Non-hotspot PIK3CA mutations are more frequent in CLOVES than in common or combined lymphatic malformations

Pascal Brouillard,Angela Queisser,Frank Hammer,Nassim Homayun Sepehr,Anne Dompmartin,Annamaria Weitz-Tuoretmaa,Matthieu J Schlögel,Philippe Clapuyt,Simon Boutry,Miikka Vikkula,Elodie Fastré,Sandra Schmitz,Jussi Laranne,Laurence M Boon,Catheline Vilain,Louise Pasquesoone,Raphaël Helaers

doi:10.1186/s13023-021-01898-y

Abstract

BackgroundTheragnostic management, treatment according to precise pathological molecular targets, requests to unravel patients’ genotypes. We used targeted next-generation sequencing (NGS) or digital droplet polymerase chain reaction (ddPCR) to screen for somatic PIK3CA mutations on DNA extracted from resected lesional tissue or lymphatic endothelial cells (LECs) isolated from lesions. Our cohort (n = 143) was composed of unrelated patients suffering from a common lymphatic malformation (LM), a combined lymphatic malformation [lymphatico-venous malformation (LVM), capillaro-lymphatic malformation (CLM), capillaro-lymphatico-venous malformation (CLVM)], or a syndrome [CLVM with hypertrophy (Klippel-Trenaunay-Weber syndrome, KTS), congenital lipomatous overgrowth-vascular malformations-epidermal nevi -syndrome (CLOVES), unclassified PIK3CA-related overgrowth syndrome (PROS) or unclassified vascular (lymphatic) anomaly syndrome (UVA)].ResultsWe identified a somatic PIK3CA mutation in resected lesions of 108 out of 143 patients (75.5%). The frequency of the variant allele ranged from 0.54 to 25.33% in tissues, and up to 47% in isolated endothelial cells. We detected a statistically significant difference in the distribution of mutations between patients with common and combined LM compared to the syndromes, but not with KTS. Moreover, the variant allele frequency was higher in the syndromes.ConclusionsMost patients with an common or combined lymphatic malformation with or without overgrowth harbour a somatic PIK3CA mutation. However, in about a quarter of patients, no such mutation was detected, suggesting the existence of (an)other cause(s). We detected a hotspot mutation more frequently in common and combined LMs compared to syndromic cases (CLOVES and PROS). Diagnostic genotyping should thus not be limited to PIK3CA hotspot mutations. Moreover, the higher mutant allele frequency in syndromes suggests a wider distribution in patients’ tissues, facilitating detection. Clinical trials have demonstrated efficacy of Sirolimus and Alpelisib in treating patients with an LM or PROS. Genotyping might lead to an increase in efficacy, as treatments could be more targeted, and responses could vary depending on presence and type of PIK3CA-mutation.

Highlights

Theragnostic management, treatment according to precise pathological molecular targets, requests to unravel patients’ genotypes
To assess the contribution of PIK3CA mutations in common, combined and syndromic lymphatic malformation (LM) (Table 1), we screened DNA extracted from frozen tissues or isolated cells from 143 patients: 105 common LM, 3 lymphatico-venous malformation (LVM), 1 capillaro-lymphatic malformation (CLM), 7 capillaro-lymphatico-venous malformation (CLVM), 4 KTS, 14 congenital lipomatous overgrowth-vascular malformations-epidermal nevi -syndrome (CLOVES), 7 PIK3CA-related overgrowth syndrome (PROS) and 2 unclassified vascular (lymphatic) anomaly syndrome (UVA)
PIK3CA mutations were found in 78/105 common LM (74.3%), 3/3 LVM (100%), 6/7 CLVM (85.7%), 4/4 KTS (100%), 12/14 CLOVES (85.7%) and 5/7 PROS (71.4%) (Table 1)

Summary

Results

To assess the contribution of PIK3CA mutations in common, combined and syndromic LM (Table 1), we screened DNA extracted from frozen tissues or isolated cells from 143 patients: 105 common LM, 3 LVM, 1 CLM, 7 CLVM, 4 KTS, 14 CLOVES, 7 PROS and 2 UVA. Strong enrichment of the somatic mutation was observed within the isolated endothelial cells (ECs) and especially lymphatic ECs. We detected a statistically significant difference in the distribution of hotspot mutations and non-hotspot mutations between common and combined LM compared to the syndromes (p value = 1.329 × 10–7), to CLOVES (p value = 1.441 × 10–5) or to unclassified PROS (p value = 1.207 × 10–4), but not to KTS (Fig. 3 and Table 3). Out of the altogether 367 patients reported by us and others, 54 (14.7%) are PIK3CA-negative: 38 LM, 1 CLM, 3 CLVM, 1 KTS, 4 CLOVES, 5 PROS and 2 UVA Some of these could just be under the threshold of detection.

Conclusions

Background

Discussion

Conclusion

Methods