title: Selective modification of antigen-specific T cells by RNA electroporation.
abstract: It has been observed that the efficient transfection of T cells by RNA electroporation requires prior activation of T cells with mitogens or by anti-CD3 antibody stimulation. We hypothesized that this requirement for T cell activation could be leveraged to express marker genes within activated T cells responding to antigen-pulsed dendritic cells and allow for the selective enrichment and modification of antigen-specific T cells. Using electroporation of mRNA encoding green fluorescent protein as a marker gene, we demonstrate that RNA electroporation can efficiently allow for the separation of cytomegalovirus-specific CD8+ and CD4+ T cells from bulk culture responding to cytomegalovirus pp65 antigen-pulsed dendritic cells. Furthermore, we demonstrate that cytomegalovirus-specific T cells can be functionally modified by RNA transfection of the C-X-C chemokine receptor, CXCR2, to migrate efficiently toward a variety of CXCR2-specific chemokines in vitro and in vivo. These studies demonstrate the utility of RNA transfection as a simple method by which to purify and selectively modify the function of antigen-specific T cells for use in adoptive immunotherapy, and importantly provide evidence that transient expression of proteins by RNA transfection is an efficient means of modulating the in vivo function of activated T cells.
authors.affiliation: Division of Neurosurgery, Department of Surgery, Preston Robert Tisch Brain Tumor Center at Duke, Duke University Medical Center, Durham, NC 27710, USA. d.mitchell@duke.edu
authors.forename: Duane A
authors.initials: DA
authors.lastname: Mitchell
authors.forename: Isaac
authors.initials: I
authors.lastname: Karikari
authors.forename: Xiuyu
authors.initials: X
authors.lastname: Cui
authors.forename: Weihua
authors.initials: W
authors.lastname: Xie
authors.forename: Robert
authors.initials: R
authors.lastname: Schmittling
authors.forename: John H
authors.initials: JH
authors.lastname: Sampson
mesh_terms: D000941:Antigens / Q000032:analysis / Q000235:genetics / Q000276:immunology*; D001932:Brain Neoplasms / Q000276:immunology / Q000628:therapy; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology / Q000637:transplantation; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology / Q000637:transplantation; D002465:Cell Movement; D002478:Cells, Cultured; D018925:Chemokines / Q000378:metabolism; D003713:Dendritic Cells / Q000187:drug effects / Q000276:immunology; D018274:Electroporation; D005910:Glioma / Q000276:immunology / Q000628:therapy; D049452:Green Fluorescent Proteins / Q000235:genetics; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D008213:Lymphocyte Activation; D010455:Peptides / Q000235:genetics / Q000276:immunology; D010750:Phosphoproteins / Q000276:immunology / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics*; D023063:Receptors, Interleukin-8B / Q000235:genetics*; D013601:T-Lymphocytes / Q000276:immunology* / Q000637:transplantation; D014162:Transfection; D014763:Viral Matrix Proteins / Q000276:immunology / Q000494:pharmacology
chemical_list: D000941:Antigens; D018925:Chemokines; D010455:Peptides; D010750:Phosphoproteins; D012333:RNA, Messenger; D023063:Receptors, Interleukin-8B; D014763:Viral Matrix Proteins; C065418:cytomegalovirus matrix protein 65kDa; D049452:Green Fluorescent Proteins