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Amphotericin B (AmpB) is an effective but toxic antifungal drug. Thus, improving its activity/toxicity relationship is of interest. AmpB disrupts fungal membranes by two proposed mechanisms: ergosterol sequestration from the membrane and pore formation. Whether these two mechanisms operate in conjunction and how they could be potentiated remains to be fully understood. Here, we report that gladiolin, a polyketide antibiotic produced by Burkholderia gladioli , is a strong potentiator of AmpB and acts synergistically against Cryptococcus and Candida species, including drug-resistant C. auris . Gladiolin also synergizes with AmpB against drug-resistant fungal biofilms, while exerting no mammalian cytotoxicity. To explain the mechanism of synergy, we show that gladiolin interacts with membranes via a previously unreported binding mode for polyketides. Moreover, gladiolin modulates lipid binding by AmpB and, in combination, causes faster and more pronounced lipid rearrangements relative to AmpB alone which include membrane thinning consistent with ergosterol extraction, areas of thickening, pore formation, and increased membrane destruction. These biophysical data provide evidence of a functional interaction between gladiolin and AmpB at the membrane interface. The data further indicate that the two proposed AmpB mechanisms (ergosterol sequestration and pore formation) act in conjunction to disrupt membranes, and that gladiolin synergizes by enhancing both mechanisms. Collectively, our findings shed light on AmpB's mechanism of action and characterize gladiolin as an AmpB potentiator, showing an antifungal mechanism distinct from its proposed antibiotic activity. We shed light on the synergistic mechanism at the membrane, and provide insights into potentiation strategies to improve AmpB's activity/toxicity relationship., Importance: Amphotericin B (AmpB) is one of the oldest antifungal drugs in clinical use. It is an effective therapeutic, but it comes with toxicity issues due to the similarities between its fungal target (the membrane lipid ergosterol) and its mammalian counterpart (cholesterol). One strategy to improve its activity/toxicity relationship is by combinatorial therapy with potentiators, which would enable a lower therapeutic dose of AmpB. Here, we report on the discovery of the antibiotic gladiolin as a potentiator of AmpB against several priority human fungal pathogens and fungal biofilms, with no increased toxicity against mammalian cells. We show that gladiolin potentiates AmpB by increasing and accelerating membrane damage. Our findings also provide insights into the on-going debate about the mechanism of action of AmpB by indicating that both proposed mechanisms, extraction of ergosterol from membranes and pore formation, are potentiated by gladiolin., Competing Interests: G.L.C. is a shareholder, non-executive director, and consultant of Erebagen Ltd. The other authors declare no competing interests.

Lenacapavir (LEN), a long-acting injectable, is the first approved human immunodeficiency virus type 1 capsid inhibitor and one of a few Food and Drug Administration-approved drugs that exhibit atropisomerism. LEN exists as a mixture of two class 2 atropisomers that interconvert at a fast rate (half-life < 2 hours) with a ratio that is stable over time and unaffected by enzymes or binding to proteins in plasma. LEN exhibits low systemic clearance (CL) in nonclinical species and humans; however, in all species, the observed CL was higher than the in vitro predicted CL. The volume of distribution was moderate in nonclinical species and consistent with the tissue distribution observed by whole-body autoradiography in rats. LEN does not distribute to brain, consistent with being a P-glycoprotein (P-gp) substrate. Mechanistic drug disposition studies with [ 14 C]LEN in intravenously dosed bile duct-cannulated rats and dogs showed a substantial amount of unchanged LEN (31%-60% of dose) excreted in feces, indicating that intestinal excretion (IE) was a major clearance pathway for LEN in both species. Coadministration of oral elacridar, a P-gp inhibitor, in rats decreased CL and IE of LEN. Renal excretion was < 1% of dose in both species. In plasma, almost all radioactivity was unchanged LEN. Low levels of metabolites in excreta included LEN conjugates with glutathione, pentose, and glucuronic acid, which were consistent with metabolites formed in vitro in H μ rel hepatocyte cocultures and those observed in human. Our studies highlight the importance of IE for efflux substrates that are highly metabolically stable compounds with slow elimination rates. SIGNIFICANCE STATEMENT: LEN is a long-acting injectable that exists as conformationally stable atropisomers. Due to an atropisomeric interconversion rate that significantly exceeds the in vivo elimination rate, the atropisomer ratio of LEN remains constant in circulation. The disposition of LEN highlights that intestinal excretion has a substantial part in the elimination of compounds that are metabolically highly stable and efflux transporter substrates., (Copyright © 2024 by The Author(s).)

Little is known about the methods and outcomes of patient-reported outcome measure (PROM) use among high-risk medical device registries. The objective of this scoping review was to assess the utility and predictive ability of PROMs in high-risk medical device registries. We searched Ovid Medline, Embase, APA PsychINFO, Cochrane Library, and Scopus databases for published literature. After searching, 4323 titles and abstracts were screened, and 262 full texts were assessed for their eligibility. Seventy-six papers from across orthopedic ( n = 64), cardiac ( n = 10), penile ( n = 1), and hernia mesh ( n = 1) device registries were identified. Studies predominantly used PROMs as an outcome measure when comparing cohorts or surgical approaches ( n = 45) or to compare time points ( n = 13) including pre- and postintervention. Fifteen papers considered the predictive ability of PROMs. Of these, 8 treated PROMs as an outcome, 5 treated PROMs as a risk factor through regression analysis, and 2 papers treated PROMs as both a risk factor and as an outcome. One paper described PROMs to study implant survival. To advance methods of PROM integration into clinical decision-making for medical devices, an understanding of their use in high-risk device registries is needed. This scoping review found that there is a paucity of studies using PROMs to predict long-term patient and clinical outcomes in high-risk medical device registries. Determination as to why PROMs are rarely used for predictive purposes in long-term data collection is needed if PROM data are to be considered suitable as real-world evidence for high-risk device regulatory purposes, as well as to support clinical decision-making., (© The Author(s) 2024. Published by Oxford University Press on behalf of The Aesthetic Society.)

