Shoulder dystocia documentation: an evaluation of a documentation training intervention.

OBJECTIVE: To evaluate the quality and content of nurse and physician shoulder dystocia delivery documentation before and after MORE training in shoulder dystocia management skills and documentation. METHODS: Approximately 384 charts at the Ottawa Hospital General Campus involving a diagnosis of shoulder dystocia between the years of 2000 and 2006 excluding the training year of 2003 were identified. The charts were evaluated for 14 key components derived from a validated instrument. The delivery notes were then scored based on these components by 2 separate investigators who were blinded to delivery note author, date, and patient identification to further quantify delivery record quality. RESULTS: Approximately 346 charts were reviewed for physician and nurse delivery documentation. The average score for physician notes was 6 (maximum possible score of 14) both before and after the training intervention. The nurses' average score was 5 before and after the training intervention. CONCLUSIONS: Negligible improvement was observed in the content and quality of shoulder dystocia documentation before and after nurse and physician training.

Toxic shock syndrome of a probable gynecologic source in an adolescent: a case report and review of the literature.

BACKGROUND: Toxic shock syndrome (TSS) is an acute toxin-mediated infectious syndrome characterized by fever, hypotension, desquamation, and multiorgan involvement. It is a rare condition (incidence of 0.79/100,000 women), particularly in the adolescent population, and it may be menstrual (mTSS) or non-menstrual (nmTSS) in origin. CASE: A 15-year-old girl developed symptoms of nausea, vomiting, and diarrhea that worsened over a 3-day period. At initial presentation, she was hypotensive, febrile, and tachycardic. Her condition deteriorated and within 36 hours she required intubation, vasopressor treatment, and antibiotic therapy. Multiple sites were cultured but only the vaginal culture, which grew Staphylococcus aureus, was positive. Recent menses with tampon use was reported. She responded to aggressive therapy and was discharged home 3 weeks after initial presentation. SUMMARY AND CONCLUSION: We describe a rare case of TSS of a probable gynecologic source in a 15-year-old female who successfully responded to aggressive intensive care treatment. mTSS should be considered in the differential diagnosis of an adolescent presenting with signs of septic shock, particularly if there is a recent history of tampon use. Early intervention is critical to improving survival.

The discovery of MK-0674, an orally bioavailable cathepsin K inhibitor.

MK-0674 is a potent and selective cathepsin K inhibitor from the same structural class as odanacatib with a comparable inhibitory potency profile against Cat K. It is orally bioavailable and exhibits long half-life in pre-clinical species. In vivo studies using deuterated MK-0674 show stereoselective epimerization of the alcohol stereocenter via an oxidation/reduction cycle. From in vitro incubations, two metabolites could be identified: the hydroxyleucine and the glucuronide conjugate which were confirmed using authentic synthetic standards.

The discovery of odanacatib (MK-0822), a selective inhibitor of cathepsin K.

Odanacatib is a potent, selective, and neutral cathepsin K inhibitor which was developed to address the metabolic liabilities of the Cat K inhibitor L-873724. Substituting P1 and modifying the P2 side chain led to a metabolically robust inhibitor with a long half-life in preclinical species. Odanacatib was more selective in whole cell assays than the published Cat K inhibitors balicatib and relacatib. Evaluation in dermal fibroblast culture showed minimal intracellular collagen accumulation relative to less selective Cat K inhibitors.

Enzyme occupancy measurement of intracellular protein tyrosine phosphatase 1B using photoaffinity probes.

Protein tyrosine phosphatase 1B (PTP1B) is believed to be one of the enzymes involved in down-regulating the insulin receptor and is a drug target for the treatment of type II diabetes. To better understand the in vitro and in vivo behavior of PTP1B inhibitors, a cell-based assay to directly measure enzyme occupancy of PTP1B by inhibitors using photoaffinity labeling was developed. Two photoaffinity probes were synthesized containing the photolabile diazirine moiety. These photoprobes were specific for PTP1B and T-cell protein tyrosine phosphatase over CD45, with the most potent photoprobe having an IC(50) value of 0.2nM for PTP1B. Activation of the photoprobes with a 40-W UV lamp in the presence of purified AspTyrLysAspAspAspAspLys (Flag)-PTP1B formed a 1:1 irreversible adduct with the enzyme. The photolabeling was competed by known PTP1B inhibitors, vanadate, and the peptide inhibitor N-benzoyl-l-glutamyl-[4-phosphono(difluoromethyl)]-l-phenylalanyl-[4-phosphono(difluoromethyl)]l-phenylalanineamide (BzN-EJJ-amide). In HepG2 (human hepatoma cell line) cells, endogenous PTP1B was labeled by the UV-activated photoprobes in both lysed and intact cells. Enzyme occupancy measurements were conducted with a series of PTP1B inhibitors using the photoprobe affinity assay. Several compounds were shown to bind to endogenous PTP1B in the HepG2 intact cells.

Detection of covalent adducts to cytochrome P450 3A4 using liquid chromatography mass spectrometry.

Protein covalent labeling can be an undesirable property of compounds being studied in drug discovery programs. Identifying such compounds relies on the use of radiolabeled material, which requires an investment in time and resources not typically expended until later in the discovery process. We describe the detection of covalent adducts to cytochrome P450 3A4, the most abundant and important P450 from a human and drug discovery viewpoint, using liquid chromatography mass spectrometry. The technique is illustrated using L-754,394 and 6',7'-dihydroxybergamottin, two known inhibitors of P450 3A4. Mass spectrometry of the intact apoprotein as well as the adducted protein is demonstrated. Such methodology may provide the means for screening compounds for covalent protein binding without the use of a radiolabel. It also provides direct information about mechanism-based inhibitors in terms of extent, stoichiometry, and nature of the adduct(s) (mass shift). This information may provide a means for understanding the mechanism of covalent labeling earlier in a drug discovery environment.

