24 research studies from the landmark BLUEPRINT project and IHEC consortia reveal how variation in blood cells’ characteristics and numbers can affect a person’s risk of developing complex diseases such as heart disease, and autoimmune diseases including rheumatoid arthritis, asthma, coeliac disease and type 1 diabetes.

The papers, along with another 17 in high-impact journals, represent the culmination of a five-year, £25 million (€30 million) project that brought together 42 leading European universities, research institutes and industry partners and the work of IHEC. The project’s goals were to explore and describe the range of epigenetic changes that take place in bone marrow as stem cells develop into different types of mature blood cell. It also sought to match epigenetic changes and genetic differences to the physical characteristics of each cell type and use this knowledge to understand how these can lead to blood disorders, cancer and other complex diseases*.

In the first study, Sanger Institute researchers worked closely with colleagues at the University of Cambridge and the University of Oxford to carry out the largest and most in-depth study of DNA and blood cell characteristics using the UK BioBank resource and the INTERVAL study. By comparing almost 30 million DNA sequence differences in more than 173,000 people with variation in the physical properties of blood cells the scientists identified 2,500 previously undiscovered locations in the genome that influence blood cell characteristics and functions. Further work showed that genetic differences affecting some of these characteristics are linked to increased risk of heart attack, or to rheumatoid arthritis and other common autoimmune diseases.

“The scale, resolution and homogeneity of our work were vital. Because we examined so many people we were able to discover important ‘rare and low frequency’ genetic differences that are present in fewer than 10 per cent of the population. We found that these can have a much larger impact on the characteristics of blood cells than the common differences studied previously. Of the more than 300 rare and low frequency difference we found, 74 appear to affect the structure of proteins. These give us important clues as to which biological pathways are involved in controlling the production, function and characteristics of blood cells.”

The team found that genetic differences that cause people to have more young red blood cells in their peripheral bloodstreams also increase the risk they will have a heart attack.

“When mature red blood cells rupture in our blood the body replaces them with new, young red cells – a process known as haemolysis. So we think that increased haemolysis and increased risk of coronary heart disease are affected by the same biological pathways. Identifying these pathways may offer new treatment possibilities.”

Dr Adam Butterworth, one of the study’s senior authors, from the University of Cambridge
‘By combining our detailed genetic information with data from the BLUEPRINT project, we were able to identify with high certainty ‘active’ regions of the human genome that are more likely to be involved in disease mechanisms.’

Heather Elding, one of the paper’s first authors, from the Sanger Institute
For example, in another new finding, the research team showed that genetic differences that increased the amount of certain white blood cells, known as eosinophils, also increased the risk of a person developing rheumatoid arthritis, asthma, coeliac disease and type 1 diabetes.

In the second paper, researchers collaborated with scientists at the University of Cambridge, McGill University in Canada and several UK and European institutions to explore the role that epigenetics plays in the development and function of three major human immune cell types: CD14+ monocytes, CD16+ neutrophils and naïve CD4+ T cells, from the genomes of 197 individuals. They studied the contributions of various genetic control mechanisms, including epigenetic changes such as methyl tags on promoter regions in the DNA and histone modifications, to understand how these different levels of regulation interacted with genetic differences to change the expression of genes, immune function and, ultimately, human disease.

The team identified 345 regions of the genome where they could pinpoint the likely molecular causes underlying a person’s predisposition to immune-related diseases such as inflammatory bowel disease, type 1 diabetes and multiple sclerosis.

“We have created an expansive, high-resolution atlas of variations that deepens our understanding of the interplay between the genetic and epigenetic machinery that drives the three primary cells of the human immune system. We have identified hundreds of genetic variations associated with autoimmune diseases that appear to affect the activity of genes in specific regions of the genome, pointing to biological pathways that may be involved in disease and which, ultimately, may be treatable with medication.”

