Search

Chenlu Gao,1– 3 Peng Li,1– 3 Christopher J Morris,1,2 Xi Zheng,1,3 Ma Cherrysse Ulsa,1,3 Lei Gao,1– 4 Frank AJL Scheer,1– 3 Kun Hu1– 3 1Division of Sleep and Circadian Disorders, Departments of Medicine and Neurology, Brigham and Women’s Hospital, Boston, MA, USA; 2Division of Sleep Medicine, Harvard Medical School, Boston, MA, USA; 3Broad Institute of MIT and Harvard, Cambridge, MA, USA; 4Department of Anesthesia, Critical Care and Pain Medicine, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USACorrespondence: Kun Hu, Brigham and Women’s Hospital, 221 Longwood Avenue, 040 BLI, Boston, MA, 02115, USA, Tel +1 617-525 8694, Fax +1 617-732 7337, Email khu1@bwh.harvard.eduPurpose: Actigraphy-based sleep detection algorithms were mostly validated using nighttime sleep, and their performance in detecting daytime sleep is unclear. We evaluated and compared the performance of Actiware and the Cole-Kripke algorithm (C-K) – two commonly used actigraphy-based algorithms – in detecting daytime and nighttime sleep.Participants and Methods: Twenty-five healthy young adults were monitored by polysomnography and actigraphy during two in-lab protocols with scheduled nighttime and/or daytime sleep (within-subject design). Mixed-effect models were conducted to compare the sensitivity, specificity, and F1 score (a less-biased measure of accuracy) of Actiware (with low/medium/high threshold setting, separately) and C-K in detecting sleep epochs from actigraphy recordings during nighttime/daytime. t-tests and intraclass correlation coefficients were used to assess the agreement between actigraphy-based algorithms and polysomnography in scoring total sleep time (TST).Results: Sensitivity was similar between nighttime (Actiware: 0.93– 0.99 across threshold settings; C-K: 0.61) and daytime sleep (Actiware: 0.93– 0.99; C-K: 0.66) for both the C-K and Actiware (daytime/nighttime×algorithm interaction: p > 0.1). Specificity for daytime sleep was lower (Actiware: 0.35– 0.54; C-K: 0.91) than that for nighttime sleep (Actiware: 0.37– 0.62; C-K: 0.93; p = 0.001). Specificity was also higher for C-K than Actiware (p < 0.001), with no daytime/nighttime×algorithm interaction (p > 0.1). C-K had lower F1 (nighttime = 0.74; daytime = 0.77) than Actiware (nighttime = 0.95– 0.98; daytime = 0.90– 0.91) for both nighttime and daytime sleep (all p < 0.05). The daytime-nighttime difference in F1 was opposite for Actiware (daytime: 0.90– 0.91; nighttime: 0.95– 0.98) and C-K (daytime: 0.77; nighttime: 0.74; interaction p = 0.003). Bias in TST was lowest in Actiware (with medium-threshold) for nighttime sleep (underestimation of 5.99 min/8h) and in Actiware (with low-threshold) for daytime sleep (overestimation of 17.75 min/8h).Conclusion: Daytime/nighttime sleep affected specificity and F1 but not sensitivity of actigraphy-based sleep scoring. Overall, Actiware performed better than the C-K algorithm. Actiware with medium-threshold was the least biased in estimating nighttime TST, and Actiware with low-threshold was the least biased in estimating daytime TST.Keywords: Actiware, Cole-Kripke algorithm, sleep scoring, shift worker, circadian rhythms

OBJECTIVE: The authors sought to identify ways in which the use of general practice computer systems could be improved to enhance safety in primary care. DESIGN: Qualitative study using semistructured interviews. PARTICIPANTS: Thirty one participants, representing a broad range of relevant disciplines and interest groups. Participants included clinicians, computer system and drug database suppliers, academics with interests in health informatics and members of governmental, professional and patient representative bodies. SETTING: UK. RESULTS: Participants identified deficiencies in current systems that pose serious threats to patient safety. To bring about improvements, providers need to supply clinicians with safe, accurate and accessible information for decision support; be aware of the importance of human ergonomics in the design of hazard alerts; consider the value of audit trails and develop mechanisms to allow for the accurate transfer of information between clinical computer systems. These improvements in computer systems will be most likely to occur if mandated through regulations. Individual practices are in need of improved education and training which focuses, in particular, on providing support with recording data accurately and using call, recall and reminders effectively. CONCLUSION: There are significant opportunities for improving the safety of general practice computer systems. Priorities include improving the knowledge base for clinical decision support, paying greater attention to human ergonomics in system design, improved staff training and the introduction of new regulations mandating system suppliers to satisfy essential safety requirements. [ABSTRACT FROM AUTHOR]

Our objective was to identify and establish consensus on the most important safety features of GP computer systems, with a particular emphasis on medicines management. We used a two-round electronic Delphi survey, completed by a 21-member multidisciplinary expert panel, all from the UK. The main outcome measure was percentage agreement of the panel members on the importance of the presence of a number of different safety features (presented as clinical statements) on GP computer systems. We found 90% or greater agreement on the importance of 32 (58%) statements. These statements, indicating issues considered to be of considerable importance (rated as important or very important), related to: computerised alerts; the need to avoid spurious alerts; making it difficult to override critical alerts; having audit trails of such overrides; support for safe repeat prescribing; effective computer-user interface; importance of call and recall management; and the need to be able to run safety reports. The high level of agreement among the expert panel members indicates clear themes and priorities that need to be addressed in any further improvement of safety features in primary care computing systems. [ABSTRACT FROM AUTHOR]

