title: In vitro evolution of a highly replicating, doxycycline-dependent HIV for applications in vaccine studies.
abstract: A major concern associated with the use of vaccines based on live-attenuated viruses is the possible and well documented reversion to pathogenic phenotypes. In the case of HIV, genomic deletions or mutations introduced to attenuate viral pathogenicity can be repaired by selection of compensating mutations. These events lead to increased virus replication rates and, eventually, disease progression. Because replication competence and degree of protection appear to be directly correlated, further attenuation of a vaccine virus may compromise the ability to elicit a protective immune response. Here, we describe an approach toward a safe attenuated HIV vaccine. The system is not based on permanent reduction of infectivity by alteration of important viral genomic sequences, but on strict control of replication through the insertion of the tetracycline (Tet) system in the HIV genome. Furthermore, extensive in vitro evolution was applied to the prototype Tet-controlled HIV to select for variants with optimized rather than diminished replication capacity. The final product of evolution has properties uniquely suited for use as a vaccine strain. The evolved virus is highly infectious, as opposed to a canonically attenuated virus. It replicates efficiently in T cell lines and in activated and unstimulated peripheral blood mononuclear cells. Most importantly, replication is strictly dependent on the nontoxic Tetanalogue doxycycline and can be turned on and off. These results suggest that this in vitro evolved, doxycycline-dependent HIV might represent a useful tool toward the development of a safer, live-attenuated HIV vaccine.
authors.affiliation: Department of Human Retrovirology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands.
authors.forename: G
authors.initials: G
authors.lastname: Marzio
authors.forename: K
authors.initials: K
authors.lastname: Verhoef
authors.forename: M
authors.initials: M
authors.lastname: Vink
authors.forename: B
authors.initials: B
authors.lastname: Berkhout
mesh_terms: D016915:AIDS Vaccines / Q000235:genetics; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001483:Base Sequence; D002478:Cells, Cultured; D019020:Directed Molecular Evolution; D004318:Doxycycline / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D013601:T-Lymphocytes / Q000166:cytology; D013997:Time Factors; D014407:Tumor Cells, Cultured; D014613:Vaccines, Attenuated; D014775:Virus Activation; D014779:Virus Replication / Q000187:drug effects*
chemical_list: D016915:AIDS Vaccines; D000900:Anti-Bacterial Agents; D014613:Vaccines, Attenuated; D004318:Doxycycline