A blood test may be able to sound early warning bells that patients with advanced melanoma skin cancer are relapsing, according to a study.

Scientists from the Cancer Research UK Manchester Institute studied the DNA shed by tumours into the bloodstream – called circulating tumour DNA – in blood samples from seven advanced melanoma patients at The Christie NHS Foundation Trust.

“Being able to track cancers in real time as they evolve following treatment has huge potential for the way we monitor cancers and intervene to stop them growing back.’ – Professor Peter Johnson, Cancer Research UK’s chief clinician
In this early work they found they could see whether a patient was relapsing by tracking levels of circulating tumour DNA. And they found that new mutations in genes like NRAS and PI3K appeared, possibly causing the relapse by allowing the tumour to become resistant to treatment.

Most melanoma patients respond to treatment at first but their cancer can become resistant within a year. It is hoped that these approaches will allow doctors to use circulating tumour DNA to tailor treatment for individual patients to get the best result.

Around 40 to 50 per cent of melanoma patients have a faulty BRAF gene and they can be treated with the targeted drugs vemurafenib or dabrafenib. But for many of these patients the treatments don’t work, or their tumours develop resistance after a relatively short time. When this happens these patients can be offered immunotherapy drugs including pembrolizumab, nivolumab and ipilimumab. Detecting this situation early could be key to improving their care and chances of survival. Cancer Research UK Manchester Institute

A group of researchers, led by Prof. Matozaki Takashi and Associate Prof. Murata Yoji at the Kobe University Graduate School of Medicine Division of Molecular and Cellular Signalling, were the first to demonstrate the role of stomach cancer–associated protein tyrosine phosphatase (SAP)-1 in the pathogenesis and prevention of Crohn’s disease, ulcerative colitis, and other inflammatory bowel disorders. Their findings are expected to accelerate the development of targeted therapies for inflammatory gastrointestinal diseases.

Inflammatory bowel diseases, such as Crohn’s disease and ulcerative colitis, are disorders of unknown etiology that are often characterized by abdominal pain, diarrhoea, bloody stool, fever, and weight loss. These symptoms frequently interfere with activities of daily living and place patients at an elevated risk of mortality. Patients are also associated with a high risk of developing colorectal cancer. In Japan, there are an estimated 200,000 patients with Crohn’s disease and ulcerative colitis, who qualify for the special Government-led medical assistance system for intractable diseases. Currently, the administration of anti-inflammatory agents only provides palliative results, and the medical community is awaiting new definitive therapies.

Although recent studies have demonstrated that intestinal epithelial cells play a critical role in regulating bowel inflammation, the underlying mechanism remains largely unknown. Previously, Prof. Matozaki, Assistant Prof. Murata, and their colleagues found that SAP-1 localizes to the microvilli of the brush border in gastrointestinal epithelial cells. The transmembrane-type tyrosine phosphatase SAP-1 has an extracellular domain that protrudes into the intestinal lumen and a cytoplasmic domain that mediates tyrosine dephosphorylation of proteins. Here, they showed that SAP-1 ablation in a mouse model of inflammatory bowel disease resulted in a marked increase in the incidence and severity of bowel inflammation, suggesting that SAP-1 plays a protective role against colitis.

In addition, carcinoembryonic antigen-related cell adhesion molecule (CEACAM) 20, an intestinal microvillus-specific membrane protein, was identified as the target of SAP-1 tyrosine dephosphorylation. Suppression of CEACAM20 functions via dephosphorylation was suggested to contribute to preventing colitis. By shedding light on the anti-inflammatory mechanism of the intestinal epithelial cells, Prof. Matozaki and colleagues believe that their findings will drive the development of drugs that target SAP-1 and CEACAM20 to overcome intractable inflammatory bowel diseases. Kobe University

The Beatson Institute of Cancer Research, Bearsden, Glasgow, hosted ‘Circulating Biomarkers 2015’, a Biotexcel conference and workshop. The role of circulating biomarkers in early diagnosis and treatment monitoring is gaining momentum with kits for analysing DNA and RNA from circulating tissue already on the market, and liquid biopsy products in development. The analysis of circulating biomarkers allows less expensive and less invasive screening of patients, and so would enable the early diagnosis of disease and hence timely treatment, for example in pancreatic cancer where presentation typically occurs too late for a cure to be achieved. Also, monitoring of treatment in, for example, breast cancer patients by screening of circulating tumour cells will allow clinicians to make better and quicker decisions about the best therapy for the patient.
In the now familiar format, the meeting included a mix of lectures, a networking workshop, a panel debate, and technology presentations.  The lectures included, among others, presentations on the analysis of circulating tumour DNA (Prof. Charles Coombes, Imperial College London, UK; Dr Gerhardt Attard, Institute of Cancer Research and the Royal Marsden, Surrey, UK), RNA (Prof. Sue Burchill, Leeds Institute of Cancer and Pathology, UK), circulating tumour cells (CTCs) (Dr Vera Cappelletti, National Cancer Institute, Milan, Italy; Dr François-Clément Bidard, Institut Curie, Paris, France) and micro RNA (Dr Alberto Rocci, Manchester Royal Infirmary, Manchester, UK). Other talks discussed the potential of metabolomic biomarkers in cancer (Dr Oliver Maddocks, Beatson Institute for Cancer Research, Glasgow, UK) and how to use a variety of biomarkers and other parameters for the evaluation of the complex  situation of ageing and lifespan (Prof. Paul Shiels, University of Glasgow, Glasgow, UK).
The technology presentations included talks on technology for the enrichment of circulating cell-free DNA (Dr Vipulkumar Patel, Analytik Jena), coupling the CellSearch system (CellSearch) and DEPArray platform (Silicon Biosystems) to isolate single CTCs (by Dr Francesca Fontana (Silicon Biosystems) and targeted biomarker detection by MassARRAY (Dr Malcolm Plant, Agena Bioscience).
The panel debate was centred around the question, ‘CTCs vs. cfDNA vs. miRNA vs. mRNA: which is better and why?’ but perhaps the more interesting discussion that evolved was on the ethics of biomarker analysis (Is it ethical to screen for a condition where there is no treatment ?) and the practicalities of screening (How do we screen for conditions that need to be caught before the presentation of symptoms?). Additionally, the meeting provided many networking opportunities and the benefit of such discussion will, no doubt, be borne out by the development and continuation of new and existing collaborations. A very profitable meeting for all involved.

Demonstrating the potential of precision medicine, an international study based at UT Southwestern Medical Center used next-generation DNA sequencing technology to identify more than 1,000 gene variants that affect susceptibility to systemic lupus erythematosus (SLE).

Precision medicine is an emerging field that aims to deliver highly personalized health care by understanding how individual differences in genetics, environment, and lifestyle impact health and disease.

SLE, commonly called lupus, is a serious, potentially fatal autoimmune disease that the National Institutes of Health reports affects nine times more women than men, and is more likely to strike young African-American, Hispanic, Asian, and Native American women. The disease often begins between the ages of 15 and 44.

“SLE starts when the immune system attacks multiple organ systems in the body, which can result in a complex array of symptoms that are difficult to manage clinically and can lead to organ damage,” said Dr. Edward Wakeland, Chair of Immunology at UT Southwestern and co-senior author of the study posted online recently in the journal eLife. “Our findings support the potential of precision medicine to provide clinically relevant information about genetic susceptibility that may ultimately improve diagnosis and treatment.”

The study also may have implications for other systemic autoimmune diseases, a category of diseases that affect multiple body systems and includes Type 1 diabetes, rheumatoid arthritis, and multiple sclerosis, he said.