OBJECTIVE: To describe the process that has been undertaken to (i) validate further UK indicators for preventable drug-related morbidity (PDRM) generated by a previous study and (ii) develop additional new indicators derived from UK clinical practice. DESIGN: A two-round Delphi questionnaire survey. SETTING: A UK study based in primary care. PARTICIPANTS: A purposively selected sample of primary care pharmacists (n = 9) and general practitioners (n = 8). OUTCOME MEASURES: The establishment of consensus among the panel as to whether an indicator represented a PDRM. RESULTS: The pre-defined level of consensus was achieved for 24 indicators (59%; 24 of 41). CONCLUSIONS: We have identified that although mechanisms exist for identifying 'the outcome' component of PDRM indicators, identifying the 'pattern of care' that is needed to prevent their occurrence is far more challenging. Nonetheless, we have taken considerable steps along the path of validating such indicators. Future operationalization in a general practice setting should help to facilitate improvements in medicine management in primary care and ultimately benefit patients. [ABSTRACT FROM AUTHOR]

Electrospraying is a processing technique that has gained much interest to prepare polymeric particles. The technique operates at ambient temperature, thereby avoiding heat induced degradation of labile therapeutics (e.g. peptides and proteins). Exposure to organic solvents can be minimised by co-axial electrospraying through separation of core (aqueous) and shell (organic) solvents. However, aqueous solutions are often difficult to electrospray due to high surface tension. Immiscibility between the core-shell solvents creates a further process challenge. Herein, we describe for the first time the use of hydrophobic ion pairing (HIP) to encapsulate a polypeptide into polymeric particles prepared by co-axial electrospraying. Peptide ion pairs were prepared to incorporate a model peptide - teriparatide - into an organic solvent, permitting facile electrospraying while also protecting the peptide from denaturation. Teriparatide loaded PLGA particles were generated by electrospraying from aqueous or ethanolic peptide solutions (core). A PLGA solution in chloroform (with and without co-solvents) was employed as the shell solution. The aqueous core solution led to a teriparatide encapsulation efficiency of 79.2 ± 19.8 %, which was not significantly different from the ethanolic core (57.1 ± 14.5 %). When aqueous solutions were used the process lacked reproducibility, resulting in low process yields (61.3 ± 4.0 %). In contrast, when an organic core was used a dry powder bed was achieved with a yield of 102.2 ± 8.8 %. The peptide's integrity and biological functionality were retained after electrospraying as ion pairs, as evidenced in a cell-based PTH1 receptor binding assay., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Teriparatide (and analogue peptides) are the only FDA approved anabolic treatments for osteoporosis. Current therapies are administered as a daily subcutaneous injection, which limits patient adherence and clinical efficacy. To achieve the desired anabolic effect, a controlled delivery system must ensure a pulsatile release profile over a prolonged period. Thermo-responsive formulations (e.g. liposomes) can undergo a temperature-related phase-transition which can allow active control of drug release. Herein, thermo-responsive liposomes were developed to permit control over teriparatide release rate through modulation of temperature. Entrapment of hydrophilic molecules, including peptides, within liposomes remains challenging due to the large volume of hydration. In this work, hydrophobic ion pairing was employed for the first time to enhance peptide entrapment within liposomes. The method resulted in a hydrophobic complex that achieved high teriparatide entrapment (>75 %) in sub-200 nm monodispersed liposomes. Hydrophobic ion pairing outperformed other entrapment approaches. Several liposomal formulations with transition temperatures between 38 and 50 °C were obtained by modulation of the phospholipid composition. In vitro assays demonstrated temperature-dependent release kinetics with faster rates of release observed at/above the transition temperature. The maintenance of biological activity of released teriparatide was demonstrated in a cell-based assay utilising the PTH1 receptor. Overall, this provides the first proof-of-concept of the suitability of thermo-responsive systems for pulsatile delivery of teriparatide and similar peptides., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

The CACHE challenges are a series of prospective benchmarking exercises to evaluate progress in the field of computational hit-finding. Here we report the results of the inaugural CACHE challenge in which 23 computational teams each selected up to 100 commercially available compounds that they predicted would bind to the WDR domain of the Parkinson's disease target LRRK2, a domain with no known ligand and only an apo structure in the PDB. The lack of known binding data and presumably low druggability of the target is a challenge to computational hit finding methods. Of the 1955 molecules predicted by participants in Round 1 of the challenge, 73 were found to bind to LRRK2 in an SPR assay with a K D lower than 150 μM. These 73 molecules were advanced to the Round 2 hit expansion phase, where computational teams each selected up to 50 analogs. Binding was observed in two orthogonal assays for seven chemically diverse series, with affinities ranging from 18 to 140 μM. The seven successful computational workflows varied in their screening strategies and techniques. Three used molecular dynamics to produce a conformational ensemble of the targeted site, three included a fragment docking step, three implemented a generative design strategy and five used one or more deep learning steps. CACHE #1 reflects a highly exploratory phase in computational drug design where participants adopted strikingly diverging screening strategies. Machine learning-accelerated methods achieved similar results to brute force (e.g., exhaustive) docking. First-in-class, experimentally confirmed compounds were rare and weakly potent, indicating that recent advances are not sufficient to effectively address challenging targets.

