Is telomere length in peripheral blood lymphocytes correlated with cancer susceptibility or radiosensitivity?

Mean terminal restriction fragment (TRF) lengths in white blood cells (WBCs) have been previously found to be associated with breast cancer. To assess whether this marker could be used as a test for breast cancer susceptibility in women, TRF length was measured in 72 treated female breast cancer patients and 1696 unaffected female controls between the ages of 45 and 77 from the Twin Research Unit at St Thomas' Hospital, as well as 140 newly diagnosed breast cancer cases and 108 mammographically screened unaffected controls from Guy's Hospital. Mean TRF was also tested for correlation with chromosome radiosensitivity and apoptotic response in the Guy's Hospital patients. After adjusting for age, smoking and body mass index, there was no significant difference in TRF lengths between the treated breast cancer patients and unaffected controls (P=0.71). A positive correlation between age-adjusted apoptotic response and mean TRF in newly diagnosed untreated breast cancer patients (P=0.008) was identified but no significant difference in TRF lengths between breast cancer patients and unaffected controls was detected (P=0.53). This suggests that TRF lengths in WBC, is not a marker of breast cancer susceptibility and does not vary significantly between affected women before and after treatment.

Biallelic mutation of MSH2 in primary human cells is associated with sensitivity to irradiation and altered RAD51 foci kinetics.

BACKGROUND: Reports of differential mutagen sensitivity conferred by a defect in the mismatch repair (MMR) pathway are inconsistent in their conclusions. Previous studies have investigated cells established from immortalised human colorectal tumour lines or cells from animal models. METHODS: We examined primary human MSH2-deficient neonatal cells, bearing a biallelic truncating mutation in MSH2, for viability and chromosomal damage after exposure to DNA-damaging agents. RESULTS: MSH2-deficient cells exhibit no response to interstrand DNA cross-linking agents but do show reduced viability in response to irradiation. They also show increased chromosome damage and exhibit altered RAD51 foci kinetics after irradiation exposure, indicating defective homologous recombinational repair. DISCUSSION: The cellular features and sensitivity of MSH2-deficient primary human cells are broadly in agreement with observations of primary murine cells lacking the same gene. The data therefore support the view that the murine model recapitulates early features of MMR deficiency in humans, and implies that the variable data reported for MMR-deficient immortalised human cells may be due to further genetic or epigenetic lesions. We suggest caution in the use of radiotherapy for treatment of malignancies in individuals with functional loss of MSH2.

Development and implementation of a new rapid aneuploidy diagnostic service within the UK National Health Service and implications for the future of prenatal diagnosis.

BACKGROUND: Prenatal diagnosis for chromosome abnormality is routinely undertaken by full karyotype analysis of chromosomes from cultured cells; pregnant women must wait on average 13-14 days for their results. Autosomal trisomies, which account for around 80% of significant abnormalities, can be detected by quantitative fluorescence (QF) PCR. We report on the development and implementation of this technique as the first such routine service within a diagnostic department of the UK National Health Service (NHS). METHODS: We designed a "one-tube test" comprising four primer pairs for polymorphic tetranucleotide repeat sequences on chromosome 21, four primer pairs for sequences on chromosome 18, three primer pairs for sequences on chromosome 13, and one primer pair to identify the sex chromosomes. All prenatal samples received by our NHS diagnostic department between April, 2000, and April, 2001, were tested. After DNA extraction, PCR amplification was done and the products separated on a capillary-based genetic analyser; the results were interpreted with dedicated software. Follow-up karyotype analysis was done on all samples. FINDINGS: 1148 amniotic fluid samples, 188 chorionic villus samples, and 37 fetal tissue samples were tested; the amplification failure rate was zero with our current protocol. QF-PCR results were obtained and reported on 1314 (98%) of the prenatal samples; the remaining 22 (2%) were uninformative because of maternal-cell contamination. One case of mosaicism in a chorionic villus sample, and two cases indicating somatic expansion of a tetranucleotide repeat were found. No false positive or false negative results were obtained. The mean reporting time for the last 4 months of data collection was 1.25 working days. INTERPRETATION: QF-PCR aneuploidy testing is an efficient and accurate technique for the detection of autosomal trisomies in prenatal samples. Implementation of this service has led to the rapid diagnosis of abnormalities and early reassurance for women with normal results.

Telomere shortening in Fanconi anaemia demonstrated by a direct FISH approach.

Analysis of telomere status in patients with Fanconi anaemia (FA) has previously been carried out by measurement of telomere restriction fragment (TRF) length by Southern blotting and densitometry. Results from these studies indicated that FA patients had significant reduction in telomere length compared with age-matched controls. This paper confirms and extends these findings using a direct FISH technique, which showed that 15 out of 16 FA patients had increased loss of telomere signals compared with controls. In 12 out of the 16 patients, decrease in telomere signal intensity could also be detected using a Q-FISH approach.

Support for linkage of autism and specific language impairment to 7q3 from two chromosome rearrangements involving band 7q31.

