CRISPR/Cas9-modified Human T Cell ( PD-1and ACE2 Knockout Engineered T Cells ) for Inducing Long-term Immunity in COVID-19 Patients

T-cell exhaustion may limit long-term immunity in COVID-19 patients. T cells can losetheir ability to fight viruses and tumors when they have prolonged exposure to theseenemies. New data suggests people who experience mild COVID-19 symptoms show themolecular signs of exhausted memory T cells and therefore could have a reduced ability tofight reinfection. On contrary people who develop severe COVID-19 symptoms may be betterprotected from reinfection. A recent study reported that the 82.1% of COVID-19 casesdisplayed low circulating lymphocyte counts . It has been reported that, in the case ofchronic viruses, continuous PD-1 expression causes T-cell exhaustion, and impairs theability of killing the infectious cells . The adumbration of patients with COVID-19 ischaracterized by a diminished lymphocyte percentage, with a similar proportion of CD4+and CD8+ T-cells. The quantity of T-cells, mostly CD8+ T-cells, presenting highexpression rates of late activity marker CD25 and exhaustion marker PD-1 increases.Therefore, SARS-CoV-2 is able to make changes by modifying the acquired immune system,including B and T cells. According to experiments, PD-1's expression, as an importantfactor in the induction and maintenance of circumferential tolerance keeping thestability of T-cells, has been found to have a higher percentage in different cells ofCOVID-19 patients. In an experiment conducted by Diao et al., on the patients withSARS-CoV-2, it was observed that the expression of PD-1 on the surface of T-cells wasincreased significantly; it was also shown that during the SARS-CoV-2 -induced disease,additional expressions of PD-1 and Tim-3 on the T-cells were directly related to thedisease's severity; the factors that were also increased in other viral infections. Tcell exhaustion" phenomenon could be reversed relatively easily, for example when the Tcells are no longer exposed to the virus or tumor. But unfortunately, although exhaustedT cells recovered from chronic infection (REC-TEX) regain some function and features ofmemory T cells (TMEM), they retain epigenetic scars indicating the control of geneexpression is "locked in" to their exhaustion history. Once T cells become exhausted,they remain fundamentally 'wired' to be exhausted-thus it may be hard to get them tobecome effective virus- and cancer-fighters again," said John Wherry, PhD, chair of thedepartment of Systems Pharmacology and Translational Therapeutics and director of thePenn Institute of Immunology in the Perelman School of Medicine at the University ofPennsylvania. Furthermore, COVID-19 may infect T lymphocyte cells and induce apoptosisand apoptotic markers. Lymphocytopenia was also found in the Middle East respiratorysyndrome (MERS) cases. MERS-CoV can directly infect human primary T lymphocytes andinduce T-cell apoptosis through extrinsic and intrinsic apoptosis pathways, but it cannotreplicate in T lymphocytes. However, it is unclear whether SARS-CoV-2 can also infect Tcells, resulting in lymphocytopenia. A study showed that T cells express a very lowexpression level of hACE2 on its cell surface and T-cell lines were significantly moresensitive to SARS-CoV-2 infection when compared with SARS-CoV . In other words, theseresults tell us that T lymphocytes may be more permissive to SARS-CoV-2 infection.Therefore, it is plausible that the S protein of SARS-CoV-2 might mediate potentinfectivity, even on cells expressing low hACE2, which would, in turn, explain why thetransmission rate of SARS-CoV-2 is so high. Through recent advances in genomic editing, Tcells can now be successfully modified via CRISPR/Cas9 technology. For instance, engaging(post-)transcriptional mechanisms to enhance T cell cytokine production, the retargetingof T cell antigen specificity or rendering T cells refractive to inhibitory receptorsignaling can augment T cell effector function. Therefore, CRISPR/Cas9-mediated genomeediting might provide novel strategies for inducing long term immunity againstCOVID-19.Immunotherapies with autologous T cells have become a powerful treatment optionfor many diseases like viral infection or cancer. These include the adoptive isolationand transfer of naturally-occurring virus/tumor-specific T cells and the transfer of Tlymphocytes that have been genetically modified . According to the investigator,exhausted virus-reactive CD8+ memory T cells will be isolated from patients with mildinfection using a modified antigen-reactive T cell enrichment (ARTE) assay. exhaustedvirus-reactive CD8+ memory T cells will be collected and both Programmed cell deathprotein 1(PDCD1) gene and ACE2 gene will be knocked out by CRISPR Cas9 in the laboratory.The lymphocytes will be selected and expanded ex vivo and infused back into patients.