Human colorectal cancer cells induce T-cell death through release of proapoptotic microvesicles: role in immune escape.
OBJECTIVE
Normal and neoplastic cells release microvesicles, whose effects on the immune system still need to be elucidated. Because human colorectal cancer cells are hypothesized to escape immune recognition by expressing proapoptotic molecules, we investigated whether microvesicles bearing Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand and inducing apoptosis of activated T cells are secreted by colorectal cancer cells both in vitro and in affected patients.


METHODS
Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand expression were analyzed in colorectal cancer cells and purified microvesicles by flow cytometry, Western blotting, and immunoelectron microscopy. Microvesicle tumor origin was assessed through simultaneous detection of lysosomal (CD63) and adenocarcinoma (carcinoembryonic antigen) markers. Proapoptotic activity of microvesicles was evaluated by annexin V/propidium iodide staining and caspase activation in T cells, including CD8+ T lymphocytes from colorectal cancer patients.


RESULTS
Colorectal cancer cells showed a granular pattern of tumor necrosis factor-related apoptosis-inducing ligand and Fas ligand expression, suggesting a secretory behavior. These proapoptotic molecules were detected on isolated microvesicles, together with class I HLA, CD63, and carcinoembryonic antigen. Microvesicles induced Fas ligand-mediated and tumor necrosis factor-related apoptosis-inducing ligand-mediated apoptosis of activated CD8+ T cells generated from colorectal cancer patients. Microvesicles with comparable phenotypes and functions were found in plasma from patients with advanced disease, whereas vesicular structures expressing Fas ligand and tumor necrosis factor-related apoptosis-inducing ligand were also detected in colorectal cancer specimens.


CONCLUSIONS
These data show that colorectal cancer induces T-cell apoptosis through the release of Fas ligand-bearing and tumor necrosis factor-related apoptosis-inducing ligand-bearing microvesicles both in vitro and in vivo. This mechanism of immune escape has potential implications as a prognostic factor and could be targeted for the development of new antitumor therapies in colorectal cancer patients.
Huber
Veronica
V
Unit of Immunotherapy of Human Tumors, Istituto Nazionale Tumori, Milan, Italy.
Fais
Stefano
S
Iero
Manuela
M
Lugini
Luana
L
Canese
Paola
P
Squarcina
Paola
P
Zaccheddu
Annamaria
A
Colone
Marisa
M
Arancia
Giuseppe
G
Gentile
Massimo
M
Seregni
Ettore
E
Valenti
Roberta
R
Ballabio
Giuseppina
G
Belli
Filiberto
F
Leo
Ermanno
E
Parmiani
Giorgio
G
Rivoltini
Licia
L
D017209:Apoptosis; D051017:Apoptosis Regulatory Proteins; D015179:Colorectal Neoplasms / Q000276:immunology* / Q000503:physiopathology*; D022162:Cytoplasmic Vesicles; D053222:Fas Ligand Protein; D006801:Humans; D008562:Membrane Glycoproteins / Q000096:biosynthesis* / Q000276:immunology / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology*; D053221:TNF-Related Apoptosis-Inducing Ligand; D014407:Tumor Cells, Cultured; D019139:Tumor Escape / Q000276:immunology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000276:immunology / Q000378:metabolism
D051017:Apoptosis Regulatory Proteins; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha