Spatial Profiles of Intratumoral PD-1+ Helper T Cells Predict Prognosis in Head and Neck Squamous Cell Carcinoma

Abstract

BackgroundFunctional interactions between immune cells and neoplastic cells in the tumor immune microenvironment have been actively pursued for both biomarker discovery for patient stratification, as well as therapeutic anti-cancer targets to improve clinical outcomes. Although accumulating evidence indicates that intratumoral infiltration of immune cells has prognostic significance, limited information is available on the spatial infiltration patterns of immune cells within intratumoral regions. This study aimed to understand the intratumoral heterogeneity and spatial distribution of immune cell infiltrates associated with cell phenotypes and prognosis in head and neck squamous cell carcinoma (HNSCC).MethodsA total of 88 specimens of oropharyngeal squamous cell carcinoma, categorized into discovery (n = 38) and validation cohorts (n = 51), were analyzed for immune contexture by multiplexed immunohistochemistry (IHC) and image cytometry-based quantification. Tissue segmentation was performed according to a mathematical morphological approach using neoplastic cell IHC images to dissect intratumoral regions into tumor cell nests versus intratumoral stroma.ResultsTissue segmentation revealed heterogeneity in intratumoral T cells, varying from tumor cell nest-polarized to intratumoral stroma-polarized distributions. Leukocyte composition analysis revealed higher ratios of TH1/TH2 in tumor cell nests with higher percentages of helper T cells, B cells, and CD66b+ granulocytes within intratumoral stroma. A discovery and validation approach revealed a high density of programmed death receptor-1 (PD-1)+ helper T cells in tumor cell nests as a negative prognostic factor for short overall survival. CD163+ tumor-associated macrophages (TAM) provided the strongest correlation with PD-1+ helper T cells, and cases with a high density of PD-1+ helper T cells and CD163+ TAM had a significantly shorter overall survival than other cases.ConclusionThis study reveals the significance of analyzing intratumoral cell nests and reports that an immune microenvironment with a high density of PD-1+ helper T cells in tumoral cell nests is a poor prognostic factor for HNSCC.

Deciphering the Immune Complexity in Esophageal Adenocarcinoma and Pre-Cancerous Lesions With Sequential Multiplex Immunohistochemistry and Sparse Subspace Clustering Approach

“…Apart from the efficiency and accuracy, it is possible that these unsupervised clustering tools will generate subsets that, if biologically relevant, are difficult to interpret for their identities. For example, unclassified immune cell subsets in this study and “CD45 + other” in other multiplex imaging studies ( 56 ). It does not necessarily mean that these cells do not have biological significance, but they are unclear if analyzed using only one clustering method and a limited antibody panel.…”

Contribution of MMP14-expressing cancer-associated fibroblasts in the tumor immune microenvironment to progression of colorectal cancer

“…Image cytometry–based analysis of cell populations allowed quantitative evaluation of the cell subsets as cell number per area ( Figure 2C ). In addition, we applied our recently developed Tissue Segmentation method ( 39 ) to quantify cell number according to different tumor areas including IF, ISA, and TN ( Supplementary Figure S2B ), thus allowing detailed evaluation of the clinical relevance of each cell subset. Given that α smooth muscle actin (αSMA), a marker for CAFs in our mIHC panel, was expressed in both smooth muscle cells and fibroblasts in the IF, this region was excluded from subsequent analysis of the association of CAFs with immune cells.…”

“…The IF was defined as the area within 300 µm external to the boundary separating the tumor cell area from the surrounding connective tissue ( Supplementary Figure S2B ), as described previously ( 38 ). The CT was defined as an intratumoral area and was further categorized into a tumor cell nest (TN) area and a non–tumor cell area comprising mostly intratumoral stromal tissue (ISA) ( Supplementary Figure S2B ) with the use of a mathematical morphological approach performed with the Tissue Segmentation platform introduced in our previous study ( 39 ). Between two and five fields of view were evaluated depending on the size of the tumor (average of 3.64 fields of view for CT and 4.37 for IF).…”

The Role of Pathology-Based Methods in Qualitative and Quantitative Approaches to Cancer Immunotherapy

“…Chromogenic-based mIHC was used for the determination of the clinicopathological significance of PD-1, LAG3, and TIM3 markers in stage II/III gastric cancer patients [ 34 ] and for the identification the phenotypes and spatial profiles of intratumoural PD-1+ helper T cells associated with the prognosis of head and neck squamous cell carcinoma (HNSCC) [ 35 ]. Some analytical validation attempts of automated multiplex chromogenic IHC for diagnostic and predictive purposes could potentially be made in the clinic for NSCLC patient care.…”