Short Communication
Prenatal diagnosis and array comparative genomic hybridization
characterization of interstitial deletions of 8q23.3-q24.11 and 8q24.13
associated with Langer-Giedion syndrome and Cornelia de Lange syndrome
and haploinsufficiency of TRPS1, RAD21 and EXT1
Chih-Ping Chen a,b,c,d,e,f *, Ming-Huei Lin a, Yi-Yung Chen a, Schu-Rern Chern b, Yen-Ni Chen a, Peih-Shan Wu g, Chen-Wen Pan a, Meng-Shan Lee a and Wayseen Wang b,h
a Department of Obstetrics and Gynecology, Mackay Memorial Hospital, Taipei, Taiwan b Department of Medical Research, Mackay Memorial Hospital, Taipei, Taiwan
c Department of Biotechnology, Asia University, Taichung, Taiwan
d School of Chinese Medicine, College of Chinese Medicine, China Medical University, Taichung, Taiwan e Institute of Clinical and Community Health Nursing, National Yang-Ming University, Taipei, Taiwan f Department of Obstetrics and Gynecology, School of Medicine, National Yang-Ming University, Taipei,
Taiwan
g Gene Biodesign Co. Ltd, Taipei, Taiwan
h Department of Bioengineering, Tatung University, Taipei, Taiwan
* Correspondence to: Chih-Ping Chen, MD
Department of Obstetrics and Gynecology, Mackay Memorial Hospital 92, Section 2, Chung-Shan North Road, Taipei, Taiwan
Tel: +886-2-25433535; Fax: +886-2-25433642, +886-2-25232448 E-mail: [email protected]
Abstract
Objective: To present prenatal diagnosis of Langer-Giedion syndrome (LGS/TRPS type II) and
Cornelia de Lange syndrome-4 (CDLS4).
Materials and methods: A 36-year-old woman underwent amniocentesis at 17 weeks of gestation
because of advanced maternal age. Conventional cytogenetic analysis of amniocentesis revealed an interstitial deletion of chromosome 8q or del(8)(q23.3q24.13). Level II prenatal ultrasound examination revealed craniofacial dysmorphism. The pregnancy was terminated, and a malformed fetus was delivered with characteristic craniofacial dysmorphism of LGS/TRPS type II and CDLS4. Whole-genome array comparative genomic hybridization (aCGH) on the DNA extracted from cultured amniocytes was performed.
Results: The analysis of aCGH analysis revealed a result of arr 8q23.3q24.11
(116,087,006-118,969,399)1, 8q24.13 (123,086,851-124,470,847)1 (NCBI build 37) with a 2.88-Mb deletion of 8q23.3-q24.11 encompassing 6 OMIM genes of TRPS1, EIF3H, RAD21,
SLC30A8, MED30 and EXT1, and a 1.383-Mb deletion of 8q24.13 encompassing 4 OMIM
genes of ZHX2, DERL1, ZHX1 and ATAD2.
Conclusions: In the present case, the conventional cytogenetic analysis of cultured amniocytes
showed the result of del(8)(q23.3q24.13), whereas aCGH analysis of cultured amniocytes showed the result of deletions of 8q23.3-q24.11 and 8q24.13 with the presence of the segment 8q24.12. Therefore, aCGH provides the advantage of better understanding of the nature of interstitial deletion and genotype-phenotype correlation in this case.
Keywords: 8q23.3-q24.13 deletion, Cornelia de Lange syndrome-4, EXT1, Langer-Giedion
Introduction
An interstitial deletion of chromosome 8q including 8q23.3 and 8q24.11 has been associated with Langer-Giedion syndrome (LGS) and Cornelia de Lange syndrome-4 (CDLS4) [1-3]. LGS or trichorhinophalangeal syndrome (TRPS) type II (OMIM 150230) is a contiguous gene syndrome associated with loss of functional copies of the TRPS1 gene (OMIM 604386) at 8q23.3 and the
EXT1 gene (OMIM 608177) at 8q24.11 [4]. CDLS4 (OMIM 614701) is associated with
heterozygous mutations or deletion of the RAD21 gene (OMIM 606462) at 8q24.11 [1-3,5-6]. We previously reported a 19-year-old male with laxity of the skin and joints, facial dysmorphism, epilepsy, gastroesophageal reflux, cardiovascular defects, mental retardation, developmental delay, exostoses, scoliosis, an 8q23.3-q24.22 deletion, haploinsufficiency of TRPS1, RAD21, EXT1 and
KCNQ3, and LGS and CDLS4 [6]. Here, we present prenatal diagnosis of interstitial deletions of
8q23.3-q24.11 and 8q24.13 associated with LGS and CDLS4, and haploinsufficiency of TRPS1,
RAD21 and EXT1.
