Quantification of lipid and peptide content in antigenic peptide-loaded liposome formulations by reversed-phase UPLC using UV absorbance and evaporative light scattering detection

Antigenic peptide-loaded cationic liposomes have shown promise as cancer vaccines. Quantification of both peptides and lipids is critical for quality control of such vaccines for clinical translation. In this work we describe a reversed phase ultra-performance liquid chromatography (RP-UPLC) method that separates lipids (DOTAP, DOPC and their degradation products) and two physicochemically different peptides within 12 min. Samples were prepared by dilution in a 1:1 (v/v) mixture of methanol and water. Peptide quantifi-cation was done via UV detection and lipids were quantified by an evaporative light scattering detector (ELSD), both coupled to the RP-UPLC system, with high precision (RSD < 3.5%). We showed that the presence of lipids and peptides did not mutually influence their quantification. Limit of detection (LOD) and limit of quantification (LOQ), as determined in the ICH guidelines, were 6 and 20 ng for DOTAP, 12 ng and 40 ng for DOPC, 3.0 ng and 8.0 ng for peptide A and 2.4 ng and 7.2 ng for the more hydrophobic peptide B. Finally, lipid degradation of DOTAP and DOPC was monitored in peptide loaded DOTAP:DOPC liposomes upon storage at 4 degrees C and 40 degrees C.(c) 2022 The Authors. Published by Elsevier Inc. on behalf of American Pharmacists Association. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)Drug Delivery Technolog

Billiards in a general domain with random reflections

We study stochastic billiards on general tables: a particle moves according to its constant velocity inside some domain ${\mathcal D} \subset {\mathbb R}^d$ until it hits the boundary and bounces randomly inside according to some reflection law. We assume that the boundary of the domain is locally Lipschitz and almost everywhere continuously differentiable. The angle of the outgoing velocity with the inner normal vector has a specified, absolutely continuous density. We construct the discrete time and the continuous time processes recording the sequence of hitting points on the boundary and the pair location/velocity. We mainly focus on the case of bounded domains. Then, we prove exponential ergodicity of these two Markov processes, we study their invariant distribution and their normal (Gaussian) fluctuations. Of particular interest is the case of the cosine reflection law: the stationary distributions for the two processes are uniform in this case, the discrete time chain is reversible though the continuous time process is quasi-reversible. Also in this case, we give a natural construction of a chord "picked at random" in ${\mathcal D}$, and we study the angle of intersection of the process with a $(d-1)$-dimensional manifold contained in ${\mathcal D}$.Comment: 50 pages, 10 figures; To appear in: Archive for Rational Mechanics and Analysis; corrected Theorem 2.8 (induced chords in nonconvex subdomains

The Counterpart Principle of Analogical Support by Structural Similarity

We propose and investigate an Analogy Principle in the context of Unary Inductive Logic based on a notion of support by structural similarity which is often employed to motivate scientific conjectures

Chronic hand eczema in Europe:Patient experiences and perspectives (CHEPEP) in qualitative interviews

Background: Chronic hand eczema (CHE) is a very common skin disease among the European population. It causes itch and pain and, in more severe cases, seriously impairs hand functioning at work and in private life. Objectives: To explore perspectives of people with lived experience on CHE-related problems, wishes and goals. Methods: Following a qualitative approach, we conducted topic-guided interviews in five European countries and applied template analysis to identify recurrent themes among patients with CHE. Results: We interviewed 60 patients in seven outpatient dermatological and occupational medicine clinics in Croatia, Denmark, Germany, the Netherlands and Spain. Five main themes were identified: (1) knowledge about the disease and its course, (2) preventive behaviour, (3) hand eczema therapy, (4) impact on everyday life and (5) attitudes towards CHE and healthcare. Participants did not feel well informed about CHE, especially about causes, triggers and treatment options. Preventive measures were experienced as more or less effective but also cumbersome. Experiences with therapy were diverse. Treatment satisfaction depended on the results and on the perceived support from the treatment teams. Participants found it important to be taken seriously, to receive practical advice, to try out additional treatments or examinations, find new hope and have occupational perspectives. They wished that others could better understand the physical and emotional burden of CHE. Patient support groups were not mentioned. Participants found it important to learn to take care of themselves and accept life with CHE. Conclusions: Due to its annoying symptoms, high visibility and impaired functioning at work and in private life, CHE has a high emotional and social impact. Some people may require support to learn coping with CHE and its prevention. Patients wish for information about causes and triggers. They value physicians who listen to them and keep looking for solutions.</p

Towards Heat-stable Oxytocin Formulations: Analysis of Degradation Kinetics and Identification of Degradation Products

