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

ABSTRACT: 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 TGF? 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 TGF?/IL-6-mediated induction of Maf.

SUBMITTER: Giordano M

PROVIDER: S-EPMC4551351 | biostudies-literature | 2015 Aug

REPOSITORIES: biostudies-literature

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

Giordano Marilyn M Henin Coralie C Maurizio Julien J Imbratta Claire C Bourdely Pierre P Buferne Michel M Baitsch Lukas L Vanhille Laurent L Sieweke Michael H MH Speiser Daniel E DE Auphan-Anezin Nathalie N Schmitt-Verhulst Anne-Marie AM Verdeil Grégory G

The EMBO journal 20150702 15

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 overexpr ...[more]

PMID: 26139534

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Project description:In chronic viral infections such as HIV-1, HBV and HCV, CD8+ T cells may become ”exhausted”, characterized by progressive functional deficiency. Recently, the underlying mechanisms have been characterized by molecular profiling of virus-specific CD8+ T cells. In contrast, only little is known of self/tumor-specific CD8+ T cells from cancer patients, likely because they are much more difficult to assess. For the first time, we determined the molecular profile of human tumor-specific CD8+ T cells, upon sorting of Melan-A/MART-1 specific T cells directly ex vivo from 19 melanoma patients after vaccination with peptide and CpG. For comparison, we sorted protective T cells specific for the two herpes viruses EBV and CMV. In peripheral blood, we found multiple features of functional effector T cells, with only small but nevertheless significant differences between the three T cell populations, resulting in clean clustering according to antigen specificity. In contrast, Melan-A/MART-1 specific T cells obtained from tumor-infiltrated lymph nodes (TILNs) expressed multiple genes associated with T cell exhaustion, compatible with the known functional deficiencies of T cells in melanoma metastases. We show that individual T cells simultaneously expressed multiple inhibitory receptors implied in functional impairment. Together, the data indicate that in circulation, human tumor-specific T cells have the potential to become competent effector cells. In the tumor environment, however, T cells are exhausted and fail to control malignant disease. Our novel resource data identify mechanisms of functional T cell deficiency in melanoma patients, providing a rational basis for the improvement of immune therapy. 52 samples were measured in two sets of experiments. 4 self/tumor-specific samples from blood were replicated between the first and second sets. Set 1 (32 samples, all from blood): 13 naive samples, 10 EBV-specific samples, 9 self/tumor-specific samples. Set 2 (20 samples): 7 CMV-specific samples from blood, 7 self/tumor-specific samples from TILN, 6 self/tumor-specific samples from blood.

Project description:BackgroundGene expression profiling has revolutionized our ability to molecularly classify primary human tumors and significantly enhanced the development of novel tumor markers and therapies; however, progress in the diagnosis and treatment of melanoma over the past 3 decades has been limited, and there is currently no approved therapy that significantly extends lifespan in patients with advanced disease. Profiling studies of melanoma to date have been inconsistent due to the heterogeneous nature of this malignancy and the limited availability of informative tissue specimens from early stages of disease.Methodology/principle findingsIn order to gain an improved understanding of the molecular basis of melanoma progression, we have compared gene expression profiles from a series of melanoma cell lines representing discrete stages of malignant progression that recapitulate critical characteristics of the primary lesions from which they were derived. Here we describe the unsupervised hierarchical clustering of profiling data from melanoma cell lines and melanocytes. This clustering identifies two distinctive molecular subclasses of melanoma segregating aggressive metastatic tumor cell lines from less-aggressive primary tumor cell lines. Further analysis of expression signatures associated with melanoma progression using functional annotations categorized these transcripts into three classes of genes: 1) Upregulation of activators of cell cycle progression, DNA replication and repair (CDCA2, NCAPH, NCAPG, NCAPG2, PBK, NUSAP1, BIRC5, ESCO2, HELLS, MELK, GINS1, GINS4, RAD54L, TYMS, and DHFR), 2) Loss of genes associated with cellular adhesion and melanocyte differentiation (CDH3, CDH1, c-KIT, PAX3, CITED1/MSG-1, TYR, MELANA, MC1R, and OCA2), 3) Upregulation of genes associated with resistance to apoptosis (BIRC5/survivin). While these broad classes of transcripts have previously been implicated in the progression of melanoma and other malignancies, the specific genes identified within each class of transcripts are novel. In addition, the transcription factor NF-KB was specifically identified as being a potential "master regulator" of melanoma invasion since NF-KB binding sites were identified as consistent consensus sequences within promoters of progression-associated genes.Conclusions/significanceWe conclude that tumor cell lines are a valuable resource for the early identification of gene signatures associated with malignant progression in tumors with significant heterogeneity like melanoma. We further conclude that the development of novel data reduction algorithms for analysis of microarray studies is critical to allow for optimized mining of important, clinically-relevant datasets. It is expected that subsequent validation studies in primary human tissues using such an approach will lead to more rapid translation of such studies to the identification of novel tumor biomarkers and therapeutic targets.

