Interleukin-6 Signaling Research Articles

Neuronal growth regulator 1 may modulate interleukin-6 signaling in adipocytes.

Interleukin-6 (IL-6) is a pleiotropic cytokine that plays both anti- and pro-inflammatory roles. Due to the restricted expression of membrane IL-6 receptor (IL-6R), most pro-inflammatory functions of IL-6 are attributed to its association with soluble IL-6R (sIL-6R). Neuronal growth regulator 1 (NEGR1) is a brain-enriched membrane protein that has recently been recognized as a risk factor for many human diseases including obesity, depression, and autism. In the present study, we report that the expression levels of IL-6 and IL-6R, as well as the phosphorylation of signal transducer and activator of transcription (STAT) 3, were significantly elevated in white adipose tissues of Negr1 knockout mice. Elevated levels of circulating IL-6 and sIL-6R have also been observed in Negr1 -/- mice. Furthermore, NEGR1 interacted with IL-6R, which was supported by subcellular fractionation and an in situ proximity ligation assay. Importantly, NEGR1 expression attenuated the phosphorylation of STAT3 by sIL-6R, suggesting that NEGR1 negatively regulates IL-6 trans-signaling. Taken together, we propose that NEGR1 may play a regulatory role in IL-6 signaling by interacting with IL-6R, which may contribute to a molecular link underlying obesity, inflammation, and the depression cycle.

The RNA editor ADAR2 promotes immune cell trafficking by enhancing endothelial responses to interleukin-6 during sterile inflammation

Immune cell trafficking constitutes a fundamental component of immunological response to tissue injury, but the contribution of intrinsic RNA nucleotide modifications to this response remains elusive. We report that RNA editor ADAR2 exerts a tissue- and stress-specific regulation of endothelial responses to interleukin-6 (IL-6), which tightly controls leukocyte trafficking in IL-6-inflamed and ischemic tissues. Genetic ablation of ADAR2 from vascular endothelial cells diminished myeloid cell rolling and adhesion on vascular walls and reduced immune cell infiltration within ischemic tissues. ADAR2 was required in the endothelium for the expression of the IL-6 receptor subunit, IL-6 signal transducer (IL6ST; gp130), and subsequently, for IL-6 trans-signaling responses. ADAR2-induced adenosine-to-inosine RNA editing suppressed the Drosha-dependent primary microRNA processing, thereby overwriting the default endothelial transcriptional program to safeguard gp130 expression. This work demonstrates a role for ADAR2 epitranscriptional activity as a checkpoint in IL-6 trans-signaling and immune cell trafficking to sites of tissue injury.

Identification of hub genes and biological mechanisms underlying the pathogenesis of asthenozoospermia and chronic epididymitis.

Objective: Asthenozoospermia (AZS) is one of the most common causes of male fertility, affecting family wellbeing and population growth. Chronic epididymitis (CE) is a common and lingering inflammatory disease in the scrotum. Inflammation in the epididymis has a severe impact on sperm motility. This study aimed to explore the genetic profile and critical pathways involved in the pathological mechanisms of AZS and CE, and discover potential biomarkers. Methods: Genomic datasets of AZS and CE were obtained from the Gene Expression Omnibus (GEO) database, and relevant differentially expressed genes (DEGs) were identified. GO and pathway enrichment analyses, construction of a protein-protein interaction network, and receiver operator characteristic curve analysis were conducted. The expression profile of hub genes was validated in immunohistochemical data and testicular cell data. Immune infiltration, miRNA-hub gene interactions, and gene-disease interactions were explored. The mRNA levels of hub genes were further measured by qRT-PCR. Results: A total of 109 DEGs were identified between the AZS/CE and healthy control groups. Pathways of the immune system, neutrophil degranulation, and interleukin-4 and interleukin-13 signaling were enriched in AZS and CE. Five hub genes (CD300LB, CMKLR1, CCR4, B3GALT5, and CTSK) were selected, and their diagnostic values were validated in AZS, CE, and independent validation sets (area under the curve >0.7). Furthermore, the five-hub gene signature was well characterized in testicular immunohistochemical staining and testicular cells from healthy controls. Immune infiltration analysis showed that infiltration of CD8+ cells and T helper cells was significantly related to the expression level of five hub genes. In addition, a miRNA-hub gene network and interaction of other diseases were displayed. The mRNA levels of hub genes (CD300LB, CMKLR1, CCR4, and B3GALT5) were significantly elevated in the patient group. The mRNA level of CTSK also showed a similar trend. Conclusion: Our study uncovered the genetic profile involved in AZS and CE, and elucidated enriched pathways and molecular associations between hub genes and immune infiltration. This finding provides novel insight into the common pathogenesis of both diseases as well as the potential biomarkers for CE-associated AZS.