Purpose. To investigate degradation kinetics of oxytocin as a function of temperature and pH, and identify the degradation products. Materials and Methods. Accelerated degradation of oxytocin formulated at pH 2.0, 4.5, 7.0 and 9.0 was performed at 40, 55, 70 and 80°C. Degradation rate constants were determined from RP-HPLC data. Formulations were characterized by HP-SEC, UV absorption and fluorescence spectroscopy. Degradation products were identified by ESI-MS/MS. Results. The loss of intact oxytocin in RP-HPLC was pH- and temperature-dependent and followed (pseudo) first order kinetics. Degradation was fastest at pH 9.0, followed by pH 7.0, pH 2.0 and pH 4.5. The Arrhenius equation proved suitable to describe the kinetics, with the highest activation energy (116.3 kJ/mol) being found for pH 4.5 formulations. At pH 2.0 deamidation of Gln 4, Asn 5, and Gly 9-NH2, as well as combinations thereof were found. At pH 4.5, 7.0 and 9.0, the formation of tri- and tetrasulfidecontaining oxytocin as well as different types of disulfide and dityrosine-linked dimers were found to occur. Beta-elimination and larger aggregates were also observed. At pH 9.0, mono-deamidation of Gln 4, Asn 5, and Gly 9-NH2 additionally occurred. Conclusions. Multiple degradation products of oxytocin have been identified unequivocally, including various deamidated species, intramolecular oligosulfides and covalent aggregates. The strongly pH dependent degradation can be described by the Arrhenius equation. KEY WORDS: aggregation; Arrhenius kinetics; degradation; mass spectrometry; oxytocin

Immune Modulation by Adjuvants Combined with Diphtheria Toxoid Administered Topically in BALB/c Mice After Microneedle Array Pretreatment

Purpose. In this study, modulation of the immune response against diphtheria toxoid (DT) by various adjuvants in transcutaneous immunization (TCI) with microneedle array pretreatment was investigated. Methods. TCI was performed on BALB/c mice with or without microneedle array pretreatment using DT as a model antigen co-administrated with lipopolysaccharide (LPS), Quil A, CpG oligo deoxynucleotide (CpG) or cholera toxin (CT) as adjuvant. The immunogenicity was evaluated by measuring serum IgG subtype titers and neutralizing antibody titers. Results. TCI with microneedle array pretreatment resulted in a 1,000-fold increase of DT-specific serum IgG levels as compared to TCI. The immune response was further improved by co-administration of adjuvants, showing a progressive increase in serum IgG titers when adjuvanted with LPS, Quil A, CpG and CT. IgG titers of the CT-adjuvanted group reached levels comparable to those obtained after DTalum subcutaneous injection. The IgG1/IgG2a ratio of DT-specific antibodies decreased in the following sequence: plain DT, Quil A, CT and CpG, suggesting that the immune response was skewed towards the Th1 direction. Conclusions. The potency and the quality of the immune response against DT administered by microneedle array mediated TCI can be modulated by co-administration of adjuvants. KEY WORDS: cholera toxin; CpG; diphtheria toxoid; microneedle array; transcutaneous immunization

Report from the fifth international consensus meeting to harmonize core outcome measures for atopic eczema/dermatitis clinical trials (HOME initiative)

This is the report from the fifth meeting of the Harmonising Outcome Measures for Eczema initiative (HOME V). The meeting was held on 12–14 June 2017 in Nantes, France, with 81 participants. The main aims of the meeting were (i) to achieve consensus over the definition of the core domain of long-term control and how to measure it and (ii) to prioritize future areas of research for the measurement of the core domain of quality of life (QoL) in children. Moderated whole-group and small-group consensus discussions were informed by presentations of qualitative studies, systematic reviews and validation studies. Small-group allocations were performed a priori to ensure that each group included different stakeholders from a variety of geographical regions. Anonymous whole-group voting was carried out using handheld electronic voting pads according to predefined consensus rules. It was agreed by consensus that the long-term control domain should include signs, symptoms, quality of life and a patient global instrument. The group agreed that itch intensity should be measured when assessing long-term control of eczema in addition to the frequency of itch captured by the symptoms domain. There was no recommendation of an instrument for the core outcome domain of quality of life in children, but existing instruments were assessed for face validity and feasibility, and future work that will facilitate the recommendation of an instrument was agreed upon. The Harmonising Outcome Measures for Eczema (HOME) initiative is an international group working together to develop a core outcome set (COS) for clinical trials in eczema (synonymous with atopic eczema and atopic dermatitis). HOME is coordinated from the Centre of Evidence Based Dermatology, University of Nottingham, U.K. Participation in HOME is open to anyone with an interest in outcomes for eczema. A COS is the agreed upon minimum set of instruments that should be included in all clinical trials for a particular condition. Use of a COS does not preclude using other instruments; other domains and instruments can also be included to meet the specific requirements of individual trials. COS initiatives are active across many fields of medicine and should enable better synthesis of trial data and reduce selective outcome reporting bias. The HOME initiative follows the best current guidance on developing a COS. Four core domains have been identified: clinician-reported signs; patient-reported symptoms; quality of life; and long-term control. The core outcome measurement instruments for clinician-reported signs and patient-reported symptoms have been established: the Eczema Area and Severity Index (EASI) for measuring clinician reported signs was agreed on at the HOME III meeting, and the Patient-Oriented Eczema Measure (POEM) was chosen to measure patient-reported symptoms at the HOME IV meeting. This is a report from the fifth consensus meeting of the HOME initiative (HOME V), which was held on 12–14 June 2017 in Nantes, France. The local organizers were Sebastien Barbarot and Jean-Francois Stalder of Nantes University Hospital, France

