Revision of interpretation criteria to define microsatellite instability in mismatch repair-deficient neoplasms with subtle electropherogram changes.

Abstract

To improve analytic performance characteristics of a microsatellite instability (MSI-V1.2) assay in endometrial cancers (ECs). Nonneoplastic and neoplastic DNA from colorectal cancers (CRCs) and ECs were compared to define MSI by calculating base shifting of the highest peak and the 5% peak (the leftmost peak with a peak height >5% of the highest peak). We first demonstrated highly precise sizing by capillary electrophoresis. However, relative intensity of multiple peaks, characteristic for microsatellite amplicons, might show a 1-base, but not a 2-base or more, shift of the highest or 5% peak among duplicate runs of nonneoplastic DNA. This inherent bias of the polymerase chain reaction-based MSI assay may lead to false-positive interpretation if MSI was defined by a 1-base shift or more. Subsequently, MSI was evaluated by a 2-base shift or more of the highest peak (original criteria) or a 2-base shift or more of either the highest or 5% peak (revised criteria) without subjective interpretation of a subtle change of electropherogram configuration (the so-called shoulder pattern). While both criteria were highly sensitive in CRCs, the revised criteria improved sensitivity (83% vs 67%) and accuracy (89% vs 79%) and maintained 100% specificity in ECs. The revised criteria provided sensitive, specific, and objective interpretation to examine subtle changes of MSI.