Targeting IL-6 trans-signalling: past, present and future prospects.

Interleukin-6 (IL-6) is a key immunomodulatory cytokine that affects the pathogenesis of diverse diseases, including autoimmune diseases, chronic inflammatory conditions and cancer. Classical IL-6 signalling involves the binding of IL-6 to the membrane-bound IL-6 receptor α-subunit (hereafter termed 'mIL-6R') and glycoprotein 130 (gp130) signal-transducing subunit. By contrast, in IL-6 trans-signalling, complexes of IL-6 and the soluble form of IL-6 receptor (sIL-6R) signal via membrane-bound gp130. A third mode of IL-6 signalling - known as cluster signalling - involves preformed complexes of membrane-bound IL-6-mIL-6R on one cell activating gp130 subunits on target cells. Antibodies and small molecules have been developed that block all three forms of IL-6 signalling, but in the past decade, IL-6 trans-signalling has emerged as the predominant pathway by which IL-6 promotes disease pathogenesis. The first selective inhibitor of IL-6 trans-signalling, sgp130, has shown therapeutic potential in various preclinical models of disease and olamkicept, a sgp130Fc variant, had promising results in phase II clinical studies for inflammatory bowel disease. Technological developments have already led to next-generation sgp130 variants with increased affinity and selectivity towards IL-6 trans-signalling, along with indirect strategies to block IL-6 trans-signalling. Here, we summarize our current understanding of the biological outcomes of IL-6-mediated signalling and the potential for targeting this pathway in the clinic.

Shared Molecular Signatures Across Zika Virus Infection and Multiple Sclerosis Highlight AP-1 Transcription Factor as a Potential Player in Post-ZIKV MS-Like Phenotypes.

Zika virus (ZIKV) is an arbovirus of the Flaviviridae genus that has rapidly disseminated from across the Pacific to the Americas. Robust evidence has indicated a crucial role of ZIKV in congenital virus syndrome, including neonatal microcephaly. Moreover, emerging evidence suggests an association between ZIKV infection and the development of an extensive spectrum of central nervous system inflammatory demyelinating diseases (CNS IDD), such as multiple sclerosis-like clinical phenotypes. However, the underlying mechanisms of host-pathogen neuro-immune interactions remain to be elucidated. This study aimed to identify common transcriptional signatures between multiple sclerosis (MS) and ZIKV infection to generate molecular interaction networks, thereby leading to the identification of deregulated processes and pathways, which could give an insight of these underlying molecular mechanisms. Our investigation included publicly available transcriptomic data from MS patients in either relapse or remission (RR-MS) and datasets of subjects acutely infected by ZIKV for both immune peripheral cells and central nervous system cells. The protein-protein interaction (PPI) analysis showed upregulated AP-1 transcription factors (JUN and FOS) among the top hub and bottleneck genes in RR-MS and ZIKV data. Gene enrichment analysis retrieved a remarkable presence of ontologies and pathways linked to oxidative stress responses, immune cell function, inflammation, interleukin signaling, cell division, and transcriptional regulation commonly enriched in both scenarios. Considering the recent findings concerning AP-1 function in immunological tolerance breakdown, regulation of inflammation, and its function as an oxidative stress sensor, we postulate that the ZIKV trigger may contribute as a boost for the activation of such AP-1-regulated mechanisms that could favor the development of MS-like phenotypes following ZIKV infection in a genetically susceptible individual.

IL-6 prevents Th2 cell polarization by promoting SOCS3-dependent suppression of IL-2 signaling

Defective interleukin-6 (IL-6) signaling has been associated with Th2 bias and elevated IgE levels. However, the underlying mechanism by which IL-6 prevents the development of Th2-driven diseases remains unknown. Using a model of house dust mite (HDM)-induced Th2 cell differentiation and allergic airway inflammation, we showed that IL-6 signaling in allergen-specific T cells was required to prevent Th2 cell differentiation and the subsequent IgE response and allergic inflammation. Th2 cell lineage commitment required strong sustained IL-2 signaling. We found that IL-6 turned off IL-2 signaling during early T-cell activation and thus inhibited Th2 priming. Mechanistically, IL-6-driven inhibition of IL-2 signaling in responding T cells was mediated by upregulation of Suppressor Of Cytokine Signaling 3 (SOCS3). This mechanism could be mimicked by pharmacological Janus Kinase-1 (JAK1) inhibition. Collectively, our results identify an unrecognized mechanism that prevents the development of unwanted Th2 cell responses and associated diseases and outline potential preventive interventions.

