Array comparative genomic hybridization for genetic evaluation of fetal loss between 10 and 20 weeks of gestation.

OBJECTIVE: To estimate genomic copy number changes in fetal loss between 10 and 20 weeks of gestation using array comparative genomic hybridization. METHODS: This was a prospective series of 35 women who experienced pregnancy loss between 10-20 weeks of gestation with either normal karyotype (n=9) or no conventional cytogenetic testing (n=26). DNA was isolated from fetal tissue and parental blood. Array comparative genomic hybridization was performed on DNA from fetal tissue using a whole genome BAC array chip. Copy number changes in fetal tissue were then compared against databases of benign copy number changes. Parental DNA was analyzed using the same BAC array in cases that contained suspected pathogenic copy number changes. In cases where de novo copy number changes were detected in fetal DNA, further characterization was performed using a 244K oligonucleotide microarray. RESULTS: DNA was successfully isolated in 30 of 35 (86%) of cases. Array comparative genomic hybridization was performed in all of these. De novo copy number changes were detected in six (20%) cases using the Spectral chip and confirmed in four (13%) cases using the Agilent chip. These ranged in size from 93-289 Kb and mapped on 5p, 13q and Xq22. In the cases with de novo copy number changes, the higher-density Agilent array detected additional changes (20-1,310 Kb). CONCLUSION: Array comparative genomic hybridization detected de novo copy number changes in 13% of cases where routine cytogenetic testing was normal or not performed. These involved large regions of DNA and may provide novel explanations for some cases of otherwise unexplained pregnancy loss. LEVEL OF EVIDENCE: III.

Detection of a de novo interstitial 2q microdeletion by CGH microarray analysis in a patient with limb malformations, microcephaly and mental retardation.

We describe the cytogenetic diagnosis using BAC- and oligonucleotide microarrays of a 16-year-old Laotian-American female, who first presented at 2 1/2 years of age with microcephaly, developmental retardation, and skeletal abnormalities of the upper limb including mild syndactyly of the second and third and the third and fourth fingers, short middle phalanges and clinodactyly of the fifth digit at the distal interphalangel joint on both hands, and symphalangism of the metacarpal-phalangeal joints of the second and fifth digits bilaterally. Her lower limbs displayed symphalangism of the metatarsal-phalangeal joint of the second, third, and fourth digits on both feet, with fusion of the middle and distal phalanges of the second and fifth digits and hallux valgus bilaterally. G-banded chromosomal study at age 4 was normal. However, comparative genomic hybridization at age 15 with the Spectral Genomics 1 Mb Hu BAC array platform indicated a microdeletion involving two BAC clones, RP11-451F14 --> RP11-12N7 at 2q31.1. The maximal deletion on initial analysis comprised the HOXD cluster, which is implicated in limb development. Fluorescence in situ hybridization (FISH) using the RP11-451F14 probe confirmed the deletion. Both parents were negative for the deletion. Additional FISH using BAC RP11-387A1, covering the HOXD cluster, limited the maximal deletion to approximately 2.518 Mb, and excluded involvement of the HOXD cluster. The Agilent 44K and 244K platforms demonstrated a deletion of approximately 2,011,000 bp, which did not include the HOXD cluster. The malformations in our patient may be caused by deletion of a regulatory element far upstream of the HOXD cluster.

Global hypomethylation is common in prostate cancer cells: a quantitative predictor for clinical outcome?

This study was designed to determine if cytological detection of 5-methylcytosine (5MC) was feasible on prostate tumor sections and to determine if levels of 5MC differed in malignant compared to normal prostate tissue. We further sought to see if 5MC levels correlated with any clinical outcome data. Thirty prostate tumor sections were obtained from patients who underwent radical prostatectomies from 1988 to 1995; these represented a mix of low to high grade tumors. Clinical data were maintained for each of these patients with a minimum of 7 years of follow up. Sections were stained with a commercially available antibody to 5MC and immunocytochemistry levels were subsequently quantified using a computer-assisted true-color imaging system. Tumor and benign regions of the same archived sections were compared, in addition to a series of 12 normal prostate samples. Prostate cancer cells exhibited a pronounced global decrease in methylation compared with benign and normal tissue. This was observed in 29 of 30 patients (96.7%) studied and densitometric scanning of methylation staining indicated that this value was quantifiable. Overall, higher methylation values were detected in men who had positive surgical margins and recurrent disease. These data suggest that loss of methylation is a feature of prostate cancer, and partial gain of methylation (presumably at promoters of specific genes) is associated with clinical outcome and is measurable using whole-cell assays.