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Abstract
TrkB agonist drugs are shown here to have a significant effect on the regeneration of afferent cochlear synapses after noise-induced synaptopathy. The effects were consistent with regeneration of cochlear synapses that we observed in vitro after synaptic loss due to kainic acid-induced glutamate toxicity and were elicited by administration of TrkB agonists, amitriptyline and 7,8- dihydroxyflavone, directly into the cochlea via the posterior semicircular canal 48 h after exposure to noise. Synaptic counts at the inner hair cell and wave 1 amplitudes in the ABR were partially restored 2 weeks after drug treatment. Effects of amitriptyline on wave 1 amplitude and afferent auditory synapse numbers in noise-exposed ears after systemic (as opposed to local) delivery were profound and long-lasting; synapses in the treated animals remained intact one year after the treatment. However, the effect of systemically delivered amitriptyline on synaptic rescue was dependent on dose and the time window of administration: it was only effective when given before noise exposure at the highest injected dose. The long-lasting effect and the efficacy of post-exposure treatment indicate a potential broad application for the treatment of synaptopathy, which often goes undetected until well after the original damaging exposure(s).
Authors
Katharine A. Fernandez, Takahisa Watabe, Mingjie Tong, Xiankai Meng, Kohsuke Tani, Sharon G. Kujawa, Albert S.B. Edge
Abstract
Interleukin-10 (IL-10) is a critical cytokine used by immune cells to suppress inflammation. Paradoxically, immune cell-derived IL-10 can drive insulin resistance in obesity by suppressing adipocyte energy expenditure and thermogenesis. However, the source of IL-10 necessary for the suppression of adipocyte thermogenesis is unknown. We show here that CD4+ Foxp3+ regulatory T cells (Tregs) are a significant source of IL-10, and that Treg-derived IL-10 can suppress adipocyte beiging. Unexpectedly, Treg-specific loss of IL-10 resulted in increased insulin sensitivity and reduced obesity in high fat diet (HFD)-fed male mice. Mechanistically, we determined that Treg-specific loss of the transcription factor Blimp-1, a driver of IL-10 expression by Tregs, phenocopied the Treg-specific IL-10-deficient mice. Loss of Blimp-1 expression in Tregs resulted in reduced ST2+, KLRG1+, IL-10-secreting Tregs, particularly in the white adipose tissue. Blimp-1-deficient mice were protected from glucose intolerance, insulin resistance and diet-induced obesity (DIO), through increased white adipose tissue browning. Taken together, our data show that Blimp-1-regulated IL-10 secretion by Tregs represses white adipose tissue beiging to maintain adipose tissue homeostasis.
Authors
Lisa Y. Beppu, Raja Mooli, Xiaoyao Qu, Giovanni J. Marrero, Christopher A. Finley, Allen N. Fooks, Zackary P. Mullen, Adolfo B. Frias Jr., Ian J. Sipula, Bingxian Xie, Katherine E. Helfrich, Simon C. Watkins, Amanda C. Poholek, Sadeesh K. Ramakrishnan, Michael J. Jurczak, Louise M. D'Cruz
Abstract
Although many HIV cure strategies seek to expand HIV-specific CD8+ T cells to control the virus, all are likely to fail if cellular exhaustion is not prevented. A loss in stem-like memory properties (i.e., the ability to proliferate and generate secondary effector cells) is a key feature of exhaustion; little is known, however, about how these properties are regulated in human virus-specific CD8+ T cells. We found that virus-specific CD8+ T cells from humans and non-human primates naturally controlling HIV/SIV infection express more of the transcription factor, TCF-1, than non-controllers. HIV-specific CD8+ T cell TCF-1 expression correlated with memory marker expression and expansion capacity and declined with antigenic stimulation. CRISPR-Cas9 editing of TCF-1 in human primary T cells demonstrated a direct role in regulating expansion capacity. Collectively, these data suggest that TCF-1 contributes to the regulation of the stem-like memory property of secondary expansion capacity of HIV-specific CD8+ T cells, and they provide a rationale for exploring the enhancement of this pathway in T cell-based therapeutic strategies for HIV.
