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Association of digital measures and self-reported fatigue: a remote observational study in healthy participants and participants with chronic inflammatory rheumatic disease.
No full textFatigue is a subjective, complex and multi-faceted phenomenon, commonly experienced as tiredness. However, pathological fatigue is a major debilitating symptom associated with overwhelming feelings of physical and mental exhaustion. It is a well-recognized manifestation in chronic inflammatory rheumatic diseases, such as Sjögren's Syndrome and Systemic Lupus Erythematosus and an important predictor of patient's health-related quality of life (HRQoL). Patient reported outcome questions are the key instruments to assess fatigue. To date, there is no consensus about reliable quantitative assessments of fatigue.Observational data for a period of one month were collected from 296 participants in the United States. Data comprised continuous multimodal digital data from Fitbit, including heart rate, physical activity and sleep features, and app-based daily and weekly questions covering various HRQoL factors including pain, mood, general physical activity and fatigue. Descriptive statistics and hierarchical clustering of digital data were used to describe behavioural phenotypes. Gradient boosting classifiers were trained to classify participant-reported weekly fatigue and daily tiredness from multi-sensor and other participant-reported data, and extract a set of key predictive features.Cluster analysis of Fitbit parameters highlighted multiple digital phenotypes, including sleep-affected, fatigued and healthy phenotypes. Features from participant-reported data and Fitbit data both contributed as key predictive features of weekly physical and mental fatigue and daily tiredness. Participant answers to pain and depressed mood-related daily questions contributed the most as top features for predicting physical and mental fatigue, respectively. To classify daily tiredness, participant answers to questions on pain, mood and ability to perform daily activities contributed the most. Features related to daily resting heart rate and step counts and bouts were overall the most important Fitbit features for the classification models.These results demonstrate that multimodal digital data can be used to quantitatively and more frequently augment pathological and non-pathological participant-reported fatigue
Convergence of distinct RNA-silencing pathways on GW182/Tnrc6
No full textThe RNA-binding protein Trim71/Lin41 is a phylogenetically conserved developmental regulator that functions in mammalian stem cell reprogramming, brain development and cancer. Trim71 recognizes target mRNAs through hairpin motifs and silences them through molecular mechanisms that await identification. Using a genome-wide CRISPR-based screen, we find that the miRNA pathway attenuates Trim71 activity in mouse embryonic stem cells (mESCs). This interaction does not involve regulation of Trim71 levels, which are not limiting for Trim71 activity in mESCs. Instead, Trim71 competes with Argonaute (Ago) proteins for GW182/Tnrc6 proteins as shared silencing factors. We show that Trim71 and Ago2 recruit Tnrc6 both to unique and to shared target transcripts. Our results provide insight into the mechanism of action of Trim71 and identify an unexpected function of Tnrc6 outside the miRNA-induced silencing complex (miRISC). By functioning in distinct silencing pathways, Tnrc6s provide a means to integrate the activity of distinct silencing pathways through competition and, potentially, cooperation
cGLRs are a diverse family of pattern recognition receptors in innate immunity.
No full textCyclic GMP-AMP synthase (cGAS) is an enzyme in human cells that controls an immune response to cytosolic DNA. Upon binding DNA, cGAS synthesizes a nucleotide signal 2'3'-cGAMP that activates STING-dependent downstream immunity. Here, we discover that cGAS-like receptors (cGLRs) constitute a major family of pattern recognition receptors in innate immunity. Building on recent analysis in Drosophila, we identify >3,000 cGLRs present in nearly all metazoan phyla. A forward biochemical screening of 150 animal cGLRs reveals a conserved mechanism of signaling including response to dsDNA and dsRNA ligands and synthesis of isomers of the nucleotide signals cGAMP, c-UMP-AMP, and c-di-AMP. Combining structural biology and in vivo analysis in coral and oyster animals, we explain how synthesis of distinct nucleotide signals enables cells to control discrete cGLR-STING signaling pathways. Our results reveal cGLRs as a widespread family of pattern recognition receptors and establish molecular rules that govern nucleotide signaling in animal immunity
Neutralizing Antibody Validation Testing and Reporting Harmonization.