Synthesis of a novel peptidic photoaffinity probe for the PTP-1B enzyme.

The synthesis of a novel radioactive peptidic photoaffinity probe for the PTP-1B enzyme as well as some SAR leading to the choice of this compound as a photoaffinity probe are presented.

Catalytic inactivation of protein tyrosine phosphatase CD45 and protein tyrosine phosphatase 1B by polyaromatic quinones.

Polyaromatic quinones, such as the environmental pollutants 9,10-phenanthrenediones, elicit a wide range of responses including growth inhibition, immune suppression, and glucose normalization in diabetic models. Yet the molecular mechanisms behind these effects remain controversial. Here we report that many of them are oxygen-dependent and catalytic inactivators of protein tyrosine phosphatases (PTP). Under aerobic conditions, the PTP inactivation by 2-nitro-9,10-phenanthrenedione followed a pseudo-first-order process, with the rate of inactivation increasing nearly linearly with increasing inhibitor concentration, yielding apparent inactivation rate constants of 4300, 387, and 5200 M(-1) s(-1) at pH 7.2 against CD45, PTP1B, and LAR, respectively. The rate of CD45 inactivation increased approximately 25-fold from pH 6.0 to 7.5, with complete inactivation achieved using a catalytic amount (0.05 molar equiv) of the inhibitor. The quinone-catalyzed CD45 inactivation was prevented by catalase or superoxide dismutase. Inactivated CD45 after (125)I-9,10-phenanthrenedione treatment carried no radioactivity, indicating the absence of a stable inhibitor/enzyme complex. The activity of inactivated CD45 was partially restored ( approximately 10%) by hydroxylamine or dithiothreitol, supporting the presence of a small population of sulfenic acid or sulfenyl-amide species. Treatment of PTP1B with 2-nitro-9,10-phenanthrenedione resulted in the specific and sequential oxidation of the catalytic cysteine to the sulfinic and sulfonic acid. These results suggest that reactive oxygen species and the semiquinone radical, continuously generated during quinone-catalyzed redox cycling, mediate the specific catalytic cysteine oxidation. Naturally occurring quinones may act as efficient regulators of protein tyrosine phosphorylation in biological systems. Aberrant phosphotyrosine homeostasis resulting from continued polyaromatic hydrocarbon quinone exposure may play a significant role in their disease etiology.

Application of microfluidic devices to proteomics research: identification of trace-level protein digests and affinity capture of target peptides.

This report describes an integrated and modular microsystem providing rapid analyses of trace-level tryptic digests for proteomics applications. This microsystem includes an autosampler, a microfabricated device comprising a large channel (2.4 microl total volume), an array of separation channels, together with a low dead volume enabling the interface to nanoelectrospray mass spectrometry. The large channel of this microfluidic device provides a convenient platform to integrate C(18) reverse phase packing or other type of affinity media such as immobilized antibodies or immobilized metal affinity chromatography beads thus enabling affinity selection of target peptides prior to electrophoretic separation and mass spectrometry analyses on a quadrupole/time-of-flight instrument. Sequential injection, preconcentration, and separation of peptide standards and tryptic digests are achieved with a throughput of up to 12 samples/per h and a concentration detection limit of approximately 5 nM (25 fmol on chip). Replicate injections of peptide mixtures indicated that reproducibility of migration time was 1.2-1.8%, whereas relative standard deviation ranging from 9.2 to 11.8% are observed on peak heights. The application of this device for trace-level protein identification is demonstrated for two-dimensional gel spots obtained from extracts of human prostatic cancer cells (LNCap) using both peptide mass-fingerprint data base searching and on-line tandem mass spectrometry. Enrichment of target peptides prior to mass spectral analyses is achieved using c-myc-specific antibodies immobilized on protein G-Sepharose beads and facilitates the identification of antigenic peptides spiked at a level of 20 ng/ml in human plasma. Affinity selection is also demonstrated for gel-isolated protein bands where tryptic phosphopeptides are captured on immobilized metal affinity chromatography beads and subsequently separated and characterized on this microfluidic system.

Hyphenation of capillary high-performance liquid chromatography with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for nano-scale screening of single-bead combinatorial libraries.

This paper focuses on the technical aspects of chemical screening from 384-well plate nano-scale single-bead combinatorial libraries. The analytical technique utilized is a combination of capillary liquid chromatography with ultraviolet detection and matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The HPLC/MALDI-MS hyphenation is achieved by means of a micro-fraction collector with a peak detection system that automatically collects the peaks onto the MALDI targets for subsequent characterization. Several experimental parameters such as type of 384-well plate, well-plate sealing foils, and a column-switching procedure were investigated using a small test library of nine components. Additionally, the influence of different MALDI matrices, different MALDI targets and sample-spotting techniques on the MALDI detection sensitivity as well as the ruggedness and sample throughput capacity of this technique were studied. Optimum results for the analytes investigated were obtained with 2,5-dihydroxybenzoic acid using on-line mixing of HPLC effluent and matrix solution. To demonstrate the potential of this capillary HPLC/MALDI-TOFMS method, its application to several single-bead libraries was investigated. The instrumental method allowed for the rapid identification and purity assessment of combinatorial libraries with detection limits down to the higher femtomole level using both UV detection and MALDI mass spectrometry.