Laser fetoscopy ablation for twin-twin transfusion syndrome may reduce the risk of ROP.

PURPOSE: Twin-twin transfusion syndrome (TTTS) is a condition wherein monochorionic twins share a common placenta with placental anastomoses between the two foetal circulations. Most infants who survive TTTS are born prematurely. This study aimed to determine whether fetoscopic laser ablation (FLA) can reduce the risk of retinopathy of prematurity (ROP) and whether TTTS was a risk factor for ROP. METHODS: This single-centre, retrospective, comparative study included 32 monochorionic twins with TTTS matched for gestational age, birthweight and sex to premature twins and singletons without TTTS (n = 68; twins, n = 34; and singletons, n = 34) born between 2003 and 2022. A single ophthalmologist recorded the fundus findings. FLA was performed using Solomon's technique to separate the vascular systems of the twins with TTTS. RESULTS: The gestational age and weight of premature infants with TTTS treated with FLA were significantly higher than those of untreated infants (p = 0.001 and p = 0.001, respectively); however, the hyaline membrane grade was lower (p = 0.004). A significant increase in weight (g/day) (p = 0.002) and lesser avascular area in the peripheral temporal retina (p = 0.045) was observed at postnatal week 4. The risk of ROP in the FLA group was 2.6 times (13.3% vs. 35.3%) lower than that in the non-FLA group; however, this difference was not significant. The incidence of any stage of ROP (25% vs. 18%) and treatment for ROP type 1 (6.25% vs. 5.9%) did not differ significantly between monochorionic twins with TTTS and premature infants without TTTS. CONCLUSION: The gestational age of premature infants with TTTS treated with FLA was higher than that of untreated infants. Moreover, a reduction in complications of prematurity was also observed. Laser fetoscopy in twin-twin transfusion syndrome may reduce the risk of ROP, but the difference was not statistically significant in this small study.

Patterns of school and college of pharmacy engagement in addressing the opioid crisis.

INTRODUCTION: The American Association of Colleges of Pharmacy (AACP) has emphasized the need to prepare pharmacy students for practicing amidst the opioid crisis. This research aimed to identify patterns and predictors of pharmacy program participation in skills-based education, research, and service activities designed to address this crisis. METHODS: Opioid-related activities were identified from the AACP opioid-related activities database and classified by two independent reviewers. The final activities included: (1) direct participation in drug disposal and/or naloxone outreach, (2) opioid-focused research, and (3) skills-based training in the doctor of pharmacy curriculum. Latent class analysis was used to identify classes of program involvement in these activities. Differences in class membership based on program and geographic characteristics were examined using multivariable logistic regression. RESULTS: Of the 106 schools included, a minority reported opioid-focused research (38.7%), drug disposal or naloxone outreach (30.2%), or hands-on learning (22.6%). A "highly engaged" class (34.9%) and a "limited engagement" class (65.1%) were identified. "Highly engaged" programs were more likely to report opioid-related research (65.9% vs. 24.6%, P < .001), drug disposal or naloxone outreach events (86.5% vs. 0%, P < .001), and skills-based education (40.5% vs. 13%, P = .001) than "limited engagement" programs. No school or geographic factors were significantly associated with class membership. CONCLUSIONS: Nearly two-thirds of schools and colleges of pharmacy reported limited involvement in skills-based education, research, and outreach efforts. Future research should explore other predictors of school-level opioid-related activities, including faculty expertise and institutional priorities.

Effect of post-operative oral steroids on bleeding rate after pediatric tonsillectomy.

