J Med Genet 1996;33:237-239

Cytogenetic and clinical characteristics of a caseinvolving complete duplication of Xpter-+Xql3

Syed M Jalal, Richard Dahl, Lisa Erickson, Donald Zimmerman, Noralane Lindor

CytogeneticsLaboratory,Mayo Clinic andMayo Foundation,200 First Street SW,Rochester, MN 55905,USAS M JalalR Dahl

Department ofReproductiveEndocrinology,Mayo Clinic andMayo Foundation,Rochester, MN, USAL Erickson

Department ofPediatrics, MayoClinic and MayoFoundation,Rochester, MN, USAD Zimmerman

Department ofMedical Genetics,Mayo Clinic andMayo Foundation,Rochester, MN, USAN Lindor

Correspondence to:Dr Jalal.Received 5 July 1995Revised version accepted forpublication 23 October 1995

AbstractTrue isochromosomes for Xp probably donot exist in a liveborn. We describe a rarecase of complete Xp duplication and re-tention of the inactivation centre at Xql3.Cytogenetically, it is described as a non-mosaic 46,X,psu idic(X)(ql3). Completeduplication ofXpter-Xql3 was confirmedby banded analysis and FISH probes for Xcentromere, Xp2l, XIST locus, and wholechromosome paints for X and Y. The ab-normal X was always late replicating.

Clinically, the patient was short stat-ured, had primary amenorrhoea, and in-complete development ofsecondary sexualcharacteristics, but otherwise was pheno-typically normal. There are no non-mosaicreported cases with complete duplicationof i(Xp) confirmed by FISH or moleculartechniques. Those cases with partial du-plication of Xp and presence of the in-activation centre share the traits ofamenorrhoea and poor secondary sexualdevelopment. To develop a clinical profileof duplication ofXp (in presence of Xql3)there is a need to study more cases.(J Med Genet 1996;33:237-239)

Key words: i(Xp) duplication characteristics.

Isochromosome Xq occurs with a frequency of1 in 50000 and accounts for 15 to 20% ofTurer syndrome patients in pure or mosaicform.' Isochromosome Xp theoretically shouldoccur in a comparable proportion, but thisis not the case. This may be in part because of

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the presence of dicentric i(Xq) in relativelyhigh proportion, shown either cytogeneticallyor molecularly, which would result in i(Xp)that is acentric and thus non-viable. There area handful of non-mosaic cases reported to bei(Xp).26 In these cases, it is difficult to dis-tinguish by only chromosome banding betweeni(Xp) and deletion of Xq resulting in a nearmetacentric chromosome.7 Therman et al pro-pose that without an intact inactivation centrein the q arm, the entire p arm will remain activeand that the disruption of genetic imbalancefrom such duplication will be severe. A 1993paper8 describes a case oftall stature and amen-orrhoea involving a rearranged X(pter-+ql3::p 11.4 -+pter) where molecular and FISH tech-niques confirmed the near duplication of Xp.We present a rare case of complete X p armduplication and proximal Xq duplication thatincluded the XIST locus, which is known tobe at Xql3.9 The clinical features of the caseand cytogenetic implications are discussed.

Case reportThe patient was a 21 year old Chilean collegestudent referred for evaluation of primaryamenorrhoea and short stature. She was ahealthy infant of a normal pregnancy born atterm to a 28 year old mother and 29 year oldfather. Although short stature was evident inearly childhood, health and development wereotherwise unremarkable.

Physical examination at the age of 21 yearsshowed a height of 143-5 cm (<5th centile or

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Figure 1 Photographs of the woman with 46,X,psu idic(X) (q13).

Figure 2 NormalX and the psu idic(X) (q13)chromosome by GTL (A), QFQ (B), and CBG (C)banding. The arrows indicate the approximate location ofband Xql3.

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28alal, Dahl, Erickson, Zimmerman, Lindor

50th centile for age 11-3 years), weight was51 kg, and head circumference was 52 cm. Thecranium was normocephalic and the facies con-sistent with ethnicity. The ear helices showeda prominent antihelix and normal position. Theposterior hairline was not low and pterygiumcoli was not evident. The thyroid was mod-erately enlarged, symmetrical, and withoutnodules. The bony thorax was normal and noscoliosis was evident. Examination of the heart,lungs, and abdomen showed no abnormalities.Breast and external genitalia were consistentwith Tanner III development. Internal pelvic

Figure 3 An alpha satellite probe specific for the X centromere (green) used as a controland a cosmid probe specific for the XIST locus (orange) had XIST signal sandwiched bytwo centromere signals in the abnormal X and single signals for the normal X

Figure 4 A cosmid probe for Xp2l had one signal on a normal X and two on theabnornalX chromosome.

examination was not permitted by the patient.Morphology of the hands, palmar flexioncreases, and dermal ridges were normal exceptfor radial loops on both index fingers. Thelower extremities showed mild genu valguswith patellae appearing laterally displaced andmild leg length and calf circumference dis-crepancy (fig 1). The neurological examinationwas unremarkable. A large hypopigmented ma-cule of irregular shape was present on theforehead (it developed at the age of 14 years).The number of pigmented naevi was average.