RETRACTED ARTICLE: Qilongtian ameliorate bleomycin-induced pulmonary fibrosis in mice via inhibiting IL-17 signal pathway

Pulmonary fibrosis (PF) is a special type of pulmonary parenchymal disease, with chronic, progressive, fibrosis, and high mortality. There is a lack of safe, effective, and affordable treatment methods. Qilongtian (QLT) is a traditional Chinese prescription that is composed of Panax notoginseng, Earthworm, and Rhodiola, and shows the remarkable clinical curative effect of PF. However, the mechanism of QLT remains to be clarified. Therefore, we studied the effectivity of QLT in treating Bleomycin (BLM) induced PF mice. 36 C57BL/6 J mice were randomized into the control group, the model group, the low-, medium- and high-dose QLT group, and Pirfenidone group. After establishing a model of pulmonary fibrosis in mice, the control and model groups were infused with a normal saline solution, and the delivery group was infused with QLT. Pulmonary function in the mice from each group was detected. Pulmonary tissue morphologies and collagen deposition were stained by HE and Masson. The content of hydroxyproline (HYP) was detected by alkaline hydrolysis and the mRNA and protein expression of related genes in pulmonary tissues were detected by using q-PCR, ELISA, and Western blot. Our studies have shown that QLT significantly reduced the inflammatory injury, hydroxy-proline content, and collagen deposition of pulmonary tissue in BLM-induced PF mice and down-regulated the cytokine related to inflammation and fibrosis and PF expression on the mRNA and protein level in PF mice. To identify the mechanism of QLT, the Transcriptome was measured and the IL-17 signal pathway was screened out for further research. Further studies indicated that QLT reduced the mRNAs and protein levels of interleukin 17 (IL-17), c–c motif chemokine ligand 12 (CCL12), c-x-c motif chemokine ligand 5 (CXCL5), fos-like antigen 1 (FOSL1), matrix metalloproteinase-9 (MMP9), and amphiregulin (AREG), which are inflammation and fibrosis-related genes in the IL-17 signal pathway. The results indicated that the potential mechanism for QLT in the prevention of PF progression was by inhibiting inflammation resulting in the IL-17 signal pathway. Our study provides the novel scientific basis of QLT and represents new therapeutics for PF in clinical.

Identifying chronic obstructive pulmonary disease from integrative omics and clustering in lung tissue

BackgroundChronic obstructive pulmonary disease (COPD) is a highly morbid and heterogenous disease. While COPD is defined by spirometry, many COPD characteristics are seen in cigarette smokers with normal spirometry. The extent to which COPD and COPD heterogeneity is captured in omics of lung tissue is not known.MethodsWe clustered gene expression and methylation data in 78 lung tissue samples from former smokers with normal lung function or severe COPD. We applied two integrative omics clustering methods: (1) Similarity Network Fusion (SNF) and (2) Entropy-Based Consensus Clustering (ECC).ResultsSNF clusters were not significantly different by the percentage of COPD cases (48.8% vs. 68.6%, p = 0.13), though were different according to median forced expiratory volume in one second (FEV1) % predicted (82 vs. 31, p = 0.017). In contrast, the ECC clusters showed stronger evidence of separation by COPD case status (48.2% vs. 81.8%, p = 0.013) and similar stratification by median FEV1% predicted (82 vs. 30.5, p = 0.0059). ECC clusters using both gene expression and methylation were identical to the ECC clustering solution generated using methylation data alone. Both methods selected clusters with differentially expressed transcripts enriched for interleukin signaling and immunoregulatory interactions between lymphoid and non-lymphoid cells.ConclusionsUnsupervised clustering analysis from integrated gene expression and methylation data in lung tissue resulted in clusters with modest concordance with COPD, though were enriched in pathways potentially contributing to COPD-related pathology and heterogeneity.

Design of cell-type-specific hyperstable IL-4 mimetics via modular de novo scaffolds.

