title: Gene therapy in HIV-infected cells to decrease viral impact by using an alternative delivery method.
abstract: The ability of dendrimer 2G-[Si{O(CH(2))(2)N(Me)(2) (+)(CH(2))(2)NMe(3) (+)(I(-))(2)}](8) (NN16) to transfect a wide range of cell types, as well as the possible biomedical application in direct or indirect inhibition of HIV replication, was investigated. Cells implicated in HIV infection such as primary peripheral blood mononuclear cells (PBMC) and immortalized suspension cells (lymphocytes), primary macrophages and dendritic cells, and immortalized adherent cells (astrocytes and trophoblasts) were analyzed. Dendrimer toxicity was evaluated by mitochondrial activity, cell membrane rupture, release of lactate dehydrogenase, erythrocyte hemolysis, and the effect on global gene expression profiles using whole-genome human microarrays. Cellular uptake of genetic material was determined using flow cytometry and confocal microscopy. Transfection efficiency and gene knockdown was investigated using dendrimer-delivered antisense oligonucleotides and small interfering RNA (siRNA). Very little cytotoxicity was detected in a variety of cells relevant to HIV infection and erythrocytes after NN16 dendrimer treatment. Imaging of cellular uptake showed high transfection efficiency of genetic material in all cells tested. Interestingly, NN16 further enhanced the reduction of HIV protein 24 antigen release by antisense oligonucleotides due to improved transfection efficiency. Finally, the dendrimer complexed with siRNA exhibited therapeutic potential by specifically inhibiting cyclooxygenase-2 gene expression in HIV-infected nervous system cells. NN16 dendrimers demonstrated the ability to transfect genetic material into a vast array of cells relevant to HIV pathology, combining high efficacy with low toxicity. These results suggest that NN16 dendrimers have the potential to be used as a versatile non-viral vector for gene therapy against HIV infection.
authors.affiliation: Laboratory of Molecular Immunobiology, Plataforma de Laboratorio, Hospital General Universitario Gregorio Marañón, 28007 Madrid, Spain.
authors.forename: Teresa
authors.initials: T
authors.lastname: Gonzalo
authors.forename: Maria Isabel
authors.initials: MI
authors.lastname: Clemente
authors.forename: Louis
authors.initials: L
authors.lastname: Chonco
authors.forename: Nick D
authors.initials: ND
authors.lastname: Weber
authors.forename: Laura
authors.initials: L
authors.lastname: Díaz
authors.forename: María Jesús
authors.initials: MJ
authors.lastname: Serramía
authors.forename: Rafael
authors.initials: R
authors.lastname: Gras
authors.forename: Paula
authors.initials: P
authors.lastname: Ortega
authors.forename: F Javier
authors.initials: FJ
authors.lastname: de la Mata
authors.forename: Rafael
authors.initials: R
authors.lastname: Gómez
authors.forename: Luis A
authors.initials: LA
authors.lastname: Lopez-Fernández
authors.forename: Maria Angeles
authors.initials: MA
authors.lastname: Muñoz-Fernández
authors.forename: José Luís
authors.initials: JL
authors.lastname: Jiménez
mesh_terms: D001253:Astrocytes / Q000378:metabolism; D045744:Cell Line, Tumor; D050091:Dendrimers / Q000737:chemistry* / Q000633:toxicity; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D005434:Flow Cytometry; D015316:Genetic Therapy; D016655:HIV Core Protein p24 / Q000235:genetics / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism; D008264:Macrophages / Q000276:immunology / Q000378:metabolism; D020320:Oligoribonucleotides, Antisense; D034741:RNA, Small Interfering; D012367:RNA, Viral; D014162:Transfection
chemical_list: D050091:Dendrimers; D016655:HIV Core Protein p24; D020320:Oligoribonucleotides, Antisense; D034741:RNA, Small Interfering; D012367:RNA, Viral