Common variants in the CYP2C19 gene are associated with susceptibility to endometriosis

Objective: To follow-up previous studies highlighting a possible role for cytochrome P450, family 2, subfamily C, 19 (CYP2C19) in susceptibility to endometriosis by searching for additional variants in the CYP2C19 gene that may be associated with the disease. Design Case-control study. Setting Academic research. Subject(s) The cases comprised 2,271 women with surgically confirmed endometriosis; the controls comprised 939 women with self-report of no endometriosis and 1,770 unscreened population samples. Intervention(s) Sequencing of the CYP2C19 region and follow-up of 80 single nucleotide polymorphisms (SNPs) in two case-control samples. Main Outcome Measure(s) Allele frequency differences between cases and controls. Result(s) Sequencing of the CYP2C19 gene region resulted in the detection of a large number of known and novel SNPs. Genotyping of 80 polymorphic SNPs in 901 endometriosis cases and 939 controls resulted in study-wide significant association signals for SNPs in moderate or complete linkage disequilibrium with rs4244285, a functional SNP in exon 5 that abrogates CYP2C19 function through the creation of an alternative splice site. Evidence of association was also detected for another functional SNP in the CYP2C19 promoter, rs12248560, which was highlighted in our previous study. Conclusion(s) Functional variants in CYP2C19 may contribute to endometriosis susceptibility in both familial and sporadic cases. © 2014 by American Society for Reproductive Medicine.

The future for genetic studies in reproduction

Genetic factors contribute to risk of many common diseases affecting reproduction and fertility. In recent years, methods for genome-wide association studies(GWAS) have revolutionized gene discovery forcommontraits and diseases. Results of GWAS are documented in the Catalog of Published Genome-Wide Association Studies at the National Human Genome Research Institute and report over 70 publications for 32 traits and diseases associated with reproduction. These include endometriosis, uterine fibroids, age at menarche and age at menopause. Results that pass appropriate stringent levels of significance are generally well replicated in independent studies. Examples of genetic variation affecting twinning rate, infertility, endometriosis and age at menarche demonstrate that the spectrum of disease-related variants for reproductive traits is similar to most other common diseases.GWAS 'hits' provide novel insights into biological pathways and the translational value of these studies lies in discovery of novel gene targets for biomarkers, drug development and greater understanding of environmental factors contributing to disease risk. Results also show that genetic data can help define sub-types of disease and co-morbidity with other traits and diseases. To date, many studies on reproductive traits have used relatively small samples. Future genetic marker studies in large samples with detailed phenotypic and clinical information will yield new insights into disease risk, disease classification and co-morbidity for many diseases associated with reproduction and infertility.

An evaluation of potential candidate genes involved in salinity tolerance in striped catfish (Pangasianodon hypopthalmus) using an RNA-SEQ approach

The project investigated the molecular response of Tra catfish (Pangasianodon hypophthalmus) to elevated salinity conditions. We employed Next generation sequencing platform to evaluate differential gene expression profiles of key genes under two salinity conditions. Results of the current project can form the basis for further studies to confirm the functional roles of specific genes that influence salinity tolerance in the target species and more broadly in other freshwater teleost fishes. Ultimately, the approach can contribute to developing superior culture stocks of the target species.

