Chronic allergic inflammatory diseases are a major cause of morbidity, allergic asthma alone affecting over 300 million people worldwide. Epidemiological studies demonstrate that environmental stimuli are associated with either promotion or prevention of disease. Major reductions in asthma prevalence are documented in European and US farming communities. Protection is associated with exposure of mothers during pregnancy to microbial breakdown products present in farm dusts and unprocessed foods, and enhancement of innate immune competence in the children. We sought to develop a scientific rationale for progressing these findings towards clinical application for primary disease prevention. Treatment of pregnant mice with a defined clinically-approved immune-modulator was shown to markedly reduce susceptibility of their offspring to development of the hallmark clinical features of allergic airway inflammatory disease. Mechanistically, offspring displayed enhanced dendritic cell-dependent airway mucosal immune surveillance function, which resulted in more efficient generation of mucosal-homing T-regulatory cells in response to local inflammatory challenge. We provide evidence that the principal target for maternal treatment effects was the fetal dendritic cell progenitor compartment, equipping the offspring for accelerated functional maturation of the airway mucosal dendritic cell network following birth. These data provide proof-of-concept supporting the rationale for development of transplacental immune reprogramming approaches for primary disease prevention.
Kyle T. Mincham, Naomi M. Scott, Jean-Francois Lauzon-Joset, Jonatan Leffler, Alexander N. Larcombe, Philip A. Stumbles, Sarah A. Robertson, Christian Pasquali, Patrick G. Holt, Deborah H. Strickland
It is suggested that subtyping of complex inflammatory diseases can be based on genetic susceptibility and relevant environmental exposure (G+E). We propose that using matched cellular phenotypes in human subjects and corresponding pre-clinical models with the same G+E combinations are useful to this end. As an example, defective Paneth cells can subtype Crohn's disease (CD) subjects; Paneth cell defects have been linked to multiple CD susceptibility genes and are associated with poor outcome. We hypothesized that CD susceptibility genes interact with cigarette smoking, a major CD environmental risk factor, to trigger Paneth cell defects. We found that both CD subjects and mice with ATG16L1T300A (T300A; a prevalent CD susceptibility allele) developed Paneth cell defects triggered by tobacco smoke. Transcriptional analysis of full thickness ileum and Paneth cell-enriched crypt base cells showed the T300A-smoking combination altered distinct pathways, including pro-apoptosis, metabolic dysregulation, and selective down-regulation of the PPARγ pathway. Pharmacologic intervention by either apoptosis inhibitor or PPARγ agonist rosiglitazone prevented smoking-induced crypt apoptosis and Paneth cell defects in T300A mice and mice with conditional Paneth cell-specific knockout of Atg16l1. This study demonstrates how explicit G+E can drive disease relevant phenotype, and provides rational strategies to identify actionable targets.
Ta-Chiang Liu, Justin T. Kern, Kelli L. VanDussen, Shanshan Xiong, Gerard E. Kaiko, Craig B. Wilen, Michael W. Rajala, Roberta Caruso, Michael J. Holtzman, Feng Gao, Dermot P.B. McGovern, Gabriel Nunez, Richard D. Head, Thaddeus S. Stappenbeck
Inflammation occurs in all tissues in response to injury or stress and is the key process underlying hepatic fibrogenesis. Targeting chronic and uncontrolled inflammation is one strategy to prevent liver injury and fibrosis progression. Here, we demonstrate that triggering receptor expressed on myeloid cells-1 (TREM-1), an amplifier of inflammation, promotes liver disease by intensifying hepatic inflammation and fibrosis. In the liver, TREM-1 expression is limited to liver macrophages and monocytes and is highly upregulated on Kupffer cells, circulating monocytes, and monocyte-derived macrophages in a mouse model of chronic liver injury and fibrosis induced by carbon tetrachloride (CCl4) administration. TREM-1 signaling promotes pro-inflammatory cytokine production and mobilization of inflammatory cells to the site of injury. Deletion of Trem1 reduced liver injury, inflammatory cell infiltration, and fibrogenesis. Reconstitution of Trem1-deficient mice with Trem1-sufficient Kupffer cells restored recruitment of inflammatory monocytes and severity of liver injury. Markedly increased infiltration of liver fibrotic areas with TREM-1-positive Kupffer cells and monocytes/macrophages was found in patients with hepatic fibrosis. Our data support a role of TREM-1 in liver injury and hepatic fibrogenesis and suggests that TREM-1 is a master regulator of Kupffer cell activation, which escalates chronic liver inflammatory responses, activates hepatic stellate cells, and reveals a novel mechanism of promotion of liver fibrosis.
