A spectrum of novel NPHS1 and NPHS2 gene mutations in pediatric nephrotic syndrome patients from Pakistan

Background

Mutations in the NPHS1 and NPHS2 genes are among the main causes of early-onset and familial steroid resistant nephrotic syndrome respectively. This study was carried out to assess the frequencies of mutations in these two genes in a cohort of Pakistani pediatric NS patients.

Methods

Mutation analysis was carried out by direct sequencing of the NPHS1 and NPHS2 genes in 145 nephrotic syndrome (NS) patients. This cohort included 36 samples of congenital or infantile onset NS cases and 39 samples of familial cases obtained from 30 families.

Results

A total of 7 homozygous (6 novel) mutations were found in the NPHS1 gene and 4 homozygous mutations in the NPHS2 gene. All mutations in the NPHS1 gene were found in the early onset cases. Of these, one patient has a family history of NS. Homozygous p.R229Q mutation in the NPHS2 gene was found in two children with childhood-onset NS.

Conclusions

Our results show a low prevalence of disease causing mutations in the NPHS1 (22% early onset, 5.5% overall) and NPHS2 (3.3% early onset and 3.4% overall) genes in the Pakistani NS children as compared to the European populations. In contrast to the high frequency of the NPHS2 gene mutations reported for familial SRNS in Europe, no mutation was found in the familial Pakistani cases. To our knowledge, this is the first comprehensive screening of the NPHS1 and NPHS2 gene mutations in sporadic and familial NS cases from South Asia.     

Spectrum of NIPBL gene mutations in Polish patients with Cornelia de Lange syndrome

Cornelia de Lange syndrome (CdLS) is a rare multi-system genetic disorder characterised by growth and developmental delay, distinctive facial dysmorphism, limb malformations and multiple organ defects. The disease is caused by mutations in genes responsible for the formation and regulation of cohesin complex. About half of the cases result from mutations in the NIPBL gene coding delangin, a protein regulating the initialisation of cohesion. To date, approximately 250 point mutations have been identified in more than 300 CdLS patients worldwide. In the present study, conducted on a group of 64 unrelated Polish CdLS patients, 25 various NIPBL sequence variants, including 22 novel point mutations, were detected. Additionally, large genomic deletions on chromosome 5p13 encompassing the NIPBL gene locus were detected in two patients with the most severe CdLS phenotype. Taken together, 42 % of patients were found to have a deleterious alteration affecting the NIPBL gene, by and large private ones (89 %). The review of the types of mutations found so far in Polish patients, their frequency and correlation with the severity of the observed phenotype shows that Polish CdLS cases do not significantly differ from other populations.     

Structure of the gene for congenital nephrotic syndrome of the finnish type (NPHS1) and characterization of mutations.

Congenital nephrotic syndrome of the Finnish type (NPHS1) is an autosomal recessive disorder that is caused by mutations in the recently discovered nephrin gene, NPHS1 ({"type":"entrez-nucleotide","attrs":{"text":"AF035835","term_id":"3025698","term_text":"AF035835"}}AF035835). The disease, which belongs to the Finnish disease heritage, exists predominantly in Finland, but many cases have been observed elsewhere in Europe and North America. The nephrin gene consists of 29 exons spanning 26 kb in the chromosomal region 19q13.1. In the present study, the genomic structure of the nephrin gene was analyzed, and 35 NPHS1 patients were screened for the presence of mutations in the gene. A total of 32 novel mutations, including deletions; insertions; nonsense, missense, and splicing mutations; and two common polymorphisms were found. Only two Swedish and four Finnish patients had the typical Finnish mutations: a 2-bp deletion in exon 2 (Finmajor) or a nonsense mutation in exon 26 (Finminor). In seven cases, no mutations were found in the coding region of the NPHS1 gene or in the immediate 5''-flanking region. These patients may have mutations elsewhere in the promoter, in intron areas, or in a gene encoding another protein that interacts with nephrin.     

