Digital pathology-aided assessment of tumor-infiltrating T lymphocytes in advanced stage, HPV-negative head and neck tumors

Abstract

Aim This study aimed to evaluate the presence and prognostic value of tumor-infiltrating T cells in the tumor epithelium in advanced stage, HPV-negative head and neck squamous cell carcinoma (HNSCC) patients treated with primary chemoradiotherapy using digital pathology. Methods Pre-treatment biopsies from 80 oropharyngeal, 52 hypopharyngeal, and 29 laryngeal cancer patients were collected in a tissue microarray (TMA) and immunohistochemically stained for T-cell markers CD3, CD4, CD8, FoxP3, and PD1, and for immune checkpoint PD-L1. For each marker, the number of positive tumor-infiltrating lymphocytes (TILs) per mm 2 tumor epithelium was digitally quantified and correlated to overall survival (OS), disease-free survival (DFS), and locoregional control (LRC), as well as to clinicopathological characteristics. Differences in clinical outcome were estimated using Cox proportional hazard analysis and visualized using Kaplan-Meier curves. Results The patient cohort had a 3-year OS of 58%, with a median follow-up of 53 months. None of the T-cell markers showed a correlation with OS, DFS or LRC. A low N stage was correlated to a better prognosis (OS: HR 0.39, p = 0.0028, DFS: HR 0.34, p = < 0.001, LRC: HR 0.24, p = 0.008). High TIL counts were more often observed in PD-L1-positive tumors (p < 0.05). Conclusion This study showed an objective, digital pathology-aided method to assess TILs in the tumor epithelium. However, it did not provide evidence for a prognostic role of the presence of CD3 + , CD4 + , CD8 + , FoxP3 + , and PD1 + TILs in the tumor epithelium of advanced stage, HPV-negative HNSCC patients treated with primary chemoradiotherapy. Keywords Head and neck squamous cell carcinoma (HNSCC) · Tumor-infiltrating lymphocytes (TILs) · T cells · Prognostic biomarkers AbbreviationsACE-27 Adult Comorbidity Evaluation-27 AUC Area under the curve CI Confidence interval DFS Disease-free survival FFPE Formalin fixed, paraffin embedded HR Hazard ratio ICC Intraclass correlation coefficient LRC Locoregional control ROC Receiver operating characteristics TMA Tissue microarray

Immune related proteins and tumor infiltratingCD8+ lymphocytes in hypopharyngeal cancer in relation to human papillomavirus (HPV) and clinical outcome

“…Birtalan et al found a high PD‐L1 expression on immune cells to be linked to an improved survival for patients with HPV‐negative tumors. In contrast, de Ruither et al did not find a correlation between number of CD8+ TILs or tumor PD‐L1 expression and survival 40 . Also Schneider et al could not show a correlation between the expression of PD‐L1 on the tumor cells and survival, although the numbers of PD‐1 positive TILs did correlate to increased survival 24 .…”

“…This indicates that the immune defense is of major importance for clinical outcome also for patients harboring HPV‐negative HPSCC. Some earlier studies have evaluated CD8+ TILs or expression of PD‐1 and PD‐L1 in HPSCC, both in relation to survival, and in some cases also tumor HPV‐status 20,21,23,24,40 . Ono et al found a positive correlation between a high number of CD8+ TILs and survival in two studies 20,21 .…”

Tumor-Infiltrating Lymphocytes in the Tumor Microenvironment of Laryngeal Squamous Cell Carcinoma: Systematic Review and Meta-Analysis

“…The studies that did not fulfill all the inclusion criteria were discarded. Of these, seven were excluded because they did not report the outcome of LSCC separately from other locations of HNSCC [12,[15][16][17][18][19][20], eight because no HRs were available [21][22][23][24][25][26][27][28][29], one study because they reported outcome in terms of local control [30], one study that evaluated Il-12R expression on tumor cells [31], and one study that evaluated lymphocytes in peripheral blood [32]. This left 11 studies for the analysis (Fig- Studies were selected if they met the following inclusion criteria: (1) prognostic value of TIL was evaluated in patients with LSCC (primary or recurrent), (2) TIL were evaluated either immunohistochemically (CD8+, CD4+, and/or CD3+) and/or in hematoxylin-and eosin (HE)-stained sections, in resected specimens or in diagnostic biopsies, (3) TIL were evaluated either in tumor epithelium and/or tumor stroma, (4) prognostic value of TIL was evaluated by time-to-event survival analysis with overall survival (OS) and/or disease-free survival (DFS), and (5) original articles published in English.…”

CD8+ T Cell Infiltration Predicts Chemoradiosensitivity in Nasopharyngeal or Oropharyngeal Cancer

“…Similarly, de Ruiter et al reported that CD8+ TIL level is not correlated with OS or disease-free survival in HPV-negative HNC patients treated with CRT. 46 In addition, several studies have demonstrated that CD8+ TILs play a more significant role in virus-related HNCs than in nonvirus-related HNCs. [47][48][49] Considering our results and those of previous reports, we suggest the potential necessity of investigating other T-cell markers to evaluate their prognostic significance in nonvirus-related naso-or oropharyngeal cancer.…”