Heine Annkristin, Grünebach Frank, Holderried Tobias, Appel Silke, Weck Markus M, Dörfel Daniela, Sinzger Christian, Brossart Peter
Department of Hematology, Oncology, and Immunology, University of Tübingen, Germany.
Mol Ther. 2006 Feb;13(2):280-8. doi: 10.1016/j.ymthe.2005.08.019. Epub 2005 Oct 10.
Transfection of dendritic cells (DCs) with RNA was shown to be effective in the generation of antigen-specific T cells, probably due to the induction of a polyclonal T cell response directed against multiple antigens. To verify this assumption we used DCs, generated from cytomegalovirus (CMV)-negative or -positive donors, that were electroporated with in vitro-transcribed RNA (in vitro transcript, IVT) coding for the CMV pp65 antigen. We found that transfection of DCs with pp65 IVT induces an expansion of polyclonal CD8(+) T lymphocytes that recognize peptide antigens presented on different HLA molecules. These T lymphocytes are able to lyse DCs pulsed with pp65-derived peptides or transfected with the cognate IVT. Furthermore, this approach allowed the identification of immunodominant epitopes presented upon IVT transfection. Interestingly, transfection of DCs with pp65 IVT resulted in the induction of CD4(+)-specific T cells. Cotransfection of DCs with IVTs coding for the CMV antigens pp65 and IE1 elicited polyclonal T lymphocytes specific for peptides derived from both antigens. More importantly, cytotoxic T cells could be generated in two of three CMV-negative donors. Finally, functional CMV-specific autologous cytotoxic T lymphocytes were successfully generated from immunosuppressed patients after allogeneic hematopoietic stem cell transplantation.
用RNA转染树突状细胞（DCs）已被证明在产生抗原特异性T细胞方面是有效的，这可能是由于诱导了针对多种抗原的多克隆T细胞反应。为了验证这一假设，我们使用了从巨细胞病毒（CMV）阴性或阳性供体产生的DCs，并用编码CMV pp65抗原的体外转录RNA（体外转录本，IVT）进行电穿孔。我们发现用pp65 IVT转染DCs可诱导识别不同HLA分子上呈递的肽抗原的多克隆CD8(+) T淋巴细胞扩增。这些T淋巴细胞能够裂解用pp65衍生肽脉冲处理或用同源IVT转染的DCs。此外，这种方法还能鉴定IVT转染后呈递的免疫显性表位。有趣的是，用pp65 IVT转染DCs可诱导产生CD4(+)特异性T细胞。用编码CMV抗原pp65和IE1的IVT共转染DCs可引发针对源自这两种抗原的肽的多克隆T淋巴细胞。更重要的是，在三个CMV阴性供体中的两个中可以产生细胞毒性T细胞。最后，在异基因造血干细胞移植后，成功地从免疫抑制患者中产生了功能性CMV特异性自体细胞毒性T淋巴细胞。
