Synthesis of esters and amides of 2-aryl-1-hydroxy-4-methyl-1H-imidazole-5-carboxylic acids and study of their antiviral activity against orthopoxviruses.

Smallpox was eradicated >40 years ago but it is not a reason to forget forever about orthopoxviruses pathogenic to humans. Though in 1980 the decision of WHO to cease vaccination against smallpox had seemed logical, it led to the decrease of cross immunity against other infections caused by orthopoxviruses. As a result, in 2022 the multi-country monkeypox outbreak becomes a topic of great concern. In spite of existing FDA-approved drugs for the treatment of such diseases, the search for new small-molecule orthopoxvirus inhibitors continues. In the course of this search a series of novel 2-aryl-1-hydroxyimidazole derivatives containing ester or carboxamide moieties in position 5 of heterocycle has been synthesized and tested for activity against Vaccinia virus in Vero cell culture. Some of the compounds under consideration revealed a selectivity index higher than that of the reference drug Cidofovir. The highest selectivity index SI = 919 was exhibited by ethyl 1-hydroxy-4-methyl-2-[4-(trifluoromethyl)phenyl]-1H-imidazole-5-carboxylate 1f. The most active compound also demonstrated inhibitory activity against the cowpox virus (SI = 20) and the ectromelia virus (SI = 46).

Anti-Influenza Activity of Cetraria islandica Lichen Extracts in In Vitro Experiments.

The toxicity and antiviral activity of extracts obtained by the methods of aqueous and ethanol extraction of bioactive substances from Cetraria islandica lichen as a raw material were studied. Aqueous and ethanol extracts of lichen were characterized by low toxicity with respect to the passaged MDCK cell culture and exhibited antiviral activity. The ethanol extract showed more potent in vitro antiviral activity against human A/H3N2 and avian A/H5N1 influenza viruses: in a concentration of 50 µg/ml, it suppressed replication of these viruses by 3.5 and 4 log10, respectively, while the aqueous extract inhibited replication of viruses by 2 and 6 log10, respectively, when taken in a concentration of 500 µg/ml that was 10-fold higher than the concentration of the ethanol extract.

Estimation of Absolute Bioavailability of the Chemical Substance of the Anti-Smallpox Preparation NIOCH-14 in Mice.

We compared absolute bioavailability of the chemical substance of the anti-smallpox preparation NIOCH-14 and chemical compound ST-246 active against orthopoxviruses after oral administration to mice in doses of 10 and 50 µg/g and intravenous administration to mice in a dose of 2 µg/g body weight. The absolute bioavailability of NIOCH-14 is comparable with the absolute bioavailability of ST-246.

[EVALUATION OF THE HUMAN SENSITIVITY TO SMALLPOX VIRUS BY THE PRIMARY CULTURES OF THE MONOCYTE-MACROPHAGES].

Studies of the primary cultures of granulocytes, mononuclear, and monocyte-macrophage cells derived from human blood were performed using variola virus (VARV) in the doses of 0.001-0.021 PFU/cell (plaques-forming units per cell). Positive dynamics of the virus accumulation was observed only in the monocyte-macrophages with maximum values of virus concentration (5.0-5.5 Ig PFU/ml) mainly within six days after the infection. The fact of VARV replication in the monocyte-macrophages was confirmed by the data of electron microscopy. At the same time, virus vaccines when tested in doses 3.3 and 4.2 Ig PFU/ml did not show the ability to reproduce in these human cells. The people sensitivity to VARV as assessed from the data obtained on human monocyte-macrophages corresponded to -1 PFU (taking into account the smooth interaction of the virus in the body to the cells of this type), which is consistent to previously found theoretical data on the virus sensitivity. The human susceptibility to VARV assessed experimentally can be used to predict the adequacy of developed smallpox models (in vivo) based on susceptible animals. This is necessary for reliable assessment of the efficiency of development of drugs for treatment and prophylaxis of the smallpox.

[THE USE OF THE MODEL MOUSE ICR--VARIOLA VIRUS FOR EVALUATION OF ANTIVIRAL DRUG EFFICACY].

