Retroviral Vector-Mediated Expression in Primary Human T Cells of an Endoplasmic Reticulum-Retained CD4 Chimera Inhibits Human Immunodeficiency Virus Type-1 Replication

Abstract

Intracellular expression of genes that inhibit key steps in the human immunodeficiency virus (HIV-1) replicative cycle could offer an alternative therapy for AIDS treatment. One of these approaches involves the inhibition of env protein maturation through the expression of CD4 molecules with added exogenous sequences that promote their retention in the endoplasmic reticulum (ER). We have tested this strategy using a CD4 chimera (CD4ϵ10) containing an ER retention sequence derived from the TCR CD3-ϵ chain. Transfection of CD4ϵ10 in the human T cell line Jurkat made it resistant to infection with two different HIV-1 isolates, which was evaluated by measuring p24 antigen production, induction of apoptosis, and syncytia formation. Furthermore, polymerase chain reaction (PCR) analysis of genomic DNA showed no traces of the proviral HIV-1 genome in CD4ϵ10-transfected cells, suggesting it was not maintained latently in these cells. To facilitate the delivery of the CD4ϵ10 chimera to primary cells from AIDS patients, a Moloney-based retroviral vector was constructed that expresses CD4ϵ10 under the transcriptional control of the HIV-1 long terminal repeat (LTR) promoter. Transduction of the MT-2 human T cell line with this vector rendered it resistant to infection with HIV-1 by a process that involved the inhibition of gp160 proteolytic processing. Finally, transduction of the CD4ϵ10 chimera into T lymphoblasts derived from asymptomatic HIV-infected individuals demonstrated a protective effect, resulting in both an increased cellular proliferation rate and an increased percentage of CD4⁺ cells. These results suggest that it is feasible to use retroviral transduction of CD4ϵ10 as a gene therapy approach for AIDS treatment.

Overview summary

The intracellular expression of genes that interfere with the human immunodeficiency virus (HIV) replicative cycle, a method known as intracellular immunization, may be an alternative strategy for AIDS therapy. We aim to develop one of these strategies consisting on the transduction of primary T cells or their progenitors with a CD4 chimera that may inhibit env protein maturation and consequently, viral spreading. Here we show that the CD4 chimera actually inhibits HIV-1 replication when expressed in human T cell lines. To facilitate delivery to primary T cells, a retroviral vector capable of transducing the CD4 chimera has been developed. Transduction of this construct protected lymphoblasts derived from the blood of HIV-infected individuals from the cytotoxic effect of HIV-1 infection and resulted in a recovery in the numbers of CD4⁺ cells. These results are the first step toward the use of this retroviral construct for gene therapy of AIDS patients.