SUBMITTED 25 NOV 21 1

REVISION REQ. 26 JAN 22; REVISION RECD. 6 MAR 22 2

ACCEPTED 15 MAR 22 3

ONLINE-FIRST: APRIL 2022 4

DOI: https://doi.org/10.18295/squmj.4.2022.033 5

First Report of a Derivative Chromosome 13 with a Duplicated 11p15 Locus 7

Associated with Silver-Russell syndrome 8

*Nishath Hamza, Musallam Al-Araimi, Kamla Al Salmani, Salwa Al 9

Obeidani 10

National Genetic Centre, Royal Hospital, Ministry of Health, Muscat, Oman 12

*Corresponding Author’s e-mail: nishath.h@moh.gov.om 13

Abstract 15

Silver-Russell Syndrome (SRS) is a disorder that is primarily characterized by intrauterine 16

growth restriction which may occur asymmetrically or in whole, leading to a fetus being small 17

relative to its gestational age. We present a female infant (proband), with severe congenital 18

anomalies. The proband carried a >25Mb duplication of the chromosomal 11p15-11pter locus of 19

chromosome 13; creating a derivative chromosome 13 [der(13)] and was reported as 20

46,XX,der(13)add(11p15-11pter). A methylation-sensitive assay confirmed a diagnosis of 21

Silver-Russell Syndrome (SRS). Although the prognosis for SRS patients is generally good, our 22

proband presented with a clinically severe phenotype culminating in death at nine months. To the 23

best of our knowledge, this is the first report of a derivative chromosome 13 with a duplicated 24

11p15 locus being reported in a patient with SRS. 25

Keywords: Silver-Russell syndrome, growth retardation, imprinting, derivative chromosome. 26

mailto:nishath.h@moh.gov.om

Introduction 28

Silver-Russell Syndrome (SRS) is a disorder that is primarily characterized by intrauterine 29

growth restriction which may occur asymmetrically or in whole, leading to a fetus being small 30

relative to its gestational age. Individuals are diagnosed with SRS when they present with growth 31

restriction, relative macrocephaly at birth (head circumference ≥1.5 SD above birth weight 32

and/or length), prominent forehead usually with frontal bossing, triangular facies,micrognathia 33

and feeding difficulties.1,2 Rarely, SRS patients may also exhibit fifth-finger clinodactyly.1 34

SRS is one of twelve imprinting disorders which are caused by epigenetic (methylation) or 36

genetic abnormalities. Among SRS patients, 35%-50% of cases are due to loss of paternal allele 37

methylation (LOM) at the imprinted control region 1 (ICR1) at 11p15.5 and 7%-10% are due to 38

maternal uniparental disomy (UPD) of chromosome 7.3,4 In rare cases, somatic mosaicism for 39

maternal UPD(11), duplication of the maternal 11p15.5, inversions and translocations affecting 40

chromosome 11, as well as maternal UPD of chromosomes 14, 16 and 20 have also been 41

reported.5 In addition to epigenetic and copy number variants (CNVs), mutations in certain genes 42

have also been reported to cause SRS. For example, maternal transmission of gain-of-function 43

mutations in the CDKN1C gene3 or paternal transmission of loss-of-function mutations in the 44

IGF2 gene4 has been described in association with SRS. Also, genes which are upstream 45

regulators of IGF2 such as HMGA2 or PLAG1 are also associated with SRS.4 46

Case Report 48

Our proband was a female infant born in 2018 at a tertiary hospital in Muscat, Oman. She 49

presented with multiple anomalies such as intrauterine growth restriction, macrocephaly, broad 50

fontanelle, feeding difficulty, low set ears and failure to thrive without ventilator support (Figure 51

1A). The proband was born to unrelated parents (family pedigree shown in Figure 1B). A history 52

of miscarriage at six weeks of pregnancy in the proband’s mother prompted the Special Care 53

Baby Unit (SCBU) physician at the referral hospital to request cytogenetic studies in the proband 54

and her parents. Informed consent was obtained from the proband’s parents prior to the referral 55

for genetic studies. Peripheral blood samples were obtained from the patient and her parents in 56

both Heparin and EDTA tubes. The proband was referred to our genetic clinic a few days after 57

her birth. However, the infant was in a critical condition and remained in the SCBU on ventilator 58

until her death at the age of nine months. As a result, a direct clinical assessment of the proband 59

could not be carried out by our genetic clinic. Instead, a description of patient phenotype was 60

communicated to our clinical geneticist by the referring physician over phone. 61

Chromosomal karyotyping of the proband yielded an abnormal karyotype with a large 63

heterozygous additional chromosomal region on the p-arm of chromosome 13. Genomic DNA 64

was extracted from the whole blood of the proband and used to perform array-based comparative 65

genomic hybridization (CGH) with the Affymetrix Cytoscan HD kit (Thermo Fisher Scientific, 66

USA). Array CGH data analysis using the ChAS software v.3.1.0.15 revealed a heterozygous 67

