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Rationale: The in‐sample calibration curve (ISCC) approach of quantification utilizes the response of isotopologue ions from spiked‐in stable isotope labeled internal standard (SIL‐IS) to build a standard curve. The quantitative analysis of the study sample is achieved based on the response of selected monoisotopic analyte ion against the calibration curve. Although this methodology has been demonstrated to be feasible by unit and high‐resolution mass spectrometers, quantitation on high‐resolution mass spectrometer with product ions has not been tested. We tested the feasibility of this approach using product ions on an high‐resolution mass spectrometer equipped with an Orbitrap detector. Methods: Using a proteomics workflow for sample preparation, two surrogate peptides were quantified from a complex matrix of protein digest from human peripheral blood mononuclear cells (hPBMCs). SIL‐IS was spiked in at different levels to construct calibration curves in a traditional manner. ISCCs were prepared using extracted ion chromatograms from isotopically resolved mass spectra and compared with traditional calibration curves. Results: A linear response was observed with ISCC approach for at least two to three orders of magnitude in MS1 as well as targeted MS2 (tMS2). From protein digests, isobaric interferences were observed for endogenous peptides on the MS1 level; this was circumvented with product‐ion‐based quantitation where for one peptide, %CV for endogenous levels was more than 20% with ISCC but higher with the traditional calibration curve approach. For the second peptide, endogenous levels could not be determined in the traditional approach as calibrant levels did not bracket the lower end, and with the ISCC approach, isotopologues at abundances lower than the endogenous level allowed for quantitative assessments. Conclusions: ISCC demonstrated improved precision across surrogate peptides from endogenous protein digests. In samples where endogenous analyte concentrations were low in abundance, ISCC rescued what would have been a non‐reportable result in a traditional bioanalytical assay as calibrant levels were not prepared at adequately low levels to bracket unknowns. ISCC using high‐resolution mass spectrometer is feasible and ideal compared to unit resolution mass spectrometers. High‐resolution mass spectrometer allows for isotopic resolution for analytes with > + 2 charge state and provides flexibility in quantification using multiple product ions. ISCC using high‐resolution mass spectrometer allows for simultaneous assaying of low abundance isotopologues, the signal acquisition of which is not constrained by limits in data acquisition or calibrant preparation as with other approaches but rather limited by platform sensitivity. In contrast to unit resolution mass spectrometers, these features offered by high‐resolution mass spectrometer could be especially useful for the drug discovery assay support where there is less lead time for assay development than for the assays to support the drug development studies. [ABSTRACT FROM AUTHOR]

Summary: Background: A novel potassium‐competitive acid blocker, DWP14012, is in clinical development as a potential alternative to proton pump inhibitors for the treatment of acid‐related diseases. Aims: To evaluate the safety, tolerability, pharmacodynamics and pharmacokinetics of DWP14012 in humans. Methods: A randomised, double‐blind, double‐dummy, placebo‐ and active‐controlled, single‐ and multiple‐ascending dose (SAD and MAD, respectively) study was conducted in healthy male subjects without Helicobacter pylori infection. Subjects randomly received a single oral dose of 10‐320 mg DWP14012, esomeprazole (active comparator) or placebo in the SAD study (n = 72) and once daily doses of 20‐160 mg DWP14012, esomeprazole or placebo for 7 days in the MAD study (n = 48; 8:2:2). Tolerability was evaluated using a microRNA‐122 assay. Pharmacodynamics were evaluated through 24‐hour gastric pH monitoring, and pharmacokinetics were evaluated plasma and urine DWP14012 concentrations. Results: DWP14012 was generally well tolerated. The liver toxicity of DWP14012 was not higher than that of placebo after multiple oral administrations. DWP14012 showed rapid and sustained suppression of gastric acid secretion for 24 hours after dosing. Clear dose‐response and exposure‐response relationships were observed. Plasma concentrations of DWP14012 increased in a dose‐proportional manner in the MAD study, whereas in the SAD study, DWP14012 did not significantly accumulate in the plasma. Conclusions: DWP14012 was well tolerated, and showed a rapid and long‐lasting gastric acid suppression effect in healthy subjects. These results justify further investigation of DWP14012 in patients with acid‐related disorders. [ABSTRACT FROM AUTHOR]