Authors
Rachel L. Rutishauser, Christian Deo T. Deguit, Joseph Hiatt, Franziska Blaeschke, Theodore L. Roth, Lynn Wang, Kyle A. Raymond, Carly E. Starke, Joseph C. Mudd, Wenxuan Chen, Carolyn P. Smullin, Rodrigo Matus-Nicodemos, Rebecca Hoh, Melissa R. Krone, Frederick M. Hecht, Christopher D. Pilcher, Jeffrey N. Martin, Richard A. Koup, Daniel C. Douek, Jason M. Brenchley, Rafick-Pierre Sékaly, Satish K. Pillai, Alexander Marson, Steven G. Deeks, Joseph M. McCune, Peter W. Hunt
Abstract
Recent in vivo tracer studies demonstrated that targeted mass spectrometry (MS) on the Q Exactive Orbitrap could determine the metabolism of HDL proteins 100s-fold less abundant than APOA1. In this study, we demonstrate that the Orbitrap Lumos can measure tracer in proteins whose abundances are 1000s-fold less than APOA1, specifically the lipid transfer proteins PLTP, CETP, and LCAT. Relative to the Q Exactive, the Lumos improved tracer detection by reducing tracer enrichment compression, thereby providing consistent enrichment data across multiple HDL sizes from six participants. We determined by compartmental modeling that PLTP is secreted in medium and large HDL (alpha2, 1, and 0), and is transferred from medium to larger sizes during circulation from where it is catabolized. CETP is secreted mainly in alpha1 and alpha2, and remains in these sizes during circulation. LCAT is secreted mainly in medium and small HDL (alpha2, 3, prebeta). Unlike PLTP and CETP, LCAT appearance on HDL is markedly delayed compared to APOA1 and the other transfer proteins, indicating that LCAT may reside for a time outside of systemic circulation before attaching to HDL in plasma. The determination of these lipid transfer proteins’ unique metabolic structures was possible due to advances in MS technologies.
Authors
Sasha A. Singh, Allison B. Andraski, Hideyuki Higashi, Lang Ho Lee, Ashisha Ramsaroop, Frank M. Sacks, Masanori Aikawa
Abstract
The cohesin complex plays an essential role in chromosome maintenance and transcriptional regulation. Recurrent somatic mutations in the cohesin complex are frequent genetic drivers in cancer including myelodysplatic syndromes (MDS) and acute myeloid leukemia (AML). Here, using genetic dependency screens of STAG2-mutant AML, we identified DNA damage repair and replication as genetic dependencies in cohesin-mutant cells. We demonstrated increased levels of DNA damage and sensitivity of cohesin-mutant cells to PARP inhibition. We developed a mouse model of MDS in which Stag2 mutations arise as clonal secondary lesions in the background of clonal hematopoiesis driven by Tet2 mutations, and demonstrated selective depletion of cohesin-mutant cells with PARP inhibition in vivo. Finally, we demonstrated a shift from STAG2- to STAG1-containing cohesin complexes in cohesin-mutant cells, which is associated with longer DNA loop extrusion, more intermixing of chromatin compartments, and increased interaction with PARP and RPA proteins. Our findings inform the biology and therapeutic opportunities for cohesin-mutant malignancies.