The empire of cancer: Gene patents and cancer voices

In his book, The Emperor of All Maladies, Siddhartha Mukherjee writes a history of cancer — "It is a chronicle of an ancient disease — once a clandestine, 'whispered-about' illness — that has metamorphosed into a lethal shape-shifting entity imbued with such penetrating metaphorical, medical, scientific, and political potency that cancer is often described as the defining plague of our generation." Increasingly, an important theme in the history of cancer is the role of law, particularly in the field of intellectual property law. It is striking that a number of contemporary policy debates over intellectual property and public health have concerned cancer research, diagnosis, and treatment. In the area of access to essential medicines, there has been much debate over Novartis’ patent application in respect of Glivec, a treatment for leukaemia. India’s Supreme Court held that the Swiss company’s patent application violated a safeguard provision in India’s patent law designed to stop evergreening. In the field of tobacco control, the Australian Government introduced plain packaging for tobacco products in order to address the health burdens associated with the tobacco epidemic. This regime was successfully defended in the High Court of Australia. In the area of intellectual property and biotechnology, there have been significant disputes over the Utah biotechnology company Myriad Genetics and its patents in respect of genetic testing for BRCA1 and BRCA2, which are related to breast cancer and ovarian cancer. The Federal Court of Australia handed down a decision on the validity of Myriad Genetics’ patent in respect of genetic testing for BRCA1 in February 2013. The Supreme Court of the United States heard a challenge to the validity of Myriad Genetics’ patents in this area in April 2013, and handed down a judgment in July 2013. Such disputes have involved tensions between intellectual property rights, and public health. This article focuses upon one of these important test cases involving intellectual property, public health, and cancer research. In June 2010, Cancer Voices Australia and Yvonne D’Arcy brought an action in the Federal Court of Australia against the validity of a BRCA1 patent — held by Myriad Genetics Inc, the Centre de Recherche du Chul, the Cancer Institute of Japan and Genetic Technologies Limited. Yvonne D’Arcy — a Brisbane woman who has had treatment for breast cancer — maintained: "I believe that what they are doing is morally and ethically corrupt and that big companies should not control any parts of the human body." She observed: "For my daughter, I've had her have [sic] mammograms, etc, because of me but I would still like her to be able to have the test to see if the mutation gene is in there from me." The applicants made the following arguments: "Genes and the information represented by human gene sequences are products of nature universally present in each individual, and the information content of a human gene sequence is fixed. Genetic variations or mutations are products of nature. The isolation of the BRCA1 gene mutation from the human body constitutes no more than a medical or scientific discovery of a naturally occurring phenomenon and does not give rise to a patentable invention." The applicants also argued that "the alleged invention is not a patentable invention in that, so far as claimed in claims 1–3, it is not a manner of manufacture within the meaning of s 6 of the Statute of Monopolies". The applicants suggested that "the alleged invention is a mere discovery". Moreover, the applicants contended that "the alleged invention of each of claims 1-3 is not a patentable invention because they are claims for biological processes for the generation of human beings". The applicants, though, later dropped the argument that the patent claims related to biological processes for the generation of human beings. In February 2013, Nicholas J of the Federal Court of Australia considered the case brought by Cancer Voices Australia and Yvonne D’Arcy against Myriad Genetics. The judge presented the issues in the case, as follows: "The issue that arises in this case is of considerable importance. It relates to the patentability of genes, or gene sequences, and the practice of 'gene patenting'. Briefly stated, the issue to be decided is whether under the Patents Act 1990 (Cth) a valid patent may be granted for a claim that covers naturally occurring nucleic acid — either deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) — that has been 'isolated'". In this context, the word "isolated" implies that naturally occurring nucleic acid found in the cells of the human body, whether it be DNA or RNA, has been removed from the cellular environment in which it naturally exists and separated from other cellular components also found there. The genes found in the human body are made of nucleic acid. The particular gene with which the patent in suit is concerned (BRCA1) is a human breast and ovarian cancer disposing gene. Various mutations that may be present in this gene have been linked to various forms of cancer including breast cancer and ovarian cancer.' The judge held in this particular case that Myriad Genetics’ patent claims were a "manner of manufacture" under s 6 of the Statute of Monopolies and s 18(1)(a) of the Patents Act 1990 (Cth). The matter is currently under appeal in the Full Court of the Federal Court of Australia. This article interprets the dispute over Myriad Genetics in light of the scholarly work of Nobel Laureate Professor Joseph Stiglitz on inequality. Such work has significant explanatory power in the context of intellectual property and biotechnology. First, Stiglitz has contended that "societal inequality was a result not just of the laws of economics, but also of how we shape the economy — through politics, including through almost every aspect of our legal system". Stiglitz is concerned that "our intellectual property regime … contributes needlessly to the gravest form of inequality." He maintains: "The right to life should not be contingent on the ability to pay." Second, Stiglitz worries that "some of the most iniquitous aspects of inequality creation within our economic system are a result of 'rent-seeking': profits, and inequality, generated by manipulating social or political conditions to get a larger share of the economic pie, rather than increasing the size of that pie". He observes that "the most iniquitous aspect of this wealth appropriation arises when the wealth that goes to the top comes at the expense of the bottom." Third, Stiglitz comments: "When the legal regime governing intellectual property rights is designed poorly, it facilitates rent-seeking" and "the result is that there is actually less innovation and more inequality." He is concerned that intellectual property regimes "create monopoly rents that impede access to health both create inequality and hamper growth more generally." Finally, Stiglitz has recommended: "Government-financed research, foundations, and the prize system … are alternatives, with major advantages, and without the inequality-increasing disadvantages of the current intellectual property rights system.’" This article provides a critical analysis of the Australian litigation and debate surrounding Myriad Genetics’ patents in respect of genetic testing for BRCA1. First, it considers the ruling of Nicholas J in the Federal Court of Australia that Myriad Genetics’ patent was a manner of manufacture as it related to an artificially created state of affairs, and not mere products of nature. Second, it examines the policy debate over gene patents in Australia, and its relevance to the litigation involving Myriad Genetics. Third, it examines comparative law, and contrasts the ruling by Nicholas J in the Federal Court of Australia with developments in the United States, Canada, and the European Union. Fourth, this piece considers the reaction to the decision of Nicholas at first instance in Australia. Fifth, the article assesses the prospects of an appeal to the Full Federal Court of Australia over the Myriad Genetics’ patents. Finally, this article observes that, whatever happens in respect of litigation against Myriad Genetics, there remains controversy over Genetic Technologies Limited. The Melbourne firm has been aggressively licensing and enforcing its related patents on non-coding DNA and genomic mapping.

