Enhanced High Mutation Rate and Natural Selection to Produce Attenuated Viral Vaccine with CRISPR Toolkit in RNA Viruses especially SARS-CoV-2

Abstract

The best and most effective way to combat pandemics is to use effective vaccines and live attenuated vaccines are among the most effective vaccines. However, one of the major problems is the length of time it takes to get the attenuated vaccines. Today, the CRISPR toolkit (Clustered Regularly Inerspaced Short Palindromic Repeats) has made it possible to make changes with high efficiency and speed. Using this toolkit to make point mutations on the RNA virus's genome in a coculture of permissive and nonpermissive cells and under controlled conditions can accelerate changes in the genome and accelerate natural selection to obtain live attenuated vaccines.

Insertion-and-Deletion Mutations between the Genomes of SARS-CoV, SARS-CoV-2, and Bat Coronavirus RaTG13

“…The CRISPR technology was originally developed from the bacterial CRISPR-Cas9 antiviral immune system, which had already existed in almost all archaea and many bacteria in the natural environment ( 40 – 43 ). The CRISPR-Cas system is useful in RNA and DNA editing that realized relatively precise genome modifications, including an induction of indel to the DNA sequences ( 44 , 45 ). The CRISPR toolkit is further expected to shorten the time for developing an effective live attenuated vaccine for some viruses or realizing viral load reduction after viral infections ( 45 , 46 ).…”

“…The CRISPR-Cas system is useful in RNA and DNA editing that realized relatively precise genome modifications, including an induction of indel to the DNA sequences ( 44 , 45 ). The CRISPR toolkit is further expected to shorten the time for developing an effective live attenuated vaccine for some viruses or realizing viral load reduction after viral infections ( 45 , 46 ). However, whether the CRISPR-Cas13 system (which targets RNA and not DNA) can realize the long insertion-and-deletion mutations in RNA molecules that were observed in the present study remains unknown.…”

Principles and Applications of CRISPR Toolkit in Virus Manipulation, Diagnosis, and Virus-Host Interactions

“…The use of CRISPR interference (CRISPRi) in CD4 + T cells has also shown that ELL2 (elongation factor for RNA polymerase II 2) plays a key role in regulating HIV latency. Elevation in the level of ELL2 and ELL2-SECs (super elongation complex) following proteasome inhibition is indicative of the proteasome-ELL2 axis's important role in regulating HIV-1 latency, making it a promising target for therapeutic intervention [112]. In a study using the CRISPR system, 121 host genes associated with influenza A virus replication were identified, and WDR7 (WD repeat-containing protein 7, which is involved in gene regulation, signal transduction, cell cycle progression, and apoptosis), TMEM199 (transmembrane protein 199, with a role in protein localization in some cells), and CCDC115 (coiled-coil domain-containing protein 115, which is required for intracellular iron homeostasis in aerobic conditions and is a component of the proton-transporting vacuolar) host factors were shown to be essential for the entry and functioning of the virus.…”

Research and Therapeutic Approaches in Stem Cell Genome Editing by CRISPR Toolkit

“…Some different stem cell types have so far been successfully used in clinical studies and have received scientific approval. Besides viral genome modification, more and more publications are confirming that CRISPR/Cas9 genome editing is a potent technique that can greatly advance biomedicine, such as in virus genome editing and in stem cell research [ 11 , 17 , 18 , 19 ]. Organoids are interesting new systems that help us improve our knowledge of diseases’ mechanisms, development, evolution, homeostasis, and therapy.…”