Vectofusin-1, a New Viral Entry Enhancer, Strongly Promotes Lentiviral Transduction of Human Hematopoietic Stem Cells

Abstract

Gene transfer into hCD34+ hematopoietic stem/progenitor cells (HSCs) using human immunodeficiency virus type 1 (HIV-1)-based lentiviral vectors (LVs) has several promising therapeutic applications. Yet, efficiency, safety, and cost of LV gene therapy could be ameliorated by enhancing target cell transduction levels and reducing the amount of LV used on the cells. Several transduction enhancers already exist such as fibronectin fragments and cationic compounds, but all present limitations. In this study, we describe a new transduction enhancer called Vectofusin-1, which is a short cationic peptide, active on several LV pseudotypes. Vectofusin-1 is used as a soluble additive to safely increase the frequency of transduced HSCs and to augment the level of transduction to one or two copies of vector per cell in a vector dose-dependent manner. Vectofusin-1 acts at the entry step by promoting the adhesion and the fusion between viral and cellular membranes. Vectofusin-1 is therefore a promising additive that could significantly ameliorate hCD34+ cell-based gene therapy.

Retrovirus-based vectors for transient and permanent cell modification

“…Furthermore, small molecules can serve to improve transduction efficiencies. In addition to a panel of well-known compounds, such as polybrene or protamine sulphate, Vectofusin-1 and phorbol 12-myristate 13-acetate (PMA) were recently identified to enhance transduction of CD34+ hematopoietic stem/progenitor cells [21,22].…”

Molecular Determinants of Vectofusin-1 and Its Derivatives for the Enhancement of Lentivirally Mediated Gene Transfer into Hematopoietic Stem/Progenitor Cells

“…Viral Vector Production and Vector Titering-LVs were generated by transient calcium phosphate transfection of HEK293T cells with four plasmids: the gagpol (pKLgagpol) and rev (pBA.rev) expression plasmids (34), the transfer plasmid (pCCLsin.cPPT.hPGK.eGFP.WPRE), and the plasmid encoding either the VSV-G (pMDG) or the GALVTR envelope glycoprotein (pBA.GALV/Ampho-Kana) (29). After 24 h of production, raw viral supernatants were harvested, filtered (0.45 m), and frozen at Ϫ80°C.…”

“…Recently, we described a new transduction enhancer, Vectofusin-1 (29), also called LAH4-A4, a short histidine-rich amphipathic peptide derived from the LAH4 family of DNA transfection agents (30 -33). Vectofusin-1 promotes the transduction of HSPCs and different cell lines with a broad range of lentiviral and ␥-retroviral pseudotypes with no apparent cytotoxicity (29).…”

“…Vectofusin-1 promotes the transduction of HSPCs and different cell lines with a broad range of lentiviral and ␥-retroviral pseudotypes with no apparent cytotoxicity (29). The enhancing effects of Vectofusin-1 are comparable with those of other viral transduction enhancers, such as semen-derived enhancer of viral infection peptide or Retronectin, a human fibronectin fragment used currently in clinical settings (29). However, the critical determinants of Vectofusin-1 playing a key role in lentiviral transduction enhancement are still unknown.…”

Concurrent Measures of Fusion and Transduction Efficiency of Primary CD34+ Cells with Human Immunodeficiency Virus 1-Based Lentiviral Vectors Reveal Different Effects of Transduction Enhancers

“…Another strategy to improve the entry step is the addition of cofactors during the transduction procedure, such as cationic polymers-for example, polybrene (Toyoshima and Vogt, 1969;Cornetta and Anderson, 1989), DEAE-Dextran (Vogt, 1967), or cationic peptides [e.g., Vectofusin-1 (Fenard et al, 2013b), Retronectin (Pollok and Williams, 1999), protamine sulfate (Cornetta and Anderson, 1989), human semen enhancer of viral infection (Wurm et al, 2010), or HIV-1 gp120-derived peptides (Yolamanova et al, 2013). The action mechanism of these cationic additives is mainly described as the capacity to neutralize membrane charges, to improve virus-cell interactions and virus-cell fusion, and/or to promote virus aggregation (Pollok and Williams, 1999;Davis et al, 2004;Roan et al, 2009;Fenard et al, 2013b;Yolamanova et al, 2013).…”