- Email: [email protected]
Molecular cytogenetic characterization of Xp22.32→pter deletion and Xq26.3→qter duplication in a male fetus associated with 46,Y,rec(X)dup(Xq) inv(X)(p22.3q26.3), a hypoplastic left heart, short stature, and maternal X chromosome pericentric inversion
Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Contents lists available at ScienceDirect
Taiwanese Journal of Obstetrics & Gynecology journal homepage: www.tjog-online.com
Short Communication
Molecular cytogenetic characterization of Xp22.32/pter deletion and Xq26.3/qter duplication in a male fetus associated with 46,Y,rec(X) dup(Xq) inv(X)(p22.3q26.3), a hypoplastic left heart, short stature, and maternal X chromosome pericentric inversion Chih-Ping Chen a, b, c, d, e, f, *, Chen-Yu Chen a, g, h, Schu-Rern Chern b, Peih-Shan Wu i, Yen-Ni Chen a, Shin-Wen Chen a, Chen-Chi Lee a, Dai-Dyi Town a, Meng-Shan Lee a, Chien-Wen Yang b, Wayseen Wang b, j a
Department of Obstetrics and Gynecology, MacKay Memorial Hospital, Taipei, Taiwan 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 Department of Medicine, MacKay Medical College, New Taipei City, Taiwan h MacKay Junior College of Medicine, Nursing and Management, Taipei, Taiwan i Gene Biodesign Co. Ltd, Taipei, Taiwan j Department of Bioengineering, Tatung University, Taipei, Taiwan b
a r t i c l e i n f o
a b s t r a c t
Article history: Accepted 19 May 2016
Objective: We present molecular cytogenetic characterization of an Xp22.32/pter deletion and an Xq26.3/qter duplication in a male fetus with congenital malformations and maternal X chromosome pericentric inversion. Materials and Methods: A 22-year-old woman underwent amniocentesis at 17 weeks of gestation because of an abnormal maternal serum screening result. Prenatal ultrasound revealed a hypoplastic left heart and short limbs. Amniocentesis revealed a karyotype of 46,Y,der(X) t(X;?)(p22.31;?). The pregnancy was subsequently terminated, and a malformed fetus was delivered with short stature and facial dysmorphism. Repeat amniocentesis was performed before termination of the pregnancy. Array comparative genomic hybridization was performed on uncultured amniocytes and maternal blood. Conventional cytogenetic analysis was performed on cultured amniocytes, cord blood, and blood from both parents. Fluorescence in situ hybridization was performed on cultured amniocytes. Results: The maternal karyotype was 46,X,inv(X)(p22.3q26.3). The fetal karyotype was 46,Y, rec(X) dup(Xq)inv(X)(p22.3q26.3) or 46,Y, rec(X)(qter/q26.3::p22.3/qter). Array comparative genomic hybridization on uncultured amniocytes revealed a 4.56-Mb deletion of Xp22.33ep22.32 encompassing SHOX, CSF2RA, and ARSE, and a 19.22-Mb duplication of Xq26.3eq28 encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2 in the fetus. The mother did not have X chromosome imbalance. Conclusion: Detection of X chromosome aberration in a male fetus should give suspicion of a recombinant X chromosome derived from maternal X chromosome pericentric inversion. Copyright © 2016, Taiwan Association of Obstetrics & Gynecology. Published by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/ 4.0/).
Keywords: hypotrophic heart short stature Xp22.32/pter deletion Xq26.3/qter duplication Yp11.31/pter deletion
Introduction * Corresponding author. Department of Obstetrics and Gynecology, MacKay Memorial Hospital, 92, Section 2, Chung-Shan North Road, Taipei 10449, Taiwan. E-mail address: [email protected] (C.-P. Chen).
Xp-Xq rearrangements with duplication/deletion events involving Xq terminal duplication and Xp terminal deletion have
http://dx.doi.org/10.1016/j.tjog.2016.05.009 1028-4559/Copyright © 2016, Taiwan Association of Obstetrics & Gynecology. Published by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
706
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Figure 1. A karyotype of 46,X,inv(X)(p22.3q26.3) in the mother.
rarely been reported. The derivative or recombinant X chromosome with Xq duplication/Xp deletion can be the result of a meiotic recombination event in a parent carrying a pericentric X inversion or the result of direct X inheritance from a phenotypically normal mother carrying the same aberrant derivative X chromosome [1e9]. In case of one of the parents with a chromosome inversion, there is about 5% recurrence risk for the recombinant chromosome in the offspring [10,11]. Affected male patients with a recombinant X chromosome with terminal Xq duplication/Xp deletion may present: terminal Xq duplication syndrome, such as growth retardation, developmental delay, hypotonia, and genital abnormalities; and terminal Xp deletion syndrome, such as short stature, mental retardation, ichthyosis, Kallmann syndrome, skeletal dysplasia, chondrodysplasia punctata, and hypogonadism [6]. Here, we present molecular cytogenetic characterization of a recombinant X chromosome with terminal Xq duplication/Xp deletion in a fetus with congenital malformations and maternal X chromosome pericentric inversion. Materials and methods Clinical description During the first pregnancy, a 22-year-old, primigravid woman underwent amniocentesis at 17 weeks of gestation because of an abnormal maternal serum screening result showing a trisomy 18 risk of 1/262. Her husband was aged 26 years, and there was no family history of congenital malformations. The woman had a body height of 147 cm, and her husband had a body height of 170 cm. Prenatal ultrasound at 20 weeks of gestation revealed a hypoplastic
left heart and short limbs. The biparietal diameter was 5.19 cm (21.36 weeks), abdominal circumference 15.34 cm (20.54 weeks), femur length 2.69 cm (18.2 weeks), tibia length 2.3 cm (18 weeks), humerus length 1.7 cm (15 weeks), and ulna length 2.4 cm (17 weeks). Amniocentesis revealed a karyotype of 46,Y,der(X) t(X;?)(p22.31;?). The pregnancy was terminated at 24 weeks of gestation, and a malformed male fetus was delivered with a body weight of 412 g, a body length of 23.5 cm, short stature, short limbs, and facial dysmorphism of hypertelorism and low-set ears. Repeat amniocentesis was performed before termination of the pregnancy. Array comparative genomic hybridization (aCGH) was performed on the DNA extracted from uncultured amniocytes and maternal blood. Conventional cytogenetic analysis was performed on cultured amniocytes, cord blood, and parental bloods. Metaphase fluorescence in situ hybridization (FISH) was performed on cultured amniocytes. During the second pregnancy, the same mother underwent amniocentesis at 23 weeks of gestation, which revealed a karyotype of 46,Y,inv(X)(p22.3q26.3). aCGH was performed on the DNA extracted from cultured amniocytes, and metaphase FISH was performed on cultured amniocytes. Preterm birth occurred at 29 weeks of gestation, and a baby was delivered with a body weight of 1489 g, a body length of 45 cm, and a head circumference of 28.5 cm. The baby was normal when followed up at age 4 months with a body weight of 4.6 kg and a body length of 45 cm with no phenotypic abnormalities. aCGH Whole-genome aCGH on the DNA extracted from uncultured amniocytes in the first pregnancy was performed using ISCA Plus Cytogenetic Array (Roche NimbleGen, Madison, WI, USA). This
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
707
Figure 2. A karyotype of 46,Y,rec(X)dup(Xq)inv(X)(p22.3q26.3) or 46,Y,rec(X)(qter/q26.3::p22.3/qter) in the fetus of the first pregnancy.