The sequence: Inside the race for the human genome

The human genome project was a grand scientific enterprise which attracted both hyperbole and ridicule alike. The project was lauded as “the moon shot of the life sciences”, the “holy grail of man”, “the code of codes”, and “the book of life”. Such rhetoric has also received scorn. President George Bush senior managed to deflate the pretensions of the project with the accidental slip that it was the “human gnome initiative”. In The Sequence, Kevin Davies seeks to go beyond such metaphors, and provide a candid and honest account of the race of the human genome project. The author is indebted to the authoritative book The Gene Wars, which considered the early struggles over the human genome project. Robert Cook-Deegan observes that there was initially much debate over whether there should be a Human Genome Project at all: The debate became one of “big” science versus “small” science. The reliance on systematic technology development and goal-directed gene-mapping efforts presaged a new style for biology, one that elicited excitement from those attracted to whiz-bang technologies but drew gasps of revulsion from those who aspired to cultivate biology on a more modest scale and with decentralized organisation. The battle was, among other things, over whose vision would control the budget and which scientific aesthetic would prevail.

The new conquistadors: Patent law and expressed sequence tags

It's akin to the old Spanish, English and Portuguese explorers. They would take their boats until they found some edge of land, then they would go up and plant the flag of their king or queen. They didn't know what they'd discovered; how big it is, where it goes to - but they would claim it anyway. David Korn of the Association of American Medical Colleges This article analyses recent litigation over patent law and expressed sequence tags (ESTs). In the case of In re Fisher, the United States Court of Appeals for the Federal Circuit engaged in judicial consideration of the revised utility guidelines of the United States Patent and Trademark Office (USPTO). In this matter, the agricultural biotechnology company Monsanto sought to patent ESTs in maize plants. A patent examiner and the Board of Patent Appeals and Interferences had doubted whether the patent application was useful. Monsanto appealed against the rulings of the USPTO. A number of amicus curiae intervened in the matter in support of the USPTO - including Genentech, Affymetrix, Dow AgroSciences, Eli Lilly, the National Academy of Sciences, and the Association of American Medical Colleges. The majority of the Court of Appeals for the Federal Circuit supported the position of the USPTO, and rejected the patent application on the grounds of utility. The split decision highlighted institutional tensions over the appropriate thresholds for patent criteria - such as novelty, non-obviousness, and utility. The litigation raised larger questions about the definition of research tools, the incremental nature of scientific progress, and the role of patent law in innovation policy. The decision of In re Fisher will have significant ramifications for gene patents, in the wake of the human genome project. Arguably, the USPTO utility guidelines need to be reinforced by a tougher application of the standards of novelty and non-obviousness in respect of gene patents.

