Airway mucins promote immunopathology in virus-exacerbated chronic obstructive pulmonary disease

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Abstract

The respiratory tract surface is protected from inhaled pathogens by a secreted layer of mucus rich in mucin glycoproteins. Abnormal mucus accumulation is a cardinal feature of chronic respiratory diseases but the relationship between mucus and pathogens during exacerbations is poorly understood. We identified elevations in airway MUC5AC and MUC5B concentrations during spontaneous and experimentally-induced chronic obstructive pulmonary disease (COPD) exacerbations. MUC5AC was more sensitive to changes in expression during exacerbation and was therefore more predictably associated with virus load, inflammation, symptom severity, decrements in lung function, and secondary bacterial infections. MUC5AC was functionally related to inflammation as Muc5ac-deficient (Muc5ac-/-) mice had attenuated rhinovirus (RV)–induced airway inflammation and exogenous MUC5AC glycoprotein administration augmented inflammatory responses and increased release of extracellular adenosine triphosphate (ATP) in mice and human airway epithelial cell cultures. Hydrolysis of ATP suppressed MUC5AC augmentation of rhinovirus-induced inflammation in mice. Therapeutic suppression of mucin production using an epidermal growth factor receptor (EGFR) antagonist ameliorated immunopathology in a mouse COPD exacerbation model. The coordinated virus induction of MUC5AC and MUC5B suggests that non-Th2 mechanisms trigger mucin hypersecretion during exacerbations. Our data identifies a pro-inflammatory role for MUC5AC during viral infection and suggest that MUC5AC inhibition may ameliorate COPD exacerbations.

Authors

Aran Singanayagam, Joseph Footitt, Matthias Marczynski, Giorgia Radicioni, Michael T. Cross, Lydia J. Finney, Maria-Belen Trujillo-Torralbo, Maria Adelaide Calderazzo, Jie Zhu, Julia Aniscenko, Thomas B. Clarke, Philip L. Molyneaux, Nathan W. Bartlett, Miriam F. Moffatt, William O. Cookson, Jadwiga A. Wedzicha, Christopher M. Evans, Richard C. Boucher, Mehmet Kesimer, Oliver Lieleg, Patrick Mallia, Sebastian L. Johnston

Targeting HIF-1α abrogates PD-L1-mediated immune evasion in tumor microenvironment but promotes tolerance in normal tissues

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Abstract

Anti-CTLA-4 + anti-PD-1/PD-L1 combination is the most effective cancer immunotherapy but causes high incidence of immune-related adverse events (irAE). Here we report that targeting of HIF-1α suppressed PD-L1 expression on tumor cells and tumor-infiltrated myeloid cells, but unexpectedly induced PD-L1 in normal tissues by an IFNγ–dependent mechanism. Targeting the HIF-1α-PD-L1 axis in tumor cells reactivated tumor-infiltrating lymphocytes (TILs) and caused tumor rejection. The HIF-1α inhibitor echinomycin potentiated cancer immunotherapeutic effects of anti-CTLA-4 therapy with efficacy comparable to anti-CTLA-4+anti-PD-1 antibodies. However, while anti-PD-1 exacerbated irAE triggered by Ipilimumab, echinomycin protected mice against irAE by increasing PD-L1 levels in normal tissues. Our data suggest that targeting HIF-1α fortifies the immune tolerance function of the PD-1:PD-L1 checkpoint in normal tissues but abrogates its immune evasion function in the tumor microenvironment (TME) to achieve safer and more effective immunotherapy.

Authors

Christopher M. Bailey, Yan Liu, Mingyue Liu, Xuexiang Du, Martin Devenport, Pan Zheng, Yang Liu, Yin Wang

Up-regulated YB-1 protein promotes glioblastoma growth through an YB-1/CCT4/mLST8/mTOR pathway

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Abstract

The Y-box binding protein 1 (YB-1) is a multi-functional RNA binding protein involved in virtually each step of RNA metabolism. However, the functions and mechanisms of YB-1 in one of the most aggressive cancers, glioblastoma, are not well understood. In this study, we identified that YB-1 protein was markedly overexpressed in glioblastoma and acted as a critical activator of both mTORC1 and mTORC2 signaling. Mechanistically, YB-1 bound the 5’ untranslated region (UTR) of the CCT4 mRNA to promote the translation of CCT4, a component of CCT chaperone complex, that in turn activated the mTOR signal pathway by promoting mLST8 folding. In addition, YB-1 autoregulated its own translation by binding to its 5’ UTR, leading to sustained activation of mTOR signaling. In glioblastoma patients, the protein level of YB-1 positively correlated with CCT4 and mLST8 expression as well as activated mTOR signaling. Importantly, the administration of RNA decoys specifically targeting YB-1 in a mouse xenograft model resulted in slower tumor growth and better survival. Taken together, these findings uncover a disrupted proteostasis pathway involving YB-1/CCT4/mLST8/mTOR axis in promoting glioblastoma growth, suggesting that YB-1 is a potential therapeutic target for the treatment of glioblastoma.

