Journal of Genetics (2019) 98:42 © Indian Academy of Sciences https://doi.org/10.1007/s12041-019-1090-5 RESEARCH ARTICLE Clinical presentation and genetic profiles of Chinese patients with velocardiofacial syndrome in a large referral centre DANDAN WU1,2, YANG CHEN1,2, QIMING CHEN1,2, GUOMING WANG1,2∗, XIAOFENG XU1,2,A.PENG3, JIN HAO4, JINGUANG HE5∗,LIHUANG6∗ and JIEWEN DAI1,2∗ 1Department of Oral and Cranio-maxillofacial Surgery, National Clinical Research Center for Oral Disease, Shanghai Ninth People’s Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, People’s Republic of China 2Shanghai Key Laboratory of Stomatology, Shanghai 200011, People’s Republic of China 3State Key Laboratory of Oral Diseases, West China School of Stomatology, Chengdu 610041, People’s Republic of China 4Harvard School of Dental Medicine, Harvard University, Boston 02125, USA 5Department of Plastic Surgery, Shanghai Ninth People’s Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200011, People’s Republic of China 6Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Fujian Medical University, Fuzhou, People’s Republic of China *For correspondence. E-mail: Jiewen Dai, [email protected]; Guoming Wang, [email protected]; Jinguang He, [email protected]; Li Huang, [email protected]. Received 29 November 2017; revised 8 January 2019; accepted 11 January 2019; published online 4 May 2019 Abstract. Diagnosis and treatment of velocardiofacial syndrome (VCFS) with variable genotypes and phenotypes are considered to be very complicated. Establishing an exact correlation between the phenotypes and genotypes of VCFS is still a challenging. In this paper, 88 Chinese VCFS patients were divided into five groups based on palatal anomalies and one or two of other four common phenotypes, and copy number variations (CNVs) were detected using multiplex ligation-dependent probe amplification (MLPA), array comparative genomic hybridization (aCGH) and quantitative polymerase chain reaction. The findings showed that palatal anomalies and characteristic malformation of face were important indicators for 22q11.2 microdeletion, and there was difference in the phenotypic spectrum between the duplication and deletion of 22q11.2. MLPA was a highly cost-effective, sensitive and preferred method for patients with 22q11.2 deletion or duplication. Our results also firstly reported that all three patients who simultaneously exhibited palatal anomalies and cognitive disorder, without other phenotypes, have Top3b duplication, which strongly suggested that Top3b may be a pathogenic gene for these patients. Further, the findings showed that patients with palatal anomalies and congenital heart disease or immune deficiency, with or without other uncommon phenotypes, exhibited heterogeneity in CNVs, including 4q34.1- qter, 6q25.3, 4q23, Xp11.4, 13q21.1, 17q23.2, 7p21.3, 2p11.2, 11q24.3 and 16q23.3, and some possible pathogenic genes, including BCOR, PRR20A, TBX2, SMYD1, KLKB1 and TULP4 have been suggested. For these patients, aCGH, whole genomic sequencing, combined with references and phenomics database to find pathogenic gene, may be choices of priority. Taking these findings together, we offered an alternative method for diagnosis of Chinese VCFS patients based on this phenotypic strategy. Keywords. diagnosis; Chinese patients; velocardiofacial syndrome patients; phenotypic strategy. Introduction Dandan Wu, Yang Chen and Qiming Chen contributed equally to Velocardiofacial syndrome (VCFS) (MIM: 192430, also this work. named DiGeorge syndrome, MIM: 188400), affects ∼1 Electronic supplementary material: The online version of this article (https://doi.org/10.1007/s12041-019-1090-5) contains supplemen- tary material, which is available to authorized users. 1 42 Page 2 of 11 Dandan Wu et al. in 2000–7000 individuals (Kobrynski and Sullivan 2007; Materials and methods Wu et al. 2013; Kruszka et al. 2017). Phenotypes in every patient with VCFS exhibited great variety, and more Patients than 180 clinical features involving almost all organs and systems have been reported. The five most common Eighty-eight patients, diagnosed as VCFS based on clini- symptoms for VCFS patients include palatal anomalies, cal and auxiliary examinations, from the Center for Cleft congenital heart disease (CHD), characteristic malfor- Lip and Palate, Shanghai Ninth People’s Hospital, Shang- mation of face (CMF), immune deficiency and cognitive hai Jiao Tong University School of Medicine were enrolled or behavioural disorder. However, no single phenotype in this study. Some patients’ samples and materials from occurs in all patients, and no patient would exhibit all our previous study also were included in this study (Wu reported phenotypes (Perez and Sullivan 2002; Kobryn- et