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We have developed and tested chimeric T-cell receptors (TCR) specific for p185HER2. In these experiments, retroviral vectors expressing the N29γ or N29ζ receptors were constructed in pRET6. Amphotropic viral producer cells were established in the GALV-based PG13 packaging cell line. Ficoll purified human peripheral blood lymphocytes (PBL) were virally transduced using an optimized protocol incorporating activation with immobilized anti-CD3/anti-CD28 monoclonal anti- bodies, followed by viral infection in the presence of fibronectin fragment CH296. Transduced cells were co-cultured with human tumor cell lines that overexpress (SK-OV-3) or underexpress (MCF7) p185HER2 to assay for antigen specific im- mune responses. Both CM+ and CD8+ T-cells transduced with the N29γ or N29ζ chTCR demonstrated HER2-specific anti- gen responses, as determined by release of Th1 like cytokines, and cellular cytotoxicity assays. Our results support the fea- sibility of adoptive immunotherapy with genetically modified T-cells expressing a chTCR specific for p185HER2.
We have developed and tested chimeric T-cell receptors (TCR) specific for p185HER2. In these experiments, retroviral vectors expressing the N29γ or N29ζ receptors were constructed in pRET6. Amphotropic viral producer cells were established in the GALV-based PG13 packaging cell line. Ficoll purified human peripheral blood lymphocytes (PBL) were virally transduced using an optimized protocol incorporating activation with immobilized anti-CD3 / anti-CD28 monoclonal anti- bodies, followed by viral infection in the presence of fibronectin fragment CH 296. Transduced cells were co-cultured Both CM + and CD8 + T-cells were transduced with the N29γ or N29ζ chTCR demonstrated HER2-specific anti-tumor activity in human tumor cell lines that overexpress (SK-OV-3) or underexpress (MCF7) p185HER2 to assay for antigen specific im- gen responses, as determined by release of Th1 like cytokines, and cellular cytotoxicity assays. Our results support the fea- sibility of adoptive immunotherapy with genetic ally modified T-cells expressing a chTCR specific for p185HER2.