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The Kinase Chemogenomic Set (KCGS): An Open Science Resource for Kinase Vulnerability Identification.
We describe the assembly and annotation of a chemogenomic set of protein kinase inhibitors as an open science resource for studying kinase biology. The set only includes inhibitors that show potent kinase inhibition and a narrow spectrum of activity when screened across a large panel of kinase biochemical assays. Currently, the set contains 187 inhibitors that cover 215 human kinases. The kinase chemogenomic set (KCGS), current Version 1.0, is the most highly annotated set of selective kinase inhibitors available to researchers for use in cell-based screens.
Inhibition of the SUV4-20 H1 histone methyltransferase increases frataxin expression in Friedreich's ataxia patient cells.
The molecular mechanisms of reduced frataxin (<i>FXN</i>) expression in Friedreich's ataxia (FRDA) are linked to epigenetic modification of the <i>FXN</i> locus caused by the disease-associated GAA expansion. Here, we identify that SUV4-20 histone methyltransferases, specifically SUV4-20 H1, play an important role in the regulation of FXN expression and represent a novel therapeutic target. Using a human <i>FXN</i>-GAA-Luciferase repeat expansion genomic DNA reporter model of FRDA, we screened the Structural Genomics Consortium epigenetic probe collection. We found that pharmacological inhibition of the SUV4-20 methyltransferases by the tool compound A-196 increased the expression of FXN by ∼1.5-fold in the reporter cell line. In several FRDA cell lines and patient-derived primary peripheral blood mononuclear cells, A-196 increased FXN expression by up to 2-fold, an effect not seen in WT cells. SUV4-20 inhibition was accompanied by a reduction in H4K20me2 and H4K20me3 and an increase in H4K20me1, but only modest (1.4-7.8%) perturbation in genome-wide expression was observed. Finally, based on the structural activity relationship and crystal structure of A-196, novel small molecule A-196 analogs were synthesized and shown to give a 20-fold increase in potency for increasing <i>FXN</i> expression. Overall, our results suggest that histone methylation is important in the regulation of <i>FXN</i> expression and highlight SUV4-20 H1 as a potential novel therapeutic target for FRDA.
<jats:title>Abstract</jats:title> <jats:p>Muscle wasting and atrophy are regulated by multiple molecular processes, including mRNA processing. Reduced levels of the polyadenylation binding protein nucleus 1 (PABPN1), a multifactorial regulator of mRNA processing, cause muscle atrophy. A proteomic study in muscles with reduced PABPN1 levels suggested dysregulation of sarcomeric and cytoskeletal proteins. Here we investigated the hypothesis that reduced PABPN1 levels lead to an aberrant organization of the cytoskeleton. MURC, a plasma membrane-associated protein, was found to be more abundant in muscles with reduced PABPN1 levels, and it was found to be expressed at regions showing regeneration. A polarized cytoskeletal organization is typical for muscle cells, but muscle cells with reduced PABPN1 levels (named as shPAB) were characterized by a disorganized cytoskeleton that lacked polarization. Moreover, cell mechanical features and myogenic differentiation were significantly reduced in shPAB cells. Importantly, restoring cytoskeletal stability, by actin overexpression, was beneficial for myogenesis, expression of sarcomeric proteins and proper localization of MURC in shPAB cell cultures and in shPAB muscle bundle. We suggest that poor cytoskeletal mechanical features are caused by altered expression levels of cytoskeletal proteins and contribute to muscle wasting and atrophy.</jats:p>
Dual role of ER stress in response to metabolic co-targeting and radiosensitivity in head and neck cancer cells
<jats:title>Abstract</jats:title><jats:p>Arginine deprivation therapy (ADT) is a new metabolic targeting approach with high therapeutic potential for various solid cancers. Combination of ADT with low doses of the natural arginine analog canavanine effectively sensitizes malignant cells to irradiation. However, the molecular mechanisms determining the sensitivity of intrinsically non-auxotrophic cancers to arginine deficiency are still poorly understood. We here show for the first time that arginine deficiency is accompanied by global metabolic changes and protein/membrane breakdown, and results in the induction of specific, more or less pronounced (severe vs. mild) ER stress responses in head and neck squamous cell carcinoma (HNSCC) cells that differ in their intrinsic ADT sensitivity. Combination of ADT with canavanine triggered catastrophic ER stress via the eIF2α-ATF4(GADD34)-CHOP pathway, thereby inducing apoptosis; the same signaling arm was irrelevant in ADT-related radiosensitization. The particular strong supra-additive effect of ADT, canavanine and irradiation in both intrinsically more and less sensitive cancer cells supports the rational of ER stress pathways as novel target for improving multi-modal metabolic anti-cancer therapy.</jats:p>
The duration, dynamics and determinants of SARS-CoV-2 antibody responses in individual healthcare workers.