Abstract: The discovery of biomarkers from high‐dimensional data is a very challenging task in cancer diagnoses. On the one hand, biomarker discovery is the so‐called high‐dimensional small‐sample problem. On the other hand, these data are redundant and noisy. In recent years, biomarker discovery from high‐throughput biological data has become an increasingly important emerging topic in the field of bioinformatics. In this study, we propose a binary differential evolution algorithm for feature selection. Firstly, we suggest using a two‐stage approach, where three filter methods including the Fisher score, T‐statistics, and Information gain are used to generate the feature pool for input to differential evolution (DE). Secondly, in order to improve the performance of differential evolution algorithm for feature selection, a new variant of binary DE called BDE is proposed. Three optimization strategies are incorporated into the BDE. The first strategy is the heuristic method in initial stage, the second one is the self‐adaptive parameter control, and the third one is the minimum change value to improve the exploration behaviour thus enhance the diversity. Finally, Support vector machine (SVM) is used as the classifier in 10 fold cross‐validation method. The experimental results of our proposed algorithm on some benchmark datasets demonstrate the effectiveness of our algorithm. In addition, the BDE forged in this study will be of great potential in feature selection problems. [ABSTRACT FROM AUTHOR]

Abstract: In this study, a new formulation for the thermo‐osmotic permeability of natural pore solutions containing monovalent and divalent cations is proposed. The mathematical formulation proposed here is based on the theoretical framework supporting thermo‐osmosis which relies on water structure alteration in the pore space of surface‐charged materials caused by solid‐fluid electrochemical interactions. The ionic content balancing the surface charge of clay minerals causes a disruption in the hydrogen bond network when more structured water is present at the clay surface. Analytical expressions based on our heuristic model are proposed and compared to the available data for NaCl solutions. It is shown that the introduction of divalent cations reduces the thermo‐osmotic permeability by one third compared to the monovalent case. The analytical expressions provided here can be used to advantage for safety calculations in deep underground nuclear waste repositories. [ABSTRACT FROM AUTHOR]

Decreased oral clopidogrel absorption caused by induction of intestinal permeability glycoprotein (P-gp) expression after aspirin administration was observed in rats. This study evaluated the effect of aspirin coadministration on the pharmacokinetics/pharmacodynamics of clopidogrel in humans. A single 75-mg dose of clopidogrel was orally administered before and after 2 and 4 weeks of once-daily 100-mg aspirin administration in 18 healthy volunteers who were recruited based on CYP2C19 and PON1 genotypes. Plasma concentrations of clopidogrel and its active metabolite, H4, and relative platelet inhibition (RPI) were determined. The P-gp microRNA miR-27a increased by up to 7.67-fold (P = 0.004) and the clopidogrel area under the concentration-time curve (AUC) decreased by 14% (P > 0.05), but the AUC of H4 remained unchanged and RPI increased by up to 15% (P = 0.002) after aspirin administration. These findings indicate low-dose aspirin coadministration may decrease clopidogrel bioavailability but does not decrease its efficacy. [ABSTRACT FROM AUTHOR]

RATIONALE Sensitivity is one major challenge limiting the application of liquid chromatography/tandem mass spectrometry (LC/MS/MS) methods for bioanalysis of proteins. A novel selective peptide derivatization (SPD) strategy was proposed to improve assay sensitivity. The main concept of the SPD strategy is to selectively derivatize surrogate peptides of the target protein in the digests, while not derivatizing the abundant background peptides, thereby improving the separation of target peptides during sample extraction and chromatography, and increasing the sensitivity. Additional benefits may help improve sensitivity include (1) increased ionization efficiency; (2) improved fragmentation pattern; and (3) increased sample extraction recovery of target peptides. METHODS Feasibility assessment of the SPD strategy was conducted using BMS-986012, a monoclonal antibody, as the model protein, and with malondialdehyde (MDA) to selectively derivatize the arginine-containing surrogate peptide SLIY in tryptic-digested monkey serum samples. RESULTS The decreased polarity and basicity of the MDA-derivatized peptide SLIY (MDA-SLIY) helped improve its separation from the background peptides during solid-phase extraction (SPE) and chromatography. The recovery of MDA-SLIY was 36.1-44.2%, which was ~3-fold higher than the recovery of peptide SLIY (11.9-16.1%). There was no significant ion suppression for MDA-SLIY. Overall, SPD improved the sensitivity ~5-fold. SPD methodology was then successfully applied to the development of a sensitive LC/MS/MS assay for BMS-986012 in monkey serum. CONCLUSIONS This work demonstrates the feasibility of the SPD strategy for sensitivity enhancement. SPD can provide a simple, cost-efficient, and antibody-free sample preparation approach to improve sensitivity. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