Methods and detection
Clinical description
A 36-year-old, gravida 3, para 0, woman underwent amniocentesis at 17 weeks of gestation because of advanced maternal age. The woman had experienced two spontaneous abortions. She and her husband were unrelated, and there was no family history of congenital malformations. Conventional cytogenetic analysis of amniocentesis revealed an interstitial deletion of chromosome 8q or del(8)(q23.3q24.13) (Fig. 1). Level II prenatal ultrasound examination at 21 weeks of gestation revealed craniofacial dysmorphism of bulbous nose, depressed nasal bridge, micrognathia, protruding upper lip, large protruding ears and large philtrum (Fig. 2). However, the fetal biometry of the head circumference, femur length and abdominal circumference were equivalent to 21 weeks of gestation. There were no short stature and no abnormalities of the long bones. The pregnancy was terminated at 22 weeks of gestation, and the 392-g malformed proband
was delivered with characteristic craniofacial dysmorphism of Langer-Giedion syndrome and Cornelia de Lange syndrome (Fig. 3).
Conventional cytogenetic analysis
Routine cytogenetic analysis by G-banding techniques was performed. Amniotic fluid, placental tissues, parental bloods and cord blood were collected, and the samples were subjected to cell culture according to the standard cytogenetic protocol.
Array comparative genomic hybridization (aCGH)
Whole-genome aCGH on the DNA extracted from cultured amniocytes was performed using NimbleGen ISCA Plus Cytogenetic Array (Roche NimbleGen, Madison, WI, USA). The NimbleGen ISCA Plus Cytogenetic Array has 630,000 probes and a median resolution of 15-20 kb across the entire genome according to the manufacturer's instruction. The DNA from cultured amniocytes was extracted first. It was done by following the manufacturer's protocol of QIAamp DNA Mini kit (Qiagen, Inc., Valencia, CA, USA). Then, the 0.5μg of the extracted DNA was labeled in Cy5 dye compared with equivalent amount of normal female gDNA (G1521, Promega, Madison, WI, USA) labeled in Cy3 dye to perform the aCGH experiment. The experiment was performed according to the procedures recommended from Roche NimbleGen ISCA plus Cytogenetic Array's user guide. The data were finally represented by using Nexus 6.1 (BioDiscovery, Hawthorne, CA, USA).
Results
G-banding chromosome analysis revealed a karyotype of 46,XY in the father, a karyotype of 45,X[3]/47,XXX[5]/46,XX[32] in the mother, and a karyotype of 46XY,del(8)(q23.3q24.13)dn in the amniotic fluid, cord blood and placental tissues. The analysis of aCGH analysis revealed a result of arr 8q23.3q24.11 (116,087,006-118,969,399)1, 8q24.13 (123,086,851-124,470,847)1 (NCBI build 37) with a 2.88-Mb deletion of 8q23.3-q24.11 encompassing 6 OMIM genes of
Discussion
Prenatal diagnosis of LGS has been previously described [7-8]. We additionally present prenatal diagnosis of interstitial deletion of 8q involving 8q23.3-q24.11 and 8q24.13 associated with LGS/TRPS type II and CDLS4. LGS/TRPS type II is characterized by a peculiar face with bulbous nose and large protruding ears, loose redundant skin, lax joints, phalangeal abnormalities of the hands and multiple exostoses, mild growth deficiency, mild to severe mental retardation, microcephaly and sparse scalp hair [4,6]. CDLS4 is characterized by a peculiar face with bushy eyebrows, synophrys, depressed nasal bridge, micrognathia, anteverted nares, microcephaly, prominent symphysis and spurs in the anterior angle of mandible, growth deficiency, mental retardation, gastroesophageal reflux, duplication of gut, malrotation of colon, volvus, pyloric stenosis, and occasional findings of seizures, congenital heart defects and inguinal hernia [5-6]. The present case shows that fetuses with LGS/TRPS type II and CDLS4 due to haploinsufficiency of TRPS1, RAD21 and EXT1 caused by 8q23.3-8q24.11 deletion may not present skeletal abnormalities and growth deficiencies on the second-trimester ultrasound but may present characteristic facial dysmorphism. The present case was prenatally diagnosed simply by routine amniocentesis because of advanced maternal age. Fairweather [7] first reported prenatal diagnosis of LGS/TRPS type II and del(8)(q24.1q24.3)dn in a fetus by amniocentesis due to abnormal prenatal ultrasound findings of a single umbilical artery, pulmonary stenosis with post-stenotic dilation, pericardial effusion and short femoral bones. Piotrowski et al [8] later reported prenatal diagnosis of LGS and 8q24 deletion at 32 weeks of gestation by amniocentesis due to fetal ascites with hydrometrocolpos presenting as an intra-abdominal cyst.