array has 630,000 probes and a median resolution of 15e20 kb across the entire genome according to the manufacturer's instruction. Whole-genome aCGH on the DNAs extracted from maternal blood and cultured amniocytes in the second pregnancy was performed using CytoChip ISCA Array (Illumina, San Diego, CA, USA), which has 60,000 probes and a median resolution of 51 kb across the entire genome according to the manufacturer's instruction. Conventional cytogenetic analysis Routine cytogenetic analysis by G-banding techniques at the 550 bands of resolution was performed. Amniotic fluid, cord blood and parental blood were collected, and the samples were subjected to cell culture according to the standard blood cytogenetic protocol. FISH Metaphase FISH analysis on cultured amniocytes was performed using the bacterial artificial chromosome probes of RP11-513H18 (Xq27.3, 142,900,963e143,081,146; dye: FITC, spectrum green), RP11-155F12 (Xp22.3, 2,456,240e2,612,008; Yp11.32-p11.31, 2,406, 240e2,562,008; dye: Texas red, spectrum red), and RP11-22P2 (Xq24, 118,252,205e118,409,958; dye: Cy5, spectrum yellow) according to the standard FISH protocol. Results During the first pregnancy, cytogenetic analysis revealed a karyotype of 46,X,inv(X)(p22.3q26.3) (Figure 1) in the maternal blood,
a karyotype of 46,XY in the paternal blood and a karyotype of 46,Y,rec(X)dup(Xq)inv(X)(p22.3q26.3) or 46,Y,rec(X)(qter/q26.3:: p22.3/qter) (Figure 2) in the cultured amniocytes and the cord blood lymphocytes. aCGH on uncultured amniocytes revealed a 4.56-Mb deletion of Xp22.33ep22.32 [arr Xp22.33 (1e2,697,663) 0.5, Xp22.33p22.32 (2,697,663e4,557,932)0.2] encompassing SHOX, CSF2RA and ARSE, and a 19.22-Mb duplication of Xq26.3eq28 [arr Xq26.3q28 (136,052,757e155,270,560)2.0] encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2 in the fetus (Figure 3). Metaphase FISH analysis on the cultured amniocytes revealed two green signals in the terminal ends of the X chromosome and absence of the red signal in the X chromosome, indicating an Xp/Xq rearrangement with terminal Xp deletion/terminal Xq duplication (Figure 4). During the second pregnancy, cytogenetic analysis of the cultured amniocytes revealed a karyotype of 46,Y,inv(X) (p22.3q26.3) (Figure 5). aCGH analysis on cultured amniocytes and maternal blood revealed no genomic imbalance in X chromosome. Metaphase FISH analysis on the cultured amniocytes revealed the order of greenecentromereeyellowered in the X chromosome, indicating a pericentric inversion of X chromosome in the fetus (Figure 6). Discussion The present case had a 19.22-Mb duplication of Xq26.3eq28 encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2. MECP2 (OMIM 300005) encodes methyl-CpG-binding protein that can both activate and repress transcription. MECP2 is a critical dosage-sensitive gene. Deletion or loss-of-function mutation of MECP2 is responsible for a progressive neurological disorder of Rett
708
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Figure 3. Array comparative genomic hybridization on uncultured amniocytes of the first pregnancy shows 4.56-Mb deletion of Xp22.33ep22.32 encompassing SHOX, CSF2RA, and ARSE, and a 19.22-Mb duplication of Xq26.3eq28 encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2. (A, B, C) Zoom-in view of X chromosome.
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Figure 4. Metaphase fluorescence in situ hybridization on the cultured amniocytes of the first pregnancy using the bacterial artificial chromosome probes of RP11-513H18 (Xq27.3, green) and RP11-155F12 (Xp22.3, red) shows two green signals in the terminal ends of the X chromosome and absence of the red signal in the X chromosome, indicating a recombinant X chromosome in the fetus with an Xp/Xq rearrangement resulting in terminal Xp deletion/terminal Xq duplication.
