Multi-Institutional Evaluation of Pathologists' Assessment Compared to Immunoscore.

BACKGROUND The Immunoscore (IS) is a quantitative digital pathology assay that evaluates the immune response in cancer patients. This study reports on the reproducibility of pathologists' visual assessment of CD3+- and CD8+-stained colon tumors, compared to IS quantification. METHODS An international group of expert pathologists evaluated 540 images from 270 randomly selected colon cancer (CC) cases. Concordance between pathologists' T-score, corresponding hematoxylin-eosin (H&E) slides, and the digital IS was evaluated for two- and three-category IS. RESULTS Non-concordant T-scores were reported in more than 92% of cases. Disagreement between semi-quantitative visual assessment of T-score and the reference IS was observed in 91% and 96% of cases before and after training, respectively. Statistical analyses showed that the concordance index between pathologists and the digital IS was weak in two- and three-category IS, respectively. After training, 42% of cases had a change in T-score, but no improvement was observed with a Kappa of 0.465 and 0.374. For the 20% of patients around the cut points, no concordance was observed between pathologists and digital pathology analysis in both two- and three-category IS, before or after training (all Kappa < 0.12). CONCLUSIONS The standardized IS assay outperformed expert pathologists' T-score evaluation in the clinical setting. This study demonstrates that digital pathology, in particular digital IS, represents a novel generation of immune pathology tools for reproducible and quantitative assessment of tumor-infiltrated immune cell subtypes

Caractérisation des lymphocytes T gamma delta et de l'expression de BTN3A dans le cancer colorectal

Γδ T cells are a sub-population of T cells, with innate and adaptive immunity functions. They are activated via a non-MHC Class I antigen presentation and have a potential anti-tumoral activity. Indeed, the human Vγ9Vδ2 T cells recognize a complex of heterodimerized butyrophilin (BTN) proteins, involving BTN3A1 and 2A1 proteins. Thus, γδ T cells are considered as potential targets for immunotherapy. Several strategies exist to potentiate their antitumoral activity. One of them is to use an antibody against BTN3A protein to selectively activated Vγ9Vδ2 T cell. Imcheck Therapeutics proposed an anti-BTN3A, already tested in clinical trial phase I/IIa. My work, in collaboration with Imcheck Therapeutics, was to assess, on colorectal cancer, γδ T cell infiltration and phenotypes, to evaluate BTN3A protein expressions and their impact on tumor development and patient survival. We worked with two cohorts of CRC patients. Thanks to a multiplexed immunostaining technology (Brightplex®, HalioDx) and a digital pathology analysis, we determined γδ T cell, and subtypes, infiltration with a spatial information. γδ T cells infiltrate tumor glands but densities are higher in the stroma zone. γδ T cells, expressing PD1, have an higher density in the invasive margin compare to tumor center. In contrast, cytotoxic γδ T cells have the same density between invasive margine and tumor center. γδ and Vγ9 T cell densities inversely correlate with tumor development. Moreover, a higher density of Vγ9 T cell is associated with a good pronostic from patient with CRC. These datas suggested that a detection of Vγ9 T cell by immunohistochemistry could be a pronostic marker for patient with CRC.Les lymphocytes T γδ sont une sous population de lymphocyte T, avec une réponse immunitaire innée et adaptative. Dans le cancer, un sous-type particulier de lymphocytes T γδ est connu pour sa réponse antitumorale : les lymphocytes T Vγ9Vδ2. Leur activation se fait par la reconnaissance d’un complexe de protéines butyrophilines, BTN3A1 et 2A1, formé en une conformation précise après fixation de résidu de la voie du mévalonate. Ces cellules sont de nouvelles cibles thérapeutiques. Mes travaux, réalisés en collaboration avec Imcheck Therapeutics, ont pour but de caractériser les lymphocytes T γδ dans le cancer colorectal (CCR). Et d’évaluer l’expression de BTN3A. Ainsi que d’estimer l’impact des lymphocytes γδ sur le développement tumoral et la survie des patients. Ces travaux ont été faits sur des cohortes de patients atteints d’un CCR. Grâce à une méthode d’immunohistochimie multiplexée couplée à une analyse numérique, nous avons analysé l’infiltrat des lymphocytes T γδ avec une information spatiale. Les lymphocytes T γδ infiltrent les glandes tumorales. Cependant, leur densité est plus élevée dans le stroma. La densité en lymphocytes T γδ exprimant PD1 est plus importante en marge de la tumeur que dans le centre. Alors que des lymphocytes T γδ cytotoxiques ont la même densité dans le centre et en marge de la tumeur. L’infiltrat en lymphocyte T γδ et Vγ9 est inversement corrélé avec évolution tumorale. Un fort infiltrat en lymphocyte T Vγ9 est lié à un meilleur pronostic chez les patients atteints d’un CCR. Ces données suggèrent que la détection des lymphocytes T Vγ9 par immunohistochimie permet d’aider à la définition du pronostic chez les patients avec un CCR