The present case had haploinsufficiency of TRPS1, EXT1 and RAD21. TRPS1 is located at 8q23.3 and encodes a GATA-type transcription factor that has 9 zinc-finger motifs involved in the development and differentiation of bones, kidneys and hair follicles [9]. Mutations or haploinsuffiency of TRPS1 causes TRPS type I and TRPS type III [10-14]. TRPS type I (OMIM 190350) is characterized by craniofacial dysmorphism of sparse scalp hair, bulbous tip of the nose,
long philtrum, protruding ears, cone-shaped epiphyses at the phalanges, hip malformations and short stature [10]. TRPS type III (OMIM 190351) is similar to TRPS type I but is characterized by severe short stature, severe brachydactyly and more severe shortening of all phalanges and metacarpals [12]. EXT1 is located at 8q24.11 and encodes exostosin 1 which is essential in the regulation of chondrocyte differentiation, ossification and apoptosis [15]. EXT1 is a tumor suppressor gene [16-18]. Mutations or haploinsufficiency of EXT1 will cause multiple exostoses type I (OMIM 133700) and chondrosarcoma (OMIM 215300) [19]. An interstitial deletion of 8q23.3-8q24.11 involving loss of the functional copies of TRPS1 and EXT1 causes LGS/TRPS type II. RAD21 is located at 8q24.11 and encodes nuclear matrix protein 1 which is essential for sister chromatid cohesion during mitosis and meiosis [1,20]. Haploinsufficiency or heterozygous loss-of-function mutations of RAD21 cause CDLS4 [1].
The peculiar aspect of the present case is the discrepancy between the conventional cytogenetics and aCGH. The aCGH analysis has the advantage of detecting microdeletion syndromes in the fetuses with a normal karyotype by conventional cytogenetic analysis [21-22] or an apparently balanced translocation by conventional cytogenetic analysis [23]. In the present case, the conventional cytogenetic analysis of cultured amniocytes showed the result of del(8) (q23.3q24.13), whereas aCGH analysis of cultured amniocytes showed the result of deletions of 8q23.3-q24.11 and 8q24.13 with the presence of the segment 8q24.12. Therefore, aCGH provides the advantage of better understanding of the nature of interstitial deletion and genotype-phenotype correlation in this case.
Acknowledgements
This work was supported by research grants NSC-101-2314-B-195-011-MY3 and MOST 103-2314-B-195-010 from the Ministry of Science and Technology and MMH-E-104-04 from Mackay
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Figure Legends
Fig. 1. A karyotype of 46,XY,del(8)(q23.3q24.13). The arrows indicate the breakpoints.
Fig. 2. Craniofacial dysmorphism on (A) two-dimensional and (B) three-dimensional ultrasound. Fig. 3. The proband at birth.
Fig. 4. Array comparative genomic hybridization on cultured amnicoytes shows a 2.88-Mb deletion at
chromosome bands 8q23.3-q24.11 [arr 8q23.3q24.11 (116,087,006-118,969,399)1] and a 1.383-Mb deletion of 8q24.13 [arr 8q24.13 (123,086,851-124,470,847)1] (NCBI build 37). (A)
