RT Journal Article SR Electronic T1 DNA methylation-based signature of CD8+ tumor-infiltrating lymphocytes enables evaluation of immune response and prognosis in colorectal cancer JF Journal for ImmunoTherapy of Cancer JO J Immunother Cancer FD BMJ Publishing Group Ltd SP e002671 DO 10.1136/jitc-2021-002671 VO 9 IS 9 A1 Qi Zou A1 Xiaolin Wang A1 Donglin Ren A1 Bang Hu A1 Guannan Tang A1 Yu Zhang A1 Meijin Huang A1 Rish K Pai A1 Daniel D Buchanan A1 Aung Ko Win A1 Polly A Newcomb A1 William M Grady A1 Huichuan Yu A1 Yanxin Luo YR 2021 UL http://jitc.bmj.com/content/9/9/e002671.abstract AB Background Tumor-infiltrating lymphocytes (TILs), especially CD8+ TILs, can be used for predicting immunotherapy responsiveness and survival outcome. However, the evaluation of CD8+ TILs currently relies on histopathological methodology with high variability. We therefore aimed to develop a DNA methylation signature for CD8+ TILs (CD8+ MeTIL) that could evaluate immune response and prognosis in colorectal cancer (CRC).Methods A CD8+ MeTIL signature score was constructed by using CD8+ T cell-specific differentially methylated positions (DMPs) that were identified from Illumina EPIC methylation arrays. Immune cells, colon epithelial cells, and two CRC cohorts (n=282 and 335) were used to develop a PCR-based assay for quantitative analysis of DNA methylation at single-base resolution (QASM) to determine CD8 + MeTIL signature score.Results Three CD8+ T cell-specific DMPs were identified to construct the CD8+ MeTIL signature score, which showed a dramatic discriminability between CD8+ T cells and other cells. The QASM assay we developed for CD8+ MeTIL markers could measure CD8+ TILs distributions in a fully quantitative, accurate, and simple manner. The CD8+ MeTIL score determined by QASM assay showed a strong association with histopathology-based CD8+ TIL counts and a gene expression-based immune marker. Furthermore, the low CD8+ MeTIL score (enriched CD8+ TILs) was associated with MSI-H tumors and predicted better survival in CRC cohorts.Conclusions This study developed a quantitative DNA methylation-based signature that was reliable to evaluate CD8+ TILs and prognosis in CRC. This approach has the potential to be a tool for investigations on CD8+ TILs and a biomarker for therapeutic approaches, including immunotherapy.Data are available upon reasonable request. All data relevant to the study are included in the article or uploaded as supplementary information. The data that support the findings of this study are available from the CCFR but restrictions apply to the availability of these data, which were used under license for the current study. The other datasets used and analyzed during the current study are available from the corresponding author on reasonable request.