Dr. Wakeland and colleagues sequenced millions of DNA base pairs from more than 1,700 people, which allowed precise identification of the genetic variations contributing to SLE, he said. Specifically, the researchers identified 1,206 DNA variations located in 16 different regions of the human genome associated with increased susceptibility to SLE. They then showed that almost all of them (1,199) modify the level of expression of specific molecules that regulate immune responses, he said.

In addition, the two-year study identified many of the specific regulatory variations that were changed in SLE patients and demonstrated that accurately identifying such so-called causal variants increased the accuracy of the genetic association of individual SLE risk genes with susceptibility to SLE.

“Prior to our study, such a comprehensive sequence analysis had not been done and little was known about the exact genetic variations that modify the functions of the genes that cause SLE,” added Dr. Wakeland, who holds the Edwin L. Cox Distinguished Chair in Immunology and Genetics.

The scientists began their comprehensive sequence analysis using the DNA samples of 1,349 American Europeans (773 with SLE disease and 576 without) from sample collections at UT Southwestern, the University of Southern California, UCLA, Oklahoma Medical Research Foundation, and the Université Catholique de Louvain in Belgium.

They then determined the precise DNA sequences at SLE-associated genetic regions scattered throughout the genome. They found that SLE risk is associated with specific clusters of DNA variations, commonly called haplotypes, and that some haplotypes increased the risk for SLE while others provided protection from SLE.

After identifying the sets of DNA variants that increased SLE susceptibility in Caucasians, they used multiple public databases, including the international 1000 Genomes Project (2,504 genomic samples from the global human population) to determine whether these haplotypes also were found in South American, South Asian, African, and East Asian populations.

They discovered that the variants and haplotypes were distributed across subpopulations worldwide. Their findings indicate that many common haplotypes in the immune system are shared at different frequencies throughout the global population, suggesting that these variations in the immune system have ancient origins and persist in populations for long periods, Dr. Wakeland said.

“We thank the many SLE patients and control participants whose sample contributions were essential for these studies,” the researchers wrote. UT Southwestern Medical Center

Tests that can distinguish whether HIV-positive people are infected with a drug-resistant strain or a non-resistant strain allow patients to get the most effective treatment as quickly as possible. A team of Brown University researchers describes a new method that works faster and more sensitively in lab testing than the current standard technologies.

The main advance enabling that improved performance is that the system operates directly on the virus’ more readily available RNA rather than requiring extra, potentially error-prone steps to examine DNA derived from RNA. In a single tube, the system can first combine two engineered probes (ligation) if a mutation is present and then make many copies of those combined probes (amplification) for detection.

“LRA (ligation on RNA amplification) uniquely optimizes two enzymatic reactions — RNA-based ligation, and quantitative PCR (polymerase chain reaction) amplification — into a single system,” said Anubhav Tripathi, professor of engineering at Brown and corresponding author on the paper. “Each HIV contains about 10,000 nucleotides, or building blocks, in its genetic material, and a drop of blood from a patient with resistant HIV can contain thousands to millions of copies of HIV. To find that one virus, out of thousands to millions, which is mutated at just a single nucleotide is like finding a needle in a haystack.”

The experiments reported in the paper show that the LRA test was sensitive enough to find a commonly sought K103N mutation in concentrations as low as one mutant per 10,000 strands of “normal” viral RNA. The LRA detection worked within two hours, while alternative technologies such as ASPCR or pyrosequencing, can take as long as eight.

LRA works by sending in many copies of a pair of short engineered probes of genetic material to complement the RNA in the HIV sample. Under optimized conditions, those pairs that perfectly match the target HIV RNA containing a mutation that causes drug resistance can rapidly become fused together, or ligated, by an enzyme. If there is a single nucleotide difference, the pair won’t fuse.

The fusing of the engineered genetic probes is designed to happen at room temperature. After a short period, the LRA system then heats the slightly alkaline solution, which shuts off the fusing reaction but turns on the amplification (copying) of fused pairs. That allows the LRA system to produce a strong signal of fused pairs, if there are any. All this happens in a single step, without any need to change solution. Brown University