Mice of the ICR outbred population were infected intranasally (i/n) with the variola virus (VARV, strain Ind-3a). Clinical signs of the disease did not appear even at the maximum possible dose of the virus 5.2 lg PFU/head (plaque-forming units per head). In this case, 50% infective dose (ID50) of VARV estimated by the presence or absence of the virus in the lungs three days after infection (p.i.) was equal to 2.7 ± 0.4 lg PFU/head. Taking into account the 10% application of the virus in the lungs during the intranasal infection of the mice, it was adequate to 1.7 lg PFU/lungs. This indicates a high infectivity of the VARV for mice comparable to its infectivity for humans. After the i/n infection of mice with the VARV at a dose 30 ID50/ head the highest concentration of the virus detected in the lungs (4.9 ± 0.0 lg PFU/ml of homogenate) and in nasal cavity tissues (4.8 ± 0.0 lg PFU/ml) were observed. The pathomorphological changes in the respiratory organs of the mice infected with the VARV appeared at 3-5 days p.i., and the VARV reproduction noted in the epithelial cells and macrophages were noticed. When the preparations ST-246 and NIOCH-14 were administered orally at a dose of 60 µg/g of mouse weight up to one day before infection, after 2 hours, 1 and 2 days p.i., the VARV reproduction in the lungs after 3 days p.i. decreased by an order of magnitude. Thus, outbred ICR mice infected with the VARV can be used as a laboratory model of the smallpox when evaluating the therapeutic and prophylactic efficacy of the antismallpox drugs.

[Role of the tissue antioxidant status in response to chronic irradiation of mice during early ontogenesis].

The response of the liver and blood erythrocyte lipids to the low intensity chronic γ-irradiation action of mice at the dose of 8 cGy during early ontogenesis immediately and 7 months after irradiation is studied. The maintainance of the changed structural state of the lipid component in liver and especially blood erythrocytes which are characterized by a lower antioxidant status compared with the liver lipids is revealed during a long time after the cessation of irradiation. This confirms the possibility of using blood erythrocyte lipids as a perspective model for the estimation of the effects of the weak impact of unfavorable factors on the organism.

[Development of the disease in marmot at the intranasal infection with the monkeypox virus].

In experimental study the sensitivity of the Marmota bobak species to the monkeypox virus (MPXV) with the intranasal (i/n) infection was tested. It was demonstrated that 50% of the infective dose (ID50) of the MPXV on external clinical signs of the disease was 2.2 Ig plaque forming units (PFU). The percentage of the marmot mortality is slightly dependent on the infecting dose of the MPXV, therefore it is not possible to correctly determine the value of 50 % fatal dose (FD50) for these animals. The most pronounced external clinical signs of the disease were obtained in the marmots: pox-like skin rash throughout the surface of the body and mucous membranes, purulent discharge from the nose, lymphadenitis, discoordination, tremor of the extremities, fever, increased aggression, and ruffled fur. In the course of experiments intended to determine the dynamics of the accumulation of the MPXV in various organs, tissues, and blood serum of marmot infected i/n with dose of 3.7 Ig PFU, it was found that the trachea, lungs, and the bifurcation lymph nodes are the primary target organs. The trachea, lungs, nasal mucosa membrane, and skin are the organs with maximal virus replication recorded at 5, 7, 9, and 12 days after the infection. The transfer of the MPXV into the secondary target organs (nasal mucosa membrane, brain, spleen, duodenum, adrenal glands, and skin) was carried out in marmots with lymphogenic and hematogenic ways of the dissemination of the infection.

[Evaluation of therapeutic-prophylactic effectiveness of chemical compound NIOC-14 against ectromelia virus in vivo].

