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Constitutive expression of chimeric neo-Rev response element transcripts suppresses HIV-1 replication in human CD4+ T
D Bevec1, B Volc-Platzer, K Zimmermann
1ART Department, Sandoz Research Institute, Vienna, Austria.
Abstract:
We have previously reported that chimeric neomycin phosphotransferase (neo)-Rev response element (RRE) transcripts suppress the function of the human immunodeficiency virus type 1 (HIV-1) Rev trans-activator protein in HeLa cells. In an extension of these experiments, human CD4+ CEM cells (G418-resistant cell populations and clonal isolates) stably expressing chimeric neo-RRE genes (2, 3, or 6 RRE copies) were generated using retroviral-mediated gene transfer. The transduced CEM clones were infected with the HIV-1 HTLVIIIB isolate and the following three phenotypes were observed: (i) the transduced CEM cells were readily infected with HIV-1 indistinguishable from the control CEM cells; (ii) the appearance of HIV-1 replication markers was significantly delayed; (iii) no signs of HIV-1 replication were detectable although proviral HIV-1 DNA sequences could be detected in these cells. Furthermore, HIV antigen expression was limited in neo-resistant CEM cell populations inoculated with the HIV-1 HTLVIIIB isolate. Only 10% of the CEM-pX17-3xRRE cells and 20% of the CEM-pX17-2xRRE cells displayed HIV-1 antigens 43 days after challenge and had retained CD4 surface expression on 47% and 64% of the cells, respectively. In sharp contrast, 80% of the CEM-pX17 or the CEM-pX17-6xRRE cells expressed HIV-1 antigens but no CD4 antigens were detectable in these cultures. These results clearly indicate that RRE decoys could be developed into an effective somatic gene therapy approach against HIV-1 induced acquired immunodeficiency syndrome (AIDS).
Insights
Chimeric neo-Rev response element (RRE) transcripts effectively suppress human immunodeficiency virus type 1 (HIV-1) replication in CD4+ cells. This gene therapy approach shows promise for treating HIV-1 induced acquired immunodeficiency syndrome (AIDS).
Area of Science:
- Molecular Biology
- Gene Therapy
- Virology
Background:
- Chimeric neomycin phosphotransferase (neo)-Rev response element (RRE) transcripts were previously shown to inhibit human immunodeficiency virus type 1 (HIV-1) Rev trans-activator function in HeLa cells.
- The Rev protein is crucial for HIV-1 replication, making it a target for therapeutic intervention.
Purpose of the Study:
- To evaluate the efficacy of stably expressing chimeric neo-RRE genes in human CD4+ CEM cells as a potential gene therapy for HIV-1.
- To assess the impact of varying RRE copy numbers on HIV-1 replication and cell survival.
Main Methods:
- Generation of G418-resistant human CD4+ CEM cell populations and clonal isolates using retroviral-mediated gene transfer to stably express chimeric neo-RRE genes (2, 3, or 6 copies).
- Infection of transduced CEM cells with the HIV-1 HTLVIIIB isolate.
- Analysis of HIV-1 replication markers, antigen expression, and CD4 surface expression in infected cells.
Main Results:
- Transduced CEM cells were susceptible to HIV-1 infection, but HIV-1 replication markers were significantly delayed.
- No detectable HIV-1 replication occurred in some cell lines, despite the presence of proviral DNA.
- Limited HIV antigen expression and retained CD4 surface expression were observed in cells with 2 or 3 RRE copies, contrasting with cells expressing 6 RRE copies or no RRE, which showed higher antigen expression and CD4 loss.
Conclusions:
- RRE decoys can be effectively developed into a somatic gene therapy strategy against HIV-1 induced acquired immunodeficiency syndrome (AIDS).
- The number of RRE copies influences the therapeutic outcome, with intermediate copy numbers showing more favorable results in preserving CD4+ T cells.