Pharmaceutical companies routinely screen compounds for hemodynamics related safety risk. In vitro secondary pharmacology is initially used to prioritize compounds while in vivo studies are later used to quantify and translate risk to humans. This strategy has shown limitations but could be improved via the incorporation of molecular findings in the animal-based toxicological risk assessment. The aim of this study is to develop a mathematical model for rat and dog species that can integrate secondary pharmacology modulation and therefore facilitate the overall pre-clinical safety translation assessment. Following an extensive literature review, we built two separate models recapitulating known regulation processes in dogs and rats. We describe the resulting models and show that they can reproduce a variety of interventions in both species. We also show that the models can incorporate the mechanisms of action of a pre-defined list of 50 pharmacological mechanisms whose modulation predict results consistent with known pharmacology. In conclusion, a mechanistic model of hemodynamics regulations in rat and dog species has been developed to support mechanism-based safety translation in drug discovery and development., Competing Interests: Author(s) CM, KH, GD, MR, LS, CV, and AP were employed by the AstraZeneca., (Copyright © 2024 Morris, Rolf, Starnes, Villar, Pointon, Kimko and Di Veroli.)

The insulin-linked polymorphic region is a variable number of tandem repeats region of DNA in the promoter of the insulin gene that regulates transcription of insulin. This region is known to form the alternative DNA structures, i-motifs and G-quadruplexes. Individuals have different sequence variants of tandem repeats and although previous work investigated the effects of some variants on G-quadruplex formation, there is not a clear picture of the relationship between the sequence diversity, the DNA structures formed, and the functional effects on insulin gene expression. Here we show that different sequence variants of the insulin linked polymorphic region form different DNA structures in vitro. Additionally, reporter genes in cellulo indicate that insulin expression may change depending on which DNA structures form. We report the crystal structure and dynamics of an intramolecular i-motif, which reveal sequences within the loop regions forming additional stabilising interactions that are critical to formation of stable i-motif structures. The outcomes of this work reveal the detail in formation of stable i-motif DNA structures, with potential for rational based drug design for compounds to target i-motif DNA., (© 2024. The Author(s).)

Background and Purpose: Decreased aortic compliance is a precursor to numerous cardiovascular diseases. Compliance is regulated by the rigidity of the aortic wall and the vascular smooth muscle cells (VSMCs). Extracellular matrix stiffening, observed during ageing, reduces compliance. In response to increased rigidity, VSMCs generate enhanced contractile forces that result in VSMC stiffening and a further reduction in compliance. Mechanisms driving VSMC response to matrix rigidity remain poorly defined., Experimental Approach: Human aortic-VSMCs were seeded onto polyacrylamide hydrogels whose rigidity mimicked either healthy (12 kPa) or aged/diseased (72 kPa) aortae. VSMCs were treated with pharmacological agents prior to agonist stimulation to identify regulators of VSMC volume regulation., Key Results: On pliable matrices, VSMCs contracted and decreased in cell area. Meanwhile, on rigid matrices VSMCs displayed a hypertrophic-like response, increasing in area and volume. Piezo1 activation stimulated increased VSMC volume by promoting calcium ion influx and subsequent activation of PKC and aquaporin-1. Pharmacological blockade of this pathway prevented the enhanced VSMC volume response on rigid matrices whilst maintaining contractility on pliable matrices. Importantly, both piezo1 and aquaporin-1 gene expression were up-regulated during VSMC phenotypic modulation in atherosclerosis and after carotid ligation., Conclusions and Implications: In response to extracellular matrix rigidity, VSMC volume is increased by a piezo1/PKC/aquaporin-1 mediated pathway. Pharmacological targeting of this pathway specifically blocks the matrix rigidity enhanced VSMC volume response, leaving VSMC contractility on healthy mimicking matrices intact. Importantly, upregulation of both piezo1 and aquaporin-1 gene expression is observed in disease relevant VSMC phenotypes., (© 2023 The Authors. British Journal of Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

The skin is a multifunctional organ, forming a barrier between the external and internal environment, thereby functioning as a safeguard against extrinsic factors. Autophagy has been implicated in epidermal differentiation and in preserving skin homeostasis. LC3-associated phagocytosis (LAP) uses some but not all components of autophagy. The Atg16l1 ( Δ WD) mouse model lacks the WD40 domain required for LAP and has been widely used to study the effects of LAP deficiency and autophagy on tissue homeostasis and response to infection. In this study, the Δ WD model was used to study the relationship between LAP and skin homeostasis by determining whether LAP-deficient mice display a cutaneous phenotype. Skin histology of wild-type and Δ WD mice aged 1 year revealed minor morphological differences in the tail skin dermal layer. RT-qPCR and western blot analysis showed no differences in key keratin expression between genotypes. Skin barrier formation, assessed by dye permeation assays, demonstrated full and proper formation of the skin barrier at embryonic day 18.5 in both genotypes. Biomechanical analysis of the skin showed decreased skin elasticity in aged Δ WD but not wild-type mice. In summary, the LAP-deficient Δ WD mice displayed subtle alterations in dermal histology and age-related biomechanical changes., (© 2024 The Authors.)

Purpose: Antiangiogenic therapies are known to cause high radiographic response rates due to reduction in vascular permeability resulting in a lower degree of contrast extravasation. In this study, we investigate the prognostic ability for model-derived parameters describing enhancing tumor volumetric dynamics to predict survival in recurrent glioblastoma treated with antiangiogenic therapy., Experimental Design: N = 276 patients in two phase II trials were used as training data, including bevacizumab ± irinotecan (NCT00345163) and cabozantinib (NCT00704288), and N = 74 patients in the bevacizumab arm of a phase III trial (NCT02511405) were used for validation. Enhancing volumes were estimated using T1 subtraction maps, and a biexponential model was used to estimate regrowth (g) and regression (d) rates, time to tumor regrowth (TTG), and the depth of response (DpR). Response characteristics were compared to diffusion MR phenotypes previously shown to predict survival., Results: Optimized thresholds occurred at g = 0.07 months-1 (phase II: HR = 0.2579, P = 5 × 10-20; phase III: HR = 0.2197, P = 5 × 10-5); d = 0.11 months-1 (HR = 0.3365, P < 0.0001; HR = 0.3675, P = 0.0113); TTG = 3.8 months (HR = 0.2702, P = 6 × 10-17; HR = 0.2061, P = 2 × 10-5); and DpR = 11.3% (HR = 0.6326, P = 0.0028; HR = 0.4785, P = 0.0206). Multivariable Cox regression controlling for age and baseline tumor volume confirmed these factors as significant predictors of survival. Patients with a favorable pretreatment diffusion MRI phenotype had a significantly longer TTG and slower regrowth., Conclusions: Recurrent glioblastoma patients with a large, durable radiographic response to antiangiogenic agents have significantly longer survival. This information is useful for interpreting activity of antiangiogenic agents in recurrent glioblastoma., (©2023 American Association for Cancer Research.)

