Kirill I. Yurlov

Pandemic preparedness is a complex of threat-agnostic countermeasures developed in advance which would be efficient against a future outbreak regardless of its causative agent, and broad-spectrum antivirals constitute a critical component of this complex. Plant polyphenols are known to suppress viruses of unrelated families by acting on multiple viral and cellular structures. We therefore searched for broad-spectrum antivirals among polyphenols that have been confirmed as safe to humans. The ellagitannin geraniin and galloylglucose constituents of the drug Rutan (1,2,3,4,6-penta-O-galloyl-β-D-glucose [R5], 3-bis-O-galloyl-1,2,4,6-tetra-O-galloyl-β-D-glucose [R6], 2,4-bis-O-galloyl-1,3,6-tri-O-galloyl-β-D-glucose [R7], 2,3,4-bis-O-galloyl-1,6-di-O-galloyl-β-D-glucose [R8]) were isolated from Geranium sanguineum and sumac (Rhus coriaria), respectively. We revealed their activity towards herpes simplex viruses (HSV-1 and HSV-2), human cytomegalovirus (CMV), and the Epstein–Barr virus (EBV). R5 suppressed HSV-1 and HSV-2 with equal efficiency, while Rutan and R7 were more active against HSV-1, and geraniin against HSV-2. Rutan and R5 also inhibited the intracellular replication of CMV and EBV (contrary to our expectations, geraniin and polyphenols R6–R8 showed no activity). Thus, we have shown for the first time that sumac polyphenols are capable of suppressing—in addition to HIV, influenza virus, and SARS-CoV-2—the reproduction of representatives of all three Orthoherpesviridae subfamilies, meeting the criteria for further development as broad-spectrum antivirals.

Introduction. Human cytomegalovirus infection can induce tumor cell resistance to chemotherapeutic agents through modulation of apoptotic pathways. In the search for alternative approaches to overcome virus-associated drug resistance, the application of nanomaterials (aqueous fullerene dC60) represents a promising strategy. The potential to overcome human cytomegalovirus mediated chemoresistance opens new avenues for developing combined therapeutic approaches in oncology. Aim – to evaluate the impact of human cytomegalovirus infection on the resistance of hepatocellular carcinoma and promyelocytic leukemia cells to doxorubicin, as well as the potential of aqueous fullerene dC60 to restore chemosensitivity in monocytic leukemia cells. Materials and methods. Hepatocellular carcinoma cells (Huh 7.5), promyelocytic leukemia cells (HL-60), monocytic leukemia cells (THP-1), and HCMV AD169 were used. The experimental procedures included standard cell culture techniques, virological methods, immunocytochemistry, Western blotting, Real-Time Polymerase Chain Reaction, Quantitative Reverse Transcription Polymerase Chain Reaction and MTT assay. Results. Human cytomegalovirus infection reduced doxorubicin cytotoxicity by 30% in both hepatocellular carcinoma and promyelocytic leukemia cells. In monocytic leukemia cells, combined treatment with doxorubicin and dC60 restored chemosensitivity to human cytomegalovirus infected cells, achieving 93% tumor cell death at half the standard doxorubicin concentration. Conclusion. Human cytomegalovirus infection induces doxorubicin resistance in both hematopoietic (promyelocytic leukemia, monocytic leukemia) and solid (hepatocellular carcinoma) tumor models. Importantly, combined treatment doxorubicin with aqueous fullerene dC60 not only overcomes virus-mediated drug resistance in monocytic leukemia cells but also enhances cytotoxicity at reduced doxorubicin concentrations, offering prospects for developing less toxic combined therapeutic regimens.

Introduction. One of the more promising developments in preventing the spread of infections, including COVID-19, is the production of vaccines based on virus-like particles (VLP). Currently, in the National Research Center for Epidemiology and Microbiology named after N.F. Gamaleya of the Ministry of Health of the Russia has developed a VLP-based vaccine against COVID-19. The aim of this study is to evaluate the tolerability, safety and immunogenicity of a new vaccine for the prevention of COVID–19 based on VLP compared with placebo for 21 days after two intramuscular injections in phase I clinical trials. Materials and methods. A double-blind, placebo-controlled study of the tolerability, safety and immunogenicity of a vaccine for the prevention of COVID-19 based on VLP was conducted with a dose of the drug containing 40 and 80 micrograms of antigen, the placebo being 0.9% NaCl. The presence or absence of adverse events (AEs) after vaccination was noted in 180 volunteers aged 18 to 55 years; clinical and biochemical blood parameters, the intensity of humoral and cellular immunity before and after vaccination were assessed using enzyme immunoassay, neutralization reactions, lymphocyte blast transformation reactions and flow cytometry. Results. An analysis of the tolerability and safety of the new COVID-19 VLP-vaccine showed that most adverse events were registered within the first 10 days after vaccination, mainly after the first vaccination. In the period from 11 to 21 days after vaccination, AEs were observed in isolated cases. No deaths, serious or other AEs have been reported. The administration of the studied vaccine to the volunteers had no negative effect on the basic vital signs. A comparative analysis of immunogenicity indicators in volunteers showed that the administration of a vaccine with both an antigen content of 40 µg and an antigen content of 80 µg leads to a pronounced and significant increase in the level of specific immunoglobulins, virus neutralizing antibodies and activation of a cell-mediated immune response. As part of the phase I clinical trials, a dose of 80 µg was selected as optimal. Conclusion. It has been shown that a new vaccine for the prevention of COVID-19 based on VLP with an antigen content of 40 and 80 µg when administered intramuscularly to volunteers does not cause serious adverse events and induces a tense humoral and cellular immune response.

