Abstract
BackgroundThe gene transduction efficiency of adenovirus to hematopoietic cells, especially T lymphocytes, is needed to be improved. The purpose of this study is to improve the transduction efficiency of T lymphocytes by using fiber-modified human adenovirus 5 (HAdV-5) vectors.ResultsFour fiber-modified human adenovirus 5 (HAdV-5) vectors were investigated to transduce hematopoietic cells. F35-EG or F11p-EG were HAdV-35 or HAdV-11p fiber pseudotyped HAdV-5, and HR-EG or CR-EG vectors were generated by incorporating RGD motif to the HI loop or to the C-terminus of F11p-EG fiber. All vectors could transduce more than 90% of K562 or Jurkat cells at an multiplicity of infection (MOI) of 500 viral particle per cell (vp/cell). All vectors except HR-EG could transduce nearly 90% cord blood CD34+ cells or 80% primary human T cells at the MOI of 1000, and F11p-EG showed slight superiority to F35-EG and CR-EG. Adenoviral vectors transduced CD4+ T cells a little more efficiently than they did to CD8+ T cells. These vectors showed no cytotoxicity at an MOI as high as 1000 vp/cell because the infected and uninfected T cells retained the same CD4/CD8 ratio and cell growth rate.ConclusionsHAdV-11p fiber pseudotyped HAdV-5 could effectively transduce human T cells when human EF1a promoter was used to control the expression of transgene, suggesting its possible application in T cell immunocellular therapy.
Highlights
The gene transduction efficiency of adenovirus to hematopoietic cells, especially T lymphocytes, is needed to be improved
Construction of adenoviral vectors We constructed 5 first-generation adenovectors, in which the E1/E3 regions were deleted and GFP expression cassette including the human EF1a promoter, GFP coding sequence and SV40 polyA signal were inserted into the E1 region
The five vectors were the same except the fiber protein: Human adenovirus 5 (HAdV5)-EG contained the original fiber of Human adenoviruses (HAdV)-5 and served as a control vector; HAdV5F35-EG contained a chimeric fiber of human adenovirus 5 (HAdV-5) tail and HAdV-35 shaft and knob; HAdV5F11p-EG contained a chimeric fiber of HAdV-5 tail and HAdV-11p shaft and knob; HAdV5F11pHR-EG was different from HAdV5F11p-EG in that RGD4C peptide was inserted into the HI loop of the knob domain; and HAdV5F11pCR-EG was different from HAdV5F11p-EG in that RGD4C peptide was fused to the C-terminal of the fiber with a [GGGGS]3 linker (Fig. 1)
Summary
The gene transduction efficiency of adenovirus to hematopoietic cells, especially T lymphocytes, is needed to be improved. The purpose of this study is to improve the transduction efficiency of T lymphocytes by using fiber-modified human adenovirus 5 (HAdV-5) vectors. T lymphocytes play an important role in adaptive immunity. T cells are important targets for gene therapy of numerous human diseases, including cancer, diabetes, arthritis and AIDS [2,3,4,5]. Gene delivery to T cells has been achieved by retroviruses including gamma-retrovirus, lentivirus and alpha-retrovirus [2, 6]. Retrovirus is able to integrate into the host’s genome, and the transgene stably expresses.
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