Objective: Shift workers typically experience misalignment between their circadian system and behavioral/environmental cycles and have an increased risk for obesity. Experimental studies in non-shift workers have suggested that circadian misalignment can disrupt energy balance regulation. This study examined the impact of circadian misalignment in the most relevant population, i.e., chronic shift workers., Methods: Seven healthy chronic night shift workers underwent a randomized crossover study with two 3-day laboratory protocols: a night work protocol including 12-hour inverted behavioral/environmental cycles (circadian misalignment) and a day work protocol (circadian alignment)., Results: Circadian misalignment led to a ~17% increase in 24-hour acylated ghrelin levels in the chronic shift workers (p = 0.009). Consistently, circadian misalignment resulted in ~14% higher hunger at breakfast in the night shift (p = 0.04). Circadian misalignment did not significantly change fasting and postprandial energy expenditure or respiratory exchange ratio (all p > 0.32). Unexpectedly, 24-hour behavioral activity levels were ~38% higher (p < 0.0001) during circadian misalignment, despite a concurrent increase in sleepiness (p = 0.03)., Conclusions: These results reveal that circadian misalignment, while carefully controlling for dietary intake, increases acylated ghrelin in chronic shift workers. Further studies should test whether the observed acute effects of circadian misalignment in chronic shift workers contribute to their increased obesity risk in the long term., (© 2023 The Obesity Society.)

A woman in her 50s with a medical history of cirrhosis, alcohol use disorder, primary biliary cholangitis and extended spectrum beta lactamase (ESBL) Klebsiella presented with weakness, cough and abdominal pain with positive blood cultures for ESBL Klebsiella , and was treated with intravenous meropenem and patient symptoms improved. Testing for Strongyloides antibodies was positive, so she was treated with ivermectin. Strongyloidiasis-associated Gram-negative rod (GNR) bacteremia are rare conditions; however, it is important to consider an underlying strongyloidiasis in recurrent GNR bacteremia to prevent recurrent hospitalisation and morbidity., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Limited 2023. No commercial re-use. See rights and permissions. Published by BMJ.)

Burkholderia pseudomallei is the causative agent of the tropical disease, melioidosis. It is intrinsically resistant to many antimicrobials and treatment requires an onerous regimen of intravenous and orally administered drugs. Relapse of disease and high rates of mortality following treatment are common, demonstrating the need for new anti- Burkholderia agents. The cationic bola-amphiphile, 12,12'-(dodecane-1,12-diyl) bis (9-amino-1,2,3,4-tetrahydroacridinium), referred to as 12-bis-THA, is a molecule with the potential to treat Burkholderia infections. 12-bis-THA spontaneously forms cationic nanoparticles that bind anionic phospholipids in the prokaryotic membrane and are readily internalized. In this study, we examine the antimicrobial activity of 12-bis-THA against strains of Burkholderia thailandensis . As B. pseudomallei produces a polysaccharide capsule we first examined if this extra barrier influenced the activity of 12-bis-THA which is known to act on the bacterial envelope. Therefore two strains of B. thailandensis were selected for further testing, strain E264 which does not produce a capsule and strain E555 which does produce a capsule that is chemically similar to that found in B. pseudomallei . In this study no difference in the minimum inhibitory concentration (MIC) was observed when capsulated (E555) and unencapsulated (E264) strains of B. thailandensis were compared, however time-kill analysis showed that the unencapsulated strain was more susceptible to 12-bis-THA. The presence of the capsule did not affect the membrane permeation of 12-bis-THA at MIC concentrations. Proteomic and metabolomic analyses showed that 12-bis-THA causes a shift in central metabolism away from glycolysis and glyoxylate cycle, and suppressed the production of the F 1 domain of ATP synthase. In summary, we provide insight into the molecular mechanisms underpinning the activity of 12-bis-THA against B. thailandensis and discuss its potential for further development., Competing Interests: MM was previously the CSO of Procarta Biosystems, the developer of 12-bis-THA. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Pattinson, Bahia, Le Gall, Morris, Harding and McArthur.)

Background: No pharmacological treatments are specifically indicated for painful small fibre neuropathy (SFN). CONVEY, a phase 2 enriched-enrolment study, evaluated the efficacy and safety of vixotrigine, a voltage- and use-dependent sodium channel blocker, in participants with idiopathic or diabetes-associated painful SFN., Methods: CONVEY was a phase 2, multicentre, placebo-controlled, double-blind (DB), enriched-enrolment, randomised withdrawal study. The study was conducted at 68 sites in 13 countries (Europe and Canada) between May 17, 2018, and April 12, 2021. Following a 4-week open-label period in which 265 adults with painful SFN (a mixture of large and small fibre neuropathy was not exclusionary) received oral vixotrigine 350 mg twice daily (BID), 123 participants (with a ≥30% reduction from baseline in average daily pain [ADP] score during the open-label period) were randomised 1:1:1 to receive 200 mg BID, 350 mg BID or placebo for a 12-week double-blind (DB) period. Primary endpoint was change from baseline in ADP at DB Week 12. Secondary endpoints included the proportion of participants with a ≥30% reduction from baseline in ADP and the proportion of Patient Global Impression of Pain (PGIC) responders at DB Week 12. Treatment-emergent adverse events (AEs) were monitored. Statistical significance was set at 0.10 (2-sided). The trial was registered on ClinicalTrials.gov (NCT03339336) and on ClinicalTrialsregister.eu (2017-000991-27)., Findings: A statistically significant difference from placebo in least squares mean reduction in ADP score from baseline to DB Week 12 was observed with vixotrigine 200 mg BID (-0.85; SE, 0.43; 95% CI, -1.71 to 0.00; p = 0.050) but not 350 mg BID (-0.17; SE, 0.43; 95% CI, -1.01 to 0.68; p = 0.70). Numerically, but not statistically significantly, more participants who received vixotrigine vs placebo experienced a ≥30% ADP reduction from baseline (68.3-72.5% vs 52.5%), and only the 350 mg BID group had significantly more PGIC responders vs placebo (48.8% vs 30.0%; odds ratio = 2.60; 95% CI, 0.97-6.99; p = 0.058) at DB Week 12. AEs were mostly mild to moderate in the vixotrigine groups. The most common AEs (≥5% of vixotrigine-treated participants) in the DB 200 mg BID and 350 mg BID vixotrigine groups were falls, nasopharyngitis, muscle spasm, and urinary tract infection., Interpretation: In our study, vixotrigine 200 mg BID, but not 350 mg BID, met the primary endpoint; more vixotrigine-treated participants experienced a ≥30% reduction from baseline in ADP at DB Week 12. Vixotrigine (at both dosages) was well tolerated in participants with SFN., Funding: Biogen, Inc., Competing Interests: C.G.F. discloses grants from the European Union's Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant for PAIN-Net, molecule-to-man pain network (grant no. 721841), from Grifols and Lamepro for a trial on IVIg in small fibre neuropathy, and from Prinses Beatrix Spierfonds and serving on steering committees and advisory boards for Biogen Inc, Vertex Pharmaceuticals Incorporated, Lilly, and OliPass for studies in small fibre neuropathy. N.A. discloses grants from DOLORisk (European Union's Horizon) and honoraria from Pfizer Inc, Sanofi, Novartis AG, Grünenthal, Air Liquide, and Merz outside the submitted work and from Biogen Inc for scientific advice on vixotrigine over the past 3 years. G.L. discloses support from the European Commission, Italian Ministry of Health, Lombardy Foundation for Biomedical Research (FRRB), and Italian Research Foundation for Amyotrophic Lateral Sclerosis (AriSLA) and scientific advisory for CSL Behring, Biogen Inc, Vertex Pharmaceuticals Incorporated, Chromocell Corporation, Janssen Pharmaceuticals, Inc, Lilly, and the Bracco Group. R.H.D. discloses research grants and contracts received from the US Food and Drug Administration and the US National Institutes of Health in the past 5 years and compensation for serving on advisory boards or consulting on clinical trial methods from Abide, Acadia, Adynxx, Analgesic Solutions, Aptinyx, Aquinox, Asahi Kasei, Astellas, Beckley, Biogen, Biohaven, Biosplice, Boston Scientific, Braeburn, Cardialen, Celgene, Centrexion, Chiesi, Chromocell, Clexio, Collegium, Concert, Confo, Decibel, Editas, Eli Lilly, Endo, Ethismos (equity), Eupraxia, Exicure, Glenmark, Gloriana, Grace, Hope, Lotus, Mainstay, Merck, Mind Medicine (equity), Neumentum, Neurana, NeuroBo, Novaremed, Novartis, OCT, OliPass, Pfizer, Q-State, Reckitt Benckiser, Regenacy (also equity), Sangamo, Sanifit, Scilex, Semnur, SIMR Biotech, Sinfonia, SK Biopharmaceuticals, Sollis, SPRIM, Teva, Theranexus, Toray, Vertex, Vizuri, and WCG. R.F. discloses personal compensation and/or stock options for serving on scientific advisory boards for AlgoRx, Allergan, Applied Therapeutics, Clexio, Collegium, CND Life Sciences, Glenmark, GW Pharma, GlaxoSmithKline, Eli Lilly, Lundbeck, Maxona, Novartis, NeuroBo, Regenacy, Vertex, and Wörwag. K.T.D., H.F., M.S., C.J.M., and M.K. are employees of and may hold stock and/or stock options in Biogen Inc. A.H. and H.N. are former employees of Biogen. A.H. is currently employed by Pfizer, Inc, and H.N. by Sumitovant Biopharma., (© 2023 The Authors.)

Urinary tract infection (UTI) afflicts millions of patients globally each year. While the majority of UTIs are successfully treated with orally administered antibiotics, the impact of oral antibiotics on the host microbiota is under close research scrutiny and the potential for dysbiosis is a cause for concern. Optimal treatment of UTI relies upon the selection of an agent which displays appropriate pharmacokinetic-pharmacodynamic (PK-PD) properties that will deliver appropriately high concentrations in the urinary tract after oral administration. Alternatively, high local concentrations of antibiotic at the urothelial surface can be achieved by direct instillation into the urinary tract. For antibiotics with the appropriate physicochemical properties, this can be of critical importance in cases for which an intracellular urothelial bacterial reservoir is suspected. In this review, we summarise the underpinning biopharmaceutical barriers to effective treatment of UTI and provide an overview of the evidence for the deployment of the intravesical administration route for antibiotics.

