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Abstract

First-generation immune checkpoint inhibitors, including anti–CTLA-4 and anti–programmed death 1 (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 sialic acid–binding immunoglobulin-like lectins (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 until recently 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 cancer (NSCLC), colorectal, and ovarian cancer patients. Siglec-9–expressing T cells coexpressed several inhibitory receptors, including PD-1. Targeting of the sialoglycan-SAMP/Siglec pathway in vitro and in vivo resulted in increased anticancer immunity. T cell expression of Siglec-9 in NSCLC patients correlated with reduced survival, and Siglec-9 polymorphisms showed association with the risk of developing lung and colorectal cancer. Our data identify the sialoglycan-SAMP/Siglec pathway as a potential target for improving T cell activation for immunotherapy.

Authors

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 von Bergwelt-Baidon, Wu Zhang, Heinz-Josef Lenz, Younghun Han, Christopher I. Amos, Mohammedyaseen Syedbasha, Adrian Egli, Frank Stenner, Daniel E. Speiser, Ajit Varki, Alfred Zippelius, Heinz Läubli

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Abstract

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 3 murine lupus models are not effector cells, as hypothesized, but rather express multiple inhibitory receptors and are highly dysfunctional, with reduced cytokine production and proliferative capacity. In other systems, this hypofunctional profile is linked directly to metabolic and specifically mitochondrial dysfunction, which we also observed in KITs. The T cell phenotype was driven by the expression of an “exhausted” transcriptional signature. Our data thus reveal that the tissue parenchyma has the capability of suppressing T cell responses and limiting damage to self. These findings suggest avenues for the treatment of autoimmunity based on selectively exploiting the exhausted phenotype of tissue-infiltrating T cells.

Authors

Jeremy S. Tilstra, Lyndsay Avery, Ashley V. Menk, Rachael A. Gordon, Shuchi Smita, Lawrence P. Kane, Maria Chikina, Greg M. Delgoffe, Mark J. Shlomchik

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Abstract

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 showed in novel autochthonous mouse models that EMT accelerated KrasG12D lung tumorigenesis by upregulating expression of key enzymes of the HBP pathway. We demonstrated that HBP was required for suppressing KrasG12D-induced senescence, and targeting HBP significantly delayed 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) posttranslational modification on intracellular proteins. O-GlcNAcylation suppressed KrasG12D oncogene-induced senescence (OIS) and accelerated lung tumorigenesis. Conversely, loss of O-GlcNAcylation delayed lung tumorigenesis. O-GlcNAcylation of proteins SNAI1 and c-MYC correlated with the EMT-HBP axis and accelerated lung tumorigenesis. Our results demonstrated that O-GlcNAcylation was sufficient and required to accelerate KrasG12D lung tumorigenesis in vivo, which was reinforced by epithelial plasticity programs.

Authors

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

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Abstract

The underlying molecular mechanisms of age-related hearing loss (ARHL) in humans and many strains of mice have not been fully characterized. This common age-related disorder is assumed to be closely associated with oxidative stress. Here, we demonstrate that mTORC1 signaling is highly and specifically activated in the cochlear neurosensory epithelium (NSE) in aging mice, and rapamycin injection prevents ARHL. To further examine the specific role of mTORC1 signaling in ARHL, we generated murine models with NSE-specific deletions of Raptor or Tsc1, regulators of mTORC1 signaling. Raptor-cKO mice developed hearing loss considerably more slowly than WT littermates. Conversely, Tsc1 loss led to the early-onset death of cochlear hair cells and consequently accelerated hearing loss. Tsc1-cKO cochleae showed features of oxidative stress and impaired antioxidant defenses. Treatment with rapamycin and the antioxidant N-acetylcysteine rescued Tsc1-cKO hair cells from injury in vivo. In addition, we identified the peroxisome as the initial signaling organelle involved in the regulation of mTORC1 signaling in cochlear hair cells. In summary, our findings identify overactive mTORC1 signaling as one of the critical causes of ARHL and suggest that reduction of mTORC1 activity in cochlear hair cells may be a potential strategy to prevent ARHL.

Authors

Xiaolong Fu, Xiaoyang Sun, Linqing Zhang, Yecheng Jin, Renjie Chai, Lili Yang, Aizhen Zhang, Xiangguo Liu, Xiaochun Bai, Jianfeng Li, Haibo Wang, Jiangang Gao

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Abstract

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, GFP-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 and motor dysfunctions, and social and object memory impairments, all of which were partially reversed by daily NAP administration (systemic/nasal). In conclusion, we have connected ADNP-related synaptic pathology to developmental and behavioral outcomes, establishing NAP in vivo target engagement and identifying potential biomarkers. Together, these studies pave a path toward the clinical development of NAP (CP201) for the treatment of ADNP syndrome.

