Polymorphisms in the vitamin D receptor and their associations with risk of schizophrenia and selected anthropometric measures.

The association between vitamin D levels and skeletal growth has long been recognized. However, exposure to low levels of vitamin D during early life is also known to alter brain development, and is a candidate risk factor for schizophrenia. This study examines the association between four polymorphisms in the vitamin D receptor (VDR) and 1) risk of schizophrenia, and 2) three anthropometric variables (height, head size, and head shape). Four single-nucleotide polymorphisms (SNPs; rs10735810/FokI, rs1544410/BsmI, rs7975232/ApaI, and rs731236/TaqI) in the VDR gene were genotyped in 179 individuals with schizophrenia and 189 healthy controls. No significant associations were detected between any of the four VDR SNPs and risk of schizophrenia. Patients were slightly but significantly shorter compared to controls. Of the four SNPs, only rs10735810/FokI was associated with any of the anthropometric measures: the M4 isoform of this SNP was significantly associated with larger head size (P = 0.002). In light of the evidence demonstrating a role for vitamin D during brain development, the association between polymorphisms in VDR and brain development warrants closer scrutiny.

Radioactive waste disposal implications of extending Part IIA of the Environmental Protection Act to cover radioactively contaminated land.

A short study has been carried out of the potential radioactive waste disposal issues associated with the proposed extension of Part IIA of the Environmental Protection Act 1990 to include radioactively contaminated land, where there is no other suitable existing legislation. It was found that there is likely to be an availability problem with respect to disposal at landfills of the radioactive wastes arising from remediation. This is expected to be principally wastes of high volume and low activity (categorised as low level waste (LLW) and very low level waste (VLLW)). The availability problem results from a lack of applications by landfill operators for authorisation to accept LLW wastes for disposal. This is apparently due to perceived adverse publicity associated with the consultation process for authorisation coupled with uncertainty over future liabilities. Disposal of waste as VLLW is limited both by questions over volumes that may be acceptable and, more fundamentally, by the likely alpha activity of wastes (originating from radium and thorium operations). Authorised on-site disposal has had little attention in policy and guidance in recent years, but may have a part to play, especially if considered commercially attractive. Disposal at BNFL's near surface disposal facility for LLW at Drigg is limited to wastes for which there are no practical alternative disposal options (and preference has been given to operational type wastes). Therefore, wastes from the radioactively contaminated land (RCL) regime are not obviously attractive for disposal to Drigg. Illustrative calculations have been performed based on possible volumes and activities of RCL arisings (and assuming Drigg's future volumetric disposal capacity is 950,000 m3). These suggest that wastes arising from implementing the RCL regime, if all disposed to Drigg, would not represent a significant fraction of the volumetric capacity of Drigg, but could have a significant impact on the radiological capacity with respect to 226Ra plus 232Th. The government's decision-making programme for managing solid radioactive wastes in the UK may possibly achieve a general consensus that the use of landfill for LLW from the RCL regime has a fundamental role to play. However, this is unlikely to change the situation within the next few years. No new national facility arising from this programme is likely to be available during the first decade of the operation of a new RCL regime. Hence it appears that Drigg will need to play an important role for some years to come.

Molecular genetics of schizophrenia.

1. Schizophrenia is a chronic, disabling brain disease that affects approximately 1% of the world's population. It is characterized by delusions, hallucinations and formal thought disorder, together with a decline in socio-occupational functioning. While the causes for schizophrenia remain unknown, evidence from family, twin and adoption studies clearly demonstrates that it aggregates in families, with this clustering largely attributable to genetic rather than cultural or environmental factors. Identifying the genes involved, however, has proven to be a difficult task because schizophrenia is a complex trait characterized by an imprecise phenotype, the existence of phenocopies and the presence of low disease penetrance. 2. The current working hypothesis for schizophrenia causation is that multiple genes of small to moderate effect confer compounding risk through interactions with each other and with non-genetic risk factors. The same genes may be commonly involved in conferring risk across populations or they may vary in number and strength between different populations. To search for evidence of such genetic loci, both candidate gene and genome-wide linkage studies have been used in clinical cohorts collected from a variety of populations. Collectively, these works provide some evidence for the involvement of a number of specific genes (e.g. the 5-hydroxytryptamine (5-HT) type 2a receptor (5-HT2a) gene and the dopamine D3 receptor gene) and as yet unidentified factors localized to specific chromosomal regions, including 6p, 6q, 8p, 13q and 22q. These data provide suggestive, but no conclusive, evidence for causative genes. 3. To enable further progress there is a need to: (i) collect fine-grained clinical datasets while searching the schizophrenia phenotype for subgroups or dimensions that may provide a more direct route to causative genes; and (ii) integrate recent refinements in molecular genetic technology, including modern composite marker maps, DNA expression assays and relevant animal models, while using the latest analytical techniques to extract maximum information in order to help distinguish a true result from a false-positive finding.

