SARS-CoV-2: a quick review of some of the available treatment options

Background: With more than 244 million cases worldwide, the SARS-CoV-2 pandemic has affected almost every country on the planet. Its impact on the health, economy, education and scientific systems, among others, has been significant. The search for therapeutic tools for the treatment and control of this new disease, COVID-19, has been intense, but it has not yet been possible to select a specific and effective drug for its treatment. Methodology: the objective of this work is to summarize the existing information, related to some of the most common drugs used in therapy against COVID-19, to achieve this a search methodology was used in scientific and general databases (Pubmed, Google Scholar) using keywords related to the topic of interest. The results show that the most common drugs used in the treatment of this disease have mostly been evaluated in clinical trials. Conclusion(s): despite almost two years passed since the first case of COVID-19, effective and specific drugs have not been developed or reassigned for the treatment of the disease and some of of the already evaluated have shown controversial results. Copyright © 2021, Revista Salus. All rights reserved.

Cellular stress and SARS-CoV-2

Background: SARS-CoV-2, the etiological agent responsible for COVID-19, is an RNA virus belonging to the Coronaviridae family. During the virus replication, viral components interact with the cellular machinery, inducing alterations in cell physiology, which contributes to viral pathogenesis. Method(s): A bibliographical research about cellular stress and SARS-CoV-2 was performed at NCBI/Pubmed. Result(s): In response to the infection, signaling pathways are activated in the host cell, the goal of these pathways being to restore homeostasis. If homeostasis is not recovered, the signaling leads to cell death activation. Among the best-characterized signaling pathways, the cellular stress pathways such as oxidative stress, UPR, and autophagy stand out, which are evolutionarily conserved and are also interconnected with each other. There is strong theoretical and experimental evidence of various interactions of some components of these pathways with different viral proteins of coronavirus, and some studies with SARS-CoV-2 have already been performed. In this review, we highlight some of the cellular pathways-virus characterized to date. Conclusion(s): The cellular pathways and their relationship to viral infections remains unclear. The study of these relationships might constitute an important target for new research and the development of antiviral therapies. Copyright © 2021, Revista Salus. All rights reserved.

Biology and genetic diversity of SARS-CoV-2, the causative agent of COVID-19

The infection generated by SARS-CoV-2 has caused more than 200 million cases and 4.5 million deaths worldwide. SARS-CoV-2 has accumulated mutations that allow it to be classified into different lineages. Some of these lineages have been designated variants by the WHO: under monitoring (VUM), of interest (VOI), or of concern (VOC). Methodology. Different strategies for genomic surveillance of these SARS-CoV-2 variants have been described in each country. In Venezuela, the strategies include the amplification of a fragment of the spike, PCR-RFLP, and sequencing of the complete viral genome, which has allowed us to monitor the introduction of VOCs and VOIs to the country. Results. By October 2021, in Venezuela, the circulation of three VOCs, Alpha, Gamma, and Delta, and the two VOIs (Lambda and Mu) have been described. Globally, the Delta variant predominates in practically all continents except some Latin American countries, although it is estimated that it will soon prevail in the region as well. Discussion. The circulation of variants in the countries is a very dynamic process and Venezuela does not escape from this reality;therefore, it is important to continue genomic surveillance of this virus. Copyright © 2021, Revista Salus. All rights reserved.

Sub-lineages of the Omicron variant of SARS-CoV-2: characteristic mutations and their relation to epidemiological behavior

By the end of 2021, the Omicron variant of SARS-CoV-2, the coronavirus responsible for COVID-19, emerges, causing immediate concern, due to the explosive increase in cases in South Africa and a large number of mutations. This study describes the characteristic mutations of the Omicron variant in the Spike protein, and the behavior of the successive epidemic waves associated to the sub-lineages throughout the world. The mutations in the Spike protein described are related to the virus ability to evade the protec-tion elicited by current vaccines, as well as with possible reduced susceptibility to host proteases for priming of the fusion process, and how this might be related to changes in tropism, a replication enhanced in nasal epithelial cells, and reduced in pulmonary tissue;traits probably associated with the apparent reduced severity of Omicron compared to other variants. © 2022, Instituto de Investigaciones Clinicas. All rights reserved.

Detection of the Omicron variant of SARS-CoV-2 by restriction analysis targeting the mutations K417N and N440K of the spike protein

By the end of 2021, the Omicron variant of concern (VOC) emerges in South Africa. This variant caused immediate concern, due to the explosive increase in cases associated with it and the large number of mutations it exhibits. In this study, the restriction sites that allow detecting the mutations K417N and N440K in the Spike gene are described. This analysis al-lows us to propose a rapid method for the identification of cases infected with the Omicron variant. We show that the proposed methodology can contribute to provide more information on the prevalence and rapid detection of cases of this new VOC. © 2022, Instituto de Investigaciones Clinicas. All rights reserved.

