Research On The Immune Mechanism Of Specific TCR-T Cells Response To HCMV Infection

Objective:Human cytomegalovirus(HCMV)infection is a leading cause of morbidity and mortality in immunocompromised patients,but no specific therapeutic strategy is effective clinically,despite recent achievements.HCMV-specific T-cell therapy was thought to be helpful for the management of HCMV infection.To conduct a deep exploration of the immune response mechanism of HCMV-specific T cells and make preparation for the future application,the possibility of engineering peripheral blood mononuclear cells(PBMCs)from immunocompetent and immunocompromised subjects with specific T-cell receptor(TCR)genes was investigated.Methods:In this study,PBMCs were collected from HCMV-infected immunocompetent subjects,and from hematopoietic stem cell transplant(HSCT)recipients in the third month post-HSCT(Month-3 PBMCs)and in the sixth month post-HSCT(Month-6 PBMCs).One HCMV-specific TCR sequence,CASSSANYGYFT,was selected as the research target.Lentivirus vectors containing the specific TCR genes and mock TCR genes were constructed and applied to infect the PBMCs.PBMCs transduced with TCR lentiviral vectors were marked as TCR-V group,PBMCs transduced with control mock lentiviral vectors were marked as TCRm-V group.Pentamer staining and IFN-y Enzyme-linked immunospot assay were applied to evaluate the quantity and specific function of the TCR-engineered T cells.Results:CD8-positive T cells that specifically bind to peptide-pentamers(CD8+/PEN+T cells)could be generated by transferring TCR genes to immunocompetent PBMCs.Besides,TCR-transduced PBMCs produced considerable amounts of IFN-y-secreting T cells when challenged with HCMV peptide(Compared with TCRm-V group,p<0.001);Compared with mock transduction,transfer of TCR genes into month-3 PBMCs increased the percentage of CD8+/PEN+T cells and significantly enhanced the frequency of IFN-y-secreting T cells(p<0.05).For month-6 PBMCs,CD8+/PEN+T cells were more abundant in the TCR-V groups than in the TCRm-V groups,whereas no difference was observed in the numbers of IFN-y-secreting T cells between the TCR-V and TCRm-V groups(p>0.05).Conclusion:TCR gene transfer is a potential therapeutic strategy for controlling HCMV infection in HSCT recipients.HCMV-specific TCR-T cells would be more meaningful in the prevention and treatment of HCMV infection in the circumstance of incomplete immune reconstitution post-HSCT.