Detection of epidermal growth factor receptor mutations in formalin fixed paraffin embedded biopsies in Malaysian non-small cell lung cancer patients

Background

Somatic mutations of the epidermal growth factor receptor (EGFR) are reportedly associated with various responses in non-small cell lung cancer (NSCLC) patients receiving the anti-EGFR agents. Detection of the mutation therefore plays an important role in therapeutic decision making. The aim of this study was to detect EGFR mutations in formalin fixed paraffin embedded (FFPE) samples using both Scorpion ARMS and high resolution melt (HRM) assay, and to compare the sensitivity of these methods.

Results

All of the mutations were found in adenocarcinoma, except one that was in squamous cell carcinoma. The mutation rate was 45.7% (221/484). Complex mutations were also observed, wherein 8 tumours carried 2 mutations and 1 tumour carried 3 mutations.

Conclusions

Both methods detected EGFR mutations in FFPE samples. HRM assays gave more EGFR positive results compared to Scorpion ARMS.     

Predatory ability of generalist predators on eggs,young nymphs and adults of the invasive Halyomorpha halys in southern Europe

Halyomorpha halys (Stål, 1855) is an invasive pest causing serious damage to agricultural crops in Europe and the USA. Very little is known about H. halys predators in Europe. This survey evaluated the potential of generalist predators/omnivorous species by means of predation bioassays in tri-dimensional arenas, where the predator had to locate H. halys prey items on the leaves of a bean plant. Eleven species of different taxa were tested and the prey items consisted in fresh eggmasses, 1st and 2nd instar nymphs. One species was also tested against adults. Some predators were species commercially available as biocontrol agents against plant pests, other predators were wild, captured in habitats shared with H. halys. All tested specimens were starved 24 h before starting the experiment. The survivorship of control prey items in predator-excluding cages was compared to that of predator treatment groups to determine the effect of predator presence. According to the results, the generalist species showed a quite low acceptance of H. halys prey items, since only two species caused 80% mortality on at least one item (Eupholidoptera chabrieri and Rhynocoris iracundus) and mortality due the other species never exceed 60%. Among commercially available species only Adalia bipunctata adults and Chrysoperla carnea larvae were effective, predating the eggs and 1st instar nymphs, respectively. Among the field collected specimens, the orthopteran E. chabrieri and the predatory hemipterans R. iracundus, Nagusta goedelii and Himacerus mirmicoides showed efficacy against 1st instar nymphs, E. chabrieri and R. iracundus showed efficacy against 2nd instar nymphs, whereas only E. chabrieri and N. goedelii predated the eggs. R. iracundus was also tested on the adults and successfully predated them. By identifying some of the species that can exploit H. halys as a suitable prey in southern Europe, the present investigation provides an important contribution for conservation biological control of this pest.

     

miRNA-148a and miRNA-30c expressions as potential biomarkers in breast cancer patients

BackgroundBreast cancer is an extensively identified malignant tumor and is a prime cause of cancer mortalities in females. It has been shown that alteration of miRNAs expression (up or down regulation) can affect the initiation and progression of many malignancies. We aimed to evaluate the role of circulating miRNA-148a and miRNA-30c in female patients with breast cancer and estimate their usage as potential biomarkers in the diagnosis, prognosis and survival of breast cancer.MethodsThis study included 75 breast cancer female patients.They were compared with 55 apparently healthy female subjects. miRNAs expression analysis was assessed via real-time PCR.ResultsTo discriminate breast cancer patients from controls, miR-30c showed the best performance at a cut off value of ≤20.6 (AUC = 0.998, 97.33% sensitivity, 96.36% specificity, p < 0.001), followed by miR-148a (AUC = 0.995, 94.67% sensitivity, 90.91% specificity, p < 0.001 at a cut off value of ≤0.1), CA 15-3 (AUC = 0.930, 88.0% sensitivity, 81.82% specificity, p < 0.001 at a cut off value of >21.3), and finally CEA (AUC = 0.751, 70.67% sensitivity, 63.64% specificity, p < 0.001 at a cut off value of >2.5).ConclusionmiRNA-148a and miRNA-30c expressions were down regulated in female patients with breast cancer and might be considered as potential blood biomarkers. Both also might have rule in disease treatment and selection of therapeutic targets. Future studies are needed to improve their role in predicting response to treatment and prognosis.     

