Enhancement of adeno-associated virus serotype 6 transduction into T cells with cell-penetrating peptides

BackgroundAdeno-associated viruses (AAVs) are gaining interest in the development of cellular immunotherapy. Compared to other viral vectors, AAVs can reduce the risk of insertional oncogenesis. AAV serotype 6 (AAV6) shows the highest efficiency for transducing T cells. Nevertheless, a multiplicity of infection (MOI) of up to one million viral genomes per cell is required to transduce the target cells effectively. Cell-penetrating peptides (CPPs) are short, positively charged peptides that easily translocate the plasma membranes and can facilitate the cellular uptake of a wide variety of cargoes, including small molecules, nucleic acids, drugs, proteins and viral vectors.MethodsThe present study evaluated five CPPs (Antp, TAT-HA2, LAH4, TAT1 and TAT2) on their effects on enhancing transduction of AAV6 packaging a green fluorescent protein transgene into Jurkat T cell line.ResultsVector incubation with peptides TAT-HA2 and LAH4 at a final concentration of 0.2 mm resulted in an approximately two-fold increase in transduced cells. At the lowest MOI tested (1.25 x 104), using LAH4 resulted in a 10-fold increase in transduction efficiency. The peptide LAH4 increased the uptake of AAV6 viral particles in both Jurkat cells and mouse primary T cells. Regardless of the large size of the AAV6-LAH4 complexes, their internalization does not appear to depend on macropinocytosis.ConclusionsOverall, the present study reports an approach to significantly improve the delivery of transgenes into T cells using AAV6 vectors. Notably, the peptides TAT-HA2 and LAH4 contribute to improving the use of AAV6 as a gene delivery vector for the engineering of T cells. The present study investigated cell-penetrating peptides (CPPs) to enhance the transduction of adeno-associated viruses (AAVs) into T cells. The CPPs TAT-HA2 and LAH4 significantly increased the uptake of AAV serotype 6 (AAV6) viral particles into both Jurkat and mouse primary T cells, thus improving the potential of AAV6 as a vector for T cell engineering. This approach offers promising avenues for developing novel cellular immunotherapies that rely on AAV6-mediated gene delivery to T cells.image