Objectives: This study aims to investigate the cost incurred by people travelling to the neurology outpatient clinic of a large metropolitan hospital. As outpatients are a substantial portion of a hospital's demographic, we aimed to understand the patient experience of various commuters., Methods: We conducted an observational study collecting demographic details and travel information for how people attended the neurology clinic of Monash Medical Centre. Statistical analysis was performed using R. 165 participants were randomly selected and interviewed in-person. Data were collected via an anonymous questionnaire. The study was approved by the Monash Health Human Ethics Research Committee., Results: 155 responses were included in the analysis. Patients paid an average of $A16.64 to travel to Monash Medical Centre. Drivers paid on average $A16.70 and those taking public transport paid on average $A9.64, with the maximum cost overall being $A120.00. For patients driving to hospital, parking accounted for 60% of their travel costs. The average to Monash Medical centre was 20.82 km with the maximum being 190.88 km. Distance from hospital was correlated with a higher cost of travel (p<0.001, Spearman's rank correlation coefficient=0.48). There was also an inverse association between distance from hospital and socioeconomic status (p<0.001, Spearman's rank correlation coefficient=-0.26)., Conclusion: Travelling to hospital can be a costly endeavour. Driving is the most popular form of transport, but a large portion of the cost involved is hospital parking. Further research should be conducted at other tertiary centres with larger samples., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2024. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Purpose: Health care is a major contributor to climate change, and critical care is one of the sector's highest carbon emitters. Health economic evaluations form an important component of critical care and may be useful in identifying economically efficient and environmentally sustainable strategies. The purpose of this scoping review was to synthesise available literature on whether and how environmental impact is considered in health economic evaluations of critical care., Methods: A robust scoping review methodology was used to identify studies reporting on environmental impact in health economic evaluations of critical care. We searched six academic databases to locate health economic evaluations, costing studies and life cycle assessments of critical care from 1993 to present., Results: Four studies met the review's inclusion criteria. Of the 278 health economic evaluations of critical care identified, none incorporated environmental impact into their assessments. Most included studies (n = 3/4) were life cycle assessments, and the remaining study was a prospective observational study. Life cycle assessments used a combination of process-based data collection and modelling to incorporate environmental impact into their economic assessments., Conclusions: Health economic evaluations of critical care have not yet incorporated environmental impact into their assessments, and few life cycle assessments exist that are specific to critical care therapies and treatments. Guidelines and standardisation regarding environmental data collection and reporting in health care are needed to support further research in the field. In the meantime, those planning health economic evaluations should include a process-based life cycle assessment to establish key environmental impacts specific to critical care., (© 2024. The Author(s).)

Lenacapavir (LEN) is a picomolar first-in-class capsid inhibitor of human immunodeficiency virus type 1 (HIV-1) with a multistage mechanism of action and no known cross resistance to other existing antiretroviral (ARV) drug classes. LEN exhibits a low aqueous solubility and exceptionally low systemic clearance following intravenous (IV) administration in nonclinical species and humans. LEN formulated in an aqueous suspension or a PEG/water solution formulation showed sustained plasma exposure levels with no unintended rapid drug release following subcutaneous (SC) administration to rats and dogs. A high total fraction dose release was observed with both formulations. The long-acting pharmacokinetics (PK) were recapitulated in humans following SC administration of both formulations. The SC PK profiles displayed two-phase absorption kinetics in both animals and humans with an initial fast-release absorption phase, followed by a slow-release absorption phase. Noncompartmental and compartmental analyses informed the LEN systemic input rate from the SC depot and exit rate from the body. Modeling-enabled deconvolution of the input rates from two processes: absorption of the soluble fraction (minor) from a direct fast-release process leading to the early PK phase and absorption of the precipitated fraction (major) from an indirect slow-release process leading to the later PK phase. LEN SC PK showed flip-flop kinetics due to the input rate being substantially slower than the systemic exit rate. LEN input rates via the slow-release process in humans were slower than those in both rats and dogs. Overall, the combination of high potency, exceptional stability, and optimal release rate from the injection depot make LEN well suited for a parenteral long-acting formulation that can be administered once up to every 6 months in humans for the prevention and treatment of HIV-1.

