par Zhang, Yi;Sun, Zhaojun;Nicolay, Hugues;Meyer, Ralf R.G.;Renkvist, Nicolina;Stroobant, Vincent;Corthals, Jurgen;Carrasco, Javier;Eggermont, Alexander A.M.M.;Marchand, Marie ;Thielemans, Kris M.;Wölfel, Thomas;Boon, Thierry;van der Bruggen, Pierre
Référence The Journal of immunology, 174, 4, page (2404-2411)
Publication Publié, 2005-02
Référence The Journal of immunology, 174, 4, page (2404-2411)
Publication Publié, 2005-02
Article révisé par les pairs
Résumé : | Quantitative evaluation of T cell responses of patients receiving antitumoral vaccination with a protein is difficult because of the large number of possible HLA-peptide combinations that could be targeted by the response. To evaluate the responses of patients vaccinated with protein MAGE-3, we have developed an approach that involves overnight stimulation of blood T cells with autologous dendritic cells loaded with the protein, sorting by flow cytometry of the T cells that produce IFN-γ, cloning of these cells, and evaluation of the number of T cell clones that secrete IFN-γ upon stimulation with the Ag. An important criterion is that T cell clones must recognize not only stimulator cells loaded with the protein, but also stimulator cells transduced with the MAGE-3 gene, so as to exclude the T cells that recognize contaminants generated by the protein production system. Using this approach it is possible to measure T cell frequencies as low as 10 -6. We analyzed the frequencies of anti-vaccine CD4 T cells in five metastatic melanoma patients who had been injected with a MAGE-3 protein without adjuvant and showed evidence of tumor regression. Anti-MAGE-3 CD4 T cells were detected in one of the five patients. The frequency of the anti-MAGE-3 CD4 T cells was estimated at 1/60,000 of the CD4 T cells in postvaccination blood samples, representing at least an 80-fold increase in the frequency found before immunization. The frequencies of one anti-MAGE-3 CD4 T cell clonotype were confirmed by PCR analysis on blood lymphocytes. The 13 anti-MAGE-3 clones, which corresponded to five different TCR clonotypes, recognized the same peptide presented by HLA-DR1. |