Authors
Zuzana Tothova, Anne-Laure Valton, Rebecca Gorelov, Mounica Vallurupalli, John M. Krill-Burger, Amie Holmes, Catherine C. Landers, J. Erika Haydu, Edyta Malolepsza, Christina R. Hartigan, Melanie Donahue, Katerina D. Popova, Sebastian H. J. Koochaki, Sergey V. Venev, Jeanne F. Rivera, Edwin Chen, Kasper Lage, Monica Schenone, Alan D. D'Andrea, Steven A. Carr, Elizabeth A. Morgan, Job Dekker, Benjamin L. Ebert
Abstract
Clinical trials of biologic therapies in type 1 diabetes (T1D) aim to mitigate autoimmune destruction of pancreatic beta cells through immune perturbation and serve as resources to elucidate immunological mechanisms in health and disease. In the T1DAL trial of alefacept (LFA3-Ig) in recent onset T1D, endogenous insulin production was preserved in 30% of subjects for two years post-therapy. Given our previous findings linking exhausted CD8 T cells to beneficial response in T1D trials, we applied unbiased analyses to sorted CD8 T cells to evaluate their potential role in T1DAL. Using RNA-seq, we found that greater insulin C-peptide preservation was associated with a module of activation- and exhaustion-associated genes. This signature was dissected into two distinct CD8 memory populations through correlation with clustered cytometry data. Both populations were hypo-proliferative, shared expanded TCR junctions, and expressed exhaustion-associated markers including TIGIT and KLRG1. The populations were distinguished by reciprocal expression of CD8 T and NK cell markers (GZMB, CD57 and inhibitory KIR genes), versus T cell activation and differentiation markers (PD1 and CD28). These findings support previous evidence linking exhausted CD8 T cells to successful immune interventions for T1D, while suggesting multiple inhibitory mechanisms can promote this beneficial cell state.
Authors
Kirsten E. Diggins, Elisavet Serti, Virginia S. Muir, Mario G. Rosasco, TingTing Lu, Elisa Balmas, Gerald T. Nepom, S. Alice Long, Peter S. Linsley
Abstract
Human lung adenocarcinoma (LUAD) in current or former smokers exhibits a high tumor mutational burden (TMB) and distinct mutational signatures. Syngeneic mouse models of clinically relevant smoking-related LUAD are lacking. We established and characterized a tobacco-associated transplantable murine LUAD cell line, designated FVBW-17, from a LUAD induced by the tobacco carcinogen 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanone (NNK) in the FVB/N mouse strain. Whole exome sequencing of FVBW-17 cells identified tobacco-associated KrasG12D and Trp53 mutations and a similar mutation profile to that of classic alkylating agents with a TMB >500. FVBW-17 cells transplanted subcutaneously, via tail vein and orthotopically generated tumors in FVB/N mice that were histologically similar to human LUAD. FVBW-17 tumors expressed PD-L1, were infiltrated with CD8+ T cells, and responsive to anti-PD-L1 therapy. FVBW-17 cells were also engineered to express green fluorescent protein and luciferase to facilitate the detection and quantification of tumor growth. Distant metastases to lung, spleen, liver, and kidney were observed from subcutaneously transplanted tumors. This novel cell line is a robust representation of human smoking-related LUAD biology and provides a much needed pre-clinical model in which to test promising new agents and combinations including immune-based therapies.
Authors
Laura P. Stabile, Vinod Kumar, Autumn Gaither-Davis, Eric H.B. Huang, Frank P. Vendetti, Princey Devadassan, Sanja Dacic, Riyue Bao, Richard A. Steinman, Timothy F. Burns, Christopher J. Bakkenist
Abstract
Computational models based on recent maps of the red blood cell proteome suggest that mature erythrocytes may harbor targets for common drugs. This prediction is relevant to red blood cell storage in the blood bank, in which the impact of small molecule drugs or other xenometabolites deriving from dietary, iatrogenic or environmental exposures (“exposome”) may alter erythrocyte energy and redox metabolism and, in so doing, affect red cell storage quality and post-transfusion efficacy. To test this prediction, here we provide a comprehensive characterization of the blood donor exposome, including the detection of common prescription and over-the-counter drugs in 250 units donated by healthy volunteers from the REDS-III RBC Omics study. Based on high-throughput drug screenings of 1,366 FDA-approved drugs, we report a significant impact of ~65% of the tested drugs on erythrocyte metabolism. Machine learning models built using metabolites as predictors were able to accurately predict drugs for several drug classes/targets (bisphosphonates, anticholinergics, calcium channel blockers, adrenergics, proton-pump inhibitors, antimetabolites, selective serotonin reuptake inhibitors, and mTOR) suggesting that these drugs have a direct, conserved, and significant impact on erythrocyte metabolism. As a proof of principle, here we show that the antiacid ranitidine – though rarely detected in the blood donor population – has a strong effect on RBC markers of storage quality in vitro. We thus show that ranitidine supplementation to blood units could improve erythrocyte metabolism and storage quality when supplemented to blood bags, through mechanisms involving sphingosine 1-phosphate-dependent modulation of erythrocyte glycolysis and/or direct binding to hemoglobin.