Second stage of a genome scan of schizophrenia: study of five positive regions in an expanded sample.

In a previous genome scan of 43 schizophrenia pedigrees, nonparametric linkage (NPL) scores with empirically derived pointwise P-values less than 0.01 were observed in two regions (chromosomes 2q12-13 and 10q23) and less than 0.05 in three regions (4q22-23, 9q22, and 11q21). Markers with a mean spacing of about 5 cM were typed in these regions in an expanded sample of 71 pedigrees, and NPL analyses carried out. No region produced significant genomewide evidence for linkage. On chromosome 10q, the empirical P-value remained at less than 0.01 for the entire sample (D10S168), evidence in the original 43 pedigrees was slightly increased, and a broad peak of positive results was observed. P-values less than 0.05 were observed on chromosomes 2q (D2S436) and 4q (D4S2623), but not on chromosomes 9q or 11q. It is concluded that this sample is most supportive of linkage on chromosome 10q, with less consistent support on chromosomes 2q and 4q. Am. J. Med. Genet. (Neuropsychiatr. Genet.) 96:864-869, 2000.

Follow-up study on a susceptibility locus for schizophrenia on chromosome 6q.

Evidence for suggestive linkage to schizophrenia with chromosome 6q markers was previously reported from a two-stage approach. Using nonparametric affected sib pairs (ASP) methods, nominal p-values of 0.00018 and 0.00095 were obtained in the screening (81 ASPs; 63 independent) and the replication (109 ASPs; 87 independent) data sets, respectively. Here, we report a follow-up study of this 50cM 6q region using 12 microsatellite markers to test for linkage to schizophrenia. We increased the replication sample size by adding an independent sample of 43 multiplex pedigrees (66 ASPs; 54 independent). Pairwise and multipoint nonparametric linkage analyses conducted in this third data set showed evidence consistent with excess sharing in this 6q region, though the statistical level is weaker (p=0.013). When combining both replication data sets (total of 141 independent ASPs), an overall nominal p-value=0.000014 (LOD=3. 82) was obtained. The sibling recurrence risk (lambdas) attributed to this putative 6q susceptibility locus is estimated to be 1.92. The linkage region could not be narrowed down since LOD score values greater than three were observed within a 13cM region. The length of this region was only slightly reduced (12cM) when using the total sample of independent ASPs (204) obtained from all three data sets. This suggests that very large sample sizes may be needed to narrow down this region by ASP linkage methods. Study of the etiological candidate genes in this region is ongoing.

Genome scan of schizophrenia.

OBJECTIVE: The goal of this study was to identify chromosomal regions likely to contain schizophrenia susceptibility genes. METHOD: A genomewide map of 310 microsatellite DNA markers with average spacing of 11 centimorgans was genotyped in 269 individuals--126 of them with schizophrenia-related psychoses--from 43 pedigrees. Nonparametric linkage analysis was used to assess the pattern of allele sharing at each marker locus relative to the presence of disease. RESULTS: Nonparametric linkage scores did not reach a genomewide level of statistical significance for any marker. There were five chromosomal regions in which empirically derived p values reached nominal levels of significance at eight marker locations. There were p values less than 0.01 at chromosomes 2q (with the peak value in this region at D2S410) and 10q (D10S1239), and there were p values less than 0.05 at chromosomes 4q (D4S2623), 9q (D9S257), and 11q (D11S2002). CONCLUSIONS: The results do not support the hypothesis that a single gene causes a large increase in the risk of schizophrenia. The sample (like most others being studied for psychiatric disorders) has limited power to detect genes of small effect or those that are determinants of risk in a small proportion of families. All of the most positive results could be due to chance, or some could reflect weak linkage (genes of small effect). Multicenter studies may be useful in the effort to identify chromosomal regions most likely to contain schizophrenia susceptibility genes.

The molecular genetics of schizophrenia: an update.

