Inherited loss-of-function mutations in BRCA1 and BRCA2 and other tumor suppressor genes predispose to ovarian carcinomas, but the overall burden of disease due to inherited mutations is not known. Using targeted capture and massively parallel genomic sequencing, we screened for germ-line mutations in 21 tumor suppressor genes in genomic DNA from women with primary ovarian, peritoneal, or fallopian tube carcinoma. Subjects were consecutively enrolled at diagnosis and not selected for age or family history. All classes of mutations, including point mutations and large genomic deletions and insertions, were detected. Of 360 subjects, 24% carried germ-line loss-of-function mutations: 18% in BRCA1 or BRCA2 and 6% in BARD1, BRIP1, CHEK2, MRE11A, MSH6, NBN, PALB2, RAD50, RAD51C, or TP53. Six of these genes were not previously implicated in inherited ovarian carcinoma. Primary carcinomas were generally characterized by genomic loss of normal alleles of the mutant genes. Of women with inherited mutations, >30% had no family history of breast or ovarian carcinoma, and >35% were 60 y or older at diagnosis. More patients with ovarian carcinoma carry cancer-predisposing mutations and in more genes than previously appreciated. Comprehensive genetic testing for inherited carcinoma is warranted for all women with ovarian, peritoneal, or fallopian tube carcinoma, regardless of age or family history. Clinical genetic testing is currently done gene by gene, with each test costing thousands of dollars. In contrast, massively parallel sequencing allows such testing for many genes simultaneously at low cost. O varian carcinoma is the most deadly of gynecological malignancies; the majority of women are diagnosed with advanced stage disease when the chance of cure is small. Inherited mutations in BRCA1 and BRCA2 create a lifetime risk of ovarian carcinoma of between 20% (for BRCA2) and 50% or even higher (for BRCA1) (1). It has been previously estimated that 13-15% of patients with ovarian carcinoma in North America carry germ-line mutations in BRCA1 or BRCA2 (2, 3). Hereditary ovarian carcinoma also occurs in the context of Lynch syndrome [hereditary nonpolyposis colorectal cancer (HNPCC)], but the proportion of ovarian carcinoma explained by germ-line mutations in the mismatch repair genes has not been determined. Inherited mutations in RAD51C, RAD51D, and PALB2 have also been reported in patients with familial ovarian carcinoma (4-6). The overall proportion of ovarian carcinoma due to germ-line mutations in these genes and the roles of other tumor suppressor genes, particularly those implicated in inherited breast cancer, remain unknown.Women with early stage ovarian carcinoma have far better survival than women whose carcinomas are diagnosed at later stages, but current methods of early detection have not proven effective (7). In contrast, risk-reducing salpingo-oophorectomy in women with BRCA1 or BRCA2 mutations dramatically reduces risk of ovarian carcinoma and significantly decreases overall mortality (8-10). It is critically i...

Massively parallel sequencing of targeted regions, exomes, and complete genomes has begun to dramatically increase the pace of discovery of genes responsible for human disorders. Here we describe how exome sequencing in conjunction with homozygosity mapping led to rapid identification of the causative allele for nonsyndromic hearing loss DFNB82 in a consanguineous Palestinian family. After filtering out worldwide and population-specific polymorphisms from the whole exome sequence, only a single deleterious mutation remained in the homozygous region linked to DFNB82. The nonsense mutation leads to an early truncation of the G protein signaling modulator GPSM2, a protein that is essential for maintenance of cell polarity and spindle orientation. In the mouse inner ear, GPSM2 is localized to apical surfaces of hair cells and supporting cells and is most highly expressed during embryonic development. Identification of GPSM2 as essential to the development of normal hearing suggests dysregulation of cell polarity as a mechanism underlying hearing loss.

Comprehensive sequencing of tumor suppressor genes to evaluate inherited predisposition to cancer yields many individually rare missense alleles of unknown functional and clinical consequence. To address this problem for CHEK2 missense alleles, we developed a yeast-based assay to assess in vivo CHEK2mediated response to DNA damage. Of 25 germline CHEK2 missense alleles detected in familial breast cancer patients, 12 alleles had complete loss of DNA damage response, 8 had partial loss and 5 exhibited a DNA damage response equivalent to that mediated by wild-type CHEK2. Variants exhibiting reduced response to DNA damage were found in all domains of the CHEK2 protein. Assay results were in agreement with epidemiologic assessments of breast cancer risk for those variants sufficiently common for case-control studies to have been undertaken. Assay results were largely concordant with consensus predictions of in silico tools, particularly for damaging alleles in the kinase domain. However, of the 25 variants, 6 were not consistently classifiable by in silico tools. An in vivo assay of cellular response to DNA damage by mutant CHEK2 alleles may complement and extend epidemiologic and genetic assessment of their clinical consequences.

Age-related hearing loss is due to death over time, primarily by apoptosis, of hair cells in the inner ear. Studies of mutant genes responsible for inherited progressive hearing loss have suggested possible mechanisms for hair cell death, but critical connections between these mutations and the causes of progressive hearing loss have been elusive. In an Israeli kindred, dominant, adult-onset, progressive nonsyndromic hearing loss DFNA51 is due to a tandem inverted genomic duplication of 270 kb that includes the entire wild-type gene encoding the tight junction protein TJP2 (ZO-2). In the mammalian inner ear, TJP2 is expressed mainly in tight junctions, and also in the cytoplasm and nuclei. TJP2 expression normally decreases with age from embryonic development to adulthood. In cells of affected family members, TJP2 transcript and protein are overexpressed, leading to decreased phosphorylation of GSK-3beta and to altered expression of genes that regulate apoptosis. These results suggest that TJP2- and GSK-3beta-mediated increased susceptibility to apoptosis of cells of the inner ear is the mechanism for adult-onset hearing loss in this kindred and may serve as one model for age-related hearing loss in the general population.

Individuals with autism are more likely to carry rare inherited and de novo copy number variants (CNVs). However, further research is needed to establish which CNVs are causal and the mechanisms by which these CNVs influence autism. We examined genomic DNA of children with autism (N¼41) and healthy controls (N¼367) for rare CNVs using a high-resolution array comparative genomic hybridization platform. We show that individuals with autism are more likely to harbor rare CNVs as small as B10 kb, a threshold not previously detectable, and that CNVs in cases disproportionately affect genes involved in transcription, nervous system development, and receptor activity. We also show that a subset of genes that have known or suspected allele-specific or imprinting effects and are within rare-case CNVs may undergo loss of transcript expression. In particular, expression of CNTNAP2 and ZNF214 are decreased in probands compared with their unaffected transmitting parents. Furthermore, expression of PRODH and ARID1B, two genes affected by de novo CNVs, are decreased in probands compared with controls. These results suggest that for some genes affected by CNVs in autism, reduced transcript expression may be a mechanism of pathogenesis during neurodevelopment.