<h4>Background</h4>SARS-CoV-2 IgG antibody measurements can be used to estimate the proportion of a population exposed or infected and may be informative about the risk of future infection. Previous estimates of the duration of antibody responses vary.<h4>Methods</h4>We present 6 months of data from a longitudinal seroprevalence study of 3276 UK healthcare workers (HCWs). Serial measurements of SARS-CoV-2 anti-nucleocapsid and anti-spike IgG were obtained. Interval censored survival analysis was used to investigate the duration of detectable responses. Additionally, Bayesian mixed linear models were used to investigate anti-nucleocapsid waning.<h4>Results</h4>Anti-spike IgG levels remained stably detected after a positive result, e.g., in 94% (95% credibility interval, CrI, 91-96%) of HCWs at 180 days. Anti-nucleocapsid IgG levels rose to a peak at 24 (95% credibility interval, CrI 19-31) days post first PCR-positive test, before beginning to fall. Considering 452 anti-nucleocapsid seropositive HCWs over a median of 121 days from their maximum positive IgG titre, the mean estimated antibody half-life was 85 (95%CrI, 81-90) days. Higher maximum observed anti-nucleocapsid titres were associated with longer estimated antibody half-lives. Increasing age, Asian ethnicity and prior self-reported symptoms were independently associated with higher maximum anti-nucleocapsid levels and increasing age and a positive PCR test undertaken for symptoms with longer anti-nucleocapsid half-lives.<h4>Conclusion</h4>SARS-CoV-2 anti-nucleocapsid antibodies wane within months, and faster in younger adults and those without symptoms. However, anti-spike IgG remains stably detected. Ongoing longitudinal studies are required to track the long-term duration of antibody levels and their association with immunity to SARS-CoV-2 reinfection.
Community prevalence of SARS-CoV-2 in England from April to November, 2020: results from the ONS Coronavirus Infection Survey.
<h4>Background</h4>Decisions about the continued need for control measures to contain the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) rely on accurate and up-to-date information about the number of people testing positive for SARS-CoV-2 and risk factors for testing positive. Existing surveillance systems are generally not based on population samples and are not longitudinal in design.<h4>Methods</h4>Samples were collected from individuals aged 2 years and older living in private households in England that were randomly selected from address lists and previous Office for National Statistics surveys in repeated cross-sectional household surveys with additional serial sampling and longitudinal follow-up. Participants completed a questionnaire and did nose and throat self-swabs. The percentage of individuals testing positive for SARS-CoV-2 RNA was estimated over time by use of dynamic multilevel regression and poststratification, to account for potential residual non-representativeness. Potential changes in risk factors for testing positive over time were also assessed. The study is registered with the ISRCTN Registry, ISRCTN21086382.<h4>Findings</h4>Between April 26 and Nov 1, 2020, results were available from 1 191 170 samples from 280 327 individuals; 5231 samples were positive overall, from 3923 individuals. The percentage of people testing positive for SARS-CoV-2 changed substantially over time, with an initial decrease between April 26 and June 28, 2020, from 0·40% (95% credible interval 0·29-0·54) to 0·06% (0·04-0·07), followed by low levels during July and August, 2020, before substantial increases at the end of August, 2020, with percentages testing positive above 1% from the end of October, 2020. Having a patient-facing role and working outside your home were important risk factors for testing positive for SARS-CoV-2 at the end of the first wave (April 26 to June 28, 2020), but not in the second wave (from the end of August to Nov 1, 2020). Age (young adults, particularly those aged 17-24 years) was an important initial driver of increased positivity rates in the second wave. For example, the estimated percentage of individuals testing positive was more than six times higher in those aged 17-24 years than in those aged 70 years or older at the end of September, 2020. A substantial proportion of infections were in individuals not reporting symptoms around their positive test (45-68%, dependent on calendar time.<h4>Interpretation</h4>Important risk factors for testing positive for SARS-CoV-2 varied substantially between the part of the first wave that was captured by the study (April to June, 2020) and the first part of the second wave of increased positivity rates (end of August to Nov 1, 2020), and a substantial proportion of infections were in individuals not reporting symptoms, indicating that continued monitoring for SARS-CoV-2 in the community will be important for managing the COVID-19 pandemic moving forwards.<h4>Funding</h4>Department of Health and Social Care.