25,109kbp (~25 Mb) duplication of the 11p15-11pter chromosomal locus (hg19:chr11:230,615-68

2,339,766). Hence the karyotype (Figure 2A) was reported as 46,XX,der(13)add(11p15-11pter). 69

To the best of our knowledge, this is the first report of a derivative chromosome 13 with a 70

duplicated 11p15 locus being reported in a patient with SRS. 71

Given the clinical feature of growth restriction in the proband, the involvement of the 11p locus 73

warranted further testing due to its association with SRS. A methylation-sensitive Multiplex 74

ligation-dependent Probe Amplification (MS-MLPA) assay6 was conducted on the DNA from 75

the proband using the ME030 (Lot No:C3-0219) kit from MRC Holland (The Netherlands). This 76

kit is a multi-disease assay which tests the 11p15 locus for both BWS and RSS, as well as the 77

5q35.3 locus (NSD1 gene) for Sotos syndrome. The assay samples were then run on the genome 78

analyzer ABI 3700 and the data generated was analyzed using the Coffalyser (v.210604.1451). 79

The MS-MLPA data (Figure 2B) confirmed the duplication of the 11p15 locus, but also revealed 80

LOM at the ICR1. 81

Meanwhile, karyotyping both parents of the proband revealed that the der(13) chromosome 83

observed in the patient was maternally inherited (Figure 3). Hence, the diagnosis of SRS due to 84

maternal 11p duplication was established in the proband. 85

The 25 years old mother (Figure 1, II.3) of the proband was found to be a carrier of a 87

heterozygous balanced non-reciprocal translocation between chromosome 11 and 13: 88

46,XX,der(13)t(11;13)(p11;p12). This phenotypically normal, but genotypically abnormal 89

karyotype (Figure 3) was characterized by one of the chromosomes 13 having an additional 90

translocated 11p15-11pter region on its p-arm creating the der(13) chromosome, and one of the 91

chromosomes 11 lacking the region from 11p15-11pter. The father of the proband was observed 92

to have a normal male karyotype. 93

The parents of the proband had not been amenable to an appointment at our clinic while their 95

child was in the SCBU. After the death of the proband, the parents met with our genetic 96

counselor and the implications of the karyotype and MS-MLPA results were explained to them. 97

During genetic counseling of the proband’s parents, it transpired that there was a family history 98

of miscarriages reported in the 52 years old maternal grandmother (Figure 1B, I.1) and a 30 years 99

old maternal aunt of the proband (Figure 1B, II.1). Fertility problems were also reported in a 100

maternal 28 years old uncle (Figure 1B, II.2) of the patient who had a single offspring after 101

treatment for infertility. The maternal grandmother of our proband, I.1 was unable to recall the 102

number of miscarriages she underwent. These individuals were then invited for genetic 103

counseling and offered karyotyping after informed consent. All three tested family members 104

carried karyotypes identical to the proband’s mother (balanced non-reciprocal translocation; 105

Figure 3). Another 19 years old maternal uncle of the proband was reported to be diagnosed with 106

unilateral kidney disease (Figure 1B, II.). However, this individual was not willing to undergo 107

genetic counseling or testing. 108

109

Informed consent for testing and publication of anonymized data was collected from all 110

patients/guardians involved in this study and appropriate ethical standards were employed in all 111

procedures. 112

113

Discussion 114

This is the first report of a case where SRS is associated with a derivative chromosome 13 115

carrying a duplicated 11p arm. In light of the fact that translocation events involving 116

chromosome 11p and chromosome 13 have never been reported before except in oncology 117

patients, this finding is quite novel. The der(13) chromosome in the proband, resulted in an extra 118

copy of the maternal 11p12 to 11pter region within the karyotype, with no apparent loss of 119

chromosome 13 regions according to array CGH analysis The der(13) chromosome was 120

transmitted through at least three generations of a family. 121

122

Although rare, maternal duplications of 11p12-11pter which include the 11p15 locus, are 123

estimated to cause the associated SRS phenotype in <1% of SRS patients.7 The cases of maternal 124

11p15 duplications reported previously were mostly interstitial duplication events with or 125

without inversions, encompassing the 11p15 locus8 or rarely, due to unbalanced translocations 126

between chromosome 11 and chromosomes 4, 9, 10, 15, 16 and 17.9-13 While most of these 127

rearrangements involved ICR1; duplications of the whole ICR2 as well as partial duplication of 128

ICR1 were also rarely reported in association with SRS. However, in all of these cases, the 129

patients survived much longer than our patient, albeit with varying degrees of prognosis7-15. 130

