Discovery and optimization of novel pyridines as highly potent and selective glycogen synthase kinase 3 inhibitors.

Glycogen synthase kinase-3 plays an essential role in multiple biochemical pathways in the cell, particularly in regards to energy regulation. As such, Glycogen synthase kinase-3 is an attractive target for pharmacological intervention in a variety of disease states, particularly non-insulin dependent diabetes mellitus. However, due to homology with other crucial kinases, such as the cyclin-dependent protein kinase CDC2, developing compounds that are both potent and selective is challenging. A novel series of derivatives of 5-nitro-N2-(2-(pyridine-2ylamino)ethyl)pyridine-2,6-diamine were synthesized and have been shown to potently inhibit glycogen synthase kinase-3 (GSK3). Potency in the low nanomolar range was obtained along with remarkable selectivity. The compounds activate glycogen synthase in insulin receptor-expressing CHO-IR cells and in primary rat hepatocytes, and have acceptable pharmacokinetics and pharmacodynamics to allow for oral dosing. The X-ray co-crystal structure of human GSK3-ß in complex with compound 2 is reported and provides insights into the structural determinants of the series responsible for its potency and selectivity.

Case report of a fatal intoxication by Nucynta.

Tapentadol (Nucynta) is a centrally acting opioid analgesic prescribed for the treatment of moderate to severe acute pain. Its efficacy is believed to be due to µ-opioid receptor agonist activity and inhibition of norepinephrine reuptake resulting in increased norepinephrine concentrations. There is only one other case in the literature relating to the toxicity of this agent or report of a fatality. This case report documents a case in which tapentadol was identified as the cause of death. The tapentadol concentration found in the heart blood submitted in this case was more than 20 times the upper limit of the therapeutic range. Possible mechanisms of death include respiratory depression, central nervous system depression, and serotonin syndrome. Based on the scene investigation and autopsy findings in this case, the medical examiner determined that the cause of death was narcotic (Nucynta) intoxication and the manner of death was undetermined.

In vivo evaluation of the third generation biodegradable stent: a novel approach to avoiding the forgotten stent syndrome.

PURPOSE: Ureteral stents are prone to irritation, encrustation and infection, and they require additional procedures for removal. Furthermore, indwelling polymer stents are often forgotten with devastating consequences to the patient. We describe the degradation time, and physiological and histological responses elicited by a novel biodegradable ureteral stent in a porcine model. MATERIALS AND METHODS: A total of 16 female Yorkshire pigs were used in the study. Ten biodegradable Uriprene™ stents and 6 biostable Polaris™ stents were cystoscopically inserted unilaterally in 2 groups of animals. Excretory urogram, and blood and urine tests were performed on different days until day 28. Biostable stents were removed on day 21. On day 28 all pigs underwent necropsy for microscopic and histological evaluation. RESULTS: Nine of the 10 biodegradable stents (90%) degraded completely by 4 weeks, while 1 pig had 3 fragments smaller than 1.5 cm in the bladder. Excretory urogram showed equivalent drainage and significantly less hydronephrosis in biodegradable stented kidneys. Blood and urine parameters were similar in the 2 groups. A transient increase in serum creatinine on day 7 in 40% of the pigs with a degradable stent resolved by day 10. There were significantly fewer abnormal histological findings in the degradable stent group. We evaluated drainage characteristics in an unobstructed ureter and results may not be representative of what develops in obstructed ureters. CONCLUSIONS: The third generation biodegradable stent is a safe, effective alternative to conventional polymer stents, resulting in equivalent drainage and less hydronephrosis.

Integration of Laboratory Data into a National Electronic Health Record (EHR).

This paper discusses the development and implementation of an API to integrate external laboratory information systems with a national e-health operator using LOINC codes as a standard measurement vocabulary. The integration provides many benefits, including reduced risk of medical errors, unnecessary tests, and administrative burden on healthcare providers. Security measures were implemented to prevent unauthorized access to sensitive patient information. The "Armed eHealth" mobile application was developed to allow patients to access their lab test results directly on their mobile devices. The implementation of the universal coding system has improved communication, reduced duplications, and improved the quality of care for patients in Armenia. Overall, the integration of the universal coding system for lab tests has had a positive impact on the healthcare system in Armenia.

Effect of two counseling interventions on self-reported alcohol consumption, alcohol biomarker phosphatidylethanol (PEth), and viral suppression among persons living with HIV (PWH) with unhealthy alcohol use in Uganda: A randomized controlled trial.