Prion Uptake in the Gut: Identification of the First Uptake and Replication Sites

After oral exposure, prions are thought to enter Peyer's patches via M cells and accumulate first upon follicular dendritic cells (FDCs) before spreading to the nervous system. How prions are actually initially acquired from the gut lumen is not known. Using high-resolution immunofluorescence and cryo-immunogold electron microscopy, we report the trafficking of the prion protein (PrP) toward Peyer's patches of wild-type and PrP-deficient mice. PrP was transiently detectable at 1 day post feeding (dpf) within large multivesicular LAMP1-positive endosomes of enterocytes in the follicle-associated epithelium (FAE) and at much lower levels within M cells. Subsequently, PrP was detected on vesicles in the late endosomal compartments of macrophages in the subepithelial dome. At 7–21 dpf, increased PrP labelling was observed on the plasma membranes of FDCs in germinal centres of Peyer's patches from wild-type mice only, identifying FDCs as the first sites of PrP conversion and replication. Detection of PrP on extracellular vesicles displaying FAE enterocyte-derived A33 protein implied transport towards FDCs in association with FAE-derived vesicles. By 21 dpf, PrP was observed on the plasma membranes of neurons within neighbouring myenteric plexi. Together, these data identify a novel potential M cell-independent mechanism for prion transport, mediated by FAE enterocytes, which acts to initiate conversion and replication upon FDCs and subsequent infection of enteric nerves

30-day morbidity and mortality of sleeve gastrectomy, Roux-en-Y gastric bypass and one anastomosis gastric bypass: a propensity score-matched analysis of the GENEVA data

Background: There is a paucity of data comparing 30-day morbidity and mortality of sleeve gastrectomy (SG), Roux-en-Y gastric bypass (RYGB), and one anastomosis gastric bypass (OAGB). This study aimed to compare the 30-day safety of SG, RYGB, and OAGB in propensity score-matched cohorts. Materials and methods: This analysis utilised data collected from the GENEVA study which was a multicentre observational cohort study of bariatric and metabolic surgery (BMS) in 185 centres across 42 countries between 01/05/2022 and 31/10/2020 during the Coronavirus Disease-2019 (COVID-19) pandemic. 30-day complications were categorised according to the Clavien–Dindo classification. Patients receiving SG, RYGB, or OAGB were propensity-matched according to baseline characteristics and 30-day complications were compared between groups. Results: In total, 6770 patients (SG 3983; OAGB 702; RYGB 2085) were included in this analysis. Prior to matching, RYGB was associated with highest 30-day complication rate (SG 5.8%; OAGB 7.5%; RYGB 8.0% (p = 0.006)). On multivariate regression modelling, Insulin-dependent type 2 diabetes mellitus and hypercholesterolaemia were associated with increased 30-day complications. Being a non-smoker was associated with reduced complication rates. When compared to SG as a reference category, RYGB, but not OAGB, was associated with an increased rate of 30-day complications. A total of 702 pairs of SG and OAGB were propensity score-matched. The complication rate in the SG group was 7.3% (n = 51) as compared to 7.5% (n = 53) in the OAGB group (p = 0.68). Similarly, 2085 pairs of SG and RYGB were propensity score-matched. The complication rate in the SG group was 6.1% (n = 127) as compared to 7.9% (n = 166) in the RYGB group (p = 0.09). And, 702 pairs of OAGB and RYGB were matched. The complication rate in both groups was the same at 7.5 % (n = 53; p = 0.07). Conclusions: This global study found no significant difference in the 30-day morbidity and mortality of SG, RYGB, and OAGB in propensity score-matched cohorts

30-Day morbidity and mortality of bariatric metabolic surgery in adolescence during the COVID-19 pandemic – The GENEVA study

Background: Metabolic and bariatric surgery (MBS) is an effective treatment for adolescents with severe obesity. Objectives: This study examined the safety of MBS in adolescents during the coronavirus disease 2019 (COVID-19) pandemic. Methods: This was a global, multicentre and observational cohort study of MBS performed between May 01, 2020, and October 10,2020, in 68 centres from 24 countries. Data collection included in-hospital and 30-day COVID-19 and surgery-specific morbidity/mortality. Results: One hundred and seventy adolescent patients (mean age: 17.75 ± 1.30 years), mostly females (n = 122, 71.8%), underwent MBS during the study period. The mean pre-operative weight and body mass index were 122.16 ± 15.92 kg and 43.7 ± 7.11 kg/m2, respectively. Although majority of patients had pre-operative testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (n = 146; 85.9%), only 42.4% (n = 72) of the patients were asked to self-isolate pre-operatively. Two patients developed symptomatic SARS-CoV-2 infection post-operatively (1.2%). The overall complication rate was 5.3% (n = 9). There was no mortality in this cohort. Conclusions: MBS in adolescents with obesity is safe during the COVID-19 pandemic when performed within the context of local precautionary procedures (such as pre-operative testing). The 30-day morbidity rates were similar to those reported pre-pandemic. These data will help facilitate the safe re-introduction of MBS services for this group of patients