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Femtosecond laser: a new intradermal DNA delivery method for efficient, long-term gene expression and genetic immunization

Evelyne Zeira^*,1, Alexandra Manevitch^*,1, Zakharia Manevitch, Eli Kedar, Michal Gropp^*, Nili Daudi, Rimma Barsuk, Menahem Harati^*, Hagit Yotvat^*, Philip J. Troilo^||, Thomas G. Griffiths, II^||, Stephen J. Pacchione^||, Dana F. Roden^||, Zhutian Niu^||, Ofer Nussbaum^¶, Gideon Zamir^#, Orit Papo^**, Izhack Hemo, Aaron Lewis and Eithan Galun^*,2

^* Goldyne Savad Institute of Gene Therapy, Hadassah-Hebrew University Hospital;

Department of Applied Physics, The Selim and Rachel Benin School of The Hebrew University of Jerusalem Engineering and Computer Sciences, Natural Science Faculty, and The Department of Ophthalmology, Hadassah Laser Center;

Liver Unit, Hadassah-Hebrew University Hospital;

The Lautenberg Center of Immunology, Hebrew University-Hadassah Medical School, Jerusalem, Israel;

^|| Department of Biologics Safety Assessment, Merck Research Laboratories, Merck & Co., Inc., West Point, Pennsylvania, USA;

^¶ XTL Biopharmaceuticals Ltd., Rehovot, Israel; and

^# Departments of Surgery and of

^** Pathology, Hadassah-Hebrew University Hospital, Jerusalem, Israel

²Correspondence: The Goldyne Savad Institute of Gene Therapy, Hadassah University Hospital, P.O. Box 12000, Jerusalem, 91120 Israel. E-mail: eithang{at}hadassah.org.il

A femtosecond laser beam gene transduction (SG-LBGT) system is described as a novel and efficient method of intradermal (i.d.) nonviral gene delivery in mice by permeabilizing cells utilizing femtosecond laser pulses. Using this approach, significant gene expression and efficient dermal transduction lasting for >7 months were obtained. The ability of this new DNA gene transfer method to enhance genetic vaccination was tested in BALB/C mice. A single i.d. injection of a plasmid (10 µg) containing the hepatitis B virus (HBV) surface antigen (HBsAg), followed by pulses of laser, induced high titers of HBsAg-specific antibodies lasting for >210 days and increased levels of IgG1, IgG2a, IFN, and IL-4, indicating the activation of both Th1 and Th2 cells. Moreover, mice vaccinated using the SG-LBGT followed by challenge with pHBV showed increased protection against viral challenge, as detected by decreased levels of HBV DNA, suggesting an efficient Th1 effect against HBV-infected replicating cells. Tumor growth retardation was induced in vaccinated mice challenged with an HBsAg-expressing syngeneic tumor. In most of the parameters tested, administration of plasmid followed by laser application was significantly more effective and prolonged than that of plasmid alone. Tissue damage was not detected and integration of the plasmid into the host genomic DNA probably did not occur. We suggest that the LBGT method is an efficient and safe technology for in vivo gene expression and vaccination and emphasizes its potential therapeutic applications for i.d. nonviral gene delivery.—Zeira, E., Manevitch, A., Manevitch, Z., Kedar, E., Gropp, M., Daudi, N., Barsuk, R., Harati, M., Yotvat, H., Troilo, P. J., Griffiths, T. G., II, Pacchione, S. J., Roden, D. F., Niu, Z., Nussbaum, O., Zamir, G., Papo, O., Hemo, I., Lewis, A., Galun, E. Femtosecond laser: a new intradermal DNA delivery method for efficient, long-term gene expression and genetic immunization.

Key Words: gene delivery • vaccination • HBV • antitumor