PURPOSE: To test the efficacy of two interventions to reduce alcohol use and increase viral suppression compared to a control in persons with HIV (PWH). METHODS: In a three-arm (1:1:1) randomized controlled trial (N = 269), we compared in-person counselling (45-70 minutes, two sessions over three months) with interim monthly booster phone calls (live call arm) or twice-weekly automated booster sessions (technology arm) to a brief advice control arm. We enrolled PWH self-reporting unhealthy alcohol use (Alcohol Use Disorders Identification Test - Consumption, prior three months, women ≥3, men ≥4). Primary outcomes were number of self-reported drinking days (NDD) in the prior 21 and biomarker phosphatidylethanol (PEth) at six and nine months and viral suppression (<40 copies/mL) at nine months; we adjusted for sex and baseline outcomes. RESULTS: At baseline, mean 21-day NDDs were 9.4 (95 % CI: 9.1-9.8), mean PEth was 407.8 ng/mL (95 % CI: 340.7-474.8), and 89.2 % were virally suppressed. At follow-up, there were significant reductions in mean NDDs for the live call versus control arm (3.5, 95 % CI:2.1-4.9, p < 0.001) and for the technology versus control arm (3.6, 95 % CI: 2.2-5.1, p < 0.001). The mean PEth differences compared to the control arm were not significant, i.e. 36.4 ng/mL (95 % CI: -117.5 to 190.3, p = 0.643) for the live call and -30.9 ng/mL (95 % CI: -194.8 to 132.9, p = 0.711) for the technology arm. Nine-month viral suppression compared to the control was similar in the live call and in the technology arm. CONCLUSION: Intervention effects were found on self-reported NDD but not PEth or viral suppression, suggesting no treatment effect. (NCT #03928418).

Design and synthesis of 6,6-fused heterocyclic amides as raf kinase inhibitors.

Compounds belonging to several scaffolds-quinazolines, quinolines and quinoxalines-were designed and synthesized as Raf kinase inhibitors. Scaffolds were assessed for in vitro Braf(V600E) inhibition, and overall kinase selectivity. Pharmacokinetic parameters for one of the scaffolds were also determined.

Diabetic ketoacidosis: a silent death.

Diabetic ketoacidosis (DKA) results from severe insulin deficiency and can be diagnosed at autopsy despite no known history of the disease. Diabetic ketoacidosis may be the initial manifestation of type 1 diabetes or may result from increased insulin requirement in type 1 diabetic patients. The purpose of this study was to determine the percentage of DKA death investigated by the Office of Chief Medical Examiner that was not associated with a known history of diabetes.Cases investigated by the Office of Chief Medical Examiner during a 6-year period whose cause of death was DKA were identified using a centralized database. To determine the percentage with known history of diabetes, investigation reports were reviewed for any documentation of this history. The toxicology reports of all DKA deaths were reviewed together with histologic slides, if available, for possible microscopic changes. Concentrations of vitreous glucose, vitreous acetone, and blood acetone were used to diagnose DKA in these autopsied cases.Nearly a third of all death from DKA (32 of 92 during a 6-year period) occurred in individuals who had no known history of diabetes, emphasizing the importance of regular physicals that include a check of glucose concentration, and especially if any warning signs are present. In a case of sudden death, it is recommended that the volatile toxicology analysis at a medical examiner's office should include tests for acetone concentration, which when elevated, together with an elevated vitreous glucose, indicates DKA.

International Collaboration to Develop a Remote Monitoring Web App for COVID-19 Patients in Armenia: Design and Development With Agile Methodology.

BACKGROUND: COVID-19 has led to over 500 million cases and 6.2 million deaths around the world. Low- and middle-income countries (LMICs) like Armenia face unique infrastructure, financial, and capacity challenges that in many cases result in worse outcomes. Health care facilities across Armenia experienced a shortage of resources, including hospital beds and oxygen, which was further exacerbated by the war with neighboring Azerbaijan. Without a framework for home-based care, health care facilities were severely strained by COVID-19 patients who had prolonged oxygen requirements but were otherwise clinically stable. OBJECTIVE: This paper describes our approach to establishing an international collaboration to develop a web app to support home monitoring of patients with COVID-19 with persistent oxygen requirements. METHODS: The app was developed using a rapid, coordinated, and collaborative approach involving an international group of clinicians, developers, and collaborators. Health screening, monitoring, and discharge forms were developed into a lightweight OpenMRS web app and customized for the local Armenian context. RESULTS: The software was designed and developed over 2 months using human-centered design and agile sprints. Once live, 5087 patient records were created for 439 unique patients. CONCLUSIONS: This project suggests a promising framework for designing and implementing remote monitoring programs in LMICs, despite pandemic and geopolitical challenges.

