Acrylamide-Based Separation Matrices

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Moving towards high density clinical signature studies with a human proteome catalogue developing multiplexing mass spectrometry assay panels

A perspective overview is given describing the current development of multiplex mass spectrometry assay technology platforms utilized for high throughput clinical sample analysis. The development of targeted therapies with novel personalized medicine drugs will require new tools for monitoring efficacy and outcome that will rely on both the quantification of disease progression related biomarkers as well as the measurement of disease specific pathway/signaling proteins

Inflammatory markers in Huntington's disease plasma—A robust nanoLC–MRM-MS assay development

AbstractThe development of an MRM assay for the measurements of six inflammatory markers is presented. We report a robust and sensitive quantitative assay with a relative standard deviation of <15% that accounts for the entire sample processing. The assay has a dynamic range with 4 orders of magnitude and the LOQs are in the attomolar range. We used plasma from Huntington's disease gene carriers and healthy controls to compare our MRM method with antibody based methods. Importantly, we found a good agreement between assays for the measurement of C-reactive protein, in contrast to complement component 3 and complement factor H

Limited Tumor Tissue Drug Penetration Contributes to Primary Resistance against Angiogenesis Inhibitors

Resistance mechanisms against antiangiogenic drugs are unclear. Here, we correlated the antitumor and antivascular properties of five different antiangiogenic receptor tyrosine kinase inhibitors (RTKIs) (motesanib, pazopanib, sorafenib, sunitinib, vatalanib) with their intratumoral distribution data obtained by matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI). In the first mouse model, only sunitinib exhibited broad-spectrum antivascular and antitumor activities by simultaneously suppressing vascular endothelial growth factor receptor-2 (VEGFR2) and desmin expression, and by increasing intratumoral hypoxia and inhibiting both tumor growth and vascularisation significantly. Importantly, the highest and most homogeneous intratumoral drug concentrations have been found in sunitinib-treated animals. In another animal model, where - in contrast to the first model - vatalanib was detectable at homogeneously high intratumoral concentrations, the drug significantly reduced tumor growth and angiogenesis. In conclusion, the tumor tissue penetration and thus the antiangiogenic and antitumor potential of antiangiogenic RTKIs vary among the tumor models and our study demonstrates the potential of MALDI-MSI to predict the efficacy of unlabelled small molecule antiangiogenic drugs in malignant tissue. Our approach is thus a major technical and preclinical advance demonstrating that primary resistance to angiogenesis inhibitors involves limited tumor tissue drug penetration. We also conclude that MALDI-MSI may significantly contribute to the improvement of antivascular cancer therapies

Separate and combined effects of advanced age and obesity on mammary adipose inflammation, immunosuppression and tumor progression in mouse models of triple negative breast cancer

Introduction: Advanced age and obesity are independent risk and progression factors for triple negative breast cancer (TNBC), which presents significant public health concerns for the aging population and its increasing burden of obesity. Due to parallels between advanced age- and obesityrelated biology, particularly adipose inflammation, we hypothesized that advanced age and obesity each accelerate mammary tumor growth through convergent, and likely interactive, mechanisms. Methods: To test this hypothesis, we orthotopically transplanted murine syngeneic TNBC cells into the mammary glands of young normoweight control (7 months), young diet-induced obese (DIO), aged normoweight control (17 months), and aged DIO female C57BL/6J mice. Results: Here we report accelerated tumor growth in aged control and young DIO mice, compared with young controls. Transcriptional analyses revealed, with a few exceptions, overlapping patterns of mammary tumor inflammation and tumor immunosuppression in aged control mice and young DIO mice, relative to young controls. Moreover, aged control and young DIO tumors, compared with young controls, had reduced abundance ofcytotoxic CD8 T cells. Finally, DIO in advanced age exacerbated mammary tumor growth, inflammation and tumor immunosuppression. Discussion: These findings demonstrate commonalities in the mechanisms driving TNBC in aged and obese mice, relative to young normoweight controls. Moreover, we found that advanced age and DIO interact to accelerate mammary tumor progression. Given the US population is getting older and more obese, age- and obesity-related biological differences will need to be considered when developing mechanism-based strategies for preventing or controlling breast cancer

The Hidden Story of Heterogeneous B-raf V600E Mutation Quantitative Protein Expression in Metastatic Melanoma-Association with Clinical Outcome and Tumor Phenotypes

In comparison to other human cancer types, malignant melanoma exhibits the greatest amount of heterogeneity. After DNA-based detection of the BRAF V600E mutation in melanoma patients, targeted inhibitor treatment is the current recommendation. This approach, however, does not take the abundance of the therapeutic target, i.e., the B-raf V600E protein, into consideration. As shown by immunohistochemistry, the protein expression profiles of metastatic melanomas clearly reveal the existence of inter-and intra-tumor variability. Nevertheless, the technique is only semi-quantitative. To quantitate the mutant protein there is a fundamental need for more precise techniques that are aimed at defining the currently non-existent link between the levels of the target protein and subsequent drug efficacy. Using cutting-edge mass spectrometry combined with DNA and mRNA sequencing, the mutated B-raf protein within metastatic tumors was quantitated for the first time. B-raf V600E protein analysis revealed a subjacent layer of heterogeneity for mutation-positive metastatic melanomas. These were characterized into two distinct groups with different tumor morphologies, protein profiles and patient clinical outcomes. This study provides evidence that a higher level of expression in the mutated protein is associated with a more aggressive tumor progression. Our study design, comprised of surgical isolation of tumors, histopathological characterization, tissue biobanking, and protein analysis, may enable the eventual delineation of patient responders/non-responders and subsequent therapy for malignant melanoma

Evaluation of SLOG/TCI-III pediatric system on target control infusion of propofol

<p>Abstract</p> <p>Background</p> <p>The target-controlled infusion-III (SLOG/TCI-III) system was derived from a model set up by the local pediatric population for target control infusion of propofol.</p> <p>Methods</p> <p>The current study aimed at evaluating the difference between target concentrations of propofol and performance, which was measured using the SLOG/TCI-III system in children. Thirty children fulfilling the I-II criteria according to American Society of Anesthesiology were enrolled in the study. The target plasma concentration of propofol was fed into the SLOG/TCI-III system and compared with the measured concentrations of propofol. Blood samples were collected and analyzed by high performance liquid chromatography with fluorescence detector. The performance error (PE) was determined for each measured blood propofol concentration. The performances of the TCI-III system were determined by the median performance error (MDPE), the median absolute performance error (MDAPE), and Wobble (the median absolute deviation of each PE from the MDPE), respectively.</p> <p>Results</p> <p>Concentration against target concentration showed good linear correlation: concentration = 1.3428 target concentration - 0.2633 (r = 0.8667). The MDPE and MDAPE of the pediatric system were 10 and 22%, respectively, and the median value for Wobble was 24%. MDPE and MDAPE were less than 15 and 30%, respectively.</p> <p>Conclusions</p> <p>The performance of TCI-III system seems to be in the accepted limits for clinical practice in children.</p