Molecular docking tools are regularly used to computationally identify new molecules in virtual screening for drug discovery. However, docking tools suffer from inaccurate scoring functions with widely varying performance on different proteins. To enable more accurate ranking of active over inactive ligands in virtual screening, we created a machine learning consensus docking tool, MILCDock, that uses predictions from five traditional molecular docking tools to predict the probability a ligand binds to a protein. MILCDock was trained and tested on data from both the DUD-E and LIT-PCBA docking datasets and shows improved performance over traditional molecular docking tools and other consensus docking methods on the DUD-E dataset. LIT-PCBA targets proved to be difficult for all methods tested. We also find that DUD-E data, although biased, can be effective in training machine learning tools if care is taken to avoid DUD-E's biases during training.

Background: Effective identification of individuals at increased risk for developing opioid use disorder (OUD) could reduce the overdose fatalities and mitigate the harm of the opioid epidemic. Early evidence has linked certain subjective experiences during the first substance exposure to subsequent substance misuse. This is consistent with anecdotal evidence that "first response" to opioids may distinguish those who later develop OUD from those who do not, further suggesting individual differences in neural activity and responses to specific substances., Objective: The objective of this scoping review was to evaluate the evidence on the relationship between the subjective first response to opioids and the risk of developing OUD., Methods: Ovid MEDLINE search (through 1/29/2021), followed by the search for articles published by the first/senior authors (PubMed) and references citing (Web of Science Citing Reference) identified eligible publications, was focused on studies involving humans, published in English, and describing the initial subjective response to opioid exposure and its association with OUD development or its risk. Systematic data extraction was completed for each eligible study., Results: 3364 title/abstracts and 48 full-text articles were reviewed. Four articles, describing six studies (526 adult participants) were eligible and included. These studies were methodologically heterogeneous, including 2 randomized controlled trials, and 1 case-control, and 3 cross-sectional studies, with small sample sizes, precluding data pooling and meta-analysis. Results suggested that individuals with OUD or at elevated risk for OUD (determined by the validated OUD risk assessment surveys) were more likely to report their "first opioid response" as euphoric compared to their counterparts., Conclusions: Limited evidence suggests that subjectively-reported euphoric response to first opioid exposure could predict the OUD risk. Prospective studies are needed to evaluate the utility of assessing first response (e.g., via brief point-of-care screening surveys) for informing clinical decisions when prescribing opioids., (Copyright © 2022 Elsevier Inc. All rights reserved.)

There are thousands of compounds shown to interact with G-quadruplex DNA, yet very few which target i-motif (iM) DNA. Previous work showed that tobramycin can interact with iM- DNA, indicating the potential for sugar-molecules to target these structures. Computational approaches indicated that the sugar-containing natural products baicalin and geniposidic acid had potential to target iM-DNA. We assessed the DNA interacting properties of these compounds using FRET-based DNA melting and a fluorescence-based displacement assay using iM-DNA structures from the human telomere and the insulin linked polymorphic region (ILPR), as well as complementary G-quadruplex and double stranded DNA. Both baicalin and geniposidic acid show promise as iM-interacting compounds with potential for use in experiments into the structure and function of i-motif forming DNA sequences and present starting points for further synthetic development of these as probes for iM-DNA., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Effective mentoring of undergraduate students is a growing requirement for the promotion of faculty at many universities. It is often challenging for young investigators to define a successful mentoring strategy, partially due to the absence of a broadly accepted definition of what mentoring should entail. To overcome this, an outcome-oriented mentoring framework was developed and used with more than 25 students over three years. It was found that a systematic mentoring approach can help students quickly realize their scientific potential and result in meaningful contributions to science. This report especially shows how the Critical Assessment of Protein Structure Prediction (CASP14) challenge was used to amplify student research efforts. As a result of this challenge, multiple publications, presentations and scholarships were awarded to the participating students. The mentoring framework continues to see much success in allowing undergraduate students, including students from underrepresented groups, to foster scientific talent and make meaningful contributions to the scientific community., (open access.)

Introduction: Progress in the knowledge of metabolic interactions between cancer and its microenvironment is ongoing and may lead to novel therapeutic approaches. Until recently, melanoma was considered a glycolytic tumour due to mutations in mitochondrial-DNA, however, these malignant cells can regain OXPHOS capacity via the transfer of mitochondrial-DNA, a process that supports their proliferation in-vitro and in-vivo. Here we study how melanoma cells acquire mitochondria and how this process is facilitated from the tumour microenvironment., Methods: Primary melanoma cells, and MSCs derived from patients were obtained. Genes' expression and DNA quantification was analysed using Real-time PCR. MSC migration, melanoma proliferation and tumour volume, in a xenograft subcutaneous mouse model, were monitored through bioluminescent live animal imaging., Results: Human melanoma cells attract bone marrow-derived stromal cells (MSCs) to the primary tumour site where they stimulate mitochondrial biogenesis in the MSCs through upregulation of PGC1a. Mitochondria are transferred to the melanoma cells via direct contact with the MSCs. Moreover, inhibition of MSC-derived PGC1a was able to prevent mitochondrial transfer and improve NSG melanoma mouse tumour burden., Conclusion: MSC mitochondrial biogenesis stimulated by melanoma cells is prerequisite for mitochondrial transfer and subsequent tumour growth, where targeting this pathway may provide an effective novel therapeutic approach in melanoma., (© 2022. The Author(s).)

