Cold-adapted SARS-CoV-2 variants with different temperature sensitivity exhibit an attenuated phenotype and confer protective immunity.

As novel SARS-CoV-2 Variants of Concern emerge, the efficacy of existing vaccines against COVID-19 is declining. A possible solution to this problem lies in the development of a live attenuated vaccine potentially able of providing cross-protective activity against a wide range of SARS-CoV-2 antigenic variants. Cold-adapted (ca) SARS-CoV-2 variants, Dubrovka-ca-B4 (D-B4) and Dubrovka-ca-D2 (D-D2), were obtained after long-term passaging of the Dubrovka (D) strain in Vero cells at reduced temperatures. Virulence, immunogenicity, and protective activity of SARS-CoV-2 variants were evaluated in experiments on intranasal infection of Syrian golden hamsters (Mesocricetus auratus). In animal model infecting with ca variants, the absence of body weight loss, the significantly lower viral titer and viral RNA concentration in animal tissues, the less pronounced inflammatory lesions in animal lungs as compared with the D strain indicated the reduced virulence of the virus variant. Single intranasal immunization with D-B4 and D-D2 variants induced the production of neutralizing antibodies in hamsters and protected them from infection with the D strain and the development of severe pneumonia. It was shown that for ca SARS-CoV-2 variants, the temperature-sensitive (ts) phenotype was not obligate for virulence reduction. Indeed, the D-B4 variant, which did not possess the ts phenotype but had lost the ability to infect human lung cells Calu-3, exhibited reduced virulence in hamsters. Consequently, the potential phenotypic markers of attenuation of ca SARS-CoV-2 variants are the ca phenotype, the ts phenotype, and the change in species specificity of the virus. This study demonstrates the great potential of SARS-CoV-2 cold adaptation as a strategy to develop a live attenuated COVID-19 vaccine.

Immunogenic properties of SARS-CoV-2 inactivated by ultraviolet light.

Vaccination against COVID-19 is the most effective method of controlling the spread of SARS-CoV-2 and reducing mortality from this disease. The development of vaccines with high protective activity against a wide range of SARS-CoV-2 antigenic variants remains relevant. In this regard, evaluation of the effectiveness of physical methods of virus inactivation, such as ultraviolet irradiation (UV) of the virus stock, remains relevant. This study demonstrates that the UV treatment of SARS-CoV-2 completely inactivates its infectivity while preserving its morphology, antigenic properties, and ability to induce the production of virus-neutralizing antibodies in mice through immunization. Thus, the UV inactivation of SARS-CoV-2 makes it possible to obtain viral material similar in its antigenic and immunogenic properties to the native antigen, which can be used both for the development of diagnostic test systems and for the development of an inactivated vaccine against COVID-19.

Viral Membrane Fusion Proteins and RNA Sorting Mechanisms for the Molecular Delivery by Exosomes.

The advancement of precision medicine critically depends on the robustness and specificity of the carriers used for the targeted delivery of effector molecules in the human body. Numerous nanocarriers have been explored in vivo, to ensure the precise delivery of molecular cargos via tissue-specific targeting, including the endocrine part of the pancreas, thyroid, and adrenal glands. However, even after reaching the target organ, the cargo-carrying vehicle needs to enter the cell and then escape lysosomal destruction. Most artificial nanocarriers suffer from intrinsic limitations that prevent them from completing the specific delivery of the cargo. In this respect, extracellular vesicles (EVs) seem to be the natural tool for payload delivery due to their versatility and low toxicity. However, EV-mediated delivery is not selective and is usually short-ranged. By inserting the viral membrane fusion proteins into exosomes, it is possible to increase the efficiency of membrane recognition and also ease the process of membrane fusion. This review describes the molecular details of the viral-assisted interaction between the target cell and EVs. We also discuss the question of the usability of viral fusion proteins in developing extracellular vesicle-based nanocarriers with a higher efficacy of payload delivery. Finally, this review specifically highlights the role of Gag and RNA binding proteins in RNA sorting into EVs.

Antiviral Activity of Umifenovir In Vitro against a Broad Spectrum of Coronaviruses, Including the Novel SARS-CoV-2 Virus.

An escalating pandemic of the novel SARS-CoV-2 virus is impacting global health, and effective antivirals are needed. Umifenovir (Arbidol) is an indole-derivative molecule, licensed in Russia and China for prophylaxis and treatment of influenza and other respiratory viral infections. It has been shown that umifenovir has broad spectrum activity against different viruses. We evaluated the sensitivity of different coronaviruses, including the novel SARS-CoV-2 virus, to umifenovir using in vitro assays. Using a plaque assay, we revealed an antiviral effect of umifenovir against seasonal HCoV-229E and HCoV-OC43 coronaviruses in Vero E6 cells, with estimated 50% effective concentrations (EC50) of 10.0 ± 0.5 µM and 9.0 ± 0.4 µM, respectively. Umifenovir at 90 µM significantly suppressed plaque formation in CMK-AH-1 cells infected with SARS-CoV. Umifenovir also inhibited the replication of SARS-CoV-2 virus, with EC50 values ranging from 15.37 ± 3.6 to 28.0 ± 1.0 µM. In addition, 21-36 µM of umifenovir significantly suppressed SARS-CoV-2 virus titers (≥2 log TCID50/mL) in the first 24 h after infection. Repurposing of antiviral drugs is very helpful in fighting COVID-19. A safe, pan-antiviral drug such as umifenovir could be extremely beneficial in combating the early stages of a viral pandemic.