Massively parallel functional testing of MSH2 missense variants conferring Lynch syndrome risk

The lack of functional evidence for the majority of missense variants limits their clinical interpretability and poses a key barrier to the broad utility of carrier screening. In Lynch syndrome (LS), one of the most highly prevalent cancer syndromes, nearly 90% of clinically observed missense variants are deemed “variants of uncertain significance” (VUS). To systematically resolve their functional status, we performed a massively parallel screen in human cells to identify loss-of-function missense variants in the key DNA mismatch repair factor MSH2. The resulting functional effect map is substantially complete, covering 94% of the 17,746 possible variants, and is highly concordant (96%) with existing functional data and expert clinicians’ interpretations. The large majority (89%) of missense variants were functionally neutral, perhaps unexpectedly in light of its evolutionary conservation. These data provide ready-to-use functional evidence to resolve the ∼1,300 extant missense VUSs in MSH2 and may facilitate the prospective classification of newly discovered variants in the clinic.