Biallelic germline nonsense variant of MLH3 underlies polyposis predisposition

Abstract

Purpose: Some 10% of familial adenomatous polyposis (FAP) and 80% of attenuated polyposis (AFAP) cases remain molecularly unexplained. We scrutinized such cases by exome-wide and targeted methods to search for novel susceptibility genes.

Methods: Exome sequencing was conducted on 40 unexplained (mainly sporadic) cases with FAP or AFAP from Finland. The DNA mismatch repair (MMR) gene MLH3 (MutL Homolog 3) was pinpointed and prompted a subsequent screen of ~1000 Swedish patients referred to clinical panel sequencing for colon tumor susceptibility.

Results: Three homozygous carriers of a truncating variant in MLH3, c.3563C>G, p.Ser1188Ter, were identified among the index cases from the Finnish series. An additional biallelic carrier of the same variant was present in the Swedish series. All four patients shared a 0.8-Mb core haplotype around MLH3, suggesting a founder variant. Colorectal polyps from variant carriers showed no instability at mono-, di-, tri-, or tetranucleotide repeats, in agreement with previous findings of a minor role of MLH3 in MMR. Multiple loci were affected by loss of heterozygosity, suggesting chromosomal instability.

Conclusion: Our results show that a biallelic nonsense variant of MLH3 underlies a novel syndrome with susceptibility to classical or attenuated adenomatous polyposis and possibly extracolonic tumors, including breast cancer.

Keywords: MLH3; biallelic germline variant; polyposis.

Fig. 1. Discovery of the pathogenic MLH3 variant in five polyposis families

(a) Flow-chart of this investigation including the Finnish and Swedish arms. MLH3 findings and methods used for their identification are shown. (b) Pedigrees of polyposis families with the MLH3 p.Ser1188Ter variant. Families 158, 168, 177, and 1007 are from Finland and family SWE is from Sweden. The pedigrees were generated with Pedigree Chart Designer. Numbers below the symbols are patient identifiers. Arrow denotes the index person. Carrier status for the MLH3 c.3563C>G, p.Ser1188Ter variant is shown (+/+, homozygous carrier, +/− heterozygous carrier). Tumor manifestations and age at diagnosis (years) are given below the patient symbol. AFAP attenuated familial adenomatous polyposis, ES exome sequencing, FAP familial adenomatous polyposis, LS Lynch syndrome.

Fig. 2. Origin and molecular characteristics of the MLH3 p.Ser1188Ter variant

(a) Shared founder haplotype in carriers of the MLH3 p.Ser1188Ter variant. The haplotypes for an 8.8-Mb region between microsatellite markers D14S268 and D14S1000 are shown for the index patients from the indicated families (158, 168, 177, and SWE, homozygous carriers; 1007, heterozygous carrier). The conserved disease haplotype associated with the p.Ser1188Ter variant (between positions 73,271,847 and 75,777,880) is shaded. Deviation from the conserved haplotype at D14S1047 in SWE could result from either mutation or recombination. A core haplotype of 0.8 Mb (between D14S1047 and D14S284) that includes the MLH3 gene and is shared by all p.Ser1188Ter variant carriers is indicated by brackets. (b) Location of the p.Ser1188Ter variant relative to the main functional domains of MLH3. A lollipop diagram of the MLH3 protein was created by MutationMapper. The functional domains are as follows: ATPase domain, MMR (DNA mismatch repair domain), and MutL_C (MutL C terminal dimerization domain). (c) Results from microsatellite instability (MSI) and allelic imbalance (AI)/loss of heterozygosity (LOH) analyses of colorectal adenomas from MLH3 p.Ser1188Ter variant carriers. The markers used were BAT25 (mononucleotide repeat from chromosome 4), D17S250 (dinucleotide repeat from chromosome 17), and D9S242 (tetranucleotide repeat from chromosome 9). All tumors were microsatellite-stable (MSS, marker BAT25). Allelic imbalance was evident with D17S250 and D9S242.