Mucosal IFNλ1 mRNA-based immunomodulation effectively reduces SARS-CoV-2 induced mortality in mice.

RNA vaccines elicit protective immunity against SARS-CoV-2, but the use of mRNA as an antiviral immunotherapeutic is unexplored. Here, we investigate the activity of lipidoid nanoparticle (LNP)-formulated mRNA encoding human IFNλ1 (ETH47), which is a critical driver of innate immunity at mucosal surfaces protecting from viral infections. IFNλ1 mRNA administration promotes dose-dependent protein translation, induction of interferon-stimulated genes without relevant signs of unspecific immune stimulation, and dose-dependent inhibition of SARS-CoV-2 replication in vitro. Pulmonary administration of IFNλ1 mRNA in mice results in a potent reduction of virus load, virus-induced body weight loss and significantly increased survival. These data support the development of inhaled administration of IFNλ1 mRNA as a potential prophylactic option for individuals exposed to SARS-CoV-2 or at risk suffering from COVID-19. Based on the broad antiviral activity of IFNλ1 regardless of virus or variant, this approach might also be utilized for other respiratory viral infections or pandemic preparedness.

[Results of Endoscopic Screening and Therapy of the Duodenum in MUTYH-associated Polyposis]. / Ergebnisse der endoskopischen Vorsorge und Therapie des Duodenums bei MUTYH-assoziierter Polyposis.

MUTYH-associated polyposis (MAP) is a very rare autosomal recessive polyposis syndrome. It is caused by a homozygous or compound heterozygous germline mutation in the MUTYH gene. MAP is characterised by numerous colorectal adenomas; furthermore there is an increased risk for colorectal cancer (CRC). However, the phenotype can be highly variable; for example, affected individuals also have an increased risk of polyps of the upper gastrointestinal tract and development of duodenal carcinomas.This study included 15 patients with evidence of a pathogenic MUTYH variant, who were screened at the National Center for Hereditary Tumor Syndromes. Oesophagogastroduodenoscopy (EGD) results were prospectively recorded in a database from 2012 to 2023.At least one EGD (median 4, range 1-15) was performed in 15 patients, seven of whom carried a homozygous and 8 a compound heterozygous pathogenic MUTYH variant. The median surveillance period was 115 months (range, 3-215 months). The median age at baseline was 44 (range 17-65) years. A total of 72 EGDs were performed (median 4; range 1-15). Five patients had duodenal adenomas; histology showed tubular adenomas with low grade intraepithelial dysplasia (LGIEN) in all of these cases. The total number of duodenal adenomas detected was 48, and the median number was 3 (range, 1-37). Neither high grade intraepithelial neoplasia (HGIEN) nor duodenal cancer was detected during the surveillance period.Patients with MUTYH-associated polyposis should be managed in a multidisciplinary centre for hereditary tumour disease. Our cohort showed more patients with duodenal adenomas than in previously published data. However, no progression to HGIEN or duodenal carcinomas was observed as a result of the endoscopic therapy performed.

Transcriptional Dynamics of NRF2 Overexpression and KEAP1-NRF2 Inhibitors in Human Cell Line and Primary Lung Cells.

Oxidative stress in the human lung is caused by both internal (e.g., inflammation) and external stressors (smoking, pollution, and infection) to drive pathology in a number of lung diseases. Cellular damage caused by oxidative damage is reversed by several pathways, one of which is the antioxidant response. This response is regulated by the transcriptional factor NRF2, which has the ability to regulate the transcription of more than 250 genes. In disease, this balance is overwhelmed, and the cells are unable to return to homeostasis. Several pharmacological approaches aim to improve the antioxidant capacity by inhibiting the interaction of NRF2 with its key cytosolic inhibitor, KEAP1. Here, we evaluate an alternative approach by overexpressing NRF2 from chemically modified RNAs (cmRNAs). Our results demonstrate successful expression of functional NRF2 protein in human cell lines and primary cells. We establish a kinetic transcriptomic profile to compare antioxidant response gene expression after treatment of primary human bronchial epithelial cells with either KEAP1 inhibitors or cmRNAs. The key gene signature is then applied to primary human lung fibroblasts and alveolar macrophages to uncover transcriptional preferences in each cell system. This study provides a foundation for the understanding of NRF2 dynamics in the human lung and provides initial evidence of alternative ways for pharmacological interference.

Modulation of Liver Inflammation and Fibrosis by Interleukin-37.