Objective: We describe COVID-19 risk reduction strategies adopted by Victorian adults during December 2021-January 2022, a period of high COVID-19 infection and limited government mandated public health measures., Methods: In February 2022, participants of a Victorian-based cohort study (Optimise) completed a cross-sectional survey on risk reduction behaviours during December 2021-January 2022. Regression modelling estimated the association between risk reduction and demographics., Results: A total of 556 participants were included (median age 47 years; 75% women; 82% in metropolitan Melbourne). Two-thirds (61%) adopted at least one risk reduction behaviour, with uptake highest among younger participants (18-34 years; adjusted relative risk (aRR): 1.20, 95% confidence interval [CI]: 1.01, 1.41) and those with a chronic health condition (aRR: 1.17, 95% CI: 1.02, 1.35)., Conclusions: Participants adopted their own COVID-19 risk reduction strategies in a setting of limited government restrictions, with young people more likely to adopt a risk reduction strategy that did not limit social mobility., Implication for Public Health: A public health response to COVID-19 that focusses on promoting personal risk reduction behaviours, as opposed to mandated restrictions, could be enhanced by disseminating information on and increasing availability of effective risk reduction strategies tailored to segments of the population., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Since the emergence of the Omicron variant in Australia in late 2021 there have been over 5.47 million cases and 4993 deaths, disproportionately impacting on people with social and structural disadvantage. However there has been increasing reluctance by governments to intervene to reduce the impact of COVID-19 transmission and its consequences. This commentary article provides a perspective on the support and guidance required to mitigate individuals and communities risk by using a harm reduction approach to highlight strategies that can reduce COVID-19 transmission and infection., Competing Interests: Conflicts of interest The authors have no competing interests to declare., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Aims: Unsupervised injectable opioid agonist therapy (iOAT) may decrease the unmet treatment needs for people who inject opioids. We aimed to model whether unsupervised iOAT may be effective in reducing fatal and non-fatal overdose, and estimate the cost per life saved., Methods: The study used a decision tree model based on Australian and international parameters for overdose risk in people who inject opioids who are: not on OAT; new/stable to methadone/buprenorphine treatment; on iOAT; or on unsupervised iOAT. We modeled scenarios of (1) current OAT only (status quo), or current OAT plus either (2) 5% supervised iOAT, (3) 5% supervised or 5.69% unsupervised iOAT (based on willingness to enroll), OR (4) 1.2% supervised and 10% unsupervised iOAT (the same cost as scenario 2). The study measured overdoses (fatal and nonfatal) and treatment costs per 10,000 people who inject opioids per annum, and cost-per deaths averted on implementation of iOAT., Results: With current OAT, the study found an estimated 1655.5 (1552.7-1705.3) overdoses, 19.3 (17.9-20.3) overdose deaths and AUD 23,335,081 in treatment costs per 10,000 people per annum. Implementation of 5% enrollment in supervised iOAT costs an additional AUD 14,807,855 and showed a reduction of 122.9 (95% UI 114.2-130.5) overdoses and 2.0 (1.8-2.0) overdose deaths per 10,000 people per annum ($7,774,172 [7,283,182-8,146,989] per death averted). For the same treatment costs, additional coverage of 10% unsupervised iOAT and 1.2% supervised iOAT could be achieved, which the study estimated to prevent 269.0 (95% UI 250.0-278.7) overdoses and 4.0 (3.7-4.2) overdose deaths per 10,000 people per annum ($3,723,340 (3,385,878-3,894,379) per death averted), alongside further benefits of treatment unaccounted for in this study., Conclusion: An implementation scenario with greater unsupervised iOAT compared to supervised iOAT allows for an increased reduction in overdose and overdose deaths per annum at the same cost, with the additional benefit of increased treatment coverage among people who inject opioids., Competing Interests: Declaration of competing interest SN and PD have received funding from Indivior. SN has received research funding from Seqirus, and PD has received an investigator-driven grant from Gilead Sciences for unrelated work on Hepatitis C and an untied educational grant from Reckitt Benckiser for unrelated work on the introduction of buprenorphine-naloxone into Australia. PD and SN and have served as unpaid members of an Advisory Board for an intranasal naloxone product., (Copyright © 2022 Elsevier Inc. All rights reserved.)