al. 2013). Patients who exhibited at least two of the ski and Sullivan 2007; Lopez-Rivera et al. 2017). five most common phenotypes, including palatal abnor- Previous evidence showed that an autosomal dom- mality, CHD, CMF, immune deficiency and cognitive or inant microdeletion on the long arm of chromosome behavioural disorder were clinically diagnosed as VCFS. 22(22q11.2) was the most common pathogenic factor for Except for these five common phenotypes, all the other VCFS, and the vast majority of individuals with 22q11.2 phenotypes were defined as uncommon phenotypes. In microdeletion carry a typical 3-Mb deletion. In addition, this study, palatal abnormalities mean patients exhibited some patients carry a proximal 1.5-Mb deletion nested to submucosal cleft palate, occult submucous cleft palate, the typically deleted region (TDR), and only a few patients complete cleft palate (CCP) or congenital velopharyngeal exhibited atypical deletions overlapping or nonoverlap- insufficiency (CVPI). CMF mean patients exhibited all ping with the TDR (Perez and Sullivan 2002; Ensenauer of the following phenotypes: vertically long face, narrow et al. 2003; Kobrynski and Sullivan 2007; Panamonta palpebral fissures, fleshy nose with a broad nasal root, flat- et al. 2016a, b). However,except for 22q11.2 microdeletion, tened malar region and retrognathia. CHD mean patients previous findings also showed that some cases exhib- with one or several conotruncal CHDs, aortic arch anoma- ited clinical phenotypes similar to those of VCFS having lies or other phenotypes: tetralogy of fallot, atrial septal 22q11.2 duplication or copy number variations (CNVs) defect, interrupted aortic arch, patent ductus arteriosus on other chromosomes. These patients were also clinically and ventricular septal defect. Immune deficiency means diagnosed as VCFS by some geneticists and doctors (Wu patients had thymic hypoplasia, T-cell deficiency or history et al. 2013). of recurrent infections. Cognitive or behavioural disorders Establishing an exact correlation between the pheno- mean mild mental retardation or intellectual disability. types and genotypes of VCFS is still a challenge. With Every enrolled patient would receive a general physical the development of biotechnology, such as whole-genome examination in detail, and the phenotypes were diagnosed sequencing, array comparative genomic hybridization by specialist physicians. The basic information and clin- (aCGH) and multiplex ligation-dependent probe ampli- ical phenotypes of these patients are listed in table 1 in fication (MLPA), detection of genetic variation in patients electronic supplementary material at http://www.ias.ac.in/ has become easier (Jalali et al. 2008; Poirsier et al. 2016). jgenet/. However, narrowing the detecting area and focus on some In this study, we tried to classify VCFS patients based on potential target based on the phenotypes may increase the five most common phenotypes. All enrolled 88 patients the precision of diagnosis and reduce patients’ cost. in this study were divided into five groups (table 2 in elec- Human phenotype ontology offers a computational bridge tronic supplementary material): (i) group 1 (patients 1 to between genotypes and phenotypes (Zemojtel et al. 2014; 66): patients exhibited palatal anomalies and CMF, with Kohler et al. 2017). Our previous preliminary findings also or without other phenotypes; (ii) group 2 (patients 67 to showed that all VCFS patients with palatal anomalies and 69): patients exhibited palatal anomalies and cognitive CMF would have a 22q11.2 microdeletion (Wu et al. 2013). or behavioural disorder, with or without other uncom- All of these findings inspired us that VCFS patients may be mon phenotypes; (iii) group 3 (patients 70 to 80): patients divided into different groups based on clinical phenotypes, exhibited palatal anomalies and CHD, with or without and these different groups may exhibit different genetic other uncommon phenotypes; (iv) group 4 (patients 81 to variations. 86): patients exhibited palatal anomalies and immune defi- In this paper, 88 Chinese patients with clinical diagnosis ciency, with or without other uncommon phenotypesand of VCFS were divided into five groups based on palatal (v) group 5 (patients 87 and 88): there were two patients anomalies and one or two of other four common pheno- exhibiting palatal anomalies, cognitive or behavioural types, and these groups exhibited different CNVs. Further, disorder and immune deficiency, with or without other some new possible pathogenic genes also were found based uncommon phenotypes. on this strategy, which offered a useful method for classi- One hundred normal patients were included in this fication diagnosis of Chinese VCFS patients based on