Authors

Jin-Zhu Wang, Hong Zhu, Pu You, Hui Liu, Wei-Kang Wang, Xiaojuan Fan, Yun Yang, Keren Xu, Yingfeng Zhu, Qunyi Li, Ping Wu, Chao Peng, Catherine C.L. Wong, Kaicheng Li, Yufeng Shi, Nu Zhang, Xiuxing Wang, Rong Zeng, Ying Huang, Liusong Yang, Zefeng Wang, Jingyi Hui

Interleukin-10 contributes to reservoir establishment and persistence in SIV-infected macaques treated with antiretroviral therapy

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Abstract

Interleukin (IL)-10 is an immunosuppressive cytokine that signals through STAT3 to regulate T follicular helper cell (TFH) differentiation and germinal center formation. In SIV-infected macaques, levels of IL-10 in plasma and lymph node (LN) were induced by infection and not normalized with ART. During chronic infection, plasma IL-10 and transcriptomic signatures of IL-10 signaling were correlated with the cell-associated SIV-DNA content within LN CD4+ memory subsets, including TFH, and predicted the frequency of CD4+ TFH and their cell-associated SIV-DNA content during ART, respectively. In ART-treated RMs, cells harboring SIV-DNA by DNAscope were preferentially found in the LN B-cell follicle in proximity to IL-10. Finally, we demonstrated that the in vivo neutralization of soluble IL-10 in ART-treated, SIV-infected macaques reduced B cell follicle maintenance and by extension LN memory CD4+ T-cells, including TFH and those expressing PD-1 and CTLA-4. Thus, these data support a role for IL-10 in maintaining a pool of target cells in lymphoid tissue that serve as a niche for viral persistence. Targeting IL-10 signaling to impair CD4+ T-cell survival and improve antiviral immune responses may represent a novel approach to limit viral persistence in ART-suppressed people living with HIV.

Authors

Justin Harper, Susan P. Ribeiro, Chi Ngai Chan, Malika Aid, Claire Deleage, Luca Micci, Maria Pino, Barbara Cervasi, Gopalan Raghunathan, Eric Rimmer, Gulesi Ayanoglu, Guoxin Wu, Neeta Shenvi, Richard J.O. Barnard, Gregory Q. Del Prete, Kathleen Busman-Sahay, Guido Silvestri, Deanna A. Kulpa, Steven E. Bosinger, Kirk Easley, Bonnie J. Howell, Dan Gorman, Daria J. Hazuda, Jacob D. Estes, Rafick-Pierre Sekaly, Mirko Paiardini

Remodeling the tumor microenvironment via blockade of LAIR-1 and TGF-β signaling enables PD-L1-mediated tumor eradication

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Abstract

Collagens in the extracellular matrix (ECM) provide a physical barrier to tumor immune infiltration, while also acting as a ligand for immune inhibitory receptors. Transforming growth factor-β (TGF-β) is a key contributor to shaping the ECM by stimulating the production and remodeling of collagens. TGF-β-activation signatures and collagen-rich environments have both been associated with T-cell exclusion and lack of responses to immunotherapy. Here we describe the effect of targeting collagens that signal through the inhibitory leukocyte-associated immunoglobulin-like receptor-1 (LAIR-1) in combination with blockade of TGF-β and programmed cell death ligand 1 (PD-L1). This approach remodeled the tumor collagenous matrix, enhanced tumor infiltration and activation of CD8+ T cells, and repolarized suppressive macrophage populations resulting in high cure rates and long-term tumor-specific protection across murine models of colon and mammary carcinoma. The results highlight the advantage of direct targeting of ECM components in combination with immune checkpoint blockade therapy.

Authors

Lucas A. Horn, Paul L. Chariou, Sofia R. Gameiro, Haiyan Qin, Masafumi Iida, Kristen Fousek, Thomas J. Meyer, Margaret Cam, Dallas Flies, Solomon Langermann, Jeffrey Schlom, Claudia Palena

Histone methyltransferase WHSC1 loss dampens MHC-I antigen presentation pathway to impair IFN-γ-stimulated anti-tumor immunity