AIM: Study pharmacodynamic parameters of anti-viral effectiveness of a chemical compound NIOC-14 in experiments in mice infected with ectromelia virus (EV). MATERIALS AND METHODS: EV (K-1 strain) was obtained from the State Collection of Viral Infections and Rickettsioses Causative Agents of the State Scientific Centre of Virology and Biotechnology "Vector". Outbred ICR mice were intranasally infected with EV at a dose of 10 LD50 per animal (10 x 50% lethal doses/animal) and per orally received NIOC-14 or ST-246 as a positive control. Chemical compound NIOC-14 (7-[N'-(4-trifluoromethylbenzoyl)-hidrazincarbonyl]-tricyclo[3.2.2.0(2,4)]non-8-en-6-carbonic acid) was synthesized in Novosibirsk Institute of Organic Chemistry (NIOC). Anti-pox preparation ST-246, developed by SIGA Technologies Inc. (USA), was synthesized in NIOC using the technique described by the authors. RESULTS: 50% effective doses against EV in vivo were shown not to differ significantly between the preparations NIOC-14 (3.59 µg/g mouse mass) and ST-246 (5.08 µg/g mouse mass). During determination of therapeutic window, administration of NIOC-14 to mice 1 day or 1 hour before EV infection, as well as 1, 2 and 4 days after EV infection and then for 9 days was found to ensure 100% animal survival. Administration of NIOC-14 as well as ST-246 resulted in the decrease relative to control of EV titers in lungs, nasal cavity, brains, liver, spleen, kidneys and pancreas. CONCLUSION: Anti-viral effectiveness of NIOC-14 against EV in vivo was thus comparable by all the studied pharmacodynamic parameters with anti-viral activity of anti-pox-virus preparation ST-246.

[A new approach to analysis of participation of oxidative processes in regulation of metabolism in animal tissues].

A new approach to the analysis of the oxidative processes participated in regulation of metabolism in norm and the assessment of the biological consequences under the action of damaging factors of different nature and intensity for animal groups is suggested using the analysis of interrelations between the different parameters of the physicochemical regulatory system of the lipid peroxidation in tissues of the laboratory mice (the phospholipid composition, amounts of the oxidation products in lipids). by changes of the scale and direction of interrelations between the reciprocal parameters of the lipid peroxidation system in norm.

[Peculiarities of antioxidants as radioprotectors under radiation damage of different extent].

Results of the own and literature data about the effectiveness of antioxidants as radioprotectors under the action of ionizing irradiation in animals are generalized dependingon the extent of the radiation damage and the linear energy loss of radiation. The initial antioxidant status of tissues is shown to make a different con- tribution in the manifestation of the radioprotective properties of natural and synthetic antioxidants owing to the non-linearity of the lipid peroxidation processes depending on the radiation dose and dose rate. Thus, a complex approach and consideration of the time factor are necessary when estimating radioprotective effectiveness of antioxidants as radoprotectors under radiation damage of different extent.

[Interrelation between structural state, physicochemical properties and lipid composition in microbial cells].

We performed complex investigations of antioxidant properties and the lipid composition of the five gram-negative bacteria depending on the cultivation season. Structural state, the lipid composition and physicochemical properties of lipids of the three gram-negative bacteria (Renobacter vacuolatum, Flectobacillus major WKM 869, Pseudomonas fluorescens) during their growth and mycelium of four species of the xylotrophic basidiomycetes (Panus tigrinus IBK-131, Fomes fomentarius M71, Laetiporus sulfureus M131, Piptoporus betulinus M60) during the lag phase were also studied. Changes in the composition of the lipid component in the studied bacteria led to considerable structural rearrangements in their membrane system during the growth of bacteria. It is shown that more hydrophilic regions of the lipid component are of paramount importance in maintaining the fluidity of the entire lipid bilayer in the basidiomycete mycelium. The scale and direction of the revealed interrelations between rotational correlation times of probes and the lipid composition depend both on localization of the probe in membrane and the composition and physicochemical properties of the microorganism lipids.

[A comparative study of the antiviral activity of chemical compounds concerning the orthopoxviruses experiments in vivo].

