Regulatory T-Cells Induce an Immune Escape in a Murine Model of Primary Intraocular B-Cell Lymphoma

Abstract

Purpose: : The role of regulatory T cells (Tregs) in the immune evasion of tumors is usually prominent. The aim of this study was to assess the presence and role of Tregs in a murine model of primary intraocular B-cell lymphoma (PIOL).

Methods: : We used a syngeneic model of PIOL developed in immunocompetent BALB/c mice by injecting a lymphomatous B-cell line in the posterior chamber. Immunohistochemistry and flow cytometric analysis have been performed to study the tumor growth and the immune infiltrate. Cytometric beads array was used to characterize the polarization of infiltrating T-lymphocytes (TILs). Depletion of Tregs was achieved using intraperitoneal injection of anti-CD25 mAb (PC61). Infiltrating T-lymphocytes were restimulated in vivo and in vitro by anti-CD3/CD28 mAbs.

Results: : Tregs were recruited in PIOL eyes (16.2% of living cells) compared to control eyes (1.2%, p<0.001). The number of infiltrating Tregs was correlated with tumor burden (r²=0.76), and inversely correlated with CD4⁺ T-cells (r²=-0.79). Recruitment of Tregs was also observed in the spleen of mice bearing tumor (2.9%) compared to controls (2.1%, p<0.001), but not in the draining lymph nodes. Depletion of Tregs resulted in a significant decrease of tumor burden (53.8% in control mice compared to 37.7% in PC61 mice, p=0.03), and an increase of the immune infiltrate. Eventhough the number of tumor cells decreased and the immune response quantitatively increased, tumor growth could not be abrogated. In vivo restimulation of TILs was performed with intravitreal injection of anti-CD3/CD28 mAbs at day 6. This resulted in decrease of tumor burden (62.9% to 50.6%, p=0.03).

Conclusions: : Several mechanisms can contribute to the tolerance towards the tumor. In our model, we demonstrated that Tregs are recruited in the tumor microenvironment. Tregs limit the recruitement of CD4+ T-cells in the eyes and are thought to inhibit their effector functions. TILs can be reactivated in vitro and in vivo to enhance the anti-tumoral immune response, but tumor growth can not be abrogated. The mechanisms by which Tregs are generated could be similar to ACAID, yet in the posterior chamber. Several escape mechanisms are thus responsible for the tolerance to the tumor in the eye. Future therapeutic strategies will have to combine tools targeting both the tumor cells and its microenvironment.

Keywords: tumors • immune tolerance/privilege • immunomodulation/immunoregulation