Higher FOXP3-TSDR demethylation rates in adjacent normal tissues in patients with colon cancer were associated with worse survival

Abstract

BackgroundThe influence of natural regulatory T cells (nTregs) on the patients with colon cancer is unclear. Demethylated status of the Treg-specific demethylated region (TSDR) of the FOXP3 gene was reported to be a potential biomarker for the identification of nTregs.MethodsThe demethylation rate of the TSDR (TSDR-DMR) was calculated by using methylation-specific quantitative polymerase chain reaction (MS-qPCR) assay. The expression of TSDR-DMR and FOXP3 mRNA was investigated in various colorectal cancer cell lines. A total of 130 colon carcinoma samples were utilized to study the DMR at tumor sites (DMRT) and adjacent normal tissue (DMRN). The correlations between DMRs and clinicopathological variables of patients with colon cancer were studied.ResultsThe TSDR-DMRs varied dramatically among nTregs (97.920 ± 0.466%) and iTregs (3.917 ± 0.750%). Significantly, DMRT (3.296 ± 0.213%) was higher than DMRN (1.605 ± 0.146%) (n = 130, p = 0.000). Higher DMRN levels were found in female patients (p = 0.001) and those with distant metastases (p = 0.017), and were also associated with worse recurrence-free survival in non-stage IV patients (low vs. high, p = 0.022). However, further Cox multivariate analysis revealed that the FOXP3-TSDR status does not have prognostic value.ConclusionMS-qPCR assays of FOXP3-TSDR can efficiently distinguish nTregs from non-nTregs. Abnormal recruitment of nTregs occurs in the local tumor microenvironment. Infiltration of tissue-resident nTregs may have a negative role in anti-tumor effects in patients with colon cancer; however, this role is limited and complicated.

Differential profile of activated regulatory T cell subsets and microRNAs in tolerant liver transplant recipients

“…In accordance with previous studies, Tol patients had a significantly longer time from transplant to weaning start and had a higher expression of FOXP3 mRNA . The analysis of the degree of FOXP3 TSDR methylation is a new tool for the quantitation of Tregs in peripheral blood or solid tissue in different diseases . Although a higher DMR at the FOXP3 locus in sorted CD4 + T cells from Tol kidney transplant patients has been previously shown, our work used a simplified technique starting from whole blood to show similar results in LT patients.…”

“…The DNA was then bisulfite converted using the Epitec Bisulfite Kit (Qiagen), and the degree of FOXP3 TSDR demethylation was determined by methyl‐specific quantitative polymerase chain reaction (MS‐qPCR). The methylation‐ and demethylation‐specific primers, cycling conditions, and demethylation rate (DMR) calculations were described previously . For female patients, DMR was corrected by a factor of 2 because 1 of the 2 TSDR alleles was methylated as a result of X‐inactivation.…”

Immune Checkpoints in Circulating and Tumor-Infiltrating CD4+ T Cell Subsets in Colorectal Cancer Patients

“…Majority of studies have associated Tregs with poor clinical outcomes in different cancers including CRC (47,48), however some studies have also associated these with better prognosis in CRC patients (49)(50)(51). Saito et al proposed that these results FIGURE 7 | Levels of CD4 + and CD4 + CTLA-4 + T cells in CRC patients with different staging and tumor budding.…”

Epigenetic dynamics during CD4+ T cells lineage commitment

“…Demethylation of Foxp3 is initiated during the thymic Treg cell development and continues when cells migrate to the periphery (Toker et al, 2013). Quantification of the methylation level in this region (called TSDR, Treg-specific demethylation region) has been successfully used to differentiate between stable and unstable Treg cell phenotypes and has been proposed as a biomarker for measuring Treg lineage commitment (Barzaghi et al, 2012;Bestard et al, 2011;Liu et al, 2010a;Lü et al, 2013;Wieczorek et al, 2009;Zhuo et al, 2014). This TSDR region is also occupied by active histone modifications (acetylation of H3 and H3K4me3), which contributes to the positive regulation of Foxp3.…”