No full textEvolving immunogenicity assay performance expectations and a lack of harmonized neutralizing antibody validation testing and reporting tools have resulted in significant time spent by health authorities and sponsors on resolving filing queries. A team of experts within the American Association of Pharmaceutical Scientists' Therapeutic Product Immunogenicity Community across industry and the Food and Drug Administration addressed challenges unique to cell-based and non-cell-based neutralizing antibody assays. Harmonization of validation expectations and data reporting will facilitate filings to health authorities and are described in this manuscript. This team provides validation testing and reporting strategies and tools for the following assessments: (1) format selection; (2) cut point; (3) assay acceptance criteria; (4) control precision; (5) sensitivity including positive control selection and performance tracking; (6) negative control selection; (7) selectivity/specificity including matrix interference, hemolysis, lipemia, bilirubin, concomitant medications, and structurally similar analytes; (8) drug tolerance; (9) target tolerance; (10) sample stability; and (11) assay robustness
Remibrutinib (LOU064) inhibits neuroinflammation driven by B cells and myeloid cells in preclinical models of Multiple Sclerosis
No full textBackground:
Bruton’s tyrosine kinase (BTK) is a key signaling node in B cell receptor (BCR) and Fc receptor
(FcR) signaling. BTK inhibitors (BTKi) are an emerging oral treatment option for patients suffering from multiple sclerosis (MS). Remibrutinib (LOU064) is a potent, highly selective covalent BTKi with a promising preclinical and clinical profile for MS and other autoimmune or autoallergic indications.
Methods:
The efficacy and mechanism of action of remibrutinib was assessed in two different experimental autoimmune encephalomyelitis (EAE) mouse models for MS. The impact of remibrutinib on B cell driven EAE pathology was determined after immunization with human myelin oligodendrocyte glycoprotein (HuMOG). The efficacy on myeloid cell and microglia driven neuroinflammation was determined in the RatMOG EAE. In addition, we assessed the relationship of efficacy to BTK occupancy in tissue, ex vivo T cell response, serum biomarkers such as total and antigen-specific immunoglobulins (Ig) and neurofilament light chain (NfL), as well as single cell RNA sequencing (scRNA-seq) in EAE brain and spinal cord tissue.
Results:
Remibrutinib inhibited B cell dependent HuMOG EAE in dose-dependent manner and strongly reduced neurological symptoms. At the efficacious oral dose of 30 mg/kg, remibrutinib showed strong BTK occupancy in the peripheral immune organs and in the brain of EAE mice. Ex vivo MOG-specific Th17 T cell recall response was reduced, but not polyclonal T cell response, indicating absence of general T cell inhibition. Remibrutinib also inhibited RatMOG EAE, suggesting that myeloid cell and microglia inhibition contribute to its efficacy in EAE. Remibrutinib did not reduce total IgG or IgM levels nor MOG-specific antibody response. In EAE brain and spinal cord tissue a clear anti-inflammatory effect in microglia was detected by scRNA-seq. Finally, remibrutinib showed potent inhibition of in vitro immune complex-driven inflammatory response in human microglia.
Conclusion:
Remibrutinib inhibited EAE models by a two-pronged mechanism based on inhibition of pathogenic B cell autoreactivity, as well as direct anti-inflammatory effects on microglia. Remibrutinib showed efficacy in both models in absence of direct B cell depletion, broad T cell inhibition or reduction of total Ig levels. These findings support the view that remibrutinib may represent a novel treatment option for patients with MS
Imaging in Small Rodents in the Context of Pharmacological Research: Significance of the 3R Principles
No full textMinimising potential pain and distress as well as the number of animals in biomedical experimentation and in the drug discovery and development process is not only an ethical imperative but also a challenging and evolving area of research. Non-invasive imaging can refine in vivo experimentation by using alternative readouts which represent early disease state or adverse drug effects and which allow harmful progress to be detected. This in contrast to animals being sacrificed at given time points and using artificial peri- or post-mortem endpoints. In many instances non-invasive imaging may replace pathohistological assessment of disease progression with anatomic and functional readouts. Furthermore, the number of animals can be substantially reduced by using each animal as its own control, thereby improving statistical power and information that may relate better to that observed in the clinical assessment of therapy efficacy. Taking as basis some of our own activities in the area of preclinical pharmacological research, we illustrate how imaging can contribute to the principles of humane experimental techniques outlined by Russell and Burch in 1959 in their publication about “The Principles of Humane Experimental Technique”
Sustainable cascade water reaction combining transition metal-biocatalysis and hydrophobic substrates: The effect of solvent structuring
No full textA cascade reaction consisting of a Heck reaction followed by an enzyme-catalyzed reaction is carried out in different aqueous solutions. In particular, the impact of the structuring of the reaction solvent is investigated. For this purpose, several ternary mixtures of water, isopropanol, and benzyl alcohol, including surfactant-free microemulsions, as well as binary mixtures of water and isopropanol, are taken into account. A micellar solution of the surfactant TPGS-750-M serves as a reference system. The coupling of the two reactions can be successfully performed in all 3 types of solvents, whereby the best result is achieved in a surfactant-free microemulsion. In addition, our system allows for reducing the temperature during the Heck reaction. The studies further reveal that the structures built up by surfactants are not necessarily the main reason allowing organic reactions to be carried out in water. Rather, it is a question of solubility and stability of the reaction components. Thereby, better solubility does not always correlate with increased reactivity. We thus provide a deeper understanding of solvent – reactivity relationship and want to advert an approach beside micellar solvents for transferring organic reactions into environmentally friendly aqueous reaction solutions
First-in-human study of safety, pharmacokinetics, and pharmacodynamics of MHV370, a dual inhibitor of Toll-like receptors 7 and 8, in healthy adults
No full textBackground and Objective: MHV370, a dual antagonist of human Toll-like receptors (TLR) 7 and 8, suppresses cytokines and interferon stimulated genes in vitro and in vivo, and demonstrates efficacy in murine models of lupus. This first-in-human study aimed to evaluate the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of single and multiple doses of MHV370, in healthy adults, as well as the effects of food consumption on a single dose of MHV370.