Background: There are ongoing concerns regarding pharmaceutical opioid-related harms, including overdose and dependence, with an associated increase in treatment demand. People dependent on pharmaceutical opioids appear to differ in important ways from people who use heroin, yet most opioid agonist treatment research has been conducted in people who use heroin.  OBJECTIVES: To assess the effects of maintenance opioid agonist pharmacotherapy for the treatment of pharmaceutical opioid dependence., Search Methods: We updated our searches of the following databases to January 2022: the Cochrane Drugs and Alcohol Group Specialised Register, CENTRAL, MEDLINE, four other databases, and two trial registers. We checked the reference lists of included studies for further references to relevant randomised controlled trials (RCTs)., Selection Criteria: We included RCTs with adults and adolescents examining maintenance opioid agonist treatments that made the following two comparisons. 1. Full opioid agonists (methadone, morphine, oxycodone, levo-alpha-acetylmethadol (LAAM), or codeine) versus different full opioid agonists or partial opioid agonists (buprenorphine) for maintenance treatment. 2. Full or partial opioid agonist maintenance versus non-opioid agonist treatments (detoxification, opioid antagonist, or psychological treatment without opioid agonist treatment)., Data Collection and Analysis: We used standard Cochrane methods., Main Results: We identified eight RCTs that met inclusion criteria (709 participants). We found four studies that compared methadone and buprenorphine maintenance treatment, and four studies that compared buprenorphine maintenance to either buprenorphine taper (in addition to psychological treatment) or a non-opioid maintenance treatment comparison. We found low-certainty evidence from three studies of a difference between methadone and buprenorphine in favour of methadone on self-reported opioid use at end of treatment (risk ratio (RR) 0.49, 95% confidence interval (CI) 0.28 to 0.86; 165 participants), and low-certainty evidence from four studies finding a difference in favour of methadone for retention in treatment (RR 1.21, 95% CI 1.02 to 1.43; 379 participants). We found low-certainty evidence from three studies showing no difference between methadone and buprenorphine on substance use measured with urine drug screens at end of treatment (RR 0.81, 95% CI 0.57 to 1.17; 206 participants), and moderate-certainty evidence from one study of no difference in days of self-reported opioid use (mean difference 1.41 days, 95% CI 3.37 lower to 0.55 days higher; 129 participants). There was low-certainty evidence from three studies of no difference between methadone and buprenorphine on adverse events (RR 1.13, 95% CI 0.66 to 1.93; 206 participants). We found low-certainty evidence from four studies favouring maintenance buprenorphine treatment over non-opioid treatments in terms of fewer opioid positive urine drug tests at end of treatment (RR 0.66, 95% CI 0.52 to 0.84; 270 participants), and very low-certainty evidence from four studies finding no difference on self-reported opioid use in the past 30 days at end of treatment (RR 0.63, 95% CI 0.39 to 1.01; 276 participants). There was low-certainty evidence from three studies of no difference in the number of days of unsanctioned opioid use (standardised mean difference (SMD) -0.19, 95% CI -0.47 to 0.09; 205 participants). There was moderate-certainty evidence from four studies favouring buprenorphine maintenance over non-opioid treatments on retention in treatment (RR 3.02, 95% CI 1.73 to 5.27; 333 participants). There was moderate-certainty evidence from three studies of no difference in adverse effects between buprenorphine maintenance and non-opioid treatments (RR 0.50, 95% CI 0.07 to 3.48; 252 participants). The main weaknesses in the quality of the data was the use of open-label study designs, and difference in follow-up rates between treatment arms., Authors' Conclusions: There is  very low- to moderate-certainty evidence supporting the use of maintenance agonist pharmacotherapy for pharmaceutical opioid dependence. Methadone or buprenorphine did not differ on some outcomes, although on the outcomes of retention and self-reported substance use some results favoured methadone. Maintenance treatment with buprenorphine appears more effective than non-opioid treatments. Due to the overall very low- to moderate-certainty evidence and small sample sizes, there is the possibility that the further research may change these findings., (Copyright © 2022 The Cochrane Collaboration. Published by John Wiley & Sons, Ltd.)

Objectives: To identify and map all trials in maternal health conducted in low and middle-income countries (LMIC) over the 10-year period from 2010 to 2019, to identify geographical and thematic trends, as well as comparing to global causes of maternal death and preidentified priority areas., Design: Systematic scoping review., Primary and Secondary Outcome Measures: Extracted data included location, study characteristics and whether trials corresponded to causes of mortality and identified research priority topics., Results: We searched the Cochrane Central Register of Controlled Trials database, a combined registry of trials from multiple sources. Our search identified 7269 articles, 874 of which were included for analysis. Between 2010 and 2019, maternal health trials conducted in LMICs more than doubled (50-114). Trials were conducted in 61 countries-231 trials (26.4%) were conducted in Iran. Only 225 trials (25.7%) were aligned with a cause of maternal mortality. Within these trials, pre-existing medical conditions, embolism, obstructed labour and sepsis were all under-represented when compared with number of maternal deaths globally. Large numbers of studies were conducted on priority topics such as labour and delivery, obstetric haemorrhage and antenatal care. Hypertensive disorders of pregnancy, diabetes and health systems and policy-despite being high-priority topics-had relatively few trials., Conclusion: Despite trials conducted in LMICs increasing from 2010 to 2019, there were significant gaps in geographical distribution, alignment with causes of maternal mortality and known research priority topics. The research gaps identified provide guidance and insight for future research conduct in low-resource settings., Trial Registration Number: 10.17605/OSF.IO/QUJP5., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Take home naloxone (THN) programs have been rapidly upscaled in response to increasing opioid-related mortality. One often cited concern is that naloxone provision could be associated with increased opioid use, due to the availability of naloxone to reverse opioid overdose. We conducted a systematic review to determine whether THN provision is associated with changes in substance use by participants enrolled in THN programs., Methods: We conducted a systematic review of the literature to assess changes in heroin or other substance use by people who use opioids following THN provision., Results: Seven studies with 2578 participants were included. Of the seven studies, there were two quasi-experimental studies and five cohort studies. Based on the Joanna Briggs Institute quality assessment, four studies were of moderate quality and three studies were of high quality. Of the five studies that reported on the primary outcome of heroin use, no study found evidence of increased heroin use across the study population. Five studies reported on other substance use (benzodiazepines, alcohol, cocaine, amphetamine, cannabis, prescription opioids), none of which found evidence of an increase in other substance use associated with THN provision. Four studies reported on changes in overdose frequency following THN provision: three studies reporting no change, and one study of people prescribed opioids finding a reduction in opioid-related emergency department attendances for participants who received naloxone., Conclusion: We found no evidence that THN provision was associated with increased opioid use or overdose. Concerns that THN supply may lead to increased substance use were not supported by data from reviewed studies., Competing Interests: Declarations of Interest In the past 5 years, SN and SL have been investigators on untied education grants from Indivior, unrelated to the current work. SN has provided training to health care professionals on identifying and treating codeine dependence for which her institution has received payment from Indivior. SN and TL have been investigators on untied educational grants from Seqirus, unrelated to the current work. PD has received an investigator-driven grant from Gilead Sciences for unrelated work on Hepatitis C and an untied educational grant from Reckitt Benckiser for unrelated work on the introduction of buprenorphine-naloxone into Australia. PD and SN have served as unpaid members of an Advisory Board for an intranasal naloxone product., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Objective: Fixed ratio blood product administration may improve outcomes in trauma patients with massive blood loss. The present study aimed to describe the impact of a major haemorrhage protocol (MHP) on the ratio of blood products administered for paediatric major trauma., Methods: Retrospective observational study in a state-designated paediatric major trauma centre in Melbourne, Australia. Children with major trauma who received blood products in the ED were identified from a hospital trauma registry. Blood product ratios before, during and after implementation of a hospital MHP were compared in consecutive 2 year blocks., Results: Over a 6 year period, 767 major trauma patients were identified, of whom 47 received blood products in the ED and were included in the analysis; 14 pre-MHP implementation, 24 during-MHP implementation and nine post-MHP implementation. No patients received blood products at a ratio of 1:1:1 for red blood cells:fresh frozen plasma:platelets, respectively, during any time period. In this cohort of predominantly blunt trauma, blood products were infrequently administered in the ED because of the low prevalence of massive blood loss. Coagulopathy and hypofibrinogenaemia were commonly observed, nearly half of included patients were managed operatively and one quarter did not survive their injuries., Conclusion: The implementation of a MHP did not change the ratio of blood product administration in this cohort of patients because of the infrequency of massive blood loss. Future studies may focus on the impact of treating coagulopathy and hypofibrinogenaemia on patient-centred outcomes., (© 2021 Australasian College for Emergency Medicine.)