Complex Symbolic Sequence Encodings for Predictive Monitoring of Business Processes

This paper addresses the problem of predicting the outcome of an ongoing case of a business process based on event logs. In this setting, the outcome of a case may refer for example to the achievement of a performance objective or the fulfillment of a compliance rule upon completion of the case. Given a log consisting of traces of completed cases, given a trace of an ongoing case, and given two or more possible out- comes (e.g., a positive and a negative outcome), the paper addresses the problem of determining the most likely outcome for the case in question. Previous approaches to this problem are largely based on simple symbolic sequence classification, meaning that they extract features from traces seen as sequences of event labels, and use these features to construct a classifier for runtime prediction. In doing so, these approaches ignore the data payload associated to each event. This paper approaches the problem from a different angle by treating traces as complex symbolic sequences, that is, sequences of events each carrying a data payload. In this context, the paper outlines different feature encodings of complex symbolic sequences and compares their predictive accuracy on real-life business process event logs.

Genome sequence of marine bacterium idiomarina sp. strain 28-8, isolated from Korean ark shells

Idiomarina sp. strain 28-8 is an aerobic, Gram-negative, flagellar bacterium isolated from the bodies of ark shells (Scapharca broughtonii) collected from underwater sediments in Gangjin Bay, South Korea. Here, we present the draft genome sequence of Idiomarina sp. 28-8 (2,971,606 bp, with a G+C content of 46.9%), containing 2,795 putative coding sequences.

Mitochondrial DNA sequence analysis from multiple gene fragments reveals genetic heterogeneity of Crassostrea ariakensis in East Asia

The native Asian oyster, Crassostrea ariakensis is one of the most common and important Crassostrea species that occur naturally along the coast of East Asia. Molecular species diagnosis is a prerequisite for population genetic analysis of wild oyster populations because oyster species cannot be discriminated reliably using external morphological characters alone due to character ambiguity. To date there have been few phylogeographic studies of natural edible oyster populations in East Asia, in particular this is true of the common species in Korea C. ariakensis. We therefore assessed the levels and patterns of molecular genetic variation in East Asian wild populations of C. ariakensis from Korea, Japan, and China using DNA sequence analysis of five concatenated mtDNA regions namely; 16S rRNA, cytochrome oxidase I, cytochrome oxidase II, cytochrome oxidase III, and cytochrome b. Two divergent C. ariakensis clades were identified between southern China and remaining sites from the northern region. In addition, hierarchical AMOVA and pairwise UST analyses showed that genetic diversity was discontinuous among wild populations of C. ariakensis in East Asia. Biogeographical and historical sea level changes are discussed as potential factors that may have influenced the genetic heterogeneity of wild C. ariakensis stocks across this region.

Transferability of cupped oyster EST (Expressed Sequence Tag)-derived SNP (Single Nucleotide Polymorphism) markers to related crassostrea and ostrea species

Single nucleotide polymorphisms (SNPs) are widely acknowledged as the marker of choice for many genetic and genomic applications because they show co-dominant inheritance, are highly abundant across genomes and are suitable for high-throughput genotyping. Here we evaluated the applicability of SNP markers developed from Crassostrea gigas and C. virginica expressed sequence tags (ESTs) in closely related Crassostrea and Ostrea species. A total of 213 putative interspecific level SNPs were identified from re-sequencing data in six amplicons, yielding on average of one interspecific level SNP per seven bp. High polymorphism levels were observed and the high success rate of transferability show that genic EST-derived SNP markers provide an efficient method for rapid marker development and SNP discovery in closely related oyster species. The six EST-SNP markers identified here will provide useful molecular tools for addressing questions in molecular ecology and evolution studies including for stock analysis (pedigree monitoring) in related oyster taxa.