Characterization of the diversity and temporal stability of bacterial communities in human milk

Recent investigations have demonstrated that human milk contains a variety of bacterial genera; however, as of yet very little work has been done to characterize the full diversity of these milk bacterial communities and their relative stability over time. To more thoroughly investigate the human milk microbiome, we utilized microbial identification techniques based on pyrosequencing of the 16S ribosomal RNA gene. Specifically, we characterized the bacterial communities present in milk samples collected from 16 women at three time-points over four weeks. Results indicated that milk bacterial communities were generally complex; several genera represented greater than 5% of the relative community abundance, and the community was often, yet not always, stable over time within an individual. These results support the conclusion that human milk, which is recommended as the optimal nutrition source for almost all healthy infants, contains a collection of bacteria more diverse than previously reported. This finding begs the question as to what role this community plays in colonization of the infant gastrointestinal tract and maintaining mammary health.     

Evaluation of methods for the extraction and purification of DNA from the human microbiome

Background

DNA extraction is an essential step in all cultivation-independent approaches to characterize microbial diversity, including that associated with the human body. A fundamental challenge in using these approaches has been to isolate DNA that is representative of the microbial community sampled.

Methodology/Principal Findings

In this study, we statistically evaluated six commonly used DNA extraction procedures using eleven human-associated bacterial species and a mock community that contained equal numbers of those eleven species. These methods were compared on the basis of DNA yield, DNA shearing, reproducibility, and most importantly representation of microbial diversity. The analysis of 16S rRNA gene sequences from a mock community showed that the observed species abundances were significantly different from the expected species abundances for all six DNA extraction methods used.

Conclusions/Significance

Protocols that included bead beating and/or mutanolysin produced significantly better bacterial community structure representation than methods without both of them. The reproducibility of all six methods was similar, and results from different experimenters and different times were in good agreement. Based on the evaluations done it appears that DNA extraction procedures for bacterial community analysis of human associated samples should include bead beating and/or mutanolysin to effectively lyse cells.     

IGF2BP2 Alternative Variants Associated with Glutamic Acid Decarboxylase Antibodies Negative Diabetes in Malaysian Subjects

Background

The association of Insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2) common variants (rs4402960 and rs1470579) with type 2 diabetes (T2D) has been performed in different populations. The aim of this study was to evaluate the association of alternative variants of IGF2BP2; rs6777038, rs16860234 and rs7651090 with glutamic acid decarboxylase antibodies (GADA) negative diabetes in Malaysian Subjects.

Methods/Principal Findings

IGF2BP2; rs6777038, rs16860234 and rs7651090 single nucleotide polymorphisms (SNPs) were genotyped in 1107 GADA negative diabetic patients and 620 control subjects of Asian from Malaysia. The additive genetic model adjusted for age, race, gender and BMI showed that alternative variants; rs6777038, rs16860234 and rs7651090 of IGF2BP2 associated with GADA negative diabetes (OR = 1.21; 1.36; 1.35, P = 0.03; 0.0004; 0.0002, respectively). In addition, the CCG haplotype and diplotype CCG-TCG increased the risk of diabetes (OR = 1.51, P = 0.01; OR = 2.36, P = 0.009, respectively).

Conclusions/Significance

IGF2BP2 alternative variants were associated with GADA negative diabetes. The IGF2BP2 haplotypes and diplotypes increased the risk of diabetes in Malaysian subject.     

Hexokinase-I protection against apoptotic cell death is mediated via interaction with the voltage-dependent anion channel-1: mapping the site of binding

In brain and tumor cells, the hexokinase isoforms HK-I and HK-II bind to the voltage-dependent anion channel (VDAC) in the outer mitochondrial membrane. We have previously shown that HK-I decreases murine VDAC1 (mVDAC1) channel conductance, inhibits cytochrome c release, and protects against apoptotic cell death. Now, we define mVDAC1 residues, found in two cytoplasmic domains, involved in the interaction with HK-I. Protection against cell death by HK-I, as induced by overexpression of native or mutated mVDAC1, served to identify the mVDAC1 amino acids required for interaction with HK-I. HK-I binding to mVDAC1 either in isolated mitochondria or reconstituted in a bilayer was inhibited upon mutation of specific VDAC1 residues. HK-I anti-apoptotic activity was also diminished upon mutation of these amino acids. HK-I-mediated inhibition of cytochrome c release induced by staurosporine was also diminished in cells expressing VDAC1 mutants. Our results thus offer new insights into the mechanism by which HK-I promotes tumor cell survival via inhibition of cytochrome c release through HK-I binding to VDAC1. These results, moreover, point to VDAC1 as a key player in mitochondrially mediated apoptosis and implicate an HK-I-VDAC1 interaction in the regulation of apoptosis. Finally, these findings suggest that interference with the binding of HK-I to mitochondria by VDAC1-derived peptides may offer a novel strategy by which to potentiate the efficacy of conventional chemotherapeutic agents.