Role of Fas and granule exocytosis pathways in tumor-infiltrating T lymphocyte-induced apoptosis of autologous human lung-carcinoma cells.
We have isolated a cytotoxic T lymphocyte (CTL) clone, Heu161, that reacts specifically with the human autologous lung carcinoma cell line IGR-Heu. We first demonstrated that IGR-Heu lacked Fas-receptor expression and was resistant to CD95-induced apoptosis. To further elucidate the role of Fas in tumor immune surveillance, we have stably transfected IGR-Heu with a Fas-expression vector and isolated CD95-sensitive and -resistant clones. Our data indicated that the resistance of 2 selected Fas-transfected clones to CD95-mediated lysis correlated with down-regulation of caspase-8 or its lack of cleavage and subsequent activation. All Fas transfectants, either sensitive or resistant to anti-Fas agonistic antibody, were as efficiently lysed by the CTL clone as the parental cell line. In addition, neither anti-Fas-blocking antibody nor Fas-Fc molecule inhibited T-cell lysis of Fas-sensitive tumor clone. This cytotoxicity was extracellular Ca(2+)-dependent and abolished in the presence of EGTA, indicating that it was mainly granzyme-mediated. Interestingly, although the caspase inhibitor z-VAD-fmk had no effect on tumor-cell lysis, it efficiently blocked target DNA damage triggered by autologous CTLs via the granule exocytosis pathway, indicating that the latter event was caspase-dependent. The present results suggest that lung carcinoma-specific CTLs use mainly a granule exocytosis-dependent pathway to lyse autologous target cells and that these effectors are able to circumvent alteration of the Fas-triggered intracellular signalling pathway via activation of a caspase-independent cytoplasmic death mechanism.
Dorothee
G
G
Laboratoire Cytokines et Immunologie des Tumeurs Humaines, U487 INSERM, Institut Gustave Roussy, 39 rue Camille-Des-moulins, F-94895 Villejuif, France.
Ameyar
M
M
Bettaieb
A
A
Vergnon
I
I
Echchakir
H
H
Bouziane
M
M
Chouaib
S
S
Mami-Chouaib
F
F
D017209:Apoptosis; D015153:Blotting, Western; D018287:Carcinoma, Large Cell / Q000276:immunology / Q000473:pathology*; D002289:Carcinoma, Non-Small-Cell Lung / Q000276:immunology / Q000473:pathology*; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D003594:Cytoplasmic Granules / Q000378:metabolism; D005089:Exocytosis / Q000502:physiology*; D053222:Fas Ligand Protein; D005434:Flow Cytometry; D006801:Humans; D008175:Lung Neoplasms / Q000276:immunology / Q000473:pathology*; D016246:Lymphocytes, Tumor-Infiltrating / Q000276:immunology*; D008562:Membrane Glycoproteins / Q000502:physiology; D015166:Monitoring, Immunologic; D015398:Signal Transduction; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*; D014162:Transfection; D014407:Tumor Cells, Cultured; D019014:fas Receptor / Q000378:metabolism / Q000502:physiology*
C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D019014:fas Receptor; C505530:CASP8 protein, human; C506433:CASP9 protein, human; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases