Mutation analysis of MEN1, HRPT2, CASR, CDKN1B, and AIP genes in primary hyperparathyroidism patients with features of genetic predisposition.

OBJECTIVE: Primary hyperparathyroidism (PHPT), a common endocrine condition, is usually caused by sporadically occurring parathyroid adenoma. A subset of patients carry germline mutations in genes such as MEN1 (multiple endocrine neoplasia type 1), HRPT2 (hyperparathyroidism 2), and CASR (calcium-sensing receptor) predisposing to syndromic forms of PHPT or familial isolated hyperparathyroidism (FIHP). Recently, germline mutations in two novel genes AIP (aryl hydrocarbon receptor-interacting protein) and CDKN1B (cyclin-dependent kinase inhibitor 1B) have been found to be associated with endocrine tumors. The purpose of this study was to evaluate the role of MEN1, HRPT2, CASR, AIP, and CDKN1B genes in PHPT patients with clinical features suggestive of genetic predisposition. PATIENTS AND DESIGN: Medical records of patients treated for PHPT from 1974 to 2001 at Oulu University Hospital were reviewed. Patients with multiglandular or recurrent/persistent disease, other MEN1- related manifestations, aged 40 yr or younger at onset or with a family history of PHPT/MEN1-related tumor were invited to the study. Twenty patients with previously diagnosed MEN1 were excluded. Participants were interviewed and blood samples obtained for biochemical screening and mutation analysis of MEN1, HRPT2, CASR, AIP, and CDKN1B. RESULTS: Of the 56 invited patients, 29 took part in the study. One patient was found to carry the c. 1356_1367del12 MEN1 founder mutation. Mutations in other genes were not detected. CONCLUSIONS: Apart from MEN1, mutations in other genes predisposing to PHPT seem to be rare or non-existing in Northern Finnish PHPT patients. No evidence was found for a role of AIP or CDKN1B in PHPT predisposition.

Multiple endocrine neoplasia type 1 in Northern Finland; clinical features and genotype phenotype correlation.

OBJECTIVE: The existence of genotype-phenotype correlation in multiple endocrine neoplasia type 1 (MEN1) is controversial. Two founder mutations of the MEN1 gene in Northern Finland gave us an opportunity to compare clinical features among heterozygotes of different mutations. DESIGN AND METHODS: Study cohort included 82 MEN1 heterozygotes who were tested for MEN1 during the years 1982-2001. Medical records were reviewed for manifestations of MEN1, other tumours and cause of death by the end of August 2003. Logistic regression analysis was used in evaluating the impact of age, gender and mutational status of affected heterozygotes on the likelihood of developing manifestations of MEN1. RESULTS: Founder mutations 1466del12 and 1657insC were found in 39 and 29 individuals, and D418N, G156R and R527X mutations in 9, 3 and 2 individuals respectively. Except for pituitary adenoma and nonfunctional pancreatic tumour (NFPT), age was a risk factor for all the disease manifestations. For NFPT, frameshift/nonsense mutations (1657insC, R527X) gave an odds ratio (OR) of 3.26 (95% confidence intervals (CI), 1.27-8.33; P = 0.014) compared with in-frame/missense mutations (1466del12, D418N, G156R); including the founder mutation carriers (n = 68) only, the 1657insC mutation gave an OR of 3.56 (CI, 1.29-9.83; P = 0.015). For gastrinoma, in-frame/missense mutations predicted the risk with an OR of 6.77 (CI, 1.31-35.0; P = 0.022), and in the founder mutations group the 1466del12 mutation gave an OR of 15.09 (CI, 1.73-131.9, P = 0.014). CONCLUSIONS: In this study population, NFPT was more common in the frameshift/nonsense or 1657insC mutation carriers, whereas gastrinoma was more common in the in-frame/missense or 1466del12 mutation carriers.

Effect of multiple endocrine neoplasia type 1 (MEN1) gene mutations on premature mortality in familial MEN1 syndrome with founder mutations.

Estimation of mortality and the natural course of a disease is usually based on information of carefully studied individuals with or at risk for a specific disease. Genealogical information has rarely been accurate enough for such studies. With the help of church records and multiple endocrine neoplasia type 1 (MEN1) family information of the two founder MEN1 mutations in Northern Finland (1466del12 and 1657insC), we could trace back common ancestors born in the beginning of the 1700s (1466del12) and approximately 1850 (1657insC) and find 67 probable gene carriers born between 1728 and 1929, which were identified among their offspring. Information was gathered from 34 obligatory MEN1 gene carriers and 31 spouses. The mean age (+/- sd) of death of affected males (n = 16) was 61.1 +/- 12.0 yr vs. 65.8 +/- 15.3 yr for unaffected males (n = 16) and for affected females (n = 16) was 67.2 +/- 10.7 yr vs. 67.7 +/- 14.7 yr for unaffected females (n = 13). The ages of death of the obligatory heterozygotes did not differ from that of the spouses in sex groups or from the sex-matched life expectancy estimates derived from Finnish national statistics. Causes of death differed significantly between female probands and spouses. In conclusion, obligatory MEN1 gene carrier status did not show a harmful effect on survival in this retrospective analysis tracing back to almost 300 yr.