709
syndrome (OMIM 312750). Duplication or gain-of-function mutation of MECP2 will cause MECP2 duplication syndrome, which is characterized by recurrent respiratory infections, severe mental retardation, seizures, autism, developmental regression, absent or delayed speech, infantile hypotonia, and progressive spasticity, and is 100% penetrant in affected male patients [12e14]. Fu et al [15] suggested that ventriculomegaly, hydrocephalus, agenesis of the corpus callosum, choroid plexus cysts, intrauterine growth restriction, and hydronephrosis might be the common ultrasound features in the fetuses with MECP2 duplication syndrome. MECP2 duplication syndrome have been reported in rec(X)dup(Xq)inv(X) offspring born to a pericentric inv(X) carrier mother [4,7e9]. Breman et al [8] identified two boys with MECP2 duplication syndrome with terminal Xp deletion, terminal Xq duplication and a recombinant X chromosome containing the duplicated material from Xq28 and Xp, resulting from a maternal pericentric X inversion as similarly presented in this case. SOX3 (OMIM 313430) encodes the SRY-BOX3 protein which is a transcription factor that is important in the regulation of nervous system development and embryogenesis such as gastrulation, neural induction, specification, and differentiation of many types of cells [16,17]. The dysfunction of the SOX3 protein disturbs pituitary development, and over- and underdosage of SOX3 is associated with infundibular hypoplasia and hypopituitarism [18]. Stankiewicz et al [19] reported a duplication of Xq26.2eq27.1 including SOX3 in a mother and daughter with short stature and dyslalia, and suggested a dosage effect of SOX3 in speech disorder in addition to short stature secondary to hypopituitarism. Stagi et al [20] reported a duplication of SOX3 in a boy with growth hormone deficiency, ocular dyspraxia, and intellectual disability, and suggested that
Figure 5. A karyotype of 46,Y,inv(X)(p22.3q26.3) in the fetus of the second pregnancy.
710
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
an int22h1/int22h2-mediated Xq28 duplication syndrome has been identified in males with X-linked intellectual disability, cognitive impairment, behavioral and psychiatric problems, recurrent infections and atopic diseases, obesity, distinctive facial features, and an Xq28 duplication overlapping the int22h1/int22h2 region and including RAB39B and CLIC2 [27e29]. The present case had a 4.56-Mb deletion of Xp22.33ep22.32 encompassing SHOX, CSF2RA, and ARSE. SHOX (OMIM 312865) encodes short stature homeobox protein. Deletions or mutations in SHOX cause LerieWeill dyschondrosteosis (OMIM 127300), Langer mesomelic dysplasia (OMIM 249700), and X-linked idiopathic familial short stature (OMIM 300582). CSF2RA (OMIM 306250) encodes colony-stimulating factor 2 receptor-a (CSF2RA). Mutations in CSF2RA cause pulmonary alveolar proteinosis due to CSF2RA deficiency. ARSE (OMIM 300180) encodes arylsulfatase E. Deletions or mutations in ARSE cause X-linked recessive chondrodysplasia punctata (OMIM 302950). In summary, we have presented molecular cytogenetic characterization of Xp deletion/Xq duplication in a malformed fetus with a recombinant X chromosome due to maternal X chromosome pericentric inversion, and we discuss the genotypeephenotype correlation. We suggest that detection of X chromosome aberration in a male fetus should give suspicion of a recombinant X chromosome derived from maternal X chromosome pericentric inversion. Conflict of interest Figure 6. Metaphase fluorescence in situ hybridization on the cultured amniocytes of the second pregnancy using the bacterial artificial chromosome probes of RP11-22P2 (Xq24, yellow), RP11-513H18 (Xq27.3, green), and RP11-155F12 (Xp22.3, red) shows the order of greenecentromereeyellowered in the X chromosome, indicating a pericentric inversion of X chromosome in the fetus.
SOX3 duplication should be considered in a male patient with short stature due to growth hormone deficiency associated with intellectual disability. Uguen et al [21] reported a duplication of SOX3 in three male fetuses with spina bifida and suggested that SOX3 duplication is a risk factor of neural tube defect. FMR1 (OMIM 309550) encodes fragile X mental retardation protein. A loss of expression of FMR1 results in fragile X mental retardation syndrome and fragile X tremor/ataxia syndrome. However, a duplication of FMR1 has been reported to be associated with a new X-linked mental retardation syndrome characterized by short stature, hypogonadism and facial dysmorphism [22,23]. Rio et al [22] reported familial interstitial Xq27.3eq28 duplication encompassing FMR1 but not MECP2 associated with an X-linked mental retardation syndrome in affected males. Hickey et al [23] reported a duplication of Xq27.3eq28 including FMR1 in association with an X-linked hypogonadism, gynecomastia, intellectual disability, short stature, and obesity syndrome in affected male patients. CLIC2 (OMIM 300138) encodes chloride intracellular channel 2 protein, which plays a role in the regulation of ryanodine receptor (RyR) intracellular Ca2þ release channels. Gain-of-function mutation of CLIC2 enhancing RyR channel activity has been associated with an X-linked intellectual disability, atrial fibrillation, cardiomegaly, congenital heart failure, and seizures [24,25]. RAB39B (OMIM 300774) encodes RAS-associated protein RAB39B which is a small GTPase involved in the regulation of vascular trafficking between membrane compartments. Vanmarsenille et al [26] found copy number gains of RAB39B in Xq28 in 4 male patients with mild intellectual disability and behavioral problems and suggested that increased dosage of RAB39B causes a disturbed neuronal development leading to cognitive impairment. Recently,