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Abstract

IFN-γ-stimulated histocompatibility complex-I (MHC-I) antigen presentation underlies the core of anti-tumor immunity. However, sustained IFN-γ signaling also enhances PD-L1 checkpoint pathway to dampen anti-tumor immunity. It remains unclear how these opposing effects of IFN-γ are regulated. Here we reported that loss of the histone dimethyl transferase WHSC1 impaired the anti-tumor effect of IFN-γ signaling by the transcriptional downregulation of the MHC-I machinery without affecting PD-L1 expression in colorectal cancer (CRC) cells. Whsc1 loss promoted tumorigenesis via a non-cell autonomous mechanism in an Apcmin/+ mouse model, CRC organoids and xenografts. Mechanistically, we identified that IFN-γ-STAT1 signal axis stimulated WHSC1 expression, and in turn WHSC1 directly interacted with NLRC5 to promote MHC-I gene expression, but not PD-L1 level. Concordantly, silencing Whsc1 diminished MHC-I levels, impaired anti-tumor immunity and blunted the effect of immune checkpoint inhibitor (ICB). Patient cohort analysis revealed that WHSC1 expression positively correlated with enhanced MHC-I expression, tumor-infiltrating T cells and favorable disease outcome. Together, our findings establish a tumor-suppressive function of WHSC1 that relays IFN-γ signaling to promote antigen presentation in CRC cells, and provide a rationale for boosting WHSC1 activity in immunotherapy.

Authors

Jiale Ren, Ni Li, Siyu Pei, Yannan Lian, Li Li, Yuchong Peng, Qiuli Liu, Jiacheng Guo, Xuege Wang, Ying Han, Guoying Zhang, Hanling Wang, Yaqi Li, Jun Jiang, Qintong Li, Minjia Tan, Junjie Peng, Guohong Hu, Yichuan Xiao, Xiong Li, Moubin Lin, Jun Qin

HRAS germline mutations impair LKB1/AMPK signaling and mitochondrial homeostasis in Costello syndrome models

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Abstract

Germline mutations that activate genes in the canonical RAS/MAPK signaling pathway are responsible for rare human developmental disorders known as RASopathies. Here, we analyzed the molecular determinants of Costello syndrome (CS) using a mouse model expressing HRAS p.G12S, patient skin fibroblasts, hiPSC-derived human cardiomyocytes, a HRAS p.G12V zebrafish model and human fibroblasts expressing lentiviral constructs carrying HRAS p.G12S or HRAS p.G12A mutations. The findings revealed alteration of mitochondrial proteostasis and defective oxidative phosphorylation in the heart and skeletal muscle of Costello mice that were also found in the cell models of the disease. The underpinning mechanisms involved the inhibition of the AMPK signaling pathway by mutant forms of HRAS, leading to alteration of mitochondrial proteostasis and bioenergetics. Pharmacological activation of mitochondrial bioenergetics and quality control restored organelle function in HRAS p.G12A and p.G12S cell models, reduced left ventricle hypertrophy in the CS mice and diminished the occurrence of developmental defects in the CS zebrafish model. Collectively, these findings highlight the importance of mitochondrial proteostasis in the pathophysiology of RASopathies and suggest that patients with Costello syndrome may benefit from treatment with mitochondrial modulators.

Authors

Laetitia Dard, Christophe Hubert, Pauline Esteves, Wendy Blanchard, Ghina Bou About, Lyla Baldasseroni, Elodie Dumon, Chloe Angelini, Mégane Delourme, Véronique Guyonnet-Duperat, Stephane Claverol, Marc Bonneu, Laura Fontenille, Karima Kissa, Pierre-Emmanuel Séguéla, Jean-Benoît Thambo, Nicolas Levy, Yann Herault, Nadège Bellance, Nivea Dias Amoedo, Frederique Magdinier, Tania Sorg, Didier Lacombe, Rodrigue Rossignol

Brown adipose TRX2 deficiency activates mtDNA-NLRP3 to impair thermogenesis and protect against diet-induced insulin resistance

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Abstract

Brown adipose tissue (BAT), a crucial heat-generating organ, regulate whole-body energy metabolism by mediating thermogenesis. BAT inflammation is implicated in the pathogenesis of mitochondrial dysfunction and impaired thermogenesis. However, the link between BAT inflammation and systematic metabolism remains unclear. Herein, we use mice with BAT deficiency of thioredoxin-2 (TRX2), a protein that scavenges mitochondrial reactive oxygen species (ROS), to evaluate the impact of BAT inflammation on metabolism and thermogenesis and its underlying mechanism. Our results describe that BAT-specific TRX2 ablation improves systematic metabolic performance via enhancing lipid uptake, which protects mice from diet-induced obesity, hypertriglyceridemia, and insulin resistance. TRX2 deficiency impairs adaptive thermogenesis by suppressing fatty acid oxidation. Mechanistically, loss of TRX2 induces excessive mitochondrial ROS, mitochondrial integrity disruption, and cytosolic release of mitochondrial DNA, which in turn activate aberrant innate immune responses in BAT, including the cGAS-STING and the NLRP3 inflammasome pathways. We identify NLRP3 as a key converging point, as its inhibition reverses both the thermogenesis defect and the metabolic benefits seen under nutrient overload in BAT-specific Trx2-deficient mice. In conclusion, we identify TRX2 as a critical hub integrating oxidative stress, inflammation, and lipid metabolism in BAT; uncovering an adaptive mechanism underlying the link between BAT inflammation and systematic metabolism.