Mutations in NPHS1 in a Chinese child with congenital nephrotic syndrome

Since the identification of the NPHS1 gene, which encodes nephrin, various investigators have demonstrated that the NPHS1 mutation is a frequent cause of congenital nephrotic syndrome (CNS); it is found in 98% of Finnish children with this syndrome and in 39-80% of non-Finnish cases. In China, compound heterozygous mutations in the NPHS1 gene have been identified in two Chinese families with CNS. To our knowledge, however, whether or not NPHS1 is the causative gene in sporadic Chinese CNS cases has not been established. We identified a homozygous mutation of NPHS1, 3250insG (V1084fsX1095), in a Chinese child with sporadic CNS. This finding leads us to suggest that NPHS1 mutations are also present in sporadic Chinese CNS cases. This gives additional support for the necessity for genetic examination of mutations in the NPHS1 gene in Chinese children with sporadic CNS.     

Two novel NPHS1 mutations in a Chinese family with congenital nephrotic syndrome

Congenital nephrotic syndrome of the Finnish type (CNF) is a lethal, autosomal recessive disorder mainly caused by mutations in the NPHS1 gene; it is found at a relatively high frequency in Finns. We investigated the disease-causing mutations in a Chinese family with CNF and developed a prenatal genetic diagnosis for their latest pregnancy. Mutation analysis was made of all exons and exon/intron boundaries of NPHS1 in the fetus, parents and 50 unrelated controls using PCR and direct sequencing. A heterozygous nonsense mutation within exon 20 (c.2783C>A) and a missense mutation within exon 17 (c.2225T>C) in NPHS1 were detected in the proband's father and mother, respectively, but were not found in the fetus or in 50 unrelated controls. Two novel mutations of c.2783C>A and c.2225T>C in NPHS1 were found to be causative in this Chinese CNF family with no known Finnish ancestry. The most recent sibling did not inherit these two mutations and hence was unaffected with CNF. Determining the cumulative number and ethnic distribution of known mutations can help expedite further study of the pathogenesis of CNF.     

Mutational screening of EXT1 and EXT2 genes in Polish patients with hereditary multiple exostoses

Hereditary multiple exostoses (HME) also known as multiple osteochondromas represent one of the most frequent bone tumor disorder in humans. Its clinical presentation is characterized by the presence of multiple benign cartilage-capped tumors located most commonly in the juxta-epiphyseal portions of long bones. HME are usually inherited in autosomal dominant manner, however de novo mutations can also occur. In most patients, the disease is caused by alterations in the EXT1 and EXT2 genes. In this study we investigated 33 unrelated Polish probands with the clinical and radiological diagnosis of HME by means of Sanger sequencing and MLPA for all coding exons of EXT1 and EXT2. We demonstrated EXT1 and EXT2 heterozygous mutations in 18 (54.6 %) and ten (30.3 %) probands respectively, which represents a total of 28 (84.9 %) index cases. Sequencing allowed for the detection of causative changes in 26 (78.8 %) probands, whereas MLPA showed intragenic deletions in two (6.1 %) further cases (15 mutations represented novel changes). Our paper is the first report on the results of exhaustive mutational screening of both EXT1/EXT2 genes in Polish patients. The proportion of EXT1/EXT2 mutations in our group was similar to other Caucasian cohorts. However, we found that EXT1 lesions in Polish patients cluster in exons 1 and 2 (55.6 % of all EXT1 mutations). This important finding should lead to the optimization of cost-effectiveness rate of HME diagnostic testing. Therefore, the diagnostic algorithm for HME should include EXT1 sequencing (starting with exons 1–2), followed by EXT2 sequencing, and MLPA/qPCR for intragenic copy number changes.     