The authors have no conflicts of interest relevant to this article. Acknowledgments This work was supported by research grants MOST-103-2314-B195-010 and MOST-104-2314-B-195-009 from the Ministry of Science and Technology and MMH-E-105-04 from MacKay Memorial Hospital, Taipei, Taiwan. References [1] Mohandas T, Geller RL, Yen PH, Rosendorff J, Bernstein R, Yoshida A, et al. Cytogenetic and molecular studies on a recombinant human X chromosome: implications for the spreading of X chromosome inactivation. Proc Natl Acad Sci USA 1987;84:4954e8. [2] Schmidt M, Du Sart D, Kalitsis P, Leversha M, Dale S, Sheffield L, et al. Duplications of the X chromosome in males: evidence that most parts of the X chromosome can be active in two copies. Hum Genet 1991;86:519e21. [3] Vasquez AI, Rivera H, Bobadilla L, Crolla JA. A familial Xpþ chromosome, dup (Xq26.3/qter). J Med Genet 1995;32:891e3. [4] Madariaga ML, Rivera H. Familial inv(X)(p22q22): ovarian dysgenesis in two sisters with del Xq and fertility in one male carrier. Clin Genet 1997;52: 180e3. [5] Lammer EJ, Punglia DR, Fuchs AE, Rowe AG, Cotter PD. Inherited duplication of Xq27.2/qter: phenocopy of infantile PradereWilli syndrome. Clin Dysmorphol 2001;10:141e4. [6] Kokalj-Vokac N, Marcun-Varda N, Zagorac A, Erjavec-Skerget A, Zagradisnik B, Todorovic M, et al. Subterminal deletion/duplication event in an affected male due to maternal X chromosome pericentric inversion. Eur J Pediatr 2004;163: 658e63. [7] Bleyl SB, Byrne JLB, South ST, Dries DC, Stevenson DA, Rope AF, et al. Brachymesomelic dysplasia with Peters anomaly of the eye results from disruptions of the X chromosome near the SHOX and SOX3 genes. Am J Med Genet 2007;143A:2785e95. [8] Breman AM, Ramocki MB, Kang SHL, Williams M, Freedenberg D, Patel A, et al. MECP2 duplications in six patients with complex sex chromosome rearrangements. Eur J Hum Genet 2011;19:409e15. [9] Sanmann JN, Bishay DL, Starr LJ, Bell CA, Pickering DL, Stevens JM, et al. Characterization of six novel patients with MECP2 duplications due to unbalanced rearrangements of the X chromosome. Am J Med Genet 2012;158A: 1285e91. ne ticiens Français. Pericentric inversions in man. A French [10] Groupe de Cytoge ne t 1986;29:129e68. collaborative study. Ann Ge [11] Goodman BK, Shaffer LG, Rutberg J, Leppert M, Harum K, Gagos S, et al. Inherited duplication Xq27-qter at Xp22.3 in severely affected males:
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
[12]
[13] [14] [15]
[16] [17]
[18]
[19]
[20]
[21]
molecular cytogenetic evaluation and clinical description in three unrelated families. Am J Med Genet 1998;80:377e84. Sanlaville D, Prieur M, de Blois MC, Genevieve D, Lapierre JM, Ozilou C, et al. Functional disomy of the Xq28 chromosome region. Eur J Hum Genet 2005;13:579e85. Sanlaville D, Schluth-Bolard C, Turleau C. Distal Xq duplication and functional Xq disomy. Orphanet J Rare Dis 2009;4:4. Ramocki MB, Tavyev YJ, Peters SU. The MECP2 duplication syndrome. Am J Med Genet 2010;152A:1079e88. Fu F, Liu HL, Li R, Han J, Yang X, Min P, et al. Prenatal diagnosis of foetuses with congenital abnormalities and duplication of the MECP2 region. Gene 2014;546:222e5. Pevny LH, Lovell-Badge R. Sox genes find their feet. Curr Opin Genet Dev 1997;7:338e44. ndez A, Han Y, Pallavi B. Control of cell fate Lefebvre V, Dumitriu B, Penzo-Me and differentiation by Sry-related high-mobility-group box (Sox) transcription factors. Int J Biochem Cell Biol 2007;39:2195e214. Woods KS, Cundall M, Turton J, Rizotti K, Mehta A, Palmer R, et al. Over- and underdosage of SOX3 is associated with infundibular hypoplasia and hypopituitarism. Am J Hum Genet 2005;76:833e49. Stankiewicz P, Thiele H, Schlicker M, Cseke-Friedrich A, Bartel-Friedrich S, Yatsenko SA, et al. Duplication of Xq26.2-q27.1, including SOX3, in a mother and daughter with short stature and dyslalia. Am J Med Genet 2005;138A: 11e7. Stagi S, Lapi E, Pantaleo M, Traficante G, Giglio S, Seminara S, et al. A SOX3 (Xq26.3e27.3) duplication in a boy with growth hormone deficiency, ocular dyspraxia, and intellectual disability: a long-term follow-up and literature review. Hormones (Athens) 2014;13:552e60. mener-Redon S, Marcorelles P, De Braekeleer M. Uguen A, Talagas M, Que Duplication of SOX3 (Xq27) may be a risk factor for neural tube defects. Am J Med Genet 2015;167A:1676e8.
711
[22] Rio M, Malan V, Boissel S, Toutain A, Royer G, Gobin S, et al. Familial interstitial Xq27.3q28 duplication encompassing the FMR1 gene but not the MECP2 gene causes a new syndromic mental retardation condition. Eur J Hum Genet 2011;18:285e90. [23] Hickey SE, Walters-Sen L, Mosher TM, Pfau RB, Pyatt R, Snyder PJ, et al. Duplication of the Xq27.3eq28 region, including the FMR1 gene, in an Xlinked hypogonadism, gynecomastia, intellectual disability, short stature, and obesity syndrome. Am J Med Genet 2013;161A:2294e9. [24] Witham S, Takano K, Schwartz C, Alexov E. A missense mutation in CLIC2 associated with intellectual disability is predicted by in silico modeling to affect protein stability and dynamics. Proteins 2011;79:2444e54. [25] Takano K, Liu D, Tarpey P, Gallant E, Lam A, Witham S, et al. An X-linked channelopathy with cardiomegaly due to a CLIC2 mutation enhancing ryanodine receptor channel activity. Hum Mol Genet 2012;21:4497e507. [26] Vanmarsenille L, Giannandrea M, Fieremans N, Verbeeck J, Belet S, Raynaud M, et al. Increased dosage of RAB39B affects neuronal development and could explain the cognitive impairment in male patients with distal Xq28 copy number gains. Hum Mutat 2014;35:377e83. [27] Andersen EF, Baldwin EE, Ellingwood S, Smith R, Lamb AN. Xq28 duplication overlapping the int22h-1/int22h-2 region and including RAB39B and CLIC2 in a family with intellectual and developmental disability. Am J Med Genet 2014;164A:1795e801. [28] El-Hattab AW, Schaaf CP, Fang P, Roeder E, Kimonis VE, Church JA, et al. Clinical characterization of int22h1/int22h2-mediated Xq28 duplication/deletion: new cases and literature review. BMC Med Genet 2015;16:12. [29] El-Hattab AW, Schaaf CP, Cheung SW. Xq28 duplication syndrome, Int22h1/ Int22h2 mediated. In: Pagon RA, Adam MP, Ardinger HH, Wallace SE, Amemiya A, Bean LJH, et al, editors. GeneReviews. Seattle (WA): University of Washington, Seattle; 1993e2016. http://www.ncbi.nlm.nih.gov/books/ NBK349624/. [accessed 10.03.16].