OBJECTIVE: This paper aims to summarise the latest molecular genetic findings in schizophrenia, while providing background information on a number of relevant methodological issues. METHOD: Accumulative genetic data indicate that schizophrenia is a genetically complex disease with an unclear mode of transmission. The development and rapid progression of molecular genetics have provided a wide variety of methods to search for genes predisposing to human disease. The genetic basis for a number of the simpler diseases has been identified and characterised using these methods. More recently, progress has been made in identifying genes predisposing to the genetically more complex diseases such as diabetes mellitus, multiple sclerosis, bipolar disorder and schizophrenia. RESULTS: The latest findings on chromosomes 3, 6, 8, 13, 18 and 22 and on the X chromosome are reviewed. CONCLUSIONS: There is now suggestive support for three susceptibility loci (6p24-22, 8p22-21 and 22q12-q13.1) for schizophrenia, and it is likely that other regions will emerge from studies now in progress. Finding and then characterising genes within these loci will require long-term commitment and systematic efforts in clinical, laboratory and analytical fields.

A linkage study of schizophrenia to markers within Xp11 near the MAOB gene.

A sex chromosome locus for psychosis has been considered on the basis of some sex differences in genetic risk and expression of illness, and an association with X-chromosome anomalies. Previous molecular genetic studies produced weak evidence for linkage of schizophrenia to the proximal short arm of the X-chromosome, while some other regions were not ruled out. Here we report an attempt to expand the Xp findings in: (i) a multicenter collaboration focusing on 92 families with a maternal pattern of inheritance (Study I), and (ii) an independent sample of 34 families unselected for parental mode of transmission (Study II). In the multicenter study, a parametric analysis resulted in positive lod scores (highest of 1.97 for dominant and 1.19 for recessive inheritance at a theta of 0.20) for locus DXS7, with scores below 0.50 for other markers in this region (MAOB, DXS228, and ARAF1). Significant allele sharing among affected sibling pairs was present at DXS7. In the second study, positive lod scores were observed at MAOB (highest of 2.16 at a theta of 0.05 for dominant and 1.64 at a theta of 0.00 for recessive models) and ALAS2 (the highest of 1.36 at a theta of 0.05 for a recessive model), with significant allele sharing (P = 0.003 and 0.01, respectively) at these two loci. These five markers are mapped within a small region of Xp11. Thus, although substantial regions of the X-chromosome have been investigated without evidence for linkage being found, a locus predisposing to schizophrenia in the proximal short arm of the X-chromosome is not excluded.

Affected-only multiplex pedigree analysis of GAW10 problem 2.

From a single extended pedigree simulation replicate, high density, affected only subpedigrees were isolated, based on the T > 40 affected status for the disease trait, Q1. On this sample of 14 pedigrees, with a range of two to six affected members (48 total), we conducted a haplotype based, multilocus, nonparametric genome-wide search of the provided data (367 markers) using the computer program GENEHUNTER. As with most genome screens in complex diseases, the objective of this strategy was to identify regions (hot-spots) which breached our predetermined threshold (p < 0.05), requiring confirmation by other groups or consortia. Of the six regions with threshold breaching scores (p < 0.05), the most promising, on chromosome 8 and chromosome 4, corresponded to the locations of MG2 and MG3. Both of these regions have multiple, consecutive markers above threshold and contained the only scores that exceeded p < 0.01. In addition, a fourth hot-spot consisting of a single marker above threshold, was less than 15 cM from MG1 on chromosome 5. The positions of the remaining three hot-spots did not correspond to the any of the major genes and are therefore false positives. An additional analysis of a single nuclear pedigree simulation replicate, using the extended transmission disequilibrium test (ETDT), was applied to markers in each of the above hot-spot regions to look for evidence of disequilibrium with the disease trait. This analysis provided weak additional support for the chromosome 8 finding, even though the sample was very small (36 pedigrees containing 44 affected offspring).

Confirmation of susceptibility locus on chromosome 13 in Australian breast cancer families.

Two major genes determining predisposition to breast cancer, termed BRCA1 and BRCA2, have been mapped to the long arms of chromosomes 17 and 13, respectively. Each locus is believed to account for approximately 40% of cases of familial breast cancer. We used linkage and haplotype analysis with simple tandem repeat polymorphisms at chromosomal bands 17q21 and 13q12 to determine the contribution of the BRCA1 and BRCA2 genes to predisposition to breast cancer in four Australian breast cancer kindreds, one of which had two male cousins with breast cancer. Surprisingly all families segregated a haplotype of markers on 13q and showed positive lod scores supporting linkage to BRCA2. In addition, haplotype analysis identified an informative recombination between D13S260 and D13S171 in one affected individual, which refines the localisation of BRCA2 to between D13S260 and D13S267; a distance of 2-3 cM. Tumours of the stomach and cervix, as well as melanoma and leukaemia/lymphoma also occur in these pedigrees but the numbers are too low to determine whether they may be significantly associated with BRCA2 carrier status. Our results confirm the existence of BRCA2 on the long arm of chromosome 13 and support previous findings that this locus is likely to confer risk in families with affected males. Furthermore, our observations suggest that the BRCA2 gene may also contribute to the development of other neoplasma.

