Differential gene expression of tumor-infiltrating CD4 ⁺ T cells in advanced versus early stage colorectal cancer and identification of a gene signature of poor prognosis

Abstract

Tumor-infiltrating lymphocytes (TILs) play indispensable roles in the progression and response to treatment of solid tumors. However, the prognostic significance of CD4 + TILs is not fully disclosed in cancers generally and in CRC in particular, mainly due to the existence of different functional subsets of CD4 + T cells. We performed transcriptomic profiling of CD4 + TILs isolated from CRC patients in order to identify differentially expressed genes and their functional pathways in early versus advanced disease stages. We found that in advanced stages, genes related to immune and inflammatory responses, in particular Th1mediated immune response and cytotoxicity-mediated genes, were downregulated; while epigeneticmediated silencing genes were upregulated. Interestingly, we identified genes, which were steadily upregulated or downregulated in CD4 + TILs with CRC progression from stage I to IV. Additionally, of the top 200 deregulated genes, 43 upregulated and 64 downregulated genes showed similar deregulation trends in the cancer genome atlas CRC dataset. From these 97 deregulated genes, we identified a "poor prognosis CD4 gene signature (ppCD4sig)". Patients with high ppCD4sig score showed shorter diseasespecific survival (DSS) and progression-free interval (PFI). The ppCD4sig was an independent prognostic indicator for DSS (HR = 1.73, 95% CI 1.32-2.27, P = 0.0001) and PFI (HR = 1.75, 95% CI 1.3-2.35, P = 0.0016). Additionally, patients at advanced stages and at a younger age (<55 years) were more likely to have a high ppCD4sig score. Altogether, our data provide novel insights and a unique prognostic gene signature of CD4 + TILs in the CRC microenvironment.

Transcriptome of Tumor-Infiltrating T Cells in Colorectal Cancer Patients Uncovered a Unique Gene Signature in CD4+ T Cells Associated with Poor Disease-Specific Survival

“…We found that CD4 + TILs in patients with high ppScores express low DNMT3B, KDM5B/6B and high HDAC5 and KAT2B levels, compared to those with low ppScores (data not shown). This is consistent with previous findings showing that the DNMT3B gene was downregulated in CD4 + TILs from CRC patients with advanced stages [ 15 ], indicating that transcriptional silencing through DNA methylation in CD4 + TILs could be associated with poor prognosis; KAT2B is important to skew Th1 differentiation into Tregs [ 42 ]; HDACs, including HDAC5, could prevent the induction and differentiation of Th1 cells [ 43 , 44 ], and reduced levels of KDM5B and KDM6B can positively skew Th2 differentiation [ 45 , 46 ]. However, IFN-γ and their receptor expression in CD8 + T cells may also be influenced by epigenetic modifications [ 47 ] and, therefore, functional investigations are necessitated to confirm impaired IFN-γ release by CD8 + TILs in CRC patients.…”

“…Various bioinformatics tools were utilized for analyses and visualization of RNA-Seq data. The raw data used in this study were generated previously in the same patient cohort [ 15 , 16 ] but were analyzed herein to uncover novel information. The study design has been clearly presented in [ 16 ] and summarized below.…”

“…Molecular and cellular profiling of immune cell populations, specifically T cells, bear great significance in understanding CRC development and progression. We have recently performed transcriptomic profiling of sorted CD4 + [ 15 ] and CD8 + [ 16 ] tumor-infiltrating lymphocytes (TILs) from bulk tumors of CRC patients with varying disease stages. We reported downregulation of Th1-mediated immune response and cytotoxicity-mediated genes but upregulation of epigenetic silencing-related genes in CD4 + TILs from CRC patients with advanced stage disease [ 15 ].…”

“…We have recently performed transcriptomic profiling of sorted CD4 + [ 15 ] and CD8 + [ 16 ] tumor-infiltrating lymphocytes (TILs) from bulk tumors of CRC patients with varying disease stages. We reported downregulation of Th1-mediated immune response and cytotoxicity-mediated genes but upregulation of epigenetic silencing-related genes in CD4 + TILs from CRC patients with advanced stage disease [ 15 ]. Moreover, epigenetic regulation-related genes and response to hypoxia were upregulated, while T cell/cell proliferation- and cell cycle-related genes were downregulated in CD8 + TILs in advanced stage CRC patients [ 16 ].…”

A Prognostic Model Based on the Immune-Related lncRNAs in Colorectal Cancer

“…These immune cells play important roles in the prognosis of CRC. Sasidharan et al identified a "poor prognosis CD4 gene signature" (ppCD4sig), found that patients with high ppCD4sig score showed shorter disease specific survival and progression-free interval, which provided novel insights and a unique prognostic gene signature of CD4 + tumor-infiltrating lymphocytes in the CRC microenvironment (Sasidharan Nair et al, 2020). Lavoie et al reported that loss of free fatty acid receptor two promoted colon tumorigenesis in mice by reducing gut barrier integrity, increasing tumor bacterial load, promoting exhaustion of CD8 + T cells, and over activating DCs, leading to their death (Lavoie et al, 2020).…”