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

Therapeutic Implications of the Immunoscore in Patients with Colorectal Cancer

International audienceFour decades were needed to progress from the first demonstration of the independent prognostic value of lymphocytes infiltration in rectal cancers to the first recommendation from the international guidelines for the use of a standardized immune assay, namely the “Immunoscore” (IS), to accurately prognosticate colon cancers beyond the TNM-system. The standardization process included not only the IS conceptualization, development, fine-tuning, and validation by a large international consortium, but also a demonstration of the robustness and reproducibility across the world and testing of international norms and their effects on the IS. This is the first step of a major change of paradigm that now perceives cancer as the result of contradicting driving forces, i.e., the tumor expansion and the immune response, interacting dynamically and influencing the prognosis and the response to therapies. This prompted us to evaluate and evidence the capacity of the tumor immune status, as reflected by the IS, to accurately predict chemotherapy responses in an international, randomized cohort study of colon cancer. Moreover, we developed a derived IS performed on initial diagnostic biopsies (ISB) to assess response levels to neoadjuvant therapies. In rectal cancer, ISB was positively correlated with the degree of histologic response to neoadjuvant chemoradiotherapy and identified - alone and even more accurately if combined with clinical data- patients eligible for a noninvasive strategy. Based on these results, we are currently setting up an international cohort for confirmation. The potential role of IS with immunotherapies must be anticipate

Prognostic association of FoxP3 regulatory T cells with tumor infiltrating CD8 cytotoxic T cells quantified on resected liver colorectal metastases (LCM)

Background: The adaptive Th1 immune response (CD3/CD8/CD45RO T-cells) is a strong prognostic factor for the survival of the patients after colorectal primary tumor and metastases resection. The prognostic impact of regulatory T-cells (FoxP3) in metastatic settings is less characterized. Methods: Metastatic colorectal patients undergoing curative liver surgery for all resected LCM were investigated. For this cohort, we previously reported the prognostic role of an immune score based (IS) for T (CD3/CD8/CD45RO) and B (CD20) cells infiltration . The FoxP3 and CD8 cells infiltrating the center (CT ) and the invasive margin (IM) of LCM were immunohistochemically stained and their densities quantified on whole slides with a dedicated image analysis software. The mean value of the 3 most infiltrated areas (0.64 mm²) was calculated. The CT and IM densities of FoxP3 and CD8 were classified into high (Hi) or low (Lo) according to the minimum p-value cut-off. The total number of Hi densities determined the IS : 0-2 Hi (low IS) or 3-4 Hi (high IS). Cumulative DFS/OS analyses were performed using the Kaplan-Meier estimator. The hazard ratio for OS/DFS comparing (IS 0-2 vs 3-4) was determined using univariate Cox regression and the significance by log-rank tests. Results: 294 LCM from 88 patients (Male/Female 1.1, mean 3.3/patient, synchronous/metachronous 5.8) were included. Inter-metastatic heterogeneity for FoxP3 and CD8 (CT/IM) is high and observed among all patients. For the least infiltrated LCM/patient: a high IS is prognostic for DFS (HR:0.31 95%IC 0.18-0.53; p = 0.0006) and OS (HR:0.29 95%IC 0.14-0.64; p = 0.001). Independently of CD8 cells, a high FoxP3 density in both regions (CT/IM) is not prognostic for DFS (HR:0.53 95%IC 0.28-0.97; p = 0.74) but for OS (HR:0.16 95%IC 0.02-1.17; p = 0.04). Conclusions: Regulatory T cells associated with cytotoxic T cells in both tumor regions (CT/IM) of the least infiltrated LCM/patient are highly prognostic after curative resection. These results confirm the important role of tumor regions and FoxP3 for modulating the tumor immune response

B cells (CD20+) associated to tumor infiltrating cytotoxic T-cells observed on resected liver colorectal metastases (LCM) are prognostic

Aim: Colorectal cancer infiltrating cytotoxic T-cells (CD8+ cells) are a strong prognostic factor for survival after primary tumor and metastases resection. The impact of B cells for prognosis is less characterized. Methods: Metastatic colorectal patients (pts) engaged for curative liver surgery with available FFPE blocks for all resected LCM, were included. The density of CD8+ and CD20+ cells in the metastasis (CT) and the invasive margin (IM) for all LCM was determined by whole-slide immunohistochemistry and quantified with dedicated image analysis software. The mean value of the 3 most infiltrated areas (0.8 mm2) was calculated. The densities of CD8 and CD20 (CT and IM regions) were classified into Hi or Lo according to cut-off values (minimal p-value approach). The total number of Hi densities was calculated to determine the immunoscore (IS) 0-2 Hi (low IS) or 3-4 Hi (high IS). For pts with multiple LCM, all LCM were quantified. The mean value of all densities, the least and the most infiltrated LCM/pt were analyzed. Cumulative DFS/OS analyses were performed using the Kaplan-Meier estimator. The hazard ratio (HR) for OS/DFS comparing (IS0-2 vs 3-4) was determined using univariate Cox regression and the significance by log-rank tests. Results: 294 LCM from 88 patients (M/F 1.1, mean 3.3/pt, synchr/metachr 5.8) were included. For the least infiltrated metastasis: a high IS is prognostic for DFS and OS. Independently of CD8 cells, a high CD20 density associated concomitantly with both regions (CT/IM) is prognostic for OS (HR: 0.36; p = 0.00004) but not for DFS (HR 0.58, p = 0.1). [...