Effect of upadacitinib on atopic hand eczema in patients with moderate-to-severe atopic dermatitis: Results from two randomized phase 3 trials.

BACKGROUND: Approximately 60% of patients with atopic dermatitis have involvement of the hands adding to the burden of disease. OBJECTIVE: This analysis aims to evaluate the effect of upadacitinib monotherapy on atopic hand eczema in patients with moderate-to-severe AD over 16 weeks in the Measure Up 1 and 2 studies. METHODS: Data from patients (ages 12-75) randomized 1:1:1 to receive upadacitinib 15 mg, 30 mg, or placebo once daily in the Measure Up 1 and 2 studies were analysed for impact on atopic hand eczema assessed using the Hand Eczema Severity Index (HECSI). The percent change from baseline in HECSI score was a prespecified additional endpoint at all visits. The proportion of patients with at least a 75% improvement in HECSI score (HECSI 75) was evaluated post hoc. RESULTS: Patients treated with upadacitinib 15 mg or 30 mg experienced greater improvement in HECSI score compared with placebo as early as Week 1, which was maintained through Week 16. At Week 16, the mean change from baseline in HECSI score for patients receiving upadacitinib 15 mg, 30 mg, and placebo was -68%, -74%, and -15% in Measure Up 1 and -68%, -74% and +21% (positive change indicates worsening for placebo) in Measure Up 2, respectively. A greater proportion of upadacitinib-treated patients achieved HECSI 75 compared with placebo at all timepoints beginning at Week 1 through Week 16. CONCLUSIONS: Upadacitinib 15 mg and 30 mg monotherapy provided rapid and sustained improvement in atopic hand eczema compared with placebo through Week 16 in patients with moderate-to-severe AD. At Week 16, the observed mean improvements in HECSI score in upadacitinib-treated patients were clinically meaningful based on previous interpretability studies. These results suggest that upadacitinib may be an effective treatment option for atopic hand eczema in patients with moderate-to-severe AD.

Budd-Chiari syndrome (BCS): a challenging diagnosis not to be overlooked-single center report and pictorial essay.

Budd-Chiari syndrome (BCS) is a rare disease with a variable clinical presentation and often late diagnosis. Doppler ultrasonography (DUS) permits to determine the site of the obstructed venous tracts, the thrombotic or non-thrombotic nature of the obstruction with its morphologic features and the flow-pattern alterations. Other non-specific findings, which are seen in most of the other liver diseases, include ascites, hepatosplenomegaly and caudate hypertrophy. The aim of this study is to show our experience in BCS reporting retrospectively 15 cases referred to our hepatology center between 2017 and 2021. Four selected cases depict the extreme heterogeneous behaviour of BCS and highlight the importance of DUS as a diagnostic tool when there is a clinical suspicion. In patients, mainly young, who present with ascites and abdominal pain, BCS has to be considered and DUS is the first imaging technique to be performed to rule it out.

Tolerability profile of topical cannabidiol and palmitoylethanolamide: a compilation of single-centre randomized evaluator-blinded clinical and in vitro studies in normal skin.

BACKGROUND: An increasing number of studies have investigated the adverse effect profile of oral cannabinoids; however, few studies have provided sufficient data on the tolerability of topical cannabinoids in human participants. AIM: To assess the tolerability profile of several commercial topical formulations containing cannabidiol (CBD) and palmitoylethanolamide (PEA) on the skin of healthy human participants. METHODS: Three human clinical trials and one in vitro study were conducted. The potential for skin irritation, sensitization and phototoxicity of several products, were assessed via patch testing on healthy human skin. The products assessed included two formulations containing CBD and PEA, one containing hemp seed oil and four concentrations of CBD alone. Ocular toxicity was tested using a traditional hen's egg chorioallantoic membrane model with three CBD, PEA and hemp seed oil formulations. RESULTS: There was no irritation or sensitization of the products evident via patch testing on healthy participants. Additionally, mild phototoxicity of a hemp seed oil product was found at the 48-h time point compared with the negative control. The in vitro experiment demonstrated comparable effects of cannabinoid products with historically nonirritating products. CONCLUSION: These specific formulations of CBD- and PEA-containing products are nonirritating and nonsensitizing in healthy adults, and further encourage similar research assessing their long-term safety and efficacy in human participants with dermatological diseases. There are some limitations to the study: (i) external validity may be limited as formulations from a single manufacturer were used for this study, while vast heterogeneity exists across unregulated, commercial CBD products on the market; and (ii) products were assessed only on normal, nondiseased human skin, and therefore extrapolation to those with dermatological diseases cannot be assumed.

