The role of rare compound heterozygous events in autism spectrum disorder.

The identification of genetic variants underlying autism spectrum disorders (ASDs) may contribute to a better understanding of their underlying biology. To examine the possible role of a specific type of compound heterozygosity in ASD, namely, the occurrence of a deletion together with a functional nucleotide variant on the remaining allele, we sequenced 550 genes in 149 individuals with ASD and their deletion-transmitting parents. This approach allowed us to identify additional sequence variants occurring in the remaining allele of the deletion. Our main goal was to compare the rate of sequence variants in remaining alleles of deleted regions between probands and the deletion-transmitting parents. We also examined the predicted functional effect of the identified variants using Combined Annotation-Dependent Depletion (CADD) scores. The single nucleotide variant-deletion co-occurrence was observed in 13.4% of probands, compared with 8.1% of parents. The cumulative burden of sequence variants (n = 68) in pooled proband sequences was higher than the burden in pooled sequences from the deletion-transmitting parents (n = 41, X2 = 6.69, p = 0.0097). After filtering for those variants predicted to be most deleterious, we observed 21 of such variants in probands versus 8 in their deletion-transmitting parents (X2 = 5.82, p = 0.016). Finally, cumulative CADD scores conferred by these variants were significantly higher in probands than in deletion-transmitting parents (burden test, ß = 0.13; p = 1.0 × 10-5). Our findings suggest that the compound heterozygosity described in the current study may be one of several mechanisms explaining variable penetrance of CNVs with known pathogenicity for ASD.

Polymorphisms in MMP-14 and MMP-2 genes and ovarian cancer survival.

BACKGROUND: Functional polymorphisms in matrix metalloproteinases can increase or decrease the risk of cancer. This study focused on ovarian cancer and investigated how polymorphisms in the coding region of MMP-14 and the promoter region of MMP-2 are related to clinical characteristics including survival. METHODS: In 144 patients with ovarian tumours from a Caucasian population, polymorphisms of MMP-14 (+7096 and +6767) and MMP-2 (-735 and -1306) were analysed. These results were then correlated to the immunohistochemical expression of MMP-14 and MMP-2 and clinical characteristics. RESULTS: In these patients, the MMP-14 +7096 polymorphism showed only TT genotype, in sharp contrast to the described MAF (minimal allele frequency) C of 27%. The MMP-14 +6767 G>A polymorphism was found to have a hazard ratio of 2.09 (CI 1.00-4.35, p 0.046) for recurrence-free survival in advanced-stage patients. However, this significance disappeared after Bonferroni correction for multiple testing. No other correlations between MMP-14 and MMP-2 polymorphisms, immunohistochemistry and clinical characteristics were found, except between the MMP-2 -1306 polymorphism and differentiation grade, with a Spearman correlation coefficient of -0.19, p 0.064. CONCLUSIONS: In ovarian cancer, the MMP-14 +6767 G>A polymorphism in the coding region seems to improve recurrence-free survival with a hazard ratio of 2.09 (CI 1.00-4.35, p 0.046). However, as this significance disappeared after correction for multiple testing, there is a need for further research on the functional effect of this change in the MMP-14 gene with larger patient sample sizes.

Detection of long repeat expansions from PCR-free whole-genome sequence data.

Identifying large expansions of short tandem repeats (STRs), such as those that cause amyotrophic lateral sclerosis (ALS) and fragile X syndrome, is challenging for short-read whole-genome sequencing (WGS) data. A solution to this problem is an important step toward integrating WGS into precision medicine. We developed a software tool called ExpansionHunter that, using PCR-free WGS short-read data, can genotype repeats at the locus of interest, even if the expanded repeat is larger than the read length. We applied our algorithm to WGS data from 3001 ALS patients who have been tested for the presence of the C9orf72 repeat expansion with repeat-primed PCR (RP-PCR). Compared against this truth data, ExpansionHunter correctly classified all (212/212, 95% CI [0.98, 1.00]) of the expanded samples as either expansions (208) or potential expansions (4). Additionally, 99.9% (2786/2789, 95% CI [0.997, 1.00]) of the wild-type samples were correctly classified as wild type by this method with the remaining three samples identified as possible expansions. We further applied our algorithm to a set of 152 samples in which every sample had one of eight different pathogenic repeat expansions, including those associated with fragile X syndrome, Friedreich's ataxia, and Huntington's disease, and correctly flagged all but one of the known repeat expansions. Thus, ExpansionHunter can be used to accurately detect known pathogenic repeat expansions and provides researchers with a tool that can be used to identify new pathogenic repeat expansions.