Introduction. The COVID-19 pandemic caused by SARS-CoV-2 has created serious health problems worldwide. The most effective way to prevent the occurrence of new epidemic outbreaks is vaccination. One of the modern and effective approaches to vaccine development is the use of virus-like particles (VLPs). The aim of the study is to develop a technology for production of VLP based on recombinant SARS-CoV-2 proteins (E, M, N and S) in insect cells. Materials and methods. Synthetic genes encoding coronavirus proteins E, M, N and S were used. VLP with various surface proteins of strains similar to the Wuhan virus, Delta, Alpha and Omicron were developed and cloned into the pFastBac plasmid. The proteins were synthesized in the baculovirus expression system and assembled into VLP in the portable Trichoplusia ni cell. The presence of insertion in the baculovirus genome was determined by PCR. ELISA and immunoblotting were used to study the antigenic activity of VLP. VLP purification was performed by ultracentrifugation using 20% sucrose. Morphology was assessed using electron microscopy and dynamic light scattering. Results. VLPs consisting of recombinant SARS-CoV-2 proteins (S, M, E and N) were obtained and characterized. The specific binding of antigenic determinants in synthesized VLPs with antibodies to SARS-CoV-2 proteins has been demonstrated. The immunogenic properties of VLPs have been studied. Conclusion. The production and purification of recombinant VLPs consisting of full-length SARS-CoV-2 proteins with a universal set of surface antigens have been developed and optimized. Self-assembling particles that mimic the coronavirus virion induce a specific immune response against SARS-CoV-2.

Introduction. SARS-CoV-2 infection causes immune disorders that create conditions for the reactivation of human herpesviruses (HHVs). However, the estimates of the HHVs effect on the course and outcome of COVID-19 are ambiguous. Аim – to study the possible relationship between the HHV reactivation and the adverse outcome of COVID-19. Materials and methods. Postmortem samples from the brain, liver, spleen, lymph nodes and lungs were obtained from 59 patients treated at the Moscow Infectious Diseases Hospital No.1 in 2021–2023. The group 1 comprised 39 patients with fatal COVID-19; group 2 (comparison group) included 20 patients not infected with SARS-CoV-2 who died from various somatic diseases. HHV DNA and SARS-CoV-2 RNA were determined by PCR. Results. HHV DNA was found in autopsy samples from all patients. In group 1, EBV was most often detected in lymph nodes (94%), HHV-6 in liver (68%), CMV in lymph nodes (18%), HSV in brain (16%), VZV in lung and spleen (3% each). The detection rates of HHVs in both groups was similar. Important differences were found in viral load. In patients with COVID-19, the number of samples containing more than 1,000 copies of HHV DNA per 100,000 cells was 52.4%, in the comparison group – 16.6% (p 0.002). An association has been established between the reactivation of HSV and HHV-6 and the severity of lung damage. Reactivation of EBV correlated with increased levels of liver enzymes. Conclusion. Reactivation of HHVs in patients with fatal COVID-19 was associated with severe lung and liver damages, which indicates a link between HHV reactivation and COVID-19 deaths.

