Efficient gene expression by self-complementary adeno-associated virus serotype 2 and 5 in various human cancer cells

Abstract

Abstract. The feasibility of various self-complementary AAV (scAAV) serotypes as efficient gene delivery vehicles in human cancer cells was evaluated. To dissect the transduction characteristics, we infected a variety of human cancer cells with scAAV1-6 or scAAV8 expressing GFP. scAAV2 led to the best transduction efficiency with nearly complete transgene expression at 1000 MOI in most cancer cells, regardless of cell/tissue origins. scAAV5 could also induce effective gene expression, even though gene transfer potency by scAAV5 was poorer than that by scAAV2. Substantial portion of transgene expression lasted over a month following gene delivery by both scAAV 2 and scAAV5, indicating that long-term gene expression can occur. Moreover, co-infection of scAAV2 and scAAV5 can induce simultaneous transgene expressions introduced via each vector. Thus, the current study provide evidence that scAAV2 and scAAV5 vectors are excellent gene transfer tools in a wide variety of human cancer cells, independently driving persistent transgene expression.

Self-complementary adeno-associated viral vectors for gene therapy of hemophilia B: progress and challenges

“…Other AAV serotypes have been shown to mediate effective gene transfer to other tissues, such as AAV6 for cardiac cells [64,79], AAV1 for hematopoietic stem cells [26,80] and AAV3 for liver tumor cells [53]. Nevertheless, the AAV2 capsid is the most effective in transduction of the majority of cell types tested thus far [52,79,81,82]. Pre-existing immunity to specific AAV serotypes, as well as the need for these serotypes to transduce a target cell type, may therefore limit the potential for AAV-mediated gene transfer.…”

Acquisition of selective antitumoral effects of recombinant adeno-associated virus by genetically inserting tumor-targeting peptides into capsid proteins

“…Cells were infected with various rAAVs, as previously described (6). To maximize the transduction of rAAV vectors, adenovirus 5 was occasionally added.…”

“…Using pSp72 plasmid containing the Rep gene of AAV2 and the Cap gene of AAV5 as an original backbone, final constructs were generated by a series of cloning steps and sequencing conformation. Self-complementary rAAV vectors were generated as described elsewhere (6). Titration of rAAVs prepared using CsCl 2 gradient ultracentrifugation was performed using TaqMan-based Q-PCRs (iQTM Supermix, Bio-Rad, Hercules, CA, USA).…”

“…Among these rAAV serotypes undergoing development as gene transfer tools, the rAAV2 vector has been analyzed most extensively, and is currently employed in clinical gene therapy (3,5). Recently, we and other groups demonstrated the potential of the rAAV5 serotype as an effective vector for antitumoral gene delivery (6,7). Other studies reported the lowest prevalence of AAV5-neutralizing antibodies and the highest prevalence of AAV2-neutralizing antibodies in the normal human population (8,9).…”

“…level of neutralizing antibodies in human populations, and superior virus production yield over other serotypes (6,9,28). Limited information is available on rAA5 modification at present (29), although several studies have focused on genetic modification of rAAV1 and 2 capsids (20,30,31).…”