Sanger Institute www.sanger.ac.uk/news/view/landmark-project-shows-heart-disease-and-rheumatoid-arthritis-risk-raised-genetic-changes

Researchers from the University of Wisconsin School of Medicine and Public Health and Carbone Cancer Center have better defined a pro-growth signalling pathway common to many cancers that, when blocked, kills cancer cells but leaves healthy cells comparatively unharmed.

The study could establish new avenues of therapeutic treatments for many types of solid tumours.

Growth signals typically come in the form of chemical agonists outside of cells that bind to protein receptors on cells. Activated receptors are responsible for transmitting the signal to the inside of the cell, ultimately generating a growth messenger called PIP3.

Two years ago, research out of UW–Madison professor Richard Anderson’s lab found that some of these agonist-stimulated receptors continue to transmit the signal even after they have been pulled into the cell, sequestered in vesicles called endosomes and presumably on their way to being degraded.

“According to dogma in the literature, receptors shouldn’t make PIP3 at these internal sites, but they were,” Anderson says. “We set out to ask, ‘Why is that?’”

In this new study, a postdoctoral fellow in Anderson’s lab, Suyong Choi, showed that the proteins known to be in this signal transmission cascade were all present on endosomes inside the cell, supporting the idea that the key growth message was being signalled from these internal compartments.

However, there was one fact which they could not biologically explain: In a typical signalling cascade, each step amplifies the signal, suggesting there should be more and more of the messenger molecules; but here, levels of PIP3 and other intermediary messengers were too low to be detected in endosomes.

“A scaffold completely solves this issue, because it acts like an assembly line, bringing together all of the proteins and passing one messenger molecule to the next protein in the cascade until the last protein, PI3K, is activated and generates PIP3,” Anderson says. “Suyong Choi found that the scaffolding protein IQGAP1 brings all of these proteins together like a happy family on the endosome. It’s an incredibly efficient mechanism.”

Choi discovered that the IQGAP1 complex pulls together all of the signalling components in the PI3K pathway. Remarkably, this assembly happens in response to nearly all agonists that switch on growth and cell survival signals in cells. Once Choi had established how the proteins in the complex interacted, he was able to block scaffold formation in cells by adding a small, competing fragment of the IQGAP1 protein.

“It worked beautifully to block assembly of IQGAP1 and PI3K complex,” Anderson says. “The really cool thing was, when we treated different cells with these inhibitory fragments, the disruption of IQGAP1 and PI3K complex formation had almost no effect on normal cells but it killed cancer cells very efficiently.”

PI3K is an essential protein, and cells (and whole organisms) die if they do not have any functional PI3K because the protein is involved in multiple signalling pathways. However, it is specifically this pathway, mediated through IQGAP1, that is required for the growth and survival of cancer cells but not normal cells. In fact, mice lacking IQGAP1 develop normally but are resistant to developing solid tumours.

“Pharmaceutical companies have developed PI3K inhibitors, but many of these have failed, likely because they’re hitting all PI3Ks and the different pathways,” Anderson said. “If you can specifically disrupt this agonist-activated PI3K pathway, the one that has a specific role in cancer, then you can effectively treat

University of Wisconsin School of Medicine and Public Health www.med.wisc.edu/news-events/cancer-signaling-pathway-could-lead-to-new-cancer-therapies/49720

A multi-institutional group of researchers, led by investigators at Children’s Hospital Los Angeles and the University of Michigan, have identified a simple and inexpensive tool for assessing the prognosis of paediatric brain tumours called ependymomas. Their study, which demonstrates the epigenetic mechanism behind these tumours, may offer future opportunities for novel therapeutic options.

Childhood posterior fossa ependymomas (PF) are tumours found largely in the hind brain (consisting of the cerebellum, pons and the brainstem) of children. Routine assessment of tumour grade and other markers in PF ependymomas do not correlate well with outcomes in these tumours, highlighting the need for new prognostic markers. Genomic sequencing efforts have not identified mutations in these tumours, and the origin of PF ependymomas remains obscure.

