title: Blocking of retroviral infection at a step prior to reverse transcription in cells transformed to constitutively express interferon beta.
abstract: We are developing methods for somatic-cell gene therapy directed against infection with human immunodeficiency virus, by enhancing antiviral resistance of target cells through the constitutive production of autocrine interferon (IFN). Using the human IFN-beta coding sequence under the constitutive low-expression control of a 0.6-kb murine H-2Kb promoter-fragment, we have constructed a retroviral vector, HMB-KbHuIFN beta, and have transformed cells of the T98G human neuroblastoma line, the U-937 human promonocytic line, and the CEM human lymphocytic line. These human IFN-beta-transformed cell populations have acquired a low, constitutive production of human IFN, while replicating at a rate similar to that of untransformed cells and of cells transformed with the control vector carrying a human IFN-beta sequence encoding an inactive, mutated protein. In the three different cell populations tested, transformation with the HMB-KbHuIFN beta vector resulted in a 1.3-2.3 log10 reduction in the number of cells infected with a defective amphotropic MFG-LaZ retrovirus. A kinetic study of the fate of the MFG-LacZ retrovirus in the culture medium and intracellularly immediately after exposure of the cells to virus revealed a significant reduction of the appearance of intracellular virus in human IFN-beta-transformed cells. A similar effect was obtained by treating untransformed T98G, U-937, and CEM cells with exogenous human IFN-beta. The blocking effect of autocrine or exogenous human IFN-beta on viral entry was not limited to virus specific for the amphotropic receptor but was also obtained in murine IFN-beta-treated NIH 3T3 mouse fibroblasts infected with an ecotropic MFG-LacZ retrovirus. Infection of human IFN-beta-transformed CEM cells with human immunodeficiency virus type 1 gave comparable results. Immediately following exposure of the cells to human immunodeficiency virus, a kinetic study of the fate of the virus failed to reveal the appearance of intracellular virus and showed that the majority of the input virus remained in the extracellular medium. We conclude that low autocrine IFN-beta synthesis, or exposure of cells to exogenous IFN-beta, prevents virus from getting inside the cells, regardless of the virus receptor involved.
authors.affiliation: Centre Nationale de la Recherche Scientifique, Institut Curie, Université Paris XI, Orsay, France.
authors.forename: V
authors.initials: V
authors.lastname: Vieillard
authors.forename: E
authors.initials: E
authors.lastname: Lauret
authors.forename: V
authors.initials: V
authors.lastname: Rousseau
authors.forename: E
authors.initials: E
authors.lastname: De Maeyer
mesh_terms: D000818:Animals; D002472:Cell Transformation, Viral; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D006678:HIV / Q000187:drug effects / Q000254:growth & development; D006801:Humans; D016899:Interferon-beta / Q000235:genetics / Q000494:pharmacology*; D012194:RNA-Directed DNA Polymerase; D011994:Recombinant Proteins / Q000494:pharmacology; D012190:Retroviridae / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D014158:Transcription, Genetic
chemical_list: D011994:Recombinant Proteins; D016899:Interferon-beta; D012194:RNA-Directed DNA Polymerase