In the experiments using intranasal (i/n) infection of mice with the ectromelia virus (EV) in a dose 10 LD50/head (10 x 50% lethal doselhead) or with the monkaypox virus (MPXV) in a dose 10 ID50/head (10 x 50% infective dose/ head) it was demonstrated that the antiviral efficiency of chemical compounds - the condensed derivatives of pyrrolidin-2,5-dion, as well as their predecessors and the nearest analogues, synthesized in Novosibirsk Institute of Organic Chemistry of the Siberian Branch of the Russian Academy of Sciences (NIOCH SB RAS) was observed. As a positive control we used the antipoxvirus chemical preparation ST-246 available from SIGA Technologies Inc. (USA), synthesized in NIOCH SB RAS by the technique suggested by the authors. It was demonstrated that the compound NIOCH-14 (7-[N'-(4-Trifluoromethylbenzoil)-hydrazidecarbonil]-tricyclo[3.2.2.02,4]non-8-en-6-carbonic acid) possessed comparable with ST-246 antiviral activity concerning EV and MPXV on all indicators used. Therefore, at infection of mice with EV (strain K-1) and peroral administration of NIOCH-14 and ST-246 in a dose 50 mkg/g of mouse weight (12-14 g) within 10 days the survival rate and average life expectancy of mice authentically exceeded the control levels. EV titers in lungs through 6 days after infection in the same groups were lower than in the control. In addition to that, after 7 days of infection of mice with MPXV (strain V79-1-005) and daily peroral administration of NIOCH-14 and ST-246 in a dose 60 mkg/g of mouse weight (9-11 g) authentic decrease in a part of infected animals and MPXV titers in lungs was observed.

[The blood erythrocyte lipid composition of mice exposed to the chronic radiation action at low doses during the early stages of ontogeny].

The comparative analysis of the blood erythrocyte lipid composition of the laboratory mice (males and females) after exposure to the chronic radiation action at the dose of 8 cGy during the early stages of the ontogeny was performed within the early and remote periods after irradiation. This action is shown to cause the marked effect both on the quantitative changes in the lipid composition and the structural state of the blood erythrocyte lipid component as follows from the numerous disturbances of the correlation interrelations between parameters of the lipid composition which are due to the structural state of the erythrocyte membrane. The most substantial changes in the blood erythrocyte lipid composition were detected for males within the remote period after the exposure, which provides evidence in favor of the disorder in the hemopoiesis process. The data obtained allow us to suggest that the erythrocyte membrane viscosity under the chronic low intensity irradiation is supported by the balance maintenance between the relative content of the phospholipid lysoforms and the molar ratio of the [sterols]/[phospholipids].

[Significance of the initial state of the murine liver for the development of consequences after the combined action of the factors of different nature].

The search of the most significant parameters of the liver initial state of mice Balb/c (males) for the development of the biological consequences under a combined action of the Tween-80 (0.3% solution) in the 10% solution of the aqueous acetone and X-rays at the doses of 4 and 5 Gy during one month after exposure was performed by means of the software program for the experimental data analysis. The study has shown both the absence of the same correlation relationships and the existence of two groups of relationships in terms of the coefficient correlation values under the alteration of the X-ray dose. The assumption has been made about different mechanisms underlying regulation of the biochemical process in the murine liver after the combined action of low-toxicity chemical agents at low doses and X-rays irradiation at sublethal doses, whose contribution to the development of effects of different factors depends on the radiation dose.

[Change of the lipid parameters in the murine liver during one month after low intensity X-ray exposure at low doses].

Parameters of the physicochemical regulatory system of lipid peroxidation in the liver of white outbred mice (females) were studied before and during one month after X-ray exposure at the doses less than 1.5 mGy in the autumn and spring-summer seasons. The initial value of parameters is found to exert the most substantial influence on the liver relative mass, the phosphatidylcholine and lysoform relative content in the liver phospholipids of mice. The reliable diminution and the substantial influence of the dose rate dynamics during irradiation are revealed for the molar ratio of [sterols]/[phospholipids], the phosphatidylcholine/phosphatidylethanolamine ratio and the ratio of sums of the more easily oxidizable to the more poorly oxidizable fractions ofphospolipids. The experimental data testify to the complicated nonlinear character of the biological effects of X-irradiation at low doses.

[Surface active properties of isobornylphenols in systems with different degrees of complexity].

