Characterization of a third generation lentiviral vector pseudotyped with Nipah virus envelope proteins for endothelial cell transduction

Abstract

Lentiviruses are becoming progressively more popular as gene therapy vectors due to their ability to integrate into quiescent cells and recent clinical trial successes. Directing these vectors to specific cell types and limiting off-target transduction in vivo remains a challenge. Replacing the viral envelope proteins responsible for cellular binding, or pseudotyping, remains a common method to improve lentiviral targeting. Here, we describe the development of a high titer, 3rd generation lentiviral vector pseudotyped with Nipah virus fusion protein (NiV-F) and attachment protein (NiV-G). Critical to high titers was truncation of the cytoplasmic domains of both NiV-F and NiV-G. As known targets of wild-type Nipah virus, primary endothelial cells are shown to be effectively transduced by the Nipah pseudotype. In contrast, human CD34+ hematopoietic progenitors were not significantly transduced. Additionally, the Nipah pseudotype has increased stability in human serum compared to VSV pseudotyped lentivirus. These findings suggest that the use of Nipah virus envelope proteins in 3rd generation lentiviral vectors would be a valuable tool for gene delivery targeted to endothelial cells.

Nipah virus fusion protein: Importance of the cytoplasmic tail for endosomal trafficking and bioactivity

“…A similar finding was made by Palomares et al (2013) for F 23. Though Witting et al (2013) reported that G tail truncation had a more pronounced effect on LV production as did F tail deletions, they also observed that F proteins with a large tail deletion (F 25) improved the LV titers.…”

“…Glycoprotein G is responsible for binding to ephrin-B2/-B3 on host cells (Bonaparte et al, 2005;Negrete et al, 2005Negrete et al, , 2006 and complexes of G and F mediate fusion between viral and cellular membranes during virus entry and cell-to-cell spread (reviewed in Diederich and Maisner, 2007). Aside of their incorporation into infectious NiV particles, NiV-F and -G can efficiently pseudotype lentiviral vectors thereby allowing specific targeting of ephrin-positive cells (Khetawat and Broder, 2010;Palomares et al, 2013;Witting et al, 2013). http://dx.doi.org/10.1016/j.ejcb.2015.05.005 0171-9335/© 2015 Elsevier GmbH.…”

“…However, this mutation was not seen to obviously affect fusion activity. Three other papers describing tail truncation mutants with or without deletion of the Y 525 RSL endocytosis signal gave inconsistent results, ranging from no effect of certain deletions to increased or decreased expression and fusion activity (Aguilar et al, 2007;Khetawat and Broder, 2010;Witting et al, 2013). The lack of a clear picture for tail truncation mutants might be due to different cell types or expression systems, or usage of tail deletion mutants containing an AU tag (DTYRYI) adding a bona fide endocytosis signal to the cytoplasmic domain.…”

Pseudotyping of lentiviral vector with novel vesiculovirus envelope glycoproteins derived from Chandipura and Piry viruses

“…During the last two decades, several heterologous viral envelopes have been tested for lentiviral vector pseudotyping to facilitate efficient gene transfer into various cell types [9–19]. Among the first to be used were the ones from amphotropic retroviruses which were soon followed by broader host tropic viral envelopes that included vesicular stomatitis virus [9, 20–22].…”

“…For example, Rabies virus glycoprotein shows more efficient gene delivery to the nervous system, compared with VSV-G pseudotypes [11]. Other envelope proteins employed for lentiviral vector pseudotyping include those from the measles virus, baboon retrovirus, filovirus, baculovirus, Nipah virus, Ross river virus and Cocal virus among others [10, 15, 16, 19, 23–25]. Relative to VSV-G, some of these alternative envelope proteins achieved comparable or better gene delivery efficiencies into certain tissues or cells, such as airway epithelia and resting hematopoietic stem cells [10, 23, 26].…”