Childhood autism is characterised by impairments in communication and reciprocal social interaction together with restricted/stereotyped interests, which are evident before 3 years of age. Specific developmental disorders of speech and language (SDDSL) are characterised by impairment in the development of expressive and/or receptive language skills which is not associated with intellectual, sensory, physical, or neurological impairment. Family and twin studies indicate a substantial genetic component in the aetiology of both disorders. They also reveal increased rates of SDDSL in relatives of autistic individuals, suggesting that this phenotype can represent one manifestation of the genetic liability for autism. Modelling of the recurrence risk for autism and milder phenotypes, such as SDDSL, suggest that three or four epistatic loci may be aetiologically involved. A recently published linkage study of an exceptional family with an apparently dominantly inherited SDDSL implicated chromosome band 7q31 as the site of the putative susceptibility locus (SPCH1). This region of chromosome 7 also shows strong linkage in multiplex families with autism. We present two individuals (one has autism, the other SDDSL) with different, apparently balanced chromosome rearrangements involving a breakpoint at 7q31.3. Fluorescence in situ hybridisation was used to localise the breakpoints to an approximately 1 cM interval between CFTR and D7S643. Our findings may be of interest and relevance to the genetic aetiology of autism, and helpful in the search for susceptibility loci for SDDSL and autism. Am. J. Med. Genet. (Neuropsychiatr. Genet. ) 96:228-234, 2000.

Chromosome 22q11 deletions are not found in autistic patients identified using strict diagnostic criteria. IMGSAC. International Molecular Genetics Study of Autism Consortium.

A group of 103 subjects with a strict diagnosis of autism were tested for deletion of band q11.2 on the long arm of chromosome 22. No deletions were found, indicating that when a patient has been diagnosed with autism using strict and consistent criteria, in the absence of other indications, it is unlikely that this individual will have a 22q11 deletion. Testing for 22q11 deletions is therefore unlikely to be necessary in these patients. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:15-17, 2000.

Clinical expression of Menkes disease in a girl with X;13 translocation.

Menkes disease is a rare X-linked recessive disorder of copper metabolism, characterised by progressive neurological degeneration, abnormal hair and connective tissue manifestations. We report on a girl with classic Menkes disease, carrying a de novo balanced translocation 46,X,t(X;13)(q13.3; q14.3). The translocation breakpoints at Xq13.3 and 13q14.3 coincide with the Menkes disease and Wilson disease loci, respectively.

A new approach to the elucidation of complex chromosome rearrangements illustrated by a case of Rieger syndrome.

A patient with a complex chromosome rearrangement and unilateral Rieger syndrome is presented. This rearrangement involves four chromosomes and six breakpoints, one of which is at 4q25, the candidate region for Rieger syndrome. We discuss a novel approach to the elucidation of this case using a multiprobe fluorescence in situ hybridisation method to show rearrangements unpredictable from G banded analysis, and the clear and unambiguous presentation of the karyotype using computer generated colour ideograms.

Rapid identification of multiple supernumerary ring chromosomes with a new FISH technique.

Multiple supernumerary ring chromosomes are a rare cytogenetic finding which is poorly understood. With the introduction of FISH techniques, their chromosomal origin can now be defined clearly. The techniques described previously are complicated and time consuming. We report a new rapid technique which has been used to investigate two new cases. Multiple probes were hybridised to a single slide by means of marking the underside with a diamond pen to form a grid of squares, pipetting fixed cell suspension into the centre of each square, forming a rubber solution grid on the denatured, dehydrated slide following the lines on the underside, adding a mixture of probes into each square, and sealing the slide with a silicone rubber rim and a covering slide. The type of probe and the size, dimensions, and number of squares in the grid can be tailored to individual cases. The two new cases examined here are mosaic for three (case 1) and four (case 2) supernumerary ring chromosomes derived from different chromosomes. Normal cell lines were also present. The karyotypes were established as 47,XY,+r(4)/47,XY,+r(17)/.../48,XY,+r(17),+r(20)/ 49,XY,+r(4),+r(17),+r(20)/46,XY for case 1 and 47,XX,+r(4)/47,XX,+r(8)/47,XX,+r (10)/48,XX,+r(X),+r(4)/... /49,XX,+r(X),+r (8),+r(10)/46,XX for case 2. Our findings suggest that the ring chromosomes were formed during meiosis, perhaps involving complex rearrangements, resulting in a germ cell containing all markers, with subsequent loss of markers during cell division. Our second case also shows that the outcome is not invariably mental or physical handicap.

Phenotypic diversity in the Smith-Lemli-Opitz syndrome.

The phenotype of four cases of Smith-Lemli-Opitz syndrome (SLO) with proven defects in cholesterol biosynthesis are compared, and shown to be markedly disparate even between sibs, and demonstrate the dilemma for the clinician. The advent of a biochemical test for SLO has been enormously valuable in determining which patients are truly affected by the condition, but because of the wide phenotypic variation, a diagnosis on clinical features alone remains problematic.

Chromosome deletion 17p11.2 (Smith-Magenis syndrome) in seven new patients, four of whom had been referred for fragile-X investigation.