Radiographs of the hips and knees showedmild osteopenia and open growth plates. Boneage was between 13 5 and 14 year standards.Ultrasonography of the pelvis showed a smalluterus (2 x 4 cm, prepubertal size), and theovaries could not be identified. No pelvicmasses were seen.

Endocrine evaluation showed a normal thyr-oxine level but raised sensitive TSH and thyroidmicrosomal antibody titre of 1:6400, luteinisinghormone 15 * 5 IU/l, and follicle stimulating hor-mone 44 IU/l, oestradiol 20 pg/ml (normal pre-menopausal levels are 30-400). Chemistrytesting was significant for alkaline phosphataseof 219 IU/l (normal 33-118), and the iso-enzyme analysis showed that the rise was at-tributable to bone enzyme.

CYTOGENETIC ANALYSISThirty metaphases from PHA stimulated bloodculture were analysed by GTL, QFQ, and CBGbanding techniques. Each metaphase had apseudoisodicentric X with the break and fusionpoint at Xql3 (fig 2). The karyotype was de-scribed as 46,X,psu idic(X)(ql3). Thirty con-secutive analysable metaphases were scored forthe replication pattern (10 jig/ml BrdU, sixhour terminal pulse). The psu idic(X)(ql3)was late replicating in each metaphase.The psu idic(X)(ql3) was analysed by fluor-

escent in situ hybridisation (FISH) using a varietyof DNA probes. Modifications of the man-ufacturing company's protocol were similar tothose described earlier.'0 1 Use of alpha satelliteprobe specific for the X centromere confirmedthat both the active and inactive centromereswere of X chromosome origin. Chromosomespecific paint probes forX and Y (that cross hy-bridise to the Xpter pseudoautosomal region)confirmed that both arms wereXp orXp derived.The cosmid probe for the XIST locus (Vysis,Downers Grove, IL) was positive (fig 3) betweenthe two centrometres, indicating the presence ofthe XIST gene. Oncor (Gaithersburg, MD) pro-vided us with a probe specific for Xp2l. Use ofthis probe resulted in one signal (fig 4) on eacharm of the abnormal X, consistent with the ban-ded cytogenetic interpretation of complete Xpduplication.

DiscussionAlthough i(Xq) reportedly occurs with a fre-quency of 1 in 50000 and is present in 15 to20% of Turner syndrome patients,' theexistence of i(Xp) without the proximal q armhas been questioned.7 Therman et al7 proposethat without the presence of the inactivation

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Cytogenetic and clinical characteristics of a case involving complete duplication of Xpter-Xql3

centre, which is now known to be at Xq 13,9the p arm duplication causes so much of agenetic imbalance that i(Xp) do not surviveeven as low level mosaics. Over the years theirprediction seems to have held. It has beenwidely recognised in the last four to five yearsthat a relatively high proportion of i(Xq) casesare demonstrably dicentric, especially by mo-lecular techniques. Thus, if a high proportionof i(Xp) are from U type isochromatid fusionand acentric, they will be non-viable. This mayin part explain the paucity of i(Xp).There are some reported cases ofnon-mosaic

i(Xp),356 mosaic i(Xp) with 45,X,412 and acase of 46,X?i(Xp).'2 Whether the small nearmetacentric chromosome in these cases is i(Xp)or simply del(Xq), where the size of the q armis about the same as the Xp, cannot be resolvedby standard banding techniques. Therman etal7 propose that these cases may well be Xqdeletions. A range of anomalies from primaryamenorrhoea, some features of Turner syn-drome, endocrine hormone anomalies, togonadal dysgenesis and essentially no otherdetectable dysmorphic features have been re-ported for these patients. Interestingly, in anextensive review, Therman and Susman" haveprovided a summary of clinical features for 67cases involving a range of deletions of the qarm and 52 cases involving deletions of the parm. In their words, "one is struck by thesimilarity of the anomalies caused by the de-letions of the two arms" as it relates to thedegree of amenorrhoea, height, and other stig-mata of Turner syndrome. Thus, the clinicalprofile may not permit a distinction between qarm deletion and those described for p armduplication in the earlier publications.A recent paper by Leppig et alt is directly

comparable to our case. Cytogenetically, theirpatient is described as having rec(X)dup p,inv (X)(pter-+q3::11.4-÷pter) from a chro-mosomally normal mother. The short arm andXIST region of q arm duplication was con-firmed by molecular and FISH techniques. Theclinical descriptions provided for their patientare primary amenorrhoea, immature secondarysexual characteristics, tall stature with an eu-nuchoid habitus (90th centile), normal psy-chomotor development and intellect, and nofeatures of Turner syndrome.A case reported by Pettigrew et all4 of a 28

year old female with primary amenorrhoeaand features of Turner syndrome had a verysimilar structural anomaly: 46,X,idic(X)(pter-+q13.2::q13.2-+pter). However, she had 6%45,X mosaicism which may explain the Turnersyndrome features.An earlier case of a 171 year old girl was re-

