Regulation of T lymphocyte function in tumors
We have recently discovered a new type of anergy of human CD8 T cells, which is observed on tumor-infiltrating lymphocytes. We are analyzing the mechanism of this type of anergy and we look for agents that reverse this anergy. The analysis of the T cell responses of melanoma patients vaccinated against tumor antigens has led us to consider the possibility that the limiting factor for therapeutic success is not the intensity of the anti-vaccine response but the degree of anergy presented by intratumoral lymphocytes. We therefore intend to pursue clinical trials involving the use of these agents in combination with anti-tumoral vaccination.
A scenario to explain the low level of clinical responses in vaccinated patients
The identification of specific tumor antigens recognized by T lymphocytes on human cancer cells has elicited numerous clinical trials involving vaccination of tumor-bearing cancer patients with defined tumor antigens. These treatments have shown a low clinical efficacy. Among metastatic melanoma patients, about 5% show a complete or partial clinical response following vaccination, whereas an additional 10% show some evidence of tumor regression without clear clinical benefit. We believe that progress depends on understanding why a small minority of patients shows tumor regression following vaccination, whereas the majority does not.
Recent analyses of the T cell responses of melanoma patients has led us to propose the following scenario. Most melanoma patients produce a spontaneous T cell response against melanoma tumor antigens at a relatively early stage of the disease (primary tumor or early metastatic tumor).
Our working hypothesis:
T cells generated by the vaccine succeed in reversing the local immunosuppression
These T cells can eliminate some tumors at an early stage, but often they do not succeed in eliminating the tumor and they become anergic. Thus, the tumors of the patients about to receive the vaccine, already contain anergic T cells directed against tumor antigens. Presumably this anergy is maintained by immunosuppressive factors present in the tumor. A few patients show tumor regression following vaccination because some T cells generated by the vaccine penetrate inside the tumor, attack some tumor cells and succeed in reversing the local immunosuppression, possibly by releasing cytokines or chemokines. This reactivates the anergic T cells located inside the tumor and elicits new anti-tumoral T cells. These T cells then proceed to the elimination of the tumor cells. Most patients do not show tumor regression because the few anti-vaccine T cells that reach the tumor are immediately overwhelmed by the immunosuppressive environment.
Accordingly, our working hypothesis is that the crucial difference between the responding and the non-responding patients is not the intensity of their direct T cell response to the vaccine but the intensity of the immunosuppression inside the tumor. It is therefore important to know which immunosuppressive mechanisms operate in human tumors.
A new mechanism causing anergy of human tumor-infiltrating lymphocytes
We have identified a novel mechanism causing anergy of human CD8 cytolytic T lymphocytes (CTL), including tumor-infiltrating lymphocytes. This type of anergy appears to occur transiently during the normal stimulation cycle of T lymphocytes.
We observed that, a few days after antigen stimulation, CTL clones lose the capacity to secrete cytokines and in some case the cytolytic activity. These functions are recovered gradually and are usually completely restored after two weeks. TCRs and CD8 co-receptors were co-localized at the cell surface of functional CTL but, on the contrary, distant at the cell surface of non-functional CTL.
TCR and CD8 do not co-localize on recently stimulated CTL without effector functions
TCR and CD8 do not co-localize on tumor-infiltrating lymphocytes
Human CD8 tumor-infiltrating T lymphocytes were isolated from tumor ascites or solid tumors and compared with T lymphocytes from blood donors. TCR were observed to be distant from CD8 on the cell surface of tumor-infiltrating lymphocytes, whereas TCR and CD8 co-localized on blood T lymphocytes. The tumor-infiltrating lymphocytes were anergic, being unable to secrete INF-γ or other cytokines after non-specific stimulation with anti-CD3 and anti-CD28 antibodies.
Glycoprotein-galectin lattices restrain mobility of TCR
On the basis of the work of other groups, we hypothesized that the absence of TCR-CD8 co-localization at the cell surface of anergic T cells is due to the loss of mobility of the TCR, which is trapped in a lattice of galectin-3. The presence of galectin-3 in ascites and solid tumors has been shown in many studies. To test this hypothesis, tumor-infiltrating lymphocytes were incubated with N-acetyllactosamine, a disaccharide ligand of galectin-3. This treatment restored the TCR-CD8 co-localization and the capacity to secrete IFN-γ and other cytokines after stimulation.
Treatment of tumor-infitrating lymphocytes with a galectin ligand reverses anergy
Towards a clinical trial combining vaccination and galectin-binding polysaccharides
These observations indicate that ex vivo human tumor-infiltrating lymphocytes can recover their effector functions with galectin ligands and suggest that treatment of cancer patients with galectin ligands could correct the anergy of tumor-infiltrating lymphocytes. It is possible that peptide vaccination combined with local injection of a galectin ligand will be more effective at producing tumor regression than vaccination alone. We have recently identified a polysaccharide already approved for clinical use that was more efficient than N-acetyllactosamine to correct the anergy of CTL clones and human tumor-infiltrating lymphocytes. We therefore intend to launch a clinical trial combining peptide vaccination and injections of galectin-binding polysaccharides in melanoma tumor-bearing metastatic patients.
Further reading
Demotte N., Stroobant V., Courtoy P.J., Van der Smissen P., Colau D., Luescher I.F., Hivroz C., Nicaise J., Squifflet J.-L., Mourad M., Godelaine D., Boon T. and van der Bruggen P. 2008. Restoring the association of the T cell receptor with CD8 reverses anergy in human tumor-infiltrating lymphocytes. Immunity, 28: 414-424.
Lurquin C., Lethé B., Corbière V., Théate I., van Baren N., Coulie P.G. and Boon T. 2005. Contrasting frequencies of anti-tumor and anti-vaccine T cells in metastases of a melanoma patient vaccinated with a MAGE tumor antigen. J. Exp. Med., 201: 249-257.
To know more... (pdf chapter of the last de Duve Institute report)
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