Molecular characterization of 13 patients with PIK3CA‐related overgrowth spectrum using a targeted deep sequencing approach

Abstract Activating variants in the PIK3CA gene cause a heterogeneous spectrum of disorders that involve congenital or early‐onset segmental/focal overgrowth, now referred to as PIK3CA ‐related overgrowth spectrum (PROS). Historically, the clinical diagnoses of patients with PROS included a range of distinct syndromes, including CLOVES syndrome, dysplastic megalencephaly, hemimegalencephaly, focal cortical dysplasia, Klippel–Trenaunay syndrome, CLAPO syndrome, fibroadipose hyperplasia or overgrowth, hemihyperplasia multiple lipomatosis, and megalencephaly capillary malformation‐polymicrogyria (MCAP) syndrome. MCAP is a sporadic overgrowth disorder that exhibits core features of progressive megalencephaly, vascular malformations, distal limb malformations, cortical brain malformations, and connective tissue dysplasia. In 2012, our research group contributed to the identification of predominantly mosaic, gain‐of‐function variants in PIK3CA as an underlying genetic cause of the syndrome. Mosaic variants are technically more difficult to detect and require implementation of more sensitive sequencing technologies and less stringent variant calling algorithms. In this study, we demonstrated the utility of deep sequencing using the Illumina TruSight Oncology 500 (TSO500) sequencing panel in identifying variants with low allele fractions in a series of patients with PROS and suspected mosaicism: pathogenic, mosaic PIK3CA variants were identified in all 13 individuals, including 6 positive controls. This study highlights the importance of screening for low‐level mosaic variants in PROS patients. The use of targeted panels with deep sequencing in clinical genetic testing laboratories would improve diagnostic yield and accuracy within this patient population.