Toll-like receptor 4 mediates vascular remodeling in hyperhomocysteinemia

Nephrotic syndrome (NS) is a kidney disease predominantly present in children with idiopathic condition; final stage of the disease progresses into end-stage renal disease. Generally, NS is treated using standard steroid therapy, however; most of the children are steroid sensitive and about 15–20% are non-responders (SRNS). Non-responsiveness of these children would be a risk with the possibility of mutational changes in podocyte genes (NPHS1, NPHS2, WT1, PLCE1). The mutation in podocyte genes is associated with SRNS. NPHS1, NPHS2, and WT1 genes are identified/directly linked to SRNS. The present study is a surveillance on the mutation analysis of WT1 (exons 8 and 9) and NPHS2 (exons 1–8) gene in SRNS followed by clinical management. In the present study, we analyzed these two genes in a total of 117 SRNS (73 boys and 44 girls) children. A total of five mutations were detected in six children. First, WT1 mutation was detected at 9th intron-IVS 9 + 4C > T position in one SRNS female patient. This WT1 mutation was identified in a girl having Frasier Syndrome (FS) with focal segmental glomerulosclerosis and a complete sex reversal found through molecular and karyological screening. In NPHS2, missense mutations of P20L (in two children), P316S, and p.R229Q, and a frame shift mutation of 42delG were detected. Thus, applying molecular investigation helped us to decide on treatment plan of SRNS patients, mainly to avoid unnecessary immunosuppressive treatment.     

Low Prevalence of CHEK2 Gene Mutations in Multiethnic Cohorts of Breast Cancer Patients in Malaysia

CHEK2 is a protein kinase that is involved in cell-cycle checkpoint control after DNA damage. Germline mutations in CHEK2 gene have been associated with increase in breast cancer risk. The aim of this study is to identify the CHEK2 gene germline mutations among high-risk breast cancer patients and its contribution to the multiethnic population in Malaysia. We screened the entire coding region of CHEK2 gene on 59 high-risk breast cancer patients who tested negative for BRCA1/2 germline mutations from UKM Medical Centre (UKMMC), Hospital Kuala Lumpur (HKL) and Hospital Putrajaya (HPJ). Sequence variants identified were screened further in case-control cohorts consisting of 878 unselected invasive breast cancer patients (180 Malays, 526 Chinese and 172 Indian) and 270 healthy individuals (90 Malays, 90 Chinese and 90 Indian). By screening the entire coding region of the CHEK2 gene, two missense mutations, c.480A>G (p.I160M) and c.538C>T (p.R180C) were identified in two unrelated patients (3.4%). Further screening of these missense mutations on the case-control cohorts unveiled the variant p.I160M in 2/172 (1.1%) Indian cases and 1/90 (1.1%) Indian control, variant p.R180C in 2/526 (0.38%) Chinese cases and 0/90 Chinese control, and in 2/180 (1.1%) of Malay cases and 1/90 (1.1%) of Malay control. The results of this study suggest that CHEK2 mutations are rare among high-risk breast cancer patients and may play a minor contributing role in breast carcinogenesis among Malaysian population.     

Plasma membrane targeting of podocin through the classical exocytic pathway: effect of NPHS2 mutations

Podocytes are specialized epithelial cells of the glomerulus in the kidney, which interconnect at the top of the glomerular basement membrane through the slit diaphragm, an adherens-like junction that plays a crucial role in the glomerular filtration process. Podocin, a plasma membrane anchored stomatin-like protein, is expressed in lipid rafts at the insertion of the slit diaphragm in podocytes. Mutations in NPHS2 , the gene encoding podocin, are associated with inherited and sporadic cases of steroid-resistant nephrotic syndrome. Here, we show that brefeldin A induces accumulation of newly synthesized podocin in the endoplasmic reticulum, suggesting that podocin biosynthesis follows the classical secretory pathway, and we study the effect of 12 NPHS2 mutations associated with steroid-resistant nephrotic syndrome on the trafficking of the protein. We found that 9 podocin mutants were not targeted to the plasma membrane, 8 being retained in the endoplasmic reticulum and one being localized in late endosomes. Furthermore, by screening our database of patients with NPHS2 mutations, we found that podocin mutants retained in the endoplasmic reticulum are associated with earlier onset of the disease than those correctly targeted to the cell membrane. Our data suggest that most of NPHS2 mutations lead to retention of podocin in the endoplasmic reticulum and therefore provide a rationale for devising therapeutic approaches aimed at correcting the protein processing defect.     