Contents lists available at ScienceDirect
Taiwanese Journal of Obstetrics & Gynecology journal homepage: www.tjog-online.com
Short Communication
Molecular cytogenetic characterization of Xp22.32/pter deletion and Xq26.3/qter duplication in a male fetus associated with 46,Y,rec(X) dup(Xq) inv(X)(p22.3q26.3), a hypoplastic left heart, short stature, and maternal X chromosome pericentric inversion Chih-Ping Chen a, b, c, d, e, f, *, Chen-Yu Chen a, g, h, Schu-Rern Chern b, Peih-Shan Wu i, Yen-Ni Chen a, Shin-Wen Chen a, Chen-Chi Lee a, Dai-Dyi Town a, Meng-Shan Lee a, Chien-Wen Yang b, Wayseen Wang b, j a
Department of Obstetrics and Gynecology, MacKay Memorial Hospital, Taipei, Taiwan 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 Department of Medicine, MacKay Medical College, New Taipei City, Taiwan h MacKay Junior College of Medicine, Nursing and Management, Taipei, Taiwan i Gene Biodesign Co. Ltd, Taipei, Taiwan j Department of Bioengineering, Tatung University, Taipei, Taiwan b
a r t i c l e i n f o
a b s t r a c t
Article history: Accepted 19 May 2016
Objective: We present molecular cytogenetic characterization of an Xp22.32/pter deletion and an Xq26.3/qter duplication in a male fetus with congenital malformations and maternal X chromosome pericentric inversion. Materials and Methods: A 22-year-old woman underwent amniocentesis at 17 weeks of gestation because of an abnormal maternal serum screening result. Prenatal ultrasound revealed a hypoplastic left heart and short limbs. Amniocentesis revealed a karyotype of 46,Y,der(X) t(X;?)(p22.31;?). The pregnancy was subsequently terminated, and a malformed fetus was delivered with short stature and facial dysmorphism. Repeat amniocentesis was performed before termination of the pregnancy. Array comparative genomic hybridization was performed on uncultured amniocytes and maternal blood. Conventional cytogenetic analysis was performed on cultured amniocytes, cord blood, and blood from both parents. Fluorescence in situ hybridization was performed on cultured amniocytes. Results: The maternal karyotype was 46,X,inv(X)(p22.3q26.3). The fetal karyotype was 46,Y, rec(X) dup(Xq)inv(X)(p22.3q26.3) or 46,Y, rec(X)(qter/q26.3::p22.3/qter). Array comparative genomic hybridization on uncultured amniocytes revealed a 4.56-Mb deletion of Xp22.33ep22.32 encompassing SHOX, CSF2RA, and ARSE, and a 19.22-Mb duplication of Xq26.3eq28 encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2 in the fetus. The mother did not have X chromosome imbalance. Conclusion: Detection of X chromosome aberration in a male fetus should give suspicion of a recombinant X chromosome derived from maternal X chromosome pericentric inversion. Copyright © 2016, Taiwan Association of Obstetrics & Gynecology. Published by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/ 4.0/).
Keywords: hypotrophic heart short stature Xp22.32/pter deletion Xq26.3/qter duplication Yp11.31/pter deletion
Introduction * Corresponding author. Department of Obstetrics and Gynecology, MacKay Memorial Hospital, 92, Section 2, Chung-Shan North Road, Taipei 10449, Taiwan. E-mail address: [email protected] (C.-P. Chen).
Xp-Xq rearrangements with duplication/deletion events involving Xq terminal duplication and Xp terminal deletion have
http://dx.doi.org/10.1016/j.tjog.2016.05.009 1028-4559/Copyright © 2016, Taiwan Association of Obstetrics & Gynecology. Published by Elsevier Taiwan LLC. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
706
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Figure 1. A karyotype of 46,X,inv(X)(p22.3q26.3) in the mother.
rarely been reported. The derivative or recombinant X chromosome with Xq duplication/Xp deletion can be the result of a meiotic recombination event in a parent carrying a pericentric X inversion or the result of direct X inheritance from a phenotypically normal mother carrying the same aberrant derivative X chromosome [1e9]. In case of one of the parents with a chromosome inversion, there is about 5% recurrence risk for the recombinant chromosome in the offspring [10,11]. Affected male patients with a recombinant X chromosome with terminal Xq duplication/Xp deletion may present: terminal Xq duplication syndrome, such as growth retardation, developmental delay, hypotonia, and genital abnormalities; and terminal Xp deletion syndrome, such as short stature, mental retardation, ichthyosis, Kallmann syndrome, skeletal dysplasia, chondrodysplasia punctata, and hypogonadism [6]. Here, we present molecular cytogenetic characterization of a recombinant X chromosome with terminal Xq duplication/Xp deletion in a fetus with congenital malformations and maternal X chromosome pericentric inversion. Materials and methods Clinical description During the first pregnancy, a 22-year-old, primigravid woman underwent amniocentesis at 17 weeks of gestation because of an abnormal maternal serum screening result showing a trisomy 18 risk of 1/262. Her husband was aged 26 years, and there was no family history of congenital malformations. The woman had a body height of 147 cm, and her husband had a body height of 170 cm. Prenatal ultrasound at 20 weeks of gestation revealed a hypoplastic
left heart and short limbs. The biparietal diameter was 5.19 cm (21.36 weeks), abdominal circumference 15.34 cm (20.54 weeks), femur length 2.69 cm (18.2 weeks), tibia length 2.3 cm (18 weeks), humerus length 1.7 cm (15 weeks), and ulna length 2.4 cm (17 weeks). Amniocentesis revealed a karyotype of 46,Y,der(X) t(X;?)(p22.31;?). The pregnancy was terminated at 24 weeks of gestation, and a malformed male fetus was delivered with a body weight of 412 g, a body length of 23.5 cm, short stature, short limbs, and facial dysmorphism of hypertelorism and low-set ears. Repeat amniocentesis was performed before termination of the pregnancy. Array comparative genomic hybridization (aCGH) was performed on the DNA extracted from uncultured amniocytes and maternal blood. Conventional cytogenetic analysis was performed on cultured amniocytes, cord blood, and parental bloods. Metaphase fluorescence in situ hybridization (FISH) was performed on cultured amniocytes. During the second pregnancy, the same mother underwent amniocentesis at 23 weeks of gestation, which revealed a karyotype of 46,Y,inv(X)(p22.3q26.3). aCGH was performed on the DNA extracted from cultured amniocytes, and metaphase FISH was performed on cultured amniocytes. Preterm birth occurred at 29 weeks of gestation, and a baby was delivered with a body weight of 1489 g, a body length of 45 cm, and a head circumference of 28.5 cm. The baby was normal when followed up at age 4 months with a body weight of 4.6 kg and a body length of 45 cm with no phenotypic abnormalities. aCGH Whole-genome aCGH on the DNA extracted from uncultured amniocytes in the first pregnancy was performed using ISCA Plus Cytogenetic Array (Roche NimbleGen, Madison, WI, USA). This
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
707
Figure 2. A karyotype of 46,Y,rec(X)dup(Xq)inv(X)(p22.3q26.3) or 46,Y,rec(X)(qter/q26.3::p22.3/qter) in the fetus of the first pregnancy.
