SARS-CoV-2 Omicron Mutation Is Faster than the Chase: Multiple Mutations on Spike/ACE2 Interaction Residues

Recently, a new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (B.1.1.529) Omicron variant originated from South Africa in the middle of November 2021. SARS-CoV-2 is also called coronavirus disease 2019 (COVID-19) since SARS-CoV-2 is the causative agent of COVID-19. Several studies already suggested that the SARS-CoV-2 Omicron variant would be the fastest transmissible variant compared to the previous 10 SARS-CoV-2 variants of concern, interest, and alert. Few clinical studies reported the high transmissibility of the Omicron variant but there is insufficient time to perform actual experiments to prove it, since the spread is so fast. We analyzed the SARS-CoV-2 Omicron variant, which revealed a very high rate of mutation at amino acid residues that interact with angiostatin-converting enzyme 2. The mutation rate of COVID-19 is faster than what we prepared vaccine program, antibody therapy, lockdown, and quarantine against COVID-19 so far. Thus, it is necessary to find better strategies to overcome the current crisis of COVID-19 pandemic.

Structure of SARS-CoV-2 Spike Glycoprotein for Therapeutic and Preventive Target

The global crisis caused by the coronavirus disease 2019 (COVID-19) led to the most significant economic loss and human deaths after World War II. The pathogen causing this disease is a novel virus called the severe acute respiratory syndrome coronavirus 2 (SARSCoV-2). As of December 2020, there have been 80.2 million confirmed patients, and the mortality rate is known as 2.16% globally. A strategy to protect a host from SARS-CoV-2 is by suppressing intracellular viral replication or preventing viral entry. We focused on the spike glycoprotein that is responsible for the entry of SARS-CoV-2 into the host cell. Recently, the US Food and Drug Administration/EU Medicines Agency authorized a vaccine and antibody to treat COVID-19 patients by emergency use approval in the absence of long-term clinical trials. Both commercial and academic efforts to develop preventive and therapeutic agents continue all over the world. In this review, we present a perspective on current reports about the spike glycoprotein of SARS-CoV-2 as a therapeutic target.

The Progression of SARS Coronavirus 2 (SARS-CoV2): Mutation in the Receptor Binding Domain of Spike Gene

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) is a positive-sense singlestranded RNA (+ssRNA) that causes coronavirus disease 2019 (COVID-19). The viral genome encodes twelve genes for viral replication and infection. The third open reading frame is the spike (S) gene that encodes for the spike glycoprotein interacting with specific cell surface receptor – angiotensin converting enzyme 2 (ACE2) – on the host cell membrane. Most recent studies identified a single point mutation in S gene. A single point mutation in S gene leading to an amino acid substitution at codon 614 from an aspartic acid 614 into glycine (D614G) resulted in greater infectivity compared to the wild type SARS-CoV2. We were interested in investigating the mutation region of S gene of SARS-CoV2 from Korean COVID-19 patients. New mutation sites were found in the critical receptor binding domain (RBD) of S gene, which is adjacent to the aforementioned D614G mutation residue. This specific sequence data demonstrated the active progression of SARS-CoV2 by mutations in the RBD of S gene.The sequence information of new mutations is critical to the development of recombinant SARS-CoV2 spike antigens, which may be required to improve and advance the strategy against a wide range of possible SARS-CoV2 mutations.

The Progression of SARS Coronavirus 2 (SARS-CoV2): Mutation in the Receptor Binding Domain of Spike Gene

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV2) is a positive-sense singlestranded RNA (+ssRNA) that causes coronavirus disease 2019 (COVID-19). The viral genome encodes twelve genes for viral replication and infection. The third open reading frame is the spike (S) gene that encodes for the spike glycoprotein interacting with specific cell surface receptor – angiotensin converting enzyme 2 (ACE2) – on the host cell membrane. Most recent studies identified a single point mutation in S gene. A single point mutation in S gene leading to an amino acid substitution at codon 614 from an aspartic acid 614 into glycine (D614G) resulted in greater infectivity compared to the wild type SARS-CoV2. We were interested in investigating the mutation region of S gene of SARS-CoV2 from Korean COVID-19 patients. New mutation sites were found in the critical receptor binding domain (RBD) of S gene, which is adjacent to the aforementioned D614G mutation residue. This specific sequence data demonstrated the active progression of SARS-CoV2 by mutations in the RBD of S gene.The sequence information of new mutations is critical to the development of recombinant SARS-CoV2 spike antigens, which may be required to improve and advance the strategy against a wide range of possible SARS-CoV2 mutations.

in vivo multiplex gene targeting with Streptococcus pyogens and Campylobacter jejuni Cas9 for pancreatic cancer modeling in wildtype animal

