ABSTRACT
Intratumoral immune profiles are related to prognosis and therapeutic efficacy, and could result in personalized treatments based on biomarkers. To develop a multiplex, quantitative, and rapid tissue evaluation method based on the clinically established standard immunohistochemistry (IHC), a 6-marker rapid multiplex IHC was developed based on our previously reported 14-marker multiplex IHC by reducing the number of labels and accelerating the staining procedure. First, fewer labels were required to identify the same immunological features linked to prognosis in 14-marker multiplex IHC analyses. The six selected markers showed a significant correlation with the 14 markers in the immune classification. Next, a rapid staining protocol was developed by optimizing the reaction temperature, chromogen, and washing time, allowing the completion of 6-marker analysis in 5 h and 49 min, as opposed to the several days required for conventional multiplex IHC. Validation of benign tonsil and head and neck cancer tissues revealed a significant correlation between rapid and conventional 6-makrer multiplex IHC in terms of staining intensities, densities of T cells, macrophages, lymphoid/myeloid immune cell ratios, and spatial profiles of intratumoral immune infiltrates. This method may enable quantitative assessment of the tumor-immune microenvironment on a clinically feasible time scale, which promotes the development of tissue biomarker-guided therapeutic strategies.
ABSTRACT
BACKGROUND: The relationship between the tumor-immune microenvironment and systemic inflammatory markers, including neutrophil-to-lymphocyte ratio (NLR), is unclear. METHODS: We examined the characteristics of systemic inflammatory markers and tumor immune microenvironments in relation to treatment outcomes in 29 consecutive patients with recurrent/metastatic head and neck squamous cell carcinoma (R/M HNSCC) who received pembrolizumab, using 14-marker multiplex immunohistochemistry and image cytometry. RESULTS: NLR ≥4.5 (high NLR) at pretreatment status significantly correlated with short overall survival (OS) and progression-free survival-2 (PFS2) and malnutrition status. High NLR in peripheral blood was significantly correlated with low lymphoid cell and high tumor-associated macrophage counts in tissues, especially myeloid-to-lymphoid cell ratios, suggesting an association between circulating and intratumoral immune complexity profiles. CONCLUSIONS: This study suggests a link between NLR in circulating blood, systemic nutritional status, and immune composition within the tumor.
Subject(s)
Antibodies, Monoclonal, Humanized , Head and Neck Neoplasms , Lymphocytes , Neutrophils , Squamous Cell Carcinoma of Head and Neck , Humans , Male , Female , Squamous Cell Carcinoma of Head and Neck/drug therapy , Squamous Cell Carcinoma of Head and Neck/immunology , Squamous Cell Carcinoma of Head and Neck/mortality , Squamous Cell Carcinoma of Head and Neck/pathology , Middle Aged , Head and Neck Neoplasms/drug therapy , Head and Neck Neoplasms/immunology , Head and Neck Neoplasms/pathology , Head and Neck Neoplasms/mortality , Aged , Antibodies, Monoclonal, Humanized/therapeutic use , Nutritional Status , Tumor Microenvironment/immunology , Adult , Antineoplastic Agents, Immunological/therapeutic use , Lymphocyte Count , Aged, 80 and over , Retrospective StudiesABSTRACT
Spatial profiles of the tumor-immune microenvironment are associated with disease progression and clinicopathological factors in various cancers. Follicular thyroid carcinoma (FTC) is the second most common thyroid cancer, where the presence of capsular invasion or angioinvasion determines the pathological diagnosis; however, little is known about the immune microenvironment profiles associated with the acquisition of invasive potential of FTC. In this study, we focused on FTC with minimal capsular invasion, and the spatially resolved immune microenvironment of FTC was studied in the discovery (n = 13) and validation cohorts (n = 40). CD8+ T cells, helper T cells, regulatory T cells, B cells, natural killer cells, tumor-associated macrophages, CD66+ granulocytes, mature dendritic cells, and mast cells were quantitatively evaluated in single tissue sections, via a 12-marker multiplex immunohistochemistry and image cytometry. Cell densities and compositions of immune cells were spatially stratified by six tissue regions including tumor center, subcapsular region, capsular invasion, adjacent stroma of capsular invasion, peritumoral stroma, and adjacent normal. Lymphoid cell lineages in the tumor center and subcapsular regions were significantly lower than those in adjacent normal and peritumoral stroma, potentially related to the lymphoid lineage exclusion from the intratumoral regions of FTC. Interestingly, immune cell composition profiles in the capsular invasive front were distinct from those of intratumoral region. The ratios of T cells to CD66b+ granulocytes with capsular invasion were significantly higher than those without capsular invasion, suggesting the presence of a unique immune microenvironment at the invasive front between tumor foci and stroma. In addition, tumor cells at the capsular invasive front showed significantly higher expression of tumor programmed cell death ligand 1 (PD-L1) than those at the tumor center. This study revealed spatial immune profiles associated with capsular invasion of FTC, providing new insights into the mechanisms underlying its development and initial invasion.