RESUMO
Therapeutic success of TCR gene transfer to treat tumors depends on the ability of redirected T cells to become activated upon tumor recognition in vivo. Help provided by tumor-specific Th1 cells is reported to relieve T cells from an anergized state and to induce tumor regression. We recently demonstrated the ability to generate melanoma-specific Th1 cells by genetic introduction of both a CD8-dependent TCR and the CD8alpha coreceptor into CD4+ T cells. In this study, we analyzed a TCR that binds Ag independently of CD8, a property generally preferred to induce tumor-specific T cell responses, and addressed the contribution of CD8alpha following introduction into TCR-transduced CD4+ T cells. To this end, primary human CD4+ T cells were gene transferred with a high-avidity TCR, and were shown not only to bind peptide/MHC class I, but also to effectively kill Ag-positive tumor cells in the absence of CD8alpha. The introduction of CD8alpha up-regulates the tumor-specific production of TNF-alpha and IL-2 to some extent, but significantly down-regulates production of IL-4, IL-5, and IL-10 in CD4+ T cells. The introduction of a mutated cysteine motif in CD8alpha, which prevents its binding to LCK and linker for activation of T cells, did not adversely affect expression and T cell cytotoxicity, but counteracted the CD8alpha-mediated down-regulation of IL-4 and IL-5, but not IL-10. In conclusion, CD8alpha down-regulates the production of major Th2-type cytokines, in part mediated by LCK and/or linker for activation of T cells, and may induce differentiation of tumor-specific Th1 cells, which makes this coreceptor an interesting candidate to improve the clinical potential of TCR gene transfer to treat cancer.
Assuntos
Antígenos CD8/genética , Regulação para Baixo/imunologia , Técnicas de Transferência de Genes , Interleucina-10/antagonistas & inibidores , Interleucina-4/antagonistas & inibidores , Interleucina-5/antagonistas & inibidores , Melanoma/terapia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Motivos de Aminoácidos/genética , Antígenos CD8/fisiologia , Antígenos CD8/uso terapêutico , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Linhagem Celular , Linhagem Celular Tumoral , Cisteína/genética , Citotoxicidade Imunológica/genética , Regulação para Baixo/genética , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/metabolismo , Antígeno HLA-A2/imunologia , Antígeno HLA-A2/metabolismo , Humanos , Interleucina-10/biossíntese , Interleucina-4/biossíntese , Interleucina-5/biossíntese , Melanoma/genética , Melanoma/imunologia , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/metabolismo , Ligação Proteica/genética , Ligação Proteica/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Receptores de Antígenos de Linfócitos T alfa-beta/uso terapêutico , Células Th1/citologia , Células Th1/imunologia , Células Th1/metabolismo , Transdução Genética/métodos , Antígeno gp100 de MelanomaRESUMO
T cells require both primary and costimulatory signals for optimal activation. The primary Ag-specific signal is delivered by engagement of the TCR. The second Ag-independent costimulatory signal is mediated by engagement of the T cell surface costimulatory molecule CD28 with its target cell ligand B7. However, many tumor cells do not express these costimulatory molecules. We previously constructed phage display derived F(AB), G8, and Hyb3, Ab-based receptors with identical specificity but distinct affinities for HLA-A1/MAGE-A1, i.e., "TCR-like" specificity. These chimeric receptors comprised the FcepsilonRI-gamma signaling element. We analyzed whether linking the CD28 costimulation structure to it (gamma + CD28) could affect the levels of MHC-restricted cytolysis and/or cytokine production. Human scFv-G8(POS) T lymphocytes comprising the gamma + CD28 vs the gamma signaling element alone produced substantially more IL-2, TNF-alpha, and IFN-gamma in response to HLA-A1/MAGE-A1(POS) melanoma cells. Also a drastic increase in cytolytic capacity of scFv-G8(POS) T cells, equipped with gamma + CD28 vs the gamma-chain alone was observed.