Circulating DNA is a non-invasive prognostic factor for survival in non-small cell lung cancer.

INTRODUCTION: Circulating plasma DNA is present in a considerably higher concentration in lung cancer patients than in controls. Conflicting data are reported about circulating DNA as a prognostic factor. The aim of this study was to prospectively analyse the relationship of circulating plasma DNA with overall survival (OS) of previously untreated non-small cell lung cancer (NSCLC) patients. METHODS: 46 untreated NSCLC patients and 21 controls with a follow-up time of 6.5 years were analyzed. Quantification of baseline circulating plasma DNA was performed by a real-time quantitative polymerase chain reaction (qPCR) targeting the human beta-globin gene. Survival analysis was performed using the Kaplan-Meier method and compared with a Cox-regression analysis. RESULTS: The median DNA concentration of the patients who died (87%) was significantly higher compared to the patients that survived at the end of follow-up (55ng/ml versus 23ng/ml, p=0.02). In patients with higher DNA concentration overall survival was significantly worse. In this study no relation of DNA concentration with tumour characteristics, age, gender or pulmonary inflammatory conditions was found. CONCLUSION: In this study a high circulating plasma DNA concentration at time of diagnosis in NSCLC patients was a prognostic factor for poorer survival. Circulating DNA may be used as a non-invasive biomarker to refine the prognostic profile in NSCLC patients.

The silent hemoglobin alpha chain variant Hb Riccarton [alpha51(CE9)Gly-->Ser] may affect HbA1c determination on the HLC-723 G7 analyzer.

BACKGROUND: Structural hemoglobin variants can affect the accuracy of hemoglobin A1c (HbA1c) testing and represent the most common pitfall in the determination of HbA1c. We here describe the characterization of an alpha chain variant in diabetic patients as the cause of an abnormal presentation of the HbA1c fraction on the HLC-723 G7 analyzer. METHODS: HbA1c analysis was performed using various HPLC-based HbA1c analyzers and by immunoassay. alpha-Globin mutation analysis was performed by GAP-PCR and DNA sequencing. RESULTS: The peak partially overlapping HbA1c in the chromatogram represents the glycated fraction of the silent alpha chain variant Hb Riccarton [alpha51(CE9)Gly-->Ser]. This aberrant peak is uniquely identified by the HLC-723 instrument, as it is not observed on other HPLC-based HbA1c analyzers. Occasionally, the HLC-723 may fail to properly integrate both glycated Hb fractions, resulting in a falsely low HbA1c result. The variant was confirmed in samples from other diabetic patients with identical chromatographic patterns. CONCLUSIONS: The silent alpha chain variant Hb Riccarton [alpha51(CE9)Gly-->Ser] leads to an abnormal chromatographic presentation on the HLC-723 analyzer with a risk of erroneous HbA1c determination. Manual validation of chromatograms to detect abnormalities caused by Hb variants is important to prevent incorrectly produced HbA1c results from being reported.

Improved real-time detection of the H63D and S65C mutations associated with hereditary hemochromatosis using a SimpleProbe assay format.

BACKGROUND: Genetic predisposition to hemochromatosis involves several different point mutations in the HFE gene. Routine testing for two such mutations (H63D and S65C) using real-time genotyping deserves special care, as these mutations are in close proximity (6 bp) of each other. METHODS: A novel assay was designed for these two mutations based on a SimpleProbe assay format that allows for more flexibility in assay design, as it requires only one detection probe instead of the two probes that are commonly used with a hybridization probe based assay. RESULTS: The SimpleProbe assay format yielded data that were easily interpreted without significant optimizing efforts. CONCLUSIONS: The SimpleProbe assay format offers some unique advantages compared to hybridization probes and offers a robust and interesting alternative to hybridization probes in the detection of genetic variations. This is the first time that the use of this assay format is described in the clinical literature.

The prognostic role of the STK15 T91A polymorphism and of STK15 mRNA expression in patients with urothelial cell carcinoma.

BACKGROUND: The prognostic role of the STK15 T91A polymorphism and of STK15 mRNA expression was investigated in patients with urothelial cell carcinoma (UCC). MATERIALS AND METHODS: The STK15 genotype with respect to the T91A polymorphism was assessed by restriction fragment length polymorphism in 135 patients. STK15 mRNA expression was measured in tumor tissues of 103 patients, using real-time quantitative PCR. RESULTS: The T91A polymorphism lacked any prognostic information in our patient cohort. Interestingly though, STK15 mRNA expression was increased in invasive and high-grade tumors (p-values of 0.009 and 0.0001, respectively). Additionally, patients with superficial UCC (n = 82) who had a tumor recurrence in the first year after surgery displayed elevated STK15 mRNA expression levels (p = 0.009). Kaplan-Meier survival analysis revealed an increased risk of tumor progression for patients with Ta tumors (n = 62) and high STK15 expression (log-rank p = 0.04). Furthermore, a decreased overall (log-rank p = 0.006) and UCC-specific survival (log-rank p = 0.001) were shown for patients with elevated STK15 mRNA levels. CONCLUSION: Patients with UCC and elevated levels of STK15 mRNA generally showed a more adverse disease course than patients with low levels. This may help in identifying patients in need of more aggressive treatment.