Comprehensive Paired Tumor/Germline Testing for Lynch Syndrome: Bringing Resolution to the Diagnostic Process

Abstract

Purpose: The current diagnostic testing algorithm for Lynch syndrome (LS) is complex and often involves multiple follow-up germline and somatic tests. We aimed to describe the results of paired tumor/germline testing performed on a large cohort of patients with colorectal cancer (CRC) and endometrial cancer (EC) to better determine the utility of this novel testing methodology.

Materials and methods: We retrospectively reviewed a consecutive series of patients with CRC and EC undergoing paired tumor/germline analysis of the LS genes at a clinical diagnostic laboratory (N = 702). Microsatellite instability, MLH1 promoter hypermethylation, and germline testing of additional genes were performed if ordered. Patients were assigned to one of five groups on the basis of prior tumor screening and germline testing outcomes. Results for each group are described.

Results: Overall results were informative regarding an LS diagnosis for 76.1% and 60.8% of patients with mismatch-repair-deficient (MMRd) CRC and EC without and with prior germline testing, respectively. LS germline mutations were identified in 24.8% of patients in the group without prior germline testing, and interestingly, in 9.5% of patients with previous germline testing; four of these were discordant with prior tumor screening. Upon excluding patients with MLH1 promoter hypermethylation and germline mutations, biallelic somatic inactivation was seen in approximately 50% of patients with MMRd tumors across groups.

Conclusion: Paired testing identified a cause for MMRd tumors in 76% and 61% of patients without and with prior LS germline testing, respectively. Findings support inclusion of tumor sequencing as well as comprehensive LS germline testing in the LS testing algorithm. Paired testing offers a complete, convenient evaluation for LS with high diagnostic resolution.

FIG 1.

Paired tumor/germline patients by prior testing and tumor screening history. Abnormal immunohistochemistry (IHC): abnormal IHC screening because of loss of expression of at least one mismatch repair (MMR) protein as reported on external final IHC result; weak/equivocal staining: MMR protein expression reported as weak, equivocal, attenuated, nearly absent, segmental, partial, focal, decreased, or indeterminate without complete loss of any other MMR proteins on external final IHC result. (*) Three patients submitted for paired testing were removed from additional analysis; two from group 1 because microsatellite instability (MSI) results from paired testing were microsatellite stable (MSS) and one from group 3 because MSI results from paired testing were MSI high (MSI-H). CRC, colorectal cancer; EC, endometrial cancer; MSI-L, low microsatellite instability.

FIG 2.

Comparison of paired tumor/germline testing outcome for patients with mismatch repair deficient (MMRd) tumors by germline testing history. (A) Overall distribution of paired testing results in patients with MMRd tumors. (B) Paired testing outcomes after removing patients with germline Lynch syndrome mutations and MLH1 promoter hypermethylation from each group. Inconclusive: patients in whom abnormal immunohistochemistry (IHC) result is unexplained by somatic or germline findings. Double somatic: patients with somatic inactivation of an MMR gene as a result of double somatic mutations (or single somatic mutation with copy-neutral loss of heterozygosity) in the same gene and concordant with IHC result. MLH1 promoter hypermethylation (MPH): patients with MLH1 promoter hypermethylation identified as explanatory for MMRd. Germline mutation: patients with a germline MMR mutation/likely pathogenic variant. Group 1 without previous testing: patients in whom germline testing of one or more MMR genes, either in whole or in part, was not previously performed. Group 2 with previous testing: patients in whom germline testing of one or more MMR genes was previously performed.