Development of a Stable Virus-Like Particle Vaccine Formulation against Chikungunya Virus and Investigation of the Effects of Polyanions

Chikungunya virus (CHIKV) is an alphavirus that infects millions of people every year, especially in the developing world. The selective expression of recombinant CHIKV capsid and envelope proteins results in the formation of self-assembled virus-like particles (VLPs) that have been shown to protect nonhuman primates against infection from multiple strains of CHIKV. This study describes the characterization, excipient screening, and optimization of CHIKV VLP solution conditions towards the development of a stable parenteral formulation. The CHIKV VLPs were found to be poorly soluble at pH 6 and below. Circular dichroism, intrinsic fluorescence, and static and dynamic light scattering measurements were therefore performed at neutral pH, and results consistent with the formation of molten globule structures were observed at elevated temperatures. A library of GRAS excipients was screened for their ability to physically stabilize CHIKV VLPs using a high-throughput turbidity based assay. Sugars, sugar alcohols, and polyanions were identified as potential stabilizers and the concentrations and combinations of select excipients were optimized. The effects of polyanions were further studied, and while all polyanions tested stabilized CHIKV VLPs against aggregation, the effects of polyanions on conformational stability varied

Reducing the environmental impact of surgery on a global scale: systematic review and co-prioritization with healthcare workers in 132 countries

Abstract Background Healthcare cannot achieve net-zero carbon without addressing operating theatres. The aim of this study was to prioritize feasible interventions to reduce the environmental impact of operating theatres. Methods This study adopted a four-phase Delphi consensus co-prioritization methodology. In phase 1, a systematic review of published interventions and global consultation of perioperative healthcare professionals were used to longlist interventions. In phase 2, iterative thematic analysis consolidated comparable interventions into a shortlist. In phase 3, the shortlist was co-prioritized based on patient and clinician views on acceptability, feasibility, and safety. In phase 4, ranked lists of interventions were presented by their relevance to high-income countries and low–middle-income countries. Results In phase 1, 43 interventions were identified, which had low uptake in practice according to 3042 professionals globally. In phase 2, a shortlist of 15 intervention domains was generated. In phase 3, interventions were deemed acceptable for more than 90 per cent of patients except for reducing general anaesthesia (84 per cent) and re-sterilization of ‘single-use’ consumables (86 per cent). In phase 4, the top three shortlisted interventions for high-income countries were: introducing recycling; reducing use of anaesthetic gases; and appropriate clinical waste processing. In phase 4, the top three shortlisted interventions for low–middle-income countries were: introducing reusable surgical devices; reducing use of consumables; and reducing the use of general anaesthesia. Conclusion This is a step toward environmentally sustainable operating environments with actionable interventions applicable to both high– and low–middle–income countries

STRUCTURE AND FUNCTION OF NATIVE AND EDITED DYSTROPHIN RODS

The purpose of this study is to examine the biophysical properties of the rod region of the dystrophin protein. This is important due to the severity of the disease Duchenne Muscular Dystrophy, (DMD), which is associated with the malfunction of this protein. DMD is one of the most serious single gene genetic defects of man. This rod region consists of a number of repeat motifs called spectrin type repeats or STRs. The thermodynamical and biochemical stability analysis shows, which single motifs are unstable on their own and which ones become more stable when linked to their appropriate tandem neighbors. This knowledge will impact strategies to produce modified mini dystrophins for gene therapy. Exon skipping therapy is an emerging approach to treat such genetic diseases. This is done by the administration of modified antisense oligonucleotides, AONs, which can interfere with exon splicing process and eliminate certain exons from the mature transcript. Furthermore, the rod region has a number of ancillary functions, such as providing secondary binding sites for actin, neuronal NO synthetase and phospholipids, which may be adversely perturbed by the edits.Ph.D. in Biological, Chemical, and Physical Sciences, May 201

Exon edited dystrophin rods in the hinge 3 region.