<h4>Purpose</h4>The Phase 3 LIBERTY ASTHMA QUEST study in patients aged ≥12 years with uncontrolled, moderate-to-severe asthma demonstrated the efficacy and safety of dupilumab 200 mg and 300 mg every 2 weeks (q2w) vs matched placebo in the overall population. This post hoc analysis assessed dupilumab efficacy by disease severity as evidenced by baseline % predicted forced expiratory volume in 1 second (FEV<sub>1</sub>) and dose of inhaled corticosteroids (ICS).<h4>Patients and methods</h4>Severe asthma exacerbation rates, change from baseline in FEV<sub>1</sub>, asthma control, quality of life, and fractional exhaled nitric oxide (FeNO) levels over the 52-week treatment period were assessed in patients with elevated type 2 inflammation biomarkers stratified by ICS dose and FEV<sub>1</sub>% predicted at baseline.<h4>Results</h4>In patients with elevated baseline eosinophils, dupilumab 200 mg and 300 mg q2w vs placebo reduced severe exacerbation rates by 50% (<i>P</i>=0.06) and 67% (<i>P</i>=0.001), respectively, in those with medium-dose ICS/FEV<sub>1</sub>% predicted 60-90%, and by 59% (<i>P</i><0.001) and 47% (<i>P</i>=0.006) in those with high-dose ICS/FEV<sub>1</sub>% predicted <60%, improved pre-bronchodilator FEV<sub>1</sub> at Week 12 by 0.16L (<i>P</i>=0.005) and 0.08L (<i>P</i>=0.13), and by 0.20L (<i>P</i>=0.003) and 0.21L (<i>P</i><0.001), respectively, in the same subgroups. Dupilumab vs placebo also improved asthma control and quality of life and suppressed FeNO levels in all patient subgroups with similar results observed irrespective of baseline biomarker status or disease severity.<h4>Conclusion</h4>Dupilumab reduced severe exacerbations and improved lung function, asthma control and quality of life in patients with elevated baseline eosinophils irrespective of baseline ICS dose or FEV<sub>1</sub>% predicted.
Mycobacterium tuberculosis (M.tb) is responsible for more deaths globally than any other pathogen. The only available vaccine, bacillus Calmette-Guérin (BCG), has variable efficacy throughout the world. A more effective vaccine is urgently needed. The immune response against tuberculosis relies, at least in part, on CD4<sup>+</sup> T cells. Protective vaccines require the induction of antigen-specific CD4<sup>+</sup> T cells via mycobacterial peptides presented by MHC class-II in infected macrophages. In order to identify mycobacterial antigens bound to MHC, we have immunoprecipitated MHC class-I and class-II complexes from THP-1 macrophages infected with BCG, purified MHC class-I and MHC class-II peptides and analysed them by liquid chromatography tandem mass spectrometry. We have successfully identified 94 mycobacterial peptides presented by MHC-II and 43 presented by MHC-I, from 76 and 41 antigens, respectively. These antigens were found to be highly expressed in infected macrophages. Gene ontology analysis suggests most of these antigens are associated with membranes and involved in lipid biosynthesis and transport. The sequences of selected peptides were confirmed by spectral match validation and immunogenicity evaluated by IFN-gamma ELISpot against peripheral blood mononuclear cell from volunteers vaccinated with BCG, M.tb latently infected subjects or patients with tuberculosis disease. Three antigens were expressed in viral vectors, and evaluated as vaccine candidates alone or in combination in a murine aerosol M.tb challenge model. When delivered in combination, the three candidate vaccines conferred significant protection in the lungs and spleen compared with BCG alone, demonstrating proof-of-concept for this unbiased approach to identifying new candidate antigens.