Interrelations between the structure of the semi-synthetic phenolic antioxidants -- isobornylphenols and their surface active properties were studied in the chemical (the lecithin aggregation in hexane) and biological (the incubation with the blood erythrocytes) model systems. It has been shown that all studied compounds are able to affect the lecithin aggregation in hexane: the share of the main fraction of the L micelles decreases with increasing the share of particles of greater size. The effect substantially depends on hindered OH group and the presence of the intramolecular hydrogen bond in molecule. The cytotoxic properties of isobornylphenols (the concentration is 100 M) are predominantly due to the molecule structure. The interrelation between the aggregate size of the main fraction of L in the presence of the studied compounds and the discocyte share during mice blood erythrocyte incubation in their presence for 4 h is revealed. Thus, this provides the possibility to assume that the ability of the different biological active substances to affect the lecithin aggregation in non-polar solvent could be used as a model system for the initial assessment of their surface active properties.

[Characteristics of erythrocyte lipids in blood of tundra voles (Microtus oeconomus Pall.) inhabiting areas with increased natural radioactivity].

Interrelations between the lipid characteristics of the blood erythrocytes and 226Ra accumulation in the body of tundra voles (Microtus oeconomus Pall.) inhabiting areas with different levels of the radiation background were investigated. It is shown that the ratio of the phospholipid (PL) fractions which cause the blood erythrocyte structure depends on the phase of the population cycle, as well as on the sex and age of tundra voles. The statistically significant interrelation between lysoforms and the sphingomielin content has been revealed in the blood erythrocyte PL of tundra voles; its scale somewhat differs for the animals from the reference and Ra areas. The peroxide concentration in the blood erythrocyte lipids of tundra voles from the Ra area exceeded the control values in all mature groups of the animals trapped at the depression phase of the population density. The 226Ra content in the bodies of the tundra voles which were trapped in the Ra area at the increased phase of the population cycle is for certain higher than that for the animals from the reference area. Interrelations between the lipid peroxidation parameters in the blood erythrocytes and the body 226Ra content for separate sex-age groups of tundra voles have been revealed.

[Participation of the lipid peroxidation processes in the adaptation mechanism of Muridae like rodents to the radioactive contamination of the Chernobyl NPP zone].

Results of the comparative analysis of the complex investigation of rodent population state caught at areas with different levels of contamination in the accident zone of the Chernobyl NPP during 1986-1993 and 2007, at areas with a practically normal radiation background in the neighborhood of the Kiev City in 1993 and at areas with the normal and increased radiation background in Ukhta region of Komi Republic in 1993-2008 are presented. It is shown that the absence of the linear dependence in changes of the lipid peroxidation (LPO) regulatory system parameters in rodent tissues on the dose of the external gamma-radiation at areas of their trap persists for a long time. Different ability to normalization of the studied indices and nonlinearity of their dose dependences allow us to suggest that changes of the scale and direction of interrelations between the reciprocal parameters of the LPO regulatory system in norm in tissues of rodents which were caught in the accident zone and at areas with an increased radiation background should be the information signal determining selection of the strategy of adaptation to the chronic radiation factor action.

[Oxazole derivatives as compounds modifying radiation effects in the body of mice].

A new class of substances exhibiting radioprotective and radiosensitizing effects depending on the concentration of the substance has been found. The radioprotective effect is probably due to the resonant absorption of radiation energy and its transformation into low-energy forms, as well as reactions with water radiolysis products. We studied the effects of 2,5-difeniloxazole and di[2-(5-feniloxazolil)]benzene in various concentrations in conjunction with irradiation on the growth of melanoma B-16 in mice and the average time of their lives. When using individual doses of irradiation and doses of preparations, we observed an increase in the average lifetime of mice and a reduced tumor size. These data allow us to conclude about the possibility of using these substances in the radiotherapy of tumors.

[Effect of physicochemical properties of murine liver lipids on the interrelation between the parameters of their composition].

The effect of properties of liver lipids on the interrelation between the contents of different lipid fractions has been studied on nine groups of intact SHK female mice. To reveal interrelations, lipids were divided into four groups depending on their antioxidative activity, prooxidant properties, antiperoxide activity, and the content of peroxide in lipids. The data obtained indicate that the physicochemical properties of lipids affect the biosynthesis and degradation of phospholipids, which plays a significant role in the origination of consequences under the action of unfavourable factors on biological objects having the different intensity of oxidative processes.