The interleukin-4 (IL-4) cytokine plays a critical role in modulating immune homeostasis. Although there is great interest in harnessing this cytokine as a therapeutic in natural or engineered formats, the clinical potential of native IL-4 is limited by its instability and pleiotropic actions. Here, we design IL-4 cytokine mimetics (denoted Neo-4) based on a de novo engineered IL-2 mimetic scaffold and demonstrate that these cytokines can recapitulate physiological functions of IL-4 in cellular and animal models. In contrast with natural IL-4, Neo-4 is hyperstable and signals exclusively through the type I IL-4 receptor complex, providing previously inaccessible insights into differential IL-4 signaling through type I versus type II receptors. Because of their hyperstability, our computationally designed mimetics can directly incorporate into sophisticated biomaterials that require heat processing, such as three-dimensional-printed scaffolds. Neo-4 should be broadly useful for interrogating IL-4 biology, and the design workflow will inform targeted cytokine therapeutic development.

Purification and identification of anti-inflammatory peptides from sturgeon (Acipenser schrenckii) cartilage

Cartilage is a nonedible byproduct with little saleable value. However, previous studies have proposed the possibility of producing peptides from cartilage with immune function modulation potential. The current study aimed to investigate the potential anti-inflammatory activity of peptides derived from sturgeon (Acipenser schrenckii) cartilage in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. Five peptide sequences, including four novel peptides, were identified from ethanol-soluble cartilage hydrolysates. Among these five peptides, LTGP, LLLE, LLEL and VGPAGPAGP reduced the production of nitric oxide (NO) and interleukin-6 (IL-6) while increasing interleukin-10 (IL-10) excretion. Transcriptome analysis suggested the inhibition of activated mitogen-activated protein kinase (MAPK) and interleukin-17 (IL-17) signaling pathways after LLEL intervention. MAPK, which is involved in the IL-17 signaling pathway, was further proved to be blocked by downregulating the phosphorylation of p38, extracellular-signal regulated protein kinase (ERK), and c-jun N-terminal kinase (JNK). This novel peptide offers an attractive approach to develop functional foods.

Abstract 4090: Regional administration of IL-12 endowed CAR T cells effectively targets systemic disease

Abstract While chimeric antigen receptor (CAR) T cells have received FDA approvals in treating hematological malignancies, clinical responses of CAR T cells for solid tumors have been underwhelming. Intense investigation is underway to improve CAR T cells trafficking, persistence and efficacy. Interleukin-12 (IL-12) is a potent inflammatory cytokine that recruits and activates various endogenous and adoptively transferred immune cells, but development of its clinical applications is hindered by toxicity of systemic IL-12 signaling. In this study, we engineered and expressed membrane-bound IL-12 (mbIL12) in CAR T cells to overcome current limitations in CAR T cell and IL-12 therapies. We evaluated CAR T cell function using co-culture assays and preclinical murine models bearing human tumor xenografts. We modeled systemic metastasis of ovarian and breast cancer and assessed the impact of mbIL12 in CAR T cells on targeting regional and systemic diseases. Syngeneic mouse models were used to evaluate toxicity induced by mbIL12 in CAR T cells. We observed a greater number of CAR T cells in the peripheral blood of mice receiving locoregionally delivered CAR T cells engineered with mbIL12, suggesting that mbIL12 improves CAR T cell trafficking and persistence. Furthermore, mice bearing regional and systemic diseases experienced more potent and durable anti-tumor activity when treated with mbIL12 endowed CAR T cells. These results were mirrored in vitro where mbIL12 improved CAR T cells activation, proliferation and IFNg secretion when co-cultured with tumor cells, recapitulating our findings in vivo. Toxicities observed in immune-competent mice receiving systemic IL-12 delivery were not detected in those administered CAR T cells with mbIL12. We demonstrated that mbIL12 enhances anti-tumor function of CAR T cells for treatment of solid tumors. This study also showed that mbIL12 is safe, providing a translatable engineering strategy to improve CAR T cell therapy. IL-12 signaling is well known to modulate function of various immune cells, but the impact of CAR T cells with mbIL12 on immune tumor microenvironments remains to be elucidated. We are currently investigating changes in the immune landscape that CAR T cells with mbIL12 induce. Citation Format: Hee Jun Lee, Yuki Yamaguchi, Jackson Gibson, Cody Cullen, John P. Murad, Anthony K. Park, Isabel Monroy, Cari Young, Lea Christian, Lauren Adkins, Lawrence Stern, Wen-Chung Chang, Catalina Martinez, Stephen J. Forman, Saul J. Priceman. Regional administration of IL-12 endowed CAR T cells effectively targets systemic disease. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4090.

