BRCA carriers after risk-reducing bilateral salpingo-oophorectomy: menopausal hormone therapy knowledge gaps, and the impact of physicians' recommendations.

OBJECTIVE: Female carriers of BRCA1/2 gene mutations are at an increased lifetime risk for breast and ovarian cancers. They are recommended to undergo risk-reducing surgery, including bilateral salpingo-oophorectomy (RR-BSO), upon completion of childbearing. RR-BSO surgery decreases morbidity and mortality but results in early menopause. Menopausal hormone therapy (MHT) is under-utilized despite being shown as safe for carriers. We aim to evaluate the factors associated with decision-making regarding MHT use following RR-BSO in healthy BRCA mutation carriers. METHODS: Female carriers aged <50 years who underwent RR-BSO and were followed in a multidisciplinary clinic completed online multiple-choice and free-text questionnaires. RESULTS: A total of 142 women met the inclusion criteria and filled the questionnaire: 83 were MHT users and 59 were non-users. MHT users underwent RR-BSO earlier than non-users (40.82 ± 3.91 vs. 42.88 ± 4.34; p < 0.0001). MHT usage was positively associated with MHT explanation (odds ratio 4.318, 95% confidence interval [CI] [1.341-13.902], p = 0.014), and knowledge regarding the safety of MHT and its effects on general health (odds ratio 2.001, 95% CI [1.443-2.774], p < 0.0001). MHT users and non-users retrospectively evaluated their comprehension of RR-BSO consequences as significantly lower than before surgery (p < 0.001). CONCLUSION: Post-RR-BSO outcomes, including the effects on women's quality of life and its possible mitigation through MHT use, need to be addressed pre surgery by healthcare providers.

Common founder BRCA2 pathogenic variants and breast cancer characteristics in Ethiopian Jews.

PURPOSE: BRCA1/2 founder pathogenic variants (PVs) occur in various populations, but data on the mutational spectrum in Africans are limited. We examined BRCA1/2 PVs in breast cancer patients of Ethiopian Jewish (EJ) origin. METHODS: We retrospectively analyzed BRCA1/2 test results and clinical features of EJ breast cancer patients from seven medical institutions. We obtained heterozygote carrier rates in affected individuals from the laboratories of the largest Israeli HMO (Clalit). Population carrier frequency was determined in EJ controls. RESULTS: We identified three recurrent BRCA2 PVs in 11 EJ breast cancer patients (9 females, 2 males): c.7579delG, c.5159C > A, and c.9693delA. Only c.5159C > A was previously reported in Africans. In women, mean age at diagnosis was 35.7y; 8/9 were diagnosed with advanced disease. All tumors were invasive, 4/9 were triple negative. Only 3/11 carriers had relevant family history. Carrier rate in high-risk breast cancer patients was 11% (3/28; 95%CI [2.3%, 28.2%]). Combined carrier rate among controls was 1.8% (5/280; 95%CI [0.6%, 4.1%]). CONCLUSION: EJs harbor 3 recurrent BRCA2 PVs presenting with relatively severe breast cancer morbidity. Combined with the high BRCA2 carrier rate in the EJ population, these findings merit increasing awareness in this community and suggest that a culturally adapted population screening approach may be warranted.

Expanding the phenotype of CRB2 mutations - A new ciliopathy syndrome?

Recessive CRB2 mutations were recently reported to cause both steroid resistant nephrotic syndrome and prenatal onset ventriculomegaly with kidney disease. We report two Ashkenazi Jewish siblings clinically diagnosed with ciliopathy. Both presented with severe congenital hydrocephalus and mild urinary tract anomalies. One affected sibling also has lung hypoplasia and heart defects. Exome sequencing and further CRB2 analysis revealed that both siblings are compound heterozygotes for CRB2 mutations p.N800K and p.Gly1036Alafs*43, and heterozygous for a deleterious splice variant in the ciliopathy gene TTCB21. CRB2 is a polarity protein which plays a role in ciliogenesis and ciliary function. Biallelic CRB2 mutations in animal models result in phenotypes consistent with ciliopathy. This report expands the phenotype of CRB2 mutations to include lung hypoplasia and uretero-pelvic renal anomalies, and confirms cardiac malformation as a feature. We suggest that CRB2-associated disease is a new ciliopathy syndrome with possible digenic/triallelic inheritance, as observed in other ciliopathies. Clinically, CRB2 should be assessed when ciliopathy is suspected, especially in Ashkenazi Jews, where we found that p.N800K carrier frequency is 1 of 64. Patients harboring CRB2 mutations should be tested for the complete range of ciliopathy manifestations.