The daily rhythm of plasma melatonin concentrations is typically unimodal, with one broad peak during the circadian night and near-undetectable levels during the circadian day. Light at night acutely suppresses melatonin secretion and phase shifts its endogenous circadian rhythm. In contrast, exposure to darkness during the circadian day has not generally been reported to increase circulating melatonin concentrations acutely. Here, in a highly-controlled simulated night shift protocol with 12-h inverted behavioral/environmental cycles, we unexpectedly found that circulating melatonin levels were significantly increased during daytime sleep (p < .0001). This resulted in a secondary melatonin peak during the circadian day in addition to the primary peak during the circadian night, when sleep occurred during the circadian day following an overnight shift. This distinctive diurnal melatonin rhythm with antiphasic peaks could not be readily anticipated from the behavioral/environmental factors in the protocol (e.g., light exposure, posture, diet, activity) or from current mathematical model simulations of circadian pacemaker output. The observation, therefore, challenges our current understanding of underlying physiological mechanisms that regulate melatonin secretion. Interestingly, the increase in melatonin concentration observed during daytime sleep was positively correlated with the change in timing of melatonin nighttime peak (p = .002), but not with the degree of light-induced melatonin suppression during nighttime wakefulness (p = .92). Both the increase in daytime melatonin concentrations and the change in the timing of the nighttime peak became larger after repeated exposure to simulated night shifts (p = .002 and p = .006, respectively). Furthermore, we found that melatonin secretion during daytime sleep was positively associated with an increase in 24-h glucose and insulin levels during the night shift protocol (p = .014 and p = .027, respectively). Future studies are needed to elucidate the key factor(s) driving the unexpected daytime melatonin secretion and the melatonin rhythm with antiphasic peaks during shifted sleep/wake schedules, the underlying mechanisms of their relationship with glucose metabolism, and the relevance for diabetes risk among shift workers., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Night work increases diabetes risk. Misalignment between the central circadian “clock” and daily behaviors, typical in night workers, impairs glucose tolerance, likely due to internal misalignment between central and peripheral circadian rhythms. Whether appropriate circadian alignment of eating can prevent internal circadian misalignment and glucose intolerance is unknown. In a 14-day circadian paradigm, we assessed glycemic control during simulated night work with either nighttime or daytime eating. Assessment of central (body temperature) and peripheral (glucose and insulin) endogenous circadian rhythms happened during constant routine protocols before and after simulated night work. Nighttime eating led to misalignment between central and peripheral (glucose) endogenous circadian rhythms and impaired glucose tolerance, whereas restricting meals to daytime prevented it. These findings offer a behavioral approach to preventing glucose intolerance in shift workers.

The lymphatics maintain fluid balance by returning interstitial fluid to veins via contraction/compression of vessel segments with check valves. Disruption of lymphatic pumping can result in a condition called lymphedema with interstitial fluid accumulation. Lymphedema treatments are often ineffective, which is partially attributable to insufficient understanding of specialized lymphatic muscle lining the vessels. This muscle exhibits cardiac-like phasic contractions and smooth muscle-like tonic contractions to generate and regulate flow. To understand the relationship between this sub-cellular contractile machinery and organ-level pumping, we have developed a multiscale computational model of phasic and tonic contractions in lymphatic muscle and coupled it to a lymphangion pumping model. Our model uses the sliding filament model (Huxley in Prog Biophys Biophys Chem 7:255-318, 1957) and its adaptation for smooth muscle (Mijailovich in Biophys J 79(5):2667-2681, 2000). Multiple structural arrangements of contractile components and viscoelastic elements were trialed but only one provided physiologic results. We then coupled this model with our previous lumped parameter model of the lymphangion to relate results to experiments. We show that the model produces similar pressure, diameter, and flow tracings to experiments on rat mesenteric lymphatics. This model provides the first estimates of lymphatic muscle contraction energetics and the ability to assess the potential effects of sub-cellular level phenomena such as calcium oscillations on lymphangion outflow. The maximum efficiency value predicted (40%) is at the upper end of estimates for other muscle types. Spontaneous calcium oscillations during diastole were found to increase outflow up to approximately 50% in the range of frequencies and amplitudes tested., (© 2021. The Author(s).)

An effective delivery vehicle of genetic materials to their target site is the key to a successful gene therapy. In many cases, nanoparticles are used as the vehicle of choice and the efficiency of the delivery relies heavily on the physicochemical properties of the nanoparticles. Microfluidics, although being a low throughput method, has been increasingly researched for the preparation of nanoparticles. A range of superior properties were claimed in the literature for microfluidic-prepared platforms, but no evidence on direct comparison of the properties of the nanoparticles prepared by microfluidics and conventional high throughput method exists, leaving the industry with little guidance on how to select effective large-scale nanoparticle manufacturing method. This study used plasmid DNA-loaded PLGA-Eudragit nanoparticles as the model system to critically compare the nanoparticles prepared by conventional and microfluidics-assisted nanoprecipitation. The PLGA-Eudragit nanoparticles prepared by microfluidics were found to be statistically significantly larger than the ones prepared by conventional nanoprecipitation. PLGA-Eudragit nanoparticle prepared conventionally showed higher DNA loading efficiency. Although the DNA-loaded nanoparticles prepared by both methods did not induce significant cytotoxicity, the transfection efficiency was found to be higher for the ones prepared conventionally which has good potential for plasmid delivery. This study for the first time provides a direct comparison of the DNA-loaded nanoparticles prepared by microfluidic and conventional methods. The findings bring new insights into critical evaluation of the selection of manufacturing methods of nanoparticles for future gene therapy., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Significance: This study establishes normative data and a testing procedure for the oculomotor assessment tool. The oculomotor assessment tool standardizes visual targets for the Vestibular/OculoMotor Screening assessment and provides additional metrics that may aid in the differentiation between those with normal and those with abnormal oculomotor function potentially caused by a concussion., Purpose: This study aimed to assess the oculomotor endurance of healthy participants with no self-reported history of concussions using the oculomotor assessment tool., Methods: Healthy participants (n = 376, average age of 20.4 years, range of 11 to 34 years, with no self-reported history of concussions) were recruited to perform the following three tasks for 60 seconds each: (1) horizontal saccades, (2) vertical saccades, and (3) vergence jumps. The participants were instructed to alternate visual fixation between two targets for each of the tasks as fast as they could without overshooting or undershooting the visual target. The differences in the number of eye movements between the initial and latter 30 seconds of the 1-minute test were analyzed., Results: A statistical difference (P < .001) was observed in the number of eye movements for all three tasks (horizontal saccades [70 ± 15 for initial 30 seconds, 63 ± 13 for latter 30 seconds], vertical saccades [68 ± 14, 63 ± 13], and vergence jumps [43 ± 11, 39 ± 10]) between the initial and latter 30 seconds. No significant differences were identified in the number of eye movements or the change in eye movements between the initial and latter 30 seconds based on sex., Conclusions: These results establish a normative database for various eye movements. These data could potentially be used to compare different patient populations who have binocular endurance dysfunctions potentially due to traumatic brain injury, such as patients with concussion(s)., Competing Interests: Conflict of Interest Disclosure: New Jersey Institute of Technology has licensed the right to manufacture and sell the OculoMotor Assessment Tool device to Gulden Ophthalmics on behalf of the authors CY and TLA., (Copyright © 2021 American Academy of Optometry.)