Spectrum of <Emphasis Type="Italic">JAG1</Emphasis> gene mutations in Polish patients with Alagille syndrome

Alagille syndrome (ALGS) is an autosomal dominant disorder characterized by developmental abnormalities in several organs including the liver, heart, eyes, vertebrae, kidneys, and face. The majority (90-94 %) of ALGS cases are caused by mutations in the JAG1 (JAGGED1) gene, and in a small percent of patients (～1 %) mutations in the NOTCH2 gene have been described. Both genes are involved in the Notch signaling pathway. To date, over 440 different JAG1 gene mutations and ten NOTCH2 mutations have been identified in ALGS patients. The present study was conducted on a group of 35 Polish ALGS patients and revealed JAG1 gene mutations in 26 of them. Twenty-three different mutations were detected including 13 novel point mutations and six large deletions affecting the JAG1 gene. Review of all mutations identified to date in individuals from Poland allowed us to propose an effective diagnostic strategy based on the mutations identified in the reported patients of Polish descent. However, the distribution of mutations seen in this cohort was not substantively different than the mutation distribution in other reported populations.     

Novel Mutations in FKBP10 and PLOD2 Cause Rare Bruck Syndrome in Chinese Patients

Bruck syndrome (BS) is an extremely rare form of osteogenesis imperfecta characterized by congenital joint contracture, multiple fractures and short stature. We described the phenotypes of BS in two Chinese patients for the first time. The novel compound heterozygous mutations c.764_772dupACGTCCTCC (p.255_257dupHisValLeu) in exon 5 and c.1405G>T (p.Gly469X) in exon 9 of FKBP10 were identified in one proband. The novel compound heterozygous mutations c.1624delT (p.Tyr542Thrfs*18) in exon 14 and c.1880T>C (p.Val627Ala) in exon 17 of PLOD2 were identified in another probrand. Intravenous zoledronate was a potent agent for these patients, confirmed the efficacy of bisphosphonates on this disease. In conclusion, the novel causative mutations identified in the patients expand the genotypic spectrum of BS.     

Novel compound heterozygous mutations in the <Emphasis Type="Italic">GPR98</Emphasis> (<Emphasis Type="Italic">USH2C</Emphasis>) gene identified by whole exome sequencing in a Moroccan deaf family

In the present work, we identified two novel compound heterozygote mutations in the GPR98 (G protein-coupled receptor 98) gene causing Usher syndrome. Whole-exome sequencing was performed to study the genetic causes of Usher syndrome in a Moroccan family with three affected siblings. We identify two novel compound heterozygote mutations (c.1054C?>?A, c.16544delT) in the GPR98 gene in the three affected siblings carrying post-linguale bilateral moderate hearing loss with normal vestibular functions and before installing visual disturbances. This is the first time that mutations in the GPR98 gene are described in the Moroccan deaf patients.     

Common polymorphism (81Val>Ile) and rare mutations (257Arg>Ser and 335Ile>Ser) of the MC3R gene in obese Polish children and adolescents

The predisposing role to human obesity of the MC3R gene polymorphism is controversial. In this report we present the first study focused on the search for the MC3R polymorphism in the Polish population. Altogether 257 obese children and adolescents (RBMI>120) and 94 adults, who were never obese or overweight (BMI<25), were studied. For all subjects the entire coding sequence was analyzed by direct DNA sequencing. One common polymorphism (81Val>Ile) and two rare mutations (257Arg>Ser and 335Ile>Ser) were identified. The common polymorphism was widely distributed in the obese and control cohorts, while the mutations were identified in four obese subjects only. In case of the 335Ile>Ser substitution a three-generation family, consisting of 20 members, was also analyzed. It was found that all carriers of the 335Ser mutation were obese, but among non-carriers obese subjects also were found. Our study suggests that the predisposing effect to obesity of the 81Ile polymorphic variant is rather unlikely. With regard to the studied rare mutations we suggest that the 335Ser allele may have a small predisposing effect.     