131

SRS patients generally have a good prognosis and can live well into adulthood with occasional 132

complications.14 However, the severity of clinical presentation in SRS patients with copy number 133

variants (CNV) appears to be dependent on the extent of 11p locus involved in the CNV.7,14 This 134

is evident in our patient who was unable to survive independently outside of the SCBU facility 135

because the ~25Mb duplicated maternal allele in our proband covered almost the entire 11p15.5 136

band, which included both the ICR1 and ICR2 regions and was bigger than the majority of the 137

previously reported CNVs involving the 11p15 locus.7-15 This was accompanied by 138

hypomethylation of the H19 gene. Hence, the classic SRS phenotype of growth restriction in the 139

proband likely reflects an increased expression of the maternally expressed H19 gene and 140

consequent down-regulation of the IGF2 gene expression.9-11 141

142

In the case of maternal inheritance, duplication of the 11p15 locus causes the SRS phenotype, 143

whereas a paternally inherited similar duplication would cause the Beckwith Wiedemann 144

syndrome (BWS) phenotype. No instances of BWS were seen within our proband’s family, 145

especially since most of the carriers of the der(13) chromosome detected in this family were 146

females. The maximum likelihood of paternal transmission of the der(13) chromosome and risk 147

for BWS is from the maternal uncle (Figure 1, II.2) of the proband, who has one normal 148

offspring. 149

150

A key point to be noted in this case is that patients suspected with SRS are usually subjected to 151

molecular genetic tests which can characterize either methylation abnormalities or copy number 152

variants (CNVs) or both; but not chromosomal translocations. However, the clinically severe 153

presentation in our proband and the history of miscarriage in the proband’s mother had prompted 154

a referral for cytogenetic studies. This was key to the der(13) translocation-derivative 155

chromosome being detected in multiple members of the family and the provision of accurate 156

genetic counseling to other members of the family who had a history of miscarriages and 157

infertility. The affected couples in the proband’s extended family had not suspected a hereditary 158

component to their history of reproductive failures prior to our proband being tested. 159

160

The parents of the proband were counseled regarding future risk for affected offspring. However, 161

the mother refused to consider prenatal genetic testing combined with in-vitro fertilization as a 162

reproductive option, since abortion is generally prohibited in Oman (with medical exceptions). 163

The mother decided to have future pregnancies monitored using first trimester ultrasonographic 164

diagnosis. 165

166

Conclusions 167

Although maternal duplications due to 11p15 translocation events are rare, they must be 168

suspected in patients with SRS phenotype who also present with severe failure to thrive. Offering 169

genetic testing to the parents of affected patients may help prevent further recurrences of affected 170

offspring. Determining whether a duplication event is due to the transmission of translocated 171

chromosomes, or due to interstitial duplications or inversions, is also crucial as individuals who 172

carry translocations are at significantly higher risk for infertility, recurrent miscarriages and birth 173

of offspring with moderate to severe disease phenotypes. 174

175

Data Availability Statement 176

Data generated in this study is the sole property of the Royal Hospital, Ministry of Health, 177

Oman. As such, any release of data from this study, outside of journal publications or scientific 178

abstracts, is subject to prior approval of the Scientific Research Committee, Royal Hospital, 179

Oman. 180

181

Authors’ Contribution 182

NH carried out molecular genetics analyses and wrote the manuscript; MA conducted clinical 183

sampling; patient counseling and manuscript review; KS carried out cytogenetic analyses and 184

manuscript review and SO conducted patient counseling and manuscript review. All authors 185

approved the final version of the manuscript. 186

187

Acknowledgements 188

The authors acknowledge the work of the cytogenetic and molecular genetics staff at the 189

National Genetic Center who carried out routine diagnostic processing of the patient samples 190

studied in this report. 191

192

References 193

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Figure 1: Clinical details of the proband 251

(A)The female infant was born with macrocephaly, broad fontanelle, low set ears, intrauterine growth restriction and presented with 252

failure to thrive without ventilator support. (B) Family pedigree of the proband 253

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Figure 2: Karyotype and MS-MLPA test results of the proband 273

(A) The derivative chromosome 13 with a duplicated 11p locus is indicated by a short black arrow in this karyotype of the proband 274

(B) MS-MLPA6 results: The upper panel shows the CNV analysis (C1) and methylation analysis (C1-D) in a normal control sample. 275

The bottom panel shows the CNV analysis (III.1) and methylation analysis (III.1-D) results for the proband. Black probe signals 276

indicate normalized results in comparison to control samples within the MS-MLPA run. In the CNV panel III.1, the blue signals 277

indicate the duplicated signals (3 copies) from all the probes targeting the 11p15 locus, at an average ratio of 1.5 on the y-axis; 278

whereas the red signal in the panel III.1-D indicates the decrease in methylation of the H19 locus. The orange regions include the 279

probe signals from the 11p15 locus, the grey regions indicate signals from reference probes and the violet regions represent probe 280

signals from the NSD1 gene at the 5q35.3 locus. 281

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Figure 3: Balanced, non-reciprocal translocation observed in the proband’s mother 284

The deletion at the 11p arm one of the chromosomes 11, and the addition of a 11p15-11pter region on the 13p arm of a chromosome 285

13 which created a der(13), are both indicated by black arrows. The abnormal chromosomes are compared against representative 286

ideograms. 287