Abstract 6762: Tumor immune microenvironment &genomic features of non-small cell lung carcinomas in patients with HIV (PWH)

Abstract Introduction: Lung cancer is the leading cause of cancer-related death among PWH with a relative risk that is over 3-fold higher compared to the uninfected population. In addition, HIV-infected patients with non-small cell lung cancer (NSCLC) have a significantly worse prognosis. The determinants for these associations remain uncertain. Methods: We investigated the immune contexture of 18 primary NSCLCs from PWH and 19 clinicopathologically matched NSCLCs from HIV-negative cases represented in tissue microarrays using 37-plex Imaging Mass Cytometry (IMC) panels including cell phenotype markers (CK,CD4,CD8,CD20,CD45RO,CD25,etc), immunomodulatory targets (PD-L1,PD-L2,B7-H3,B7-H4,B2M,etc) and DNA damage response/repair (DDR) indicators (γH2AX, RAD51,BRCA1,53BP1,etc). The samples were analyzed using single-cell tissue segmentation and spatial analysis. A subset of cases was also studied using whole exome DNA sequencing (WES) and mRNA sequencing (RNAseq). Results: NSCLCs from PWH showed comparable levels of tumor infiltrating lymphocytes (TILs) relative to HIV-negative cases with a trend towards a lower CD4+/CD8+ cell ratio. However, CD4+ and CD8+ TILs in PWH showed higher levels of markers associated with T-cell activation and dysfunction including GZB, CD25, Ki-67, PD-1, LAG-3, and TIM-3 than non-HIV tumors. The epithelial tumor cells in HIV-associated cases showed higher levels of the immunomodulatory ligands PD-L1, CD47, B7-H4 and VISTA; and altered levels of the DDR markers γH2AX, BRCA1 and 53BP1 relative to the non-HIV controls. Differences were also seen in tumor-associated macrophages consistent with an immune regulatory environment. The spatial analysis of tumor and immune cells revealed a distinct topographic distribution of these cells in HIV-associated malignancies. WES analysis showed a higher nonsynonymous tumor mutational burden in HIV-associated tumors (400 vs 202 mean mutations/exome) with increased predicted HLA class-I neoantigens and a numerically higher immunoediting score than the non-HIV group. Exploratory analysis identified enrichment in RYR2 and KIAA1671 gene mutations in HIV-associated malignancies. RNAseq revealed distinct mRNA expression profiles of HIV-positive and negative NSCLCs with a fraction of transcripts showing differential expression associated with tumor suppressor functions, interleukin signaling and DDR. Conclusion: NSCLCs in PWH display profound immunological and genomic alterations suggesting defective immunosurveillance. Our study reveals novel characteristics of HIV-associated NSCLC with possible clinical implications for this growing and understudied population. Citation Format: Shruti Desai, Kishu Ranjan, Syim Salahuddin, Ramsey Yusuf, Jianlei Gu, Daiwei Tang, Yong Kong, Barani Kumar Rajendran, Hongyu Zhao, Yuval Kluger, Sarah Goldberg, Brinda Emu, Kurt Alex Schalper. Tumor immune microenvironment &genomic features of non-small cell lung carcinomas in patients with HIV (PWH) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6762.

Abstract 525: Targeting interleukin-1 receptor-associated kinase 1/4 in acute myeloid leukemia stem cells

Abstract Therapies for acute myeloid leukemia (AML) patients are suboptimal due mainly to relapse of disease after treatment. Leukemia stem cell (LSC) populations maintain the disease and serve as a reservoir for relapse. Therefore, treatments that can eradicate LSCs will improve the outcomes of patients with AML. Our preliminary data have shown that the interleukin-1 (IL-1) pathway is upregulated following treatment and disease progression in AML and plays an important role in LSCs in vitro and in vivo. To target IL-1 signaling, we disclose the structure-activity relationship guided development of UR241-2, a novel small molecule compound of interleukin-1 receptor-associated kinase 1/4 (IRAK1/4), which are critical downstream factors in IL-1/Toll-Like Receptor (TLR) signaling. UR241-2, at single dose (5 nM) screening exhibited high target selectivity; other than IRAK1/4 only 9 kinases in a panel of 682 kinases were inhibited by >25% activity. The NanoBRET assay using HEK293 cells transfected with targeted kinase vectors showed notably selective target engagement of UR241-2 against IRAK4 (IC50 = 6.07 nM). By using the in vitro NF-kB reporter assay and immunoblotting analysis, we identified that UR241-2 robustly inhibits IL-1/TLR signaling in AML cells including the activation of NF-κB and the phosphorylation of P65 and P38, following IL-1 stimulation. Next, we treated murine MLL-AF9 AML cells and human MDSL cells with UR241-2 and two IRAK4 inhibitors (PF-06650833 and CA-4948) that are currently in clinical or preclinical studies. We found that UR241-2 significantly inhibits colony-forming ability in AML and myelodysplastic syndromes, with similar or better efficacy compared to PF-06650833 or CA-4948. We also determined that UR241-2 suppresses LSC clonogenicity in primary human AML cells at diagnosis and in relapse while minimally impacting normal hematopoietic stem and progenitor cell function. Last, we investigated the impacts of UR241-2 on LSC engrafted ability in vivo. We treated murine MLL-AF9 cells with UR241-2 and PF-06650833 ex vivo for three days and transplanted them into recipient mice. After four weeks, we observed that UR241-2 significantly suppresses the engraftment levels of AML cells in bone marrow. It also decreases the WBC in peripheral blood and increases the RBC, HGB and PLT counts compared to the vehicle control group. Taken together, our findings demonstrated that targeting IRAK1/4 using the novel compound UR241-2 is able to impair LSC clonogenicity and engraftment capability in AML following treatment and disease progression. This provides a potential therapeutic strategy to eradicate the LSC population and improve the outcome of AML patients. Citation Format: Tzu-Chieh Ho, Rakesh K. Singh, Hiroki Kawano, Mark W. LaMere, Jian Wang, Paula M. Vertino, Nikolay V. Dokholyan, Laura M. Calvi, Craig T. Jordan, Michael W. Becker. Targeting interleukin-1 receptor-associated kinase 1/4 in acute myeloid leukemia stem cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 525.

