Restriction of HIV-1-based lentiviral vectors in adult primary marrow-derived and peripheral mobilized human CD34+ hematopoietic stem and progenitor cells occurs prior to viral DNA integration.

Daniel O Griffin,Stephen P Goff

doi:10.1186/s12977-016-0246-0

Abstract

BackgroundGene therapy is currently being attempted using a number of delivery vehicles including lentiviral-based vectors. The delivery and insertion of a gene using lentiviral-based vectors involves multiple discrete steps, including reverse transcription of viral RNA into DNA, nuclear entry, integration of viral DNA into the host genome and expression of integrated genes. Transduction of murine stem cells by the murine leukemia viruses is inefficient because the expression of the integrated DNA is profoundly blocked. Transduction of human stem cells by lentivirus vectors is also inefficient, but the cause and specific part of the retroviral lifecycle where this block occurs is unknown.ResultsHere we demonstrate that the dominant point of restriction of an HIV-1-based lentiviral vector in adult human hematopoietic stem and progenitor cells (HSPCs) from bone marrow and also those obtained following peripheral mobilization is prior to viral DNA integration. We specifically show that restriction of HSPCs to an HIV-1-based lentiviral vector is prior to formation of nuclear DNA forms.ConclusionsMurine restriction of MLV and human cellular restriction of HIV-1 are fundamentally different. While murine restriction of MLV occurs post integration, human restriction of HIV-1 occurs before integration.

Highlights

Gene therapy is currently being attempted using a number of delivery vehicles including lentiviralbased vectors
Our study evaluates infection of primary adult marrow-derived and peripheral mobilized human CD34+ cells, a mixed population composed of true human HSCs (Lin−34+38−90+45RAdim) as well as several hematopoietic progenitor cell (HPC) types, with a vesicular stomatitis virus glycoprotein (VSV-G) pseudotyped full-length nef-negative HIV-1based vector
Transduction of primary adult human marrow‐derived CD34+ cells by lentiviral vectors is inefficient at low multiplicity of infection (MOI) We investigated the efficiency of lentiviral transduction of primary adult human marrow-derived CD34+ cells under various conditions

Summary

Introduction

Gene therapy is currently being attempted using a number of delivery vehicles including lentiviralbased vectors. Other investigations have suggested a limited role of interferonstimulated genes in the restriction of HIV-1 and related vectors in the human system [25, 26]. It is unclear whether the restriction of lentiviruses in HSPCs is due to a specific restriction factor such as TRIM5α, as has been found in other cell types, or to the absence of a specific essential co-factor of infection [27,28,29]. When human HSPCs are pre-stimulated with cytokines a large number of genome-wide modifications occur that allow for successful lentiviral transduction, but the essential changes responsible for this transformation remain unexplored [30]

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