Melanoma-associated fibroblasts impair CD8+ T cell function and modify expression of immune checkpoint regulators via increased arginase activity

Abstract

This study shows that melanoma-associated fibroblasts (MAFs) suppress cytotoxic T lymphocyte (CTL) activity and reveals a pivotal role played by arginase in this phenomenon. MAFs and normal dermal fibroblasts (DFs) were isolated from surgically resected melanomas and identified as Melan-A-/gp100-/FAP+ cells. CTLs of healthy blood donors were activated in the presence of MAF-and DF-conditioned media (CM). Markers of successful CTL activation, cytotoxic degranulation, killing activity and immune checkpoint regulation were evaluated by flow cytometry, ELISPOT, and redirected killing assays. Soluble mediators responsible for MAF-mediated effects were identified by ELISA, flow cytometry, inhibitor assays, and knock-in experiments. In the presence of MAF-CM, activated/non-naïve CTLs displayed dysregulated ERK1/2 and NF-κB signaling, impeded CD69 and granzyme B production, impaired killing activity, and upregulated expression of the negative immune checkpoint receptors TIGIT and BTLA. Compared to DFs, MAFs displayed increased amounts of VISTA and HVEM, a known ligand of BTLA on T cells, increased l-arginase activity and CXCL12 release. Transgenic arginase overexpression further increased, while selective arginase inhibition neutralized MAF-induced TIGIT and BTLA expression on CTLs. Our data indicate that MAF interfere with intracellular CTL signaling via soluble mediators leading to CTL anergy and modify immune checkpoint receptor availability via l-arginine depletion. Cellular and Molecular Life Sciences Barbara Érsek and Pálma Silló contributed equally to this work. Zoltán Pós and Krisztián Németh contributed equally to this work.

The Role of Cancer-Associated Fibroblasts in Tumor Progression

“…Ersek et al were able to determine that melanoma-associated fibroblasts (MAFs), or CAFs in melanoma cancer tissues identified by their FAP expression, impaired cytotoxic T lymphocyte function [ 42 ]. Their findings demonstrated that in the presence of MAF-conditioned media, cytotoxic T lymphocytes displayed dysregulated ERK1/2 and NF-κB signaling, impeded CD69 and granzyme B production, impaired killing activity, and upregulated expression of negative immune checkpoint receptors T cell immunoreceptor with Ig and ITIM domains (TIGIT) and B- and T- lymphocyte attenuator (BTLA).…”

In Vitro Suppression of T Cell Proliferation Is a Conserved Function of Primary and Immortalized Human Cancer-Associated Fibroblasts

“…Moreover, ECM remodeling by CAFs may impede trafficking of T cells leading to more restrained tumor immunity [ 45 ]. CAFs also suppress the function of T cells and inhibit their tumor cell killing capacity by limiting T cell infiltration into the tumor and inhibition of the T cells cytotoxic activity within the TME [ 46 , 47 , 48 , 49 ].…”

Influence of Tumor Microenvironment and Fibroblast Population Plasticity on Melanoma Growth, Therapy Resistance and Immunoescape

“…After antigen presentation, CAFs directly interact with activated CD8+ T cells, thus promoting T cell dysfunction and death via PD-L2 and FASL engagement [ 186 ]. Furthermore, Érsek et al demonstrated that MAFs can hamper cytotoxic T cell-mediated killing by altering ERK1/2 and NF-κB signalling pathways, impeding the expression of the early T cell activation marker CD69 and granzyme B production, and enhancing the expression of two potent negative regulators of T cell activity, T cell immunoreceptor with Ig and ITIM domains (TIGIT) and B and T lymphocyte attenuator (BTLA) [ 187 ]. As previously discussed, the reduction of glucose availability in the melanoma microenvironment is associated with the generation of a glucose-deficient and immunosuppressive milieu, where different cells compete for glucose uptake [ 3 ].…”

“…The increase of L-Arginine metabolism observed in melanoma and other human cancers, is involved in tumor progression, associated with neovascolarization and immune suppression induction [ 189 ]. In particular, L-arginine depletion in the TME induces T cell suppression facilitating the melanoma escape from immunosurveillance [ 3 , 187 ]. Of note, Ersek et al showed an increased L-arginase activity in MAFs.…”