Prenatal diagnosis of sub-microscopic partial trisomy 10q using chromosomal microarray analysis in a phenotypically abnormal fetus with normal karyotype

Case report

The patient was a 36-year-old Caucasian female gravida 3, para 0-0-2-0 referred at 13 weeks gestation for first trimester screening secondary to advanced maternal age. The patient’s nuchal translucency measurement and first trimester screening study were within normal limits. The patient declined chorionic villus sampling. At 17 weeks’ gestation, ultrasound showed widening of the palatal shelf and unilateral talipes (clubfoot). Following genetic counseling, an amniocentesis was performed and the karyotype was normal (46, XX). Chromosomal microarray analysis (CMA) was requested. Ultrasound at 21 weeks’ gestation showed microcephaly, a broad nasal bridge and widening of the paranasal sinuses (Fig. 1). Three-dimensional (3D) ultrasound confirmed severe hypertelorism (“fish-eye deformity”) and a broad nasal bridge (Fig. 2). Fetal magnetic resonance imaging (MRI) -identified the widening of the paranasal sinuses in conjunction with hypertelorism and broad nasal bridge (Fig. 3). Pregnancy termination at 21 weeks’ gestation by dilatation and evacuation was performed at the patient’s request.

The chromosomal microarray analysis or array-based comparative genomic hybridization (array CGH), on cultured fibroblasts from amniotic fluid revealed 2 copy number variants: a10q22.3–10q23.2 duplication and an Xp21.1 deletion. Parental CMA studies were performed and the duplication on 10q was determined to be de novo. The Xp21.1 deletion was found in the father, who had no phenotypic abnormalities or intellectual disability. The array-based comparative genomic hybridization (array CGH), can detect submicroscopic chromosome alterations at a resolution of 100 kb. The smallest reported genomic alterations of partial 10q trisomy previously reported using array CGH had a genomic size <12 Mb [11]. In our case, we identified a smaller genomic duplication of 7.1 Mb (Fig. 5). The deletion on Xp was identified in the paternal sample. The father had no clinical features of DMD, despite the 87.9 Kb deletion in this region. The Pedigree Structure of duplication in the 10q22.3–10q23.2 regions is shown in Fig. 5.

Discussion

Distal trisomy 10q syndrome is a well-defined but rare syndrome. Only 4 prior cases of partial trisomy 10q have been reported. All prior cases involved identification of microscopic karyotype abnormalities. The phenotypic features of partial Trisomy 10 are collected in (Table 1). Our patient had craniofacial features consistent with those previously described. The 10q24⟶qter region has been proposed as the critical region for distal trisomy 10q [12].

The current case represents the first prenatal diagnosis of a 10q22–10q23 duplication syndrome with a normal microscopic karyotype. This case highlights the importance of chromosomal microarray analysis in patients who have a prenatal diagnosis of a fetal abnormality and a subsequent normal karyotype [13].

Cytologically visible deletions of Xp21.1 are known to be associated with Duchenne Muscular Dystrophy (DMD). DMD represents an X-linked recessive disorder related to mutations in the dystrophin gene which is located on chromosome Xp21.1 [14]. There are no reported cases of co-existing 10q22.3–10q23.2 and Xp21.1 CNVs. This fetus was found to have a familial deletion on Xp21.1. DMD is associated with mild to severe cognitive deficits, which are independent from the muscular dysfunction [15]. However, in contrast to previously-reported cases [10] the father of this fetus did not have cognitive deficits.

The combination of microcephaly, craniofacial abnormalities and talipes (clubfoot) encompasses a wide differential diagnosis. Patients and clinicians consider a normal karyotype to be reassuring. However, they must be reminded that a normal microscopic karyotype only eliminates whole chromosome aneuploidy, chromosomal rearrangements or marker chromosomes. Although, the conventional karyotyping has a resolution of 5–10 million bases, it detects only 5% of chromosomal alterations in children with unexplained mental retardation [15–19]. As originally described, chromosomal microarray (MCA) uses hybridization to metaphase chromosomes on a slide, and produce resolution of >5–10 Mb [20]. By using dynamic standard reference intervals in high-resolution MCA, the resolution can be improved to 3 Mb [21]. The advantages of chromosomal microarray analysis are illustrated in this case where CNVs on two different chromosomes were identified despite a normal microscopic karyotype.

In the past, some clinicians have incorrectly assumed a normal karyotype meant a genetically “normal” fetus. With the advent of chromosomal microarray analysis, geneticists can now identify a larger number of well-defined genetic syndromes. In a 2012 NICHD multicenter trial comparing prenatal chromosomal microarray analysis to prenatal microscopic karyotype, CMA was able to detect all clinically significant aneuploidies and unbalanced translocations diagnosed with karyotyping [22].

