MLH1 promoter methylation analysis

1. 10 unstained sections of tumour and a corresponding H&E-stained histological section, OR

2. 1 paraffin block of tumour

Background

Endometrial cancers with loss of the DNA mismatch protein MLH1 by immunohistochemistry require further testing to determine the methylation status of the MLH1 gene promoter. A positive result for MLH1 promoter methylation will in most situations be consistent with a sporadic MSI-high tumour. A negative result for MLH1 promoter methylation necessitates further genetics referral and assessment.

Purpose of test

This test assesses the methylation status of the 3’ clinically significant region of the MLH1 gene promoter by methylation-specific melting curve analysis.

Expected test result

Methylated / Unmethylated

Caveats

DNA integrity and concentration must meet assay requirements. Low tumour content (<30%) may result in an inaccurate result. Rare cases of Lynch syndrome may have methylation of MLH1 as the second hit in Lynch syndrome tumorigenesis. Mutations to the MLH1 promoter sequence unrelated to methylation will result in amplicons that exhibit a different melt curve compared to the methylated and unmethylated wild-type promoter samples. Identification of the exact mutations present in the sequence of the MLH1 promoter amplicon requires additional sequencing studies which is not offered as part of this test.

Proficiency testing

College of American Pathologists proficiency testing programme.

Reference

Wong A, Ngeow J. Hereditary syndromes manifesting as endometrial carcinoma: how can pathological features aid risk assessment? Biomed Res Int 2015; 2015: 219012.