Construction Of PR1-HLA-A~*0201Single Chain Trimer Lentiviral Vector And Preliminary Research

Lentiviral vector developed based on human immunodeficiency virus type I (HIV-1). Gene transfer vectors based on lentiviruses provide effective means for the delivery, integration and resulting in long-term expression of exogenous genes in neuronal cells, liver cells, cardiomyocytes, endothelial cells, stem cells and other types of mammalian cells.It is attractive gene delivery vehicles in the context of non-dividing cells.Besides, lentiviral vector holds the characteristics of larger capacity of transfer gene fragments, lower rate of immunological response therefore it becomes a more powerful tool in the cellular level and overall level of transgenic research.Cellular immunity play a major role in the tumor immunity. CD8+T lymphocytes recognize tumor antigens by human leukocyte antigen class I molecules (HLA-I). Firstly, T cell antigen receptor recognition of antigen presenting cells or tumor cells of the antigen peptide presented by HLA-1molecule, and it’s the first signal of actovation. Then costimulatory molecules by ligand binding receptor and it’s the second signal. But tumor adoptive immunotherapy limiting its use in clinical by autologous activated T cell antigen presenting cells. therefore the artificial antigen presenting cell research has become increasingly activited in recent years, because of the advantages of the activation conditions easy to control, short cycle of activation and proliferation T cell.The objective of our study the selection of MHC-I molecule is HLA-A*0201. PR1(VLQELNVTV), which is an HLA-A*0201restricted antigenie peptide derived from myeloid leukemia associated antigen protease3, can prime specific CTLs responses against myeloid leukemia cells. The antigenic peptide, β2m, and heavy chain are attached by flexible linkers with genetic engineering, and cloned it into the gene transfer vector,and Sequencing it.Then we use three plasmids co-transfect the293T cells, after48h harvest the supernatant which contains the virus particles.After that,infected293T cells with recombinant lentivirus,respectively, the viral titer was checked by observing the expression of green fluorescent protein (GFP) in293T cells,and the titer is1×104TU/mL. This study lay a foundation for further provides of dual-signal artificial antigen presenting cells.