array has 630,000 probes and a median resolution of 15e20 kb across the entire genome according to the manufacturer's instruction. Whole-genome aCGH on the DNAs extracted from maternal blood and cultured amniocytes in the second pregnancy was performed using CytoChip ISCA Array (Illumina, San Diego, CA, USA), which has 60,000 probes and a median resolution of 51 kb across the entire genome according to the manufacturer's instruction. Conventional cytogenetic analysis Routine cytogenetic analysis by G-banding techniques at the 550 bands of resolution was performed. Amniotic fluid, cord blood and parental blood were collected, and the samples were subjected to cell culture according to the standard blood cytogenetic protocol. FISH Metaphase FISH analysis on cultured amniocytes was performed using the bacterial artificial chromosome probes of RP11-513H18 (Xq27.3, 142,900,963e143,081,146; dye: FITC, spectrum green), RP11-155F12 (Xp22.3, 2,456,240e2,612,008; Yp11.32-p11.31, 2,406, 240e2,562,008; dye: Texas red, spectrum red), and RP11-22P2 (Xq24, 118,252,205e118,409,958; dye: Cy5, spectrum yellow) according to the standard FISH protocol. Results During the first pregnancy, cytogenetic analysis revealed a karyotype of 46,X,inv(X)(p22.3q26.3) (Figure 1) in the maternal blood,
a karyotype of 46,XY in the paternal blood and a karyotype of 46,Y,rec(X)dup(Xq)inv(X)(p22.3q26.3) or 46,Y,rec(X)(qter/q26.3:: p22.3/qter) (Figure 2) in the cultured amniocytes and the cord blood lymphocytes. aCGH on uncultured amniocytes revealed a 4.56-Mb deletion of Xp22.33ep22.32 [arr Xp22.33 (1e2,697,663) 0.5, Xp22.33p22.32 (2,697,663e4,557,932)0.2] encompassing SHOX, CSF2RA and ARSE, and a 19.22-Mb duplication of Xq26.3eq28 [arr Xq26.3q28 (136,052,757e155,270,560)2.0] encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2 in the fetus (Figure 3). Metaphase FISH analysis on the cultured amniocytes revealed two green signals in the terminal ends of the X chromosome and absence of the red signal in the X chromosome, indicating an Xp/Xq rearrangement with terminal Xp deletion/terminal Xq duplication (Figure 4). During the second pregnancy, cytogenetic analysis of the cultured amniocytes revealed a karyotype of 46,Y,inv(X) (p22.3q26.3) (Figure 5). aCGH analysis on cultured amniocytes and maternal blood revealed no genomic imbalance in X chromosome. Metaphase FISH analysis on the cultured amniocytes revealed the order of greenecentromereeyellowered in the X chromosome, indicating a pericentric inversion of X chromosome in the fetus (Figure 6). Discussion The present case had a 19.22-Mb duplication of Xq26.3eq28 encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2. MECP2 (OMIM 300005) encodes methyl-CpG-binding protein that can both activate and repress transcription. MECP2 is a critical dosage-sensitive gene. Deletion or loss-of-function mutation of MECP2 is responsible for a progressive neurological disorder of Rett
708
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Figure 3. Array comparative genomic hybridization on uncultured amniocytes of the first pregnancy shows 4.56-Mb deletion of Xp22.33ep22.32 encompassing SHOX, CSF2RA, and ARSE, and a 19.22-Mb duplication of Xq26.3eq28 encompassing SOX3, FMR1, MECP2, RAB39B, and CLIC2. (A, B, C) Zoom-in view of X chromosome.
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
Figure 4. Metaphase fluorescence in situ hybridization on the cultured amniocytes of the first pregnancy using the bacterial artificial chromosome probes of RP11-513H18 (Xq27.3, green) and RP11-155F12 (Xp22.3, red) shows two green signals in the terminal ends of the X chromosome and absence of the red signal in the X chromosome, indicating a recombinant X chromosome in the fetus with an Xp/Xq rearrangement resulting in terminal Xp deletion/terminal Xq duplication.
709
syndrome (OMIM 312750). Duplication or gain-of-function mutation of MECP2 will cause MECP2 duplication syndrome, which is characterized by recurrent respiratory infections, severe mental retardation, seizures, autism, developmental regression, absent or delayed speech, infantile hypotonia, and progressive spasticity, and is 100% penetrant in affected male patients [12e14]. Fu et al [15] suggested that ventriculomegaly, hydrocephalus, agenesis of the corpus callosum, choroid plexus cysts, intrauterine growth restriction, and hydronephrosis might be the common ultrasound features in the fetuses with MECP2 duplication syndrome. MECP2 duplication syndrome have been reported in rec(X)dup(Xq)inv(X) offspring born to a pericentric inv(X) carrier mother [4,7e9]. Breman et al [8] identified two boys with MECP2 duplication syndrome with terminal Xp deletion, terminal Xq duplication and a recombinant X chromosome containing the duplicated material from Xq28 and Xp, resulting from a maternal pericentric X inversion as similarly presented in this case. SOX3 (OMIM 313430) encodes the SRY-BOX3 protein which is a transcription factor that is important in the regulation of nervous system development and embryogenesis such as gastrulation, neural induction, specification, and differentiation of many types of cells [16,17]. The dysfunction of the SOX3 protein disturbs pituitary development, and over- and underdosage of SOX3 is associated with infundibular hypoplasia and hypopituitarism [18]. Stankiewicz et al [19] reported a duplication of Xq26.2eq27.1 including SOX3 in a mother and daughter with short stature and dyslalia, and suggested a dosage effect of SOX3 in speech disorder in addition to short stature secondary to hypopituitarism. Stagi et al [20] reported a duplication of SOX3 in a boy with growth hormone deficiency, ocular dyspraxia, and intellectual disability, and suggested that
Figure 5. A karyotype of 46,Y,inv(X)(p22.3q26.3) in the fetus of the second pregnancy.