Anh Thu Nguyen-Lefebvre, Ashwin Ajith, Vera Portik-Dobos, Daniel David Horuzsko, Ali Syed Arbab, Amiran Dzutsev, Ramses Sadek, Giorgio Trinchieri, Anatolij Horuzsko
Human endogenous retroviruses (hERVs) are remnants of exogenous retroviruses that have integrated into the genome throughout evolution. We developed a computational workflow, hervQuant, which identified over 3,000 transcriptionally active hERVs within The Cancer Genome Atlas (TCGA) pan-cancer RNA-seq database. hERV expression was associated with clinical prognosis in several tumor types, most significantly clear cell renal cell carcinoma (ccRCC). We explored two mechanisms by which hERV expression may influence the tumor-immune microenvironment in ccRCC: through 1) RIG-I-like signaling, and 2) retroviral antigen activation of adaptive immunity. We demonstrated the ability of hERV signatures associated with these immune mechanisms to predict patient survival in ccRCC, independent of clinical staging and molecular subtyping. We identified potential tumor-specific hERV epitopes with evidence of translational activity through the use of a ccRCC Ribo-seq dataset, validated their ability to bind HLA in vitro, and identified presence of MHC tetramer-positive T cells against predicted epitopes. hERV sequences identified through this screening approach were significantly more highly expressed in ccRCC tumors responsive to treatment with programmed death receptor-1 (PD-1) inhibition. hervQuant provides new insights into the role of hERVs within the tumor-immune microenvironment as well as evidence for hERV expression-based biomarkers for patient prognosis and response to immunotherapy.
Christof C. Smith, Kathryn E. Beckermann, Dante S. Bortone, Aguirre A. de Cubas, Lisa M. Bixby, Samuel J. Lee, Anshuman Panda, Shridar Ganesan, Gyan Bhanot, Eric M. Wallen, Matthew I. Milowsky, William Y. Kim, W. Kimryn Rathmell, Ronald Swanstrom, Joel S. Parker, Jonathan S. Serody, Sara R. Selitsky, Benjamin G. Vincent
Renin cells are crucial for survival: they control fluid-electrolyte and blood pressure homeostasis, vascular development, regeneration, and oxygen delivery to tissues. During embryonic development, renin cells are progenitors for multiple cell types which retain the memory of the renin phenotype. When there is a threat to survival, those descendants are transformed and reenact the renin phenotype to restore homeostasis. We tested the hypothesis that the molecular memory of the renin phenotype resides in unique regions and states of these cells’ chromatin. Using renin cells at various stages of stimulation, we identified regions in the genome where the chromatin is open for transcription, mapped histone modifications characteristic of active enhancers such as H3K27ac and deposition of transcriptional activators such a Med1 whose deletion results in ablation of Renin expression and low blood pressure. Using the rank ordering of super-enhancers, epigenetic re-writing, and enhancer deletion analysis, we found that renin cells harbor a unique set of super-enhancers that determine their identity. The most prominent Renin super-enhancer may act as a chromatin sensor of signals that convey the physiologic status of the organism and is responsible for the transformation of renin cell descendants to the renin phenotype, a fundamental process to ensure homeostasis.