We report seven new patients with clinical features of the Smith-Magenis syndrome (SMS) and small de novo interstitial deletions of 17p11.2. Four of these patients had been referred for fragile-X studies, but standard G-banded chromosome analysis routinely carried out in addition to the fragility tests revealed the microdeletion in chromosome 17. A relatively high proportion (approximately 2%) of patients referred to this Centre for fragile-X investigation are found to have a chromosome abnormality other than fra(X)(q27.3), half of them (approximately 1%) with an unbalanced chromosome complement. The four of our seven patients with deletion 17p11.2 constitute 25% of those with an unbalanced karyotype, and establish this microdeletion as the most common chromosome abnormality-except for fra(X)(q27.3)-in patients referred for fragile-X screening. The data indicate that standard karyotyping should be offered to patients with this referral indication, in addition to any molecular or chromosome fragility tests for fragile X. We also recommend that the short arm of chromosome 17 be examined critically in these patients. Moderate quality and resolution of banding (450-550 bands per haploid chromosome set) are adequate for detection of the 17p11.2 deletion.

Down syndrome with partial duplication and del (21) syndrome: study protocol and call for collaboration. Study I: Clinical assessment.

We report on the clinical and cytogenetic assessment of five cases of Down syndrome phenotype with either a partial duplication of chromosome 21 or a normal karyotype, and we quote a case of del (21q) syndrome. Down syndromes with a partial duplication of chromosome 21 (as well as cases of del (21q), which are partly the phenotypic countertype of trisomy 21) are of paramount importance in the understanding of genes involved in the phenotype of Down syndrome. The goal is to find the relevant genes implicated in the main traits of Down syndrome (i.e. mental retardation, Alzheimer disease, and serious visceral malformations). Such a goal, in our opinion, cannot be reached just by publishing the genotype and the phenotype of a small cohort of patients: 1. a sufficient number of accurate cases is needed, and 2. data have to be computerized for definite conclusions to be reached. The main aims of this report are to present our study protocol and to invite colleagues to participate in a collaborative study in order to collect a maximum of these (rare) cases.

A fetus with an abnormal chromosome 7 and possible hydrolethalus syndrome.

A fetus with multiple abnormalities phenotypically similar to hydrolethalus syndrome, but also with broad thumbs, was found to have a de novo interstitial deletion of the terminal segment of the long arm of chromosome 7 and a small rearrangement within the proximal half of the p arm of the same chromosome. Terminal deletions of chromosome 7q fall into two broad phenotypic groups, neither of which resembles this fetus, but the digital anomalies are attributed to the 7p rearrangement disrupting the Greig syndrome gene.

Deletion 9p and sex reversal.

We report a case of a female infant with a de novo deletion of the short arm of chromosome 9, sex reversal, and an apparently intact SRY gene. Sex reversal has been reported in a number of subjects with a normal Y chromosome and a deletion of the terminal segment of the short arm of chromosome 9. The factors controlling early development of the male testes are unknown. There are likely to be many genes involved and we present additional evidence that one of these is situated on the end of the short arm of chromosome 9.

Where have all the fragile X boys gone?

A four-year retrospective survey of individuals referred for fragile X testing to South East Thames Regional Genetics Service was carried out to determine the accuracy of clinical diagnosis of fragile X syndrome among routine referrals for cytogenetic confirmation. 680 individuals from 565 pedigrees were tested for fragile X. Five affected males were identified in previously unknown families and 17 new pedigrees were diagnosed. Using the accepted prevalence data, a total of 80 affected males would have been expected in this period. The most likely explanation for the low diagnosis rate is failure of referral of affected males.

Prenatal diagnosis of chromosome abnormalities. A comparison of the results of various techniques, with special emphasis on mosaicism.

Prenatal diagnosis of chromosome abnormalities can be performed on three different samples; chorion villi (CVS), amniotic fluid (AFS) and fetal blood (FBS). We are presenting data from our own experience on the chromosome analysis of 957 CVS, 1000 AFS and 927 FBS. A total of 69 chromosome abnormalities have been detected in the CVS, 38 in the AFS and 115 in the FBS. The type of abnormalities and their frequencies are compared between the three sampling methods. Our findings are in agreement with published data, and the higher incidence of chromosomal aberrations in the FBS group reflect the greater efficiency with which aneuploidies associated with more severe congenital malformation can be detected by ultrasound. Finally, we reported 18 cases of mosaicism in CVS, 76 in AFS and 31 in FBS. Of these cases, only 10 represented a true mosaicism of the fetus, 98 cases have been classified as pseudomosaicisms and 7 identified as maternal contamination. We have encountered 9 cases of mosaicism confined to the chorionic villi and 1 case limited to the amniotic fluid cells. There appeared to be a similarity between trisomies involved in chorion confined mosaicisms and pseudomosaicism cases of the AFS. The chromosome complement of the placenta may play an important role in the development of the pregnancy, and although a mosaic result in prenatal diagnosis could present difficulties in in the prediction of the fetal karyotype, it might give important information on the general condition of the fetus.