18818 Med Genet 1992; 29: 188-190

Absence of linkage of Noonan syndrome to the 1 neurofibromatosis type locus J Med Genet: first published as 10.1136/jmg.29.3.188 on 1 March 1992. Downloaded from

M Sharland, R Taylor, M A Patton, S Jeffery

Abstract 1, and his father with definite neurofibromato- Eleven families with Noonan syndrome sis type 1 and a mild Noonan syndrome pheno- in either two or three generations have type. One possible cause of this close associ- been identified. Following the reports of ation is that the genes for Noonan syndrome subjects with features of both Noonan and neurofibromatosis type 1 are contiguous, syndrome and neurofibromatosis type 1, and that a large deletion involving both genes these pedigrees have been studied using produces the combined phenotype. In this a number of probes at the neuro- first report of linkage analysis in Noonan syn- fibromatosis type 1 locus (17q11). A signi- drome, we therefore studied families with ficantly negative lod score was obtained Noonan syndrome and no clinical evidence of with the intragenic probe NF1-C2, sug- neurofibromatosis, with a number of probes gesting that the genes for Noonan syn- from the neurofibromatosis type 1 locus on drome and neurofibromatosis type 1 are chromosome 17ql 1. not contiguous. Materials and methods FAMILY SAMPLE Noonan syndrome is a condition characterised As part of a clinical and genetic study of by , dysmorphic facies, and con- Noonan syndrome, over 100 families have genital heart disease. Following the original been seen at St George's Hospital, London. paper by Noonan and Ehmke,l it has been Detailed pedigrees were obtained and all avail- recognised that Noonan syndrome is a major able first degree relatives of the proband were cause of congenital heart disease, and is inher- carefully examined for clinical stigmata of ited as an autosomal dominant trait.2 The Noonan syndrome. From these families, 11 Noonan syndrome phenotype becomes less pedigrees were selected where subjects with obvious with age,3 and clear familial transmis- Noonan syndrome could be clearly identified sion can be established in only a proportion of in either two or three generations. In each of cases.4 In other families, Noonan syndrome in these pedigrees, the child proband had classi-

the child may arise either as the result of a new cal features of Noonan syndrome, a normal http://jmg.bmj.com/ or be inherited from an affected , and was also diagnosed as having parent. Because of the variability of the adult Noonan syndrome on the scoring system phenotype, it is very difficult to discriminate devised by Duncan et al.15 The diagnosis was between these possibilities in a proportion of made in the adult cases on clinical evaluation families attending for genetic counselling. The of a typical Noonan syndrome phenotype. localisation and subsequent cloning of the Echocardiography was performed on all sub-

gene for Noonan syndrome would be of great jects with an abnormal cardiac examination. on September 27, 2021 by guest. Protected copyright. benefit both for providing genetic counselling The pedigrees are shown in the figure. The to families, and to aid understanding of the pedigree structures confirm autosomal domin- pathogenesis of the condition. ant inheritance. Sporadic cases of apparent Noonan syn- drome have been described with varying chro- mosomal but none of the DNA ANALYSIS rearrangements,97 to 20 ml of EDTA reported cases has had a classical Noonan DNA was extracted from 10 Watson8 de- blood by the method of Kunkel et al.'6 Each syndrome phenotype. originally DNA sample was digested with the appropri- scribed three families with the combination of ate restriction enzyme as recommended in the valvular pulmonary stenosis, dull intelligence, manufacturers' instructions (Anglian). After and cafe au lait in whom subsequent South West Thames spots,' digestion, the sample was electrophoresed in a Regional Genetic analysis has shown linkage to the neurofibro- 1% agarose gel in TBE buffer. The DNA was Service, St George's matosis type 1 locus.9 The association of neuro- transferred from the gel to a nylon membrane Hospital Medical a Noonan School, Cranmer fibromatosis type 1 with syndrome (Hybond N, Amersham) using the method of Terrace, London phenotype was then extended by Allanson et Southern."7 Filters were then prehybridised in SW17 ORE. al,'0 and the possible causes of the association SDS M Sharland 0-5 mol/I phosphate buffer, pH 7-0/7% R Taylor have been clearly summarised by Opitz et al.11 solution at 65°C. Hybridisation was performed M A Patton At least 11 further cases have been reported of using probes labelled with 32P [dCTP] (NEN/ S Jeffery subjects with both neurofibromatosis type 1 DuPont) by the random primer method'8 Correspondence to and Noonan syndrome.'2-'4 One family is par- at 65°C for 16 hours. The filters were then Dr Sharland. ticularly convincing,'4 consisting of an 18 year washed to a stringency of 1 x SSC, 0 1% Received 23 April 1991. man with both the classical features of SDS for 20 minutes at 65'C and were auto- Revised version accepted old 1 July 1991. Noonan syndrome and neurofibromatosis type radiographed for one to three days at - 70°C. Absence of linkage of Noonan syndrome to the neurofibromatosis type 1 locus 189 118 9 37 110 2.2 1.2 1.1 1.1 1.2 1.1 1.1 J Med Genet: first published as 10.1136/jmg.29.3.188 on 1 March 1992. Downloaded from 1.2 1.2 1.1 1.2 2.2

