Improved structural variant interpretation for hereditary cancer susceptibility using long-read sequencing.

PURPOSE: Structural variants (SVs) may be an underestimated cause of hereditary cancer syndromes given the current limitations of short-read next-generation sequencing. Here we investigated the utility of long-read sequencing in resolving germline SVs in cancer susceptibility genes detected through short-read genome sequencing. METHODS: Known or suspected deleterious germline SVs were identified using Illumina genome sequencing across a cohort of 669 advanced cancer patients with paired tumor genome and transcriptome sequencing. Candidate SVs were subsequently assessed by Oxford Nanopore long-read sequencing. RESULTS: Nanopore sequencing confirmed eight simple pathogenic or likely pathogenic SVs, resolving three additional variants whose impact could not be fully elucidated through short-read sequencing. A recurrent sequencing artifact on chromosome 16p13 and one complex rearrangement on chromosome 5q35 were subsequently classified as likely benign, obviating the need for further clinical assessment. Variant configuration was further resolved in one case with a complex pathogenic rearrangement affecting TSC2. CONCLUSION: Our findings demonstrate that long-read sequencing can improve the validation, resolution, and classification of germline SVs. This has important implications for return of results, cascade carrier testing, cancer screening, and prophylactic interventions.

Pan-cancer analysis of advanced patient tumors reveals interactions between therapy and genomic landscapes.

Advanced and metastatic tumors with complex treatment histories drive cancer mortality. Here we describe the POG570 cohort, a comprehensive whole-genome, transcriptome and clinical dataset, amenable for exploration of the impacts of therapies on genomic landscapes. Previous exposure to DNA-damaging chemotherapies and mutations affecting DNA repair genes, including POLQ and genes encoding Polζ, were associated with genome-wide, therapy-induced mutagenesis. Exposure to platinum therapies coincided with signatures SBS31 and DSB5 and, when combined with DNA synthesis inhibitors, signature SBS17b. Alterations in ESR1, EGFR, CTNNB1, FGFR1, VEGFA and DPYD were consistent with drug resistance and sensitivity. Recurrent noncoding events were found in regulatory region hotspots of genes including TERT, PLEKHS1, AP2A1 and ADGRG6. Mutation burden and immune signatures corresponded with overall survival and response to immunotherapy. Our data offer a rich resource for investigation of advanced cancers and interpretation of whole-genome and transcriptome sequencing in the context of a cancer clinic.

Effects of high fructose intake on the development of hypertension in the spontaneously hypertensive rats: the role of AT1R/gp91PHOX signaling in the rostral ventrolateral medulla.

Both genetic and dietary factors determine the development of hypertension. Whether dietary factor impacts the development of hereditary hypertension is unknown. Here, we evaluated the effect of daily high-fructose diet (HFD) on the development of hypertension in adolescent spontaneously hypertensive rats (SHR). Six-week-old SHR were randomly divided into two groups to receive HFD or normal diet (ND) for 3 weeks. The temporal profile of systolic blood pressure, alongside the sympathetic vasomotor activity, in the SHR-HFD showed significantly greater increases at 9-12 weeks of age compared with the age-matched SHR-ND group. Immunofluorescence was used to identify the distribution of reactive oxygen species (ROS), oxidants and antioxidants in rostral ventrolateral medulla (RVLM) where sympathetic premotor neurons reside. In RVLM of SHR-HFD, the levels of ROS accumulation and lipid peroxidation were elevated. The changes in protein expression were measured by Western blot. NADPH oxidase subunit gp91phox and angiotensin II type I receptor were up-regulated in RVLM neuron. On the other hand, the expression of extracellular superoxide dismutase was suppressed. Both molecular and hemodynamic changes in the SHR-HFD were rescued by oral pioglitazone treatment from weeks 7 to 9. Furthermore, central infusion with tempol, a ROS scavenger, effectively ameliorated ROS accumulation in RVLM and diminished the heightened pressor response and enhanced sympathetic activity in the SHR-HFD. Together, these results suggest that HFD intake at adolescent SHR may impact the development of hypertension via increasing oxidative stress in RVLM which could be effectively attenuated by pioglitazone treatment.

Nontranscriptional activation of PI3K/Akt signaling mediates hypotensive effect following activation of estrogen receptor ß in the rostral ventrolateral medulla of rats.