A role for TENM1 mutations in congenital general anosmia.

Congenital general anosmia (CGA) is a neurological disorder entailing a complete innate inability to sense odors. While the mechanisms underlying vertebrate olfaction have been studied in detail, there are still gaps in our understanding of the molecular genetic basis of innate olfactory disorders. Applying whole-exome sequencing to a family multiply affected with CGA, we identified three members with a rare X-linked missense mutation in the TENM1 (teneurin 1) gene (ENST00000422452:c.C4829T). In Drosophila melanogaster, TENM1 functions in synaptic-partner-matching between axons of olfactory sensory neurons and target projection neurons and is involved in synapse organization in the olfactory system. We used CRISPR-Cas9 system to generate a Tenm1 disrupted mouse model. Tenm1(-/-) and point-mutated Tenm1(A) (/A) adult mice were shown to have an altered ability to locate a buried food pellet. Tenm1(A) (/A) mice also displayed an altered ability to sense aversive odors. Results of our study, that describes a new Tenm1 mouse, agree with the hypothesis that TENM1 has a role in olfaction. However, additional studies should be done in larger CGA cohorts, to provide statistical evidence that loss-of-function mutations in TENM1 can solely cause the disease in our and other CGA cases.

Alpha-thalassemia intellectual disability: variable phenotypic expression among males with a recurrent nonsense mutation - c.109C>T (p.R37X).

Alpha-thalassemia intellectual disability, one of the recognizable X-linked disability syndromes, is characterized by short stature, microcephaly, distinctive facies, hypotonic appearance, cardiac and genital anomalies, and marked skewing of X-inactivation in female carriers. With the advent of next generation sequencing, mutations have been identified that result in less severe phenotypes lacking one or more of these phenotypic manifestations. Here we report five unrelated kindreds in which a c.109C>T (p.R37X) mutation segregates with a variable but overall milder phenotype. The distinctive facial appearance of alpha-thalassemia intellectual disability was present in only one of the 18 affected males evaluated beyond the age of puberty, although suggestive facial appearance was present in several during infancy or early childhood. Although the responsible genetic alteration is a nonsense mutation in exon 2 of ATRX, the phenotype appears to be partially rescued by the production of alternative transcripts and/or other molecular mechanisms.

Familial haplotyping and embryo analysis for Preimplantation genetic diagnosis (PGD) using DNA microarrays: a proof of principle study.

PURPOSE: Development of PGD assays for molecular disorders is based on analysis of a familial mutation together with linked polymorphic STR markers; a process which is lengthy and requires the identification of multiple informative markers prior to PGD analysis. On the other hand, whole genome amplification (WGA), in conjunction with microarray platforms, allows the use of a universal assay for the analysis of a very large number of SNP markers at once. The aim of this study was to test high throughput pre-PGD familial haplotyping for in-case blastomere analysis in order to eliminate time-consuming pre-case preparations for each family. METHODS: A PGD cycle was performed for a couple with paternal Charcot Marie Tooth 1A (CMT1A) using a classic multiplex nested PCR approach. Mutant embryos from the case were blindly reanalyzed, as single or multi-cell biopsies, using a multiple displacement amplification-based WGA protocol and microarray SNP analysis. In parallel, relevant genomic DNA samples from the family were also analyzed by SNP microarray. RESULTS: After applying a 'unique informative allele' selection algorithm to the data, this array-based assay reconfirmed the initial diagnosis in all samples. CONCLUSIONS: We describe a PGD method that is both accurate and feasible during the time-frame required for embryo transfer. This strategy greatly reduces the time for pre-case haplotype preparation.

