Comparative Genomic Hybridization in the Study of Human Disease

Abstract

Microarray‐based comparative genomic hybridization (CGH) has made a significant impact on the ability to diagnose human constitutional disease by detecting genomic copy number changes that were previously undetectable by other types of cytogenetic and molecular technologies. Not only can hundreds of well‐characterized genetic syndromes be detected in a single assay, but new genomic disorders and disease‐causing genes are also being discovered through the utilization of this technology. Clinical implementation of array CGH Hybridization has been extended to the prenatal setting, where it is also proving to enhance the diagnostic capabilities in the perinatal period. However, the clinical interpretation of the increasing number of copy number variations detected as the resolution of the microarrays is improved still poses a formidable challenge to laboratorians, health care providers and families.

Key Concepts:

  • Array comparative genomic Hybridization (aCGH) has several advantages over traditional cytogenetic methods for diagnosing human diseases: higher resolution, more robust and automated, and shorter turn around time since no cell culture is required.

  • Copy number variation (CNV) is common in the genome making it challenging for the clinical significance to be determined. Parental testing and utilisation of several internet‐based databases assist with the interpretation of CNVs.

  • The detection rate for clinically relevant CNVs by aCGH (10–20%) is significantly higher than traditional chromosome analysis (3%) making it the recommended first‐tier cytogenetic diagnostic test for patients with unexplained developmental delay/intellectual disability, autism spectrum disorders and multiple congenital anomalies by the International Standard Cytogenomic Array Consortium.

  • The resolution of aCGH has evolved to be sensitive enough to aid in the diagnosis of single gene disorders.

  • Array CGH is a useful tool for discovering new disease‐causing genes.

  • Array CGH has led to the recognition of many new genomic disorders, many of which cannot be diagnosed clinically due to lack cardinal features, variable expressivity and reduced penetrance.

  • CNVs, particularly those that occur de novo, are increasingly being recognised as important in the etiology of both syndromic and nonsyndromic autism as well as other neuropsychiatric disorders.

  • SNP‐based microarrays can be used to diagnosis uniparental disomy.

  • Array CGH is increasingly being used in prenatal diagnosis and has demonstrated its usefulness in clarifying the significance of karyotype findings and providing diagnoses not identifiable by chromosome analysis alone.

  • It is the opinion of The American College of Obstetrician and Gynecologists (2009) that aCGH not currently replace classic cytogenetics for prenatal diagnosis, but that targeted aCGH can be offered as an adjunct tool in prenatal cases with abnormal anatomical findings and normal karyotype, as well as in cases of fetal demise with congenital anomalies and the inability to obtain a conventional karyotype.

Keywords: array comparative genomic Hybridization; copy number variation; genomic disorders; autism; uniparental disomy; prenatal diagnosis; microarray; microdeletion; microduplication

Figure 1.

Array comparative genomic hybridization. A schematic of the array CGH process is shown at the top of the slide. The bottom left portion of the slide shows a typical representation of the array data generated by in‐house software for copy number analysis with an image of the FISH confirmation on the right. (a) Genomic DNA from the patient is labelled with a green fluorescent dye (Cy5) and genomic DNA from a normal control is labelled with a red fluorescent dye (Cy3). (b) The two samples are mixed and co‐hybridized to the array of DNA fragments. (c) A laser scanner reads the fluorescent signals and the intensities of each color are quantified using special software. A copy number loss is indicated by a red spot (more control and less patient DNA) and a copy number gain is indicated by a green spot (more patient and less control DNA). (d) The graph is arranged so that chromosomal data are presented in order from chromosome 1 on the left to chromosome Y on the right. The log2 ratio scale is shown on the Y‐Axis. The centre line represents neutral copy number, and gains and losses are plotted above and below this line, respectively. In this example, a loss in copy number on chromosome 17p12 is shown in red. The table below shows the location, size of deletion, log ratio as well as the number of the oligos within this region. (e) FISH confirmation shows lack of signal (red oval) for target probe on one chromosome 17, confirming the deletion (green signal is control probe, red signal is target probe) Deletions in this region have been reported in patients with hereditary neuropathy with liability to pressure palsies (HNPP, OMIN 162500).

Figure 2.

Detection of genomic disorders. Detection of 22q.11.2 microdeletion syndrome and reciprocal 22q11.2 microduplication syndrome by array CGH with FISH confirmation. (a1) Array CGH showing a loss in copy number of chromosome band 22q11.2 involving the 22q11.2 deletion syndrome region (red circle). (a2) FISH analysis shows lack of signal (red oval) for target probe on one chromosome 22, confirming the deletion (green signal is control probe, red signal is target probe). Insert‐G‐banded chromosome analysis showing the deletion on one chromosome 22 (black arrow). (b1) Array CGH showing a gain in copy number of chromosome band 22q11.2 involving the 22q11.2 duplication syndrome region (red circle). (b2) FISH analysis shows three signals for the target probe, confirming the duplication (green signal is control probe, red signal is target probe). Insert‐G‐banded chromosome analysis showing the duplication on one chromosome 22 (black arrow).

Figure 3.

Detection of UPD and IBD by SNP array analysis. SNP array analysis showing evidence of uniparental disomy (UPD) and identity by descent (IBD). (a) UPD for the entire chromosome 15 is indicated by absence of heterozygosity (top panel‐lack of signal at 0.5 B allele frequency (BAF) which represents genotype A/B) and no change in copy number (bottom panel‐all signals are at 0 Log ratio). (b) Blocks of absence of heterozygosity (AOH) of the proximal regions of chromosome 9p and 9q as demonstrated by lack of signals at the 0.5 BAF. Within the block of AOH at 9p (red oval) is the gene for galactosaemia, GALT. The patient is affected with galactosaemia due to a homozygous mutation in the GALT gene. The parents are consanguineous, which is consistent with the multiple blocks of AOH.

Figure 4.

Prenatal diagnosis of TAR syndrome. Prenatal diagnosis of TAR syndrome by array CGH, FISH and ultrasound. (a) Array CGH showing a loss in copy number of chromosome band 1q21.1 involving the TAR syndrome region. (b) FISH analysis shows lack of signal for the target probe on one chromosome 1, confirming the deletion (green signal is control probe, red signal is target probe). (c) Ultrasound performed at 17 weeks showed bilateral absence of radii with the hands attached directly to the humeri (indicated by red arrows). (d) Ultrasound performed at 19 weeks showed humeri appeared markedly shortened and more curved but symmetrical and hands include thumbs bilaterally (indicated with red arrow).

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Cheung, Sau Wai, and Pursley, Amber Nolen(Nov 2011) Comparative Genomic Hybridization in the Study of Human Disease. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0005955.pub2]