Identification of copper as a cell culture media component causing metabolite depletion and product sequence variants

The level of peptide sequence variants in a biologic drug substance batch is a critical product quality attribute that should be monitored and controlled. These sequence variants are typically caused by DNA single nucleotide variants that arise in cloning and amplification, mistranscription due to unstable vector DNA or cell age/production stresses, or mistranslation via tRNA wobble or mischarging. In this work, a low frequency of monoclonal antibody sequence variants was detected by mass spectrometry in a drug substance batch. The variants were distributed throughout the heavy and light chains at average levels of under 1% per site with no apparent codon bias. No product-coding DNA mutations were detected via deep sequencing data. This pattern of low level, widely-distributed variation strongly suggested a misincorporation mechanism via mischarging of aminoacyl-tRNA, presumably due to amino acid depletion during the process. Copper is a critical cell culture media component that can be modulated in fed-batch processes to induce lactate consumption via its role as a cofactor for mitochondrial function and respiration. However, complete consumption of lactate can also trigger reduced levels of other metabolites required for recombinant protein assembly, which can lead to product sequence variants. To investigate the potential relationship between media copper supplementation and sequence variants, various levels of copper were supplemented into the basal media for fed-batch cultures at the 250 mL bioreactor scale. Mass spectrometry analysis of the partially purified antibody indicated a positive correlation between the amount of copper supplemented and the level of detected sequence variants as well as a mechanism for sequence variant reduction via targeted nutrient feeding. This work has identified a potential mechanism of sequence variant generation related to cell culture media copper levels as well as process alterations to prevent such variation in future batches, highlighting the importance of carefully controlling trace metal levels. Additional studies may be required to validate the potential mechanism

Decarbonising off grid homes: A rapid evidence review

Currently, there are circa 2,000 properties across the UK that are ‘off-grid’, meaning they are without access to both electricity and gas mains / grid connections. A further 4.3 million UK domestic properties are off the mains gas grid (BEIS, 2022d). A range of socio-economic challenges are faced by people living ‘off-grid’, including being at greater risk of fuel poverty, and struggling to adequately heat their homes. This evidence review forms part of a wider research project about the realities and practicalities of decarbonising off grid properties. It aims to understand what is currently known about approaches to decarbonising (or improving the energy performance of) homes that are completely or partially off grid

Fatal breakthrough mucormycosis in a multivisceral transplant patient receiving micafungin: Case report and literature review.

INTRODUCTION: Antifungal agents are routinely used in the post-transplant setting for both prophylaxis and treatment of presumed and proven fungal infections. Micafungin is an echinocandin-class antifungal with broad antifungal cover and favorable side effect profile but, notably, it has no activity against molds of the order Mucorales. PRESENTATION OF CASE: A 47-year-old woman underwent multivisceral transplantation for intestinal failure-associated liver disease. She had a prolonged post-operative recovery complicated by invasive candidiasis and developed an intolerance to liposomal amphotericin B. In view of her immunosuppression, she was commenced on micafungin as prophylaxis to prevent invasive fungal infection. However, she developed acute graft versus host disease with bone marrow failure complicated by disseminated mucormycosis which was only diagnosed post mortem. DISCUSSION: Non-Aspergillus breakthrough mold infections with micafungin therapy are rare with only eight other cases having been described in the literature. Breakthrough infections have occurred within one week of starting micafungin. Diagnosis is problematic and requires a high degree of clinical suspicion and microscopic/histological examination of an involved site. The management of these aggressive infections involves extensive debridement and appropriate antifungal cover. CONCLUSION: A high level of suspicion of invasive fungal infection is required at all times in immunosuppressed patients, even those receiving antifungal prophylaxis. Early biopsy is required. Even with early recognition and aggressive treatment of these infections, prognosis is poor

Early management of acute severe UC in the biologics era: development and international validation of a prognostic clinical index to predict steroid response

