Objective: This trial was to test whether mycophenolate mofetil (MMF) alone or with daclizumab (DZB) could arrest the loss of insulin-producing beta cells in subjects with new onset type 1 diabetes (T1D). Research Design and Methods: A multicenter, randomized, placebo-controlled, double-masked trial was initiated by Type 1 Diabetes TrialNet at 13 sites in North America and Europe. Subjects diagnosed with type 1 diabetes and with sufficient C-peptide within 3 months of diagnosis were randomized to either: MMF-alone, MMF and DZB, or placebo, and then followed for 2 years. The primary outcome was the geometric mean area under the curve (AUC) C-peptide from the 2 hour mixed meal tolerance test. Results: One hundred and twenty-six subjects were randomized and treated during the trial. The geometric mean C-peptide AUC at 2 years was unaffected by MMF-alone, or MMF and DZB, versus placebo. Adverse events were more frequent in the active therapy groups relative to the control group, but not significantly. Conclusions: Neither MMF-alone nor MMF in combination with DZB had an effect on the loss of C-peptide in subjects with new onset T1D. Higher doses or more targeted immunotherapies may be needed to affect the autoimmune process.

Objective- To describe a laparoscopic-sutured gastropexy technique in dogs and evaluate the tensile strength of the adhesion and effects on gastric function. Study Design- Experimental study. Animals- Female beagle dogs (n=7). Methods- A laparoscopic-sutured gastropexy technique was evaluated by ex vivo tensile distraction tests 10 weeks after surgery. The effect of the adhesion on gastric emptying, mucosal permeability, and systemic inflammation were evaluated by monitoring the C-reactive protein (CRP) and sucrose permeability, and by radiographic evaluation of gastric emptying 2 weeks before and 10 weeks after surgery. Results- Mean (+/-SD) tensile force to disrupt adhesions was 51.1+/-16.4 N. There was no significant postoperative increase in CRP concentration or change in sucrose permeability. The area under the curve representing the postprandial decrease in gastric radiographic area increased by 11% after gastropexy. Conclusions- This laparoscopic gastropexy technique had appropriate mechanical and functional characteristics with limited morbidity. Clinical Relevance- This laparoscopic-sutured gastropexy provides adhesion strength comparable with other gastropexy techniques tested at 10 weeks postoperatively. Only minor changes in gastric emptying were observed 10 weeks after surgery.

Clinical proteomics has the potential to enable the early detection of cancer through the development of multiplex assays that can inform clinical decisions. However, there has been some uncertainty among translational researchers and developers as to the specific analytical measurement criteria needed to validate protein-based multiplex assays. To begin to address the causes of this uncertainty, a day-long workshop titled "Interagency Oncology Task Force Molecular Diagnostics Workshop" was held in which members of the proteomics and regulatory communities discussed many of the analytical evaluation issues that the field should address in development of protein-based multiplex assays for clinical use. This meeting report explores the issues raised at the workshop and details the recommendations that came out of the day's discussions, such as a workshop summary discussing the analytical evaluation issues that specific proteomic technologies should address when seeking US Food and Drug Administration approval.

As a part of ongoing efforts of the NCI-FDA Interagency Oncology Task Force subcommittee on molecular diagnostics, members of the Clinical Proteomics Technology Assessment for Cancer program of the National Cancer Institute have submitted 2 protein-based multiplex assay descriptions to the Office of In Vitro Diagnostic Device Evaluation and Safety, US Food and Drug Administration. The objective was to evaluate the analytical measurement criteria and studies needed to validate protein-based multiplex assays. Each submission described a different protein-based platform: a multiplex immunoaffinity mass spectrometry platform for protein quantification, and an immunological array platform quantifying glycoprotein isoforms. Submissions provided a mutually beneficial way for members of the proteomics and regulatory communities to identify the analytical issues that the field should address when developing protein-based multiplex clinical assays.