Population specificity of the DNAI1 gene mutation spectrum in primary ciliary dyskinesia (PCD)

Background

Mutations in the DNAI1 gene, encoding a component of outer dynein arms of the ciliary apparatus, are the second most important genetic cause of primary ciliary dyskinesia (PCD), the genetically heterogeneous recessive disorder with the prevalence of ~1/20,000. The estimates of the DNAI1 involvement in PCD pathogenesis differ among the reported studies, ranging from 4% to 10%.

Methods

The coding sequence of DNAI1 was screened (SSCP analysis and direct sequencing) in a group of PCD patients (157 families, 185 affected individuals), the first ever studied large cohort of PCD patients of Slavic origin (mostly Polish); multiplex ligation-dependent probe amplification (MLPA) analysis was performed in a subset of ~80 families.

Results

Three previously reported mutations (IVS1+2-3insT, L513P and A538T) and two novel missense substitutions (C388Y and G515S) were identified in 12 families (i.e. ~8% of non-related Polish PCD patients). The structure of background SNP haplotypes indicated common origin of each of the two most frequent mutations, IVS1+2-3insT and A538T. MLPA analysis did not reveal any significant differences between patients and control samples. The Polish cohort was compared with all the previously studied PCD groups (a total of 487 families): IVS1+2-3insT remained the most prevalent pathogenetic change in DNAI1 (54% of the mutations identified worldwide), and the increased global prevalence of A538T (14%) was due to the contribution of the Polish cohort.

Conclusions

The worldwide involvement of DNAI1 mutations in PCD pathogenesis in families not preselected for ODA defects ranges from 7 to 10%; this global estimate as well as the mutation profile differs in specific populations. Analysis of the background SNP haplotypes suggests that the increased frequency of chromosomes carrying A538T mutations in Polish patients may reflects local (Polish or Slavic) founder effect. Results of the MLPA analysis indicate that no large exonic deletions are involved in PCD pathogenesis.     

Hereditary deafness in Kirov oblast: Estimation of the incidence rate and DNA diagnosis in children

Genetic analysis of hereditary deafness (HD) has been performed in the city of Kirov and ten rural districts of Kirov oblast (administrative region). The analysis employed the methods used in audiology, medical genetic counseling, and DNA diagnosis. Deafness has been established to be hereditary in 143 children from 100 unrelated families. The incidence rates of isolated and syndromic HDs in the period studied (1995–2001) have been estimated at 1.25 and 0.36 per 1000 newborns, respectively, the total incidence rate of all HD forms being 1.61 per 1000 newborns (1 case per 621 newborns). DNA analysis for the detection of seven frequent mutations in the genes GJB2 (the 35delG, 167delT, 235delC, and M34T mutations), GJB6 (the del(GJB6-D13S1854) and del(GJB6-D13S1830) mutations), and TMC1 (the R34X mutation) has been performed in families with isolated neurosensory deafness. Molecular genetic analysis has detected mutations in 51 children (48.6%); in 54 children (51.4%), no mutations have been found. The following genotypes have been identified in children with HD: 35delG/35delG in 32 probands (30.5%), 35delG/+ in 16 probands (15.2%), 35delG/235delC in 1 proband (0.95%), M34T/+ in 1 proband (0.95%), and M34T/35delG in 1 proband (0.95%). The 167delT mutation has not been found. The frequency of the 35delG mutation in the GJB2 gene has been estimated to be 39.05%. In the group with a family history of HD, mutations have been found in 66.7% of patients; in the group without a family history of HD, in 37.5% of patients. No mutation has been found in the GJB6 or TMC1 gene. Molecular genetic analysis has been performed in a family with clinically diagnosed Treacher Collins-Franceschetti syndrome. Sequencing has been used to find the 748–69C>T polymorphism in intron 6 (in the homozygous state) and the 3635C>G mutation in exon 23 leading to the substitution of glycine for alanine at position 1176 of the amino acid sequence (Ala1176Gly, in the heterozygous state), which have not been described before.     