Sequence and structural variations determining the recruitment of WNK kinases to the KLHL3 E3 ligase
<jats:title>Abstract</jats:title><jats:p>The BTB-Kelch protein KLHL3 is a Cullin3-dependent E3 ligase that mediates the ubiquitin-dependent degradation of kinases WNK1-4 to control blood pressure and cell volume. A crystal structure of KLHL3 has defined its binding to an acidic degron motif containing a PXXP sequence that is strictly conserved in WNK1, WNK2 and WNK4. Mutations in the second proline abrograte the interaction causing the hypertension syndrome pseudohypoaldosteronism type II. WNK3 shows a diverged degron motif containing 4 amino acid substitutions that remove the PXXP motif raising questions as to the mechanism of its binding. To understand this atypical interaction, we determined the crystal structure of the KLHL3 Kelch domain in complex with a WNK3 peptide. The electron density enabled the complete 11-mer WNK-family degron motif to be traced for the first time revealing several conserved features not captured in previous work, including additional salt bridge and hydrogen bond interactions. Overall, the WNK3 peptide adopted a conserved binding pose except for a subtle shift to accommodate bulkier amino acid substitutions at the binding interface. At the centre, the second proline was substituted by WNK3 Thr541, providing a unique phosphorylatable residue among the WNK-family degrons. Fluorescence polarisation and structural modelling experiments revealed that its phosphorylation would abrogate the KLHL3 interaction similarly to hypertension-causing mutations. Together, these data reveal how the KLHL3 Kelch domain can accommodate the binding of multiple WNK isoforms and highlight a potential regulatory mechanism for the recruitment of WNK3.</jats:p>
<jats:p>Ubiquitin is a versatile post-translational modification which is covalently attached to protein targets either as a single moiety or as a ubiquitin chain. In contrast to K48 and K63-linked chains which have been extensively studied, the regulation and function of most atypical ubiquitin chains is only starting to emerge. The deubiquitinase TRABID/ZRANB1 is tuned for the recognition and cleavage of K29 and K33-linked chains. Yet, substrates of TRABID and the cellular functions of these atypical ubiquitin signals remain unclear. We determined the interactome of two TRABID constructs rendered catalytic dead either through a point mutation in the catalytic cysteine residue or through removal of the OTU catalytic domain. We identified 50 proteins trapped by both constructs and which therefore represent candidate substrates of TRABID. We then validated the E3 ubiquitin ligase HECTD1 as a substrate of TRABID and used UbiCREST and Ub-AQUA proteomics to show that HECTD1 preferentially assembles K29- and K48-linked ubiquitin chains. Further in vitro autoubiquitination assays using ubiquitin mutants established that while HECTD1 can assemble short homotypic K29 and K48-linked chains, it requires branching at K29/K48 in order to achieve its full ubiquitin ligase activity. We next used transient knockdown and genetic knock out of TRABID in mammalian cells in order to determine the functional relationship between TRABID and HECTD1. This revealed that upon TRABID depletion, HECTD1 is readily degraded. Thus, this study identifies HECTD1 as a mammalian E3 ligase which assembles branched K29/K48 chains and also establishes TRABID-HECTD1 as a DUB/E3 pair regulating K29 linkages.</jats:p>
Safety and immunogenicity of ChAdOx1 nCoV-19 vaccine administered in a prime-boost regimen in young and old adults (COV002): a single-blind, randomised, controlled, phase 2/3 trial.
