title: Dendritic cell function after gene transfer with adenovirus-calcium phosphate co-precipitates.
abstract: Dendritic cells (DCs) are essential for initiating and directing antigen-specific T-cell responses. Genetic modification of DC is under study for cancer immunotherapy, vaccine development, and antigen-targeted immunosuppression. Adenovirus (Ad) type 5 (Ad5)-mediated gene transfer to mouse bone marrow DCs and human monocyte-derived DCs is inefficient because neither express the cognate high-affinity Ads receptor. We show that co-precipitating adenoviral vectors with calcium phosphate (CaPi) increased gene expression (2000-fold) and transduction efficiency (50-fold) in mouse DC, primarily owing to receptor-independent viral uptake. Moreover, Ad5:CaPi-treated DCs were activated to express the maturation surface molecules CD40 and CD86, and to secrete proinflammatory cytokines tumor necrosis factor-alpha and interleukin 6. However, neither DC transduction nor maturation was dependent on viral protein interactions with cell surface integrin. Ad5:CaPi also transduced human DC more efficiently than Ad5 alone, similar to a genetically modified vector (Ad5f35) targeted to the CD46 receptor. As such, this approach combines the efficiency of adenoviral-mediated endosomal escape and nuclear trafficking with the receptor independence of nonviral gene delivery. Importantly, CaPi co-precipitation could be used to functionally modify DC to activate and expand cytomegalovirus-specific memory cytotoxic T lymphocytes. This study identifies a simple technique to improve the efficacy of current Ad5 gene transfer, in support of clinical adoptive immunotherapy.
authors.affiliation: Interdepartmental Program in Cell and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Seiler
authors.forename: Stephen
authors.initials: S
authors.lastname: Gottschalk
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Cerullo
authors.forename: Maheshika
authors.initials: M
authors.lastname: Ratnayake
authors.forename: Viraj P
authors.initials: VP
authors.lastname: Mane
authors.forename: Christian
authors.initials: C
authors.lastname: Clarke
authors.forename: Donna J
authors.initials: DJ
authors.lastname: Palmer
authors.forename: Philip
authors.initials: P
authors.lastname: Ng
authors.forename: Cliona M
authors.initials: CM
authors.lastname: Rooney
authors.forename: Brendan
authors.initials: B
authors.lastname: Lee
mesh_terms: D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology / Q000378:metabolism; D002130:Calcium Phosphates / Q000494:pharmacology*; D036022:Capsid Proteins / Q000235:genetics / Q000378:metabolism; D003713:Dendritic Cells / Q000187:drug effects / Q000378:metabolism*; D005260:Female; D005434:Flow Cytometry; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D016023:Integrins / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D011485:Protein Binding; D013601:T-Lymphocytes / Q000187:drug effects / Q000378:metabolism; D014162:Transfection / Q000379:methods*
chemical_list: D002130:Calcium Phosphates; D036022:Capsid Proteins; D016023:Integrins; C020243:calcium phosphate