X-linked neonatal diabetes mellitus, enteropathy and endocrinopathy syndrome is the human equivalent of mouse scurfy.

To determine whether human X-linked neonatal diabetes mellitus, enteropathy and endocrinopathy syndrome (IPEX; MIM 304930) is the genetic equivalent of the scurfy (sf) mouse, we sequenced the human ortholog (FOXP3) of the gene mutated in scurfy mice (Foxp3), in IPEX patients. We found four non-polymorphic mutations. Each mutation affects the forkhead/winged-helix domain of the scurfin protein, indicating that the mutations may disrupt critical DNA interactions.

Secreted amyloid beta-protein similar to that in the senile plaques of Alzheimer's disease is increased in vivo by the presenilin 1 and 2 and APP mutations linked to familial Alzheimer's disease.

To determine whether the presenilin 1 (PS1), presenilin 2 (PS2) and amyloid beta-protein precursor (APP) mutations linked to familial Alzheimer's disease (FAD) increase the extracellular concentration of amyloid beta-protein (A beta) ending at A beta 42(43) in vivo, we performed a blinded comparison of plasma A beta levels in carriers of these mutations and controls. A beta 1-42(43) was elevated in plasma from subjects with FAD-linked PS1 (P < 0.0001), PS2N1411 (P = 0.009), APPK670N,M671L (P < 0.0001), and APPV7171 (one subject) mutations. A beta ending at A beta 42(43) was also significantly elevated in fibroblast media from subjects with PS1 (P < 0.0001) or PS2 (P = 0.03) mutations. These findings indicate that the FAD-linked mutations may all cause Alzhelmer's disease by increasing the extracellular concentration of A beta 42(43), thereby fostering cerebral deposition of this highly amyloidogenic peptide.

A familial Alzheimer's disease locus on chromosome 1.

The Volga German kindreds are a group of seven related families with autosomal dominant early-onset Alzheimer's disease (AD). Linkage to known AD-related loci on chromosomes 21 and 14 has been excluded. Significant evidence for linkage to AD in these families was obtained with D1S479 and there was also positive evidence for linkage with other markers in the region. A 112-base pair allele of D1S479 co-segregated with the disease in five of seven families, which is consistent with a common genetic founder. This study demonstrates the presence of an AD locus on chromosome 1q31-42.

Candidate gene for the chromosome 1 familial Alzheimer's disease locus.

A candidate gene for the chromosome 1 Alzheimer's disease (AD) locus was identified (STM2). The predicted amino acid sequence for STM2 is homologous to that of the recently cloned chromosome 14 AD gene (S182). A point mutation in STM2, resulting in the substitution of an isoleucine for an asparagine (N141l), was identified in affected people from Volga German AD kindreds. This N141l mutation occurs at an amino acid residue that is conserved in human S182 and in the mouse S182 homolog. The presence of missense mutations in AD subjects in two highly similar genes strongly supports the hypothesis that mutations in both are pathogenic.

Attitudes of couples identified through screening as carriers of Gaucher disease type 1.

Gaucher disease (GD) type 1 is the most frequent autosomal recessive disorder among Ashkenazi Jews, but because the phenotype is tremendously variable, including it in the 'Ashkenazi Panel' of carrier screening is controversial. As part of a nationwide study conducted in Israel to evaluate the outcomes of carrier screening for GD, we studied the experience of 65/82 (79%) of the couples identified as being at risk for an affected child. We found that pre-test information was regarded as insufficient and improved in post-result counseling. About 70% of the subjects interpreted the genetic counseling as directive, mostly toward prenatal diagnosis (PND) but against pregnancy termination of affected fetuses. We evaluated the various motivations that had led couples to utilize PND. Subjects' attitudes toward pregnancy termination correlated with their specific genotypes, with their perception of the severity of GD and with attending additional medical consultation. Of the 30 interviewed participants who were faced with having an affected fetus, 80% came to terms with their decision to utilize PND, but about half of the few who terminated the pregnancy regret their decision. Despite questionable benefits of screening, most of the participants did not regret having been tested and supported the continuation of this program. We offer explanations for these findings and suggest extensive genetic and medical counseling for any future carrier screening for low penetrance, treatable disease.

