Page 30 - Clinical variability in Noonan syndrome with emphasison ear and eye

CHAPTER 2
Materials and Methods
For this observational study, we included family members of a large NS family, known in the Radboud University Hospital in the Netherlands. After informed consent was obtained, the family members were asked for prospective clinical evaluation of NS characteristics, photographs and genetic analyses. In total 21 family members participated in the clinical evaluation and in 14 family members a genetic analysis was performed. In one patient, the proband, all described NS and NS-like genes were analyzed and solely a SOS1 mutation was found.
In all 21 family members the clinical characteristics were scored and photographs were taken. The results were reviewed by two of the researchers including an experienced clinical geneticist. The characteristics were compared to the NS scoring system, as described by van der Burgt (2).
Genetic analysis
In total seventeen genes involved in RASopathies (including NS) were analyzed in the index person either by using NGS gene panel or Sanger sequencing (Ion AmpliSeq kit 2.0, Thermo Fisher Scientific). The sequenced genes are: A2ML1, BRAF, CBL, HRAS, KRAS, LZTR1, MAP2K1, MAP2K2, NRAS, PTPN11, RAF1, RIT1, RRAS, SOS1, SOS2, SHOC2 and SPRED1. In other family members only exon 20 of the SOS1 gene (NM_005633.3) containing the variant of interest was sequenced.
Results
Clinical results
Three generations of this family with ages ranging from 8 to 73 years old were included. Clinical examination revealed three individuals with the clinical diagnosis of definitive NS based on the van der Burgt criteria (2) among the family members with genetic analyses. In these three individuals, number III-7, IV-8 and IV-10 in Table 1, two had typical facial features including ptosis and one had a suggestive face. In all three a pectus excavatum was present. Based on the genetic analysis they all had a first-degree relative with NS. No cardiac abnormalities, short stature, cryptorchidism or lymphatic dysplasia were found in these individuals. Individual number IV-8 had a mild intellectual disability with a verbal intelligence quotient of 91 and a performance intelligence quotient of 76. The other seven affected family members had different NS characteristics, with milder phenotypes. No congenital heart diseases or short stature (below -2 standard deviation) were seen in the affected family members. In three NS
28