Background: Antithrombotic medications (antiplatelets and anticoagulants) reduce the risk of cardiovascular disease (CVD), but with the disadvantage of increasing bleeding risk. Ethnicity and socioeconomic deprivation are independent predictors of major bleeds among patients without CVD, but it is unclear whether they are also predictors of major bleeds among patients with CVD or atrial fibrillation (AF) after adjustment for clinical variables., Methods: Prospective cohort study of 488,107 people in New Zealand Primary Care (including 64,420 Māori, the indigenous people of New Zealand) aged 30-79 years who had their CVD risk assessed between 2007 and 2016. Participants were divided into three mutually exclusive subgroups: (1) AF with or without CVD (n = 15,212), (2) CVD and no AF (n = 43,790), (3) no CVD or AF (n = 429,105). Adjusted hazards ratios (adjHRs) were estimated from Cox proportional hazards models predicting major bleeding risk for each of the three subgroups to determine whether ethnicity and socioeconomic deprivation are independent predictors of major bleeds in different cardiovascular risk groups., Results: In all three subgroups (AF, CVD, no CVD/AF), Māori (adjHR 1.63 [1.39-1.91], 1.24 [1.09-1.42], 1.57 [95% CI 1.45-1.70], respectively), Pacific people (adjHR 1.90 [1.58-2.28], 1.30 [1.12-1.51], 1.62 [95% CI 1.49-1.75], respectively) and Chinese people (adjHR 1.53 [1.08-2.16], 1.15 [0.90-1.47], 1.13 [95% CI 1.01-1.26], respectively) were at increased risk of a major bleed compared to Europeans, although for Chinese people the effect did not reach statistical significance in the CVD subgroup. Compared to Europeans, Māori and Pacific peoples were generally at increased risk of all bleed types (gastrointestinal, intracranial and other bleeds). An increased risk of intracranial bleeds was observed among Chinese and Other Asian people and, in the CVD and no CVD/AF subgroups, among Indian people. Increasing socioeconomic deprivation was also associated with increased risk of a major bleed in all three subgroups (adjHR 1.07 [1.02-1.12], 1.07 [1.03-1.10], 1.10 [95% CI 1.08-1.12], respectively, for each increase in socioeconomic deprivation quintile)., Conclusion: Ethnicity and socioeconomic status should be considered in bleeding risk assessments to guide the use of antithrombotic medication for the management of AF and CVD.

Aim: To evaluate the frequency and predictors of poor outcome in febrile children presenting to the Emergency Department., Methods: Retrospective observational study from the Emergency Department of The Royal Children's Hospital, Melbourne, Australia. All children with presenting complaint of fever or triage temperature >38°C over a 6-month period were included. Poor outcome was defined as: new organ dysfunction or the requirement for organ support therapy (inotrope infusion, mechanical ventilation, renal replacement therapy and extra-corporeal life support). Predictors evaluated were as follows: initial vital signs, blood tests and clinical scores. Odds ratio, sensitivity, specificity and area under the receiver-operating characteristics curve were calculated for each predictor variable., Results: Between Jan-June 2019, 6217 children met inclusion criteria. Twenty-seven (0.4%) developed new organ dysfunction, 10 (0.2%) required organ support therapy (inotrope infusion in 5, mechanical ventilation in 6, renal replacement therapy in 1, extra-corporeal life support in 1). Odds of new organ dysfunction, requirement for inotropic support and mechanical ventilation were higher with abnormal initial vital signs, blood tests and clinical scores, though overall test characteristics were poor due to infrequency., Conclusion: Poor outcomes were uncommon among febrile children presenting to the Emergency Department. Vital signs, blood tests and clinical scores were poor predictors., (©2020 Foundation Acta Paediatrica. Published by John Wiley & Sons Ltd.)