A simple method for detection of mutations in amino acid 452 of the Spike protein of SARS-CoV-2 using restriction enzyme analysis

Variants of Concern or Interest of SARS-CoV-2 (VOC or VOI), the coronavirus responsible for COVID-19, have emerged in several countries. Mutations in the amino acid 452 of the Spike protein are particularly important and associated with some of these variants: L452R, present in Delta VOC, and L452Q, present in Lambda VOI. These mutations have been associated with both increased infectivity and evasion of protective immune response. A search on GISAID to detect the number of sequences harboring the L452R mutation and the frequency of Delta VOC among them, showed that since August 2021, most of these sequences belong to the Delta VOC. Restriction enzyme analysis is proposed as a rapid method to detect L452R. A small amplicon from the Spike gene was digested with MspI. A 100% concordance was observed between digestion and sequencing results. The mutation L452Q can also be detected by restriction analysis, allowing the identification of putative Lambda VOIs. The proposed methodology, which allows screening of a great number of samples, could provide a faster information on the prevalence of Delta VOC cases.

SARS-CoV-2 genetic diversity in Venezuela: predominance of D614G variants and analysis of one outbreak

SARS-CoV-2 is the new coronavirus responsible for COVID-19 disease. The first two cases of COVID-19 were detected in Venezuela on March 13, 2020. The aim of this study was the genetic characterization of Venezuelan SARS-CoV-2 isolates. A total of 7 full SARS-CoV-2 genome sequences were obtained by Sanger sequencing, from patients of different regions of Venezuela, mainly from the beginning of the epidemic. Ten out of 11 isolates (6 complete genomes and 4 partial spike genomic regions) belonged to lineage B, bearing the D614G mutation in the Spike protein. Isolates from the first outbreak that occurred in the Margarita Island harbored an in-frame deletion in its sequence, without amino acids 83-85 of the NSP1 of the ORF1. The search for deletions in 48,635 sequences showed that the NSP1 gene exhibit the highest frequency of deletions along the whole genome. Structural analysis suggests a change in the N-terminal domain with the presence of this deletion. In contrast, isolates circulating later in this island lacked the deletion, suggesting new introductions to the island after this first outbreak. In conclusion, a high diversity of SARS-CoV-2 isolates were found circulating in Venezuela, with predominance of the D614G mutation. The first small outbreak in Margarita Island seemed to be associated with a strain carrying a small deletion in the NSP1 protein, but these isolates do not seem to be responsible for the larger outbreak which started in July.

Computational Modeling of Protease Inhibitors for the Development of Drugs Against Coronaviruses

In late 2019, SARS-CoV-2, a new coronavirus, emerged from Wuhan, China, and caused a world pandemic in a few months. There are no US Food and Drugs Administration (USFDA)-approved antiviral drugs or vaccines that could be used to prevent or treat this viral infection. However, several clinical trials are ongoing, searching for therapeutic alternatives. As time is crucial in a pandemic, the scientific community has used drug repurposing and data obtained from in silico models to identify possible lead compound inhibitors of SARS-CoV-2. Direct-acting antivirals are the most promising tools to control viral infections. One of the targets for direct-acting antivirals is the main protease (Mpro), a key enzyme in the SARS-CoV-2 replication cycle. This protease is a cysteine protease that shares high homology with the SARS-CoV Mpro and could be susceptible to blocking its activity by compounds such as HIV protease inhibitors. In this book chapter, we focused on the structural features of the SARS-CoV-2 Mpro that could be applied to developing new therapies using computational aid, although it is not certain whether the outcomes of such computational studies will immediately result in effective anti-SARS-CoV-2 therapy.

Importance of mutations in amino acid 484 of the Spike protein of SARS-CoV-2: rapid detection by restriction enzyme analysis

Variants of Concern of SARS-CoV-2 (VOCs), the new coronavirus responsible for COVID-19, have emerged in several countries. Mutations in the amino acid 484 of the Spike protein are particularly important and associated with some of these variants: E484K or E484Q. These mutations have been associated with evasion to neutralizing antibodies. Restriction enzyme analysis is proposed as a rapid method to detect these mutations. A search on GISAID was performed in April 2021 to detect the frequency of these two mutations in the sequence available and their association with other lineages. E484K, present in some VOCs, has emerged in several other lineages and is frequently found in recent viral isolates. A small amplicon from the Spike gene was digested with two enzymes: HpyAV, and MseI. The use of these two enzymes allows the detection of mutations at position 484, and to differentiate between these three conditions: non-mutated, and the presence of E484K or E484Q. A 100% correlation was observed with sequencing results. The proposed methodology, which allows for the screening of a great number of samples, will probably help to provide more information on the prevalence and epidemiology of these mutations worldwide, to select the candidates for whole-genome sequencing.