Objectives We aimed to determine whether changes in acute severe colitis (ASC) management have translated to improved outcomes and to develop a simple model predicting steroid non-response on admission. Design Outcomes of 131 adult ASC admissions (117 patients) in Oxford, UK between 2015 and 2019 were compared with data from 1992 to 1993. All patients received standard treatment with intravenous corticosteroids and endoscopic disease activity scoring (Ulcerative Colitis Endoscopic Index of Severity (UCEIS)). Steroid non-response was defined as receiving medical rescue therapy or surgery. A predictive model developed in the Oxford cohort was validated in Australia and India (Gold Coast University Hospital 2015–2020, n=110; All India Institute of Medical Sciences, New Delhi 2018–2020, n=62). Results In the 2015–2019 Oxford cohort, 15% required colectomy during admission vs 29% in 1992–1993 (p=0.033), while 71 (54%) patients received medical rescue therapy (27% ciclosporin, 27% anti-tumour necrosis factor, compared with 27% ciclosporin in 1992–1993 (p=0.0015). Admission C reactive protein (CRP) (false discovery rate, p=0.00066), albumin (0.0066) and UCEIS scores (0.015) predicted steroid non-response. A four-point model was developed involving CRP of ≥100 mg/L (one point), albumin of ≤25 g/L (one point), and UCEIS score of ≥4 (1 point) or ≥7 (2 points). Patients scoring 0, 1, 2, 3 and 4 in the validation cohorts had steroid response rates of 100, 75.0%, 54.9%, 18.2% and 0%, respectively. Scoring of ≥3 was 84% (95% CI 0.70 to 0.98) predictive of steroid failure (OR 11.9, 95% CI 10.8 to 13.0). Colectomy rates in the validation cohorts were were 8%–11%. Conclusions Emergency colectomy rates for ASC have halved in 25 years to 8%–15% worldwide. Patients who will not respond to corticosteroids are readily identified on admission and may be prioritised for early intensification of therapy

The motion of trees in the wind : a data synthesis

Interactions between wind and trees control energy exchanges between the atmosphere and forest canopies. This energy exchange can lead to the widespread damage of trees, and wind is a key disturbance agent in many of the world’s forests. However, most research on this topic has focused on conifer plantations, where risk management is economically important, rather than broadleaf forests, which dominate the forest carbon cycle. This study brings together tree motion time-series data to systematically evaluate the factors influencing tree responses to wind loading, including data from both broadleaf and coniferous trees in forests and open environments. Wefoundthatthetwomostdescriptive features of tree motion were (a) the fundamental frequency, which is a measure of the speed at which a tree sways and is strongly related to tree height, and (b) the slope of the power spectrum, which is related to the efficiency of energy transfer from wind to trees. Intriguingly, the slope of the power spectrum was found to remain constant from medium to high wind speeds for all trees in this study. This suggests that, contrary to some predictions, damping or amplification mechanisms do not change dramatically at high wind speeds, and therefore wind damage risk is related, relatively simply, to wind speed. Conifers from forests were distinct from broadleaves in terms of their response to wind loading. Specifically, the fundamental frequency of forest conifers was related to their size according to the cantilever beam model (i.e. vertically distributed mass), whereas broadleaves were better approximated by the simple pendulum model (i.e. dominated by the crown). Forest conifers also had a steeper slope of the power spectrum. We interpret these finding as being strongly related to tree architecture; i.e. conifers generally have a simple shape due to their apical dominance, whereas broadleaves exhibit a much wider range of architectures with more dominant crowns

Management of patients with biliary sphincter of Oddi disorder without sphincter of Oddi manometry

<p>Abstract</p> <p>Background</p> <p>The paucity of controlled data for the treatment of most biliary sphincter of Oddi disorder (SOD) types and the incomplete response to therapy seen in clinical practice and several trials has generated controversy as to the best course of management of these patients. In this observational study we aimed to assess the outcome of patients with biliary SOD managed without sphincter of Oddi manometry.</p> <p>Methods</p> <p>Fifty-nine patients with biliary SOD (14% type I, 51% type II, 35% type III) were prospectively enrolled. All patients with a dilated common bile duct were offered endoscopic retrograde cholangiopancreatography (ERCP) and sphincterotomy whereas all others were offered medical treatment alone. Patients were followed up for a median of 15 months and were assessed clinically for response to treatment.</p> <p>Results</p> <p>At follow-up 15.3% of patients reported complete symptom resolution, 59.3% improvement, 22% unchanged symptoms, and 3.4% deterioration. Fifty-one percent experienced symptom resolution/improvement on medical treatment only, 12% after sphincterotomy, and 10% after both medical treatment/sphincterotomy. Twenty percent experienced at least one recurrence of symptoms after initial response to medical and/or endoscopic treatment. Fifty ERCP procedures were performed in 24 patients with an 18% complication rate (16% post-ERCP pancreatitis). The majority of complications occurred in the first ERCP these patients had. Most complications were mild and treated conservatively. Age, gender, comorbidity, SOD type, dilated common bile duct, presence of intact gallbladder, or opiate use were not related to the effect of treatment at the end of follow-up (p > 0.05 for all).</p> <p>Conclusions</p> <p>Patients with biliary SOD may be managed with a combination of endoscopic sphincterotomy (performed in those with dilated common bile duct) and medical therapy without manometry. The results of this approach with regards to symptomatic relief and ERCP complication rate are comparable to those previously published in the literature in cohorts of patients assessed by manometry.</p

