The Clinical Proteomic Tumor Analysis Consortium (CPTAC), under the auspices of the National Cancer Institute’s Office of Cancer Clinical Proteomics Research, is a comprehensive and coordinated ...effort to accelerate the understanding of the molecular basis of cancer through the application of proteomic technologies and workflows to clinical tumor samples with characterized genomic and transcript profiles. The consortium analyzes cancer biospecimens using mass spectrometry, identifying and quantifying the constituent proteins and characterizing each tumor sample’s proteome. Mass spectrometry enables highly specific identification of proteins and their isoforms, accurate relative quantitation of protein abundance in contrasting biospecimens, and localization of post-translational protein modifications, such as phosphorylation, on a protein’s sequence. The combination of proteomics, transcriptomics, and genomics data from the same clinical tumor samples provides an unprecedented opportunity for tumor proteogenomics. The CPTAC Data Portal is the centralized data repository for the dissemination of proteomic data collected by Proteome Characterization Centers (PCCs) in the consortium. The portal currently hosts 6.3 TB of data and includes proteomic investigations of breast, colorectal, and ovarian tumor tissues from The Cancer Genome Atlas (TCGA). The data collected by the consortium is made freely available to the public through the data portal.
Genetic engineering of crop plants has been successful in transferring traits into elite lines beyond what can be achieved with breeding techniques. Introduction of transgenes originating from other ...species has conferred resistance to biotic and abiotic stresses, increased efficiency, and modified developmental programs. The next challenge is now to combine multiple transgenes into elite varieties via gene stacking to combine traits. Generating stable homozygous lines with multiple transgenes requires selection of segregating generations which is time consuming and labor intensive, especially if the crop is polyploid. Insertion site effects and transgene copy number are important metrics for commercialization and trait efficiency. We have developed a simple method to identify the sites of transgene insertions using T-DNA-specific primers and high-throughput sequencing that enables identification of multiple insertion sites in the T.sub.1 generation of any crop transformed via Agrobacterium. We present an example using the allohexaploid oil-seed plant Camelina sativa to determine insertion site location of two transgenes. This new methodology enables the early selection of desirable transgene location and copy number to generate homozygous lines within two generations.
This study is one of the first COVID-19 related bus studies to fully characterize cough aerosol dispersion and control in the highly turbulent real-world environment of driving regular bus routes on ...both a school bus and a transit bus. While several other bus studies have been conducted, they were limited to clinical contact tracing, simulation, or partial characterization of aerosol transmission in the passenger areas with constraint conditions. When considering the risk of transmission of SARS-CoV-2 (COVID-19) and other highly infectious airborne diseases, ground based public transportation systems are high-risk environments for airborne transmission particularly since social distancing of six feet is not practical on most buses. This study demonstrates that wearing of masks reduced the overall particle count released into the bus by an average of 50% or more depending on mask quality and reduced the dispersion distance by several feet. The study also demonstrates an 84.36% reduction in aerosol particles and an 80.28% reduction in the mean aerosol residence time for some test cases. We conducted 84 experimental runs using nebulized 10% sodium chloride and a mechanical exhalation simulator that resulted in 78.3 million data points and 124 miles of on-the-road testing. Our study not only captures the dispersion patterns using 28 networked particle counters, it also quantifies the effectiveness of using on-board fans, opening of various windows, use of face coverings or masks, and the use of the transit bus HVAC system. This work additionally provides empirical observations of aerosol dispersion in a real-world turbulent air environment, which are remarkably different than many existing fluid dynamics simulations, and also offers substantial discussion on the implications for inclement weather conditions, driver safety, retrofit applications to improve bus air quality, and operational considerations for public transportation organizations.
This paper investigates adiabatic shear bands (ASBs) in 2024-T351 aluminium and their effect on the mechanical properties and microstructure of this aluminium alloy. Using hat-shaped specimens in a ...split Hopkinson pressure bar (SHPB) and universal testing machine, resolved shear stress-nominal plastic shear strain curves were presented at shear strain rates between 1 × 10−4 and 15.4 × 103 s−1. A revised formula for shear stress in hat specimens is proposed. Microstructural observations were completed on deformed specimens, with hardness results, scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDS) presented. A critical shear strain rate range for ASB initiation in these specimens of approximately 8500–10500 s−1 was determined within impacted specimens.