Impact of baricitinib in combination with topical steroids on atopic dermatitis symptoms, quality of life and functioning in adult patients with moderate-to-severe atopic dermatitis from the BREEZE-AD7 Phase 3 randomized trial.

BACKGROUND: Baricitinib is an oral, selective, reversible Janus kinase 1/2 inhibitor approved in the European Union and Japan and under investigation in the United States for treatment of atopic dermatitis (AD). OBJECTIVES: To evaluate the impact of baricitinib plus background topical corticosteroids (TCS) on health-related quality of life (HRQoL), how AD symptoms impact work productivity and life functioning, and treatment benefit using patient-reported outcome (PRO) assessments in patients with moderate-to-severe AD previously experiencing inadequate response to TCS. METHODS: Adult patients with AD in BREEZE-AD7, a Phase 3, multicentre, double-blind trial, were randomised 1 : 1 : 1 to daily oral placebo (control) or baricitinib 4- or 2-mg plus TCS. PROs reported Week 1 through Week 16: Dermatology Life Quality Index (DLQI), Work Productivity and Activity Impairment-AD (WPAI-AD); Patient-Reported Outcomes Measurement Information System (PROMIS) Itch and Sleep measures, and Patient Benefit Index (PBI). Data were analysed using logistic regression (categorical) and mixed model repeated measures (continuous). PBI scores were analysed using analysis of variance. RESULTS: A total of 329 patients were randomised. Treatment with baricitinib 4-mg (N = 111) or 2 mg (N = 109) plus TCS led to rapid, statistically significant improvements [vs. TCS plus placebo (N = 109)] in DLQI ≥4-point improvement starting at Week 2 (4-mg plus TCS, P ≤ 0.001; 2-mg plus TCS P ≤ 0.05), change from baseline in WPAI-AD presenteeism at Week 1 (4-mg plus TCS, P ≤ 0.01; 2-mg plus TCS P ≤ 0.05) and PROMIS itch interference at Week 2 (4-mg plus TCS P ≤ 0.01). Improvements were sustained through Week 16 for baricitinib 4-mg. Statistically significant improvements were observed at Week 16 for PBI global score (4-mg plus TCS, P ≤ 0.001; 2-mg plus TCS P ≤ 0.05). CONCLUSIONS: Baricitinib plus TCS vs. placebo plus TCS showed significant improvements in treatment benefit at Week 16 and rapid significant improvements in HRQoL and impact of AD symptoms on work productivity and functioning through 16 weeks.

Systems based analysis of human embryos and gene networks involved in cell lineage allocation.

BACKGROUND: Little is understood of the molecular mechanisms involved in the earliest cell fate decision in human development, leading to the establishment of the trophectoderm (TE) and inner cell mass (ICM) stem cell population. Notably, there is a lack of understanding of how transcriptional networks arise during reorganisation of the embryonic genome post-fertilisation. RESULTS: We identified a hierarchical structure of preimplantation gene network modules around the time of embryonic genome activation (EGA). Using network models along with eukaryotic initiation factor (EIF) and epigenetic-associated gene expression we defined two sets of blastomeres that exhibited diverging tendencies towards ICM or TE. Analysis of the developmental networks demonstrated stage specific EIF expression and revealed that histone modifications may be an important epigenetic regulatory mechanism in preimplantation human embryos. Comparison to published RNAseq data confirmed that during EGA the individual 8-cell blastomeres are transcriptionally primed for the first lineage decision in development towards ICM or TE. CONCLUSIONS: Using multiple systems biology approaches to compare developmental stages in the early human embryo with single cell transcript data from blastomeres, we have shown that blastomeres considered to be totipotent are not transcriptionally equivalent. Furthermore we have linked the developmental interactome to individual blastomeres and to later cell lineage. This has clinical implications for understanding the impact of fertility treatments and developmental programming of long term health.

Topical corticosteroid phobia in atopic dermatitis: International feasibility study of the TOPICOP score.