Abstract 2569: Investigation of the effect of chronic IL-1 exposure on breast cancer cell line response to external signaling factors

Abstract As part of our immune response, the body produces anti-tumorigenic inflammatory proteins to kill cancer cells. However, cancer cells can usurp chronic inflammation for their own survival. Interleukin-1 (IL-1) is an inflammatory protein found in breast cancer (BCa) tumors that is elevated in the BCa patient tumors and serum and promotes metastasis, indicating IL-1 is clinically relevant. But the molecular mechanisms underlying the effect of chronic IL-1 on BCa progression remain unknown. We previously discovered that acute exposure to IL-1 represses essential hormone receptors in cancers cells, but concomitantly upregulates other compensatory pro-survival proteins that contribute to tumor progression. To determine if chronic IL-1 inflammation has the same effect, we generated cell subline models using the human breast cancer cell line, T47D, by exposing T47D cells chronically to IL-1 for several months. Notably, we found that the T47D chronic IL-1 sublines stably evolve insensitivity to exogenous IL-1. This result suggests that chronic IL-1 causes stable molecular changes that may affect cancer cell response to exogenous factors.Therapeutic strategies include blocking exogenous hormone or growth factor signals in the tumor microenvironment to attenuate cancer cell growth, proliferation, and survival. Therefore, we grew T47D parental and chronic IL-1 sublines in the absence of serum to determine the effect of chronic IL-1 exposure on BCa cell response to hormone and growth factor inhibition. Preliminarily, we do not observe differences in parental versus subline response to serum starvation, suggesting that differential response to exogenous factors is specific to IL-1 signaling. To better understand how chronic IL-1 -induced molecular changes effect BCa cell response to the tumor microenvironment, we will perform sequencing and bioinformatics analysis to identify IL-1 -dependent and/or independent signaling pathways stably altered in the chronic IL-1 sublines. Citation Format: Roopal Dhar, Ola M. Olaleye, Rana Tarig Abdelaziz, Jyotsna Tera Reddy, Nikki Delk. Investigation of the effect of chronic IL-1 exposure on breast cancer cell line response to external signaling factors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2569.

Abstract 3864: Targeting IL-6/STAT3 signaling in EGFR-mutant drug tolerant persister cells

