Prenatal diagnosis of Pallister‐Killian syndrome and literature review

Abstract Pallister‐Killian syndrome (PKS) is a rare sporadic genetic disorder usually caused by mosaicism of an extra isochromosome of 12p (i(12p)). This retrospective study analysed the prenatal ultrasound manifestations and molecular and cytogenetic results of five PKS foetuses. Samples of amniotic fluid and/or cord blood, skin biopsy and placenta were collected. Conventional karyotyping and single nucleotide polymorphism array (SNP array) were performed on all the amniotic fluid or cord blood samples. Copy number variants sequencing (CNV‐seq) and fluorescence in situ hybridization (FISH) were also used for the validation for one foetus. All the five foetuses were from pregnancies with advanced parental age. Two foetuses involved structural abnormalities and one foetus had only soft markers, all of which included increased nuchal translucency. The rest two foetuses had normal ultrasounds in the second trimester, which has rarely been reported before. The karyotype revealed typical i(12p) in four cases and a small supernumerary marker chromosome consisting of 12p and 20p in the remaining one case. The proportion of cells with i(12p) ranged from 0 to 100% in cultural cells, while SNP array results suggested 2−4 copies of 12p. For one foetus, metaphase FISH showed normal results, but the interphase FISH suggested cell lines with two, three and four copies of 12p in the amniotic fluid. Advanced parental age may be an important risk factor for PKS, and there were no typical ultrasound manifestations related to PKS. A combination of karyotype analysis and molecular diagnosis is an effective method for the diagnosis of PKS.


| INTRODUC TI ON
Pallister-Killian syndrome (PKS) (OMIM:#601803), also known as 12p tetraploid syndrome or 12p isobrachial (i(12p)) chromatid syndrome, was initially described by Pallister et al. in 1977 1 and estimated to affect 1 in 20,000 live births. 2 With the advance of molecular diagnostics, the incidence was re-evaluated as 5.1 per 1,000,000 live births. 3 Generally, it is cytogenetically characterized by tetrasomy of 12p through mosaic supernumerary isochromosomes 12p. The clinical features are highly variable, involving a variety of phenotypes associated with multiple systems, such as craniofacial dysmorphism characterized as bitemporal alopecia and palpebral fissures, skin pigmentary anomalies, variable developmental and neurodevelopmental delays, epilepsy, congenital diaphragmatic hernia, congenital heart defects, gastrointestinal malformations and genitourinary malformations. [3][4][5][6][7] In prenatal settings, the diagnosis of PKS is challenging. It was first diagnosed prenatally in 1985 via amniocytes as a tetrasomy of 12p. Since then, no more than 100 prenatal cases have been published. 4 The i(12p) cells were generally tissue-limited, and the percentages of cells with i(12p) differed significantly between different tissues in the same individual or between cultured and uncultured prenatal specimens. 2 During amniocyte sub-culturing, the supernumerary marker of i(12p) may decrease rapidly. Furthermore, there were no specific ultrasound abnormalities related to PKS; thus, the detection is usually incidental in prenatal settings. The clinical indicators that have been reported in foetus with PKS were diverse, including advanced maternal age, 3,8,9 increased nuchal translucency (NT), thickened nuchal fold, increased prenasal thickness, 3,8,[10][11][12] nasal bone hypoplasia, polyhydramnios, diaphragmatic hernia, [13][14][15] short long bones, cerebral ventriculomegaly and cardiac abnormalities. 10 Among them, polyhydramnios, diaphragmatic hernia and rhizomelic limb shortening were the most common prenatal ultrasound abnormalities reported in PKS. 16 With awareness about prenatal diagnosis and the development of molecular detection technology, the diagnosis of PKS prenatally has become more common. Single nucleotide polymorphism array (SNP array) can detect 10% of abnormal cells or even less if mosaicism involves the introduction of a new haplotype. 8 Next-generation sequencing is emerging as a viable alternative to chromosome microarray analysis, and copy number variants sequencing (CNVseq) was considered to have comparable detection efficiency as SNP array. 17

| Conventional karyotyping
Conventional karyotyping consisted of cell culture, and G-banded karyotyping was performed on cultured amniotic fluid and foetal cord blood according to the standard protocols in our laboratory. The karyotype was determined at a resolution of 320-500 band level.

| SNP array
Genomic DNA was extracted from uncultured amniotic fluid, foetal cord blood, skin biopsy and placenta using a QIAGEN kit (Qiagen,

| FISH
FISH was performed on the amniotic fluid sampled from the second amniocentesis of foetus 4. Chromosome 12p-specific probe (12pter) and chromosome 12q-specific probe (12qter) were used for FISH studies.

| Ultrasound findings
Abnormal ultrasound findings were recorded in three foetuses (foetuses 1 and 2 and foetus 4;

| Cytogenetic and molecular test results
The details of the results are summarized in Table 2. ( Figure 1E), which were confirmed by SNP array.

| DISCUSS ION
The PKS phenotype could be caused by a mosaicism of partial tetrasomy of 12p or a duplication of 12p due to the dosage-sensitive genes of 12p. 19,20 In the present study, the karyotypes revealed mosaic tetrasomy of 12p in three cases, mosaic trisomy of 12p in one case and non-mosaic tetrasomy of 12p in one case. It was reported that a minimal critical region that was located at 12p13. 31

F I G U R E 1 Abnormal karyotype of the 5 foetuses (A) (B) (C) (D) (E)
Advanced maternal age (≥35 years) was observed in the majority of previously reported cases. 2,8,14,26 The exact mechanism behind i(12p) cell line generation has not been clearly illustrated. Several theories have been proposed, majority of which suggested maternal meiosis II non-disjunction as a mechanism of mosaic tetrasomy of 12p 3,27 similar to the effect of the maternal age on aneuploidy pregnancies, although there is another theory of a paternal origin. 28,29 In our study, all the five cases had advanced maternal and paternal ages, which supported the association between advanced parental age and PKS. The population-based data by Moira Blyth showed a   Our study has several limitations. It has a small sample size and FISH validation was not performed on all samples to determine the abnormal cell composition.
In conclusion, our study showed the atypical manifestations of PKS in prenatal diagnosis such as ultrasound finding and diagnostic specimens. PKS may occur even if the ultrasound of the second trimester is normal. Advanced parental age is the most important factor for PKS. Only by a combination of cytogenetics and molecular techniques can an accurate diagnosis of PKS be made.

CO N FLI C T O F I NTE R E S T
The authors declare they have no conflict of interest.

DATA AVA I L A B I L I T Y S TAT E M E N T
All data supporting the findings of this study are available within the article.