Cooperative action of CD8 T lymphocytes and natural killer cells controls tumour growth under conditions of restricted T-cell receptor diversity

In mice expressing a transgenic T-cell receptor (TCR; TCRP1A) of DBA/2 origin with reactivity towards a cancer-germline antigen P1A, the number of TCRP1A CD8+ T cells in lymphoid organs is lower in DBA/2 than in B10.D2 or B10.D2(× DBA/2)F1 mice. This reduction results from haemopoietic cell autonomous differences in the differentiation of the major histocompatibility complex class I-restricted TCRP1A thymocytes controlled by DBA/2 versus B10.D2-encoded genes. We report here that the lower number of TCRP1A CD8+ T cells in DBA/2 mice correlated with their poor resistance to P1A-expressing mastocytoma solid tumours. Functional potency of CD8+ cytolytic T lymphocytes (CTL) from the above strains was not compromised, but their number after expansion appeared to be influenced by their genetic background. Intriguingly, non-transgenic DBA/2 mice resisted P1A+ tumours more efficiently despite poor representation of P1A-specific CTL. This was partly the result of their more heterogeneous TCR repertoire, including reactivity to non-P1A tumour antigens because mice that had rejected a P1A+ tumour became resistant to a P1A− variant of the tumour. Such ‘cross-resistance’ did not develop in the TCRP1A transgenic mice. Nonetheless, reconstitution of RAGº/º mice with TCRP1A CD8+ T cells, with or without CD4+ T cells, or exclusive representation of TCRP1A CD8+ T cells in RAGº/º TCRP1A transgenic mice efficiently resisted the growth of P1A-expressing tumours. Natural killer cells present at a higher number in RAGº/º mice also contributed to tumour resistance, in part through an NKG2D-dependent mechanism. Hence, in the absence of a polyclonal T-cell repertoire, precursor frequencies of natural killer cells and tumour-specific CTL affect tumour resistance

Distinct patterns of cytolytic T-cell activation by different tumour cells revealed by Ca2+ signalling and granule mobilization

International audienceCancer-germline genes in both humans and mice have been shown to encode antigens susceptible to targeting by cytotoxic CD8 T effector cells (CTL). We analysed the ability of CTL to kill different tumour cell lines expressing the same cancer-germline gene P1A (Trap1a). We previously demonstrated that CTL expressing a T-cell receptor specific for the P1A(35-43) peptide associated with H-2L(d), although able to induce regression of P1A-expressing P815 mastocytoma cells, were much less effective against P1A-expressing melanoma cells. Here, we analysed parameters of the in vitro interaction between P1A-specific CTL and mastocytoma or melanoma cells expressing similar levels of the P1A gene and of surface H-2L(d). The mastocytoma cells were more sensitive to cytolysis than the melanoma cells in vitro. Analysis by video-microscopy of early events required for target cell killing showed that similar patterns of increase in cytoplasmic Ca2+ concentration ([Ca2+]i) were induced by both types of P1A-expressing tumour cells. However, the use of CTL expressing a fluorescent granzyme B (GZMB-Tom) showed a delay in the migration of cytotoxic granules to the tumour interaction site, as well as a partially deficient GZMB-Tom exocytosis in response to the melanoma cells. Among surface molecules possibly affecting tumour-CTL interactions, the mastocytoma cells were found to express intercellular adhesion molecule-1, the ligand for LFA-1, which was not detected on the melanoma cells

CD8 T cell help for innate antitumor immunity.

International audienceInnate immunity is considered to initiate adaptive antitumor responses. We demonstrate that monoclonal CD8 T lymphocytes reactive to tumor Ag P1A on P815 mastocytoma cells provide essential "help" to NK cells for rejection of P1A-deficient tumors. RAG-deficient mice have normal NK cells but do not reject either tumor. Reconstitution of these mice with P1A-specific T cells conferred resistance to both P1A-expressing and -deficient tumor cells provided they were present at the same site. Elimination of Ag-negative tumor variants required both activated T and NK cells. Gene expression profiling of NK cells infiltrating P1A-positive tumors in mice with specific CD8 T cells demonstrated an activated effector phenotype. However, CD8 T cell help to NK cells appeared ineffective for P1A-negative variants separated from the P1A-positive tumor. Local tumor Ag-specific T cell-NK cell collaboration results in the elimination of tumor cells whether they express or not the T cell tumor Ag epitope, thus containing the emergence of tumor escape variants before metastasis

Minimal tolerance to a tumor antigen encoded by a cancer-germline gene

Central tolerance toward tissue-restricted Ags is considered to rely on ectopic expression in the thymus, which was also observed for tumor Ags encoded by cancer-germline genes. It is unknown whether endogenous expression shapes the T cell repertoire against the latter Ags and explains their weak immunogenicity. We addressed this question using mouse cancer-germline gene P1A, which encodes antigenic peptide P1A(35-43) presented by H-2L(d). We made P1A-knockout (P1A-KO) mice and asked whether their anti-P1A(35-43) immune responses were stronger than those of wild-type mice and whether P1A-KO mice responded to other P1A epitopes, against which wild-type mice were tolerized. We observed that both types of mice mounted similar P1A(35-43)-specific CD8 T cell responses, although the frequency of P1A(35-43)-specific CD8 T cells generated in response to P1A-expressing tumors was slightly higher in P1A-KO mice. This higher reactivity allowed naive P1A-KO mice to reject spontaneously P1A-expressing tumors, which progressed in wild-type mice. TCR-Vβ usage of P1A(35-43)-specific CD8 cells was slightly modified in P1A-KO mice. Peptide P1A(35-43) remained the only P1A epitope recognized by CD8 T cells in both types of mice, which also displayed similar thymic selection of a transgenic TCR recognizing P1A(35-43). These results indicate the existence of a minimal tolerance to an Ag encoded by a cancer-germline gene and suggest that its endogenous expression only slightly affects diversification of the T cell repertoire against this Ag

Molecular profiling of CD8 T cells in autochthonous melanoma identifies Maf as driver of exhaustion.

T cells infiltrating neoplasms express surface molecules typical of chronically virus-stimulated T cells, often termed "exhausted" T cells. We compared the transcriptome of "exhausted" CD8 T cells infiltrating autochthonous melanomas to those of naïve and acutely stimulated CD8 T cells. Despite strong similarities between transcriptional signatures of tumor- and virus-induced exhausted CD8 T cells, notable differences appeared. Among transcriptional regulators, Nr4a2 and Maf were highly overexpressed in tumor-exhausted T cells and significantly upregulated in CD8 T cells from human melanoma metastases. Transduction of murine tumor-specific CD8 T cells to express Maf partially reproduced the transcriptional program associated with tumor-induced exhaustion. Upon adoptive transfer, the transduced cells showed normal homeostasis but failed to accumulate in tumor-bearing hosts and developed defective anti-tumor effector responses. We further identified TGFβ and IL-6 as main inducers of Maf expression in CD8 T cells and showed that Maf-deleted tumor-specific CD8 T cells were much more potent to restrain tumor growth in vivo. Therefore, the melanoma microenvironment contributes to skewing of CD8 T cell differentiation programs, in part by TGFβ/IL-6-mediated induction of Maf