PURPOSE: In 2013, the FDA placed a black box warning on the usage of opioid pain medications in the post-operative setting after pediatric adenotonsillectomy. Since then, alternative pain management regimens have been employed. Some have advocated for post-operative oral steroids, in part due to the effectiveness of intraoperative intravenous steroids in reducing post-operative pain and nausea. The evidence regarding the efficacy and safety of post-operative oral steroids is not as clear. The purpose of this study was to examine whether post-tonsillectomy hemorrhage rates in pediatric patients were affected by post-operative oral steroid usage. MATERIALS AND METHODS: Case-control retrospective chart review using a deidentified data set of patients undergoing tonsillectomy with or without adenoidectomy at a single academic medical center between June 2012 and November 2015. RESULTS: A total of 1416 patients were included in the study, with 704 in the no post-operative oral steroids group and 712 in the group who did receive post-operative oral steroids. The rate of post-tonsillectomy hemorrhage in the post-operative oral steroid group was 3.1 % compared to 1.8 % in the group who did not receive post-operative oral steroids, however, this was not a statistically significant difference (P = .132). CONCLUSIONS: Our study suggests that post-operative oral steroids are safe and do not increase the risk of post-operative hemorrhage after pediatric tonsillectomy.

Utilizing Probiotics for the Prevention and Treatment of Gastrointestinal Diseases.

Probiotics are heavily advertised to promote a healthy gastrointestinal tract and boost the immune system. This review article summarizes the history and diversity of probiotics, outlines conventional in vitro assays and in vivo models, assesses the pharmacologic effects of probiotic and pharmaceutical co-administration, and the broad impact of clinical probiotic utilization for gastrointestinal disease indications.

Tracheoinnominate Artery Fistula.

Audience: This simulation provides training for emergency medicine residents in the stepwise management of a patient who presents with bleeding from a tracheoinnominate artery fistula. Additional learners who might benefit from this simulation are otolaryngology and general surgery residents as well as critical care fellows. Introduction: Hemorrhage from a tracheoinnominate artery fistula (TIAF) is a rare but life-threatening complication in a patient with a recent tracheostomy. This complication occurs in 0.7% of tracheostomy patients with a mortality of 50-70%.1 Seventy-five percent of patients with a TIAF will present within the first three weeks of surgery and 50% of patients will present with a sentinel bleed that briefly resolves.1 Key elements of a history and exam that should raise a provider's concern for this diagnosis include a recent tracheostomy (within the last 4 weeks), a percutaneous tracheostomy, prior radiation, chronic steroid use, a neck or chest deformity or a sentinel bleed.2 Survival from a TIAF hinges upon emergent, operative repair by an otolaryngologist and cardiothoracic surgeon. Cuff hyperinflation and the Utley Maneuver are critical bedside interventions to temporize this massive bleed and stabilize the patient for definitive, operative repair. Educational Objectives: By the end of this simulation, learners will be able to: 1) perform a focused history and physical exam on any patient who presents with bleeding from the tracheostomy site, 2) describe the differential diagnosis of bleeding from a tracheostomy site, including a TIAF, 3) demonstrate the stepwise management of bleeding from a suspected TIAF, including cuff hyperinflation and the Utley Maneuver, 4) verify that definitive airway control via endotracheal intubation is only feasible in the tracheostomy patient when it is clear, upon history and exam, that the patient can be intubated from above, 5) demonstrate additional critical actions in the management of a patient with a TIAF, including early consultation with otolaryngology and cardiothoracic surgery as well as emergent blood transfusion and activation of a massive transfusion protocol. Educational Methods: This case was written with a modified, low-fidelity manikin, traditionally used for training in nasogastric tube placement and tracheostomy care. We modified this manikin to simulate a hemorrhage from the tracheostomy site.3 The patient in our case had a history of laryngeal cancer, and thus we occluded his larynx for this simulation. As a result of this obstruction, he was unable to be intubated from above. We provided confederates, a bedside nurse and family member, to assist the learners throughout the case. We also utilized a simulation technician to operate dynamic vital signs on a simulated cardiac monitor. It would be technically challenging to adapt this case to a high-fidelity simulator due to potential for damage of the internal electrical elements by the large amount of artificial blood from the tracheostomy tube. However, a mechanical pump provided a useful means of active bleeding in this low-fidelity manikin. Research Methods: We provided a pre- and post-simulation questionnaire for the 33 emergency medicine residents who participated in this simulation. There were 11 residents from each of the PGY-1, PGY-2 and PGY-3 year-groups. Thirty-two residents (97%) completed the pre-survey and 33 residents (100%) completed the post-survey. For our questions, we used a 5-point Likert Scale to assess a resident's knowledge of the learning objectives within this simulation. Results: Responses from our pre- and post- survey indicated a significant improvement in knowledge about a tracheoinnominate artery fistula as well as the general management of tracheostomy complications in the emergency department. Discussion: This simulation is a useful educational tool for instructing emergency medicine residents on optimal management of tracheostomy emergencies such as a TIAF. The interprofessional teaching by an emergency medicine attending and mid-level (PGY-3) otolaryngology resident allowed for a richer and more detailed discussion during the debriefing. Throughout the case, the emergency medicine attending played the role of a bedside nurse and offered supportive, clinical cues when bleeding recurred. The otolaryngology resident played the role of a family member and offered helpful cues during the history and exam portion of the case. Following the case, both content experts provided useful clinical insight during the debriefing. If staffing availability permits, it might be advantageous to use additional simulation-trained personnel to play the roles of the nurse and family member, thus allowing the emergency medicine attending and otolaryngology content experts to simply view the case from the control room and perform the debriefing. Topics: Tracheostomy, surgical airway, tracheoinnominate artery fistula, bleeding from tracheostomy site, complications with tracheostomies, hemorrhagic shock.

