Northern Ireland Regional Genetics Service First patient with 16p11.2 submicroscopic deletion detected by array CGH in Northern Ireland Regional Genetics Service Lisa Bradley, Simon McCullough, Peter McGrattan, Susan McNerlan, Geoff Smith, Mervyn Humphreys, Vivienne McConnell Abstract The 15 month old female proband is the first child of non-consanguineous parents born after a pregnancy using assisted conception. Complex congenital cardiac disease, dysmorphism, cleft palate, right strabismus, growth retardation and developmental delay, are the cardinal features observed in the proband. Echocardiogram showed complete AVSD, supracardiac TAPVD and PDA. The following genetic investigations were normal: karyotype, 22q11 and 9q34.3 FISH, subtelomere (P036D) MLPA screen and microdeletion/duplication (P245A2) MLPA screen. Both parents showed dysmorphic features. The 41 year old mother also had learning difficulties and short stature, while the 34 year old father had two brothers with learning difficulties and epilepsy and reported a great-uncle dying at 3 days old of an unknown cardiac condition. Array CGH analysis using the Agilent Oligo 4x44k platform detected an approximate 0.5Mb deletion within the short arm of chromosome 16, region 16p11.2, from base pair 29581456 to base pair 30106254. This ~0.5 Mb deletion was confirmed using the Illumina HumanCytoSNP-12 platform and microdeletion/duplication (P297B1) MLPA screen. Both parental karyotypes and microdeletion/duplication (P245A2 and P297B1) MLPA screens were normal, excluding an inherited abnormality. Parental array based CGH results are pending. The case presented is the first positive microarray analysis result from the NIRGS following the recent introduction of array CGH to our repertoire of clinical genetic investigations. Proband At 3days (refer Fig 1) Complete AVSD, supracardiac TAPVD and PDA Large cleft palate Nasogastric tube (NG) feeding Growth parameters: wt 2nd, OFC 25th centile Dysmorphism: Worried expression, flat facies Hypertelorism, downslanting palpebral fissures Broad nasal bridge, upturned nose Smooth philtrum, small, downturned mouth Micrognathia Low set posteriorly rotated ears At 11mths (refer Fig 2) NG tube and spoon feeding Motor delay Right convergent strabismus Dysmorphism more pronounced: High anterior hairline Long, straight eyebrows, long eyelashes Epicanthic folds Prominent cupid’s bow, thick lower lip Philtrum now centrally grooved Digits:spatula- like ends Methods DNA was extracted from peripheral blood lymphocytes using the Puregene method and concentration and purity determined using a nanophotometer (Implen). DNA was analysed on two microarray systems according to the manufacturer’s protocol: 1) Agilent 4x44K oligonucleotide array CGH platform using DNA analytics software, 2) Illumina Human CytoSNP-12 bead chip system and Karyostudio software. Results Array CGH using the Agilent Oligo 4x44k platform detected an approximate 0.5Mb deletion within the short arm of chromosome 16 at band 16(p11.2) from base pair 29581456 to 30106254 (Figure 3). This deleted region not only harbours a range of known genes but also includes several Redon CNVs, genomic variants and 10 reported Decipher features (Figure 4). This deletion was confirmed using the Illumina cytoSNP-12 platform and microdeletion/microduplication (P297B1) MLPA (MRC Holland). Figure 1: Proband at 3 days Figure 2: Proband at 11 months Figure 3. Array CGH profile of chromosome 16 from Agilent 4x44k array using DNA analytics software showing deleted region at 16p11.2. Upper panel shows an enlargement of the deleted region Figure 4. ENSEMBL overview of deleted region at 16p11.2, size ~0.5Mb. Shows other known decipher cases with duplications (green; n=3) and deletions (pink/red; n=10) including regions of genomic variation. Discussion The development of array comparative genomic hybridisation (array CGH), which enables the simultaneous testing of multiple loci for copy number differences is revolutionising cytogenetic testing and facilitating a ‘reverse dysmorphology’ approach, with discovery of the cytogenic abnormality preceding the definition of the corresponding clinical phenotype (1). This is aptly illustrated in the literature in recent years with regard to chromosome 16p which is especially rich in segmentally duplicated sequences (2). Genomic instability created by misalignment of these segmental duplications during meiosis followed by nonallelic homologous recombination can create novel microdeletion/duplication syndromes (3). This mechanism accounts for our reported cytogenetic results. We suggest, from the clinical description of our patient and several others reported in the literature (3,4,5,7) that a phenotype and facial gestalt is emerging for this 16p11.2 microdeletion. Although the earliest case description reported multiple congenital malformations and death at 5mths (4) the picture now emerging is of a variable clinical outcome, including a normal phenotype (5) (analogous to the 22q11 del syndrome). Developmental delay, learning difficulties (especially speech), feeding difficulties, growth retardation, cardiac defects and ocular anomalies are sufficiently recurrent to be regarded as significant components of the phenotype, while orofacial clefting and minor digit anomalies are occasionally observed. Frequent dysmorphic features include flat facies, micrognathia, wide nasal bridge with anteverted nares, low set, posteriorly rotated ears, downslanting palpebral fissures, epicanthic folds, hypertelorism, a longish philtrim, thin upper lip and high arched palate (1,3,4,5,7). It remains to be seen if autism is part of the 16p11.2 spectrum. In an Icelandic cohort the carrier frequency among patients with autism was 1% (and 0.1% among patients with a psychiatric or language disorder) (6) while screening of 4284 patients with MR/MCA detected 22pts with a 16p11.2 del, none of whom had autism (5). Acknowledgements Thanks to Agilent Technologies for loan of the microarray scanner used in this investigation l Conclusion The continued application of array CGH coupled with descriptions of 16p11.2 del patients should clarify the spectrum of this emerging syndrome 4. Hernando et al J Med Genet 2002; 39:e24 5. Bijlsma et al Eur J Med Genet 2009; 52:77-87 6. Weiss et al N Engl J Med 2008; 358:667-675 7. Ghebranious et al Am J Med Genet 2007; 143A:1462-71 References 1. Slavotinek Hum Genet 2008 124:1-17 2. Hannes et al J Med Genet 2009; 46:223-232 3. Ballif et al Nat Genet 2007; 39:1071-1073