Augmenting Traditional Cardiac and Medical Care in Africa via Telemedicine: A Pilot Study.

Background A critical decrease in the number of healthcare providers in developing countries is one of the major burdens to healthcare access in these countries. Many factors contribute to the lack of healthcare providers, including low doctor-to-population ratio, emigration of doctors to other countries, long travel distances to hospitals, increasing cost of healthcare, and concentration of doctors in urban cities. Several measures have been taken by both governmental and nongovernmental organizations in these countries to mitigate this crisis with varying outcomes. In this study, we investigate the use of technology in the form of telemedicine in a developing country. We evaluate patient predisposition to the use of telemedicine, their experience, and some challenges involved in the use of telemedicine in this setting. Methodology We set up an electronic medical record system, OpenMRS, and added telemedicine modules to the system. Then, we recruited doctors and gave them privileges on OpenMRS after carefully vetting their credentials. Finally, we set up a website through which patients could request telemedicine consultations. We registered a telephone number in Nigeria so that patients could also request consultations via SMS. Consult requests were then entered into OpenMRS. Doctors logged in periodically and checked for patients awaiting consults. They called patients, diagnosed them, requested further diagnostics, and/or sent prescriptions to patients via SMS directly from OpenMRS. Results Data were collected over the first year of telemedicine service in Nigeria. These data were then analyzed to understand the effectiveness, patient experience, cost efficiency, and general utilization of this service. In the first year, there were 510 new patient registrations and 572 total consultations. Patient age ranged from less than one year to 77 years, with a median of 29 years. Among the users of the service, 51.8% (264) were female. For consult requests, 52.2% of requests were via the web, and others were via SMS requests. There were over 50 reviews of the service on the website and social media, and 95% of users reported a positive experience. Conclusions From preliminary data, telemedicine can potentially be a good adjunct to help doctors reach their patients, especially in rural areas where there is an immense shortage of healthcare professions. Although most patients reported a positive experience, further investigations are needed to validate our experience.

Importance of the C-terminus of the human 5-HT3A receptor subunit.

Amongst the family members of Cys-loop LGICs, the atypical ability of the 5-HT3A subunit to form functional homomeric receptors allowed a direct investigation of the role of the C-terminus. Deletion of the three C-terminal amino acids (DeltaGln453-DeltaTyr454-DeltaAla455) from the h5-HT3A subunit prevented formation of a specific radioligand binding site as well as expression within the cell membrane. Removal of merely the C-terminal residue (DeltaAla455) reduced specific radioligand binding (to 4+/-1% relative to the wild-type; cells grown at 37 degrees C) and also cell membrane expression; these reductions were less evident when the DeltaAla455 expressing cells were grown at 27 degrees C (specific radioligand binding levels 27+/-5% relative to wild-type also grown at 27 degrees C). Mutation of the h5-HT3A C-terminal amino acid, alanine, for either glycine (Ala455Gly), valine (Ala455Val) or leucine (Ala455Leu) reduced specific radioligand binding levels by 24+/-23%, 32+/-12% and 88+/-1%, respectively; the latter mutant also displaying reduced membrane expression. In contrast, mutation to alanine of the two amino acids preceding the C-terminal alanine (Gln453Ala and Tyr454Ala) had no detrimental effects on specific radioligand binding or cell membrane expression levels. The present study demonstrates an important role for the C-terminus in the formation of the functional h5-HT3A receptor. The partial restoration of 5-HT3 receptor binding and cell membrane expression when cells expressing C-terminal mutant 5-HT3A subunits were grown at a lower temperature (27 degrees C) suggests that the C-terminus stabilises the 5-HT3 receptor allowing subunit folding and subsequent maturation.

Detection of 1-benzylpiperazine and 1-(3-trifluoromethylphenyl)-piperazine in urine analysis specimens using GC-MS and LC-ESI-MS.

Designer piperazines, such as 1-benzylpiperazine (BZP) and 1-(3-trifluoromethylphenyl)-piperazine (TFMPP), are widely available and have become popular party drugs throughout the world. Used in many countries as legal alternatives to methamphetamine and ecstasy, these designer piperazines exhibit several of the same stimulant and psychoactive properties of their illicit counterparts. Presented is a case study of seven urine analysis specimens analyzed for designer piperazines. A full scan gas chromatography-mass spectrometry screen detected the presence of BZP and TFMPP in all seven specimens. Confirmation using liquid chromatography-electrospray ionization-mass spectrometry operating in selected ion monitoring mode (SIM) yielded urinary concentrations ranging from 13.0 to 429.1 mg/L and 0.79 to 25.4 mg/L for BZP and TFMPP, respectively.