Authors

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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Abstract

Graft-versus-host disease (GVHD) in the gastrointestinal (GI) tract remains the major cause of morbidity and nonrelapse mortality after BM transplantation (BMT). The Paneth cell protein regenerating islet-derived 3α (REG3α) is a biomarker specific for GI GVHD. REG3α serum levels rose in the systematic circulation as GVHD progressively destroyed Paneth cells and reduced GI epithelial barrier function. Paradoxically, GVHD suppressed intestinal REG3γ (the mouse homolog of human REG3α), and the absence of REG3γ in BMT recipients intensified GVHD but did not change the composition of the microbiome. IL-22 administration restored REG3γ production and prevented apoptosis of both intestinal stem cells (ISCs) and Paneth cells, but this protection was completely abrogated in Reg3g−/− mice. In vitro, addition of REG3α reduced the apoptosis of colonic cell lines. Strategies that increase intestinal REG3α/γ to promote crypt regeneration may offer a novel, nonimmunosuppressive approach for GVHD and perhaps for other diseases involving the ISC niche, such as inflammatory bowel disease.

Authors

Dongchang Zhao, Yeung-Hyen Kim, Seihwan Jeong, Joel K. Greenson, Mohammed S. Chaudhry, Matthias Hoepting, Erik R. Anderson, Marcel R.M. van den Brink, Jonathan U. Peled, Antonio L.C. Gomes, Ann E. Slingerland, Michael J. Donovan, Andrew C. Harris, John E. Levine, Umut Ozbek, Lora V. Hooper, Thaddeus S. Stappenbeck, Aaron Ver Heul, Ta-Chiang Liu, Pavan Reddy, James L.M. Ferrara

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Abstract

The E3 ubiquitin ligase Pellino 1 (Peli1) is a microglia-specific mediator of autoimmune encephalomyelitis. Its role in neurotropic flavivirus infection is largely unknown. Here, we report that mice deficient in Peli1 (Peli1–/–) were more resistant to lethal West Nile virus (WNV) infection and exhibited reduced viral loads in tissues and attenuated brain inflammation. Peli1 mediates chemokine and proinflammatory cytokine production in microglia and promotes T cell and macrophage infiltration into the CNS. Unexpectedly, Peli1 was required for WNV entry and replication in mouse macrophages and mouse and human neurons and microglia. It was also highly expressed on WNV-infected neurons and adjacent inflammatory cells from postmortem patients who died of acute WNV encephalitis. WNV passaged in Peli1–/– macrophages or neurons induced a lower viral load and impaired activation in WT microglia and thereby reduced lethality in mice. Smaducin-6, which blocks interactions between Peli1 and IRAK1, RIP1, and IKKε, did not inhibit WNV-triggered microglia activation. Collectively, our findings suggest a nonimmune regulatory role for Peli1 in promoting microglia activation during WNV infection and identify a potentially novel host factor for flavivirus cell entry and replication.

Authors

Huanle Luo, Evandro R. Winkelmann, Shuang Zhu, Wenjuan Ru, Elizabeth Mays, Jesus A. Silvas, Lauren L. Vollmer, Junling Gao, Bi-Hung Peng, Nathen E. Bopp, Courtney Cromer, Chao Shan, Guorui Xie, Guangyu Li, Robert Tesh, Vsevolod L. Popov, Pei-Yong Shi, Shao-Cong Sun, Ping Wu, Robyn S. Klein, Shao-Jun Tang, Wenbo Zhang, Patricia V. Aguilar, Tian Wang

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Abstract

MASTL, a Ser/Thr kinase that inhibits PP2A-B55 complexes during mitosis, is mutated in autosomal dominant thrombocytopenia. However, the connections between the cell cycle machinery and this human disease remain unexplored. We report here that, whereas Mastl ablation in megakaryocytes prevented proper maturation of these cells, mice carrying the thrombocytopenia-associated mutation developed thrombocytopenia as a consequence of aberrant activation and survival of platelets. Activation of mutant platelets was characterized by hyper-stabilized pseudopods mimicking the effect of PP2A inhibition and actin polymerization defects. These aberrations were accompanied by abnormal hyper-phosphorylation of multiple components of the actin cytoskeleton and were rescued both in vitro and in vivo by inhibiting upstream kinases such as PKA, PKC, or AMPK. These data reveal an unexpected role of Mastl in actin cytoskeleton dynamics in postmitotic cells, and suggest that the thrombocytopenia-associated mutation in MASTL is a pathogenic dominant mutation that mimics decreased PP2A activity resulting in altered phosphorylation of cytoskeletal regulatory pathways.