A source localization analysis was carried out to provide brain functional and structural assessments of individuals with poor reading skills. Standardized low-resolution brain electromagnetic tomography was used to locate sources of P2 and P3 event-related potential components in normal readers and in poor reader children performing a cued continuous performance task. Cue-elicited P2 sources in the right superior parietal gyrus were smaller in 37 poor readers than in 40 normal readers. Poor readers showed a higher P3 activation in response to a false target in frontal and frontorbital regions than normal readers. These results suggest that reading disabilities may be attributed to failures in attentional focalization for incoming stimuli.

BACKGROUND: The immunopathogenesis of type 1 diabetes mellitus is associated with T-lymphocyte autoimmunity. However, there is growing evidence that B lymphocytes play a role in many T-lymphocyte-mediated diseases. It is possible to achieve selective depletion of B lymphocytes with rituximab, an anti-CD20 monoclonal antibody. This phase 2 study evaluated the role of B-lymphocyte depletion in patients with type 1 diabetes. METHODS: We conducted a randomized, double-blind study in which 87 patients between 8 and 40 years of age who had newly diagnosed type 1 diabetes were assigned to receive infusions of rituximab or placebo on days 1, 8, 15, and 22 of the study. The primary outcome, assessed 1 year after the first infusion, was the geometric mean area under the curve (AUC) for the serum C-peptide level during the first 2 hours of a mixed-meal tolerance test. Secondary outcomes included safety and changes in the glycated hemoglobin level and insulin dose. RESULTS: At 1 year, the mean AUC for the level of C peptide was significantly higher in the rituximab group than in the placebo group. The rituximab group also had significantly lower levels of glycated hemoglobin and required less insulin. Between 3 months and 12 months, the rate of decline in C-peptide levels in the rituximab group was significantly less than that in the placebo group. CD19+ B lymphocytes were depleted in patients in the rituximab group, but levels increased to 69% of baseline values at 12 months. More patients in the rituximab group than in the placebo group had adverse events, mostly grade 1 or grade 2, after the first infusion. The reactions appeared to be minimal with subsequent infusions. There was no increase in infections or neutropenia with rituximab. CONCLUSIONS: A four-dose course of rituximab partially preserved beta-cell function over a period of 1 year in patients with type 1 diabetes. The finding that B lymphocytes contribute to the pathogenesis of type 1 diabetes may open a new pathway for exploration in the treatment of patients with this condition.(ClinicalTrials.gov number, NCT00279305.) Copyright 2009 Massachusetts Medical Society.

We have found that 1-alkyl-3-methylimidazolium chloride ionic liquids (ILs) can form immiscible liquid mixtures with some polyethylene glycols (PEGs). Binary mixtures of 1-ethyl-3-methylimidazolium chloride with PEG of molecular weight 1500, 2000, or 3400 g mol(-1), or of 1-butyl-3-methylimidazolium chloride with PEG of molecular weight 2000 or 3400 g mol(-1), have been found to give rise to entirely liquid, stable biphasic systems over a significant temperature range (from 333.15 K to 413.15 K), while mixtures of 1-ethyl-3-methylimidazolium chloride with PEG-1000 and 1-butyl-3-methylimidazolium chloride with PEG-1000 and PEG-1500 are miscible. The mutual immiscibility of the IL and the PEG increases as the temperature increases. The evolution of the composition of the phases in equilibrium with the molecular weight of the PEG, or with the variation of the length of the alkyl substituent chain of the imidazolium cation of the IL, has been explored. The trends observed are explained through the complexity of interactions present within the binary system. A thermodynamic analysis of the liquid-liquid equilibrium data indicates negative values for the change of enthalpy and entropy of mixing. The potential application of these biphasic, entirely liquid systems, with low volatility and good solvation properties, for the dissolution and separation of cellulose and lignin at elevated temperature has been preliminarily explored, although only modest results have been achieved to date.