Two cases of rare subglottic MALT lymphoma of the larynx.

BACKGROUND: Primary MALT lymphoma of the larynx is a rare condition first described in 1990. There have been only 43 reported cases as of 2015. The disease appears to be indolent in nature and responds well to radiation therapy. Symptoms are non-specific and may be limited to a combination of hoarseness, sore throat, shortness of breath, or cough. METHODS: We describe two cases of subglottic laryngeal MALT lymphoma identified from one academic medical center within five years of each other. Though identical in pathology, the presentation of the two cases were distinct in both patient demographic and tumor appearance. One patient required dilation of a subglottic stenosis caused by tumor, and the other required surgical debulking of a ball-valve-like mass. Neither patient presented with B-symptoms (fever, night sweats, weight loss) that often characterize other lymphomas. RESULTS: In both cases, histopathological exam revealed extensive infiltration of mucosa with atypical monomorphous lymphocytes, consistent with MALT lymphoma. CONCLUSION: MALT lymphoma of the larynx may present with non-specific symptoms such as cough and/or hoarseness. Thorough evaluation including flexible laryngoscopy should be performed should these symptoms persist without a known cause. Surgical biopsy and histopathological exam are crucial to determine the etiology of unknown subglottic masses.

Broadband THz absorption spectrometer based on excitonic nonlinear optical effects.

A broadly tunable THz source is realized via difference frequency generation, in which an enhancement to χ(3) that is obtained via resonant excitation of III-V semiconductor quantum well excitons is utilized. The symmetry of the quantum wells (QWs) is broken by utilizing the built-in electric-field across a p-i-n junction to produce effective χ(2) processes, which are derived from the high χ(3). This χ(2) media exhibits an onset of nonlinear processes at ~4 W cm-2, thereby enabling area (and, hence, power) scaling of the THz emitter. Phase matching is realized laterally through normal incidence excitation. Using two collimated 130 mW continuous wave (CW) semiconductor lasers with ~1-mm beam diameters, we realize monochromatic THz emission that is tunable from 0.75 to 3 THz and demonstrate the possibility that this may span 0.2-6 THz with linewidths of ~20 GHz and efficiencies of ~1 × 10-5, thereby realizing ~800 nW of THz power. Then, transmission spectroscopy of atmospheric features is demonstrated, thereby opening the way for compact, low-cost, swept-wavelength THz spectroscopy.

Frailty as a Predictor of Postoperative Outcomes among Patients with Head and Neck Cancer.