International audienceWe have studied the properties of a panel of proteins engineered to be end-products of envisioned exon skipping therapy by antisense oligonucleotides, AONs, directed at exon 51 applied to relevant dystrophin defects causing Duchenne muscular dystrophy, DMD. Exon skipping therapy is a leading therapeutic strategy being investigated for the treatment of this devastating genetic disease. AONs targeting exon 51 have progressed furthest in human clinical trials. Exon 51 skipping is applicable to a variety of dystrophin defects found in different patients. Due to the differences in original defect, the end result of the therapy will be different in each case. An open question is whether these differences will produce significant differences in the dystrophin protein so edited. In this study we have identified differences in the stability, structure and lipid binding properties of these end-product proteins produced by exon 51 skipping repair

Metronidazole-Induced Encephalopathy in a Patient with End-Stage Liver Disease

Purpose. Metronidazole-induced encephalopathy (MIE) has been rarely reported. We report a case in a patient with end-stage liver disease (ESLD). Summary. A 63-year-old male with ESLD secondary to hepatitis C virus presented with progressively worsening fatigue, slurred speech, aphasia, vomiting, and left-sided facial droop after completing a 2-week course of metronidazole for recurrent Clostridium difficile-associated diarrhea. He completed a previous course of metronidazole 3 weeks prior to presentation. He is on the liver transplant waiting list and has known hepatic encephalopathy. MRI revealed hyperintense T2 signals involving the bilateral dentate nuclei, inferior colliculi and splenium of the corpus callosum, and increased diffusion restriction at the splenium of the corpus callosum. His neurological function improved over the next several days. He underwent liver transplantation 6 days after admission. A follow-up MRI 6 weeks after presentation revealed resolution of abnormalities; however, paresthesias persisted 6 months after MIE diagnosis. Conclusion. An ESLD patient with hepatic encephalopathy developed MIE after a relatively short course of metronidazole. Metronidazole has been shown to accumulate in patients with ESLD. Increased awareness for neurotoxicity when using metronidazole in ESLD patients is warranted, especially in those with potentially confounding hepatic encephalopathy

Threading the Pieces Together: Integrative Perspective on SARS-CoV-2

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has challenged the research community globally to innovate, interact, and integrate findings across hierarchies. Research on SARS-CoV-2 has produced an abundance of data spanning multiple parallels, including clinical data, SARS-CoV-2 genome architecture, host response captured through transcriptome and genetic variants, microbial co-infections (metagenome), and comorbidities. Disease phenotypes in the case of COVID-19 present an intriguing complexity that includes a broad range of symptomatic to asymptomatic individuals, further compounded by a vast heterogeneity within the spectrum of clinical symptoms displayed by the symptomatic individuals. The clinical outcome is further modulated by the presence of comorbid conditions at the point of infection. The COVID-19 pandemic has produced an expansive wealth of literature touching many aspects of SARS-CoV-2 ranging from causal to outcome, predisposition to protective (possible), co-infection to comorbidity, and differential mortality globally. As challenges provide opportunities, the current pandemic’s challenge has underscored the need and opportunity to work for an integrative approach that may be able to thread together the multiple variables. Through this review, we have made an effort towards bringing together information spanning across different domains to facilitate researchers globally in pursuit of their response to SARS-CoV-2

Amino-Terminal Fusion of Epidermal Growth Factor 4,5,6 Domains of Human Thrombomodulin on Streptokinase Confers Anti-Reocclusion Characteristics along with Plasmin-Mediated Clot Specificity

<div><p>Streptokinase (SK) is a potent clot dissolver but lacks fibrin clot specificity as it activates human plasminogen (HPG) into human plasmin (HPN) throughout the system leading to increased risk of bleeding. Another major drawback associated with all thrombolytics, including tissue plasminogen activator, is the generation of transient thrombin and release of clot-bound thrombin that promotes reformation of clots. In order to obtain anti-thrombotic as well as clot-specificity properties in SK, cDNAs encoding the EGF 4,5,6 domains of human thrombomodulin were fused with that of streptokinase, either at its N- or C-termini, and expressed these in <i>Pichia pastoris</i> followed by purification and structural-functional characterization, including plasminogen activation, thrombin inhibition, and Protein C activation characteristics. Interestingly, the N-terminal EGF fusion construct (EGF-SK) showed plasmin-mediated plasminogen activation, whereas the C-terminal (SK-EGF) fusion construct exhibited ‘spontaneous’ plasminogen activation which is quite similar to SK i.e. direct activation of systemic HPG in absence of free HPN. Since HPN is normally absent in free circulation due to rapid serpin-based inactivation (such as alpha-2-antiplasmin and alpha-2-Macroglobin), but selectively present in clots, a plasmin-dependent mode of HPG activation is expected to lead to a desirable fibrin clot-specific response by the thrombolytic. Both the N- and C-terminal fusion constructs showed strong thrombin inhibition and Protein C activation properties as well, and significantly prevented re-occlusion in a specially designed assay. The EGF-SK construct exhibited fibrin clot dissolution properties with much-lowered levels of fibrinogenolysis, suggesting unmistakable promise in clot dissolver therapy with reduced hemorrhage and re-occlusion risks.</p></div