Estimating the population abundance of tissue-infiltrating immune and stromal cell populations using gene expression

International audienceWe introduce the Microenvironment Cell Populations-counter (MCP-counter) method, which allows the robust quantification of the absolute abundance of eight immune and two stromal cell populations in heterogeneous tissues from transcriptomic data. We present in vitro mRNA mixture and ex vivo immunohistochemical data that quantitatively support the validity of our method's estimates. Additionally, we demonstrate that MCP-counter overcomes several limitations or weaknesses of previously proposed computational approaches. MCP-counter is applied to draw a global picture of immune infiltrates across human healthy tissues and non-hematopoietic human tumors and recapitulates microenvironment-based patient stratifications associated with overall survival in lung adenocarcinoma and colorectal and breast cancer

Immune evasion before tumour invasion in early lung squamous carcinogenesis

Early detection and treatment are critical for improving the outcome of patients with cancer1. Understanding the largely uncharted biology of carcinogenesis requires deciphering molecular processes in premalignant lesions, and revealing the determinants of the intralesional immune reaction during cancer development. The adaptive immune response within tumours has previously been shown to be strongest at the earliest stage of carcinoma2,3. Here we show that immune activation and immune escape occur before tumour invasion, and reveal the relevant immune biomarkers of the pre-invasive stages of carcinogenesis in the lung. We used gene-expression profiling and multispectral imaging to analyse a dataset of 9 morphological stages of the development of lung squamous cell carcinoma, which includes 122 well-annotated biopsies from 77 patients. We identified evolutionary trajectories of cancer and immune pathways that comprise (1) a linear increase in proliferation and DNA repair from normal to cancerous tissue; (2) a transitory increase of metabolism and early immune sensing, through the activation of resident immune cells, in low-grade pre-invasive lesions; (3) the activation of immune responses and immune escape through immune checkpoints and suppressive interleukins from high-grade pre-invasive lesions; and, ultimately, (4) the activation of the epithelial–mesenchymal transition in the invasive stage of cancer. We propose that carcinogenesis in the lung involves a dynamic co-evolution of pre-invasive bronchial cells and the immune response. These findings highlight the need to develop immune biomarkers for early detection as well as immunotherapy-based chemopreventive approaches for individuals who are at high risk of developing lung cancer.SCOPUS: le.jinfo:eu-repo/semantics/publishe

Evolution of Metastases in Space and Time under Immune Selection.