1.1 2.2 1.1 1.2 2.2 2.2 2.2 1.1 1.1 1.1 51 21 41 16 1.2 1.1 1.1 1.2 1.1 1.1 1.2

1.2 1.2 1.1 1.2 1.1 1.1 1.2 1.2 1.2 1.1

200 108 220 1.2 1.1 1.1 1.2 1.2 1.1

1.1 1.2 1.1 1.2 1.2 1.1 Pedigrees showing affected status and NFI-C2 allelic distribution.

The probes used were THH32, EW204, closely linked to the neurofibromatosis locus EW206, and EW207. Unfortunately, all of on 17qll.2. There are a number of explana- these probes were uninformative in all of the tions for this finding. pedigrees available. We subsequently obtained Subjects with both neurofibromatosis and a the intragenic probe NFl-C2, recognising an convincing Noonan syndrome phenotype are EcoRI polymorphism with alleles at 9 5 and rare. A significant number of patients with 7-3 kb,'9 which was kindly provided by Dr F neurofibromatosis type have an altered body Collins. habitus that clinically overlaps with Noonan http://jmg.bmj.com/ syndrome.2' Out of the 150 subjects with Noo- nan syndrome we have seen in our clinical project, only two sporadic cases had any signs LINKAGE ANALYSIS The marker data for all pedigrees using NFl- of both Noonan syndrome and neurofibroma- tosis type 1. However, two families studied C2 is shown in the figure. Of the 11 families with both neurofibromatosis type 1 and a mild studied, three showed recombination events. A Noonan phenotype appear to have large scale on September 27, 2021 by guest. Protected copyright. two point lod score analysis was carried out in the neurofibromatosis gene (T using the program LINKAGE,20 to test the possible association of Noonan syndrome with Strachan, personal communication). It is pos- sible that a large deletion in the neurofibroma- RFLP alleles generated by NFI-C2. A gene tosis gene produces an alteration in the gene frequency of in 2000 was assumed. At product that may have an effect on fetal de- 0 = 0-05 a lod of -2 91 was generated. velopment, causing a Noonan-like phenotype. A further explanation of the absence of linkage is that Noonan syndrome is a genetic- Discussion ally heterogeneous condition. A wide variation In only 10% of the families seen was the in the clinical picture of Noonan syndrome is clinical phenotype sufficiently clear in all recognised, both within and between families. generations to enable accurate assessment of We have, however, been impressed by the each subject's phenotypic status to be made. In clinical similarity of children from different the majority of families, two or more children families at the same age. We have not been able were affected. The cardiac lesion in all affected to identify subgroups of Noonan syndrome members was dysplastic pulmonary stenosis. within our patient group as yet, but it is still No subjects with hypertrophic cardiomyo- possible that in some families the gene for pathy were included. No patient had any clini- Noonan syndrome is linked to the neurofibro- cal evidence of neurofibromatosis type 1. matosis type locus. When the segregation of the NFl-C2 alleles The most likely explanation of these find- and Noonan syndrome is considered, a signi- ings, however, is that the locus of the gene for ficantly negative lod score is obtained from the Noonan syndrome is not at 17q 11, and further pedigrees. This study suggests that the gene linkage studies are required to locate the gene for Noonan syndrome in these families is not for this autosomal dominant condition. 190 Sharland, Taylor, Patton, Jfeffery

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