Oral antiretroviral agents provide life-saving treatments for millions of people living with HIV, and can prevent new infections via pre-exposure prophylaxis 1-5 . However, some people living with HIV who are heavily treatment-experienced have limited or no treatment options, owing to multidrug resistance 6 . In addition, suboptimal adherence to oral daily regimens can negatively affect the outcome of treatment-which contributes to virologic failure, resistance generation and viral transmission-as well as of pre-exposure prophylaxis, leading to new infections 1,2,4,7-9 . Long-acting agents from new antiretroviral classes can provide much-needed treatment options for people living with HIV who are heavily treatment-experienced, and additionally can improve adherence 10 . Here we describe GS-6207, a small molecule that disrupts the functions of HIV capsid protein and is amenable to long-acting therapy owing to its high potency, low in vivo systemic clearance and slow release kinetics from the subcutaneous injection site. Drawing on X-ray crystallographic information, we designed GS-6207 to bind tightly at a conserved interface between capsid protein monomers, where it interferes with capsid-protein-mediated interactions between proteins that are essential for multiple phases of the viral replication cycle. GS-6207 exhibits antiviral activity at picomolar concentrations against all subtypes of HIV-1 that we tested, and shows high synergy and no cross-resistance with approved antiretroviral drugs. In phase-1 clinical studies, monotherapy with a single subcutaneous dose of GS-6207 (450 mg) resulted in a mean log 10 -transformed reduction of plasma viral load of 2.2 after 9 days, and showed sustained plasma exposure at antivirally active concentrations for more than 6 months. These results provide clinical validation for therapies that target the functions of HIV capsid protein, and demonstrate the potential of GS-6207 as a long-acting agent to treat or prevent infection with HIV.

Objectives: To characterise the community pharmacy supply of naloxone by supply type - individual prescription, prescriber bag, and non-dispensed (supplied over the counter or expired) - during 2014-2018; to examine whether the 2016 rescheduling of naloxone as an over-the-counter drug influenced non-dispensed naloxone supply volume., Design, Setting: Analysis of monthly naloxone prescriptions (Pharmaceutical Benefits Scheme) and sales data (IQVIA), 2014-2018, for Australia and by state and territory; time series analysis of non-dispensed naloxone supply to assess effect of rescheduling on naloxone supply., Major Outcomes: Total naloxone supply to community pharmacies; prescribed and non-dispensed naloxone supply., Results: During 2014-2018, 372 351 400 μg units of naloxone were sold to community pharmacies: non-dispensed naloxone accounted for 205 866.5 units (55.3%), prescriber bags for 155 841 units (41.8%), and individual prescriptions for 10 643.5 units (2.9%). Population-adjusted national naloxone sales to community pharmacies increased between 2014 and 2018 (per year: incidence rate ratio [IRR], 1.15; 95% CI, 1.09-2.22). This increase was primarily attributable to increased volumes of prescriber bag naloxone (IRR, 1.63; 95% CI, 1.50-1.78) and, to a lesser extent, increased individual prescription supply (IRR, 2.04; 95% CI, 1.85-2.26). Non-dispensed naloxone supply volume was unchanged at the national level (IRR, 0.93; 95% CI, 0.85-1.01); changes in non-dispensed supply immediately following rescheduling and subsequently were not statistically significant in time series analyses for most jurisdictions., Conclusions: Total naloxone supply to community pharmacies in Australia increased between 2014 and 2018, but rescheduling that enabled over-the-counter access did not significantly influence the volume of non-dispensed naloxone., (© 2020 AMPCo Pty Ltd.)

People living with HIV (PLWH) have expressed concern about the life-long burden and stigma associated with taking pills daily and can experience medication fatigue that might lead to suboptimal treatment adherence and the emergence of drug-resistant viral variants, thereby limiting future treatment options 1-3 . As such, there is strong interest in long-acting antiretroviral (ARV) agents that can be administered less frequently 4 . Herein, we report GS-CA1, a new archetypal small-molecule HIV capsid inhibitor with exceptional potency against HIV-2 and all major HIV-1 types, including viral variants resistant to the ARVs currently in clinical use. Mechanism-of-action studies indicate that GS-CA1 binds directly to the HIV-1 capsid and interferes with capsid-mediated nuclear import of viral DNA, HIV particle production and ordered capsid assembly. GS-CA1 selects in vitro for unfit GS-CA1-resistant capsid variants that remain fully susceptible to other classes of ARVs. Its high metabolic stability and low solubility enabled sustained drug release in mice following a single subcutaneous dosing. GS-CA1 showed high antiviral efficacy as a long-acting injectable monotherapy in a humanized mouse model of HIV-1 infection, outperforming long-acting rilpivirine. Collectively, these results demonstrate the potential of ultrapotent capsid inhibitors as new long-acting agents for the treatment of HIV-1 infection.

Background: Despite the non-invasive nature of non-invasive prenatal testing (NIPT), there is still a need for a separate informed consent process before testing. The objectives of this study are to assess (a) knowledge and preferences of Chinese women in a major public hospital in Hong Kong who underwent NIPT, and (b) whether their knowledge and preferences differ depending on womens' characteristics and sources of information., Methods: Setting: Prenatal diagnosis and counselling clinic. Between February 2012 and September 2013, a questionnaire survey was distributed to all women who underwent NIPT after positive aneuploidy screening. As a pilot study, ten knowledge questions were designed based on the rapid response statement on Prenatal Detection of Down Syndrome using Massively Parallel Sequencing from the International Society for Prenatal Diagnosis in 2011. The source of women's knowledge and their preferences were also evaluated. While conventional screening was publicly funded, NIPT was not. Differences between subgroups were compared using chi square tests and logistic regression analysis., Results: Of 152 women who underwent NIPT, 135 (88.8%) completed their questionnaires. More than 90% of women recognised the possibility of false positive and false negative results. Slightly more than 70% of women knew the inferior sensitivity of NIPT compared to an invasive test, and the possibility of an uninformative test result, but were not aware of the complicated aspects of NIPT. Pregnant women with an advanced level of education or those who underwent NIPT before 15 weeks provided answers that was more accurate by around 10-20% in two to three knowledge questions than those without. These associations were confirmed by multivariate logistic regression analysis. The women received information on NIPT largely from their private doctors (47.4%) and web (41.5%). In their future pregnancies, more women would opt for NIPT (a self-financed item) after positive screening ('free' in a public hospital) (57.8%) than as a primary screening (30.4%)., Conclusions: It is feasible to use a questionnaire based on the ISPD statement on NIPT to assess women's knowledge of the test. The Chinese women who underwent NIPT recognised the limitations, but did not understand the complicated aspects. More information should be provided by health care professionals in order to facilitate an informed choice by patients. More women preferred NIPT as a contingent test than as a primary screening probably because of its high cost.

