Pan-cancer proteomic map of 949 human cell lines

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The usefulness of measuring n-butyric acid concentration as a new indicator of blood decomposition in forensic autopsy

In forensic medicine, although various alcohols have been reported as indicators of decomposition in collected blood, no studies have examined short-chain fatty acids as indicators. In this study, the blood n-butyric acid concentration was quantified, and the association between n-butyric acid and decomposition was investigated to determine whether the detection of n-butyric acid could be a new indicator of decomposition. Among the forensic autopsies performed from 2016 to 2018 in our laboratory, the cases were divided into decomposed (n = 20) and non-decomposed (n = 20) groups based on macroscopic findings. Blood samples collected at the time of autopsy were derivatized with 3-nitrophenylhydrazine hydrochloride after solid-phase extraction. The n-butyric acid concentration was measured using liquid chromatography–tandem mass spectrometry. In addition, ethanol and n-propanol were measured using a gas chromatography-flame ionization detector. There was a significant difference (p < 0.01) in the concentrations of n-butyric acid between the decomposed and non-decomposed groups (0.343 ± 0.259 [0.030–0.973] and 0.003 ± 0.002 [0.001–0.007] mg/mL, respectively). In the decomposed group, n-butyric acid was detected at high concentrations, even in cases where n-propanol was low. These results suggest that n-butyric acid is more likely to be an indicator of blood decomposition than n-propanol.     

The bioavailability of ingested 26Al-labelled aluminium and aluminium compounds in the rat

The fractional uptake of ingested aluminium and aluminium compounds (aluminium citrate, aluminium nitrate, aluminium chloride, aluminium sulphate, aluminium hydroxide, aluminium oxide, aluminium metal, powdered aluminium pot electrolyte, acidic sodium aluminium phosphate (SALP), basic sodium aluminium phosphate (Kasal), sodium aluminium silicate and FD&C red 40 aluminium lake) from the gastro-intestinal tract of adult female rats was measured. This was determined by comparing retained body burden of ²⁶Al at seven days post-admistration of an i.v. injection of ²⁶Al-labelled aluminium citrate with that retained following the gastric admistration of ²⁶Al-labelled test compounds as either solutions or suspended solid. The calculated percentage uptake of ²⁶Al for all the aluminium solutions was similar: aluminium citrate 0.08%, aluminium chloride 0.05%, aluminium nitrate 0.05% and aluminium sulphate 0.21%. The uptake of ²⁶Al administered as insoluble particulates was lower: 0.03% for aluminium hydroxide; 0.02% for aluminium oxide; 0.04% for powdered pot electrolyte; 0.12% for sodium aluminium silicate; and 0.09% for FD&C red 40 aluminium lake. For aluminium metal, SALP and Kasal the amount of ²⁶Al present in the rats was insufficient to determine uptake and was less than 0.03%. The results produced for aluminium citrate, aluminium hydroxide and aluminium sulphate are close to those published for man.     

Redox active or thiol reactive? Optimization of rapid screens to identify less evident nuisance compounds

Compounds that exhibit assay interference or undesirable mechanisms of bioactivity are routinely encountered in assays at various stages of drug discovery. We observed that assays for the investigation of thiol-reactive and redox-active compounds have not been collected in a comprehensive review. Here, we review these assays and subject them to experimental optimization to improve their reliability. We demonstrate the usefulness of our assay cascade by assaying a library of bioactive compounds, chemical probes, and a set of approved drugs. These high-throughput assays should complement the array of wet-lab and in silico assays during the initial stages of hit discovery campaigns to pursue only hit compounds with tractable mechanisms of action.     

Proteomic genotyping of fingermark donors with genetically variant peptides

Proteomic genotyping detects single amino acid polymorphisms to infer the genotype of corresponding non-synonymous SNPs. Like any DNA genotype, these inferences can be used to estimate random match probability. Fingermarks are a common source of biological evidence that is sample limited and a highly variable source of identifying DNA. Genetically variant peptides from fingermarks, that contain single amino acid polymorphisms, are an additional source of identifying genetic information. To discover these peptide biomarkers epidermal corneocytes from 9 subjects were isolated, processed, digested with trypsin and applied to mass spectrometry. The resulting proteomic and matching exome datasets were used to discover, characterize and validate 60 genetically variant peptides. An average of 28.8 ± 4.4 genetically variant peptides were detected from each subject resulting in a total of 264 SNP allele inferences with 260 true and 4 false positives, a false discovery rate of 1.5%. Random match probabilities were estimated using the genotype frequencies from the matching major populations in the 1000 Genomes Project. Estimates ranged up to a value of 1 in 1.7 × 10⁸, with a median probability of 1 in 2.4 × 10⁶. Furthermore, the proteomically-inferred genotypes are likely to be compatible with the STR-based random match probability estimates since the closest STR locus was 2.2 Mb from the nearest GVP-inferred SNP. This project represents a novel mode of genetic information that can be obtained from fingermarks and has the potential to complement other methods of human identification including analysis of ridge patterns or touch DNA.     