BACKGROUND: Adherence to topical corticosteroids (TCS) is essential for the effective treatment of atopic dermatitis but can be limited by concerns about their use. This study examined the feasibility of applying the validated TOPICOP score for assessing TCS phobia across different countries. METHODS: This was a prospective multicentre feasibility study conducted in 21 hospitals in 17 countries. Patients >3 months of age with atopic dermatitis or their parents or legal representatives completed a validated translation of the TOPICOP questionnaire in the country's native language. Respondents also completed questionnaires collecting opinions about the feasibility and acceptability of the TOPICOP questionnaire. RESULTS: A total of 1564 participants in 15 countries were included in the analysis. 81% of respondents considered the questions clear or very clear, and 79% reported that it took less than 5 minutes to complete. Each of the individual items in the TOPICOP questionnaire was considered to be not at all difficult to answer by 49% to 74% of participants. The mean global TOPICOP score was 44.7%±20.5. Mean TOPICOP subscores were 37.0±22.8% for knowledge and beliefs, 54.7±27.8% for fears and 50.1±29.1% for behaviours. Global scores and subscores differed between countries, although the subscores did not always vary in parallel, suggesting different levels of TCS phobia and different drivers for each country. CONCLUSIONS: The TOPICOP score can be feasibly applied across countries and may therefore be useful for obtaining qualitative and quantitative data from international studies and for adapting patient education and treatment.

Stochastic analysis of a miRNA-protein toggle switch.

Within systems biology there is an increasing interest in the stochastic behavior of genetic and biochemical reaction networks. An appropriate stochastic description is provided by the chemical master equation, which represents a continuous time Markov chain (CTMC). In this paper we consider the stochastic properties of a toggle switch, involving a protein compound (E2Fs and Myc) and a miRNA cluster (miR-17-92), known to control the eukaryotic cell cycle and possibly involved in oncogenesis, recently proposed in the literature within a deterministic framework. Due to the inherent stochasticity of biochemical processes and the small number of molecules involved, the stochastic approach should be more correct in describing the real system: we study the agreement between the two approaches by exploring the system parameter space. We address the problem by proposing a simplified version of the model that allows analytical treatment, and by performing numerical simulations for the full model. We observed optimal agreement between the stochastic and the deterministic description of the circuit in a large range of parameters, but some substantial differences arise in at least two cases: (1) when the deterministic system is in the proximity of a transition from a monostable to a bistable configuration, and (2) when bistability (in the deterministic system) is "masked" in the stochastic system by the distribution tails. The approach provides interesting estimates of the optimal number of molecules involved in the toggle switch. Our discussion of the points of strengths, potentiality and weakness of the chemical master equation in systems biology and the differences with respect to deterministic modeling are leveraged in order to provide useful advice for both the bioinformatician and the theoretical scientist.

Monitoring Chinese hamster ovary cell culture by the analysis of glucose and lactate metabolism.

Monitoring cell growth is crucial to the success of an animal cell culture process that can be accomplished by a variety of direct or indirect methodologies. Glucose is a major carbon and energy source for cultured mammalian cells in most cases, but glycolytic metabolism often results in the accumulation of lactate. Glucose and lactate levels are therefore routinely measured to determine metabolic activities of a culture. Typically, neither glucose consumption rate nor lactate accumulation rate has a direct correlation with cell density due to the changes in culture environment and cell physiology. We discovered that although the metabolic rate of glucose or lactate varies depending on the stages of a culture, the cumulative consumption of glucose and lactate combined (Q(GL)) exhibits a linear relationship relative to the integral of viable cells (IVC), with the slope indicating the specific consumption rate of glucose and lactate combined (q(GL)). Additional studies also showed that the q(GL) remains relatively constant under different culture conditions. The insensitivity of the q(GL) to process variations allows a potentially easy and accurate determination of viable cell density by the measurement of glucose and lactate. In addition, the more predictable nature of a linear relationship will aid the design of better forward control strategies to improve cell culture processes.

Analysing gene function after duplication.

After gene duplication, mutations cause the gene copies to diverge. The classical model predicts that these mutations will generally lead to the loss of function of one gene copy; rarely, new functions will be created and both duplicate genes are conserved. In contrast, under the subfunctionalization model both duplicates are preserved due to the partition of different functions between the duplicates. A recent study provides support for the subfunctionalization model, identifying several expressed gene duplicates common to humans and mice that contain regions conserved in one duplicate but variable in the other (and vice versa). We discuss both the methodology used in this study and also how gene phylogeny may lead to additional evidence for the importance of subfunctionalization in the evolution of new genes.

Molecular variation in plant cell populations evolving in vitro in different physiological contexts.

