A recent study suggests the use of new protocols for improving the diagnostic accuracy of miRNA measurement in human blood samples.
Research carried out by scientists from Chicago Medical School (IL, USA), the University of California (CA, USA), Rosalind Franklin University of Medicine and Science (IL, USA) and Duke University produced recommendations to be considered when using protocols to study miRNA measurement. The protocols could improve sensitivity of miRNA detection up to 30-fold. The paper was recently published in the January 2012 issue of the Journal of Molecular Diagnostics.
Measurement of miRNA biomarkers could aid disease diagnosis; however, a number of factors currently impair the accuracy of miRNA testing. “A fundamental challenge to making miRNA diagnostics broadly available has been the inability to isolate enough high-quality material to analyze,” explains coauthor Sarah Linnstaedt from the Department of Molecular Genetics and Microbiology at Duke University Medical Center (NC, USA). “Our paper outlines ways of effectively collecting blood plasma samples, thus bringing us one step closer to the goal of making miRNA disease diagnostics a reality.”
In this study, miR-16 and miR-223 were quantified in healthy adult blood samples. These molecules have been identified as possible biomarkers for various diseases and conditions; miR-16 is a tumor suppressor found to be lost or deregulated in various cancers, including breast cancer, whilst miR-223 may be associated with some malignant diseases and pregnancy.
Blood collection tubes containing sodium fluoride and potassium oxalate were found to give the most accurate miRNA measurement. Quantitation of miR-223 was more accurate when quantified in plasma samples as opposed to serum samples, highlighting that the type of blood sample can affect miRNA measurement.
The protocol also includes extra purification steps and an optimum dilution level in order to reduce interference from natural components of blood plasma, which can copurify with the target miRNA. “By reducing the starting material, inhibitors were presumed to be diluted below a threshold of interference. Careful titration of starting material yields more accurate miRNA quantitation,” explains corresponding author Dominik Duelli of the Department of Cellular and Molecular Pharmacology at Chicago Medical School, and Rosalind Franklin University of Medicine and Science.
Researchers also found that interindividual differences in plasma composition yielded different miRNA measurements, suggesting that factors which affect blood chemistry (such as diet, exercise and circadian rhythms) may also confound miRNA measurement.
“The implications of this work are that without consideration of the variables we have identified, miRNA quantitation of human samples may not be reliable for the purpose of biomarker development,” concludes Duelli. “We provide approaches that enable faithful quantitation of miRNA abundance in body fluid.”
– Written by Sarah Miller
Source: Kim DJ, Linnstaedt S, Palma J et al. Plasma components affect accuracy of circulating cancer-related microRNA quantitation. J. Mol. Diagn. 14(1), 71–80 (2012).
In a novel, proof-of-concept study, researchers at the Hebrew University of Jerusalem (Israel) have revealed, for the first time, a clear-cut epigenetic signature for Type 2 diabetes (T2D). The genome-wide survey also revealed tell-tale methylation signature marks on the DNA of young subjects who later developed impaired glucose metabolism, prior to the appearance of clinical diabetic manifestations. The research is published in the journal Human Molecular Genetics.
The research team was led by Asaf Hellman of the Hebrew University‘s Institute of Medical Research Israel–Canada, who spoke to Epigenomics about the significance of the study: “This research has provided the first proof of molecular epigenetic risk factors leading to T2D, providing an ‘early warning‘ sign that could lead to new approaches to treating this, and other human disease conditions.”
Hellman argues that despite the extensive research being made into the molecular basis of variance in interindividual susceptibility to common diseases, the subject is still poorly understood, with the prime example being T2D. The aim of the present study was therefore to characterize susceptibility to the disease.
Taking an innovative research direction, the Hebrew University research team decided to map DNA methylation variation rather than traditional DNA sequence variation in 1169 T2D patients and nondiabetic controls. The results demonstrated the unique abilities of this novel research approach by revealing a clear-cut, disease predisposing DNA methylation signature. This is, as Hellman points out, one of the first reports in the scientific literature of an epigenetic risk factor for T2D.
Hellman is hopeful that by using this and similar research tools, “we are closing in on providing a clear test for the long-standing hypothesis that risk for common human diseases includes a significant epigenetic component”. He continues: “Given that epigenetic marks could be modified by drugs, this means that 10 years from now we will either be deeply involved in developing new epigenetic diagnostic/prevention/intervention agents, or perhaps be recognizing that epigenetics is not so important after all and move on in other directions. Right now I would say that both scenarios are equally possible”.
What is clear is that these findings will shed new light on the mechanism of individual predisposition to T2D and will pave the way for elucidating similar mechanisms in a long list of common human diseases, including many metabolic, autoimmune and psychiatric disorders.
– Written by Natalie Harrison
Source: Toperoff G, Aran D, Kark JD et al. Genome-wide survey reveals predisposing diabetes Type 2-related DNA methylation variations in human peripheral blood. Hum. Mol. Genet. 21(2), 371–383 (2012).
SciClips (WI, USA), a web-based company that provides a worldwide scientific platform to facilitate research collaboration and open innovation among scientists, has recently launched a new comprehensive oncology biomarker database with cancer companion diagnostic pathway maps, that is hoped to aid the discovery and prognostic potential, as well as clinical validation, of cancer biomarkers. As well as including protein, miRNA, SNP, gene-expression, epigenetic and metabolomic biomarkers, the database will help develop innovative theranostics tools (i.e., strategies and tools that can be used for individualized diagnosis and treatment).