BACKGROUND:Older adults (aged ≥70 years) are at increased risk of severe disease and death if they develop COVID-19 and are therefore a priority for immunisation should an efficacious vaccine be developed. Immunogenicity of vaccines is often worse in older adults as a result of immunosenescence. We have reported the immunogenicity of a novel chimpanzee adenovirus-vectored vaccine, ChAdOx1 nCoV-19, in young adults, and now describe the safety and immunogenicity of this vaccine in a wider range of participants, including adults aged 70 years and older. METHODS:In this report of the phase 2 component of a single-blind, randomised, controlled, phase 2/3 trial (COV002), healthy adults aged 18 years and older were enrolled at two UK clinical research facilities, in an age-escalation manner, into 18-55 years, 56-69 years, and 70 years and older immunogenicity subgroups. Participants were eligible if they did not have severe or uncontrolled medical comorbidities or a high frailty score (if aged ≥65 years). First, participants were recruited to a low-dose cohort, and within each age group, participants were randomly assigned to receive either intramuscular ChAdOx1 nCoV-19 (2·2 × 1010 virus particles) or a control vaccine, MenACWY, using block randomisation and stratified by age and dose group and study site, using the following ratios: in the 18-55 years group, 1:1 to either two doses of ChAdOx1 nCoV-19 or two doses of MenACWY; in the 56-69 years group, 3:1:3:1 to one dose of ChAdOx1 nCoV-19, one dose of MenACWY, two doses of ChAdOx1 nCoV-19, or two doses of MenACWY; and in the 70 years and older, 5:1:5:1 to one dose of ChAdOx1 nCoV-19, one dose of MenACWY, two doses of ChAdOx1 nCoV-19, or two doses of MenACWY. Prime-booster regimens were given 28 days apart. Participants were then recruited to the standard-dose cohort (3·5-6·5 × 1010 virus particles of ChAdOx1 nCoV-19) and the same randomisation procedures were followed, except the 18-55 years group was assigned in a 5:1 ratio to two doses of ChAdOx1 nCoV-19 or two doses of MenACWY. Participants and investigators, but not staff administering the vaccine, were masked to vaccine allocation. The specific objectives of this report were to assess the safety and humoral and cellular immunogenicity of a single-dose and two-dose schedule in adults older than 55 years. Humoral responses at baseline and after each vaccination until 1 year after the booster were assessed using an in-house standardised ELISA, a multiplex immunoassay, and a live severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) microneutralisation assay (MNA80). Cellular responses were assessed using an ex-vivo IFN-γ enzyme-linked immunospot assay. The coprimary outcomes of the trial were efficacy, as measured by the number of cases of symptomatic, virologically confirmed COVID-19, and safety, as measured by the occurrence of serious adverse events. Analyses were by group allocation in participants who received the vaccine. Here, we report the preliminary findings on safety, reactogenicity, and cellular and humoral immune responses. This study is ongoing and is registered with ClinicalTrials.gov, NCT04400838, and ISRCTN, 15281137. FINDINGS:Between May 30 and Aug 8, 2020, 560 participants were enrolled: 160 aged 18-55 years (100 assigned to ChAdOx1 nCoV-19, 60 assigned to MenACWY), 160 aged 56-69 years (120 assigned to ChAdOx1 nCoV-19: 40 assigned to MenACWY), and 240 aged 70 years and older (200 assigned to ChAdOx1 nCoV-19: 40 assigned to MenACWY). Seven participants did not receive the boost dose of their assigned two-dose regimen, one participant received the incorrect vaccine, and three were excluded from immunogenicity analyses due to incorrectly labelled samples. 