title: Delivery of tumor-derived RNA for the induction of cytotoxic T-lymphocytes.
abstract: Dendritic cells (DC) are professional antigen-presenting cells playing a central role in the induction of antigen-specific cytotoxic T-lymphocytes (CTL). We analyzed the efficiency of tumor RNA transfection into DC using different sources of RNA as well as delivery strategies including electroporation, lipofection and CD71-receptor-based delivery. To evaluate the sensitivity of these approaches, we utilized in vitro transcribed enhanced green fluorescence protein (EGFP)-RNA and whole tumor RNA from EGFP-transfected renal cell carcinoma cell line N43. We demonstrate that electroporation was the most effective way yielding about 30% EGFP positive cells while less than 1% of DC expressed EGFP using the transferrin receptor transfection system. Delivery of RNA with liposomes resulted in 17.5% of EGFP positive cells depending on the RNA amount. However, when these approaches were applied to transduce DC with RNA derived from the A498 cell line for T-cell priming, tumor-specific CTL could be induced using all delivery strategies suggesting that this technology has the potential to induce cytotoxic T-cell response even when low level of antigen is delivered. Furthermore, we demonstrate that amplification of whole tumor messenger RNA (mRNA) as well as the use of total instead of purified mRNA can be utilized for stimulating tumor-specific CTL responses.
authors.affiliation: Department of Hematology, Oncology and Immunology, University of Tübingen, Tübingen, Germany.
authors.forename: F
authors.initials: F
authors.lastname: Grünebach
authors.forename: M R
authors.initials: MR
authors.lastname: Müller
authors.forename: A
authors.initials: A
authors.lastname: Nencioni
authors.forename: P
authors.initials: P
authors.lastname: Brossart
mesh_terms: D002292:Carcinoma, Renal Cell; D003713:Dendritic Cells / Q000276:immunology; D015870:Gene Expression; D015316:Genetic Therapy / Q000379:methods*; D049452:Green Fluorescent Proteins; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D007680:Kidney Neoplasms; D008164:Luminescent Proteins / Q000235:genetics; D008213:Lymphocyte Activation; D012334:RNA, Neoplasm / Q000235:genetics*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014162:Transfection / Q000379:methods*; D014407:Tumor Cells, Cultured
chemical_list: D008164:Luminescent Proteins; D012334:RNA, Neoplasm; D049452:Green Fluorescent Proteins