Project description:In chronic viral infections such as HIV-1, HBV and HCV, CD8+ T cells may become M-bM-^@M-^]exhaustedM-bM-^@M-^], characterized by progressive functional deficiency. Recently, the underlying mechanisms have been characterized by molecular profiling of virus-specific CD8+ T cells. In contrast, only little is known of self/tumor-specific CD8+ T cells from cancer patients, likely because they are much more difficult to assess. For the first time, we determined the molecular profile of human tumor-specific CD8+ T cells, upon sorting of Melan-A/MART-1 specific T cells directly ex vivo from 19 melanoma patients after vaccination with peptide and CpG. For comparison, we sorted protective T cells specific for the two herpes viruses EBV and CMV. In peripheral blood, we found multiple features of functional effector T cells, with only small but nevertheless significant differences between the three T cell populations, resulting in clean clustering according to antigen specificity. In contrast, Melan-A/MART-1 specific T cells obtained from tumor-infiltrated lymph nodes (TILNs) expressed multiple genes associated with T cell exhaustion, compatible with the known functional deficiencies of T cells in melanoma metastases. We show that individual T cells simultaneously expressed multiple inhibitory receptors implied in functional impairment. Together, the data indicate that in circulation, human tumor-specific T cells have the potential to become competent effector cells. In the tumor environment, however, T cells are exhausted and fail to control malignant disease. Our novel resource data identify mechanisms of functional T cell deficiency in melanoma patients, providing a rational basis for the improvement of immune therapy. 52 samples were measured in two sets of experiments. 4 self/tumor-specific samples from blood were replicated between the first and second sets. Set 1 (32 samples, all from blood): 13 naive samples, 10 EBV-specific samples, 9 self/tumor-specific samples. Set 2 (20 samples): 7 CMV-specific samples from blood, 7 self/tumor-specific samples from TILN, 6 self/tumor-specific samples from blood.

Project description:BackgroundDeveloping analytical methodologies to identify biomarkers in easily accessible body fluids is highly valuable for the early diagnosis and management of cancer patients. Peripheral whole blood is a "nucleic acid-rich" and "inflammatory cell-rich" information reservoir and represents systemic processes altered by the presence of cancer cells.Methodology/principal findingsWe conducted transcriptome profiling of whole blood cells from melanoma patients. To overcome challenges associated with blood-based transcriptome analysis, we used a PAXgene™ tube and NuGEN Ovation™ globin reduction system. The combined use of these systems in microarray resulted in the identification of 78 unique genes differentially expressed in the blood of melanoma patients. Of these, 68 genes were further analyzed by quantitative reverse transcriptase PCR using blood samples from 45 newly diagnosed melanoma patients (stage I to IV) and 50 healthy control individuals. Thirty-nine genes were verified to be differentially expressed in blood samples from melanoma patients. A stepwise logit analysis selected eighteen 2-gene signatures that distinguish melanoma from healthy controls. Of these, a 2-gene signature consisting of PLEK2 and C1QB led to the best result that correctly classified 93.3% melanoma patients and 90% healthy controls. Both genes were upregulated in blood samples of melanoma patients from all stages. Further analysis using blood fractionation showed that CD45(-) and CD45(+) populations were responsible for the altered expression levels of PLEK2 and C1QB, respectively.Conclusions/significanceThe current study provides the first analysis of whole blood-based transcriptome biomarkers for malignant melanoma. The expression of PLEK2, the strongest gene to classify melanoma patients, in CD45(-) subsets illustrates the importance of analyzing whole blood cells for biomarker studies. The study suggests that transcriptome profiling of blood cells could be used for both early detection of melanoma and monitoring of patients for residual disease.

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

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

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