Comprehensive molecular and clinicopathological analysis of vascular malformations: A study of 319 cases

Abstract

Vascular malformations are part of overgrowth syndromes characterized by somatic mosaic mutations or rarely by germline mutations. Due to their similarities and diversity, clinicopathological classification can be challenging. A comprehensive targeted Next Generation Sequencing screen using Unique Molecular Identifiers with a technical sensitivity of 1% mutant alleles was performed for frequently mutated positions in ≥21 genes on 319 formalin-fixed paraffin-embedded samples. In 132 out of 319 cases pathogenic mosaic mutations were detected affecting genes previously linked to vascular malformations e.g. PIK3CA (n=80), TEK (TIE2) (n=11), AKT1 (n=1), GNAQ (n=7), GNA11 (n=4), IDH1 (n=3), KRAS (n=9), and NRAS (n=1). Six cases harbored a combination of mutations in PIK3CA and in GNA11 (n=2), GNAQ (n=2), or IDH1 (n=2). Aberrations in PTEN and RASA1 with a variant allele frequency approaching 50% suggestive of germline origin were identified in six out of 102 cases tested; four contained a potential second hit at a lower allele frequency. Ninety-one of the total 142 pathogenic mutations were present at a variant allele frequency <10% illustrating the importance of sensitive molecular analysis. Clinicopathological characteristics showed a broad spectrum and overlap when correlated with molecular data. Sensitive screening of recurrently mutated genes in vascular malformations may help to confirm the diagnosis and reveals potential therapeutic options with a significant contribution of PIK3CA/mTOR and RAS-MAPK pathway mutations. The co-existence of two activating pathogenic mutations in parallel pathways illustrates potential treatment challenges and underlines the importance of multigene testing. Detected germline mutations have major clinical impact.

Keywords: molecular genetics; mosaic mutations; overgrowth syndromes; vascular malformations.

Figure 1

Overview of prospective and retrospective vascular malformation cohort. NGS‐based sequence analyses of a total of 319 cases of vascular malformations was performed in a routine diagnostic (prospective) and retrospective cohort consisting of histologically confirmed vascular anomalies. Molecular results grouped per mutated gene (top) and the clinicopathological entities in both cohorts (bottom). Abbreviations: AVM, arteriovenous malformation; SOGS, somatic overgrowth syndrome

Figure 2

Mutant allele frequencies of identified mutations. The mutant allele frequencies of all (likely) pathogenic mutations in both, the prospective (top) and retrospective (bottom) cohort, grouped per gene

Figure 3

Mutant allele frequencies in cases with two pathogenic hotspot mutations. In 6 cases, a combination of one PIK3CA with an additional pathogenic activating (GNA11 and GNAQ) or transforming (IDH1) hotspot mutation was identified. The mutant allele frequencies of both mutations are depicted and connected per case

Figure 4

Mutant allele frequencies in cases with disrupting mutations. In 10 cases of the retrospective cohort, disrupting mutations were identified in PTEN (9 cases) and RASA1 (1 case). In a subset of cases, an additional mutation was found. The mutant allele frequencies are depicted per case, with connected data points in case two mutations were identified

Figure 5

Venous/cavernous malformation. HE, ×100 [Color figure can be viewed at wileyonlinelibrary.com]

Figure 6

Arteriovenous malformation showing variation in wall thickness. HE, ×20 [Color figure can be viewed at wileyonlinelibrary.com]