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Anti-human immunodeficiency virus type 1 gene therapy in human central nervous system-based cells: an initial approach against a potential viral reservoir.

Studies have demonstrated that human immunodeficiency virus type 1 (HIV-1) infection of central nervous system (CNS)-based cells in vivo results in a series of devastating clinical conditions collectively termed acquired immune deficiency syndrome (AIDS) dementia complex (ADC). Gene therapy for these neurovirological disorders necessitates utilization of a vector system that can mediate in vivo delivery and long-term expression of an antiretroviral transgene in nondividing/postmitotic CNS cellular elements. The present studies focus on the transfer of an anti-HIV-1 gene to primary isolated CNS microvascular endothelial cells (MVECs) and neuronal-based cells, for its effects in protecting these cells from HIV-1 infection. By using an HIV-1-based vector system, it was possible to efficiently transduce and maintain expression of a marker transgene, beta-galactosidase (beta-Gal), in human CNS MVECs, human fetal astrocytes, plus immature and mature (differentiated) NT2 cells. Significant transduction of the marker gene, beta-Gal, in CNS-based cells prompted the utilization of this system with an anti-HIV-1 gene therapeutic construct, RevM10, a trans-dominant negative mutant Rev protein. Initially, it was not possible to generate any HIV-1 vector particles with the RevM10 gene in the transducing construct, because of inhibitory effects on the HIV-1 vector by this gene product. However, the vector could be partially rescued by adding an additional construct that supplied wild-type rev, in trans, during a multiple construct transfection in the packaging 293T cells. Thus, it was possible to significantly improve the titer of RevM10-expressing viral particles generated from these cells. Moreover, this RevM10 vector transduced the neuronal precursor cell line NT2, retinoic acid-differentiated human neurons (hNT) from the precursor cells, and primary isolated human brain MVECs with high efficiency. RevM10 generated from the HIV-1-based vector system potently inhibited replication of diverse HIV-1 strains in human CNS MVECs and neuronal cells. The data generated from these studies represent an initial approach for future development of anti-HIV-1 gene therapy in the CNS.