An unexpected recurrence of Angelman syndrome suggestive of maternal germ-line mosaicism of del(15)(q11q13) in a Finnish family.

Angelman syndrome is a neuro-developmental disorder caused by genetic abnormalities affecting the maternal gene expression in the chromosome region 15q11-q13. In a study group of 45 Finnish Angelman patients, a recurrence of a del(15)(q11q13) was detected in one family. The mother's chromosomes 15 were structurally normal, whereas the patients and their unaffected brother shared an identical maternally derived haplotype outside the deletion region. These findings are suggestive of maternal germ-line mosaicism of del(15)(q11q13).

Identification of the familial cylindromatosis tumour-suppressor gene.

Familial cylindromatosis is an autosomal dominant genetic predisposition to multiple tumours of the skin appendages. The susceptibility gene (CYLD) has previously been localized to chromosome 16q and has the genetic attributes of a tumour-suppressor gene (recessive oncogene). Here we have identified CYLD by detecting germline mutations in 21 cylindromatosis families and somatic mutations in 1 sporadic and 5 familial cylindromas. All mutations predict truncation or absence of the encoded protein. CYLD encodes three cytoskeletal-associated-protein-glycine-conserved (CAP-GLY) domains, which are found in proteins that coordinate the attachment of organelles to microtubules. CYLD also has sequence homology to the catalytic domain of ubiquitin carboxy-terminal hydrolases (UCH).

Deletion including the oligophrenin-1 gene associated with enlarged cerebral ventricles, cerebellar hypoplasia, seizures and ataxia.

Non-specific X-linked mental retardation is a heterogeneous group of disorders with an incidence of approximately 1 in 500 males. A recently identified gene in Xq12, encoding a Rho-GTPase-activating protein, was found to be mutated in individuals with mental retardation. We describe here two sisters with a 46,XY karyotype and a microdeletion of the oligophrenin-1 gene and 1.1 Mb of flanking DNA. We have characterised the molecular interval defining this microdeletion syndrome with the fibre-FISH technique. A visual physical map of 1.2 Mb was constructed which spans the oligophrenin-1 gene and the androgen receptor gene. The analysis of the patients revealed a deletion which extended from the 5' end of the AR gene to a region approximately 80 kb proximal to the EPLG2 gene. The clinical manifestations of the two sisters include psychomotor retardation, seizures, ataxia, hypotonia and complete androgen insensitivity. Cranial MRI scans show enlargement of the cerebral ventricles and cerebellar hypoplasia. Our findings give further support for the involvement of the oligophrenin-1 gene in specific morphological abnormalities of the brain which is of importance in the investigation of male patients presenting with mental retardation. In combination with our results from physical mapping we suggest that a region around the oligophrenin-1 locus is relatively bereft of vital genes.

Founder mutations and the high prevalence of myotonia congenita in northern Finland.

OBJECTIVE AND BACKGROUND: To find an explanation at the molecular level for the high prevalence of myotonia congenita in northern Finland and the exceptional pattern of inheritance of the disease in many families, and to study genotype-phenotype correlation in the patients. METHODS: Forty-six patients with myotonia congenita and 16 unaffected relatives from 24 families were studied. All 23 exons and their flanking regions of the gene for the chloride channel protein (ClC-1) were sequenced from at least one patient from all families. RESULTS: There were three different mutations of ClC-1 in the patients: one in exon 11, a T-to-G transversion that resulted in the substitution of cysteine for phenylalanine at amino acid position 413 (F413C); one in exon 15, a C-to-T transition that resulted in the substitution of valine for alanine at amino acid position 531 (A531V); and one in exon 23, a C-to-T transition that resulted in the substitution of a stop codon for an arginine codon at amino acid position 894 (R894X). CONCLUSIONS: Molecular studies showed that even in families with apparent dominant inheritance, the actual mode of inheritance was autosomal recessive. This was explained not only by the observed consanguinity in some families but by an enrichment of three different mutations of the ClC-1 gene and a consequent high number of compound heterozygotes in the population. One of the mutations is unique to northern Finland. The conspicuous enrichment of the mutations is likely due to the founder effect and isolation by distance, as in other diseases in the Finnish heritage.

Germ-line TP53 mutations in Finnish cancer families exhibiting features of the Li-Fraumeni syndrome and negative for BRCA1 and BRCA2.