Polar body-based preimplantation genetic diagnosis for N-acetylglutamate synthase deficiency.

OBJECTIVE: We describe a sensitive and highly reliable preimplantation genetic diagnosis (PGD) assay for N-acetylglutamate synthetase (NAGS) deficiency using polar body (PB) analysis in conjunction with multiple markers flanking the gene. This rare autosomal recessive mitochondrial disorder is characterized by hyperammonemia, uncontrollable movements, developmental delay, visual impairment, failure to thrive and vomiting and is caused by mutations in the NAGS gene located on chromosome 17q21.31. METHODS: For a family with an affected child we have developed a multiplex fluorescent PCR protocol that included detection of the specific familial mutation (2729insC) in conjunction with the analysis of five informative polymorphic markers flanking the gene: D17S902, D17S965, D17S1861, D17S791 and D17S1868. Following successful amplification in single-cell fibroblasts, this protocol was used in the couple carriers of NAGS mutation. RESULTS: Of 18 retrieved eggs, 16 were at the M2 stage and 9 fertilized. 12 polar body 1s (PB1) were heterozygotes, 1 homozygote wild-type, 1 total amplification failure, and two showed inconclusive results. Three oocytes that had heterozygote PB1s showed mutant polar body 2 (PB2) indicating a wild-type oocyte. Despite the fact that the specific 2729insC mutation did not amplify in the PGD cycle, analysis of linked markers in PBs was sufficient to ensure an accurate diagnosis in 5 out of 9 oocytes. This cycle resulted in the transfer of 3 embryos originating from oocytes diagnosed as wild-type by PB analysis, with the subsequent birth of healthy twin girls. Postnatal genetic testing revealed that both girls harbored the healthy maternal allele and carried the mutant paternal allele. CONCLUSIONS: Our multiplex-nested PCR protocol based on several linked microsatellite markers offers an efficient and accurate method for PGD for NAGS syndrome even when the mutation is not amplified.

Low-dose enzyme replacement therapy for Gaucher's disease: effects of age, sex, genotype, and clinical features on response to treatment.

Although alglucerase therapy has become the treatment of choice for symptomatic patients with Gaucher's disease, the low-dose/high-frequency regimen introduced as a means to reduce the high cost of treatment has raised major controversy. We evaluated the efficacy and safety of low-dose alglucerase in 29 patients with Gaucher's disease who completed 6 to 28 months of therapy. All received intravenous alglucerase at a monthly dose of 30 units/kg, given usually in equal doses 3 times a week. All patients responded well to treatment. The hematological improvement and the reduction in organomegaly were satisfactory. No correlation was found between age, sex, genotype, previous splenectomy, or severity score index and the response to treatment. Patients with a greater degree of hepatomegaly tended to have a more pronounced decrease in liver size, although this reduction did not reach statistical significance. We confirmed that a low-dose/high-frequency regimen of alglucerase was as effective as a high-dose/low-frequency protocol in the treatment of Gaucher's disease, even in the severely ill. Whenever cost is an issue, we recommend using this low-dose regimen.

Neonatal hyperbilirubinemia in glucose-6-phosphate dehydrogenase-deficient heterozygotes.

