A tandem duplication of BRCA1 exons 1-19 through DHX8 exon 2 in four families with hereditary breast and ovarian cancer syndrome.

PURPOSE: The purpose of this study is to characterize a novel structural variant, a large duplication involving exons 1-19 of the BRCA1 gene in four independent families, and to provide diagnostically valuable information including the position of the breakpoints as well as clues to its clinical significance. METHODS: The duplication of exons 1-19 of the BRCA1 gene was initially detected by routine laboratory testing including MLPA analysis and next generation sequencing. For detailed characterization we performed array-comparative genome hybridization analysis, fluorescent in situ hybridization, next generation mapping, and long-distance PCR for break-point sequencing. RESULTS: Our data revealed a tandem duplication on chromosome 17 that encompassed 357 kb and included exons 1-19 of the BRCA1 gene and the genes NBR2, NBR1, TMEM106A, LOC100130581, ARL4D, MIR2117 up to parts of the DHX8 gene. This structural variant appeared as a tandem duplication with breakpoints in intron 19 of the BRCA1 gene and in intron 3 of the DHX8 gene (HGVS:chr17(hg19):g.41210776_41568516dup). Segregation analysis indicated that this structural rearrangement is phased in trans with a known pathogenic exon deletion of the BRCA1 gene in one family. CONCLUSIONS: The copy number variation initially recognized as duplication of exon 1-19 of the BRCA1 gene by MLPA analysis is a structural variation with breakpoints in the BRCA1 and DHX8 genes. Although currently to be classified as a variant of unknown significance, our family data indicates that this duplication may be a benign variation or at least of markedly reduced penetrance since it occurs in trans with another known fully pathogenic variant in the BRCA1 gene.

[Targeted Early Detection and Prevention of Hereditary Colorectal Carcinomas]. / Gezielte Früherkennung und Prävention von erblichen kolorektalen Karzinomen.

Every year, about 60 000 people in Germany contract colo-rectal carcinoma. Hereditary factors are the cause in approx. 5 % and those affected often fall ill at a young age. Often there are concrete indications of an individual high risk in affected families. The identification of persons at risk enables a targeted early detection and prevention of cancer as an important interdisciplinary medical task. The current AWMF guideline "Colorectal carcinoma" makes concrete statements in this regard.

Short stature before puberty: which children should be screened for SHOX deficiency?

OBJECTIVE: We studied the prevalence of deficiency in the short stature homeobox containing gene (SHOX) in prepubertal short-statured children and analyzed the clinical and radiological signs. METHODS: Screening for SHOX deficiency was performed in 449 prepubertal short-statured children (54% females, aged 4-10 years) by direct sequencing and multiplex ligation probe-dependent amplification. Children with SHOX deficiency were compared to 1:2 age- and gender-matched prepubertal children without SHOX deficiency with respect to left-hand radiographs and anthropometrics including different ratios to height and proposed scores. RESULTS: We identified 22 (4.9%) patients with SHOX deficiency (64% point mutations). Children with SHOX deficiency demonstrated a mesomelic shortening of extremities. Lower leg lengths but not forearm length was reduced in children <8 years with SHOX deficiency. 36% of all children and none of the children <8 years with SHOX deficiency demonstrated any typical radiologic sign. Increased sitting height-to-height ratio and decreased extremities-to-trunk ratio demonstrated the best positive and negative predictive values to identify SHOX deficiency. CONCLUSIONS: Screening for SHOX deficiency seems rational, especially in children with increased sitting height-to-height ratio or decreased extremities-to-trunk ratio. These criteria were also valid in young children.

Diagnostic biochip array for fast and sensitive detection of K-ras mutations in stool.

BACKGROUND: Tumor cells that shed into stool are attractive targets for molecular screening and early detection of colon or pancreatic malignancies. We developed a diagnostic test to screen for 10 of the most common mutations of codons 12 and 13 of the K-ras gene by hybridization to a new biochip array. METHODS: DNA was isolated from 26 stool samples by column-based extraction from 9 cell lines. Peptide nucleic acid (PNA)-mediated PCR clamping was used for mutant-specific amplification. We used a biochip, consisting of a small plastic support with covalently immobilized 13mer oligonucleotides. The read out of the biochip was done by confocal time-resolved laser scanning. Hybridization, scanning, and data evaluation could be performed in <2 h. RESULTS: Approximately 80 ng of DNA was obtained from 200-mg stool samples. No inhibition of the PCR by remaining impurities from stool was observed. Mutation detection was possible in 1000-fold excess of wild-type sequence. Discrimination ratios between the mutations were >19 as demonstrated by hybridization with tumor cell line DNA. Stool samples (n = 26) were analyzed in parallel with PNA-PCR, restriction assay for K-ras codon 12 mutations, sequencing, and hybridization to the biochip. Nine mutations were found by hybridization, all confirmed by sequencing. PNA-PCR alone leads to an overestimation of mutations because suppression of the wild type is not effective enough with high concentrations of wild-type DNA. The restriction assay found only four mutations. CONCLUSIONS: The K-ras biochip is well suited for fast mutation detection from stool in colorectal cancer screening.