Arteriovenous malformation from a patient with JP-HHT harbours two second-hit somatic DNA alterations in SMAD4

Background Hereditary haemorrhagic telangiectasia (HHT) is an inherited disorder of vascular malformations. It is caused by inherited loss-of- function mutations in one of three genes, ENG, ACVRL1 or SMAD4. We recently showed that HHT-associated vascular malformations from liver, lung, brain and skin develop via a two-hit genetic mechanism resulting from biallelic loss-of- function mutations in either ENG or ACVRL1. Second-hit somatic mutations in SMAD4 have not been reported in HHT-associated vascular malformations. Here, we investigate a large, aggressively growing craniofacial arteriovenous malformation (AVM) from an individual with juvenile polyposis-HHT caused by a germline mutation in SMAD4. Methods We sequenced DNA from the AVM using a targeted gene sequencing panel to at least 1000X to identify somatic mutations that might contribute to the development of the AVM. We analysed whole genome SNP genotyping data using the algorithm Mosaic Chromosomal Alterations (MoChA) to identify somatic loss of heterozygosity. Results We confirmed the germline mutation in SMAD4 (c.1610A>T, p.Asp537Val) and identified a second-hit somatic mutation also in SMAD4 (c.350dup, p.Tyr117*) that occurred in trans relative to the germline mutation. We also identified somatic loss of heterozygosity on the q arm of chromosome 18, including SMAD4. Additionally, we confirmed that the loss of heterozygosity causes loss of the wild-type allele. Thus, we identified two independent somatic alterations in SMAD4 causing biallelic loss of SMAD4 function in the AVM tissue. Conclusion We identified biallelic loss of function of SMAD4 in a craniofacial AVM, evidence that SMAD4 also follows the two-hit mutation mechanism of HHT-associated vascular malformation pathogenesis.