735. Femtosecond Laser- A New Intradermal DNA Delivery Method for Gene Expression and Vaccination

The skin is an ideal target for naked DNA delivery for gene expression and vaccination, due to ease of administration and presence of a large number of antigen-presenting cells within the tissue. There are several different methods to transfer naked DNA to the skin. However, some of these delivery methods encounter low level of transfection efficiency and lack of sustained expression. We propose a new and highly efficient method of dermal gene transduction - the Laser Beam Gene Transduction method (LBGT). Methods. Using a femtosecond infrared titanium sapphire laser beam as a laser source illumination, the shaved back skin of Balb/c mice was exposed for assessment of LBGT. The pCAG/Luc construct was used for luc expression. For in vivo real time continuous luc expression, the bioluminescence CCCD system was used (Zeira et al. Mol. Ther. 4:239, 2001). For assessment of this delivery method for vaccination, we used the pCMV/HBsAg vector. Results. A single intra-dermal injection of 10mg of pCAG/Luc followed by laser beam illumination for 2 minutes, resulted in an increase of transgene expression for up to an average of 1000-fold on day 43, relative to naked DNA injection. Characterization studies revealed the optimal conditions for LBGT into the mice dermis. Gene expression following LBGT continued for > 50 days. In mice immunized with the HBsAg plasmid, (10|[mu]|g) on day 0 and 21, HBsAg antibody titers were detected shortly after the first injection and continued to increase over one month, and were significantly higher than titers obtained by naked DNA administration without LBGT. Conclusion. Delivery of DNA plasmid into the dermis by LBGT is highly efficient for transgene expression and for elicitation of immune response. These results provide a proof of principle that femtosecond laser may appear as a novel method for the administration of DNA-based vaccines.