National Jewish Health

TRIM21 is a multifunctional E3 ubiquitin ligase and intracellular antibody receptor, yet its role during viral infection remains unclear, with reports describing both antiviral and proviral activities. Here, we show that TRIM21 regulates influenza infection in an expression-dependent manner by functioning as a molecular rheostat rather than a binary restriction factor. This graded activity of TRIM21, which leads to both suppression and promotion of influenza replication, couples linkage-specific ubiquitination of viral nucleoprotein with modulation of innate immune signaling. Additionally, loss of TRIM21 unmasks a compensatory antiviral program centered on PRKDC, which is a ubiquitination target of TRIM21. This positions PRKDC as a latent restriction factor selectively engaged when primary TRIM21 control is lost. Together, these findings reveal a hierarchical and plastic antiviral network in which TRIM21 sets an adjustable threshold for host defense while restraining secondary restriction pathways. This framework highlights the sophisticated layers of regulation of the host ubiquitin-mediated antiviral immunity.

Progressive fibrosing interstitial lung diseases (PF-ILDs) have a large global impact and there is a pressing need to develop new therapies. We investigate if nintedanib augments apoptosis of pro-fibrotic fibroblasts induced by BCL-2 inhibition with ABT-199, and if combination therapy reverses pulmonary fibrosis. Healthy and PF-ILD fibroblasts were treated with nintedanib and BCL-2 family member expression was measured. Fibroblasts and precision cut lung slices (PCLS) were analyzed for apoptosis after ABT-199 and nintedanib treatment. Therapeutic intervention with ABT-199 and nintedanib in mice with repetitive bleomycin-induced progressive pulmonary fibrosis were assessed for pro-fibrotic fibroblast and aberrant transitional epithelial cell apoptosis, lung collagen content, longitudinal oxygen saturation and micro-CT disease burden. Nintedanib treatment increased expression of pro-apoptotic BIM and anti-apoptotic BCL-2 in PF-ILD primary fibroblasts. There was significantly increased apoptosis of fibroblasts in vitro after ABT-199. This was augmented after nintedanib co-treatment both in PF-ILD fibroblasts and PCLS. ABT-199 significantly improved fibrosis in mice, which was further enhanced after nintedanib co-treatment, by targeted apoptosis of pathogenic fibroblasts and transitional epithelial cells. The pro-apoptotic effects of ABT-199 were increased with nintedanib co-treatment and reversed fibrosis in mice. Combination treatment induced pro-fibrotic fibroblast and aberrant transitional epithelial cell apoptosis. These studies highlight a new mechanistic strategy to treat PF-ILD.

INTEGUMENT-PED/NCT04845620, a 4-week, phase 3 trial, demonstrated efficacy and safety of roflumilastcream 0.05% in children aged 2-5 years with mild-to-moderate atopic dermatitis (AD). A phase 3 open-label extensiontrial(INTEGUMENT-OLE [NCT04804605]) investigated long-term outcomes continuing roflumilast cream for ≤ 56 weeks. Caregivers of eligible patients from INTEGUMENT-PED applied roflumilast cream 0.05% once-daily in INTEGUMENT-OLE. Patients achieving Validated Investigator Global Assessment for AD (vIGA-AD) clear (0) at/afterweek4 of INTEGUMENT-OLE switched to twice-weekly (BIW) application. Evaluation of safety (adverse events [AEs] and application-site tolerability) was the primary objective. Efficacy endpoints assessed from INTEGUMENT-PED baseline included vIGA-AD success (clear/almost clear [0/1] plus ≥ 2-point improvement), vIGA-AD 0/1, ≥ 75% improvement in Eczema Area and Severity Index, and Worst Itch-Numeric Rating Scale success (≥ 4-point improvement in patients with baseline score ≥ 4). Duration of vIGA-AD 0/1 and sign/symptom control ("disease control") with BIW application was determined. Among 562 patients, 14 (2.5%) had treatment-related AEs reported. Roflumilast was well tolerated; application-site pain AEs were reported for four (0.7%) patients. At treatment-week 56, AD signs/symptoms continued to improve (63.1% of patients achieved vIGA-AD 0/1). Of 170 (30.2%) patients who switched to BIW application, the median Kaplan-Meier duration of "disease control" was 238 days. Roflumilast cream 0.05% demonstrated a favorable long-term safety profile and durable efficacy for up to 56 weeks of treatment in children aged 2-5 years with AD, and "disease control" was maintained with BIW application. These results are consistent with previous studies, including patients aged ≥ 6 years from INTEGUMENT-OLE. Clinicaltrials.gov identifier: NCT04845620.

Chromobox protein-7 (CBX7) functions as a gene repressor. We, unexpectedly, found that CBX7 formed a methylation-dependent transcriptional complex, which induced gene transcription by binding to cytokine gene promoters. CBX7 translocated to the cytosol and formed a methylation-dependent signaling complex with c-Raf, MAPK (mitogen-activated protein kinase) kinase 1/2 (MEK1/2), and casein kinase-2 (CK2)–α to generate and sustain extracellular signal–regulated kinase 1/2 (ERK1/2) signaling. CBX7 is an allergen-inducible gene. Genetic and pharmacologic interventions established an essential role for CBX7 for the production of cytokines by mouse and asthmatic patient lymphoid cells, and for the induction of allergic asthma in multiple mouse models. RNA sequencing demonstrated a large-scale loss and gain in gene transcription in Cbx7−/− T cells. The top down-regulated pathways included cytokine-cytokine receptor interaction, asthma, and T helper cell differentiation. CBX7 induction of the transcriptional activation complex and methylation of the ERK1/2 signalosome was specific for lymphoid cells as they were absent in epithelial cells. Our studies established a previously unknown paradigm of CBX7-generated methylation-dependent signaling complexes regulating inflammation.

Rationale Activated pulmonary CD4+ T lymphocytes are a critical part of sarcoidosis pathogenesis. Objectives To identify molecular changes associated with sarcoidosis risk primarily and progression secondarily, we profiled and integrated DNA methylome and transcriptome data in lung CD4+ T cells. Methods Bronchoalveolar lavage (BAL) CD4+ T cells were isolated from 29 sarcoidosis (16 progressive [SarcP] and 13 non-progressive [SarcNP]) cases and 19 healthy controls. mRNA was sequenced and DNA methylation profiled on Illumina HumanMethylationEPIC arrays. Linear models were fit to test for effect of diagnosis, adjusting for age and sex. Methylation models were adjusted for unknown batch effects, inflation and bias. We used Data Integration Analysis for Biomarker discovery using Latent cOmponents (DIABLO) to integrate methylome and transcriptome datasets and validated targets using single cell RNA-seq. Measurements and Main Results Using transcriptome-wide significance, differentially expressed (DE) transcripts were identified in sarcoidosis (39), SarcNP (23), and SarcP (7) compared to controls. Using genome-wide significance, 37 CpGs and 47 regions were identified as differentially methylated (DM) in sarcoidosis compared to controls. Focusing on DE genes, we identified DM CpGs in 11 genes associated with sarcoidosis, 3 with SarcNP and 7 with SarcP. DIABLO first latent component distinguished cases and controls based on transcripts and CpGs associated with T cell signaling, T-cell differentiation, apoptosis, and epigenetic regulation. Among DE/DM transcripts identified by integrative omic analysis and validated by single cell RNA-seq is IL18 receptor accessory protein ( IL18RAP ). Conclusions Integrative methylome–transcriptome profiling of lung CD4⁺ T cells identified coordinated immune and epigenetic alterations, including IL18RAP , as candidate biomarkers.