OBJECTIVES: To understand measures of frailty among preoperative patients and explain how these can predict perioperative outcomes among patients with head and neck cancer. STUDY DESIGN: Retrospective cross-sectional case series with chart review. SETTING: Academic tertiary medical center. SUBJECTS AND METHODS: A retrospective review was performed of patients presenting to an academic hospital following a surgical procedure for a head and neck cancer diagnosis. Charts were queried for preoperative medical diagnoses to calculate 2 frailty scores: the American College of Surgeons National Surgical Quality Improvement Program modified frailty index and the Johns Hopkins Adjusted Clinical Groups frailty index. The American Society of Anesthesiologists classification system was also analyzed as a predictor. Primary outcomes were mortality, 30-day readmission, and length of stay. Perioperative complications and discharge disposition were also evaluated. RESULTS: A total of 410 charts were queried between January 2014 and December 2017. Mortality was 11%; mean ± SD length of stay was 7.4 ± 5.5 days; and the readmission rate was 17%. The modified frailty index score significantly increased the odds of mortality (odds ratio = 1.475, P = .012) and readmission (odds ratio = 1.472, P = .004), the length of stay (relative risk = 1.136, P = .001), and the number of perioperative complications. The American Society of Anesthesiologists classification was also significantly associated with poor outcomes, including readmission, length of stay, and perioperative complications. The Adjusted Clinical Groups index was not a significant predictor of outcomes in this study population. CONCLUSIONS: This study demonstrated a significant increase in poor perioperative outcomes and mortality among patients with head and neck cancer and increased frailty, as measured by the modified frailty index.

Active monitoring of formaldehyde diffusion into histological tissues with digital acoustic interferometry.

The preservation of certain labile cancer biomarkers with formaldehyde-based fixatives can be considerably affected by preanalytical factors such as quality of fixation. Currently, there are no technologies capable of quantifying a fixative's concentration or the formation of cross-links in tissue specimens. This work examined the ability to detect formalin diffusion into a histological specimen in real time. As formaldehyde passively diffused into tissue, an ultrasound time-of-flight (TOF) shift of several nanoseconds was generated due to the distinct sound velocities of formalin and exchangeable fluid within the tissue. This signal was resolved with a developed digital acoustic interferometry algorithm, which compared the phase differential between signals and computed the absolute TOF with subnanosecond precision. The TOF was measured repeatedly across the tissue sample for several hours until diffusive equilibrium was realized. The change in TOF from 6-mm thick ex vivo human tonsil fit a single-exponential decay ([Formula: see text]) with rate constants that varied drastically spatially between 2 and 10 h ([Formula: see text]) due to substantial heterogeneity. This technology may prove essential to personalized cancer diagnostics by documenting and tracking biospecimen preanalytical fixation, guaranteeing their suitability for diagnostic assays, and speeding the workflow in clinical histopathology laboratories.

Collaborative analysis of multi-gigapixel imaging data using Cytomine.

MOTIVATION: Collaborative analysis of massive imaging datasets is essential to enable scientific discoveries. RESULTS: We developed Cytomine to foster active and distributed collaboration of multidisciplinary teams for large-scale image-based studies. It uses web development methodologies and machine learning in order to readily organize, explore, share and analyze (semantically and quantitatively) multi-gigapixel imaging data over the internet. We illustrate how it has been used in several biomedical applications. AVAILABILITY AND IMPLEMENTATION: Cytomine (http://www.cytomine.be/) is freely available under an open-source license from http://github.com/cytomine/ A documentation wiki (http://doc.cytomine.be) and a demo server (http://demo.cytomine.be) are also available. CONTACT: info@cytomine.be SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

A potentiator of orthosteric ligand activity at GLP-1R acts via covalent modification.

We report that 4-(3-(benzyloxy)phenyl)-2-ethylsulfinyl-6-(trifluoromethyl)pyrimidine (BETP), which behaves as a positive allosteric modulator at the glucagon-like peptide-1 receptor (GLP-1R), covalently modifies cysteines 347 and 438 in GLP-1R. C347, located in intracellular loop 3 of GLP-1R, is critical to the activity of BETP and a structurally distinct GLP-1R ago-allosteric modulator, N-(tert-butyl)-6,7-dichloro-3-(methylsulfonyl)quinoxalin-2-amine. We further show that substitution of cysteine for phenylalanine 345 in the glucagon receptor is sufficient to confer sensitivity to BETP.