Deletion mapping of the short arm of chromosome 3 in Merkel cell carcinoma.

Little is known about the biology of Merkel cell carcinoma (MCC), also called small cell carcinoma of the skin. MCC has similarities with small cell lung cancer (SCLC): both are neuroendocrine malignancies with early metastasis to distant sites and a poor prognosis. Small cell lung cancer biopsies are known to have frequent losses on chromosome 3 in the region 3p21, yet MCCs have not been reported to have 3p deletions by karyotypic analysis. Considering the similarities between SCLC and MCC, we investigated 26 MCC tumours for loss of heterozygosity (LOH) on 3p. First, RFLP analysis was performed using PCR with nine primer sets from six loci. Second, 25 tumours were examined by microsatellite analysis for 3p markers D3S1289 and D3S1285 and SST on 3q. All 26 tumours were informative at one or more loci; of these, 18 (69%) demonstrated LOH for at least one marker on the short arm. For all informative loci the frequency of LOH was greater than 30% (range 33-75%). In a cell line derived from one tumour, it was possible to demonstrate rearrangement of chromosome 3 by in situ hybridisation. No LOH was seen in 15 informative cases for the 3q locus SST. A region 3p13-p21.1, centered on the marker D3S2, was deleted in all tumours demonstrating LOH, with a secondary deletion involving D3S30 detected in some tumours at 3p13. Our results indicate that LOH on 3p is a common occurrence in MCC; however, three tumours for which DNA was also available from a corresponding cell line suggest there may be a subset of MCC whose genesis is independent of deletions of 3p.

The contribution of the DFNB1 locus to neurosensory deafness in a Caucasian population.

Classical studies have demonstrated genetic heterogeneity for nonsyndromic autosomal recessive congenital neurosensory deafness, with at least six loci postulated. Linkage analysis in two consanguineous Tunisian kindreds has demonstrated that one such deafness locus, DFNB1, maps near chromosome 13 markers D13S175, D13S143, and D13S115. We tested these markers for cosegregation with deafness in 18 New Zealand and 1 Australian nonconsanguineous kindreds, each of which included at least two siblings with nonsyndromic presumed congenital sensorineural deafness and that had a pedigree structure consistent with autosomal recessive inheritance. When all families were combined, a peak two-point lod score of 2.547 (theta = .1) was obtained for D13S175, 0.780 (theta = .2) for D13S143, and 0.664 (theta = .3) for D13S115. While there was no statistically significant evidence for heterogeneity at any of the three loci tested, nine families showed cosegregation of marker haplotypes with deafness. These observations suggest that the DFNB1 locus may make an important contribution to autosomal recessive neurosensory deafness in a Caucasian population. In the nine cosegregating families, phenotypic variation was observed both within sibships (in four families), which indicates that variable expressivity characterizes some genotypes at the DFNB1 locus, and between generations (in two families), which suggests allelic heterogeneity.

Linkage analysis in familial melanoma kindreds to markers on chromosome 6p.

Malignant melanoma occurs as a familial cancer in 5%-10% of cases, where it segregates in a manner consistent with autosomal dominant inheritance. Evidence from cytogenetics, fine mapping studies of deletions in melanomas and recent linkage studies supports the location of a human melanoma predisposition gene on the short arm of chromosome 9. Evidence also exists for a melanoma gene on Ip, indicating genetic heterogeneity for melanoma predisposition. Previous studies have also reported findings suggestive of linkage of some melanoma families to the HLA region on the short arm of chromosome 6 (6p), indicating the possibility of even greater heterogeneity. To further define the possible effect of a gene within the HLA region on melanoma susceptibility, we have typed 7 simple tandem repeat polymorphisms (STRPs) from 6p in 16 Australian melanoma kindreds. Maximum 2-point LOD scores ranged from 1.13 (theta = 0.2) to 2.03 (theta = 0.15) for 4 contiguous markers flanking the HLA complex, and multi-point analysis gave a peak LOD score of 1.64, 24 centimorgans telomeric to D6S109. However, extended haplotype analysis of these markers showed that a region between D6S105 and HLAF segregated with melanoma in 5/16 families. These results are surprising given that the same cohort of families has previously been shown to be linked to chromosome 9. One interpretation of the current findings is that melanoma susceptibility in some families may result from a gene mapping within the HLA region of chromosome 6p.

