Retrovirus molecular conjugates: a versatile and efficient gene transfer vector system for primitive human hematopoietic progenitor cells

Abstract

In principle, transient nongenetic modification of a noninfectious gene transfer virus enabling a one time infection and transduction of human cells could eliminate the risk of formation of replication competent virus. Formation of a molecular conjugate vector by conjugation of noninfective ecotropic murine Moloney leukemia virus to polylysine (eMMLV-PL) enabled high-efficiency transduction of human HPC using in vitro and in vivo assays. Xenotransplanted NOD-SCID mice durably expressed the transgene in human leukocytes and human progenitor cells with eMMLV-PL achieving three-fold increased transduction efficiency when directly compared to optimized amphotropic MMLV (aMMLV) transduction. Both aMMLV and eMMLV assembled conjugate vectors showed similar transduction efficiency indicating predominant polylysine-mediated uptake. Integration of retroviral sequences was determined from individual human HPC recovered from eMMLV-PL-xenotransplanted animals. This simple and versatile concept of conjugate gene transfer vectors has the potential to enhance transduction efficiency as well as to improve certain safety aspects of human gene therapy. Moreover, because it permits effective cellular internalization of particles, this concept of molecular conjugates can be used as research tool to investigate the interactions of otherwise noninfectious viruses or modified viral particles at the genomic level.

PEGylated Cationic Serum Albumin for Boosting Retroviral Gene Transfer

“…For example, charged polymers, which act at an early, receptor‐independent step of infection, can alter the transduction efficiencies . Cationic polymers such as poly‐ l ‐lysine, polybrene or protamine enhance transduction rates by reducing the electrostatic repulsion between retroviral particles and cells.…”

PostexitSurface Engineering of Retroviral/Lentiviral Vectors

“…However, the modification resulted in severely reduced infectivity of R/LV particles. Direct chemical biotinylation of retroviral vectors has also been demonstrated, using sulfo-N-hydroxysuccinimide-biotin on MoMLV derived vectors [11]. Neutravidin was covalently linked to polylysine.…”

“…Other approaches lead to the biotinylation of the lentiviral vector. This can be achieved by direct chemical modification [11] or after addition of a biotin-adaptor peptide (BAP), a site for specific enzymatic biotin ligation [22, 23]. The bacterial enzyme, biotin ligase, has to be provided as a form of metabolic engineering to allow the modification of the BAP-containing protein.…”