Methods: This was a Phase 1, first-in-human, randomised, placebo-controlled study conducted in 3 parts. In study Part A, participants received (3:1) a single ascending dose (SAD) of 1, 3, 10, 20, 40, 80, 160, 320, 640, and 1000 mg MHV370 or placebo. In study Part B, participants received (3:1) multiple ascending doses (MAD) of 25, 50, 100, 200, and 400 mg MHV370 twice daily or placebo, for 14 days. In study Part C, participants received an open-label single dose of 200 mg MHV370 under fasted or fed conditions. Safety, PK, and PD parameters were evaluated.
Results: MHV370 was well tolerated, and no safety signal was observed in the study. No dose limiting adverse events occurred across the dose range evaluated. Plasma concentrations of MHV370 increased with dose. The intake of food did not have an impact on the PK of MHV370. PD data indicated a time- and dose-dependent inhibition of TLR7-mediated CD69 expression on B cells and TLR8-mediated TNF release, after ex-vivo stimulation.
Conclusion: The safety, PK and PD data support the further development of MHV370 in systemic autoimmune diseases driven by overactivation of TLR7 and TLR8
Discovery of a selective and biologically active low-molecular weight antagonist of human IL-1b
No full textHuman interleukin-1b (hIL-1b) is a pro-inflammatory cytokine involved in many diseases. While hIL-1b directed antibodies have shown clinical benefit, an orally available low-molecular weight antagonist is still elusive, limiting the applications of hIL-1b-directed therapies. Here we describe the discovery of the first low-molecular weight hIL-1b antagonist that blocks the interaction with the IL-1R1 receptor. Starting from a low affinity fragment-based screening hit 1, structure-based optimization resulted in a compound (S)-2 that binds and antagonizes hIL-1b with single-digit micromolar activity in biophysical, biochemical, and cellular assays. X-ray analysis reveals an allosteric mode of action that involves a hitherto unknown binding site in hIL-1b encompassing two loops involved in hIL-1R1/hIL-1b interactions. We show that residues of this binding site are part of a conformationally excited state of the mature cytokine. The compound antagonizes hIL-1b function in cells, including primary human fibroblasts, demonstrating the relevance of this discovery for future development of novel hIL-1b directed therapeutics
Establishing the clinical bioequivalence safe space via physiologically-based population pharmacokinetics absorption modeling. Case study: Fevipiprant/QAW039
No full textPhysiologically based pharmacokinetics (PBPK) and absorption modeling has increasingly been implemented for biopharmaceutics applications to define the bioequivalence safe space for drug product quality attributes such as dissolution. For fevipiprant/QAW039, PBPK analyses were performed to assess the impact of in vitro dissolution on the in vivo PK performance of immediate release (IR) film coated tablets during development and scaling-up to commercial scale.
A fevipiprant dissolution safe space was established using observed clinical intravenous and oral PK data from bioequivalent and non-bioequivalent formulations. Quality control tablet dissolution profiles were used as GastroPlusTM model inputs to estimate the in vivo dissolution in the GI tract, and to predict human exposure. The model was used to evaluate the intraluminal performance of the dosage forms and to determine the absorption rate limits for the 450 mg dose.
The predictive model performance was demonstrated for various oral dosage forms (150‒500 mg), including the non-bioequivalent batches in fasted healthy adults. To define the dissolution safe space boundaries at 450 mg, simulations were performed using theoretical dissolution profiles. A specification of Q=80% dissolved after 60 min for an IR oral solid dosage form reflected the limitations of the safe space. The dissolution profile of the 450 mg commercial-scale batch was within a dissolution region where bioequivalence is anticipated, not near an edge of failure for dissolution, providing additional confidence to the proposed acceptance criteria. Thus, the PBPK dissolution safe space allowed for a wider than 10% dissolution difference for bioequivalent batches, superseding f2 similarity analyses