Previous work has shown the fixation of context-specific random amplified polymorphic DNA (RAPD) patterns in tomato cell cultures grown for 2 years in different hormonal contexts. In this work, RAPD sequences were characterised and RAPD-derived molecular markers used for a further study of variation between and within auto- and auxo-trophic tomato cultures grown in different hormonal equilibria. Results were then compared with those obtained using microsatellite markers located in noncoding regions of differentiation- and hormone-related genes and with those obtained with the external transcribed spacer (ETS) from tomato rDNA. Hybridisation of RAPDs on a tomato genomic DNA bank, or on total DNA after enzymatic digestion, suggested that the markers were repetitive in nature. Sequence analysis. however, showed that the homology between different fragments was due mainly to the presence of homo-AT nucleotide stretches. Moreover, a series of computational methods, such as an information-theory algorithm coupled with AG estimates, suggested that the RAPD fragments isolated in our experiments are noncoding. The amplification of SSR-containing RAPD-derived markers, and of other SSRs located in noncoding regions of tomato functional genes, consistently showed polymorphism between auxo- and auto-trophic somaclones (the latter being either habituated or transgenic for Agrobacterium tumefaciens oncogenes) but not within these same clones. Differences were also found between auxotrophic clones and the differentiated tissue. These findings were confirmed by restriction fragment length polymorphism (RFLP) analysis with the REII repetitive element of the ETS from tomato rDNA, which was isolated during this study. The results obtained suggest a possible role for physiological context in the selection of RAPD patterns during the evolution of tomato cells with different endogenous hormonal equilibria. The results are discussed in terms of a possible role for variation in noncoding regions of hormone-related genes in the adaptation to different physiological contexts.

Molecular phylogenetics: state-of-the-art methods for looking into the past.

As the amount of molecular sequence data in the public domain grows, so does the range of biological topics that it influences through evolutionary considerations. In recent years, a number of developments have enabled molecular phylogenetic methodology to keep pace. Likelihood-based inferential techniques, although controversial in the past, lie at the heart of these new methods and are producing the promised advances in the understanding of sequence evolution. They allow both a wide variety of phylogenetic inferences from sequence data and robust statistical assessment of all results. It cannot remain acceptable to use outdated data analysis techniques when superior alternatives exist. Here, we discuss the most important and exciting methods currently available to the molecular phylogeneticist.

Activation and habituation in olfaction--an fMRI study.

This study investigated human BOLD responses in primary and higher order olfactory cortices following presentation of short- and long-duration odorant stimuli using a 3-T MR scanner. The goal was to identify temporal differences in the course of the response that might underlie habituation. A short-duration stimulus (9 s) consistently activated the primary olfactory cortex (POC). After a long stimulus (60 s), the temporal form of the response differed in different parts of the olfactory network: (1) The POC (piriform, entorhinal cortex, amygdala) and, interestingly, the hippocampus and, to a certain degree, the anterior insula show a short, phasic increase in the signal, followed by a prolonged decrease below baseline. (2) In the orbitofrontal cortex a sustained increase in activation was seen. This increase lasted approximately as long as the duration of odorant presentation ( approximately 60 s). (3) The mediodorsal nucleus of the thalamus and the caudate nucleus responded with an increase in signal which returned to baseline after approximately 15 to 30 s. The correlated biphasic hemodynamic response in the POC, hippocampus, and anterior insula during prolonged olfactory stimulation suggests that these three areas may interact closely with each other in the control of habituation. These results extend recent data which showed habituation of the rat piriform cortex and dissociation between the POC and the orbitofrontal cortex.

Development and improvement of a serum-free suspension process for the production of recombinant adenoviral vectors using HEK293 cells.

Human Embryonic Kidney 293 (HEK293) cells were adapted into a serum-free suspension medium through steps of gradual serum weaning for the production of adenoviral (AdV) gene therapy vectors. The presence of sodium heparin in the medium formulation reduced cell clumping dramatically in suspension culture. The adapted cells were ready to grow either in serum-containing medium as an attached culture or in serum-free medium in suspension culture. A scalable production process was developed in shake flasks and was then evaluated in stirred tank bioreactors. This process includes a growth phase in batch-mode followed by a production phase involving medium perfusion and supplementation. Fortification with calcium chloride post viral inoculation resulted in an increase in virus production by at least one fold. Addition of stimulating agents such as sodium butyrate, N-acetyl-L-cysteine (NAC), dimethyl sulfoxide(DMSO), or ethyl alcohol post infection was shown to further improve virus production in a dose-dependent manner. The serum-free suspension process described here should be suitable for the manufacturing of other E1-deleted AdV vectors and could potentially be used for the production of recombinant proteins by HEK293 cells.