BACKGROUND: Estrogen acts on the rostral ventrolateral medulla (RVLM), where sympathetic premotor neurons are located, to elicit vasodepressor effects via an estrogen receptor (ER)ß-dependent mechanism. We investigated in the present study nontranscriptional mechanism on cardiovascular effects following activation of ERß in the RVLM, and delineated the involvement of phosphatidylinositol 3-kinase (PI3K)/serine/threonine kinase (Akt) signaling pathway in the effects. METHODS: In male Sprague-Dawley rats maintained under propofol anesthesia, changes in arterial pressure, heart rate and sympathetic neurogenic vasomotor tone were examined after microinjection bilaterally into RVLM of 17ß-estradiol (E2ß) or a selective ERα or ERß agonist. Involvement of ER subtypes and PI3K/Akt signaling pathway in the induced cardiovascular effects were studied using pharmacological tools of antagonists or inhibitors, gene manipulation with antisense oligonucleotide (ASON) or adenovirus-mediated gene transfection. RESULTS: Similar to E2ß (1 pmol), microinjection of ERß agonist, diarylpropionitrile (DPN, 1, 2 or 5 pmol), into bilateral RVLM evoked dose-dependent hypotension and reduction in sympathetic neurogenic vasomotor tone. These vasodepressive effects of DPN (2 pmol) were inhibited by ERß antagonist, R,R-tetrahydrochrysene (50 pmol), ASON against ERß mRNA (250 pmol), PI3K inhibitor LY294002 (5 pmol), or Akt inhibitor (250 pmol), but not by ERα inhibitor, methyl-piperidino-pyrazole (1 nmol), or transcription inhibitor, actinomycin D (5 or 10 nmol). Gene transfer by microinjection into bilateral RVLM of adenovirus encoding phosphatase and tensin homologues deleted on chromosome 10 (5 × 10(8) pfu) reversed the vasodepressive effects of DPN. CONCLUSIONS: Our results indicate that vasodepressive effects following activation of ERß in RVLM are mediated by nongenomic activation of PI3K/Akt signaling pathway. This study provides new insight in the intracellular signaling cascades involved in central vasodepressive functions of estrogen.

Activation of estrogen receptor beta-dependent nitric oxide signaling mediates the hypotensive effects of estrogen in the rostral ventrolateral medulla of anesthetized rats.

BACKGROUND: Apart from their well-known peripheral cardiovascular effects, emerging evidence indicates that estrogen acts as a modulator in the brain to regulate cardiovascular functions. The underlying mechanisms of estrogen in central cardiovascular regulation, however, are poorly understood. The present study investigated the cardiovascular effects of 17beta-estradiol (E2beta) in the rostral ventrolateral medulla (RVLM), where sympathetic premotor neurons are located, and delineated the engagement of nitric oxide (NO) in E2beta-induced cardiovascular responses. METHODS: In male Sprague-Dawley rats maintained under propofol anesthesia, the changes of blood pressure, heart rate and sympathetic vasomotor tone after microinjection bilaterally into the RVLM of a synthetic estrogen, E2beta were examined for at least 120 min. The involvement of ERalpha and/or ERbeta subtypes was determined by microinjection of selective ERalpha or ERbeta agonist into bilateral RVLM. Different NO synthase (NOS) inhibitors were used to evaluate the involvement of differential of NOS isoforms in the cardiovascular effects of E2beta. RESULTS: Bilateral microinjection of E2beta (0.5, 1, or 5 pmol) into the RVLM dose-dependently decreased systemic arterial pressure (SAP) and the power density of the vasomotor components of SAP signals, our experimental index for sympathetic neurogenic vasomotor tone. These cardiovascular depressive effects of E2beta (1 pmol) were abolished by co-injection of ER antagonist ICI 182780 (0.25 or 0.5 pmol), but not a transcription inhibitor actinomycin D (10 nmol). Like E2beta, microinjection bilaterally into the RVLM of a selective ERbeta agonist 2,3-bis(4-hydroxyphenyl) propionitrile (DPN, 1, 2, or 5 pmol) induced significant decreases in these hemodynamic parameters in a dose-dependent manner. In contrast, the selective ERalpha agonist 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole (5 pmol) did not influence the same cardiovascular parameters. Co-administration bilaterally into the RVLM of NOS inhibitor NG-nitro-L-arginine methyl ester (5 nmol) or selective inducible NOS (iNOS) inhibitor S-methylisothiourea (25 pmol), but not selective neuronal NOS inhibitor 7-nitroindazole (0.5 pmol) or endothelial NOS inhibitor N5-(1-Iminoethyl)-L-ornithine (2.5 pmol), significantly attenuated the cardiovascular depressive effects elicited by DPN (2 pmol). CONCLUSION: Our results indicate that E2beta in the RVLM elicited short-term cardiovascular depressive effects via an ERbeta-dependent nontranscriptional mechanism. These vasodepressor effects of E2beta are likely to be mediated by the iNOS-derived NO in the RVLM.

PolySearch: a web-based text mining system for extracting relationships between human diseases, genes, mutations, drugs and metabolites.

A particular challenge in biomedical text mining is to find ways of handling 'comprehensive' or 'associative' queries such as 'Find all genes associated with breast cancer'. Given that many queries in genomics, proteomics or metabolomics involve these kind of comprehensive searches we believe that a web-based tool that could support these searches would be quite useful. In response to this need, we have developed the PolySearch web server. PolySearch supports >50 different classes of queries against nearly a dozen different types of text, scientific abstract or bioinformatic databases. The typical query supported by PolySearch is 'Given X, find all Y's' where X or Y can be diseases, tissues, cell compartments, gene/protein names, SNPs, mutations, drugs and metabolites. PolySearch also exploits a variety of techniques in text mining and information retrieval to identify, highlight and rank informative abstracts, paragraphs or sentences. PolySearch's performance has been assessed in tasks such as gene synonym identification, protein-protein interaction identification and disease gene identification using a variety of manually assembled 'gold standard' text corpuses. Its f-measure on these tasks is 88, 81 and 79%, respectively. These values are between 5 and 50% better than other published tools. The server is freely available at http://wishart.biology.ualberta.ca/polysearch.