OBJECTIVES: We assessed the incidence of hyperbilirubinemia, defined as serum total bilirubin >/=15 mg/dL (256 micromol/L), in a cohort of Sephardic Jewish female neonates at risk for glucose-6-phosphate dehydrogenase (G-6-PD) deficiency with especial emphasis on the heterozygotes. We studied the roles of hemolysis by blood carboxyhemoglobin (COHb) determinations and of the variant promoter of the gene for the bilirubin-conjugating enzyme uridine 5'-diphosphate glucuronosyltransferase 1 (UGT1A1) seen in Gilbert's syndrome in the pathogenesis of the hyperbilirubinemia. METHODS: Consecutively born, healthy, term, female neonates were screened for G-6-PD deficiency and observed clinically with serum bilirubin evaluations as indicated for hyperbilirubinemia. On day 3, blood was sampled for COHb, total hemoglobin (tHb), and a mandatory serum bilirubin determination. COHb, determined by gas chromatography, was expressed as percentage of tHb and corrected for inspired carbon monoxide (COHbc). DNA was analyzed for the G-6-PD Mediterranean563T mutation and for the variant UGT1A1 gene. RESULTS: The cohort included 54 G-6-PD-deficient heterozygotes, 19 deficient homozygotes, and 112 normal homozygotes. More heterozygotes (12/54, 22%; relative risk: 2.26; 95% CI: 1.07-4.80) and deficient homozygotes (5/19, 26.3%; relative risk: 2.68; 95% CI: 1.05-6.90) developed hyperbilirubinemia, than did normal homozygotes (11/112, 9.8%). Third-day serum bilirubin values that were obtained from 144 neonates were significantly higher in both heterozygotes (11.2 +/- 3. 7 mg/dL [192 +/- 64 micromol/L]) and G-6-PD-deficient homozygotes (12.0 +/- 3.0 mg/dL [206 +/- 52 micromol/L]) than in the G-6-PD normal homozygotes (9.4 +/- 3.4 mg/dL [160 +/- 58 micromol/L). In contrast, COHbc values were higher only in G-6-PD-deficient homozygotes (0.74% +/- 0.14%) and not in heterozygotes (0.69% +/- 0. 19%, not statistically significant), compared with control values (0. 63% +/- 0.19%). High COHbc values were not a prerequisite for the development of hyperbilirubinemia in any of the G-6-PD genotypes. A greater incidence of hyperbilirubinemia was found among the G-6-PD-deficient heterozygotes, who also had the variant UGT1A1 gene, in both heterozygous (6/20, 30%) and homozygous (4/8, 50%) forms, than was found in their counterparts with the normal UGT1A1 gene (2/26, 7.7%). This effect was not seen in the G-6-PD normal homozygote group. A color reduction screening test for G-6-PD deficiency identified only 20.4% (11/54) of the heterozygotes. CONCLUSIONS: We showed that G-6-PD-deficient heterozygotes, categorically defined by DNA analysis, are at increased risk for neonatal hyperbilirubinemia. The screening test that was used was unable to detect most heterozygotes. Increased bilirubin production was not crucial to the development of hyperbilirubinemia, but presence of the variant UGT1A1 gene did confer increased risk.

Incidence of thrombophilia in patients with Gaucher disease.

An inherited risk for thrombosis, including mutant thermolabile variant of methylenetetrahydrofolate reductase (MTHFR), factor V Leiden, or prothrombin may be the co-factor(s) for avascular necrosis (AVN) in patients with sickle cell disease. Similarly, heterozygosity for factor V Leiden is sufficient to explain the increased blood viscosity observed in children with Legg-Calve-Perthes disease who develop AVN. Because there are no laboratory tests or clinical markers that are helpful in predicting which patients with Gaucher disease may develop AVN, the current study was undertaken to ascertain if there exists an inherited predilection to hypercoagulability in patients with Gaucher disease and AVN. Analysis was performed on genomic DNA extracted from 56 adult patients with type I Gaucher disease. In this cohort of Ashkenazi Jewish patients, the frequency of mutations in the MTHFR, prothrombin, and factor V Leiden genes was found to be low, as was the presence of anticardiolipin antibodies; and none was correlated with increased incidence of AVN. Splenectomy, that may be a predisposing factor to AVN in patients with Gaucher disease, was factored out. Hence the presence of any of the above thrombophilic factors, and which by extension may be risk factors for AVN in other diseases, are not more common in patients with Gaucher disease who develop AVN. Studies in larger cohorts and possibly inclusion of additional factors may be needed to ascertain whether a correlation exists.

BRCA1 germline mutations in women with uterine serous papillary carcinoma.

