Gene dose imbalances in children with mental retardation

Submicroscopic chromosome aberrations can cause mental retardation (MR), congenital malformations and miscarriage. It is difficult to estimate the contribution of cytogenetic abnormalities to MR because of relatively low detection sensitivity of conventional cytogenetic methods used in the diagnostic setting. The focus of this thesis has been to improve the detection sensitivity and to study small chromosome aberrations in detail. Using FISH probes covering the 41 unique subtelomeric regions of the human genome, 111 patients with idiopathic MR and dysmorphic features were screened. Ten cryptic rearrangements (9%) were detected - five de novo deletions, one unbalanced de novo translocation, three unbalanced inherited translocations and one unbalanced inherited recombinant chromosome. Fifty of the 111 patients were also analyzed by Spectral Karyotyping (SKY) to detect interstitial abnormalities, but no additional rearrangements were found. In one case an unbalanced translocation between chromosome 12q and 17q was detected and this patient together with her family were further investigated. The fragments involved were FISH mapped for accurate size determination and genotype-phenotype correlation was performed. Four family members had inherited unbalanced variants - two cases inherited the derivative chromosome 12 and the other two the derivative chromosome 17. This family illustrates that small chromosome imbalances can cause a mild phenotype and normal, or near-normal, cognitive functions. The robustness and simplicity of different methods for comparative genomic hybridization (CGH) were tested on a set of patients with well-defined sub-microscopic chromosome abnormalities, in order to determine if the technique can be used as a reliable and sensitive diagnostic procedure. We optimized and evaluated CGH performed on metaphase chromosomes and compared the performance to a commercially available high resolution software using a standard dynamic reference interval (HR-CGH). In addition, the performance of CGH on microarrays containing cDNA clones or BAC clones was investigated. From this study, we concluded that the BAC array with an average resolution of approximately 1 Mb is reliable and would add important information in selected cases in a diagnostic setting. Based on these results, two patient groups were screened for submicroscopic chromosome imbalances by array based CGH using a BAC microarray. The first group of patients consisted of 10 children diagnosed with Kabuki syndrome (KS). This syndrome is a rare multiple congenital anomaly/mental retardation syndrome which was recently reported to be caused by a microduplication on the short arm of chromosome 8. We investigated the implicated region by FISH and microsatellite markers and were unable to confirm the duplication. In addition, the whole genome at a resolution of approximately 1 Mb was screened, without detecting any microdeletions or duplications in this group of patients. Our data demonstrate that the etiology of KS remains unknown. The second group of patients consisted of 49 children with MR and dysmorphism, where G-banding, and in the majority of the cases subtelomeric FISH and SKY, had failed to detect chromosome imbalances. Five abnormalities (10.2%) were detected - four de novo interstitial deletions and one de novo terminal duplication. In addition, large scale copy number variations (LCVs) were frequently observed. Array-CGH proved to be a reliable and sensitive tool for the detection of submicroscopic chromosome imbalances and has the advantage over FISH that it enables whole genome screening. ArrayCGH will in the future most likely be more widely implemented in the diagnostic setting and the detection of genomic imbalances of clinical significance will increase. Thorough genotypephenotype correlations needs to be performed for the identification of candidate genes involved in developmental delay