Chemical isotope labeling liquid chromatography mass spectrometry for investigating acute dietary effects of cow milk consumption on human urine metabolome

Biomarker discovery has been increasingly important in the field of metabolomics for the detection and understanding of diseases. Of the many biofluids available for metabolomics, urine is a preferred option as it is non-invasive to collect and contains a wide range of metabolites reflective of the health status of the testing individual. However, urine also contains many exogenous metabolites which are introduced through various sources such as diet. This complicates the data interpretation when searching the metabolome for disease-related endogenous metabolites. Since diet is difficult to control, this work aims to study the acute effects of diet (particularly cow milk) consumption on the human urine amine/phenol submetabolome by utilizing differential chemical isotope labeling (CIL) liquid chromatography mass spectrometry (LC-MS). LC-MS analysis of 62 urine samples collected before and after (1 hour and 2 hours) milk intake resulted in the detection of 4985 metabolites with an average of 3815 ± 206 (n = 62) detected per sample. The work aims to differentiate the exogenous “food” metabolites from the endogenous metabolite pool and to determine any dietary effects from milk intake on the human urine metabolome.     

Current status of MALDI-TOF mass spectrometry in clinical microbiology

Mass spectrometry (MS) is a type of analysis used to determine what molecules make up a sample, based on the mass spectrum that are created by the ions. Mass spectrometers are able to perform traditional target analyte identification and quantitation; however, they may also be used within a clinical setting for the rapid identification of bacteria. The causative agent in sepsis is changed over time, and clinical decisions affecting the management of infections are often based on the outcomes of bacterial identification. Therefore, it is essential that such identifications are performed quickly and interpreted correctly. Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometer is one of the most popular MS instruments used in biology, due to its rapid and precise identification of genus and species of an extensive range of Gram-negative and -positive bacteria. Microorganism identification by Mass spectrometry is based on identifying a characteristic spectrum of each species and then matched with a large database within the instrument. The present review gives a contemporary perspective on the challenges and opportunities for bacterial identification as well as a written report of how technological innovation has advanced MS. Future clinical applications will also be addressed, particularly the use of MALDI-TOF MS in the field of microbiology for the identification and the analysis of antibiotic resistance.     

Identification of IgG1 Aggregation Initiation Region by Hydrogen Deuterium Mass Spectrometry

Antibody aggregates are a potential risk for immunogenicity; therefore, rational approaches to improve associated aggregation properties need to be developed. Here, we report the amino acid region responsible for aggregation initiation. Two types of therapeutic IgG1 antibody monomer samples were prepared: IgG1 mAb40-3M stored at 40°C for 3 months, which existed in monodisperse state, and the monomer mAb65-5m, which was dissociated from small soluble aggregates by heating at 65°C for 5 min. Hydrogen deuterium exchange mass spectrometry of mAb40-3M identified 2 sites in the Fc region (site 1, F239-M256; site 2, S428-G450) with increased exchange rates. Site 1 includes a region reported as being susceptible to structural change induced by stress. Exposure of site 1 was undetected after 2 months of storage at 40°C but was subsequently detectable after 3 months. As site 2 is spatially close to site 1, the structural change of site 1 could propagate site 2. Besides these 2 regions, hydrogen deuterium exchange mass spectrometry of mAb65-5m identified an exposure of I257-W281 in Fc (site 3), within which a peptide sequence with high aggregation tendency was discovered. We thus concluded that exposure of site 3 is a trigger for the association of a partially denatured antibody.     

Forced Oxidative Degradation Pathways of the Imidazole Moiety of Daclatasvir

Daclatasvir hydrochloride (DCV) is the active pharmaceutical ingredient of Daklinza, a marketed product for the treatment of hepatitis C viral infection. The intrinsic stability of daclatasvir was evaluated via a forced degradation study. DCV was found to be stable in the solid state. In solution, its carbamate moiety is susceptible to basic hydrolysis, whereas its imidazole is liable to base-mediated autoxidation to form degradants 1 and 3, 7-8, respectively. The imidazole moiety can also be oxidized to form degradants 6-7 in the presence of hydrogen peroxide or azobisisobutyronitrile. The chloro-adduct degradant 9 was also observed in hydrogen peroxide solution. Furthermore, the imidazole moiety is sensitive to photodegradation in solution. Degradants 2-8 were observed in a solution of DCV exposed to high intensity light/UV light; the formation of degradants 2 and 5-8 was postulated through 4 degradation pathways. The degradants 3 and 4 were deemed to be secondary degradants of 7 and 5, respectively.