MAPK inhibitors induce serine peptidase inhibitor Kazal type 1 (SPINK1) secretion in BRAF V600E-mutant colorectal adenocarcinoma

The mitogen-activated protein kinase (MAPK) pathway plays a central role in colorectal cancers (CRC). In particular, BRAF V600E-mutant tumors, which represent around 10% of CRCs, are refractory to current therapies. Overexpression and secretion of serine peptidase inhibitor Kazal type 1 (SPINK1) are observed in around 50% of CRCs, and its serum level can be used as a biomarker for poor prognosis. Utilizing a recently developed extendable blocking probe assay, we analyzed the BRAF mutation status in a CRC patient cohort (N=571) using tissue-derived RNA as the starting material. From the same RNA samples, we measured the relative SPINK1 expression levels using a quantitative real-time PCR method. Expression of mutant BRAF V600E correlated with poor prognosis, as did low expression of SPINK1 mRNA. Further, BRAF V600E correlated negatively with SPINK1 levels. In order to investigate the effect of MAPK pathway-targeted therapies on SPINK1 secretion, we conducted invitro studies using both wild-type and V600E CRC cell lines. BRAF inhibitor vemurafenib, and subsequent MAPK pathway inhibitors trametinib and SCH772984, significantly increased SPINK1 secretion in V600E CRC cell lines Colo205 and HT-29 with a concomitant decrease in trypsin-1 and -2 secretion. Notably, no SPINK1 increase or trypsin-1 decrease was observed in BRAF wild-type CRC cell line Caco-2 in response to MAPK pathway inhibitors. In further mechanistic studies, we observed that only trametinib was able to diminish completely both MEK and ERK phosphorylation in the V600E CRC cells. Furthermore, the key regulator of integrated stress response, activating transcription factor 4 (ATF-4), was downregulated both at mRNA and at protein level in response to trametinib treatment. In conclusion, these data suggest that sustained inhibition of not only MAPK pathway activation, but also ATF-4 and trypsin, might be beneficial in the therapy of BRAF V600E-mutant CRC and that SPINK1 levels may serve as an indicator of therapy response.Peer reviewe

Antigenic characterization of highly pathogenic avian influenza A(H5N1) viruses with chicken and ferret antisera reveals clade-dependent variation in hemagglutination inhibition profiles.

Highly pathogenic avian influenza (HPAI) A(H5N1) viruses pose a significant economic burden to the poultry industry worldwide and have pandemic potential. Poultry vaccination against HPAI A(H5N1) viruses has been an important component of HPAI control measures and has been performed in Vietnam since 2005. To systematically assess antigenic matching of current vaccines to circulating field variants, we produced a panel of chicken and ferret antisera raised against historical and contemporary Vietnamese reference viruses representing clade variants that were detected between 2001 and 2014. The antisera were used for hemagglutination inhibition (HI) assays to generate data sets for analysis by antigenic cartography, allowing for a direct comparison of results from chicken or ferret antisera. HI antigenic maps, developed with antisera from both hosts, revealed varying patterns of antigenic relationships and clustering of viruses that were dependent on the clade of viruses analyzed. Antigenic relationships between existing poultry vaccines and circulating field viruses were also aligned with in vivo protection profiles determined by previously reported vaccine challenge studies. Our results establish the feasibility and utility of HPAI A(H5N1) antigenic characterization using chicken antisera and support further experimental and modeling studies to investigate quantitative relationships between genetic variation, antigenic drift and correlates of poultry vaccine protection in vivo

Population density, water supply, and the risk of dengue fever in Vietnam: cohort study and spatial analysis.

BACKGROUND: Aedes aegypti, the major vector of dengue viruses, often breeds in water storage containers used by households without tap water supply, and occurs in high numbers even in dense urban areas. We analysed the interaction between human population density and lack of tap water as a cause of dengue fever outbreaks with the aim of identifying geographic areas at highest risk. METHODS AND FINDINGS: We conducted an individual-level cohort study in a population of 75,000 geo-referenced households in Vietnam over the course of two epidemics, on the basis of dengue hospital admissions (n = 3,013). We applied space-time scan statistics and mathematical models to confirm the findings. We identified a surprisingly narrow range of critical human population densities between around 3,000 to 7,000 people/km² prone to dengue outbreaks. In the study area, this population density was typical of villages and some peri-urban areas. Scan statistics showed that areas with a high population density or adequate water supply did not experience severe outbreaks. The risk of dengue was higher in rural than in urban areas, largely explained by lack of piped water supply, and in human population densities more often falling within the critical range. Mathematical modeling suggests that simple assumptions regarding area-level vector/host ratios may explain the occurrence of outbreaks. CONCLUSIONS: Rural areas may contribute at least as much to the dissemination of dengue fever as cities. Improving water supply and vector control in areas with a human population density critical for dengue transmission could increase the efficiency of control efforts. Please see later in the article for the Editors' Summary

Extendable blocking probe in reverse transcription for analysis of RNA variants with superior selectivity