Background and Aims: Chronic inflammation induces liver fibrosis, cirrhosis and potentially liver cancer. Kupffer cells modulate hepatic stellate cells by secreting immunologically active proteins as TGF-ß. TGF-ß promotes liver fibrosis via the activation of Sma- and Mad-related protein 3. IL-37 broadly suppresses innate and adaptive immune responses. Intracellular IL-37 interacts with Smad3. We hypothesize that IL-37 downregulates the activation of hepatic Kupffer and stellate cells and interferes with the TGF-ß signaling cascade to modulate liver fibrogenesis. Methods: The role of IL-37 on liver inflammation and fibrogenesis was assessed in three mouse models as well as isolated Kupffer- and stellate cells. Serum IL-37 was tested by ELISA in a clinical cohort and correlated with liver disease severity. Results: Transgene expression of IL-37 in mice extends survival, reduces hepatic damage, expression of early markers of fibrosis and histologically assessed liver fibrosis after bile duct ligation. IL-37tg mice were protected against CCl4-induced liver inflammation. Colitis-associated liver inflammation and fibrosis was less severe in IL-10 knockout IL-37tg mice. Spontaneous and LPS/TGF-ß-induced cytokine release and profibrogenic gene expression was lower in HSC and KC isolated from IL-37tg mice and IL-37 overexpressing, IL-1ß stimulated human LX-2 stellate cells. However, administration of recombinant human IL-37 did not modulate fibrosis pathways after BDL in mice, LX2 cells or murine HSCs. In a large clinical cohort, we observed a positive correlation of serum IL-37 levels with disease severity in liver cirrhosis. Conclusions: Predominantly intracellular IL-37 downregulates liver inflammation and fibrosis. The correlation of serum IL-37 with disease severity in cirrhosis suggests its potential as a novel target modulating the course of liver fibrosis.

P-selectin as a candidate target in atherosclerosis.

P-selectin is of critical importance in early atherogenesis by initiating leukocyte rolling at the site of endothelial injury. In order to validate P-selectin as a candidate target for the development of anti-atherogenic strategies, we wanted to obtain quantitative information on P-selectin expression, and identify novel peptide-based lead structures that interact with P-selectin. P-selectin mRNA expression in the aortic arch and in other tissues of apoE-deficient (apoE-/-) mice was determined by real-time PCR technology. P-selectin mRNA expression of apoE-/- mice increased steadily with age to levels 14-fold higher than that of control animals. The onset and level of P-selectin expression correlated well with the extent of lesion development, and was more specific for atherosclerotic tissue as compared with other adhesion molecules. Phage display technology was used to obtain novel P-selectin antagonists. Phage display selections resulted in the isolation of a highly P-selectin-specific phage clone. Synthetic peptide-equivalents of this clone displaced the binding of the parent phage and antagonized the binding of a sialyl Lewis(x) analogue to P-selectin. In conclusion, P-selectin expression correlates with early and advanced atherosclerotic lesion development. P-selectin ligands, like the lead structure we have developed here, can therefore be considered as promising tools to identify, target or antagonize P-selectin function within the chronically inflamed arterial wall.

Uptake and processing of modified bacteriophage M13 in mice: implications for phage display.

Internalization and degradation of filamentous bacteriophage M13 by a specific target cell may have major consequences for the recovery of phage in in vivo biopanning of phage libraries. Therefore, we investigated the pharmacokinetics and processing of native and receptor-targeted phage in mice. (35)S-radiolabeled M13 was chemically modified by conjugation of either galactose (lacM13) or succinic acid groups (sucM13) to the coat protein of the phage to stimulate uptake by galactose recognizing hepatic receptors and scavenger receptors, respectively. Receptor-mediated endocytosis of modified phage reduced the plasma half-life of native M13 (t(1/2) = 4.5 h) to 18 min for lactosylated and 1.5 min for succinylated bacterophage. Internalization of sucM13 was complete within 30 min after injection and resulted in up to 5000-fold reduction of bioactive phage within 90 min. In conclusion, these data provide information on the in vivo behavior of wild-type and receptor-targeted M13, which has important implications for future in vivo phage display experiments and for the potential use of M13 as a viral gene delivery vehicle.

Specific inhibition of P-selectin-mediated cell adhesion by phage display-derived peptide antagonists.

P-selectin is a leukocyte adhesion receptor expressed on activated vascular endothelium and platelets that mediates leukocyte rolling and attachment. Because P-selectin is critically involved in inflammation, we used phage display libraries to identify P-selectin-specific peptides that might interfere with its proinflammatory function. Isolated phage contained a highly conserved amino acid motif. Synthetic peptides showed calcium-dependent binding to P-selectin, with high selectivity over E-selectin and L-selectin. The peptides completely antagonized adhesion of monocyte-derived HL60 cells to P-selectin and increased their rolling velocities in flow chamber experiments. Peptide truncation and alanine-scanning studies indicated that an EWVDV (single-letter amino acid codes) consensus motif sufficed for effective inhibition. Intriguingly, the apparent avidity of the peptides was increased 200-fold when presented in a tetrameric form (2 microM versus 10 nM), which is consistent with the proposed divalent interaction of P-selectin glycoprotein ligand 1 (PSGL-1) with P-selectin. As the EWVDV peptides inhibit the binding of an established glycoside ligand for P-selectin (sulfated Lewis A), it is conceivable that EWVDV interacts with or in close proximity to the actual carbohydrate recognition domain of P-selectin, without being a direct structural mimic of sialyl Lewis(x). These ligands are among the most potent antagonists of P-selectin yet designed. Their high affinity, selectivity, and accessible synthesis provide a promising entry to the development of new anti-inflammatory therapeutics and might be a powerful tool to provide important information on the binding site of P-selectin.