We examined how the immune microenvironment molds tumor evolution at different metastatic organs in a longitudinal dataset of colorectal cancer. Through multiplexed analyses, we showed that clonal evolution patterns during metastatic progression depend on the immune contexture at the metastatic site. Genetic evidence of neoantigen depletion was observed in the sites with high Immunoscore and spatial proximity between Ki67 tumor cells and CD3 cells. The immunoedited tumor clones were eliminated and did not recur, while progressing clones were immune privileged, despite the presence of tumor-infiltrating lymphocytes. Characterization of immune-privileged metastases revealed tumor-intrinsic and tumor-extrinsic mechanisms of escape. The lowest recurrence risk was associated with high Immunoscore, occurrence of immunoediting, and low tumor burden. We propose a parallel selection model of metastatic progression, where branched evolution could be traced back to immune-escaping clones. The findings could inform the understanding of cancer dissemination and the development of immunotherapeutics

Multi-Institutional Evaluation of Pathologists Assessment Compared to Immunoscore.

BACKGROUND: The Immunoscore (IS) is a quantitative digital pathology assay that evaluates the immune response in cancer patients. This study reports on the reproducibility of pathologists visual assessment of CD3+- and CD8+-stained colon tumors, compared to IS quantification. METHODS: An international group of expert pathologists evaluated 540 images from 270 randomly selected colon cancer (CC) cases. Concordance between pathologists T-score, corresponding hematoxylin-eosin (H&amp;E) slides, and the digital IS was evaluated for two- and three-category IS. RESULTS: Non-concordant T-scores were reported in more than 92% of cases. Disagreement between semi-quantitative visual assessment of T-score and the reference IS was observed in 91% and 96% of cases before and after training, respectively. Statistical analyses showed that the concordance index between pathologists and the digital IS was weak in two- and three-category IS, respectively. After training, 42% of cases had a change in T-score, but no improvement was observed with a Kappa of 0.465 and 0.374. For the 20% of patients around the cut points, no concordance was observed between pathologists and digital pathology analysis in both two- and three-category IS, before or after training (all Kappa &lt; 0.12). CONCLUSIONS: The standardized IS assay outperformed expert pathologists T-score evaluation in the clinical setting. This study demonstrates that digital pathology, in particular digital IS, represents a novel generation of immune pathology tools for reproducible and quantitative assessment of tumor-infiltrated immune cell subtypes

Clinical Performance of the Consensus Immunoscore in Colon Cancer in the Asian Population from the Multicenter International SITC Study.

In this study, we evaluated the prognostic value of Immunoscore in patients with stage I-III colon cancer (CC) in the Asian population. These patients were originally included in an international study led by the Society for Immunotherapy of Cancer (SITC) on 2681 patients with AJCC/UICC-TNM stages I-III CC. CD3+ and cytotoxic CD8+ T-lymphocyte densities were quantified in the tumor and invasive margin by digital pathology. The association of Immunoscore with prognosis was evaluated for time to recurrence (TTR), disease-free survival (DFS), and overall survival (OS). Immunoscore stratified Asian patients (n = 423) into different risk categories and was not impacted by age. Recurrence-free rates at 3 years were 78.5%, 85.2%, and 98.3% for a Low, Intermediate, and High Immunoscore, respectively (HR[Low-vs-High] = 7.26 (95% CI 1.75-30.19); = 0.0064). A High Immunoscore showed a significant association with prolonged TTR, OS, and DFS ( &lt; 0.05). In Cox multivariable analysis stratified by center, Immunoscore association with TTR was independent (HR[Low-vs-Int+High] = 2.22 (95% CI 1.10-4.55) = 0.0269) of the patient's gender, T-stage, N-stage, sidedness, and MSI status. A significant association of a High Immunoscore with prolonged TTR was also found among MSS (HR[Low-vs-Int+High] = 4.58 (95% CI 2.27-9.23); ≤ 0.0001), stage II (HR[Low-vs-Int+High] = 2.72 (95% CI 1.35-5.51); = 0.0052), low-risk stage-II (HR[Low-vs-Int+High] = 2.62 (95% CI 1.21-5.68); = 0.0146), and high-risk stage II patients (HR[Low-vs-Int+High] = 3.11 (95% CI 1.39-6.91); = 0.0055). A High Immunoscore is significantly associated with the prolonged survival of CC patients within the Asian population