The complexity of proteomic instrumentation for LC-MS/MS introduces many possible sources of variability. Data-dependent sampling of peptides constitutes a stochastic element at the heart of discovery proteomics. Although this variation impacts the identification of peptides, proteomic identifications are far from completely random. In this study, we analyzed interlaboratory data sets from the NCI Clinical Proteomic Technology Assessment for Cancer to examine repeatability and reproducibility in peptide and protein identifications. Included data spanned 144 LC-MS/MS experiments on four Thermo LTQ and four Orbitrap instruments. Samples included yeast lysate, the NCI-20 defined dynamic range protein mix, and the Sigma UPS 1 defined equimolar protein mix. Some of our findings reinforced conventional wisdom, such as repeatability and reproducibility being higher for proteins than for peptides. Most lessons from the data, however, were more subtle. Orbitraps proved capable of higher repeatability and reproducibility, but aberrant performance occasionally erased these gains. Even the simplest protein digestions yielded more peptide ions than LC-MS/MS could identify during a single experiment. We observed that peptide lists from pairs of technical replicates overlapped by 35-60%, giving a range for peptide-level repeatability in these experiments. Sample complexity did not appear to affect peptide identification repeatability, even as numbers of identified spectra changed by an order of magnitude. Statistical analysis of protein spectral counts revealed greater stability across technical replicates for Orbitraps, making them superior to LTQ instruments for biomarker candidate discovery. The most repeatable peptides were those corresponding to conventional tryptic cleavage sites, those that produced intense MS signals, and those that resulted from proteins generating many distinct peptides. Reproducibility among different instruments of the same type lagged behind repeatability of technical replicates on a single instrument by several percent. These findings reinforce the importance of evaluating repeatability as a fundamental characteristic of analytical technologies.

Optimal performance of LC-MS/MS platforms is critical to generating high quality proteomic data. Although individual laboratories have developed quality control samples, there is no widely available performance standard of biological complexity (and associated reference datasets) for benchmarking of platform performance for analysis of complex biological proteomes across different laboratories in the community. Individual preparations of the yeast Saccharomyces cerevisiae proteome have been used extensively by laboratories in the proteomic community to characterize LC-MS platform performance. The yeast proteome is uniquely attractive as a performance standard since it is the most extensively characterized complex biological proteome, and the only one associated with several large-scale studies estimating the abundance of all detectable proteins. In this study, we describe a standard operating protocol for large-scale production of the yeast performance standard, and offer aliquots to the community through the National Institute of Standards and Technology, where the yeast proteome is under development as a certified reference material to meet the long-term needs of the community. Using a series of metrics that characterize LC-MS performance, we provide a reference dataset demonstrating typical performance of commonly used ion trap instrument platforms in expert laboratories; the results provide a basis for laboratories to benchmark their own performance, to improve upon current methods, and to evaluate new technologies. Additionally, we demonstrate how the yeast reference, spiked with human proteins, can be used to benchmark the power of proteomic platforms for detection of differentially expressed proteins at different levels of concentration in a complex matrix, thereby providing a metric to evaluate and minimize pre-analytical and analytical variation in comparative proteomics experiments.

A major unmet need in liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analyses is a set of tools for quantitative assessment of system performance and evaluation of technical variability. Here we describe 46 system performance metrics for monitoring chromatographic performance, electrospray source stability, MS1 and MS2 signals, dynamic sampling of ions for MS/MS and peptide identification. Applied to datasets from replicate LC-MS/MS analyses, these metrics display consistent, reasonable responses to controlled perturbations. The metrics typically display variations less than 10% and thus can reveal even subtle differences in performance of system components. Analyses of data from interlaboratory studies conducted under a common standard operating procedure identified outlier data and provided clues to specific causes. Moreover, interlaboratory variation reflected by the metrics indicates which system components vary the most between laboratories. Application of these metrics enables rational, quantitative quality assessment for proteomics and other LC-MS/MS analytical applications.