Authors

Begoña Hurtado, Marianna Trakala, Pilar Ximénez-Embún, Aicha El Bakkali, David Partida, Belén Sanz-Castillo, Mónica Álvarez-Fernández, María Maroto, Ruth Sánchez-Martínez, Lola Martínez, Javier Muñoz, Pablo García de Frutos, Marcos Malumbres

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Abstract

Acute Myeloid Leukemia and Myelodysplastic Syndromes are associated with disease-initiating stem cells that are not eliminated by conventional therapies. Transcriptomic analysis of stem and progenitor populations in MDS and AML demonstrated overexpression of STAT3 that was validated in an independent cohort. STAT3 overexpression was predictive of a shorter survival and worse clinical features in a large MDS cohort. High STAT3 expression signature in MDS CD34+ cells was similar to known pre-leukemic gene signatures. Functionally, STAT3 inhibition by a clinical, antisense oligonucleotide, AZD9150, led to reduced viability and increased apoptosis in leukemic cell lines. AZD9150 was rapidly incorporated by primary MDS/AML stem and progenitor cells and led to increased hematopoietic differentiation. STAT3 knockdown also impaired leukemic growth in vivo and led to decreased expression of MCL1 and other oncogenic genes in malignant cells. These studies demonstrate that STAT3 is an adverse prognostic factor in MDS/AML and provide a pre-clinical rationale for studies using AZD9150 in these diseases.

Authors

Aditi Shastri, Gaurav Choudhary, Margarida Teixeira, Shanisha Gordon-Mitchell, Nandini Ramachandra, Lumie Bernard, Sanchari Bhattacharyya, Robert Lopez, Kith Pradhan, Orsolya Giricz, Goutham Ravipati, Li-Fan Wong, Sally Cole, Tushar D. Bhagat, Jonathan Feld, Yosman Dhar, Matthias Bartenstein, Victor J. Thiruthuvanathan, Amittha Wickrema, B. Hilda Ye, David A. Frank, Andrea Pellagatti, Jacqueline Boultwood, Tianyuan Zhou, Youngsoo Kim, A. Robert MacLeod, Pearlie K. Epling-Burnette, Minwei Ye, Patricia McCoon, Richard Woessner, Ulrich Steidl, Britta Will, Amit K. Verma

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Abstract

Obesity is a major risk factor for developing nonalcoholic fatty-liver disease (NAFLD). NAFLD is the most common form of chronic liver disease and closely associated with insulin resistance, ultimately leading to cirrhosis and hepatocellular carcinoma. However, knowledge of the intracellular regulators of obesity-linked fatty-liver disease remains incomplete. Here we showed that hepatic Rho-kinase 1 (ROCK1) drives obesity-induced steatosis in mice through stimulation of de novo lipogenesis. Mice lacking ROCK1 in the liver were resistant to diet-induced obesity due to increased energy expenditure and thermogenic gene expression. Constitutive expression of hepatic ROCK1 was sufficient to promote adiposity, insulin resistance, and hepatic lipid accumulation in mice fed a high-fat diet. Correspondingly, liver-specific ROCK1 deletion prevented the development of severe hepatic steatosis and reduced hyperglycemia in obese diabetic (ob/ob) mice. Of pathophysiologic significance, hepatic ROCK1 was markedly up-regulated in humans with fatty-liver disease and correlated with risk factors clustering around NAFLD and insulin resistance. Mechanistically, we found that hepatic ROCK1 suppresses AMPK activity and a ROCK1-AMPK pathway is necessary to mediate cannabinoid-induced lipogenesis in the liver. Furthermore, treatment with metformin, the most widely used anti-diabetes drug, reduced hepatic lipid accumulation by inactivating ROCK1, resulting in activation of AMPK downstream signaling. Taken together, our findings establish a ROCK1-AMPK signaling axis that regulates de novo lipogenesis, providing a unique target for treating obesity-related metabolic disorders such as NAFLD.

Authors

Hu Huang, Seung-Hwan Lee, Inês Sousa-Lima, Sang Soo Kim, Won Min Hwang, Yossi Dagon, Won-Mo Yang, Sungman Cho, Min-Cheol Kang, Ji A Seo, Munehiko Shibata, Hyunsoo Cho, Getachew Debas Belew, Jinhyuk Bhin, Bhavna N. Desai, Min Jeong Ryu, Minho Shong, Peixin Li, Hua Meng, Byung-Hong Chung, Daehee Hwang, Min Seon Kim, Kyong Soo Park, Paula Macedo, Morris White, John Jones, Young-Bum Kim