Conserved gene structure and function of interleukin-10 in teleost fish

Interleukin-10 (IL-10) is an important immunoregulatory cytokine produced by various types of cells. Researchers describe here the isolation and characterization of olive flounder IL-10 (ofIL-10) cDNA and genomic organization. The ofIL-10 gene encodes a 187 amino acid protein and is composed of a five exon/four intron structure, similar to other known IL-10 genes. The ofIL-10 promoter sequence analysis shows a high level of homology in putative binding sites for transcription factors which are sufficient for transcriptional regulation ofIL-10. Important structural residues are maintained in the ofIL-10 protein including the four cysteines responsible for the two intra-chain disulfide bridges reported for human IL-10 and two extra cysteine residues that exist only in fish species. The phylogenetic analysis clustered ofIL-10 with other fish IL-10s and apart from mammalian IL-10 molecules. Quantitative real-time Polymerase Chain Reaction (PCR) analysis demonstrated ubiquitous ofIL-10 gene expression in the 13 tissues examined. Additionally, the induction of ofIL-10 gene expression was observed in the kidney tissue from olive flounder infected with bacteria (Edawardsiella tarda) or virus (Viral Hemorrhagic Septicemia Virus; VHSV). These data indicate that IL-10 is an important immune regulator that is conserved strictly genomic organization and function during the evolution of vertebrate immunity.

The IFITM5 mutation c.-14C > T results in an elongated transcript expressed in human bone; And causes varying phenotypic severity of osteogenesis imperfecta type V

Background The genetic mutation resulting in osteogenesis imperfecta (OI) type V was recently characterised as a single point mutation (c.-14C > T) in the 5' untranslated region (UTR) of IFITM5, a gene encoding a transmembrane protein with expression restricted to skeletal tissue. This mutation creates an alternative start codon and has been shown in a eukaryotic cell line to result in a longer variant of IFITM5, but its expression has not previously been demonstrated in bone from a patient with OI type V. Methods Sanger sequencing of the IFITM5 5' UTR was performed in our cohort of subjects with a clinical diagnosis of OI type V. Clinical data was collated from referring clinicians. RNA was extracted from a bone sample from one patient and Sanger sequenced to determine expression of wild-type and mutant IFITM5. Results: All nine subjects with OI type V were heterozygous for the c.-14C > T IFITM5 mutation. Clinically, there was heterogeneity in phenotype, particularly in the manifestation of bone fragility amongst subjects. Both wild-type and mutant IFITM5 mRNA transcripts were present in bone. Conclusions The c.-14C > T IFITM5 mutation does not result in an RNA-null allele but is expressed in bone. Individuals with identical mutations in IFITM5 have highly variable phenotypic expression, even within the same family.

Gene expression profiling reveals a downregulation in immune-associated genes in patients with AS

Objective: To identify differentially expressed genes in peripheral blood mononuclear cells (PBMCs) from patients with ankylosing spondylitis (AS) compared with healthy individuals. Methods: RNA was extracted from PBMCs collected from 18 patients with active disease and 18 gender-matched and age-matched controls. Expression profiles of these cells were determined using microarray. Candidate genes with differential expressions were confirmed in the same samples using quantitative reverse transcription-PCR (qRT-PCR). These genes were then validated in a different sample cohort of 35 patients with AS and 18 controls by qRT-PCR. Results: Microarray analysis identified 452 genes detected with 485 probes which were differentially expressed between patients with AS and controls. Underexpression of NR4A2, tumour necrosis factor AIP3 (TNFAIP3) and CD69 was confirmed. These genes were further validated in a different sample group in which the patients with AS had a wider range of disease activity. Predictive algorithms were also developed from the expression data using receiver-operating characteristic curves, which demonstrated that the three candidate genes have ∼80% power to predict AS according to their expression levels. Conclusions: The findings show differences in global gene expression patterns between patients with AS and controls, suggesting an immunosuppressive phenotype in the patients. Furthermore, downregulated expression of three immune-related genes was confirmed. These candidate genes were also shown to be strong predictive markers for AS.