Targeted Exome Sequencing Integrated with Clinicopathological Information Reveals Novel and Rare Mutations in Atypical,Suspected and Unknown Cases of Alport Syndrome or Proteinuria

We applied customized targeted next-generation exome sequencing (NGS) to determine if mutations in genes associated with renal malformations, Alport syndrome (AS) or nephrotic syndrome are a potential cause of renal abnormalities in patients with equivocal or atypical presentation. We first sequenced 4,041 exons representing 292 kidney disease genes in a Caucasian woman with a history of congenital vesicoureteral reflux (VUR), recurrent urinary tract infections and hydronephrosis who presented with nephrotic range proteinuria at the age of 45. Her biopsy was remarkable for focal segmental glomerulosclerosis (FSGS), a potential complication of longstanding VUR. She had no family history of renal disease. Her proteinuria improved initially, however, several years later she presented with worsening proteinuria and microhematuria. NGS analysis revealed two deleterious COL4A3 mutations, one novel and the other previously reported in AS, and a novel deleterious SALL2 mutation, a gene linked to renal malformations. Pedigree analysis confirmed that COL4A3 mutations were nonallelic and compound heterozygous. The genomic results in conjunction with subsequent abnormal electron microscopy, Collagen IV minor chain immunohistochemistry and progressive sensorineural hearing loss confirmed AS. We then modified our NGS approach to enable more efficient discovery of variants associated with AS or a subset of FSGS by multiplexing targeted exome sequencing of 19 genes associated with AS or FSGS in 14 patients. Using this approach, we found novel or known COL4A3 or COL4A5 mutations in a subset of patients with clinically diagnosed or suspected AS, APOL1 variants associated with FSGS in African Americans and novel mutations in genes associated with nephrotic syndrome. These studies demonstrate the successful application of targeted capture-based exome sequencing to simultaneously evaluate genetic variations in many genes in patients with complex renal phenotypes and provide insights into etiology of conditions with equivocal clinical and pathologic presentations.     

<Emphasis Type="Italic">GJB2</Emphasis> and <Emphasis Type="Italic">GJB6</Emphasis> gene mutations found in Indian probands with congenital hearing impairment

Genetically caused deafness is a common trait affecting one in 1000 children and is predominantly inherited in an autosomal-recessive fashion. Several mutations in the GJB2 gene and a deletion of 342 kb in GJB6 gene (delGJB6-D13S1830) have been identified worldwide in patients with hearing impairment. In the present study, 303 nonsyndromic hearing-impaired patients (140 familial; 163 sporadic) were examined clinically and screened for mutations in GJB2 and GJB6 genes. Mutations in GJB2 gene were found in 33 (10.9%) patients of whom six (18.2%) were carriers for the mutant allele. The most frequent mutation was p.W24X accounting for 87% of the mutant alleles. In addition, six other sequence variations were identified in the GJB2 gene viz., c.IVS1+1G>A, c.167delT, c.235delC, p.W77X, p.R127H (polymorphism), p.M163V. None of the samples showed del(GJB6-D13S1830) or any point mutations in GJB6 gene.     

