Tc1 and Th1 T-lymphocyte rich tumor microenvironment is a hallmark of MSI colorectal cancer

Abstract

Microsatellite instability (MSI) is a strong biomarker to predict response to immune checkpoint therapy and patient’s outcome in colorectal cancer. Although enrichment of distinct T-cell subpopulations, such as type 1 cytotoxic T-cells (Tc1) or type 1 helper T-cells (Th1), have been identified to impact patient’s outcome and response to immune checkpoint therapy, only little is known about the underlying changes in the composition of the immune tumor microenvironment. To assess the density, composition, degree of functional marker expression, and spatial interplay of T-cell subpopulations in 79 MSI and 1045 microsatellite stable (MSS) colorectal cancers, a tissue microarray as well as large sections were stained with 19 antibodies directed against T-cell, antigen presenting cells, functional markers, and structural proteins using our BLEACH&STAIN multiplex fluorescence immunohistochemistry approach. A deep learning-based framework comprising > 20 different convolutional neuronal networks (U-Net and DeepLabv3+) was developed for image analysis. The composition of Type 1 (T-bet+), Type 2 (GATA3+), Type 17 (RORγT+), NKT-like (CD56+), regulatory (FOXP3+), and follicular (BCL6+) cytotoxic (CD3+CD8+) or helper (CD3+CD4+) T-cells showed marked differences between MSI and MSS patients. For instance, the fraction of Tc1 as well as Th1 was significantly higher (p<0.001 each) while the fraction of Tregs, Th2, Th17 T-cells was significantly lower (p<0.05) in MSI compared to MSS patients. The degree of TIM3, CTLA-4, and PD-1 expression on most T-cell subpopulations was significantly higher in MSI compared to MSS patients (p<0.05 each). Spatial analysis revealed increased interactions between Th1, Tc1, and dendritic cells in MSI patients while in MSS patients the strongest interactions were found between Tregs, Th17, Th2 and dendritic cells. The additional analysis of 12 large sections confirmed the observations from the TMA analysis in the center of the tumor and showed a divergent immune composition in the invasive margin. Therefore, this study identified a higher fraction of Tc1 T-cells and Th1 T-cells accompanied by a paucity of regulatory T-cell, Th17 and Th2 T-cell subpopulations along with a distinct interaction profile as a hallmark of MSI compared to MSS colorectal cancers.