November 2023 – Gynecological Pathology / Laboratory Genetics and Genomics

A 72-year-old woman presents with vaginal bleeding and is subsequently diagnosed with endometrioid adenocarcinoma (FIGO grade 2/3) (Figure 1). Immunohistochemistry (IHC) for mismatch repair (MMR) proteins is notable for an absence of MLH1, PMS2, MSH2, and MSH6 expression (Figure 2). 

Figure 1
Figure 2

What underlying molecular genetic results may adequately account for this rare immunophenotypic pattern?

  • Failure of IHC positive control
  • POLE alteration (variant of undetermined significance, VUS) and PMS2 pathogenic alteration
  • MLH1 promoter hypermethylation and MSH2 pathogenic alteration (c.560T>C, p.Leu187Pro)
  • MLH1 promoter hypermethylation and MSH6 alteration (variant of undetermined significance, VUS)

The correct answer is ...

MLH1 promoter hypermethylation and MSH2 pathogenic alteration (c.560T>C, p.Leu187Pro).

MLH1 promoter hypermethylation leads to silencing of MLH1 gene expression thus preventing subsequent protein formation. Consistent with the patient’s age, this form of epigenetic inactivation occurs most commonly in the context of sporadic (versus hereditary) tumor development. Loss of MLH1 would also account for the loss of PMS2 protein expression given the existence of the two proteins as a dimer. Independently, a pathogenic missense variant in MSH2 resulting from the instability created by MLH1/PMS2 loss would lead to the loss of MSH2, which would in turn lead to concomitant loss of MSH6 expression. 

POLE alteration (variant of undetermined significance) and PMS2 pathogenic alteration 

Although a POLE pathogenic alteration could, in theory, result in additional downstream alterations (including in the MMR genes) that could, in turn, result in loss of IHC staining for all MMR proteins. However, only a VUS was identified in POLE and the overall mutation spectrum of the tumor was not hypermutated. In addition, there is limited evidence available to support this theory. Although a PMS2 pathogenic alteration could explain the loss of PMS2 by IHC, not all PMS2 mutations cause the loss of MLH1. The PMS2 mutation would not explain the loss of MSH2/MSH6.

MLH1 promoter hypermethylation and MSH6 alteration (variant of undetermined significance, VUS)

MLH1 promoter hypermethylation would explain the loss of MLH1 and PMS2 by IHC. However, a VUS in MSH6 would not necessarily explain the loss of MSH6. Furthermore, MSH2 can be expressed by IHC in the absence of MSH6

Failure of IHC positive control

Although a failure of the positive control can certainly explain the loss of any protein by IHC, background staining of non-neoplastic cells (stroma/lymphocytes) in this case rules out this possibility (Figure 2).


  1. Nielsen SV, Stein A, Dinitzen AB, et al. Predicting the impact of Lynch syndrome-causing missense mutations from structural calculations. PLoS Genet. 2017;13(4):e1006739. Published 2017 Apr 19. doi:10.1371/journal.pgen.1006739
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Mazen Atiq, M.B.B.S.

Fellow, Molecular Genetic Pathology
Mayo Clinic

Kevin Halling, M.D., Ph.D.

Consultant, Laboratory Genetics and Genomics
Mayo Clinic
Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

Ande Rumilla, M.D.

Consultant, Laboratory Genetics and Genomics
Mayo Clinic
Assistant Professor of Laboratory Medicine and Pathology
Mayo Clinic College of Medicine and Science

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