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Abstract

Sugar- and lipid-derived aldehydes are reactive carbonyl species (RCS) frequently used as surrogate markers of oxidative stress in obesity. A pathogenic role for RCS in metabolic diseases of obesity remains controversial, however, due in part to their highly diffuse and broad reactivity, and to lack of specific RCS-scavenging therapies. Naturally occurring histidine dipeptides (e.g., anserine and carnosine) possess RCS reactivity, but their therapeutic potential in humans is limited by serum carnosinases. Here we present the rational design, characterization and pharmacological evaluation of ‘carnosinol’ (i.e. (2S)-2-(3-amino propanoylamino)-3-(1H-imidazol-5-yl)propanol) a derivative of carnosine with high oral bioavailability that is resistant to carnosinases. Carnosinol displayed a suitable ADMET profile and was determined to have the greatest potency and selectivity toward α,β-unsaturated aldehydes (e.g. 4-hydroxynonenal, HNE, acrolein) among all others so far reported. In rodent models of diet-induced obesity and metabolic syndrome, carnosinol dose-dependently attenuated HNE-adduct formation in liver and skeletal muscle while simultaneously mitigating inflammation, dyslipidemia, insulin resistance, and steatohepatitis. These improvements in metabolic parameters with carnosinol were not due to changes in energy expenditure, physical activity, adiposity or body weight. Collectively, our findings illustrate a pathogenic role for RCS in obesity-related metabolic disorders, and provide validation for a promising new class of carbonyl-scavenging therapeutic compounds rationally derived from carnosine.

Authors

Ethan J. Anderson, Giulio Vistoli, Lalage A. Katunga, Katsuhiko Funai, Luca Regazzoni, T. Blake Monroe, Ettore Gilardoni, Luca Cannizzaro, Mara Colzani, Danilo De Maddis, Giuseppe Rossoni, Renato Canevotti, Stefania Gagliardi, Marina Carini, Giancarlo Aldini

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Abstract

Heart failure (HF) remains a major source of morbidity and mortality in the U.S. The multifunctional Ca2+/calmodulin-dependent kinase II (CaMKII) has emerged as a critical regulator of cardiac hypertrophy and failure, although the mechanisms remain unclear. Previous studies have established that the cytoskeletal protein βIV-spectrin coordinates local CaMKII signaling. Here we sought to determine the role of a spectrin/CaMKII complex in maladaptive remodeling in HF. Chronic pressure overload (6 weeks transaortic constriction, TAC) induced a decrease in cardiac function in WT mice but not in animals expressing truncated βIV-spectrin lacking spectrin/CaMKII interaction (qv3J). Underlying observed differences in function was an unexpected differential regulation of STAT3-related genes in qv3J TAC hearts. In vitro experiments demonstrate that βIV-spectrin serves as a target for CaMKII phosphorylation, which regulates its stability. Cardiac-specific βIV-spectrin knockout (βIV-cKO) mice show STAT3 dysregulation, fibrosis and decreased cardiac function at baseline similar to WT TAC. STAT3 inhibition restored normal cardiac structure and function in βIV-cKO and WT TAC hearts. Our studies identify a novel spectrin-based complex essential for regulation of the cardiac response to chronic pressure overload. We anticipate that strategies targeting the new spectrin-based “statosome” will be effective at suppressing maladaptive remodeling in response to chronic stress.

Authors

Sathya D. Unudurthi, Drew M. Nassal, Amara Greer-Short, Nehal J. Patel, Taylor Howard, Xianyao Xu, Birce Onal, Tony Satroplus, Deborah Y. Hong, Cemantha M. Lane, Alyssa Dalic, Sara N. Koenig, Adam C. Lehnig, Lisa A. Baer, Hassan Musa, Kristin I. Stanford, Sakima A. Smith, Peter J. Mohler, Thomas J. Hund

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August 2018

128 9 cover

August 2018 Issue

On the cover:
Wnt and MAPK interactions calibrate intestinal regeneration

In this issue of the JCI, Kabiri et al. uncover an unexpected interaction between Wnt and MAPK signaling in regulating intestinal stem cell proliferation (ISC). They report that the Wnt pathway suppresses MAPK-driven proliferation in intestinal crypts to maintain stem cell self-renewal. On the cover, ISCs and their progeny are highlighted in yellow and red, respectively.

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Jci tm 2018 09

September 2018 JCI This Month

JCI This Month is a digest of the research, reviews, and other features published each month.

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Review Series - More

Mitochondrial dysfunction in disease

Series edited by Michael Sack

Mitochondria transform nutrients and oxygen into chemical energy that powers a multitude of cellular functions. While mitochondrial aerobic glycolysis generates the majority of a cell’s ATP, its byproducts also have wide-ranging influences on cellular health and longevity. This review series, edited by Dr. Michael Sack, focuses on the many contributions of mitochondria to disease and aging. The reviews highlight evidence linking altered mitochondrial metabolism and oxidative stress to a range of pathophysiological phenomena: inflammation and immune dysfunction, heart failure, cancer development, metabolic disease, and more. In many diseases and conditions, mitochondrial dysfunction is considered the tipping point toward pathological progression. However, as these reviews discuss, therapeutic targeting of mitochondria may be a powerful strategy to subvert disease and aging processes.

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