Predictors of naloxone use for respiratory depression and oversedation in hospitalized adults.

PURPOSE: Results of a study to determine the established risk factors most closely associated with the use of naloxone to reverse adverse effects of opioid analgesia in a hospital population are presented. METHODS: In a retrospective case-control study at a community hospital, pharmacy dispensing records were used to identify 65 cases over a one-year period that involved the use of naloxone for the treatment of oversedation or respiratory depression and met the other inclusion criteria; another 65 patients who received opioid analgesia during the same period but did not require naloxone were identified as controls. The influence of demographics and clinical variables on the likelihood of naloxone use was analyzed by linear regression and chisquare testing. RESULTS: Patients in the naloxone group had an average of 5 risk factors for opioid-induced oversedation or respiratory depression, compared with an average of 3.3 risk factors in the control group (p < 0.001). Five factors were significantly associated with naloxone use: comorbid renal disease (odds ratio [OR], 6.034; 95% confidence interval [CI], 2.565-14.195), cardiac disease (OR, 5.829; 95% CI, 2.687-12.642), respiratory disease (OR, 3.600; 95% CI, 1.742-7.441), concurrent use of central nervous system-sedating medication (OR, 4.750; 95% CI, 1.949-11.578), and positive smoking status (OR, 4.7421; 95% CI, 2.114-9.256). CONCLUSION: Hospitalized patients on general medical units who required naloxone to reverse opioid-induced oversedation or respiratory depression had significantly more risk factors than matched patients who did not require naloxone.

Pharmacokinetics and metabolism studies on the glucagon-like peptide-1 (GLP-1)-derived metabolite GLP-1(9-36)amide in male Beagle dogs.

Glucagon-like peptide-1 (GLP-1)(7-36)amide is a 30-amino acid peptide hormone that is secreted from intestinal enteroendocrine L-cells in response to nutrients. GLP-1(7-36)amide possesses potent insulinotropic actions in the augmentation of glucose-dependent insulin secretion. GLP-1(7-36)amide is rapidly metabolized by dipeptidyl peptidase-IV to yield GLP-1(9-36)amide as the principal metabolite. Contrary to the earlier notion that peptide cleavage products of native GLP-1(7-36)amide [including GLP-1(9-36)amide] are pharmacologically inactive, recent studies have demonstrated cardioprotective and insulinomimetic effects with GLP-1(9-36)amide in mice, dogs and humans. In the present work, in vitro metabolism and pharmacokinetic properties of GLP-1(9-36)amide have been characterized in dogs, since this preclinical species has been used as an animal model to demonstrate the in vivo vasodilatory and cardioprotective effects of GLP-1(9-36)amide. A liquid chromatography tandem mass spectrometry assay was developed for the quantitation of the intact peptide in hepatocyte incubations as opposed to a previously reported enzyme-linked immunosorbent assay. Although GLP-1(9-36)amide was resistant to proteolytic cleavage in dog plasma and bovine serum albumin (t1/2>240 min), the peptide was rapidly metabolized in dog hepatocytes with a t1/2 of 110 min. Metabolite identification studies in dog hepatocytes revealed a variety of N-terminus cleavage products, most of which, have also been observed in human and mouse hepatocytes. Proteolysis at the C-terminus was not observed in GLP-1(9-36)amide. Following the administration of a single intravenous bolus dose (20 µg/kg) to male Beagle dogs, GLP-1(9-36)amide exhibited a mean plasma clearance of 15 ml/min/kg and a low steady state distribution volume of 0.05 l/kg, which translated into a short elimination half life of 0.05 h. Following subcutaneous administration of GLP-1(9-36)amide at 50 µg/kg, systemic exposure of GLP-1(9-36)amide as ascertained from maximal plasma concentrations and area under the plasma concentration-time curve from zero to infinity was 44 ng/ml and 32 ng h/ml, respectively. The subcutaneous bioavailability of GLP-1(9-36)amide in dogs was 57%. Our findings raise the possibility that the cardioprotective effects of GLP-1(9-36)amide in the conscious dog model of pacing-induced heart failure might be due, at least in part, to the actions of additional downstream metabolites, which are obtained from proteolytic cleavage of the peptide backbone in the parent compound in the liver.