Abstract While the majority of epidermal growth factor receptor (EGFR) mutant non-small cell lung cancer (NSCLC) patients initially respond to treatment with an EGFR tyrosine kinase inhibitor (TKI), most will eventually develop acquired resistance to these agents. The evolution of tumor cells with acquired resistance to targeted agents initially involves the persistence of a drug-tolerant subpopulation of tumor cells which display a transient and flexible resistant state. These persister cells can remain quiescent for prolonged periods of time, but eventually give rise to fully drug resistant tumor cells that resume growth and metastatic spread. The signaling pathways that allow persister cells to remain viable or to re-activate expansive growth are poorly understood. We and others have reported that EGFR TKI resistance that occurs independent of secondary EGFR mutations or MET amplification is associated with an epithelial to mesenchymal transition (EMT). Our analysis of tumor cells that had undergone EMT-mediated resistance to the EGFR TKI osimertinib revealed that interleukin-6 (IL-6) was highly upregulated in resistant cells as compared to parental cells. We find that in the setting of EGFR mutant NSCLC, IL-6 suppresses the anti-tumor cell activity of T cells and NK cells. To assess the role of IL-6 in the early events of EGFR TKI resistance, we generated drug-tolerant persister cells by treating EGFR mutant HCC4006 and HCC827 NSCLC cells with the EGFR TKI osimertinib for 10 days and then assessed IL-6 expression. We observed that IL-6 was upregulated at both the RNA and protein levels in EGFR-TKI persister cells as compared to parental cells. While the cell surface expression of IL6R was not significantly increased in EGFR-TKI persister cells as compared to parental cells, we observed increased secretion of soluble IL6R in persister cells by ELISA assay. We next assessed changes in protein expression and activation in drug tolerant persister cells by reverse phase protein array (RPPA) and observed that p-STAT3, a key component of the IL-6 signaling pathway, was significantly elevated in persister cells as compared parental cells or cells treated with osimertinib for 24 hours. Using genetically engineered mouse models (GEMM) of EGFRL858R mutant lung cancer, we also observed a similar upregulation of both IL-6 and p-STAT3 expression at the RNA and protein levels in EGFR-mutant tumors treated with osimertinib for 2 or 4 weeks. Additionally, circulating levels of IL-6 also are elevated in EGFRL858R GEMMs treated with osimertinib for 2 or 4 weeks. Collectively, our data indicates that upregulation of IL-6 is an early event in the evolution of drug resistance. Given the role of IL-6 in promoting EGFR TKI resistance, further studies investigating whether targeting of IL-6 signaling may impair the survival or outgrowth of persister populations are warranted. Citation Format: Sonia A. Patel, Monique Nilsson, Yan Yang, Li Shen, Jing Wang, Alissa Poteete, Xiaoxing Yu, Xiaoyang Ren, Xiuning Le, John Heymach. Targeting IL-6/STAT3 signaling in EGFR-mutant drug tolerant persister cells. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3864.

Src-FAK Signaling Mediates Interleukin 6-Induced HCT116 Colorectal Cancer Epithelial–Mesenchymal Transition

Colorectal cancer is one of the most prevalent and lethal malignancies, affecting approximately 900,000 individuals each year worldwide. Patients with colorectal cancer are found with elevated serum interleukin-6 (IL-6), which is associated with advanced tumor grades and is related to their poor survival outcomes. Although IL-6 is recognized as a potent inducer of colorectal cancer progression, the detail mechanisms underlying IL-6-induced colorectal cancer epithelial-mesenchymal transition (EMT), one of the major process of tumor metastasis, remain unclear. In the present study, we investigated the regulatory role of IL-6 signaling in colorectal cancer EMT using HCT116 human colorectal cancer cells. We noted that the expression of epithelial marker E-cadherin was reduced in HCT116 cells exposed to IL-6, along with the increase in a set of mesenchymal cell markers including vimentin and α-smooth muscle actin (α-SMA), as well as EMT transcription regulators-twist, snail and slug. The changes of EMT phenotype were related to the activation of Src, FAK, ERK1/2, p38 mitogen-activated protein kinase (p38MAPK), as well as transcription factors STAT3, κB and C/EBPβ. IL-6 treatment has promoted the recruitment of STAT3, κB and C/EBPβ toward the Twist promoter region. Furthermore, the Src-FAK signaling blockade resulted in the decline of IL-6 induced activation of ERK1/2, p38MAPK, κB, C/EBPβ and STAT3, as well as the decreasing mesenchymal state of HCT116 cells. These results suggested that IL-6 activates the Src-FAK-ERK/p38MAPK signaling cascade to cause the EMT of colorectal cancer cells. Pharmacological approaches targeting Src-FAK signaling may provide potential therapeutic strategies for rescuing colorectal cancer progression.

Oral feeding of nanoplastics affects brain function of mice by inducing macrophage IL-1 signal in the intestine

Nanoplastics (NPs) as contaminants in food and water have drawn increasing public attention. However, little is known about how NPs shape the gut immune landscape after injection. In this study, we fabricate NPs (∼500nm) and microplastics (MPs) (∼2μm) and evaluate their invivo effects by feeding them to mice. The results suggest that NPs show a better ability to induce gut macrophage activation than MPs. In addition, NPs trigger gut interleukin-1 (IL-1)-producing macrophage reprogramming via inducing lysosomal damage. More importantly, IL-1 signaling from the intestine can affect brain immunity, leading to microglial activation and Th17 differentiation, all of which correlates with a decline in cognitive and short-term memory in NP-fed mice. Thus, this study provides insight into the mechanism of action of the gut-brain axis, delineates the way NPs reduce brain function, and highlights the importance of fixing the plastic pollution problem worldwide.