Mutations in BRCA1 and BRCA2 account for a large portion of the inherited predisposition to breast and ovarian cancer. It was recently discovered that mutations in these two genes are less common in the Finnish population than expected. Because the genetic background of breast cancer, in particular, is largely obscure, it became necessary to search for mutations in other susceptibility genes. Because seven of our BRCA1 and BRCA2 mutation-negative families fulfilled the criteria of either Li-Fraumeni syndrome (LFS) or Li-Fraumeni-like syndrome (LFL), we decided to screen them for germ-line TP53 mutations in exons 5-8 using a dual-temperature single-strand conformation polymorphism assay (SSCP). Two missense mutations (Asn235Ser and Tyr220Cys) were identified. The clinical significance of these findings was evaluated by comparison to previously reported germ-line TP53 mutation data, and by using the tumor loss of heterozygosity (LOH) analysis. In addition, an immunohistochemical analysis of tumor specimens from mutation-positive individuals was performed. Our results suggest that the observed missense mutations confer susceptibility to cancer, and that germ-line TP53 mutations would therefore explain an additional fraction of hereditary breast cancer in Finland.

Prenatal diagnosis of non-ketotic hyperglycinaemia: enzymatic diagnosis in 28 families and DNA diagnosis detecting prevalent Finnish and Israeli-Arab mutations.

Prenatal diagnosis for non-ketotic hyperglycinaemia (NKH) was performed by enzymatic analysis of chorionic villus samples in 28 families and by DNA analysis in two families. In 26 families, enzymatic analysis of the glycine cleavage multi-enzyme system (GCS) yielded an unambiguous diagnosis; inconclusive results in two families were due to borderline GCS activity. We analysed a second chorionic sample in these two families. In one case, GCS activity was normal in the second specimen, and the baby did not have NKH. In the other case, we again found extremely low GCS activity in the second specimen, but a healthy baby was born. The cause of this false-positive result is unknown. Molecular analysis of NKH has identified two prevalent mutations to date; the S564I mutation in a gene encoding the P-protein, a component of the GCS, in a Finnish population, and the H42R mutation in a gene encoding the T-protein in the Israeli-Arab population. These prevalent mutations allow us to obtain unambiguous prenatal diagnoses in both Finnish and Israeli-Arab families. GCS activity in samples from a Finnish family demonstrated a good agreement with DNA analysis, but the fetus of the Israeli-Arab family had an upper limit activity of the affected range, suggesting an advantages for DNA analysis.

Rapid FMR1-protein analysis of fetal blood: an enhancement of prenatal diagnostics.

Fragile-X syndrome, a frequent cause of inherited mental retardation, is characterised in almost all cases by a CGG-repeat expansion that is located within the FMR-1 gene and that prevents the expression of fragile-X mental retardation protein (FMRP). We describe a test that simultaneously allows the rapid detection of FMRP in fetal lymphocytes and distinguishes these from fetal erythrocytes. Routine molecular genetic methods fail in the rare cases where protein expression is blocked, although there is no repeat expansion. Furthermore, they are unsuitable in cases of advanced pregnancy. Our test proves extremely valuable under both these circumstances.

Comparative genomic hybridization studies in tumours from a patient with multiple endocrine neoplasia type 1.

OBJECTIVE: To identify genetic changes, other than the MEN1 gene, that might be involved in the tumorigenesis and progression of multiple endocrine neoplasia type 1 (MEN1)-related tumours. METHODS: We used comparative genomic hybridization (CGH) and loss of heterozygosity (LOH) to study tumours from various sites in a patient with MEN1. RESULTS: Gain of genetic material was not found. Frequent losses of genetic material were found in chromosomes 1, 4, 5, 6, 9, 11 and 18. Besides the chromosome 11 where the MEN1 gene is located, the other regions are known to harbour important tumour suppressor genes. CONCLUSIONS: These results suggest the involvement of other cancer-related genes in the tumorigenesis and progression of MEN1 tumours that warrant further investigations.

Epidemiology of moderate to profound childhood hearing impairments in northern Finland. Any changes in ten years?

The epidemiology of childhood hearing impairments was studied in a clinical series of the birth cohorts for 1973-82 and 1983-92 from a geographically well-defined area. The overall prevalence of hearing impairments with PTA0.5-4 kHz > or = 40 dB was 1.2/1000 live births. Even in the later cohort, the median age ascertainment was 2.6 years for the congenital or early acquired group and 3.0 years for all impairments. Risk indicators for hearing impairment were present in half of the children, and those referred for the risk were ascertained at the median age of 1.1 years. The delay from suspicion to ascertainment was over a year in about one fifth of cases, while the median time from referral to ascertainment was only 1.8 months. The planning and implementation of neonatal hearing screening are suggested, and general information is given on paedoaudiology for both parents and professionals. A database could be beneficial in increasing the systemacy of the diagnostic process.

