ZEB1 hypermethylation is associated with better prognosis in patients with colon cancer.

BACKGROUND: Colon cancer (CC) is a heterogeneous disease that is categorized into four Consensus Molecular Subtypes (CMS) according to gene expression. Patients with loco-regional CC (stages II/III) lack prognostic factors, making it essential to analyze new molecular markers that can delineate more aggressive tumors. Aberrant methylation of genes that are essential in crucial mechanisms such as epithelial mesenchymal transition (EMT) contributes to tumor progression in CC. We evaluate the presence of hyper- and hypomethylation in subrogate IHC markers used for CMS classification (CDX2, FRMD6, HTR2B, ZEB1) of 144 stage II/III patients and CC cell lines by pyrosequencing. ZEB1 expression was also studied in control and shRNA-silenced CC cell lines and in paired normal tissue/tumors by quantitative PCR. The pattern of ZEB1 staining was also analyzed in methylated/unmethylated tumors by immunohistochemistry. RESULTS: We describe for the first time the hypermethylation of ZEB1 gene and the hypomethylation of the FRMD6 gene in 32.6% and 50.9% of tumors, respectively. Additionally, we confirm the ZEB1 re-expression by epigenetic drugs in methylated cell lines. ZEB1 hypermethylation was more frequent in CMS1 patients and, more importantly, was a good prognostic factor related to disease-free survival (p = 0.015) and overall survival (p = 0.006) in our patient series, independently of other significant clinical parameters such as patient age, stage, lymph node involvement, and blood vessel and perineural invasion. CONCLUSIONS: Aberrant methylation is present in the subrogate genes used for CMS classification. Our results are the first evidence that ZEB1 is hypermethylated in CC and that this alteration is an independent factor of good prognosis.

A Novel Prognostic Biomarker Panel for Early-Stage Colon Carcinoma.

Molecular characterization of colorectal cancer has helped us understand better the biology of the disease. However, previous efforts have yet to provide significant clinical value in order to be integrated into clinical practice for patients with early-stage colon cancer (CC). The purpose of this study was to assess PD-L1, GLUT-1, e-cadherin, MUC2, CDX2, and microsatellite instability (dMMR) and to propose a risk-panel with prognostic capabilities. Biomarkers were immunohistochemically assessed through tissue microarrays in a cohort of 144 patients with stage II/III colon cancer. A biomarker panel consisting of PD-L1, GLUT-1, dMMR, and potentially CDX2 was constructed that divided patients into low, medium, and high risk of overall survival or disease-free survival (DFS) in equally sized groups. Compared with low-risk patients, medium-risk patients have almost twice the risk of death (HR = 2.10 (0.99-4.46), p = 0.054), while high-risk patients have almost four times the risk (HR = 3.79 (1.77-8.11), p = 0.001). The multivariate goodness of fit was 0.756 and was correlated with Kaplan-Meier curves (p = 0.002). Consistent results were found for DFS. This study provides a critical basis for the future development of an immunohistochemical assessment capable of discerning early-stage CC patients as a function of their prognosis. This tool may aid with treatment personalization in daily clinical practice and improve survival outcomes.

PD-L1 as a Prognostic Factor in Early-Stage Colon Carcinoma within the Immunohistochemical Molecular Subtype Classification.

BACKGROUND: There is a patent need to better characterize early-stage colorectal cancer (CRC) patients. PD-1 ligand (PD-L1) expression has been proposed as a prognostic factor but yields mixed results in different settings. The Consensus Molecular Subtype (CMS) classification has yet to be integrated into clinical practice. We sought to evaluate the prognostic value of PD-L1 expression overall and within CMS in early-stage colon cancer patients, in the hope of assisting treatment choice in this setting. METHODS: Tissue-microarrays were constructed from tumor samples of 162 stage II/III CRC patients. They underwent automatic immunohistochemical staining for PD-L1 and the proposed CMS panel. Primary endpoints were overall survival (OS) and disease-free survival (DFS). RESULTS: PD-L1 expression was significantly and independently associated with better prognosis (HR = 0.46 (0.26-0.82), p = 0.009) and was mostly seen in immune cells of the tumor-related stroma. CMS4 five-folds the risk of mortalitycompared with CMS1 (HR = 5.58 (1.36, 22.0), p = 0.034). In the subgroup CMS2/CMS3 analysis, PD-L1 expression significantly differentiated individuals with better OS (p = 0.004) and DFS (p < 0.001). CONCLUSIONS: Our study suggests that PD-L1 expression is an independent prognostic factor in patients with stage II/III colon cancer. Additionally, it successfully differentiates patients with better prognosis in the CMS2/CMS3 group and may prove significant for the clinical relevance of the CMS classification.