710
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
an int22h1/int22h2-mediated Xq28 duplication syndrome has been identified in males with X-linked intellectual disability, cognitive impairment, behavioral and psychiatric problems, recurrent infections and atopic diseases, obesity, distinctive facial features, and an Xq28 duplication overlapping the int22h1/int22h2 region and including RAB39B and CLIC2 [27e29]. The present case had a 4.56-Mb deletion of Xp22.33ep22.32 encompassing SHOX, CSF2RA, and ARSE. SHOX (OMIM 312865) encodes short stature homeobox protein. Deletions or mutations in SHOX cause LerieWeill dyschondrosteosis (OMIM 127300), Langer mesomelic dysplasia (OMIM 249700), and X-linked idiopathic familial short stature (OMIM 300582). CSF2RA (OMIM 306250) encodes colony-stimulating factor 2 receptor-a (CSF2RA). Mutations in CSF2RA cause pulmonary alveolar proteinosis due to CSF2RA deficiency. ARSE (OMIM 300180) encodes arylsulfatase E. Deletions or mutations in ARSE cause X-linked recessive chondrodysplasia punctata (OMIM 302950). In summary, we have presented molecular cytogenetic characterization of Xp deletion/Xq duplication in a malformed fetus with a recombinant X chromosome due to maternal X chromosome pericentric inversion, and we discuss the genotypeephenotype correlation. We suggest that detection of X chromosome aberration in a male fetus should give suspicion of a recombinant X chromosome derived from maternal X chromosome pericentric inversion. Conflict of interest Figure 6. Metaphase fluorescence in situ hybridization on the cultured amniocytes of the second pregnancy using the bacterial artificial chromosome probes of RP11-22P2 (Xq24, yellow), RP11-513H18 (Xq27.3, green), and RP11-155F12 (Xp22.3, red) shows the order of greenecentromereeyellowered in the X chromosome, indicating a pericentric inversion of X chromosome in the fetus.
SOX3 duplication should be considered in a male patient with short stature due to growth hormone deficiency associated with intellectual disability. Uguen et al [21] reported a duplication of SOX3 in three male fetuses with spina bifida and suggested that SOX3 duplication is a risk factor of neural tube defect. FMR1 (OMIM 309550) encodes fragile X mental retardation protein. A loss of expression of FMR1 results in fragile X mental retardation syndrome and fragile X tremor/ataxia syndrome. However, a duplication of FMR1 has been reported to be associated with a new X-linked mental retardation syndrome characterized by short stature, hypogonadism and facial dysmorphism [22,23]. Rio et al [22] reported familial interstitial Xq27.3eq28 duplication encompassing FMR1 but not MECP2 associated with an X-linked mental retardation syndrome in affected males. Hickey et al [23] reported a duplication of Xq27.3eq28 including FMR1 in association with an X-linked hypogonadism, gynecomastia, intellectual disability, short stature, and obesity syndrome in affected male patients. CLIC2 (OMIM 300138) encodes chloride intracellular channel 2 protein, which plays a role in the regulation of ryanodine receptor (RyR) intracellular Ca2þ release channels. Gain-of-function mutation of CLIC2 enhancing RyR channel activity has been associated with an X-linked intellectual disability, atrial fibrillation, cardiomegaly, congenital heart failure, and seizures [24,25]. RAB39B (OMIM 300774) encodes RAS-associated protein RAB39B which is a small GTPase involved in the regulation of vascular trafficking between membrane compartments. Vanmarsenille et al [26] found copy number gains of RAB39B in Xq28 in 4 male patients with mild intellectual disability and behavioral problems and suggested that increased dosage of RAB39B causes a disturbed neuronal development leading to cognitive impairment. Recently,
The authors have no conflicts of interest relevant to this article. Acknowledgments This work was supported by research grants MOST-103-2314-B195-010 and MOST-104-2314-B-195-009 from the Ministry of Science and Technology and MMH-E-105-04 from MacKay Memorial Hospital, Taipei, Taiwan. References [1] Mohandas T, Geller RL, Yen PH, Rosendorff J, Bernstein R, Yoshida A, et al. Cytogenetic and molecular studies on a recombinant human X chromosome: implications for the spreading of X chromosome inactivation. Proc Natl Acad Sci USA 1987;84:4954e8. [2] Schmidt M, Du Sart D, Kalitsis P, Leversha M, Dale S, Sheffield L, et al. Duplications of the X chromosome in males: evidence that most parts of the X chromosome can be active in two copies. Hum Genet 1991;86:519e21. [3] Vasquez AI, Rivera H, Bobadilla L, Crolla JA. A familial Xpþ chromosome, dup (Xq26.3/qter). J Med Genet 1995;32:891e3. [4] Madariaga ML, Rivera H. Familial inv(X)(p22q22): ovarian dysgenesis in two sisters with del Xq and fertility in one male carrier. Clin Genet 1997;52: 180e3. [5] Lammer EJ, Punglia DR, Fuchs AE, Rowe AG, Cotter PD. Inherited duplication of Xq27.2/qter: phenocopy of infantile PradereWilli syndrome. Clin Dysmorphol 2001;10:141e4. [6] Kokalj-Vokac N, Marcun-Varda N, Zagorac A, Erjavec-Skerget A, Zagradisnik