Maria Florencia Martinez, Silvia Medrano, Evan A. Brown, Turan Tufan, Stephen Shang, Nadia Bertoncello, Omar Guessoum, Mazhar Adli, Brian C. Belyea, Maria Luisa S. Sequeira Lopez, R. Ariel Gomez
Mutant KRAS drives glycolytic flux in lung cancer, potentially impacting aberrant protein glycosylation. Recent evidence suggests aberrant KRAS drives flux of glucose into the hexosamine biosynthetic pathway (HBP). HBP is required for various glycosylation processes, such as protein N- or O-glycosylation and glycolipid synthesis. However, its function during tumorigenesis is poorly understood. One contributor and proposed target of KRAS driven cancers is a developmentally conserved epithelial plasticity program called epithelial-mesenchymal transition (EMT). Here we show in novel autochthonous mouse models that EMT accelerates KrasG12D lung tumorigenesis by upregulating expression of key enzymes of the HBP pathway. We demonstrate that HBP is required for suppressing KrasG12D-induced senescence, and targeting HBP significantly delays KrasG12D lung tumorigenesis. To explore the mechanism, we investigated protein glycosylation downstream of HBP and found elevated levels of O-linked β-N-acetylglucosamine (O-GlcNAcylation) post-translational modification on intracellular proteins. O-GlcNAcylation suppressed KrasG12D oncogene-induced senescence (OIS) and accelerates lung tumorigenesis. Conversely, loss of O-GlcNAcylation delays lung tumorigenesis. O-GlcNAcylation of proteins SNAI1 and c-Myc correlates with the EMT-HBP axis and accelerated lung tumorigenesis. Our results demonstrate that O-GlcNAcylation is sufficient and required to accelerate KrasG12D lung tumorigenesis in vivo, which is reinforced by epithelial plasticity programs.
Kekoa Taparra, Hailun Wang, Reem Malek, Audrey Lafargue, Mustafa A. Barbhuiya, Xing Wang, Brian W. Simons, Matthew Ballew, Katriana Nugent, Jennifer Groves, Russell D. Williams, Takumi Shiraishi, James Verdone, Gokben Yildirir, Roger Henry, Bin Zhang, John Wong, Ken Kang-Hsin Wang, Barry D. Nelkin, Kenneth J. Pienta, Dean Felsher, Natasha E. Zachara, Phuoc T. Tran
First generation immune checkpoint inhibitors including anti-CTLA-4 and anti-PD-1 antibodies have led to major clinical progress, yet resistance frequently leads to treatment failure. Thus, new targets acting on T cells are needed. CD33-related Siglecs are pattern recognition immune receptors binding to a range of sialoglycan ligands, which appear to function as self-associated molecular patterns (SAMPs) that suppress autoimmune responses. Siglecs are expressed at very low levels on normal T cells, and these receptors were not yet considered as interesting targets on T cells for cancer immunotherapy. Here, we show an upregulation of Siglecs including Siglec-9 on tumor-infiltrating T cells from non-small cell lung (NSCLC), colorectal and ovarian cancer patients. Siglec-9 expressing T cells co-expressed several inhibitory receptors including PD-1. Targeting of the sialoglycan-SAMP/Siglec pathway in vitro and in vivo resulted in increased anti-cancer immunity. T cell expression of Siglec-9 in NSCLC patients correlated with a reduced survival, and Siglec-9 polymorphisms showed associations with the risk of developing lung and colorectal cancer. Our data identify the sialoglycan-SAMP/Siglec pathway as new potential target to improve T cell activation for immunotherapy.
Michal A. Stanczak, Shoib S. Siddiqui, Marcel P. Trefny, Daniela S. Thommen, Kayluz Frias Boligan, Stephan von Gunten, Alexandar Tzankov, Lothar Tietze, Didier Lardinois, Viola Heinzelmann-Schwarz, Michael S. von Bergwelt-Baildon, Wu Zhang, Heinz-Josef Lenz, Younghan Han, Christopher I. Amos, Mohammedyaseen Syedbasha, Adrian Egli, Frank Stenner, Daniel E. Speiser, Ajit Varki, Alfred Zippelius, Heinz Läubli
While T cells are important for the pathogenesis of systemic lupus erythematosus (SLE) and lupus nephritis, little is known about how T cells function after infiltrating the kidney. The current paradigm suggests that kidney infiltrating T cells (KITs) are activated effector cells contributing to tissue damage and ultimately organ failure. Herein, we demonstrate that the majority of CD4+ and CD8+ KITs in three murine lupus models are not effector cells, as hypothesized, but rather expressed multiple inhibitory receptors and proved highly dysfunctional with reduced cytokine production and proliferative capacity. Mechanistically this was linked directly to metabolic and specifically mitochondrial dysfunction. This was driven by the expression of an “exhausted” transcriptional signature. Our data thus reveal that the tissue parenchyma has the capability to suppress T cell responses and limit damage to self. These findings open novel avenues for the treatment of autoimmunity based on selectively exploiting the exhausted phenotype of tissue-infiltrating T cells.