New lethal disease involving type I and III collagen defect resembling geroderma osteodysplastica, De Barsy syndrome, and Ehlers-Danlos syndrome IV.

We describe the clinical findings and biochemical features of a male child suffering from a so far undescribed lethal connective tissue disorder characterised by extreme hypermobility of the joints, lax skin, cataracts, severe growth retardation, and insufficient production of type I and type III procollagens. His features are compared with Ehlers-Danlos type IV, De Barsy syndrome, and geroderma osteodysplastica, as these disorders show some symptoms and signs shared with our patient. The child died because of failure of the connective tissue structures joining the skull and the spine, leading to progressive spinal stenosis. The aortic valve was translucent and insufficient. The clinical symptoms and signs, together with histological findings, suggested a collagen defect. Studies on both skin fibroblast cultures and the patient's serum showed reduced synthesis of collagen types I and III at the protein and RNA levels. The sizes of the mRNAs and newly synthesised proteins were normal, excluding gross structural abnormalities. These findings are not in accordance with any other collagen defect characterised so far.

Myotonia congenita in northern Finland: an epidemiological and genetic study.

An epidemiological and genetic investigation of myotonia congenita was carried out in northern Finland. Altogether 58 patients were identified (of whom 54 lived in the study area) in 23 families, with a prevalence of 7.3 per 100000. The majority of the families originated from a sparsely populated area in western Lapland. The mean age at onset of the disease was 11 years with a range of 2 to 45 years. The mean time that had passed before verification of the clinical disease was 18 (SD 14) years. The sex ratio M/F was 2.2/1.0. Forty-seven cases were familial and 11 were sporadic. In six families/pedigrees the inheritance was compatible with autosomal recessive and in two families with autosomal dominant inheritance. In five additional families, in which autosomal recessive inheritance seemed most plausible, vertical transmission was also noticed. This could be explained either by consanguinity of the parents or by variant expression of the mutation(s) involved. Our results suggest that myotonia congenita is unusually frequent in northern Finland, most probably as a consequence of an enrichment of the gene mutation(s) in the population.

Improved prenatal diagnosis of the congenital nephrotic syndrome of the Finnish type based on DNA analysis.

Haplotype analysis and alpha-fetoprotein quantitation comprise a prenatal diagnosis of congenital nephrosis. Congenital nephrotic syndrome of the Finnish type (CNF) is an autosomal recessive disease characterized by massive proteinuria and nephrotic syndrome from birth. Prenatal diagnosis of CNF has previously been based on the quantitation of alpha-fetoprotein (AFP) in the amniotic fluid and maternal serum, but an increased AFP is not specific for the disease. We have recently localized the CNF gene to the chromosome 19q13.1 region and observed a strong linkage disequilibrium to the genetic markers D19S610, D19S608, D19S224 and D19S220 in this chromosomal area. Four main CNF-haplotypes have been observed in Finnish kindreds. In the present study, linkage and haplotype analyses have been applied to prenatal diagnosis of six families with a history of CNF. The results diminish the risk of false positive diagnosis and abortions of healthy fetuses in families at risk.

Hereditary spinal neurofibromatosis: a rare form of NF1?

We describe a family in which seven members in three generations were affected with a rare spinal neurofibromatosis. The affected adults showed, at the ages of 32, 37, 38, and 61, respectively, multiple spinal neurofibromas symmetrically affecting all spinal roots. Two patients were operated on for histopathologically proven cervical spinal neurofibromas. All patients had café au lait spots, one had several freckles in the axillary area, and two had possible dermal neurofibromas, but iris Lisch-nodules were not present. Other signs of neurofibromatosis types 1 and 2 were absent. A linkage study of the family suggested close linkage to the NF1 locus and excluded it from the NF2 locus. The DNA analysis of histopathologically verified spinal neurofibromas in two patients showed no evidence of LOH at 17q11.2. The findings in the present family, together with those in a family previously described, suggest a clinically distinct form of neurofibromatosis with extensive spinal neurofibromas and café au lait macules, which may be allelic to the NF1 gene.

Characterization of the full fragile X syndrome mutation in fetal gametes.

Fragile X syndrome results from the expansion of the CGG repeat in the FMR1 gene. Expansion has been suggested to be a postzygotic event with the germline protected. From an analysis of intact ovaries of full mutation fetuses, we now show that only full expansion alleles can be detected in oocytes (but in the unmethylated state). Similarly, the testes of a 13-week full mutation fetus show no evidence of premutations while a 17-week full mutation fetus exhibits some germ cells with attributes of premutations. These data discount the hypothesis that the germline is protected from full expansion and suggest full mutation contraction in the immature testis. Thus, full expansion may already exist in the maternal oocyte, or postzygotic expansion, if it occurs, arises quite early in development prior to germline segregation.