B, Todorovic M, et al. Subterminal deletion/duplication event in an affected male due to maternal X chromosome pericentric inversion. Eur J Pediatr 2004;163: 658e63. [7] Bleyl SB, Byrne JLB, South ST, Dries DC, Stevenson DA, Rope AF, et al. Brachymesomelic dysplasia with Peters anomaly of the eye results from disruptions of the X chromosome near the SHOX and SOX3 genes. Am J Med Genet 2007;143A:2785e95. [8] Breman AM, Ramocki MB, Kang SHL, Williams M, Freedenberg D, Patel A, et al. MECP2 duplications in six patients with complex sex chromosome rearrangements. Eur J Hum Genet 2011;19:409e15. [9] Sanmann JN, Bishay DL, Starr LJ, Bell CA, Pickering DL, Stevens JM, et al. Characterization of six novel patients with MECP2 duplications due to unbalanced rearrangements of the X chromosome. Am J Med Genet 2012;158A: 1285e91. ne ticiens Français. Pericentric inversions in man. A French [10] Groupe de Cytoge ne t 1986;29:129e68. collaborative study. Ann Ge [11] Goodman BK, Shaffer LG, Rutberg J, Leppert M, Harum K, Gagos S, et al. Inherited duplication Xq27-qter at Xp22.3 in severely affected males:
C.-P. Chen et al. / Taiwanese Journal of Obstetrics & Gynecology 55 (2016) 705e711
[12]
[13] [14] [15]
[16] [17]
[18]
[19]
[20]
[21]
molecular cytogenetic evaluation and clinical description in three unrelated families. Am J Med Genet 1998;80:377e84. Sanlaville D, Prieur M, de Blois MC, Genevieve D, Lapierre JM, Ozilou C, et al. Functional disomy of the Xq28 chromosome region. Eur J Hum Genet 2005;13:579e85. Sanlaville D, Schluth-Bolard C, Turleau C. Distal Xq duplication and functional Xq disomy. Orphanet J Rare Dis 2009;4:4. Ramocki MB, Tavyev YJ, Peters SU. The MECP2 duplication syndrome. Am J Med Genet 2010;152A:1079e88. Fu F, Liu HL, Li R, Han J, Yang X, Min P, et al. Prenatal diagnosis of foetuses with congenital abnormalities and duplication of the MECP2 region. Gene 2014;546:222e5. Pevny LH, Lovell-Badge R. Sox genes find their feet. Curr Opin Genet Dev 1997;7:338e44. ndez A, Han Y, Pallavi B. Control of cell fate Lefebvre V, Dumitriu B, Penzo-Me and differentiation by Sry-related high-mobility-group box (Sox) transcription factors. Int J Biochem Cell Biol 2007;39:2195e214. Woods KS, Cundall M, Turton J, Rizotti K, Mehta A, Palmer R, et al. Over- and underdosage of SOX3 is associated with infundibular hypoplasia and hypopituitarism. Am J Hum Genet 2005;76:833e49. Stankiewicz P, Thiele H, Schlicker M, Cseke-Friedrich A, Bartel-Friedrich S, Yatsenko SA, et al. Duplication of Xq26.2-q27.1, including SOX3, in a mother and daughter with short stature and dyslalia. Am J Med Genet 2005;138A: 11e7. Stagi S, Lapi E, Pantaleo M, Traficante G, Giglio S, Seminara S, et al. A SOX3 (Xq26.3e27.3) duplication in a boy with growth hormone deficiency, ocular dyspraxia, and intellectual disability: a long-term follow-up and literature review. Hormones (Athens) 2014;13:552e60. mener-Redon S, Marcorelles P, De Braekeleer M. Uguen A, Talagas M, Que Duplication of SOX3 (Xq27) may be a risk factor for neural tube defects. Am J Med Genet 2015;167A:1676e8.
711
[22] Rio M, Malan V, Boissel S, Toutain A, Royer G, Gobin S, et al. Familial interstitial Xq27.3q28 duplication encompassing the FMR1 gene but not the MECP2 gene causes a new syndromic mental retardation condition. Eur J Hum Genet 2011;18:285e90. [23] Hickey SE, Walters-Sen L, Mosher TM, Pfau RB, Pyatt R, Snyder PJ, et al. Duplication of the Xq27.3eq28 region, including the FMR1 gene, in an Xlinked hypogonadism, gynecomastia, intellectual disability, short stature, and obesity syndrome. Am J Med Genet 2013;161A:2294e9. [24] Witham S, Takano K, Schwartz C, Alexov E. A missense mutation in CLIC2 associated with intellectual disability is predicted by in silico modeling to affect protein stability and dynamics. Proteins 2011;79:2444e54. [25] Takano K, Liu D, Tarpey P, Gallant E, Lam A, Witham S, et al. An X-linked channelopathy with cardiomegaly due to a CLIC2 mutation enhancing ryanodine receptor channel activity. Hum Mol Genet 2012;21:4497e507. [26] Vanmarsenille L, Giannandrea M, Fieremans N, Verbeeck J, Belet S, Raynaud M, et al. Increased dosage of RAB39B affects neuronal development and could explain the cognitive impairment in male patients with distal Xq28 copy number gains. Hum Mutat 2014;35:377e83. [27] Andersen EF, Baldwin EE, Ellingwood S, Smith R, Lamb AN. Xq28 duplication overlapping the int22h-1/int22h-2 region and including RAB39B and CLIC2 in a family with intellectual and developmental disability. Am J Med Genet 2014;164A:1795e801. [28] El-Hattab AW, Schaaf CP, Fang P, Roeder E, Kimonis VE, Church JA, et al. Clinical characterization of int22h1/int22h2-mediated Xq28 duplication/deletion: new cases and literature review. BMC Med Genet 2015;16:12. [29] El-Hattab AW, Schaaf CP, Cheung SW. Xq28 duplication syndrome, Int22h1/ Int22h2 mediated. In: Pagon RA, Adam MP, Ardinger HH, Wallace SE, Amemiya A, Bean LJH, et al, editors. GeneReviews. Seattle (WA): University of Washington, Seattle; 1993e2016. http://www.ncbi.nlm.nih.gov/books/ NBK349624/. [accessed 10.03.16].