Stability of cyanide in cadavers and in postmortem stored tissue specimens: a review.

The toxicological and postmortem analysis of fire victims' blood and tissue can disclose the type and quantity of toxic species, such as carbon monoxide or hydrogen cyanide, that they inhaled prior to death. For fire cases, these toxicological data can reveal objective data about the nature and circumstances of a fire, and thus assist both the Medical Examiner and the Fire Investigator in their investigations. Assigning a level of significance to cyanide concentrations found in the blood and tissue of fire victims is often hampered by the fact that cyanide is inherently unstable in cadavers and in stored tissue samples. Numerous researchers have provided insight into and characterized the stability of cyanide in the body and in collected biological specimens. Based on studies by these researchers, the rate of transformation of cyanide in blood and tissue specimens is dependent on the initial cyanide concentration in the sample at time of death, the length of time that a sample remains in the cadaver, the length of time that a sample remains in storage, and the preservation (e.g., addition of sodium fluoride to sample) and storage conditions (e.g., temperature) of the sample.

Delta9-tetrahydrocannabinol content of commercially available hemp products.

Delta9-Tetrahydrocannabinol (THC) is the main psychoactive compound present in marijuana. THC can also be found, as a contaminant, in some commercially available hemp products marketed in health food stores and on the internet as a good source of essential fatty acids. The products range from oil to alcoholic beverages to nutritional bars to candies, with oil being the most popular and commonly available. The analytical results are separated into two groups, products tested prior to and after publication of 21 CFR Part 1308, "clarification of listing of tetrahydrocannabinols." The data presented are a summary of 79 different hemp products tested for THC. THC was separated by a liquid-liquid or solid-liquid extraction, depending upon the product matrix. THC concentrations range from none detected to 117.5 microg THC/g material. Typical limits of detection for the assay (depending on matrix) are 1.0-2.5 microg THC/g material. Products that were of aqueous base (beer, tea) had much lower limits of detection (2.5 ng/mL). No THC was detected in 58% of the products from group 1 and 86% of the products from group 2. The amounts indicate that THC levels in currently marketed hemp products are significantly lower than in those products available before 2003 and reported in previous studies. The results reported here may be used as a general guideline for the THC content of hemp products recently found in the marketplace today.

The detection of hydromorphone in urine specimens with high morphine concentrations.

A previous study suggested that small amounts of morphine are metabolically converted to hydromorphone. In the present study, morphine positive urine specimens obtained from a postmortem laboratory and a random urinalysis program were tested for morphine, codeine, hydromorphone, hydrocodone, oxymorphone, and oxycodone to assess the possibility that small amounts of hydromorphone are produced from the metabolism of morphine. The opioids were analyzed by gas chromatography-mass spectrometry as their respective trimethylsilyl derivatives following solid phase extraction. The limit of detection for hydromorphone was 5 ng/mL. A total of 73 morphine positive urine specimens were analyzed, with morphine concentrations ranging from 131 to 297,000 ng/mL. Hydromorphone was present at a concentration > or =5 ng/mL in 36 of these specimens at concentrations ranging from 0.02% to 12% of the morphine concentration. Hydrocodone was not detected in these specimens at the assay detection limit of 25 ng/mL. These results support earlier work suggesting that the detection of hydromorphone in urine specimens does not necessarily mean that exogenous hydromorphone or hydrocodone was used.

Distribution of phencyclidine into vitreous humor.

Vitreous humor is a fluid contained in the eye that is largely composed of water. The advantages of vitreous humor as a specimen for postmortem drug analysis include its relatively low susceptibility to contamination and the ability to analyze vitreous humor with little or no pretreatment. The postmortem analysis of ethanol in vitreous humor has been well established. However, studies of drug disposition into vitreous humor are limited. Heart blood, subclavian blood, and vitreous humor specimens from 26 phencyclidine-positive postmortem cases were analyzed to evaluate the distribution of phencyclidine into vitreous humor. Phencyclidine intoxication was not the cause of death in any of the cases analyzed. Specimens were analyzed by solid-phase extraction followed by gas chromatography-mass spectrometry. All positive blood specimens were associated with a positive vitreous humor specimen. On average, the blood phencyclidine concentrations were greater than the vitreous humor phencyclidine concentrations, with average blood/vitreous ratios of 2.85 for heart blood and 2.51 for subclavian blood. However, there was considerable variability between cases, which indicates that although vitreous humor is an appropriate specimen for the detection of phencyclidine in postmortem cases, its interpretative value is limited.

A fatality from sevoflurane abuse.

