CLINICS 2012;67(8):981-985 CASE REPORT DOI:10.6061/clinics/2012(08)23 MSX2 copy number increase and craniosynostosis: copy number variation detected by array comparative genomic hybridization Karla de Oliveira Pelegrino,I Sofia Sugayama,II Karina Lezirovitz,III Ana Lu´ cia Catelani,I Fernando Kok,I Maria de Lourdes ChauffailleI I Instituto de Pesquisa e Desenvolvimento, Grupo Fleury, Sa˜ o Paulo/SP, Brazil. II Hospital das Cl´ınicas da Faculdade de Medicina da Universidade de S a˜o Paulo, Departamento de Pediatria, S a˜o Paulo/SP, Brazil. III Hospital das Cl´ınicas da Faculdade de Medicina da Universidade de S a˜o Paulo, Departamento de Otorrinolaringologia (LIM32), S a˜o Paulo/SP, Brazil. Email: mlourdes.chauffaille@grupofleury.com.br Tel.: 55 11 5014-7621. INTRODUCTION Craniosynostosis is a disorder characterized by the premature fusion of the calvarial sutures, causing an abnormal skull shape. Because this disorder occurs with a relatively high frequency, estimated at 1 in 2500 individuals, craniosynostosis represents a relevant medical problem (1). More than 100 syndromes have been shown to be associated with this disorder. It is believed that at least 20% of cases are due to single gene mutations or chromosome abnormalities (2). Although most cases can be considered both clinically and genetically heterogeneous, there is evidence that six genes are involved in many cases: MSX2, FGFR1, FGFR2, FGFR3, FBN1, and TWISTY (1). The Msh homeobox 2 (Msx2) gene encodes a home- odomain transcription factor protein and is expressed in migrating cranial neural crest cells during development (3). Msx2 has central roles in craniofacial development (4) and limb and tissue formation (6). Furthermore, Msx2 over- expression was demonstrated to be associated with cranio- synostosis in mice (5,7), indicating that normal craniofacial formation is dependent on the Msx2 dosage. Corroborating these findings, increases in the copy number in the MSX2 region have been reported in craniosynostosis patients (8-11). In the present study, we describe the findings from a whole-genome array comparative genomic hybridization (aCGH) analysis of a four-year-old patient exhibiting cranio- synostosis, microcephaly, psychomotor development delay, short stature, and cognitive impairment, among other ab- normalities. We found a gain in a region of chromosome 5q35.2 that contains MSX2 and has been shown to be associated with craniosynostosis. Quantitative PCR con- firmed the increase in the MSX2 copy number. CASE DESCRIPTION This study was approved by the Hospital das Cl´ınicas Institutional Ethics Committee, and written informed consent Copyright 2012 CLINICS – This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http:// creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non- commercial use, distribution, and reproduction in any medium, provided the original work is properly cited. No potential conflict of interest was reported. 981 was obtained from the family. The propositus (a girl) was the first live born child of a healthy and non-consanguineous couple (the mother was 38 years old and the father was 40 years old at the time of conception). The mother is of Japanese descent, and the father is of African descent. The mother had experienced two previous miscarriages due to incontinence of the endocervical isthmus. The pregnancy was uneventful except for mild vaginal bleeding during the first trimester. The patient was born preterm (31 weeks) after vaginal delivery, with Apgar scores of 8 and 9, a weight of 1550 g, a length of 38.5 cm, and an OFC of 28.0 cm. At birth, the patient exhibited respiratory distress and underwent orotracheal intubation (3 days). The patient was hypotonic, had poor suction, and exhibited neonatal jaundice and metabolic disturbances (hyponatremia and hypocalcemia). She was discharged at 2 months of age at a weight of 1910 g. The patient also exhibited failure to thrive, recurrent otitis and global developmental delay. She was able to sit without support at 15 months and able to walk with no support and say simple words at 24 months. Physical examination at 19 months showed a weight of 6500 g, a length of 68.5 cm, and an OFC of 42.5 cm. The patient’s face was peculiar, with a narrowing of the frontal diameter, upslanting palpebral fissures, an enlarged nasal root, low and posteriorly rotated ears, bilateral convergent strabismus, a high arched palate and mild retrognathia. Neuroimaging studies (CT) revealed coronary craniosynos- tosis. No heart defects were observed. Abdominal ultra- sonography showed a mild abnormality of the renal parenchyma, but the patient’s kidney function is normal. Phytohemagglutinin (PHA)-stimulated lymphocytes were subjected to G-banding karyotyping (500 bands) of the propositus and progenitors. The results were described according to the recommendations of the ISCN (2008). G- banding karyotype analysis of the patient showed a normal 46,XX pattern with no structural or numeric variations. The same was observed for both progenitors. For the aCGH experiments, DNA from the patient and progenitors was extracted from blood samples using a QIAamp DNA Blood Midi kit (Qiagen, Germany). Labeling and hybridization reactions were performed as recommended by the manufacturer (Perkin Elmer, Norwalk). aCGH was performed with Constitutional Chip 4.0 (Perkin Elmer, Norwalk), which includes 5000 BAC clones spotted in