Biomaterial-mediated retroviral gene transfer using self-assembled monolayers

Abstract

Biomaterial-mediated gene delivery has recently emerged as a promising alternative to conventional gene transfer technologies that focus on direct delivery of viral vectors or DNA-polymer/matrix complexes. However, biomaterial-based strategies have primarily targeted transient gene expression vehicles, including plasmid DNA and adenovirus particles. This study expands on this work by characterizing biomaterial properties conducive to the surface immobilization of retroviral particles and subsequent transduction of mammalian cells at the cell-material interface. Self-assembled monolayers (SAMs) of functionally-terminated alkanethiols on gold were used to establish biomaterial surfaces of defined chemical composition. Gene transfer was observed to be greater than 90% on NH(2)-terminated surfaces, approximately 50% on COOH-functionalized surfaces, and undetectable on CH(3)-terminated SAMs, similar to controls of tissue culture-treated polystyrene. Gene delivery via the NH(2)-SAM was further characterized as a function of retrovirus coating time, virus concentration, and cell seeding density. Finally, SAM-mediated gene delivery was comparable to fibronectin- and poly-l-lysine-based methods for gene transfer. This work is significant to establishing safe and effective gene therapy strategies, developing efficient methods for gene delivery, and supporting recent progress in the field of biomaterial-mediated gene transfer.

Scaffold-mediated lentiviral transduction for functional tissue engineering of cartilage

“…Our findings extend previous work showing biomaterialmediated gene delivery using viral vectors (20,(40)(41)(42)(43)(44)(45)(46)(47)(48)(49). The majority of this work demonstrated the ability to specify the spatial location of delivered vectors, rather than the induction of tissuespecific differentiation.…”

“…Importantly, we showed that material-mediated transduction of hMSCs remained >90% after rinsing the substrates upon which lentivirus was immobilized. Moreover, we have also shown that incubation of PLL-coated surfaces with retrovirus for 16 h results in high levels of transduction of fibroblasts (∼70%) (20), suggesting that the half-life of the vector does not preclude efficient transduction. Collectively, these results support the potential of viral transduction in the joint environment.…”

“…Our findings suggest that scaffold-mediated gene delivery could provide an efficient means of in situ transduction, owing to the ability of PLL to immobilize vectors to the biomaterial, as well as the efficiency of transduction of the VSV-G pseudotyped lentivirus. In addition, we have previously demonstrated that passive adsorption of ECM onto highly cationic surfaces can direct both viral immobilization and cell adhesion (20), opening the possibility of functionalizing polymeric scaffolds with gene delivery vehicles as well as ECM components or synthetic peptides capable of enhancing cell differentiation and tissue formation (12,14). However, important questions, such as whether endogenous cells that infiltrate the scaffold at the site of implantation can be driven to differentiate toward the chondrocyte lineage, remain unanswered.…”

“…Films were then incubated in PLL solution (Sigma-Aldrich) diluted to 0.002% with DPBS for 1 h to allow for passive adsorption. PLL was aspirated, and LV-eGFP supernatant was pipetted into the well plate and allowed to incubate for 4 h at 37°C (20). DPBS was used in place of PLL in untreated control wells.…”

“…Moreover, as poly-L-lysine (PLL)-coated biomaterials effectively immobilize other enveloped viruses and facilitate efficient transduction of cells (20), PLL was used to functionalize poly (e-caprolactone) (PCL)-derived biomaterials with lentivirus (LV). In this study, we first tested whether PLL could effectively immobilize LV to PCL films and lead to efficient transduction of human mesenchymal stem cells (hMSCs).…”