280 (50%) of 552 analysable participants were female. Local and systemic reactions were more common in participants given ChAdOx1 nCoV-19 than in those given the control vaccine, and similar in nature to those previously reported (injection-site pain, feeling feverish, muscle ache, headache), but were less common in older adults (aged ≥56 years) than younger adults. In those receiving two standard doses of ChAdOx1 nCoV-19, after the prime vaccination local reactions were reported in 43 (88%) of 49 participants in the 18-55 years group, 22 (73%) of 30 in the 56-69 years group, and 30 (61%) of 49 in the 70 years and older group, and systemic reactions in 42 (86%) participants in the 18-55 years group, 23 (77%) in the 56-69 years group, and 32 (65%) in the 70 years and older group. As of Oct 26, 2020, 13 serious adverse events occurred during the study period, none of which were considered to be related to either study vaccine. In participants who received two doses of vaccine, median anti-spike SARS-CoV-2 IgG responses 28 days after the boost dose were similar across the three age cohorts (standard-dose groups: 18-55 years, 20 713 arbitrary units [AU]/mL [IQR 13 898-33 550], n=39; 56-69 years, 16 170 AU/mL [10 233-40 353], n=26; and ≥70 years 17 561 AU/mL [9705-37 796], n=47; p=0·68). Neutralising antibody titres after a boost dose were similar across all age groups (median MNA80 at day 42 in the standard-dose groups: 18-55 years, 193 [IQR 113-238], n=39; 56-69 years, 144 [119-347], n=20; and ≥70 years, 161 [73-323], n=47; p=0·40). By 14 days after the boost dose, 208 (>99%) of 209 boosted participants had neutralising antibody responses. T-cell responses peaked at day 14 after a single standard dose of ChAdOx1 nCoV-19 (18-55 years: median 1187 spot-forming cells [SFCs] per million peripheral blood mononuclear cells [IQR 841-2428], n=24; 56-69 years: 797 SFCs [383-1817], n=29; and ≥70 years: 977 SFCs [458-1914], n=48). INTERPRETATION:ChAdOx1 nCoV-19 appears to be better tolerated in older adults than in younger adults and has similar immunogenicity across all age groups after a boost dose. Further assessment of the efficacy of this vaccine is warranted in all age groups and individuals with comorbidities. FUNDING:UK Research and Innovation, National Institutes for Health Research (NIHR), Coalition for Epidemic Preparedness Innovations, NIHR Oxford Biomedical Research Centre, Thames Valley and South Midlands NIHR Clinical Research Network, and AstraZeneca.
Glycosylated Siglec-6 expression in syncytiotrophoblast-derived extracellular vesicles from preeclampsia placentas.
INTRODUCTION:Preeclampsia (PE) is associated with an exaggerated maternal systemic inflammatory response. Throughout gestation, the placenta releases extracellular vesicles through the syncytiotrophoblast layer (STB) into the maternal circulation and this is increased in PE. Expression of Siglec-6, a transmembrane receptor of molecular weight 50 KDa, is upregulated in PE placental tissue. METHODS:Here we investigated respective abundance of Siglec-6 in PE -and normal pregnancy- (NP) derived placental lysates (PL) and syncytiotrophoblast-derived extracellular vesicles (STBEV). STBEV from PE and NP placentas were isolated through dual-lobe placental perfusion and serial ultracentrifugation. Siglec-6 was characterized by immunohistochemistry, immunoblotting, mass spectrometry (MS), and deglycosylation. RESULTS:Immunoblotting revealed the expected Siglec-6 (50 KDa) band present in both PE and NP PL, however an additional heavier band was observed at 70 KDa only in PE PL, but not in NP. When interrogating STBEV we saw an absence of the expected 50 KDa band but the 70 KDa was present predominantly only in the PE STBEV. Deglycosylation of PL and STBEV from PE showed that the 70 KDa and the 50 KDa bands were reduced to 48 KDa, suggesting glycosylation. Both 48 KDa and 70 KDa bands were subjected to MS, confirming Siglec-6 expression in both. DISCUSSION:Our data shows that the inability to detect Siglec-6 in circulation might be due to the placenta secreting STBEV carrying a modified glycosylated form of Siglec-6 with a 70 KDa molecular weight, significantly and uniquely upregulated in PE STBEV.
Dynamics of Transforming Growth Factor (TGF)-β Superfamily Cytokine Induction During HIV-1 Infection Are Distinct From Other Innate Cytokines.