Jeremy S. Tilstra, Lyndsay Avery, Ashley V. Menk, Rachael A. Gordon, Shuchi Smita, Lawrence P. Kane, Maria Chikina, Greg M. Delgoffe, Mark J. Shlomchik
Regulatory T-cells (Treg) are critical for maintaining immune homeostasis. However, current Treg immunotherapies do not optimally treat inflammatory diseases in patients. Understanding the cellular processes that control Treg function may allow for the augmentation of therapeutic efficacy. In contrast to activated conventional T-cells, where protein kinase C-θ (PKC-θ) localizes to the contact-point between T-cells and antigen-presenting cells, in human and mouse Treg, PKC-θ localizes to the opposite end of the cell in the distal pole complex (DPC). Here, using a phosphoproteomic screen, we identified the intermediate filament vimentin as a PKC-θ phospho-target and show that vimentin forms a DPC superstructure on which PKC-θ accumulates. Treatment of mouse Treg with either a clinically relevant PKC-θ inhibitor or vimentin siRNA disrupted vimentin and enhanced Treg metabolic and suppressive activity. Moreover, vimentin-disrupted mouse Treg were significantly better than controls in suppressing alloreactive T-cell priming in graft-versus-host disease, and graft-versus-host disease lethality, using a complete MHC mismatch mouse model of acute graft-versus-host disease (C57BL/6 donor in to BALB/c host). Interestingly, vimentin disruption augmented suppressor function of PKC-θ-deficient mouse Treg. This suggests that enhanced Treg activity after PKC-θ inhibition is secondary to effects on vimentin, not just PKC-θ kinase activity inhibition. Our data demonstrated that vimentin is a key metabolic and functional controller of Treg activity, and provide proof-of-principle that disrupting vimentin is a feasible, translationally relevant method to enhance Treg potency.
Cameron McDonald-Hyman, James T. Muller, Michael Loschi, Govindarajan Thangavelu, Asim Saha, Sudha Kumari, Dawn K. Reichenbach, Michelle J. Smith, Guoan Zhang, Brent H. Koehn, Jiqiang Lin, Jason S. Mitchell, Brian T. Fife, Angela Panoskaltsis-Mortari, Colby J. Feser, Andrew Kemal Kirchmeier, Mark J. Osborn, Keli L. Hippen, Ameeta Kelekar, Jonathan S. Serody, Laurence A. Turka, David H. Munn, Hongbo Chi, Thomas A. Neubert, Michael L. Dustin, Bruce R. Blazar
Previous findings showed that in mice, complete knockout of activity-dependent neuroprotective protein (ADNP) abolishes brain formation, while haploinsufficiency (Adnp+/–) causes cognitive impairments. We hypothesized that mutations in ADNP lead to a developmental/autistic syndrome in children. Indeed, recent phenotypic characterization of children harboring ADNP mutations (ADNP syndrome children) revealed global developmental delays and intellectual disabilities, including speech and motor dysfunctions. Mechanistically, ADNP includes a SIP motif embedded in the ADNP-derived snippet, drug candidate NAP (NAPVSIPQ also known as CP201), which binds to microtubule end binding protein 3, essential for dendritic spine formation. Here, we established a unique neuronal membrane tagged green fluorescent protein expressing Adnp+/– mouse line allowing in vivo synaptic pathology quantification. We discovered that Adnp deficiency reduced dendritic spine density and altered synaptic gene expression, both of which were partly ameliorated by NAP treatment. Adnp+/– mice further exhibited global developmental delays, vocalization impediments, gait/motor dysfunctions and social/object memory impairments, all partially reversed by daily NAP administration (systemic/nasal). In conclusion, we now connected ADNP-related synaptic pathology to developmental/behavioral outcomes, establishing NAP in vivo target engagement and identifying potential biomarkers. Together, these studies pave the path toward clinical development of NAP (CP201) in the ADNP syndrome.
Gal Hacohen-Kleiman, Shlomo Sragovich, Gidon Karmon, Andy Y. L. Gao, Iris Grigg, Metsada Pasmanik-Chor, Albert Le, Vlasta Korenková, R. Anne McKinney, Illana Gozes
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