Identification of a novel conformationally constrained glucagon receptor antagonist.

Identification of orally active, small molecule antagonists of the glucagon receptor represents a novel treatment paradigm for the management of type 2 diabetes mellitus. The present work discloses novel glucagon receptor antagonists, identified via conformational constraint of current existing literature antagonists. Optimization of lipophilic ligand efficiency (LLE or LipE) culminated in enantiomers (+)-trans-26 and (-)-trans-27 which exhibit good physicochemical and in vitro drug metabolism profiles. In vivo, significant pharmacokinetic differences were noted with the two enantiomers, which were primarily driven through differences in clearance rates. Enantioselective oxidation by cytochrome P450 was ruled out as a causative factor for pharmacokinetic differences.

Discovery of an intravenous hepatoselective glucokinase activator for the treatment of inpatient hyperglycemia.

Glucokinase (hexokinase IV) continues to be a compelling target for the treatment of type 2 diabetes given the wealth of supporting human genetics data and numerous reports of robust clinical glucose lowering in patients treated with small molecule allosteric activators. Recent work has demonstrated the ability of hepatoselective activators to deliver glucose lowering efficacy with minimal risk of hypoglycemia. While orally administered agents require a considerable degree of passive permeability to promote suitable exposures, there is no such restriction on intravenously delivered drugs. Therefore, minimization of membrane diffusion in the context of an intravenously agent should ensure optimal hepatic targeting and therapeutic index. This work details the identification a hepatoselective GKA exhibiting the aforementioned properties.

In vitro metabolism of the glucagon-like peptide-1 (GLP-1)-derived metabolites GLP-1(9-36)amide and GLP-1(28-36)amide in mouse and human hepatocytes.

Previous studies have revealed that the glucoincretin hormone glucagon-like peptide-1 (GLP-1)(7-36)amide is metabolized by dipeptidyl peptidase-IV (DPP-IV) and neutral endopeptidase 24.11 (NEP) to yield GLP-1(9-36)amide and GLP-1(28-36)amide, respectively, as the principal metabolites. Contrary to the previous notion that GLP-1(7-36)amide metabolites are pharmacologically inactive, recent studies have demonstrated cardioprotective and insulinomimetic effects with both GLP-1(9-36)amide and GLP-1(28-36)amide in animals and humans. In the present work, we examined the metabolic stability of the two GLP-1(7-36)amide metabolites in cryopreserved hepatocytes, which have been used to demonstrate the in vitro insulin-like effects of GLP-1(9-36)amide and GLP-1(28-36)amide on gluconeogenesis. To examine the metabolic stability of the GLP-1(7-36)amide metabolites, a liquid chromatography-tandem mass spectrometry assay was developed for the quantitation of the intact peptides in hepatocyte incubations. GLP-1(9-36)amide and GLP-1(28-36)amide were rapidly metabolized in mouse [GLP-1(9-36)amide: t(1/2) = 52 minutes; GLP-1(28-36)amide: t(1/2) = 13 minutes] and human hepatocytes [GLP-1(9-36)amide: t(1/2) = 180 minutes; GLP-1(28-36)amide: t(1/2) = 24 minutes), yielding a variety of N-terminal cleavage products that were characterized using mass spectrometry. Metabolism at the C terminus was not observed for either peptides. The DPP-IV and NEP inhibitors diprotin A and phosphoramidon, respectively, did not induce resistance in the two peptides toward proteolytic cleavage. Overall, our in vitro findings raise the intriguing possibility that the insulinomimetic effects of GLP-1(9-36)amide and GLP-1(28-36)amide on gluconeogenesis and oxidative stress might be due, at least in part, to the actions of additional downstream metabolites, which are obtained from the enzymatic cleavage of the peptide backbone in the parent compounds.