OBJECTIVE: To determine the possible effects and incidence of BRCA1 and BRCA2 germline mutations in uterine serous papillary carcinoma. METHODS: We screened DNA from 12 women with uterine serous papillary carcinoma for BRCA1 and BRCA2 germline mutations common in the Jewish population (BRCA1-185delAG and 5382insC, BRCA2-6174delT). In women with germline mutations, tumor DNA was screened for loss of heterozygosity at the appropriate loci. RESULTS: Nine women were of Jewish Ashkenazi origin and three were non-Ashkenazi. Two of nine Ashkenazi women were carriers of germline mutations: one 185delAG mutation and one 5382insC mutation. Five women had histories of breast carcinoma before diagnosis of uterine serous papillary carcinoma. Family histories of seven women had at least one first-degree relative with malignant disease. Of those, four had at least one first-degree relative with breast, ovarian, or colon carcinoma. Both carriers had strong family histories of breast-ovarian carcinoma. Loss of heterozygosity analysis found loss of the wild-type BRCA1 allele in the primary uterine tumors. CONCLUSION: BRCA1 germline mutations were observed in two of nine of the women in this series. The loss of heterozygosity in the tumor tissue of the carriers, coupled with the high frequency of family and patient histories of breast or ovarian malignancies, suggest that uterine serous papillary carcinoma might be a manifestation of familial breast-ovarian cancer.

Recent advances in the genetics of Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disorder that is the most common cause of dementia in the elderly. It is a clinical-pathologic entity characterized by progressive dementia associated with the neuropathologic hallmarks of Abeta amyloid plaques, neurofibrillary tangles (NFTs), neuronal loss, and amyloid angiopathy. Three "causative" AD genes (i.e., genes in which a mutation is sufficient to result in clinical AD) for early-onset familial Alzheimer's disease (FAD) and one "susceptibility" gene that affects risk and age of onset of AD in familial and sporadic late-onset AD have been identified. The three causative genes are the amyloid precursor protein (APP gene) on chromosome 21, the presenilin-1 gene on chromosome 14, and the presenilin-2 gene on chromosome 1. The susceptibility gene is the apolipoprotein E (APOE) gene on chromosome 19. Investigations of the normal and aberrant function of these genes will provide insights into the mechanisms underlying AD and will suggest new strategies for therapeutic intervention.

The founder mutations in the BRCA1, BRCA2, and ATM genes in Moroccan Jewish women with breast cancer.

To gain insight into the molecular mechanisms underlying the inherited predisposition to breast cancer in non-Ashkenazi Jews, we genotyped 54 Jewish Moroccan women with breast cancer, unselected for family history of cancer, for the predominant Jewish mutations in BRCA1, BRCA2, and ATM. One patient (2%) was found to have the 185de1AG BRCA1 mutation, none was a carrier of the 6174delT BRCA2 mutation, and 2/54 (4%) were heterozygous for the ATM mutation. These rates were not significantly different from the rates in the general non-Ashkenazi population. These preliminary data may indicate that the predominant Jewish mutations in BRCA1, BRCA2, and ATM genes contribute little, if any, to breast cancer predisposition and risk among Moroccan Jews.

[Combination of heterozygote blood coagulation factor V Leiden carriage with erythrocytosis as a cause of the hip deep vein thrombosis]. / Sochetanie geterozigotnogo nositel'stva leidenskoi mutatsii faktora svertyvaniia krovi s éritrotsitozom kak prichina tromboza glubokikh ven bedra.

AIM: To reject one of the variants of inherited thrombophylia in a 64-year-old patient with deep thrombosis of leg veins and high hemoglobin and red cell levels. MATERIAL AND METHODS: The study was made of antithrombin III and protein C, protein S levels; resistance to activated protein C; molecular structure of DNA coding factor 5; methylenetetrahydrofolate reductase. RESULTS: The patient was diagnosed to have heterozygote factor V Leiden mutation. The replacement of arginine by glutamine in position 506 of factor V molecule raises the risk of thrombosis. This risk was aggravated by high hemoglobin, red cells, hematocrit, low volume of circulating plasma, smoking. The patient had normal levels of leukocytes and platelets, normal spleen size, slightly lowered level of erythropoietin. CONCLUSION: The presence of thrombosis in patients with erythremia or erythrocytosis rejects one of the thrombophilia forms.