Authors

Yanrui Huang, Jenny H. Zhou, Haifeng Zhang, Alberto Canfrán-Duque, Abhishek K. Singh, Rachel J. Perry, Gerald Shulman, Carlos Fernandez-Hernando, Wang Min

CADASIL mutations sensitize the brain to ischemia via spreading depolarizations and abnormal extracellular potassium homeostasis

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Abstract

Cerebral autosomal dominant arteriopathy, subcortical infarcts and leukoencephalopathy (CADASIL) is the most common monogenic form of small vessel disease characterized by migraine with aura, leukoaraiosis, strokes and dementia. CADASIL mutations cause cerebrovascular dysfunction in both animal models and humans. Here, we show that two different human CADASIL mutations (Notch3 R90C or R169C) worsen ischemic stroke outcomes in transgenic mice, explained by a higher blood flow threshold to maintain tissue viability. Both mutants developed larger infarcts and worse neurological deficits compared with wild type regardless of age or sex after filament middle cerebral artery occlusion. However, full-field laser speckle flowmetry during distal middle cerebral artery occlusion showed comparable perfusion deficits in mutants and their respective wild type controls. Circle of Willis anatomy and pial collateralization also did not differ among the genotypes. In contrast, mutants had a higher cerebral blood flow threshold below which infarction ensued, suggesting increased sensitivity of brain tissue to ischemia. Electrophysiological recordings revealed a 1.5- to 2-fold higher frequency of peri-infarct spreading depolarizations in CADASIL mutants. Higher extracellular K+ elevations during spreading depolarizations in the mutants implicated a defect in extracellular K+ clearance. Altogether, these data reveal a novel mechanism of enhanced vulnerability to ischemic injury linked to abnormal extracellular ion homeostasis and susceptibility to ischemic depolarizations in CADASIL.

Authors

Fumiaki Oka, Jeong Hyun Lee, Izumi Yuzawa, Mei Li, Daniel von Bornstaedt, Katharina Eikermann-Haerter, Tao Qin, David Y. Chung, Homa Sadeghian, Jessica L. Seidel, Takahiko Imai, Doga Vuralli, Rosangela F.M. Platt, Mark T. Nelson, Anne Joutel, Sava Sakadzic, Cenk Ayata

Tumor Treating Fields dually activate STING and AIM2 inflammasomes to induce adjuvant immunity in glioblastoma

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Abstract

Tumor Treating Fields (TTFields), an approved therapy for glioblastoma (GBM) and malignant mesothelioma, employ non-invasive application of low-intensity, intermediate-frequency, alternating electric fields to disrupt the mitotic spindle, leading to chromosome mis-segregation and apoptosis. Emerging evidence suggest that TTFields may also induce inflammation. However, the mechanism of this property and whether it can be harnessed therapeutically are unclear. Here, we report that TTFields induced focal disruption of the nuclear envelope, leading to cytosolic release of large micronuclei clusters that intensely recruited and activated 2 major DNA sensors – cGAS (cyclic GMP-AMP synthase) and AIM2 (absent-in-melanoma-2) – and their cognate cGAS/STING (stimulator-of-interferon-genes) and AIM2/Caspase-1 inflammasomes to produce pro-inflammatory cytokines (PICs), type-1 interferons (T1IFNs), and T1IFN-responsive genes (T1IRGs). In syngeneic murine GBM models, TTFields-treated GBM cells induced anti-tumor memory immunity and cure rate of 42% to 66% in a STING- and AIM2-dependent manner. Using single-cell and bulk RNA-sequencing of peripheral blood mononuclear cells (PBMCs), we detected robust post-TTFields activation of adaptive immunity in patients with GBM via a T1IFN-based trajectory and identified a gene panel signature of TTFields effects on T cell activation and clonal expansion. Collectively, these studies defined a therapeutic strategy using TTFields as cancer immunotherapy in GBM and potentially other solid tumors.

Authors

Dongjiang Chen, Son B. Le, Tarun E. Hutchinson, Anda-Alexandra Calinescu, Mathew Sebastian, Dan Jin, Tianyi Liu, Ashley Ghiaseddin, Maryam Rahman, David D. Tran