Deciphering the Pathogenicity of the PMS2 p.His701Gln Variant in a Family with Elderly-Onset Extracolonic Lynch Syndrome

Missense variants in PMS2 remain difficult to interpret in Lynch Syndrome (LS), particularly when clinical evidence is limited. We investigated the PMS2, NM_000535.7: c.2103T>A; p.His701Gln (p.H701Q) variant, currently classified as a variant of uncertain significance (VUS), identified in a 71-year-old patient with small intestinal adenocarcinoma. To clarify its clinical significance, we combined clinical, in silico, and functional analyses. Computational predictions consistently pointed to a deleterious effect, with AlphaMissense assigning a highly damaging score and most independent predictors supporting the same interpretation. H701Q affects the highly conserved C-terminal region of PMS2, close to the functionally critical Glu705 residue. In HEK-293T cells, PMS2-H701Q showed markedly reduced protein levels compared with wild-type PMS2, supporting an effect on protein stability. Co-immunoprecipitation experiments further showed a substantial reduction in the interaction between PMS2-H701Q and MLH1, indicating impaired MutLα complex formation. Both effects were comparable to those observed for the pathogenic PMS2-E705K control. Overall, the agreement between computational and functional findings provides compelling evidence for a deleterious effect of H701Q and strengthens its clinical interpretation. Collectively, these findings demonstrate the utility of functional evidence under the ClinGen-InSiGHT PMS2-specific ACMG/AMP variant classification guidelines and support reclassifying the PMS2, NM_000535.7: c.2103T>A; p.His701Gln (p.H701Q) variant from a Variant of Uncertain Significance (VUS) to likely pathogenic.