Eye Movements Affect Postural Control in Young and Older Females

Visual information is used for postural stabilization in humans. However, little is known about how eye movements prevalent in everyday life interact with the postural control system in older individuals. Therefore, the present study assessed the effects of stationary gaze fixations, smooth pursuits, and saccadic eye movements, with combinations of absent, fixed and oscillating large-field visual backgrounds to generate different forms of retinal flow, on postural control in healthy young and older females. Participants were presented with computer generated visual stimuli, whilst postural sway and gaze fixations were simultaneously assessed with a force platform and eye tracking equipment, respectively. The results showed that fixed backgrounds and stationary gaze fixations attenuated postural sway. In contrast, oscillating backgrounds and smooth pursuits increased postural sway. There were no differences regarding saccades. There were also no differences in postural sway or gaze errors between age groups in any visual condition. The stabilizing effect of the fixed visual stimuli show how retinal flow and extraocular factors guide postural adjustments. The destabilizing effect of oscillating visual backgrounds and smooth pursuits may be related to more challenging conditions for determining body shifts from retinal flow, and more complex extraocular signals, respectively. Because the older participants matched the young group's performance in all conditions, decreases of posture and gaze control during stance may not be a direct consequence of healthy aging. Further research examining extraocular and retinal mechanisms of balance control and the effects of eye movements, during locomotion, is needed to better inform fall prevention interventions

Patient-reported Outcomes: the ICHOM Standard Set for Inflammatory Bowel Disease in Real-life Practice Helps Quantify Deficits in Current Care

BackgroundPatient reported outcome measures (PROMs) are key to documenting outcomes that matter most to patients and are increasingly important to commissioners of healthcare seeking value. We report the first series of the ICHOM Standard Set for Inflammatory Bowel Disease (IBD).MethodsPatients treated for ulcerative colitis (UC) or Crohn's disease (CD) in our centre were offered enrolment into the web-based TrueColours-IBD programme. Through this programme, email prompts linking to validated questionnaires were sent for symptoms, quality of life, and ICHOM IBD outcomes.ResultsThe first 1,299 consecutive patients enrolled (779 UC, 520 CD) were studied with median 270 days of follow up (IQR 116-504). 671 (52%) were female, mean age 42 years (sd 16), mean BMI 26 (sd 5.3). 483 (37%) were using advanced therapies at registration. Median adherence to fortnightly quality of life reporting and quarterly outcomes was 100% [IQR48-100%] and 100% [IQR75-100%], respectively. In the previous 12 months, prednisolone use was reported by 229 (29%) patients with UC vs. 81 (16%) with CD, p3 months. 174 (13%) patients reported an IBD-related intervention and 80 (6%) reported an unplanned hospital admission. There were high rates of fatigue (50%) and mood disturbance (23%).ConclusionOutcomes reported by patients illustrate the scale of the therapeutic deficit in current care. Proof of principle is demonstrated that PROM data can be collected continuously with little burden on healthcare professionals. This may become a metric for quality improvement programmes, or to compare outcomes

The state of the Martian climate

60°N was +2.0°C, relative to the 1981–2010 average value (Fig. 5.1). This marks a new high for the record. The average annual surface air temperature (SAT) anomaly for 2016 for land stations north of starting in 1900, and is a significant increase over the previous highest value of +1.2°C, which was observed in 2007, 2011, and 2015. Average global annual temperatures also showed record values in 2015 and 2016. Currently, the Arctic is warming at more than twice the rate of lower latitudes

A qualitative exploration of the human resource policy implications of voluntary counselling and testing scale-up in Kenya: applying a model for policy analysis

Background: Kenya experienced rapid scale up of HIV testing and counselling services in government health services from 2001. We set out to examine the human resource policy implications of scaling up HIV testing and counselling in Kenya and to analyse the resultant policy against a recognised theoretical framework of health policy reform (policy analysis triangle). Methods: Qualitative methods were used to gain in-depth insights from policy makers who shaped scale up. This included 22 in-depth interviews with Voluntary Counselling and Testing (VCT) task force members, critical analysis of 53 sets of minutes and diary notes. We explore points of consensus and conflict amongst policymakers in Kenya and analyse this content to assess who favoured and resisted new policies, how scale up was achieved and the importance of the local context in which scale up occurred. Results: The scale up of VCT in Kenya had a number of human resource policy implications resulting from the introduction of lay counsellors and their authorisation to conduct rapid HIV testing using newly introduced rapid testing technologies. Our findings indicate that three key groups of actors were critical: laboratory professionals, counselling associations and the Ministry of Health. Strategic alliances between donors, NGOs and these three key groups underpinned the process. The process of reaching consensus required compromise and time commitment but was critical to a unified nationwide approach. Policies around quality assurance were integral in ensuring standardisation of content and approach. Conclusion: The introduction and scale up of new health service initiatives such as HIV voluntary counselling and testing necessitates changes to existing health systems and modification of entrenched interests around professional counselling and laboratory testing. Our methodological approach enabled exploration of complexities of scale up of HIV testing and counselling in Kenya. We argue that a better understanding of the diverse actors, the context and the process, is required to mitigate risks and maximise impact