Myeloid-derived suppressor cells (MDSC) are immature myeloid cells that accumulate in the circulation and the tumor microenvironment of most cancer patients. There, MDSC suppress both adaptive and ...innate immunity, hindering immunotherapies. The inflammatory milieu often present in cancers facilitates MDSC suppressive activity, causing aggressive tumor progression and metastasis. MDSC from tumor-bearing mice release exosomes, which carry biologically active proteins and mediate some of the immunosuppressive functions characteristic of MDSC. Studies on other cell types have shown that exosomes may also carry RNAs which can be transferred to local and distant cells, yet the mRNA and microRNA cargo of MDSC-derived exosomes has not been studied to date. Here, the cargo of MDSC and their exosomes was interrogated with the goal of identifying and characterizing molecules that may facilitate MDSC suppressive potency. Because inflammation is an established driving force for MDSC suppressive activity, we used the well-established 4T1 mouse mammary carcinoma system, which includes “conventional” as well as “inflammatory” MDSC. We provide evidence that MDSC-derived exosomes carry proteins, mRNAs, and microRNAs with different quantitative profiles than those of their parental cells. Several of these molecules have known or predicted functions consistent with MDSC suppressive activity, suggesting a potential mechanistic redundancy.
Protein identification from tandem mass spectra is one of the most versatile and widely used proteomics workflows, able to identify proteins, characterize post-translational modifications, and ...provide semiquantitative measurements of relative protein abundance. This manuscript describes the concepts, prerequisites, and methods required to analyze a tandem mass spectrometry dataset in order to identify its proteins, by using a tandem mass spectrometry search engine to search protein sequence databases. The discussion includes instructions for extraction, preparation, and formatting of spectral datafiles, selection of appropriate search parameter settings, and basic interpretation of the results.
Cirrhosis of the liver is associated with increased fucosylation of proteins in the plasma. We describe a data-independent (DIA) strategy for comparative analysis of the site-specific glycoforms of ...plasma glycoproteins. A library of 161 glycoforms of 25 N-glycopeptides was established by data-dependent LC-MS/MS analysis of a tryptic digest of 14 human protein groups retained on a multiple affinity removal column. The collision-induced dissociation conditions were adjusted to maximize the yield of selective Y-ions which were quantified by a data-independent mass spectrometry workflow using a 10-Da acquisition window. Using this workflow, we quantified 125 glycoforms of 25 glycopeptides, covering 10 of the 14 proteins, without any further glycopeptide enrichment. Comparison of the proteins in the plasma of healthy controls and cirrhotic patients shows an average 1.5-fold increase in the fucosylation of bi-antennary glycoforms and 3-fold increase in the fucosylation of tri- and tetra- antennary glycoforms. These results show that the adjusted glycopeptide DIA workflow using soft collision-induced fragmentation of glycopeptides is suitable for site-specific analysis of protein glycosylation in complex mixtures of analytes without glycopeptide enrichment.
We provide evidence at the molecular level that ubiquitinated proteins are present in exosomes shed by myeloid-derived suppressor cells (MDSC). Ubiquitin was selected as a post-translational ...modification of interest because it is known to play a determinant role in the endosomal trafficking that culminates in exosome release. Enrichment was achieved by two immunoprecipitations, first at the protein level and subsequently at the peptide level. Fifty ubiquitinated proteins were identified by tandem mass spectrometry filtering at a 5% spectral false discovery rate and using the conservative requirement that glycinylglycine-modified lysine residues were observed in tryptic peptides. Thirty five of these proteins have not previously been reported to be ubiquitinated. The ubiquitinated cohort spans a range of protein sizes and favors basic pI values and hydrophobicity. Five proteins associated with endosomal trafficking were identified as ubiquitinated, along with pro-inflammatory high mobility group protein B1 and proinflammatory histones.
Patients with neurofibromatosis type 1 (NF1) frequently develop plexiform neurofibromas (PNs), which can cause significant morbidity. We performed a phase II trial of the MAPK/ERK kinase inhibitor, ...mirdametinib (PD-0325901), in patients with NF1 and inoperable PNs. The primary objective was response rate based on volumetric magnetic resonance imaging analysis.
Inclusion criteria included age ≥ 16 years and a PN that was either progressive or causing significant morbidity. First-dose pharmacokinetics were performed. Patients completed patient-reported outcome measures. Patients received mirdametinib by mouth twice a day at 2 mg/m
/dose (maximum dose = 4 mg twice a day) in a 3-week on/1-week off sequence. Each course was 4 weeks in duration. Evaluations were performed after four courses for the first year and then after every six courses. Patients could receive a maximum of 24 total courses.
Nineteen patients were enrolled, and all 19 received mirdametinib. The median age was 24 years (range, 16-39 years); the median baseline tumor volume was 363.8 mL (range, 3.9-5,161 mL). Eight of the 19 patients (42%) achieved a partial response of the target PN by course 12, and 10 (53%) had stable disease. One patient (5%) developed progressive disease at course 8. Significant and durable decreases were observed in pain ratings.
To our knowledge, this analysis represents the first characterization of the activity and pharmacokinetics of mirdametinib in patients with NF1 and PNs and is the first published response study for MAPK/ERK kinase inhibitors in adults with NF1 and PNs. Mirdametinib given at 2 mg/m
/dose (maximum dose, 4 mg) twice daily in a 3-week on/1-week off sequence resulted in a 42% partial response rate with preliminary evidence of reduction in pain.
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