Demonstration of the innate electrophilicity of 4-(3-(benzyloxy)phenyl)-2-(ethylsulfinyl)-6-(trifluoromethyl)pyrimidine (BETP), a small-molecule positive allosteric modulator of the glucagon-like peptide-1 receptor.

4-(3-(Benzyloxy)phenyl)-2-(ethylsulfinyl)-6-(trifluoromethyl)pyrimidine (BETP) represents a novel small-molecule activator of the glucagon-like peptide-1 receptor (GLP-1R), and exhibits glucose-dependent insulin secretion in rats following i.v. (but not oral) administration. To explore the quantitative pharmacology associated with GLP-1R agonism in preclinical species, the in vivo pharmacokinetics of BETP were examined in rats after i.v. and oral dosing. Failure to detect BETP in circulation after oral administration of a 10-mg/kg dose in rats was consistent with the lack of an insulinotropic effect of orally administered BETP in this species. Likewise, systemic concentrations of BETP in the rat upon i.v. administration (1 mg/kg) were minimal (and sporadic). In vitro incubations in bovine serum albumin, plasma, and liver microsomes from rodents and humans indicated a facile degradation of BETP. Failure to detect metabolites in plasma and liver microsomal incubations in the absence of NADP was suggestive of a covalent interaction between BETP and a protein amino acid residue(s) in these matrices. Incubations of BETP with glutathione (GSH) in buffer revealed a rapid nucleophilic displacement of the ethylsulfoxide functionality by GSH to yield adduct M1, which indicated that BETP was intrinsically electrophilic. The structure of M1 was unambiguously identified by comparison of its chromatographic and mass spectral properties with an authentic standard. The GSH conjugate of BETP was also characterized in NADPH- and GSH-supplemented liver microsomes and in plasma samples from the pharmacokinetic studies. Unlike BETP, M1 was inactive as an allosteric modulator of the GLP-1R.

The design and synthesis of a potent glucagon receptor antagonist with favorable physicochemical and pharmacokinetic properties as a candidate for the treatment of type 2 diabetes mellitus.

A novel and potent small molecule glucagon receptor antagonist for the treatment of diabetes mellitus is reported. This candidate, (S)-3-[4-(1-{3,5-dimethyl-4-[4-(trifluoromethyl)-1H-pyrazol-1-yl]phenoxy}butyl)benzamido]propanoic acid, has lower molecular weight and lipophilicity than historical glucagon receptor antagonists, resulting in excellent selectivity in broad-panel screening, lower cytotoxicity, and excellent overall in vivo safety in early pre-clinical testing. Additionally, it displays low in vivo clearance and excellent oral bioavailability in both rats and dogs. In a rat glucagon challenge model, it was shown to reduce the glucagon-elicited glucose excursion in a dose-dependent manner and at a concentration consistent with its rat in vitro potency. Its properties make it an excellent candidate for further investigation.

FRET-based identification of mRNAs undergoing translation.

We present proof-of-concept in vitro results demonstrating the feasibility of using single molecule fluorescence resonance energy transfer (smFRET) measurements to distinguish, in real time, between individual ribosomes programmed with several different, short mRNAs. For these measurements we use either the FRET signal generated between two tRNAs labeled with different fluorophores bound simultaneously in adjacent sites to the ribosome (tRNA-tRNA FRET) or the FRET signal generated between a labeled tRNA bound to the ribosome and a fluorescent derivative of ribosomal protein L1 (L1-tRNA FRET). With either technique, criteria were developed to identify the mRNAs, taking into account the relative activity of the mRNAs. These criteria enabled identification of the mRNA being translated by a given ribosome to within 95% confidence intervals based on the number of identified FRET traces. To upgrade the approach for natural mRNAs or more complex mixtures, the stoichiometry of labeling should be enhanced and photobleaching reduced. The potential for porting these methods into living cells is discussed.