Clinical,Molecular, and Computational Analysis in two cases with mitochondrial encephalomyopathy associated with SUCLG1 mutation in a consanguineous family

Deficiency of the mitochondrial enzyme succinyl COA ligase (SUCL) is associated with encephalomyopathic mtDNA depletion syndrome and methylmalonic aciduria. This disorder is caused by mutations in both SUCL subunits genes: SUCLG1 (α subnit) and SUCLA2 (β subnit). We report here, two Tunisian patients belonging to a consanguineous family with mitochondrial encephalomyopathy, hearing loss, lactic acidosis, hypotonia, psychomotor retardation and methylmalonic aciduria. Mutational analysis of SUCLG1 gene showed, for the first time, the presence of c.41T > C in the exon 1 at homozygous state. In-silico analysis revealed that this mutation substitutes a conserved methionine residue to a threonine at position 14 (p.M14T) located at the SUCLG1 protein mitochondrial targeting sequence. Moreover, these analysis predicted that this mutation alter stability structure and mitochondrial translocation of the protein. In Addition, a decrease in mtDNA copy number was revealed by real time PCR in the peripheral blood leukocytes in the two patients compared with controls.     

Spectrum of the GJB2 mutations in Belarussian patients with hearing loss. Findings of pilot genetic screening of hearing impairment in newborns

A total of 111 unrelated probands and their 8 sibs from Grodno oblast (Belarus) with bilateral isolated sensorineural hearing impairment were studied for the presence of mutations in the connexin 26 (GJB2) gene. Mutations were detected in 51 probands (46% of the sample). A significantly higher frequency of the GJB2 gene mutations was observed in familial cases of the disease with the autosomal recessive mode of inheritance (in 78% of families). Detected characteristics of the GJB2 gene mutation spectrum demonstrated that the using the algorithm, which was designed for Russian patients, is optimal for the molecular study of patients from Belarus. In the sample of patients with hearing loss, the highest (among other similar samples studied in the world) allele frequency of c.313_326del14 mutation (7% of all pathological GJB2 alleles) was registered; Polish origin of this deletion was suggested. It was demonstrated that detection of the GJB2 gene mutation on one patient’s chromosome only is insufficient to confirm a molecular genetic diagnosis of hearing loss of the DFNB1 genetic type (autosomal recessive hearing loss caused by the GJB2 gene mutations). Pilot screening for the GJB2 gene mutations in newborns from Grodno oblast was performed. The material from 235 children was studied during the screening; nine heterozygous carriers of the mutation were found. The c.35delG mutation was detected in a homozygous state in a single newborn (hearing loss of moderate severity was subsequently audiologically confirmed in this child).     

Inherited<Emphasis Type="Italic"> BRCA2</Emphasis> mutations in African Americans with breast and/or ovarian cancer: a study of familial and early onset cases

In order to identify the spectrum of BRCA2 mutations in African Americans, breast or ovarian cancer patients from 74 independent families at elevated risk of germline mutations were investigated. The entire coding regions and flanking introns of BRCA2 were screened for germline mutations by single-stranded conformation polymorphism, protein truncation test, or denaturing high performance liquid chromatography followed by DNA sequencing. Eight distinct protein-truncating mutations were detected in six female patients (average age of onset of breast cancer: 37.6 years) and two male patients, but not in 163 unrelated disease-free controls. Two (1993delAA, 8643delAT) of the eight pathogenic mutations observed in African Americans have not been previously described. The other six pathogenic mutations (1882delT, 1991delATAA, 2001delTTAT, 2816insA, 4075delGT, 4088delA) have been detected in Caucasians; only the 2816insA mutation has been reported previously in African Americans. There were no significant differences in the frequency of deleterious BRCA2 mutations in African Americans compared with Caucasians. Six rare variations, not previously reported, were identified in five breast cancer patients but not in 163 disease-free control subjects. Of 11 different polymorphisms identified in high-risk African-American breast cancer patients, four may be unique to African Americans. An intron 10 polymorphism observed in patients was not detected in 163 disease-free African-American control subjects; this difference is statistically significant. Since many different pathogenic mutations and variants of unknown significance are observed in African Americans, BRCA2 genetic testing in high-risk African-American families must include the entire coding and flanking non-coding regions of the gene.