Multiple endocrine neoplasia type 1 (MEN1) in two Asian families.

Multiple endocrine neoplasia type 1 (MEN1), an autosomal dominant disease characterized by neoplasia of the parathyroid glands, anterior pituitary and endocrine pancreas, is rarely reported in Asian populations. The MEN1 gene, mapped to chromosome 11q13 but yet to be cloned, has been found to be homogeneous in Caucasian populations through linkage analysis. Here, two previously unreported Asian kindreds with MEN1 are described; linkage analysis using microsatellite polymorphic markers in the MEN1 region was carried out. The first kindred, of Mongolian-Chinese origin, is a multigeneration family with over 150 living members, eight of whom are affected to date. The second kindred is of Chinese origin consisting of four affected members. Linkage to chromosome 11q13 was confirmed in both kindreds, supporting evidence for genetic homogeneity. A recombination in the larger kindred localizes the gene distal to marker D11S956, consistent with its placement from previous studies. We also show that it is feasible to use these markers for predictive testing, as four gene carriers were detected in 13 family members with unknown disease status in the first kindred.

Simple tandem repeat allelic deletions confirm the preferential loss of distal chromosome 6q in melanoma.

Karyotypic analysis, loss of somatic heterozygosity, microcell fusion and cDNA transfection studies have provided compelling evidence that at least one tumour suppressor gene for melanoma resides on chromosome 6. In an attempt to further define the regions to which these putative suppressor genes map, we have carried out loss of heterozygosity (LOH) studies on DNA from 25 fresh melanoma tumours for 9 simple tandem repeat (STR) polymorphism markers spanning chromosome 6. Four samples displayed LOH or homozygosity for all markers studied, indicating that they had lost one homologue of chromosome 6. An additional 3 samples showed LOH for all markers on 6q. Furthermore, 30 melanoma cell lines, for which there were no matching somatic DNA samples, were analyzed for hemizygosity of markers on 6q. One cell line had a homozygous deletion of all markers tested and a further 12 cell lines displayed only one allele for 3 or 4 contiguous markers, indicating that most, if not all of these samples were hemizygous for the region of 6q distal to D6S87. Overall, the rate of LOH on 6q in the 55 melanoma DNAs was 35%, and there were no losses of markers on 6p without concomitant loss of markers on 6q. Two of 5 samples derived from primary melanomas showed LOH, which indicates that LOH for the melanoma suppressor gene on 6q, which maps to a region that contains the SOD2 locus, is a frequent and early event in melanoma tumorigenesis.

Refined localization of the melanoma (MLM) gene on chromosome 9p by analysis of allelic deletions.

Various lines of evidence including linkage analysis, frequent homozygous and heterozygous deletions in melanoma DNAs, and the finding of a patient with multiple primary melanomas who harbours a 5p/9p translocation involving loss of several 9p markers, have indicated that the 9p22-p13 region harbours a gene important for the development of melanoma (MLM). We have used eight short tandem repeat polymorphism (STRP) markers mapping to this region to look for allelic losses in DNA from melanoma biopsies and cell lines. Heterozygous losses were found in 8/14 (57%) fresh melanoma biopsy DNAs with the smallest region of overlap (SRO) being between IFNA and D9S169. In addition, when DNA from 30 melanoma cell lines was studied, four cell lines (13%) were found to be homozygously deleted for various 9p markers. Two of these cell lines define the borders of overlapping homozygous deletions within a 4cM region of 9p21 between IFNA and D9S171. Moreover, a further 14 melanoma cell lines were hemizygous for the IFNA/D9S171/D9S126 region. These data support the hypothesis that the MLM gene acts as a tumour suppressor, and provide a refinement of its localization on 9p.

Haplotype analysis limits the position of the familial melanoma locus on 9p to the D9S169-D9S156 interval.

A gene for familial melanoma (MLM) has been mapped to 9p22-p13 by linkage analysis using simple tandem repeat polymorphisms (STRPs) at the IFNA and D9S126 loci. This localization is consistent with the finding of homozygous deletions of these markers in DNA from two melanoma cell lines, which suggest that the locus has the properties of a tumour suppressor gene. In an attempt to further define the position of the MLM locus we have typed 10 STRPs from the short arm of chromosome 9 in 15 Australian melanoma kindreds. Extended haplotype analysis of these markers and identification of recombinants in our pedigrees indicate that the MLM gene is flanked on the centromeric side by D9S169 and on the telomeric side by D9S156. These results limit the location of the MLM locus to an interval of about 16 centimorgans.