Pancreatic ductal adenocarcinoma is a lethal cancer type that is associated with multiple gene mutations in somatic cells. Genetically engineered mouse is hardly applicable for developing a pancreatic cancer model, and the xenograft model poses a limitation in the reflection of early stage pancreatic cancer. Thus, in vivo somatic cell gene engineering with clustered regularly interspaced short palindromic repeats is drawing increasing attention for generating an animal model of pancreatic cancer. In this study, we selectedKras, Trp53, Ink4a, Smad4, and Brca2 as target genes, and applied Campylobacter jejuni Cas9 (CjCas9) andStreptococcus pyogens Cas9 (SpCas9) for developing pancreatic cancer using adeno associated virus (AAV) transduction. After confirming multifocal and diffuse transduction of AAV2, we generated SpCas9 overexpression mice, which exhibited high double-strand DNA breakage (DSB) in target genes and pancreatic intraepithelial neoplasia (PanIN) lesions with two AAV transductions; however, wild-type (WT) mice with three AAV transductions did not develop PanIN. Furthermore, small-sized Cjcas9 was applied to WT mice with two AAV system, which, in addition, developed high extensive DSB and PanIN lesions. Histological changes and expression of cancer markers such as Ki67, cytokeratin, Mucin5a, alpha smooth muscle actin in duct and islet cells were observed. In addition, the study revealed several findings such as 1) multiple DSB potential of AAV-CjCas9, 2) peri-ductal lymphocyte infiltration, 3) multi-focal cancer marker expression, and 4) requirement of > 12 months for initiation of PanIN in AAV mediated targeting. In this study, we present a useful tool for in vivo cancer modeling that would be applicable for other disease models as well.

Establishment and identification of cell lines from type O blood Korean native pigs and their efficiency in supporting embryonic development via somatic cell nuclear transfer

Due to their similarities with humans in anatomy, physiology, and genetics miniature pigs are becoming an attractive model for biomedical research. We aim to establish and evaluate blood type O cells derived from Korean native pig (KNP), a typical miniature pig breed in Korea. Ten cell lines derived from 8 KNP piglets and one adult female KNP (kidney and ear tissues) were established. To confirm the presence of blood type O, genomic DNA, fucosyltransferase (FUT) expression, and immunofluorescence staining were examined. Additionally, fluorescence-activated cell sorting and somatic cell nuclear transfer were performed to investigate the normality of the cell lines and to evaluate their effectiveness in embryo development. We found no significant bands corresponding to specific blood group A, and no increase in FUT expression in cell lines derived from piglets No. 1, No. 4, No. 5, No. 8, and the adult female KNP; moreover, they showed normal levels of expression of α 1,3-galactosyltransferase and cytidine monophosphate-N-acetylneuraminic acid hydroxylase. There was no significant difference in embryo development between skin and kidney fibroblasts derived from the blood type O KNPs. In conclusion, we successfully established blood type O KNP cell lines, which may serve as a useful model in xenotransplantation research.

Proteomic analysis of pancreas in miniature pigs according to developmental stages using two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry / 한국실험동물학회지

Organ transplantation is limited by the shortage of human organs. Many studies have sought to overcome this hurdle by using animal organs. Porcine organs, especially from miniature pigs, have been used for organ xenotransplantation rather than nonhuman primates. While the molecular profiling for transplantation is well known in humans and rodents, the situation for pigs is almost completely unknown. The present study examined protein regulation of the developing stages of the pancreatic proteome (4 day-old miniature neonate, 19 day-old miniature piglet, and 14 month-old miniature adult pigs) using two-dimensional gel electrophoresis and matrix assisted laser desorption/ionization-time of flight mass spectrometry. Thirteen different expressed spots were observed and nine were identified. The data presented within this study provides critical direction relating to the development of pancreas of miniature pigs, which will assist future proteome analysis of the pancreas, and advance our understanding of the hurdles facing xenotransplantation.

Proteomic analysis of liver in miniature pigs according to developmental stages using two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry / 한국실험동물학회지

Due to the shortage of human organ donors for transplant, various studies of xenotransplantation, or the use of animal organs instead of human organs, have been carried out. The organs of porcine are thought to be safer and of a more suitable size for xenotransplantationthan those of nonhuman primates. Understanding the levels of expression of proteins, and their post-translational regulation, would be very practical between different species and among developing stages, though the molecular profiling for xenotransplantation has been rarely studied for porcine, while that of human and rodent is well known. Here, in this present study, we report protein regulation of the developing stages of liver (4-day old neonate, 19-day old piglet and 14-month old adult miniature pigs) using 2-DE and MALDI-TOF. From images of the three different stages, a total of 8 spotswhich were differently regulated were identified, and 5 spots were identified with MALDI-TOF MS. The data presented within this study provides critical direction relating to the development of livers of miniature pigs, which will assist future proteome analysis of the liver, and advance our understanding of the hurdles facing xenotransplantaion.

Analysis of reference interval and age-related changes in serum biochemistry and hematology in the specific pathogen free miniature pig / 한국실험동물학회지

We analyzed 72 items related to serum biochemistry and hematology in 85 specific pathogen-free (SPF) Seoul National University (SNU) miniature pigs aged 1- to 36-months which originated from a Minnesota miniature pig. Almost all examined items were similar between male and female pigs. However, some items such as Cr level, B/C ratio, C.R.F, LDH, LAP and T4 were significantly different between male and female pigs (P<0.05). Thirty four examined items showed age-related changes, and the significant changes were observed in animals less than six months old. The values for BUN, K, uric acid, Ca, Ca++, and Pi were significantly higher in pigs younger than six months of age, which might reflect poor kidney function in young pigs. Additionally, TIBC, UIBC and RDW were significantly higher in young pigs, and RBC, Hb, HCT, MCHC and MCV were significantly lower in young pigs, thus indicating a similar physiology of iron deficiency anemia. These age-related specific phenotypes seemed to be normal, but it should be considered in the long-term experiment using the young pigs. In conclusion, in this study, we defined the normal reference intervals for SPF SNU miniature pigs, and we also determined that there are some physiological differences between the pig genders and ages. This study provides fundamental data for use in experiments involving SPF SNU miniature pigs.