Genome-wide association analyses identify new risk variants and the genetic architecture of amyotrophic lateral sclerosis.

To elucidate the genetic architecture of amyotrophic lateral sclerosis (ALS) and find associated loci, we assembled a custom imputation reference panel from whole-genome-sequenced patients with ALS and matched controls (n = 1,861). Through imputation and mixed-model association analysis in 12,577 cases and 23,475 controls, combined with 2,579 cases and 2,767 controls in an independent replication cohort, we fine-mapped a new risk locus on chromosome 21 and identified C21orf2 as a gene associated with ALS risk. In addition, we identified MOBP and SCFD1 as new associated risk loci. We established evidence of ALS being a complex genetic trait with a polygenic architecture. Furthermore, we estimated the SNP-based heritability at 8.5%, with a distinct and important role for low-frequency variants (frequency 1-10%). This study motivates the interrogation of larger samples with full genome coverage to identify rare causal variants that underpin ALS risk.

Large-scale screening in sporadic amyotrophic lateral sclerosis identifies genetic modifiers in C9orf72 repeat carriers.

Sporadic amyotrophic lateral sclerosis (ALS) is considered to be a complex disease with multiple genetic risk factors contributing to the pathogenesis. Identification of genetic risk factors that co-occur frequently could provide relevant insight into underlying mechanisms of motor neuron degeneration. To dissect the genetic architecture of sporadic ALS, we undertook a large sequencing study in 755 apparently sporadic ALS cases and 959 controls, analyzing 10 ALS genes: SOD1, C9orf72, TARDBP, FUS, ANG, CHMP2B, ATXN2, NIPA1, SMN1, and UNC13A. We observed sporadic cases with multiple genetic risk variants in 4.1% compared with 1.3% in controls. The overall difference was not in excess of what is to be expected by chance (binomial test, p = 0.59). We did, however, observe a higher frequency than expected of C9orf72 repeat carriers with co-occurring susceptibility variants (ATXN2, NIPA1, and SMN1; p = 0.001), which is mainly because of the co-occurrence of NIPA1 repeats in 15% of C9orf72 repeat carriers (p = 0.006).

Endogenous fluorescence spectroscopy of cell suspensions for chemopreventive drug monitoring.

Cancer chemopreventive agents such as N-4-(hydroxyphenyl)retinamide (4HPR) are thought to prevent cancers by suppressing growth or inducing apoptosis in precancerous cells. Mechanisms by which these drugs affect cells are often not known, and the means to monitor their effects is not available. In this study endogenous fluorescence spectroscopy was used to measure metabolic changes in response to treatment with 4HPR in ovarian and bladder cancer cell lines. Fluorescence signals consistent with nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD) and tryptophan were measured to monitor cellular activity through redox status and protein content. Cells were treated with varying concentrations of 4HPR and measured in a stable environment with a sensitive fluorescence spectrometer. Results suggest that redox signal of all cells changed in a similar dose-dependant manner but started at different baseline levels. Redox signal changes depended primarily on changes consistent with NADH fluorescence, whereas the FAD fluorescence remained relatively constant. Similarly, tryptophan fluorescence decreased with increased drug treatment, suggesting a decrease in protein production. Given that each cell line has been shown to have a different apoptotic response to 4HPR, fluorescence redox values along with changes in tryptophan fluorescence may be a response as well as an endpoint marker for chemopreventive drugs.

Imaging of the ovary.

Epithelial ovarian cancer has the highest mortality rate among the gynecologic cancers and spreads beyond the ovary in 90% of the women diagnosed with ovarian cancer. Detection before the disease has spread beyond the ovary would significantly improve the survival from ovarian cancer, which is currently only 30% over 5 years, despite extensive efforts to improve the survival. This study describes initial investigation of the use of optical technologies to improve the outcome for this disease by detecting cancers at an earlier and more treatable stage. Women undergoing oophorectomy were recruited for this study. Ovaries were harvested for fluorescence spectroscopy, confocal microscopy, and optical coherence tomography. Fluorescence spectroscopy showed large diagnostic differences between normal and abnormal tissue at 270 and 340 nm excitation. Optical coherence tomography was able to image up to 2mm deep into the ovary with particular patterns of backscattered intensity observed in normal versus abnormal tissue. Fluorescence confocal microscopy was able to visualize sub-cellular structures of the surface epithelium and underlying cell layers. Optical imaging and/or spectroscopy has the potential to improve the diagnostic capability in the ovary, but extended systematic investigations are needed to identify the unique signatures of disease. The combination of optical technologies supported by modern molecular biology may lead to an instrument that can accurately detect early carcinogenesis.