Constitutional microsatellite instability, genotype, and phenotype correlations in Constitutional Mismatch Repair Deficiency

Background & Aims: Constitutional mismatch repair deficiency (CMMRD) is a rare recessive childhood cancer predisposition syndrome caused by germline mismatch repair (MMR) variants. Constitutional microsatellite instability (cMSI) is a CMMRD diagnostic hallmark and may associate with cancer risk. We quantified cMSI in a large CMMRD patient cohort to explore genotype-phenotype correlations, using novel MSI markers selected for instability in blood.

Methods: Three CMMRD, one Lynch syndrome (LS), and two control blood samples were genome sequenced to >120x depth. A pilot cohort of eight CMMRD and 38 control blood samples, and a blinded cohort of 56 CMMRD, eight suspected CMMRD, 40 LS, and 43 control blood samples were amplicon sequenced to 5000x depth. Sample cMSI score was calculated using a published method comparing microsatellite reference allele frequencies to 80 controls.

Results: Thirty-two mononucleotide repeats were selected from blood genome and pilot amplicon sequencing data. cMSI scoring using these MSI markers achieved 100% sensitivity (95% CI: 93.6-100.0%) and specificity (95% CI: 97.9-100.0%), was reproducible, and was superior to an established tumour MSI marker panel. Lower cMSI scores were found in CMMRD patients with MSH6 deficiency and patients with at least one MMR missense variant, whilst patients with biallelic truncating/copy number variants had higher scores. cMSI score did not correlate with age at first tumour.

Conclusions: We present a cheap and scalable cMSI assay that enhances CMMRD detection relative to existing methods. cMSI score is associated with MMR genotype but not phenotype, suggesting it is not a useful predictor of cancer risk.