Pathogenic mobile element insertion in the MEN1 gene mimicking a deletion in MLPA: characterisation by long-read...

Mobile element insertions (MEIs) disrupting coding sequences are rare but clinically significant mutations that are often missed by standard next-generation sequencing (NGS) workflows. Multiple endocrine neoplasia type 1 (MEN1) is an autosomal dominant tumour predisposition syndrome caused by pathogenic variants in the MEN1 gene.

Patients included in this study were referred for genetic testing of hereditary cancer syndromes. In a validation of our NGS-based copy number variant detection pipeline, four cases showed discrepancies between NGS and multiplex ligation-dependent probe amplification (MLPA) results. Further investigation using soft-clipped read analysis and the MEI detection tool Scramble revealed an Alu insertion in the MEN1 gene. Long-read whole genome sequencing (LR-WGS) was used for validation and complete characterisation of the variant. Additionally, 2014 retrospective NGS samples were analysed with Scramble. Relatives of probands were tested with MLPA and PCR. Positive samples were analysed with Sanger sequencing.

The apparent exon 2 deletion detected by MLPA was shown to be a false positive caused by an Alu insertion within the probe binding site. LR-WGS confirmed the pathogenic variant (MEN1:c.143_144insAlu) and resolved its complete sequence. Screening of the retrospective cohort revealed no additional MEIs. Altogether, we identified 10 affected individuals from two Polish families carrying hereditary MEN1:c.143_144insAlu, which segregates with MEN1 syndrome.

We report a novel pathogenic Alu insertion in the MEN1 gene, which is associated with MEN1 syndrome. Our findings underscore the importance of incorporating MEI detection into routine diagnostics. It also emphasises the necessity of interpreting single-exon MLPA deletions cautiously, as recommended in the MLPA protocol.