The database includes more than 8700 discovery-phase (i.e., preclinical or clinical) cancer biomarkers from a variety of patient-derived samples, such as blood/serum, tumor, urinary, fecal, saliva and breath cancer biomarkers, for more than 100 different cancer types. Furthermore, patient/sample details, experimental results/observations, biological and molecular functional analysis, biological process analysis, chromosomal location and protein–protein interaction networks of each biomarker are also listed.
For each cancer type, biomarkers are classified into numerous categories: diagnostic biomarkers, disease predictive/risk assessment biomarkers (e.g., for predicting the risk or genetic susceptibility in developing cancer), drug efficacy/response biomarkers (i.e., predictive of anticancer treatments), prognostic biomarkers (e.g., for determining the clinical outcome in cancer patients) and cancer companion diagnostics biomarker pathway.
Founder of SciClips, Sanchayita Kar, commented that, “Future success in developing innovative personalized cancer drugs lies in adopting new differential diagnostic procedures along with companion biomarker assays to identify the most appropriate cancer patient, tumor type and disease state. Our database will be a useful tool for the discovery of new cancer biomarkers as well as clinical validation of biomarkers.”
The need for a comprehensive cancer biomarker database has evolved in accordance with the increased expenditure into biomarker research and personalized medicine.
– Written by Thomas Payne
Source: Cancer theranostics: potential applications of cancer biomarker database: www.sciclips.com/sciclips/blogArticle.do?id=1017&blog=Cancer
Researchers at The Scripps Research Institute (FL, USA) have, for the first time, shown that gene-expression changes in the brain associated with psychiatric disorders can result from histone acetylation. The finding that DNA stays too tightly wound in certain brain cells of schizophrenic subjects was found to be especially pronounced in younger subjects, suggesting that treatment might be most effective early on at minimizing or even reversing the symptoms of schizophrenia, a potentially devastating mental disorder associated with hallucinations, delusions and emotional difficulties. The findings are available online in the new Nature journal, Translational Psychiatry.
Speaking to Epigenomics about why this research is so significant, Scripps Research Associate Elizabeth Thomas, a neuroscientist who led the study, states: “Overall, our studies support the notion that schizophrenia etiology involves not only genetic factors, but epigenetic factors as well. This has enormous implications for novel therapeutic approaches for schizophrenia, and possibly other psychiatric disorders.”
Thomas worked with lead author Bin Tang, a postdoctoral fellow in her laboratory, and Brian Dean, an Australian colleague at the University of Melbourne (Australia), to obtain post-mortem brain samples from both schizophrenic and healthy brains from medical ‘brain banks‘ in the USA and Australia. The group then developed a technique to maintain histone–DNA interactions.
Not only did the researchers prove that these interactions are preserved in post-mortem brain samples, but they found that in comparison to healthy brains, the samples from schizophrenic subjects had lower levels of acetylation in certain histone portions that would block gene expression. As Thomas explains, “because the deficits in histone acetylation were detected most robustly in subjects with early-stage illness (4 years or less from initial diagnosis), this implicates an imbalance in epigenetic gene regulation as pathogenic to disease and supports a role for early treatment (with histone decetylase [HDAC] inhibitors) to prevent or even reverse symptoms of the disease.”
Thomas argues that “such treatment approaches would target a root cause of the disease, rather than simply treating symptoms,” and would represent a “much improved therapeutic approach over currently used antipsychotic drugs,” which are associated with numerous side effects (i.e., movement disorders, dystonias, metabolic disturbances and weight gain). Furthermore, because novel HDAC inhibitors are being widely explored as therapeutics for many neurological disorders, their clinical use for schizophrenia could be rapidly accelerated.
Speaking to Epigenomics about the wider impact of these results, Thomas speculates: “The translational aspect of epigenomics is greater now than ever before, as more and more human disorders are being linked to epigenetic imbalances, and increasing numbers of researchers are investigating what role these modifications play in health and disease. Our results directly merge with this translational appeal by implicating schizophrenia as another human disease associated with epigenomic disturbances”.
“I think the next 10 years will witness an explosion of epigenetic drugs that will be used to treat disorders from Huntington‘s disease to obesity,” predicts Thomas.
The Scripps neuroscientist would now like to expand her team‘s findings to identify differences in genome-wide histone acetylation marks in subjects with schizophrenia, rather than just specific genomic loci, as they did in the present study. She would also like to validate the current findings in additional cohorts of subjects, which could then provide a basis for clinical application of HDAC inhibitors for human patients.
Furthermore, it is Thomas‘ aim to further measure histone acetylation marks in peripheral tissues (i.e., blood cells and fibroblasts) of subjects with schizophrenia and other psychiatric disorders, in order to identify potential biomarkers that could be used to classify subsets of patients that may have more pronounced epigenetic imbalances. Such biomarkers could also be useful to monitor treatment paradigms using epigenetic drugs.
– Written by Natalie Harrison
Source: Tang B, Dean B, Thomas EA. Disease- and age-related changes in histone acetylation at gene promoters in psychiatric disorders. Transl. Psychiatry doi:10.1038/tp.2011.61 (2011) (Epub ahead of print).