ported by Daniel et all5 as a non-mosaici(X) (pter-÷q2 102 -pter) and had gonadaldysgenesis, short stature (157 cm), poorly de-veloped secondary sexual development, butotherwise without most stigmata associatedwith Turner syndrome. The clinical features ofthis case are remarkably identical to our patient.Our patient had some Turner-like features

including short stature, hypogonadism with

primary amenorrhoea, and Hashimoto'sthyroiditis, but was otherwise phenotypicallyunremarkable. We cannot rule out the pos-sibility of a 45,X cell line that may be tissuerestricted, although there was no evidence of45,X mosaicism in the lymphocyte culture.Other tissue sources were not available foranalysis. The difference between Leppig et al8and our patient may be in part because of thelack of complete duplication of Xp in theircase. Though the break and fusion point in thecase of Daniel et a114 is at Xq2102 and theduplication of the Xp arm has not been con-firmed by FISH or molecular techniques, thephenotype shows remarkable similarity to ourpatient. Duplication ofthe short arm with breakand fusion at Xql3 or Xq2102 may well resultin a similar phenotype as long as the XISTlocus is present.

Cytogenetically, the present case with a karyo-type of non-mosaic 46,X,psu idic(X)(ql3) isindeed rare with a complete duplication of Xpthat has been confirmed by a variety of DNAfluorescent probes. The late replication studyin our case confirms that the abnormal X isselectively inactivated. The presence of the in-activation centre (Xq13) in our case and thatofLeppig et al8 bears out the Therman prophecythat complete expression of genes in the X parm in duplicate may indeed be lethal. Thereis a need to study more cases involving Xpduplication in the presence of the XIST gene,confirmed by molecular cytogenetics, to de-velop the associated clinical phenotypic profile.

1 Hook EB, Warburton D. The distribution of chromosomalgenotype associated with Turner's syndrome: livebirth pre-valence rates and evidence ofdiminished fetal mortality andseverity in genotypes associated with structural X ab-normalities or mosaicism. Hum Genet 1983;64:24-7.

2 Caspersson C, Lindsten J, Zech L. The nature of structuralX chromosome aberrations in Turner's syndrome as re-vealed by quinacrine mustard analysis. Hereditas 1970;66:287-92.

3 de la Chapelle A, Schroder J. Isochromosome for the shortarm of X, a human 46,XXpi syndrome. Ann Hum Genet1972;36:79-87.

4 Fitzgerald PH, Donald RA. Isochromosome for the short armof X: a human mosaic 45,X/46,XXpi, Clin Genet 1975;7:148-54.

5 Keogh EJ, de Krester DM, Fitzgerald MG. Isochromosomefor short arm ofX with primary amenorrhea and pituitarytumor. AustNZ3tMed 1973;3:617-9.

6 Van den Berghe H, Fryns JP, Devos F. 46,XXip karyotypein a woman with normnal stature and gonadal dysgenesiswithout other congenital anomalies. Humangenetik 1973;20:163-6.

7 Therman E, Sarto GE, Patau K. Center for Barr body con-densation on the proximal part of Xq: a hypothesis.Chromosoma (Berl) 1974;44:361-6.

8 Leppig KA, Brown CJ, Bressler SL, et al. Mapping the distalboundary of the X-inactivation center in a rearranged Xchromosome from a female XIST. Hum Mol Genet 1993;7:883-7.

9 Brown CJ, Ballabio A, RupertJL, et al. A gene from the regionofthe human X inactivation centre is expressed exclusivelyfrom the inactive X chromosome. Nature 199 1;349:38-44.

10 Jalal SM, Law ME, Christensen ER, Spurbeck JL, DewaldGW. Method for sequential staining ofGTL-banded meta-phases with fluorescent-labeled chromosome specific paintprobes. AmJ7Med Genet 1993;46:98-103.

11 Schad CR, Kraker WA, Jalal SM, et al. Use of fluorescent insitu hybridization for marker chromosomes in congenitaland neoplastic disorders. AmJ7Clin Pathol 1991;96:203-1 0.

12 Abulhasan SJ, Teebi AS, Zaki M, Hammad I, Ai-Awadi SA,Krishna Murty DS. Mosaicism 45,X/46,X tdic(Xp:Xp) ina girl with short stature. Am Genet (Paris) 1990;33:234-8.

13 Therman E, Susman B. The similarity of phenotypic effectscaused by Xp and Xq deletions in the human female: ahypothesis. Hum Genet 1990;85:175-83.

14 Pettigrew AL, McCabe ERB, Elder FFB, LedbetterDH. Iso-dicentric X chromosome in a patient with Turner syn-drome-implications for localization of the X-inactivationcenter. Hum Genet 199 1;87:498-502.

15 Daniel A, Saville T, Southall DB. Further dicentric X iso-chromosomes and deletions, and a new structurei(X)(pter--q2102 --pter). JMed Genet 1979;16:278-84.

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