Tetranucleotide and Low Microsatellite Instability Are Inversely Associated with the CpG Island Methylator Phenotype in Colorectal Cancer.

Sabine Meessen,Daniel D Buchanan,Zeenat Jahan,Mark A Jenkins,Maija R J Kohonen-Corish,Jane E Dahlstrom,John L Hopper,Nicola Currey,Eva Segelov,Hannah W Parker

doi:10.3390/cancers13143529

Abstract

Simple SummaryA type of DNA mismatch repair defect known as “elevated microsatellite alterations at selected tetranucleotide repeats” (EMAST) is found across many different cancers. Tetranucleotide microsatellite instability, which is caused by MSH3 mismatch repair gene/protein loss-of-function, shares a molecular basis with “low microsatellite instability” (MSI-L) in colorectal cancer. Tetranucleotide microsatellite instability is also a byproduct of “high microsatellite instability” (MSI-H) that arises from deficiency of mismatch repair due to MSH2, MSH6, MLH1 or PMS2 gene alterations. MSH3-related EMAST is emerging as a biomarker of poor prognosis in colorectal cancer and needs to be clearly differentiated from MSI-H. Here, we show that tumours with non-MSI-H-related EMAST or MSI-L rarely show concordant promoter methylation of multiple marker genes. Colorectal tumours that are positive for a single (1/5) tetranucleotide repeat marker are an important subset of the EMAST spectrum.MSH3 gene or protein deficiency or loss-of-function in colorectal cancer can cause a DNA mismatch repair defect known as “elevated microsatellite alterations at selected tetranucleotide repeats” (EMAST). A high percentage of MSI-H tumors exhibit EMAST, while MSI-L is also linked with EMAST. However, the distribution of CpG island methylator phenotype (CIMP) within the EMAST spectrum is not known. Five tetranucleotide repeat and five MSI markers were used to classify 100 sporadic colorectal tumours for EMAST, MSI-H and MSI-L according to the number of unstable markers detected. Promoter methylation was determined using methylation-specific PCR for MSH3, MCC, CDKN2A (p16) and five CIMP marker genes. EMAST was found in 55% of sporadic colorectal carcinomas. Carcinomas with only one positive marker (EMAST-1/5, 26%) were associated with advanced tumour stage, increased lymph node metastasis, MSI-L and lack of CIMP-H. EMAST-2/5 (16%) carcinomas displayed some methylation but MSI was rare. Carcinomas with ≥3 positive EMAST markers (13%) were more likely to have a proximal colon location and be MSI-H and CIMP-H. Our study suggests that EMAST/MSI-L is a valuable prognostic and predictive marker for colorectal carcinomas that do not display the high methylation phenotype CIMP-H.

Highlights

Microsatellite instability (MSI) in cancer is caused by deficient repair of insertion– deletion loop mismatches, which occur during DNA replication in tandem repeat sequences throughout the genome [1,2]
We first determined the prevalence of instability for each of these five EMAST markers across the four cohorts of carcinomas where MSI-H is found at a high frequency (100%, Lynch syndrome, cohort 2), intermediate (13–15%, sporadic colorectal and colitis-associated carcinomas, cohorts 1 and 3) or low frequency
This study has shown that the high methylation phenotype CpG island methylator phenotype (CIMP)-H and concurrent MCC/CDKN2A methylation are rare in colorectal carcinomas that are positive for only one EMAST marker or MSI-L but more common in cancers with ≥2 markers

Summary

Introduction

Microsatellite instability (MSI) in cancer is caused by deficient repair of insertion– deletion loop mismatches, which occur during DNA replication in tandem repeat sequences throughout the genome [1,2]. MSI-H is caused by DNA mismatch repair (MMR) deficiency as a result of alterations in the MLH1, MSH2, MSH6 or PMS2 genes. Another form of MSI is “elevated microsatellite alterations at selected tetranucleotide repeats” (EMAST) that has been identified in up to 60% of colorectal cancers [3,4,5,6], but no standard marker panel has been adopted yet for its detection. EMAST is caused by a deficiency or dysfunction of MMR involving the MSH3 gene or protein [5,7]. The MSH3 gene is silenced through promoter methylation in gastric cancer [11], but the relevance of promoter methylation for the generation of EMAST has not been reported yet in colorectal cancer

Objectives

Methods

Results

Discussion

Conclusion