As an effective DPP-IV inhibitor, 2-(4-((2-(2 S,5 R)-2-Cyano-5-ethynyl-1-pyrrolidinyl)-2-oxoethylamino)-4-methyl-1-piperidinyl)-4-pyridinecarboxylic acid (ABT-279), is an investigational drug candidate under development at Abbott Laboratories for potential treatment of type 2 diabetes. In order to support the development of ABT-279, multiple analytical methods for an accurate, precise and selective concentration determination of ABT-279 in different matrices were developed and validated in accordance with the US Food and Drug Administration Guidance on Bioanalytical Method Validation. The analytical method for ABT-279 in dog plasma was validated in parallel to other validations for ABT-279 determination in different matrices. In order to shorten the sample preparation time and increase method precision, an automated multi-channel liquid handler was used to perform high-throughput protein precipitation and all other liquid transfers. The separation was performed through a Waters YMC ODS-AQ column (2.0 × 150 mm, 5 µm, 120 Å) with a mobile phase of 20 m m ammonium acetate in 20% acetonitrile at a flow rate of 0.3 mL/min. Data collection started at 2.2 min and continued for 2.0 min. The validated linear dynamic range in dog plasma was between 3.05 and 2033.64 ng/mL using a 50 µL sample volume. The achieved r2 coefficient of determination from three consecutive runs was between 0.998625 and 0.999085. The mean bias was between −4.1 and 4.3% for all calibration standards including lower limit of quantitation. The mean bias was between −8.0 and 0.4% for the quality control samples. The precision, expressed as a coefficient of variation (CV), was ≤4.1% for all levels of quality control samples. The validation results demonstrated that the high-throughput method was accurate, precise and selective for the determination of ABT-279 in dog plasma. The validated method was also employed to support two toxicology studies. The passing rate was 100% for all 49 runs from one validation study and two toxicology studies. Copyright © 2007 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Tumour-associated lymphatics contribute to a key component of metastatic spread, however, the biological interaction of tumour cells with intratumoural and peritumoural lymphatics (ITLs and PTLs) has remained unclear. To address this important issue, we have focused on the morphological and molecular aspects of newly formed lymphatics (lymphangiogenesis) and pre-existing lymphatics in the intratumoural and peritumoural tissues by using a hybridoma-induced tumour model. In the present study, ITLs with very high vessel density within the tumour mass showed small and flattened contours that varied from non-solid-to-solid tumours, whereas PTLs were relatively disorganized and tortuous, and packed with a cluster of tumour cells at the tumour periphery. Lymphatic endothelial cells (LECs) both in ITLs and PTLs were expressed with LYVE-1 and podoplanin in various tumour tissues, in which initial lymphatics were extremely extended and dilated. The tumour cells were frequently detected adhering to or penetrating lymphatic walls, especially near the open junctions. In the metastatic tissues, lymphangiogenic vasculatures occurred within the tumour matrix, and collecting PTLs represented abnormal twisty valve leaflets. The Western blot and RT-PCR analysis showed local variations of LEC proliferating potentials and lymphatic involvement in metastasis by a distinct profile of the protein and mRNA expression by LYVE-1, podoplanin, Prox-1 and vascular endothelial growth factor-3 (VEGFR-3). These findings indicated that both ITLs and PTLs, including enlarged pre-existing and newly formed lymphatics, may play a crucial role in metastasis with an active tumour cell adhesion, invasion, migration and implantation. [ABSTRACT FROM AUTHOR]

Atrophic body gastritis (ABG) is common in China. Although histology via endoscopy is an efficient and reliable means of diagnosing ABG, it is an invasive procedure. Therefore, in the present study serum pepsinogen (PG) was used as a biomarker to develop a novel noninvasive test as the first option for screening of ABG in certain groups of Chinese. The study population consisted of 81 selected dyspeptic patients (mean age, 64.8 ± 0.7 years; M:F, 43:38) who underwent diagnostic gastroscopy. At least four biopsy specimens were taken from the antrum and corpus of the stomach (two specimens from each site) for histological diagnosis. Blood samples for ELISA assays of serum pepsinogen I (PGI), pepsinogen II (PGII) and IgG antibodies against Helicobacter pylori (Hp IgG) were drawn after endoscopy. Cut-off points were calculated using receiver operating curves (ROC). There was no correlation between serum PG and atrophy in the antral mucosa. The mean serum concentration of PGI was lower ( P < 0.05) in patients with ABG (89.9 µg/L) than in those with normal mucosa (NM) and non-ABG (123.7 µg/L and 139.1 µg/L). The mean ratio of PGI:PGII was also lower ( P < 0.01) in patients with ABG (6.2) than in those with NM and non-ABG (11.6 and 11.7). There was no difference in serum PGI or the PGI:PGII ratio between patients with and without H. pylori infection. For diagnosing ABG, the area under the ROC of PGI and the PGI:PGII ratio was 0.741(95% CI: 0.627–0.856) and 0.874(95% CI: 0.788–0.961), respectively. The maximum of the Youden's index (YI) of PGI and the PGI:PGII ratio was 0.426 and 0.722, respectively. The best cut-off point for PGI was 97.1 µg/L with sensitivity of 67% and specificity of 76%, and for PGI:PGII ratio was 8.1 µg/L, with sensitivity of 89% and specificity of 83%. The serum PGI:PGII ratio appears to be a sensitive and specific assay for corpus atrophy, thus providing a noninvasive and indicative test for diagnosis of atrophic gastritis. [ABSTRACT FROM AUTHOR]