While lacking recurrent genetic mutations, a subset of these tumours exhibit alterations in DNA methylation. In this study, the researchers looked at modification of histones – protein components of the chromatin around which DNA winds, and which play a role in gene regulation – in particular, histone H3.

Co-lead investigator, Sriram Venneti, MD, PhD, of the Department of Pathology at the University of Michigan, observed that histone H3 is modified differently in paediatric posterior fossa ependymoma. Specifically, 80 percent of these tumours exhibited loss of the H3K27me3 a repressive mark, while 20 percent of tumours retained H3K27me3.  Researchers went back and looked at MRIs and outcomes of children treated for these tumours and identified that tumours with loss of H3K27me3 tumours behaved more aggressively and showed poor overall survival.  This suggests that reduced H3K27me3 may be a prognostic indicator in PF ependymomas.

“Detection of H3K27me3 by immunohistochemical staining is a widely available and cost effective surrogate molecular marker.  This test can be readily implemented in most departments of pathology and provides a much-needed tool to risk stratify and identify ependymoma patients who would potentially benefit from epigenetic therapies,” said co-lead investigator Alexander R. Judkins, MD, of the Department of Pathology and Laboratory Medicine at CHLA and Keck School of Medicine of the University of Southern California.

This loss in H3K27me3, along with other epigenetic changes, was similar to that observed in another type of paediatric brain tumour of the hind brain region termed diffuse intrinsic pontine gliomas (DIPGs).  This suggests that both of these tumours arise from similar epigenetic states. Intriguingly, researchers found that certain progenitor cells in this part of the brain also showed low H3K27me3, suggesting – as both tumours share epigenetic similarities – that low methylation of H3K27me3 is important to the development of tumours in this region of the brain.

Children’s Hospital of Los Angeles www.chla.org/press-release/biomarker-identified-aid-prognosis-pediatric-ependymomas

Bleeding disorders could one day be diagnosed by putting platelets through strength tests, researchers have proposed. Biomedical engineers from Emory University and the Georgia Institute of Technology have devised a microfluidic testing ground where platelets can demonstrate their strength by squeezing two protein dots together. Imagine rows and rows of strength testing machines from a carnival, but very tiny. A platelet is capable of exerting forces that are several times larger, in relation to its size, than a muscle cells.
After a blood clot forms, it contracts, promoting wound closure and restoration of normal blood flow. This process can be deficient in a variety of blood clotting disorders. Previously, it was difficult to measure an individual platelet’s contributions to contraction, because clots’ various components got in the way.
“We discovered that platelets from some patients with bleeding disorders are ‘wimpier’ than platelets from healthy people,” says Wilbur Lam, an assistant professor in the Department of Pediatrics at Emory University School of Medicine and in the Wallace H. Coulter Department of Biomedical Engineering at Georgia Tech and Emory University. “Our device may function as a new physics-based method to test for bleeding disorders, complementary to current methods.”
The first author of the paper is David Myers, an instructor at Emory’s medical school. Lam is also a physician in the Aflac Cancer and Blood Disorders Center, Children’s Healthcare of Atlanta.
The scientists infer how strong or wimpy someone’s platelets are by measuring how far the protein dots move, taking a picture of the rows of dots, and then analysing the picture on a computer.
The dots are made of fibrinogen, a sticky protein that is the precursor for fibrin, which forms a mesh of insoluble strands in a blood clot.
In addition to detecting problems with platelet contraction in patients with known inherited disorders such as Wiskott Aldrich syndrome, Myers, Lam and colleagues could also see differences in some patients who had bleeding symptoms, but who performed normally on standard diagnostic tests.
The researchers also used chemical tools to dissect the process of platelet contraction. They showed that inhibitors of Rho/ ROCK enzymes shut down platelet contraction, but inhibitors of a related pathway, MLCK (myosin light chain kinase), did not. Individual platelet contraction could become an assay for development or refinement of blood thinning drugs, Lam says.

Georgia Techhttp://tinyurl.com/j8byzzg