Human immunodeficiency virus type 1 (HIV-1) infection triggers rapid induction of multiple innate cytokines including type I interferons, which play important roles in viral control and disease pathogenesis. The transforming growth factor (TGF)-β superfamily is a pleiotropic innate cytokine family, some members of which (activins and bone morphogenetic proteins (BMPs)) were recently demonstrated to exert antiviral activity against Zika and hepatitis B and C viruses but are poorly studied in HIV-1 infection. Here, we show that TGF-β1 is systemically induced with very rapid kinetics (as early as 1-4 days after viremic spread begins) in acute HIV-1 infection, likely due to release from platelets, and remains upregulated throughout infection. Contrastingly, no substantial systemic upregulation of activins A and B or BMP-2 was observed during acute infection, although plasma activin levels trended to be elevated during chronic infection. HIV-1 triggered production of type I interferons but not TGF-β superfamily cytokines from plasmacytoid dendritic cells (DCs) in vitro, putatively explaining their differing in vivo induction; whilst lipopolysaccharide (but not HIV-1) elicited activin A production from myeloid DCs. These findings underscore the need for better definition of the protective and pathogenic capacity of TGF-β superfamily cytokines, to enable appropriate modulation for therapeutic purposes.
A clinical study was designed to record a wide range of physiological values from patients undergoing haemodialysis treatment in the Renal Unit of the Churchill Hospital in Oxford. Video was recorded for a total of 84 dialysis sessions from 40 patients during the course of 1 year, comprising an overall video recording time of approximately 304.1 h. Reference values were provided by two devices in regular clinical use. The mean absolute error between the heart rate estimates from the camera and the average from two reference pulse oximeters (positioned at the finger and earlobe) was 2.8 beats/min for over 65% of the time the patient was stable. The mean absolute error between the respiratory rate estimates from the camera and the reference values (computed from the Electrocardiogram and a thoracic expansion sensor-chest belt) was 2.1 breaths/min for over 69% of the time for which the reference signals were valid. To increase the robustness of the algorithms, novel methods were devised for cancelling out aliased frequency components caused by the artificial light sources in the hospital, using auto-regressive modelling and pole cancellation. Maps of the spatial distribution of heart rate and respiratory rate information were developed from the coefficients of the auto-regressive models. Most of the periods for which the camera could not produce a reliable heart rate estimate lasted under 3 min, thus opening the possibility to monitor heart rate continuously in a clinical environment.
Patient experiences of as-needed budesonide-formoterol by Turbuhaler® for treatment of mild asthma; a qualitative study.
BACKGROUND: Combination low-dose budesonide-formoterol, taken as-needed for symptom relief reduces exacerbation risk and is recommended for treatment of mild asthma. The NovelQ qualitative study explored patients' attitudes toward using this novel therapy. METHODS: Adults with mild asthma using reliever-only treatment were randomised to as-needed budesonide-formoterol Turbuhaler® in a multinational, 52-week open-label randomised controlled trial (NovelSTART-ACTRN12615000999538). A subgroup were interviewed to explore their attitudes to use of as-needed budesonide-formoterol after receiving it for ≥10 months. Semi-structured interviews were conducted until saturation, audio-recorded, and thematically analysed. RESULTS: Analysis of 35 participants (66% female; mean age 43.5 [range 18-74]; mean Asthma Control Questionnaire score 1.09 ± SD0.55) interviews identified 5 themes, each including both barriers and facilitators to therapy use. Themes were: 'Treatment effectiveness' i.e. how well symptoms were relieved and/or prevented; 'Lifestyle fit of the regimen' e.g. the extent to which the treatment regimen integrated into the patient's daily life; 'Attitudes toward medication use and safety' e.g. openness for new reliever treatments, beliefs about treatment necessity or side effects; 'Device attributes' e.g. perceived ease of use; and 'Doctor-patient relationship' e.g. impact of health professional support on new treatment acceptance. CONCLUSIONS: A wide range of factors seem to drive the opinions of mild asthma patients on as-needed budesonide-formoterol therapy. Many patients perceived both positive and negative treatment attributes, and their individual evaluation of these attributes determined their likelihood of using it after the study. Supportive patient-physician interactions appear key to addressing patient barriers. Recommendations for patient-centred discussions, developed from this research, are provided.