Development of an Rev-Independent, Minimal Simian Immunodeficiency Virus-Derived Vector System

Abstract

Lentiviral vectors are attractive candidates for gene therapy because of their ability to integrate into nondividing cells. To date, conventional HIV-1-based vectors can be produced at higher titers, but concerns regarding their safety for human use exist because of the possibility of recombination leading to production of infectious virions with pathogenic potential. Development of lentivirus vectors based on nonhuman lentiviruses constitutes an active area of research. We described a novel HIV-SIV hybrid vector system in which an HIV-1-derived transfer vector is encapsidated by SIVmac1A11 core particles and pseudotyped with VSV glycoprotein G. In an effort to further develop this vector system, we modified the packaging plasmid by deletion of the SIV accessory genes. Specifically, versions of the packaging plasmid (SIVpack) lacking vif, vpr, vpx, and/or nef were constructed. Our results indicate that, as with HIV-1-based packaging plasmids, deletion of accessory genes has no significant effect on transduction in either dividing or nondividing cells. The SIV packaging plasmid was also modified with regard to the requirement for RRE and rev. Deletion of the RRE and rev from SIVpack led to dramatic loss of transduction ability. Introduction of the 5' LTR from the spleen necrosis virus to packaging plasmids lacking RRE/Rev was then sufficient to fully restore vector titer. A minimal SIV transfer vector was also developed, which does not require RRE/Rev and exhibits no reduction in transduction efficiency in two packaging systems. The SIV-based vector system described here recapitulates the biological properties of minimal HIV-1-derived systems and is expected to provide an added level of safety for human gene transfer. We suggest that the SIV-derived vector system will also be useful to deliver anti-HIV-1 gene therapy reagents that would inhibit an HIV-1-derived vector.

FIV: from lentivirus to lentivector

“…The potential to exploit this property has been the principal incentive for developing viral vectors from these pathogenic complex retroviruses. After HIV-1 vectors were described, replication-defective systems were engineered from molecular clones of non-primate and other primate lentiviruses [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19]. In addition to their use in pre-clinical gene therapy research, lentiviral vectors have been important tools for basic science [20][21][22][23][24][25][26][27][28].…”

Silencing of HIV-1 Gene Expression by Two Types of siRNA Expression Systems

“…(Naldini et al, 1996;Manganini et al, 2002) and deliver genes into dividing cells at high efficiency (Tavernarakis et al, 2000;Yu et al, 2002). (Follenzi et al, 2000;Li et al, 2003;Mautino & Morgan, 2002;Mukhtar et al, 2000;Pandya et al, 2001;Qin et al, 2003;Schroers et al, 2002 Figure 5A). Furthermore, siRNA and GFP expression was observed in the CS-env-shRNA plasmid-transfected SupT1 cells for 15 days (Figures 5B,C).…”

Production of Retroviral and Lentiviral Gene Therapy Vectors: Challenges in the Manufacturing of Lipid Enveloped Virus

“…The envelope cassette encodes typically, but not necessarily, for VSV-G envelope glycoprotein. The development of a fourth generation of lentiviral vectors, rev independent, has also been claimed by means of replacing RRE (rev responsive element) with heterologous viral sequences or by codon-optimization (Bray et al 1994;Delenda 2004;Kotsopoulou et al 2000;Pandya et al 2001;Roberts and Boris-Lawrie 2000). However, its use is not widespread since, contrary to the other generations of lentiviral vectors, these packaging systems have not been made available for the research community; also the reported titers are typically one to two logs bellow the maximum titers obtained with the second or third generation systems.…”