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

Etude de la réponse immunitaire cellulaire T dans les cancers colorectaux à instabilité microsatellitaire en vue du développement de nouvelles stratégies d'immunothérapies

ROUEN-BU Sciences (764512102) / SudocSudocFranceF

License to kill: microsatellite instability and immune contexture

International audienceColorectal cancers (CRCs) with microsatellite instability (MSI) are due to a defect in the DNA mismatch repair (MMR) system resulting in an accumulation of frame-shift mutations. They are characterized by a tumor microenvironment richer in cytotoxic CD8 T-cells (CTLs) and a better prognosis compared to microsatellite stable (MSS) CRCs. The mechanisms by which defective MMR system may influence tumor-infiltrating immune cells and their impact on patient survival were still unclear. Thus, we performed a comprehensive analysis of MSI colorectal tumors. We found that the numbers of frame-shift mutations potentially resulting in neo-epitopes were positively correlated to the density of tumor infiltrating CD8 T-cells but were lower than expected at random. We also evidenced that MSI patients could naturally harbor CTLs targeting frame-shift mutation-derived antigens. This favors the hypothesis of an active immunosurveillance in MSI colorectal tumors leading to the genetic evidence of an immunoediting. To evaluate the link between MSI tumor immune contexture and prognosis, we took advantage of a standardized assay that we developed to quantify tumor-infiltrating T-cells, the Immunoscore. Multivariate analyses revealed an advantage of Immunoscore over MSI in predicting recurrence and survival. Our data suggests that the prognostic value of MSI could be attributed to major underlying differences of infiltrating immune cells. Immunotherapeutic treatments, that are more efficient in patients with a preexisting anti-tumor immunity, were approved in MSI patients following successful clinical trials. We suggest that the Immunoscore could be used not only for colorectal tumor prognosis but also for predicting responses to immunotherapies

Regulatory T lymphocytes are associated with less aggressive histologic features in microsatellite-unstable colorectal cancers.

BACKGROUND: Colorectal cancers (CRCs) with microsatellite instability (MSI) are associated with a good prognosis and a high density of tumor-infiltrating lymphocytes (TILs). We have undertaken to determine the link between TIL densities and MSI CRC histologic features. PATIENTS AND METHODS: Using tissue microarrays, T-cell sub-population infiltration, including T cells (CD3), cytotoxic T cells (CD8) and regulatory T cells (FoxP3) were studied in 86 MSI CRCs. We separately analyzed TILs of the stromal and epithelial compartments in the tumor center, the tumoral invasion margin and associated normal tissue. RESULTS: For FoxP3+ TIL density in the tumor center stromal compartment, we found a strong negative correlation with T4 stage (p = 0.01), node invasion (p<0.001) and VELIPI (vascular emboli, lymphatic invasion and perinervous invasion) criteria (p = 0.002). CONCLUSION: The strong correlation between regulatory T cell density and the absence of VELIPI criteria suggests that this sub-group of T cells is preferentially associated with less invasive tumors

Correlation between FoxP3+ tumor-infiltrating lymphocyte (TIL) density in stromal compartment of tumor center and clinicopathological variables.

<p>TILs: tumor-infiltrating lymphocytes, VELIPI: vascular emboli or lymphatic invasion or perineural invasion.</p

Determination of tumor-infiltrating lymphocyte density.

<p>Examples of tumor center chips with CD3 (A), CD8 (B) and FoxP3 (C) staining. Left panels are examples of high TIL densities and right panels are examples of low TIL densities. The stroma is circled in red, the epithelium in blue, and black areas are excluded. The marked areas that are taken into account to calculate the lymphocyte infiltrate density are circled in yellow. All data were acquired with Chip’s N Cheap TMA analysis program.</p

Percentages of tumor-infiltrating lymphocytes in the different areas studied.

<p>The bar charts represent the percentages of surfaces marked by CD3, CD8 and FoxP3 immunostaining in the stromal (stro) and epithelial (ep) compartments, in the tumor center (TC) and the invasion front (IF) tissues. Standard errors are given by the bars. CD3+ TIL densities were respectively, 2.2±2.5% in tumor center epithelium compartment, 6.4±5.1% in tumor center stromal compartment, 2.3±2.5% in invasion front epithelium compartment, 7.2±6.1% in invasion front stromal compartment. CD8+ TIL densities were respectively, 1.52±2.0% in tumor center epithelium compartment, 2.9±2.7% in tumor center stromal compartment, 1.8±2.4% in invasion front epithelium compartment, 3.3±3.0% in invasion front stromal compartment. FoxP3+ TIL densities were respectively, 0.13±0.12% in tumor center epithelium compartment, 0.59±0.53% in tumor center stromal compartment, 0.14±0.14% in invasion front epithelium compartment, 0.63±0.58% in invasion front stromal compartment.</p

Table <b>1.</b> Patient and tumors characteristics.

<p>n: number of patients, VELIPI: vascular emboli or lymphatic invasion or perineural invasion.</p

Correlation between Density of CD8+ T-cell Infiltrate in Microsatellite Unstable Colorectal Cancers and Frameshift Mutations: A Rationale for Personalized Immunotherapy

International audienceColorectal cancers with microsatellite instability (MSI) represent 15% of all colorectal cancers, including Lynch syndrome as the most frequent hereditary form of this disease. Notably, MSI colorectal cancers have a higher density of tumor-infiltrating lymphocytes (TIL) than other colorectal cancers. This feature is thought to reflect the accumulation of frameshift mutations in sequences that are repeated within gene coding regions, thereby leading to the synthesis of neoantigens recognized by CD8(+) T cells. However, there has yet to be a clear link established between CD8(+) TIL density and frameshift mutations in colorectal cancer. In this study, we examined this link in 103 MSI colorectal cancers from two independent cohorts where frameshift mutations in 19 genes were analyzed and CD3(+), CD8(+), and FOXP3(+) TIL densities were quantitated. We found that CD8(+) TIL density correlated positively with the total number of frameshift mutations. TIL densities increased when frameshift mutations were present within the ASTE1, HNF1A, or TCF7L2 genes, increasing even further when at least one of these frameshift mutations was present in all tumor cells. Through in vitro assays using engineered antigen-presenting cells, we were able to stimulate peripheral cytotoxic T cells obtained from colorectal cancer patients with peptides derived from frameshift mutations found in their tumors. Taken together, our results highlight the importance of a CD8(+) T cell immune response against MSI colorectal cancer-specific neoantigens, establishing a preclinical rationale to target them as a personalized cellular immunotherapy strategy, an especially appealing goal for patients with Lynch syndrome