Alterations In The Interleukin-1 Signaling Pathway Contribute To Increased Spontaneous Pain In Spinal Cord Injury

Alterations In The Interleukin-1 Signaling Pathway Contribute To Increased Spontaneous Pain In Spinal Cord Injury

The pathogenetic influence of smoking on SARS-CoV-2 infection: Integrative transcriptome and regulomics analysis of lung epithelial cells

Corona virus disease (COVID-19) has been emerged as pandemic infectious disease. The recent epidemiological data suggest that the smokers are more vulnerable to infection with COVID-19; however, the influence of smoking (SMK) on the COVID-19 infected patients and the mortality is not known yet. In this study, we aimed to discern the influence of SMK on COVID-19 infected patients utilizing the transcriptomics data of COVID-19 infected lung epithelial cells and transcriptomics data smoking matched with controls from lung epithelial cells. The bioinformatics based analysis revealed the molecular insights into the level of transcriptional changes and pathways which are important to identify the impact of smoking on COVID-19 infection and prevalence. We compared differentially expressed genes (DEGs) between COVID-19 and SMK and 59 DEGs were identified as consistently dysregulated at transcriptomics levels. The correlation network analyses were constructed for these common genes using WGCNA R package to see the relationship among these genes. Integration of DEGs with network analysis (protein–protein interaction) showed the presence of 9 hub proteins as key so called ”candidate hub proteins” overlapped between COVID-19 patients and SMK. The Gene Ontology and pathways analysis demonstrated the enrichment of inflammatory pathway such as IL-17 signaling pathway, Interleukin-6 signaling, TNF signaling pathway and MAPK1/MAPK3 signaling pathways that might be the therapeutic targets in COVID-19 for smoking persons. The identified genes, pathways, hubs genes, and their regulators might be considered for establishment of key genes and drug targets for SMK and COVID-19.

Improvement of Suspected Eosinophilic Otitis Media with Targeted Biologic Therapy.

To compare the responses of suspected eosinophilic otitis media to treatment with or without a targeted biologic therapy against interleukin-4 (IL-4), IL-5, or IL-13 signaling. Retrospective review. Tertiary referral center. Subjects with type 2 chronic rhinosinusitis with nasal polyposis (CRSwNP), asthma, and otitis media who underwent treatment between 2005 and 2021. Treatment with targeted biologic therapy. Pre- and posttreatment nasal endoscopy, ear examination, and audiologic evaluation. Four hundred seventy-seven subjects with type 2 CRSwNP were treated between 2005 and 2021. Sixty-two had otitis media with pre- and posttreatment evaluation. Retrospective chart review assessed pre- and posttreatment exam findings, nasal endoscopy, audiometry, and tympanometry. Nineteen subjects received a biologic therapy, whereas 43 did not. Exam, endoscopy, and tympanometry were graded for severity and compared pre- and posttreatment. Subjective ear exam and tympanometry were significantly improved with biologic therapy (control = 0.05, biologic = 0.84, p = 9.3 × 10 -5 ; control = -0.1, biologic = 0.62, p = 0.0002). Conductive hearing loss as assessed by air-bone gaps did not change between groups (control = 1.2 dB better, biologic = 1.2 dB worse, p = 0.32). Nasal endoscopy findings improved with biologic therapy relative to the control group, although not statistically significant (control = 1.04, biologic = 1.36, p = 0.22). Biologic therapies targeting interleukin-4 (IL-4), IL-5, and IL-13 signaling are potential new treatments for eosinophilic otitis media. This is the largest study demonstrating improvement in subjects with suspected eosinophilic otitis media in response to biologic therapy, and immune modulation represents a novel treatment strategy for this challenging condition. Current treatment strategies for otologic symptoms in eosinophilic disease are not tremendously effective or durable, resulting in a need for improved treatment options. To determine if targeted biologic therapy, often used for eosinophilic asthma and type 2 chronic rhinosinusitis with nasal polyposis, improves coexistent suspected eosinophilic otitis media. Treatment of suspected eosinophilic otitis media with targeted biologic therapy will result in improvement of otologic symptoms with a durable response compared with current treatment options. Level IV. Exempt. HUM00182703.