Objectives: To investigate how the introduction of noninvasive prenatal testing (NIPT) influenced women's testing choices following a positive Down syndrome screening., Methods: A retrospective study was conducted to compare differences in the uptake rates of invasive prenatal diagnosis (IPD) or no testing in one public hospital 1 year before (pre-NIPT) and 1 and 2 years after the introduction of NIPT in private in August 2011 using descriptive analysis and a χ² test. Conventional screening was funded publicly, but NIPT was not. Multivariable binary logistic regression was used to determine factors affecting choices., Results: In pre-NIPT and in years 1 and 2 after the introduction of NIPT, 306, 362 and 401 women who screened positive were seen, respectively. In year 1 and year 2, 12.6 and 26.7% of them underwent NIPT while IPD was decreased by 16.3 and 25.6%, respectively (p < 0.001). Both chorionic villus sampling and amniocentesis decreased in year 1, but only the former in year 2. However, the rate of declining further testing was similar before and after NIPT (p = 0.213). In multivariable analysis, first trimester screening, nulliparity and working women were significant predictors of accepting NIPT, while only nulliparity was a predictor of declining IPD (OR = 0.61)., Conclusions: Introduction of NIPT resulted in a significant decrease in IPD for 2 consecutive years.., (© 2014 S. Karger AG, Basel)

A protocol for a fluorescent intercalator displacement (FID) assay useful for establishing DNA binding selectivity, affinity, stoichiometry, and binding site size, and for distinguishing modes of DNA binding is presented.

Biologically active, therapeutically useful, DNA binding natural products continue to reveal new paradigms for sequence-selective recognition, to enlist beautiful mechanisms of in situ activation for DNA modification, to define new therapeutic targets, to exploit new mechanisms to achieve cellular selectivity, and to provide a rich source of new drugs. These attributes arise in compact structures of complex integrated function.

A summary of the qualitative and quantitative elements of a fluorescent intercalator displacement (FID) assay useful for establishing the DNA binding selectivity, affinity, stoichiometry, and binding site size and distinguishing modes of DNA binding is provided.

Protein titration displacement of ethidium bromide bound to hairpin deoxyoligonucleotides containing any sequence of interest provides a well-defined titration curve (measuring the loss of fluorescence derived from the DNA bound ethidium bromide) that provides both absolute binding constants (K(a)) and stoichiometry of binding. This use of a fluorescent intercalator displacement (FID) assay for establishing protein DNA binding affinity and selectivity is demonstrated with the examination of the LEF-1 HMG domain binding to hairpin deoxyoligonucleotides containing its commonly accepted consensus sequence 5'-CTTTGWW (W=A or T) and those modified (5'-CTNTGWW) to examine sequences implicated in early studies (5'-CTNTG). The effectiveness of the FID assay coupled with its technically non-demanding experimental use makes it an attractive alternative or complement to selection screening, footprinting or affinity cleavage, and electrophoretic mobility shift assays for detecting, characterizing, and quantitating protein DNA binding affinity and selectivity.

The binding affinities of several triplex forming oligonucleotides were determined using a fluorescent intercalator displacement (FID) assay.

Four hairpin polyamides bearing subtle N- and C-terminal substitutions were examined in a fluorescent intercalator displacement (FID) assay enlisting a library of 512 DNA hairpins that contain all possible five base pair sequences in a challenging probe of its capabilities for establishing DNA binding sequence selectivity. Not only did the assay define the global sequence selectivity expected based on known structural interactions and Dervan's pairing rules establishing the utility of the method for characterizing such polyamides, but previously unappreciated subtle substituent effects on global sequence selectivity were also revealed. Thus, we report the discovery of a novel five base pair high affinity binding site of the form 5'-WWCWW (vs 5'-WGWWW) for the polyamide ImPyPy-gamma-PyPyPy-beta-Dp and its structural basis.

The viability of using thiazole orange as an alternative to ethidium bromide in a fluorescent intercalator displacement (FID) assay is explored by profiling the DNA binding affinity and sequence selectivity of netropsin. Utilizing a library of hairpin deoxyoligonucleotides containing all possible four base-pair sequences, the method provides a high resolution profile of the DNA binding properties of small molecules in a high throughput format.

