as ratio of CC50/IC50 of these ureas are 47 and 59 correspondingly. It was also found, that derivatives obtained from allyl isocyanate and (-)-cytisine or 9,11-dibromocytisine are able to inhibit a reproduction of HPIV3 with SI = 58 and 95. Moreover, last compound - (1 R,5R)-N-allyl-9,11-dibromo-8-oxo-1,5,6,8-tetrahydro-2H-1,5-methanopyrido[1,2-a][1,5]diazocine-3(4H)-carboxamide with two bromine atom in 2-pyridone core of starting (-)-cytisine molecule, demonstrated high activity against HPIV3 (SI = 95) and moderate activity against IAV (SI = 16).

Synthesis of Camphecene and Cytisine Conjugates Using Click Chemistry Methodology and Study of Their Antiviral Activity.

A series of camphecene and quinolizidine alkaloid (-)-cytisine conjugates has been obtained for the first time using 'click' chemistry methodology. The cytotoxicity and virus-inhibiting activity of compounds were determined against MDCK cells and influenza virus A/Puerto Rico/8/34 (H1N1), correspondingly, in in vitro tests. Based on the results obtained, values of 50 % cytotoxic dose (CC50 ), 50 % inhibition dose (IC50 ) and selectivity index (SI) were determined for each compound. It has been shown that the antiviral activity is affected by the length and nature of linkers between cytisine and camphor units. Conjugate 13 ((1R,5S)-3-(6-{4-[(2-{(E)-[(1R,4R)-1,7,7-trimethylbicyclo[2.2.1]heptan-2-ylidene]amino}ethoxy)methyl]-1H-1,2,3-triazol-1-yl}hexyl)-1,2,3,4,5,6-hexahydro-8H-1,5-methanopyrido[1,2-a][1,5]diazocin-8-one), which contains cytisine fragment separated from triazole ring by -C6 H12 - aliphatic linker, showed the highest activity at relatively low toxicity (CC50 =168 µmol, IC50 =8 µmol, SI=20). Its selectivity index appeared higher than that of reference compound, rimantadine. According to theoretical calculations, the antiviral activity of the lead compound 13 can be explained by its influence on the functioning of neuraminidase.

5-Chloro-2-thiophenyl-1,2,3-triazolylmethyldihydroquinolines as dual inhibitors of Mycobacterium tuberculosis and influenza virus: Synthesis and evaluation.

This study describes synthesis and evaluation of novel 5-Chloro-2-thiophenyl-1,2,3-triazolylmethyldihydroquinolines 7a-o as dual inhibitors of Mycobacterium tuberculosis and influenza virus. Huisgen's [3+2] dipolar cycloaddition of 6-(azidomethyl)-5-chloro-2-(thiophen-2-yl)-7,8-dihydroquinoline 5 with various alkynes 6a-o using sodium ascorbate and copper sulphate gave new dihydroquinoline-1,2,3-triazoles 7a-o in good to excellent yields. The new compounds were evaluated for in vitro antimycobacterial against M. tuberculosis H37Rv (Mtb) and antiviral activity against influenza virus A/Puerto Rico/8/34 (H1N1). Among the fifteen new analogs, compounds 7a (MIC: 3.12 µg/mL), 7j and 7k (MIC: 6.25 µg/mL) were identified as potent antitubercular agents. The virus-inhibiting activity of all the fifteen compounds was found to be moderate, and among them the compound 7l, bearing thiophene moiety appeared the most active with good selectivity index (IC50 = 19.5 µg/mL; SI = 15). The results presented here will help developing newer dual inhibitors of tuberculosis and influenza virus.