The prediction of amino acid contacts from protein sequence is an important problem, as protein contacts are a vital step towards the prediction of folded protein structures. We propose that a powerful concept from deep learning, called ensembling, can increase the accuracy of protein contact predictions by combining the outputs of different neural network models. We show that ensembling the predictions made by different groups at the recent Critical Assessment of Protein Structure Prediction (CASP13) outperforms all individual groups. Further, we show that contacts derived from the distance predictions of three additional deep neural networks-AlphaFold, trRosetta, and ProSPr-can be substantially improved by ensembling all three networks. We also show that ensembling these recent deep neural networks with the best CASP13 group creates a superior contact prediction tool. Finally, we demonstrate that two ensembled networks can successfully differentiate between the folds of two highly homologous sequences. In order to build further on these findings, we propose the creation of a better protein contact benchmark set and additional open-source contact prediction methods.

Normal growth and development of lymphatic structures depends on mechanical forces created by accumulating interstitial fluid. However, prolonged exposure to pathologic mechanical stimuli generated by chronically elevated lymph flow results in lymphatic dysfunction. The mechanisms that transduce these mechanical forces are not fully understood. Our objective was to investigate molecular mechanisms that alter the growth and metabolism of isolated lymphatic endothelial cells (LECs) exposed to prolonged pathologically elevated lymph flow in vivo within the anatomic and physiologic context of a large animal model of congenital heart disease with increased pulmonary blood flow using in vitro approaches. To this end, late gestation fetal lambs underwent in utero placement of an aortopulmonary graft (shunt). Four weeks after birth, LECs were isolated and cultured from control and shunt lambs. Redox status and proliferation were quantified, and transcriptional profiling and metabolomic analyses were performed. Shunt LECs exhibited hyperproliferative growth driven by increased levels of Hypoxia Inducible Factor 1α (HIF-1α), along with upregulated expression of known HIF-1α target genes in response to mechanical stimuli and shear stress. Compared to control LECs, shunt LECs exhibited abnormal metabolism including abnormalities of glycolysis, the TCA cycle and aerobic respiration. In conclusion, LECs from lambs exposed in vivo to chronically increased pulmonary lymph flow are hyperproliferative, have enhanced expression of HIF-1α and its target genes, and demonstrate altered central carbon metabolism in vitro. Importantly, these findings suggest provocative therapeutic targets for patients with lymphatic abnormalities.

Guanine- and cytosine-rich nucleic acid sequences have the potential to form secondary structures such as G-quadruplexes and i-motifs, respectively. We show that stabilization of G-quadruplexes using small molecules destabilizes the i-motifs, and vice versa, indicating these gene regulatory controllers are interdependent in human cells. This has important implications as these structures are predominately considered as isolated structural targets for therapy, but their interdependency highlights the interplay of both structures as an important gene regulatory switch.

The development of new therapeutic strategies against multidrug resistant Gram-negative bacteria is a major challenge for pharmaceutical research. In this respect, it is increasingly recognized that an efficient treatment for resistant bacterial infections should combine antimicrobial and anti-inflammatory effects. Here, we explore the multifunctional therapeutic potential of nanostructured self-assemblies from a cationic bolaamphiphile, which target bacterial lipopolysaccharides (LPSs) and associates with an anti-bacterial nucleic acid to form nanoplexes with therapeutic efficacy against Gram-negative bacteria. To understand the mechanistic details of these multifunctional antimicrobial-anti-inflammatory properties, we performed a fundamental study, comparing the interaction of these nanostructured therapeutics with synthetic biomimetic bacterial membranes and live bacterial cells. Combining a wide range of experimental techniques (Confocal Microscopy, Fluorescence Correlation Spectroscopy, Microfluidics, NMR, LPS binding assays), we demonstrate that the LPS targeting capacity of the bolaamphiphile self-assemblies, comparable to that exerted by Polymixin B, is a key feature of these nanoplexes and one that permits entry of therapeutic nucleic acids in Gram-negative bacteria. These findings enable a new approach to the design of efficient multifunctional therapeutics with combined antimicrobial and anti-inflammatory effects and have therefore the potential to broadly impact fundamental and applied research on self-assembled nano-sized antibacterials for antibiotic resistant infections., (Copyright © 2020 Elsevier B.V. All rights reserved.)