Introduction. In Russia, almost half of the cases of acute intestinal infections of established etiology in 2022 are due to rotavirus infection (RVI). There is no specific treatment for rotavirus gastroenteritis. There is a need to develop modern, effective and safe vaccines to combat rotavirus infection that are not capable of multiplying (replicating) in the body of the vaccinated person. A promising approach is to create vaccines based on virus-like particles (VLPs). Objective. Study of the safety and immunogenicity of a vaccine against rotavirus infection based on virus-like particles of human rotavirus A in newborn minipigs with multiple intramuscular administration. Materials and methods. Newborn minipigs were used as an animal model in this study. The safety of the tested vaccine was assessed based on thermometry data, clinical examination, body weight gain, clinical and biochemical blood parameters, as well as necropsy and histological examination. When studying the immunogenic properties of the Gam-VLP-rota vaccine in doses of 30 and 120 µg, the cellular, humoral and secretory immune response was studied. Results. The results of assessing the general condition of animals during the immunization period, data from clinical, laboratory and pathomorphological studies indicate the safety of the vaccine against human rotavirus infection based on VLP (Gam-VLP-rota) when administered three times intramuscularly. Good local tolerance of the tested vaccine was demonstrated. The results of the assessment of humoral immunity indicate the formation of a stable immune response after three-time immunization with Gam-VLP-rota, stimulation of the production of antigen-specific IgG antibodies and their functional activity to neutralize human rotavirus A. It was shown that following the triple immunization with the minimum tested concentration of 30 µg/dose, animals developed a cell-mediated immune response. The results of the IgA titer in blood serum and intestinal lavages indicate the formation of both a systemic immunological response and the formation of specific secretory immunity to human rotavirus A. Conclusion. Thus, three-time intramuscular immunization of minipigs with the Gam-VLP-rota vaccine forms stable protective humoral and cellular immunity in experimental animals. Evaluated vaccine is safe and has good local tolerability.

Hepatitis C virus (HCV) is one of the basic culprits behind chronic liver disease, which may result in cirrhosis and hepatocarcinoma. In spite of the extensive research conducted, a vaccine against HCV has not been yet created. We have obtained human mesenchymal stem cells (hMSCs) and used them for expressing the HCV NS5A protein as a model vaccination platform. Sixteen hMSC lines of a different origin were transfected with the pcNS5A-GFP plasmid to obtain genetically modified MSCs (mMSCs). The highest efficiency was obtained by the transfection of dental pulp MSCs. C57BL/6 mice were immunized intravenously with mMSCs, and the immune response was compared with the response to the pcNS5A-GFP plasmid, which was injected intramuscularly. It was shown that the antigen-specific lymphocyte proliferation and the number of IFN-γ-synthesizing cells were two to three times higher after the mMSC immunization compared to the DNA immunization. In addition, mMSCs induced more CD4+ memory T cells and an increase in the CD4+/CD8+ ratio. The results suggest that the immunostimulatory effect of mMSCs is associated with the switch of MSCs to the pro-inflammatory phenotype and a decrease in the proportion of myeloid derived suppressor cells. Thus, the possibility of using human mMSCs for the creation of a vaccine against HCV has been shown for the first time.

Introduction. The role of human papilloma viruses is well-studied in a number of cancers. However, the role of urogenital HPV infection and its association with pathospermia, male infertility, and genitourinary tract inflammatory diseases are understudied. Objective. To determine the prevalence of HPV in patients with chronic prostatitis and infertility. Materials and methods. Quantitative PCR was used to detect high carcinogenic risk (HCR) HPV in the ejaculate of 489 males aged 22–65. Group 1 included infertile males with chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS), Group 2 included males with idiopathic infertility (without CP/CPPS), Group 3 included fertile males with CP/CPPS, Group 4 included healthy fertile males (control group). Results. 64 (14.6%) of the examined males had HPV. None of the males from the control group had HCR HPV in the ejaculate. Group 1 had a significantly higher number of all HPV genotypes (40 (22.2%) of 180) than Group 2 (25 (14.3%) of 175) and Group 3 (13 (15.9%) of 82). HPV viral load was highest in Group 3 (~2500 copies/100.000 cells). Conclusion. HCR HPV is associated with male infertility and chronic prostatitis/chronic pelvic pain syndrome. The association of mixed infection with various HPV types with infertility and high HPV viral load with CP/CPPS was shown.

Acute and chronic liver diseases are a major global public health problem; nevertheless, the etiology of 12–30% of cases remains obscure. The purpose of this research was to study the incidence of human herpesviruses (HHVs) cytomegalovirus, Epstein–Barr virus (EBV), and HHV-6 in patients with hepatitis and to examine the effect of HHV on the disease severity. We studied the clinical materials of 259 patients with hepatitis treated in Infectious Clinic n.1 (Moscow) and the archived materials of 118 patients with hepatitis C. HHV DNA was detected in the whole blood in 13.5% of patients with hepatitis B or C and in 10.1% of patients with hepatitis of unspecified etiology. EBV demonstrated the highest incidence (58.1%). Cirrhosis was diagnosed in 50% of patients with HHV and in 15.6% of patients without HHV. In patients with hepatitis C, the frequency of HHV was higher in liver biopsy (38.7%) compared to blood. The clinical and virological indicators of hepatitis were considerably higher in patients with coinfection. Conclusion: HHV detected in patients with viral hepatitis has been associated with a significant effect on the severity of the disease, and we suggest monitoring HHV DNA in patients with severe hepatitis and/or poor response to antiviral drugs.