Peer reviewe

Long-Term Weekly Iron-Folic Acid and De-Worming Is Associated with Stabilised Haemoglobin and Increasing Iron Stores in Non-Pregnant Women in Vietnam

BACKGROUND: The prevalence of anaemia and iron deficiency in women remains high worldwide. WHO recommends weekly iron-folic acid supplementation where anaemia rates in non-pregnant women of reproductive age are higher than 20%. In 2006, a demonstration project consisting of weekly iron-folic acid supplementation and regular de-worming was set up in two districts in a northern province in Vietnam where anaemia and hookworm rates were 38% and 76% respectively. In 2008 the project was expanded to all districts in the province, targeting some 250,000 women. The objectives of this study were to: 1) examine changes in haemoglobin, iron stores and soil transmitted helminth infection prevalence over three years and 2) assess women's access to and compliance with the intervention. METHODS AND FINDINGS: The study was a semi-cross-sectional, semi-longitudinal panel design with a baseline survey, three impact surveys at three-, twelve- and thirty months after commencement of the intervention, and three compliance surveys after ten weeks, eighteen and thirty six months. RESULTS: After thirty months, mean haemoglobin stabilised at 130.3 g/L, an increase of 8.2 g/L from baseline, and mean serum ferritin rose from 23.9 µg/L to 52 µg/L. Hookworm prevalence fell from 76% to 22% over the same period. After thirty six months, 81% of the target population were receiving supplements and 87% were taking 75% or more of the supplements they received. CONCLUSIONS: Weekly iron-folic acid supplementation and regular de-worming was effective in significantly and sustainably reducing the prevalence of anaemia and soil transmitted helminth infections and high compliance rates were maintained over three years

Viral pathogens associated with acute respiratory infections in central vietnamese children.

Hospitalized Vietnamese children with acute respiratory infection were investigated for 13 viral pathogens using multiplex-polymerase chain reaction. We enrolled 958 children of whom 659 (69%) had documented viral infection: rhinovirus (28%), respiratory syncytial virus (23%), influenza virus (15%), adenovirus (5%), human metapneumo virus (4.5%), parainfluenza virus (5%), and bocavirus (2%). These Vietnamese children had a range of respiratory viruses which underscores the need for enhanced acute respiratory infection surveillance in tropical developing countries

Evaluation of microscopic observation drug susceptibility assay for diagnosis of multidrug-resistant Tuberculosis in Viet Nam

<p>Abstract</p> <p>Background</p> <p>Early diagnosis of tuberculosis (TB) and multidrug resistant tuberculosis (MDR TB) is important for the elimination of TB. We evaluated the microscopic observation drug susceptibility (MODS) assay as a direct rapid drug susceptibility testing (DST) method for MDR-TB screening in sputum samples</p> <p>Methods</p> <p>All adult TB suspects, who were newly presenting to Pham Ngoc Thach Hospital from August to November 2008 were enrolled into the study. Processed sputum samples were used for DST by MODS (DST-MODS) (Rifampicin (RIF) 1 μg/ml and Isoniazid (INH) 0.4 μg/ml), MGIT culture (Mycobacterial Growth Indicator Tube) and Lowenstein Jensen (LJ) culture. Cultures positive by either MGIT or LJ were used for proportional DST (DST-LJ) (RIF 40 μg/ml and INH 0.2 μg/ml). DST profiles on MODS and LJ were compared. Discrepant results were resolved by multiplex allele specific PCR (MAS-PCR).</p> <p>Results</p> <p>Seven hundred and nine TB suspects/samples were enrolled into the study, of which 300 samples with DST profiles available from both MODS and DST-LJ were analyzed. Cording in MODS was unable to correctly identify 3 Mycobacteria Other Than Tuberculosis (MOTT) isolates, resulting in 3 false positive TB diagnoses. None of these isolates were identified as MDR-TB by MODS. The sensitivity and specificity of MODS were 72.6% (95%CI: 59.8, 83.1) and 97.9% (95%CI: 95.2, 99.3), respectively for detection of INH resistant isolates, 72.7% (95%CI: 30.9, 93.7) and 99.7% (95%CI: 98.1, 99.9), respectively for detecting RIF resistant isolates and 77.8% (95%CI: 39.9, 97.1) and 99.7% (95%CI: 98.1, 99.9), respectively for detecting MDR isolates. The positive and negative predictive values (PPV and NPV) of DST-MODS were 87.5% (95%CI: 47.3, 99.6) and 99.3% (95%CI: 97.5, 99.9) for detection of MDR isolates; and the agreement between MODS and DST-LJ was 99.0% (kappa: 0.8, <it>P </it>< 0.001) for MDR diagnosis. The low sensitivity of MODS for drug resistance detection was probably due to low bacterial load samples and the high INH concentration (0.4 μg/ml). The low PPV of DST-MODS may be due to the low MDR-TB rate in the study population (3.8%). The turnaround time of DST-MODS was 9 days and 53 days for DST-LJ.</p> <p>Conclusion</p> <p>The DST-MODS technique is rapid with low contamination rates. However, the sensitivity of DST-MODS for detection of INH and RIF resistance in this study was lower than reported from other settings.</p