Full details of the development of a simple, nondestructive, and high-throughput method for establishing DNA binding affinity and sequence selectivity are described. The method is based on the loss of fluorescence derived from the displacement of ethidium bromide or thiazole orange from the DNA of interest or, in selected instances, the change in intrinsic fluorescence of a DNA binding agent itself and is applicable for assessing relative or absolute DNA binding affinities. Enlisting a library of hairpin deoxyoligonucleotides containing all five base pair (512 hairpins) or four base pair (136 hairpins) sequences displayed in a 96-well format, a compound's rank order binding to all possible sequences is generated, resulting in a high-resolution definition of its sequence selectivity using this fluorescent intercalator displacement (FID) assay. As such, the technique complements the use of footprinting or affinity cleavage for the establishment of DNA binding selectivity and provides the information at a higher resolution. The merged bar graphs generated by this rank order binding provide a qualitative way to compare, or profile, DNA binding affinity and selectivity. The 96-well format assay (512 hairpins) can be conducted at a minimal cost (presently ca. $100 for hairpin deoxyoligonucleotides/assay with ethiduim bromide or less with thiazole orange), with a rapid readout using a fluorescent plate reader (15 min), and is adaptable to automation (Tecan Genesis Workstation 100 robotic system). Its use in generating a profile of DNA binding selectivity for several agents including distamycin A, netropsin, DAPI, Hoechst 33258, and berenil is described. Techniques for establishing binding constants from quantitative titrations are compared, and recommendations are made for use of a Scatchard or curve fitting analysis of the titration binding curves as a reliable means to quantitate the binding affinity.

Full details of the total syntheses of thiocoraline (1) and BE-22179 (2), C(2) symmetric bicyclic octadepsipeptides possessing two pendant 3-hydroxyquinoline chromophores, are described in which their relative and absolute stereochemistry were established. Key elements of the approach include the late-stage introduction of the chromophore, symmetrical tetrapeptide coupling, macrocyclization of the 26-membered octadepsipeptide conducted at the single secondary amide site following disulfide formation, and a convergent assemblage of the tetradepsipeptide with introduction of the labile thiol ester linkage in the final coupling reaction under near racemization free conditions. By virtue of the late-stage introduction of the chromophore and despite the challenges this imposes on the synthesis, this approach provides ready access to a range of key chromophore analogues. Thiocoraline and BE-22179 were shown to bind to DNA by high-affinity bisintercalation analogous to echinomycin, but with little or no perceptible sequence selectivity. Both 1 and 2 were found to exhibit exceptional cytotoxic activity (IC(50) = 200 and 400 pM, respectively, L1210 cell line) comparable to echinomycin and one analogue, which bears the luzopeptin chromophore, was also found to be a potent cytotoxic agent.

The series of four dimers derived from head to tail coupling of the two enantiomers of the duocarmycin SA alkylation subunit are described.

Yuehchukene (YCK) is a novel bis-indole alkaloid with weak estrogenic activity. Biochemical studies showed that YCK could attenuate estrogenic action. In this study, the response of MCF-7, an estrogen-receptor-positive breast cancer cell line, under different combinations of estradiol, cyclophosphamide and YCK, was tested. From the dose-response curve, we discovered that 10(-2) M cyclophosphamide, even in its so-called 'bio-inert' form, could inhibit MCF-7 cell growth. However, the cytotoxic effect of cyclophosphamide was lost by reducing its concentration to approximately 1 x 10(-3) M. On the other hand, a low concentration ( approximately 10(-8)-10(-9) M) of YCK was found to potentiate the cytotoxic effect of cyclophosphamide on the MCF-7 cell line. Such an effect was absent in the estrogen-receptor-negative cell line MDA-MB-231. These findings, together with the dual role of a mixed estrogen and anti-estrogen effect, suggested that YCK and cyclophosphamide can be a potential combination in chemo-hormonal therapy for breast cancer., (Copyright 2000 S. Karger AG, Basel)

This paper focuses on the inhibitory mechanism of E-64 and its derivatives (epoxysuccinyl-based inhibitors) with some cysteine proteases, based on the binding modes observed in the x-ray crystal structures of their enzyme-inhibitor complexes. E-64 is a potent irreversible inhibitor against general cysteine proteases, and its binding modes with papain, actinidin, cathepsin L, and cathepsin K have been reviewed at the atomic level. E-64 interacts with the Sn subsites of cysteine proteases. Although the Sn-Pn (n = 1-3) interactions of the inhibitor with the main chains of the active site residues are similar in respective complexes, the significant difference is observed in the side-chain interactions of S2-P2 and S3-P3 pairs because of different residues constituting the respective subsites. E-64-c and CA074 are representative derivatives developed from E-64 as a clinical usable and a cathepsin B-specific inhibitors, respectively. In contrast with similar binding/inhibitory modes of E-64-c and E-64 for cysteine proteases, the inhibitory mechanism of cathepsin B-specific CA074 results from the binding to the Sn' subsite.

The antigen binding fragment from an IgG2a kappa murine monoclonal antibody with specificity for alpha-(2-->8)-linked sialic acid polymers has been prepared and crystallized in the absence of hapten. Crystals were grown by vapor diffusion equilibrium with 16-18% polyethylene glycol 4000 solutions. The structure was solved by molecular replacement methods and refined to a conventional R factor of 0.164 for data to 2.8 A. The binding site is observed to display a shape and distribution of charges that is complementary to that of the predicted conformation of the oligosaccharide epitope. A thermodynamic description of ligand binding has been compiled for oligosaccharides ranging in length from 9 to 41 residues, and the data for the largest ligand has been used in a novel way to estimate the size of the antigen binding site. A model of antigen binding is presented that satisfies this thermodynamic data, as well as a previously reported requirement of conformational specificity of the oligosaccharide. X-ray crystallographic and thermodynamic evidence are consistent with a binding site that accommodates at least eight sialic acid residues.