Association of sporadic chondrocalcinosis with a -4-basepair G-to-A transition in the 5′-untranslated region of ANKH that promotes enhanced expression of ANKH protein and excess generation of extracellular inorganic pyrophosphate

Objective Certain mutations in ANKH, which encodes a multiple-pass transmembrane protein that regulates inorganic pyrophosphate (PPi) transport, are linked to autosomal-dominant familial chondrocalcinosis. This study investigated the potential for ANKH sequence variants to promote sporadic chondrocalcinosis. Methods ANKH variants identified by genomic sequencing were screened for association with chondrocalcinosis in 128 patients with severe sporadic chondrocalcinosis or pseudogout and in ethnically matched healthy controls. The effects of specific variants on expression of common markers were evaluated by in vitro transcription/translation. The function of these variants was studied in transfected human immortalized CH-8 articular chondrocytes. Results Sporadic chondrocalcinosis was associated with a G-to-A transition in the ANKH 5′-untranslated region (5′-UTR) at 4 bp upstream of the start codon (in homozygotes of the minor allele, genotype relative risk 6.0, P = 0.0006; overall genotype association P = 0.02). This -4-bp transition, as well as 2 mutations previously linked with familial and sporadic chondrocalcinosis (+14 bp C-to-T and C-terminal GAG deletion, respectively), but not the French familial chondrocalcinosis kindred 143-bp T-to-C mutation, increased reticulocyte ANKH transcription/ANKH translation in vitro. Transfection of complementary DNA for both the wild-type ANKH and the -4-bp ANKH protein variant promoted increased extracellular PPi in CH-8 cells, but unexpectedly, these ANKH mutants had divergent effects on the expression of extracellular PPi and the chondrocyte hypertrophy marker, type X collagen. Conclusion A subset of sporadic chondrocalcinosis appears to be heritable via a -4-bp G-to-A ANKH 5′-UTR transition that up-regulates expression of ANKH and extracellular PPi in chondrocyte cells. Distinct ANKH mutations associated with heritable chondrocalcinosis may promote disease by divergent effects on extracellular PPi and chondrocyte hypertrophy, which is likely to mediate differences in the clinical phenotypes and severity of the disease.

Brief report: Intestinal dysbiosis in ankylosing spondylitis

Objective Ankylosing spondylitis (AS) is a common, highly heritable immune-mediated arthropathy that occurs in genetically susceptible individuals exposed to an unknown but likely ubiquitous environmental trigger. There is a close relationship between the gut and spondyloarthritis, as exemplified in patients with reactive arthritis, in whom a typically self-limiting arthropathy follows either a gastrointestinal or urogenital infection. Microbial involvement in AS has been suggested; however, no definitive link has been established. The aim of this study was to determine whether the gut in patients with AS carries a distinct microbial signature compared with that in the gut of healthy control subjects. Methods Microbial profiles for terminal ileum biopsy specimens obtained from patients with recent-onset tumor necrosis factor antagonist-naive AS and from healthy control subjects were generated using culture-independent 16S ribosomal RNA gene sequencing and analysis techniques. Results Our results showed that the terminal ileum microbial communities in patients with AS differ significantly (P < 0.001) from those in healthy control subjects, driven by a higher abundance of 5 families of bacteria (Lachnospiraceae [P = 0.001], Ruminococcaceae [P = 0.012], Rikenellaceae [P = 0.004], Porphyromonadaceae [P = 0.001], and Bacteroidaceae [P = 0.001]) and a decrease in the abundance of 2 families of bacteria (Veillonellaceae [P = 0.01] and Prevotellaceae [P = 0.004]). Conclusion We show evidence for a discrete microbial signature in the terminal ileum of patients with AS compared with healthy control subjects. The microbial composition was demonstrated to correlate with disease status, and greater differences were observed between disease groups than within disease groups. These results are consistent with the hypothesis that genes associated with AS act, at least in part, through effects on the gut microbiome.