Production of equine sera as a potential immunotherapy against COVID-19

Emerging viruses such as the COVID-19-inducing virus, SARS-CoV-2, represent a threat to human health, unless effective vaccines, drugs or alternative treatments, such as passive immunization, become accessible. Animal-derived immunoglobulins, such as equine immunoglobulins might be useful as immunoprophylaxis or immunotherapy against this viral disease. Therapeutic antibodies (Abs) for SARS-CoV-2 were obtained from hyperimmune equine plasma using the Spike protein receptor binding domain (RBD) as an immunogen. The presence of anti-RBD antibodies was evaluated by ELISA and the titres of neutralizing antibodies were determined in viral cell culture. Immunized horses generated high-titre of anti-RBD antibodies with antiviral neutralizing activity on Vero-E6 cells of 1/1,000. To minimize potential adverse effects, the immunoglobulins were digested with pepsin, and purified to obtain the F(ab')2 fragments with the protocol standardized by Biotecfar C.A (R) for the production of snake antivenom. Pre-immune serum displayed an unexpected anti-RBD reactivity by ELISA (titre up to 1/900) and Western Blot, but no neutralizing activity. Modelling of the RBD of equine coronavirus showed that some of the known epitopes of SARS-CoV-2 RBD were structurally conserved in the equine coronavirus protein. This might suggest that some of the reactivity observed in the pre-immune serum to the SARS-CoV-2 RBD might be due to a previous exposure to equine coronavirus.

Biology and evolution of the coronavirus that causes COVID-19

The new coronavirus that causes COVID-19 is called SARS-CoV-2 and belongs to the subgenus Sarbecovirus, like its predecessor SARS-CoV. Bats appear to be the hosts of the ancestral viruses that originated these viruses, through recombination with the virus of an intermediate animal, which might be the pangolin. The virus interacts with the ACE2 receptor (angiotensin converting enzyme 2) and enters the cell by the endocytic pathway, through an early or late endosome. Viral RNA serves as messenger RNA for the translation of the first reading frame and the rest of the messenger RNAs are produced by discontinuous transcription. This peculiarity confers to this viral family a high frequency of recombination, which is associated to the high frequency of species jumping. Viruses belonging to the order Nidovirales are the only known RNA viruses with a polymerase with proof correction capacity;therefore their mutation rate is reduced. However, these genomes appear to be susceptible to be deaminated by cellular enzymes. All these mechanisms of generation of diversity leads to the existence of lineages, including those with the D614G mutation in the spike associated with a higher transmissibility. However, mutations associated with greater severity are not known to date. Just as there are different viral variants, the clinical manifestation of the disease is also highly variable. Some genetic, physiological and metabolic factors are being known that could be determining a more severe clinical presentations, often associated with the immunopathology of this disease.

SARS-CoV-2 host tropism: An in silico analysis of the main cellular factors.

Recent reports have shown that small and big felines could be infected by SARS-CoV-2, while other animals, like swines and mice, are apparently not susceptible to this infection. These findings raise the question of the role of cell factors associated with early stages of the viral infection in host selectivity. The cellular receptor for SARS-CoV-2 is the Angiotensin Converting Enzyme (ACE2). Transmembrane protease serine 2 (TMPRSS2) has been shown to prime the viral spike for its interaction with its receptor. GRP78 has also been proposed as a possible co-receptor. In this study, we used several bioinformatics approaches to bring clues in the interaction of ACE2, TMPRSS2, and GRP78 with SARS-CoV-2. We selected several mammalian hosts that could play a key role in viral spread by acting as secondary hosts (cats, dogs, pigs, mice, and ferrets) and evaluated their predicted permissiveness by in silico analysis. Results showed that ionic pairs (salt bridges, N-O pair, and long-range interactions) produced between ACE2 and the viral spike has an essential function in the host interaction. On the other hand, TMPRSS2 and GRP78 are proteins with high homology in all the evaluated hosts. Thus, these proteins do not seem to play a role in host selectivity, suggesting that other factors may play a role in the non-permissivity in some of these hosts. These proteins represent however interesting cell targets that could be explored in order to control the virus replication in humans and in the intermediary hosts.