In a Journal of Molecular Diagnostics paper published online in advance this week, the University of Toronto's Hilmi Ozcelik and his colleagues show that a next-generation sequencing-based approach — based on long-range PCR and deep sequencing — is suitable for BRCA1/2 mutation analysis. "The first-generation of BRCA1/2 mutation analysis targeted only the coding exons and has implicated protein-truncating mutations — indel, nonsense — in BRCA1/2 inactivation," Ozcelik et al. write. In this study, they describe and validate their next-generation sequencing-based approach using genomic DNA from 12 familial breast cancer patients. "NGS successfully identified all 19 distinct — 51 total — BRCA1 and 35 distinct — 63 total — BRCA2 sequence alterations detectable by the Sanger sequencing, with no false-negative or positive results," the researchers add. "These results illustrate that NGS can provide comprehensive genetic information more quickly, accurately, and at a lower cost than conventional approaches, and we propose NGS to be a more effective method forBRCA1/2 mutational analysis."

The Sample's sister publication, Cancer Minute, has more on this study here.

In Experimental and Molecular Pathology, researchers at the Geisel School of Medicine at Dartmouth College report on chimerism analyses for recipients of allogeneic stem cell transplants. PCR analysis of short tandem repeats are commonly used in allogeneic transplant recipients, the team says, and are generally performed on blood and marrow aspirates, but it is unknown whether it is necessary to look at both. In this study, the researchers performed a retrospective analysis of 42 consecutive adult allogeneic stem cell transplant recipients, and found that PCR analyses of STRs for chimerism performed on unfractionated blood highly "correlated with results obtained using unfractionated marrow aspirates at 30, 60, or 90 days following transplant." Further, the authors add, "overall and relapse-free survival of patients experiencing full donor chimerism was not statistically different from patients demonstrating mixed chimerism at days 30, 60, and 90 following [stem cell transplant]." This suggests that chimerism analyses may be assessed on peripheral blood alone, they write.

Also in Experimental and Molecular Pathology, researchers in Argentina report on the ability of N-(2-mercaptopropionyl)-glycine, or MPG, to protect against ischemia-reperfusion injury in hypertrophied hearts. The team isolated hearts from spontaneously hypertensive rats and age-matched normotensive rats, and the subjected the hearts to 50-minute global ischemia and two-hour reperfusion. "The treatment with MPG decreased infarct size, preserved [glutathione] levels and decreased SOD and MnSOD cytosolic activities, [thiobarbituric acid reactive substances] concentration, and H2O2 induced-mPTP opening in both rat strains," the authors write. "Our results show that, in both hypertrophied and normal hearts, an attenuation of mPTP opening via reduction of oxidative stress appears to be the predominant mechanism involved in the cardioprotection against reperfusion injury MPG-mediated."

Carlos Gómez-Martin at the Spanish National Cancer Research Center and his colleagues write in the Journal of Clinical Pathology that HER2 gene amplification is associated with gastric cancer patient survival. They examined HER2 gene amplification and overexpression in tumor samples from 148 gastric cancer patients and correlated that data with patient characteristics. That HER2 amplification correlated with patient survival suggests that "HER2 gene amplification approaches might be an optimal HER2/neu testing strategy for the selection of HER2+ GC patients who are candidates to be treated with anti-HER2 therapies," Gómez-Martin et al. write.

Also in the Journal of Clinical Pathology, researchers led by Ki-Seok Jang from South Korea's Hanyang University report that the loss of SIRT1 histone deacetylase expression is associated with colorectal adenocarcinoma tumor progression. Jang and his colleagues found that normal mucosa expressed SIRT1, while about 80 percent of adenomatous polyps, around 40 percent of colorectal adenocarcinoma, and about 35 percent of metastatic tissue expressed SIRT1. "SIRT1 expression decreases stepwise during colorectal carcinogenesis ... and as a function of adverse prognostic factors such as regional LN metastasis and tumor stage," the researchers write. "Our data also suggest that loss of SIRT1 is associated with the MSI [microsatellite instability]-high phenotype."

Researchers led by Ming Guan from Shanghai Medical School report in Modern Pathology that DLC1 methylation and mutations in PIK3CA may play important roles in the development of Paget's disease, a rare cutaneous malignant neoplasm of the breast. They examined tissue biopsies from 132 patients with extramammary Paget's disease for methylation levels of DLC1 CpG islands using both high-resolution melting analysis and bisulfite DNA sequencing, and for PIK3CA mutations using direct DNA sequencing. From that, they "found a high prevalence of downregulation and hypermethylation of the DLC1 gene in cases of this rare disease (33 and 39%, respectively). In addition, activating PIK3CA mutations were detected in 24% of the extramammary Paget's disease cases." This, the researchers add, shows "evidence that epigenetic and genetic alterations are common in extramammary Paget's disease and may be involved in the disease pathogenesis."

Also in Modern Pathology, the National University of Singapore's Chee-Seng Ku and colleagues review how advances in sequence capture and next-generation sequencing have led to progress in understanding familial diseases and how such approaches can be applied to better characterize colorectal cancer type X, a familial cancer. "In the context of familial colorectal cancer type X, we believe that the disease model and its genetic basis are likely to become more apparent when the approaches that we have outlined and discussed are applied in practice," Ku et al. write. "This should facilitate the iterative interrogation of the genetics of familial colorectal cancer type X and other familial cancers of similar nature before embarking on either a comprehensive genome-wide association studies or whole-genome sequencing approach."

Researchers led by Paul Ridker at Brigham and Women's Hospital show that lipoprotein-associated phospholipase A₂ mass, but not activity, is independently associated with cardiovascular disease. As they report in Clinical Chemistry, the researchers examined the link between Lp-PLA2 mass and activity with cardiovascular disease in 1,821 cases and 1,992 controls. However, the researchers note that the "the clinical utility of Lp-PLA2 mass remains questionable due to a lack of [risk] model improvement. More important are the high absolute levels for Lp-PLA2 mass seen in these data. The lack of standardization for this assay severely limits its potential clinical use."

In a Clinical Chemistry editorial, Boston Children's Hospital's Greg Miller and colleagues caution that using commercially available immunoassays for biomarker research can be problematic, especially if manufacturers do not provide adequate information about their assays. Miller and his colleagues say that researchers should examine inserts posted to company's websites prior to purchase and that such inserts should include information regarding calibration and assay performance, which users should confirm. "Both the manufacturers of assay kits and the researcher who uses them are responsible for assuring that the analytical quality of the assay is suitable for the intended use," the authors write, adding that "failure to address these matters will hinder our ability to conduct valid studies of biomarkers, and that may lead to serious errors in the evaluation of candidate biomarkers."