A case is presented of a 47-year-old man who died as a result of sevoflurane abuse. Sevoflurane was identified and confirmed by headspace gas chromatography-mass spectrometry. The heart blood sevoflurane concentration was 16 mg/L, and the peripheral blood sevoflurane concentration was 8.0 mg/L. No drugs or other volatile substances were found in the heart blood. The medical examiner ruled that the cause of death was cardiac arrhythmia due to sevoflurane toxicity. Cardiomegaly was listed on Part II of the death certificate. The manner of death was undetermined.

Synthesis, Binding Mode, and Antihyperglycemic Activity of Potent and Selective (5-Imidazol-2-yl-4-phenylpyrimidin-2-yl)[2-(2-pyridylamino)ethyl]amine Inhibitors of Glycogen Synthase Kinase 3.

In an effort to identify new antidiabetic agents, we have discovered a novel family of (5-imidazol-2-yl-4-phenylpyrimidin-2-yl)[2-(2-pyridylamino)ethyl]amine analogues which are inhibitors of human glycogen synthase kinase 3 (GSK3). We developed efficient synthetic routes to explore a wide variety of substitution patterns and convergently access a diverse array of analogues. Compound 1 (CHIR-911, CT-99021, or CHIR-73911) emerged from an exploration of heterocycles at the C-5 position, phenyl groups at C-4, and a variety of differently substituted linker and aminopyridine moieties attached at the C-2 position. These compounds exhibited GSK3 IC50s in the low nanomolar range and excellent selectivity. They activate glycogen synthase in insulin receptor-expressing CHO-IR cells and primary rat hepatocytes. Evaluation of lead compounds 1 and 2 (CHIR-611 or CT-98014) in rodent models of type 2 diabetes revealed that single oral doses lowered hyperglycemia within 60 min, enhanced insulin-stimulated glucose transport, and improved glucose disposal without increasing insulin levels.

Tizanidine distribution in a postmortem case.

The case of a 57-year-old white female who was found deceased at home by her husband is presented. A suicide note was found at the scene. No remarkable findings were observed at autopsy. Comprehensive toxicological analysis of the heart blood identified ethanol (0.16 g/dL), diazepam (1.1 mg/L), and tizanidine (2.3 mg/L). Blood concentrations of tizanidine following therapeutic use do not exceed 0.025 mg/L. The medical examiner ruled that the cause of death was combined ethanol and multiple drug intoxication, and the manner of death was suicide.

Two cases of fatal amlodipine overdose.

Two fatal overdoses of the calcium channel blocker amlodipine are described. Postmortem samples were screened for volatiles and therapeutic and abused drugs. Amlodipine was measured by liquid chromatography-atmospheric pressure photoionization-mass spectrometry. The heart blood amlodipine concentrations for the two cases were 2.4 and 0.95 mg/L, and amlodipine was quantified in all other tissues. In the first case, venlafaxine and norvenlafaxine were also found, and the angiotensin receptor antagonist olmesartan was tentatively identified. The concentrations of amlodipine are compared with previously reported fatal and nonfatal overdoses. The medical examiners ruled in both cases that the manner of death was suicide and the causes of death were mixed drug intoxication and amlodipine intoxication.

A comprehensive study on the variations in urinary concentrations of endogenous gamma-hydroxybutyrate (GHB).

This study was designed to supplement previous attempts to establish an accurate range of normal endogenous gamma-hydroxybutyrate (GHB) concentrations in random antemortem urine samples. Furthermore, its purpose was to ascertain the effect of gender, race, age, medications, and select medical conditions on endogenous concentrations of GHB in urine and the proposed endogenous urinary GHB cutoff of 10 microg/mL. Urine samples (n = 207) were provided by subjects who reported that they had never used GHB. As part of the collection process, subjects also completed a short survey to collect information about gender, race, age, orally ingested medications, and select medical conditions. All specimens were analyzed in duplicate for the presence of endogenous GHB using a previously reported headspace gas chromatography-mass spectrometry method. The data were analyzed for tendencies among different population groups. GHB concentrations ranged from 0.00 to 2.70 microg/mL in all specimens, with a median concentration of 0.24 microg/mL. Males (n = 130) had an average endogenous GHB concentration of 0.27 microg/mL (0.00-2.70 microg/mL), whereas females (n = 77) averaged 0.29 microg/mL (0.00-0.98 microg/mL). Select medical conditions and participants' race, age ranges, and medications that were used within 48 h prior to collection were also evaluated. We believe this to be the most comprehensive study on endogenous GHB concentrations in urine to date. The results of this study will aid the interpretation of low GHB concentrations measured in urine samples, particularly in investigations of drug-facilitated crimes.