The American Congress of Obstetricians and Gynecologists (ACOG) recommends chromosomal microarray analysis as the next step in management when an ultrasound reveals one or more major fetal abnormalities, especially after a negative karyotype analysis [22]. CMA can detect aneuploidy as well as submicrosomal abnormalities that are too small to be detected by conventional karyotype. CMA may be superior to microscopic karyotype for evaluation of a stillbirth, based on the larger number of genetic abnormalities which can be detected.

CMA still has some limitations. MCA cannot detect the balanced inversions, balanced translocation, tissue mosaicism or triploidy, so it is not recommended for couples who suffer from recurrent miscarriage. CMA also commonly reports “variants of uncertain significance” (VUS), which describe identified DNA changes that either have not yet been identified as pathogenic or that are associated with a variable phenotype (variable penetrance) [23].

CMA can detect several DNA alterations such as deleted or duplicated sections of DNA which are known as copy number variants (CNV), and these abnormalities are expected to be as high as 15% [17]. CMA also can detect “copy number variants” and mostly associated with structural fetal abnormalities. Most of the previously reported cases revealed either a mosaic trisomy of the long arm of chromosome 1 (unbalanced autosomal translocation between chromosome 1 and 2) [23] or a mosaic terminal 7q monosomy/distal 2p trisomy [24]. The former was the result of an unbalanced autosomal translocation between the chromosomes 1 and 15 [23]. However, the significant copy number variants can also be found in structurally normal fetuses that mostly been undiagnosed. It is estimated that the MCA detects 1.7% of DNA alterations of fetuses with a structurally normal ultrasonic examination result and a normal karyotype [20]. Another type of DNA alteration is a single-nucleotide polymorphism (SNP) in which a single nucleotide in the genome sequence is altered. This can occur between two different individuals or between paired chromosomes of the same individual and may or may not cause disease. In contrast with Down syndrome and other common trisomies. CMA also commonly diagnose “variants of uncertain significance” (VUS), which describe identified DNA changes that either have not yet been identified as pathogenic or benign or that are associated with a variable phenotype (variable penetrance) [18]. In addition, some of the genetic abnormalities associated with adult-onset disorders (eg, BRCA mutations or Charcot-Marie-Tooth disease), inherited from an asymptomatic parent can be detected by MCA and also certain types of MCA can provide an evidence of nonpaternity and consanguinity.

Because such a large number of potential findings can to be diagnosed with microarray technology, ACOG recommended to use MCA as a routine perinatal screening test for all cases of with structurally abnormal fetal ultrasonic examination and normal karyotype. When counseling patients whose fetus has been diagnosed with structural abnormalities on prenatal ultrasound, patients should be offered prenatal diagnosis using chorionic villus sampling or amniocentesis. If the microscopic karyotype analysis is abnormal, the patient can be counseled regarding the likely outcome of the pregnancy. When the microscopic karyotype is normal, it is strongly recommended that the patient should be offered chromosomal microarray analysis. Studies in the pediatric population have demonstrated that MCA can identify clinically significant abnormalities in approximately 6% of fetuses [25] and up to 15% of children with a normal microscopic karyotype abnormal chromosomal microarray analysis [15, 17].

Some perinatal studies have used MCA to differentiate between the syndromic and non-syndromic mental retardation in children with idiopathic mental retardation [13]. MCA detected chromosomal abnormalities in up to 17% of cases who had a normal karyotype. This study used a subtelomeric testing using genome-wide microarray platforms with around 2000 to >30,000 (tiling-path) interrogating BAC/PAC probes. Surprisingly, some of the detected anomalies were mosaic that were not detected by karyotyping. The author suggested that the commercially available genome-wide microarrays with >300,000 synthesized oligonucleotide probe provides a higher resolution with higher sensitivity and may eventually replace the BAC/PAC arrays in clinical laboratories [15].

Chromosomal microarray analysis is not routine in perinatal testing. It is used reflexively (after a normal standard karyotype) in cases of an abnormal ultrasound, despite the fact that array CMA abnormalities have been identified in 1.7% of fetuses with a normal prenatal ultrasound examination and a normal karyotype [17].

Summary

The current case describes the importance of prenatal chromosomal microarray analysis when evaluating a fetus with an abnormal prenatal ultrasound. The fetus was found to have microcephaly, severe hypertelorism and talipes (clubfoot). Microscopic karyotype performed on amniocentesis was normal. Chromosomal microarray analysis revealed 2 sub-microscopic chromosomal abnormalities, a partial 10q trisomy (10q22.3–10q23.2) and a deletion in the X chromosome (Xp21.1). Without the additional CMA, the fetus would have been presumed to be genetically normal.

The American Congress of Obstetricians and Gynecologists (ACOG) recommended that all patients with a structurally abnormal fetus undergoing invasive prenatal diagnostic testing. A normal karyotype should be followed by additional chromosomal microarray analysis (CMA). Patients should be advised of the limitations of microscopic karyotype. The additional information obtained from CMA can assist the clinician in the management of these difficult cases. Despite the superiority of microarray technology on karyotype in detecting a potential chromosomal abnormality in structurally abnormal fetuses, there is still some limitations for microarray technology itself.