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196 GENE THERAPY USING RECOMBINANT ADENO-ASSOCIATED VIRUS/HER-2/NEU LOADING OF DENDRITIC CELLS FOR A POTENT CELLULAR MEDIATE IMMUNE RESPONSE AGAINST LYMPHOMA PRIMARY TUMORS.
  1. E. White1,
  2. M. Chiriva-Internati1,2,
  3. F. Grizzi3,
  4. E. Cobos2
  1. 1Department of Microbiology and Immunology
  2. 2Internal Medicine, Texas Tech University Health Sciences Center and Southwest Cancer Treatment and Research Center, Lubbock, TX
  3. 3Scientific Direction, Istituto Clinico Humanitas, Rozzano, Milan, Italy

Abstract

Purpose Recent studies demonstrate that recombinant adeno-associated virus (rAAV)-based antigen-loading of dendritic cells (DC) generates significant and rapid cytotoxic T lymphocyte (CTL) responses in vitro. As a more extensive analysis of the rAAV system, we used a self-antigen, Her-2, expressed in many cancers including breast and ovarian cancer, in particular, lymphoma.

Methods The AAV vectors were found to be able to transduce up to 85% of DC and the transduced DC displayed higher levels of CD80, CD83, CD86, and CD1a over controls. Autologous PBMC/LCL targets and Her-2/neu positive lymphoma primary cancer cell. Generation of CTL and test their function by Cr(51) release assay against LCL/Her-2/neu target and relative controls.

Summary We used the Her-2/neu gene, divided into 3 overlapping segments for insertion into AAV. The three Her-2 subgenes are aa 153-653, 403-906, and 76-1255. Using only one stimulation, significant MHC class I-restricted, anti-Her-2-specific CTL killing was demonstrated against a Her-2/neu-positive lymphoma primary cancer cell line. Using synthetic antigen-positive target cells with the three Her-2 subgenes and the primary lymphoma positive for Her-2/neu for a target, we stimulated highest CTL killing. The killing was done using specific Her-2(403-903)CTL against a target present in the particular peptide and inserted by rAAV AAV/Her-2(403-906)/Neo in the LCL.

Conclusion These data suggest that AAV-based antigen loading of DC is highly effective for generating a CTL response against lymphoma tumor.

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