Authors
Travis Nemkov, Davide Stefanoni, Aarash Bordbar, Aaron Issaian, Bernhard O. Palsson, Larry J. Dumont, Ariel M. Hay, Anren Song, Yang Xia, Jasmina S. Redzic, Elan Z. Eisenmesser, James C. Zimring, Steve Kleinman, Kirk C. Hansen, Michael Busch, Angelo D’Alessandro
Abstract
Asymmetric cell division (ACD) enables the maintenance of a stem cell population while simultaneously generating differentiated progeny. Cancer stem cells (CSCs) undergo multiple modes of cell division during tumor expansion and in response to therapy, yet the functional consequences of these division modes remain to be determined. Using a fluorescent reporter for cell surface receptor distribution during mitosis, we found that ACD generated a daughter cell with enhanced therapeutic resistance and increased co-enrichment of epidermal growth factor receptor (EGFR) and neurotrophin receptor (p75NTR) from a glioblastoma CSC. Stimulation of both receptors antagonized differentiation induction and promoted self-renewal capacity. p75NTR knockdown enhanced the therapeutic efficacy of EGFR inhibition, indicating that co-inheritance of p75NTR and EGFR promotes resistance to EGFR inhibition through a redundant mechanism. These data demonstrate that ACD produces progeny with co-enriched growth factor receptors, which contributes to the generation of a more therapeutically resistant CSC population.
Authors
Masahiro Hitomi, Anastasia P. Chumakova, Daniel J. Silver, Arnon M. Knudsen, W. Dean Pontius, Stephanie Murphy, Neha S. Anand, Bjarne Winther Kristensen, Justin Lathia
Abstract
Paucity of the Glucose Transporter1 (Glut1) protein resulting from haploinsufficiency of the SLC2A1 gene arrests cerebral angiogenesis and disrupts brain function to cause Glucose Transporter1 Deficiency Syndrome (Glut1 DS). Restoring Glut1 to Glut1 DS model mice prevents disease but the precise cellular sites of action of the transporter, its temporal requirements and the mechanism(s) linking scarcity of the protein to brain cell dysfunction remain poorly understood. Here we show that Glut1 functions in a cell-autonomous manner in the cerebral microvasculature to affect endothelial tip cells and thus brain angiogenesis. Moreover, brain endothelial cell-specific Glut1 depletion not only triggers a severe neuro-inflammatory response in the Glut1 DS brain but also reduces levels of brain-derived neurotrophic factor (BDNF) and causes overt disease. Reduced BDNF correlated with fewer neurons in the Glut1 DS brain. Controlled depletion of the protein demonstrated that brain pathology and disease severity was greatest when Glut1 scarcity was induced neonatally, during brain angiogenesis. Reducing Glut1 at later stages had mild or little effect. Our results suggest that targeting brain endothelial cells during early development is important to ensure proper brain angiogenesis, prevent neuro-inflammation, maintain BDNF levels and preserve neuron numbers. This requirement will be essential for any disease-modifying therapeutic strategy for Glut1 DS.
Authors
Maoxue Tang, Sarah H. Park, Sabrina Petri, Hang Yu, Carlos B. Rueda, E. Dale Abel, Carla Y. Kim, Elizabeth M. C. Hillman, Fanghua Li, Yeojin Lee, Lei Ding, Smitha Jagadish, Wayne N. Frankel, Darryl C. De Vivo, Umrao R. Monani
Abstract
Aberrant activation of NLRP3 inflammasome has been implicated in a variety of human inflammatory diseases, however currently no pharmacological NLRP3 inhibitor has been approved in clinic. In this study, we showed that echinatin, the ingredient of the traditional herbal medicine licorice, effectively suppresses the activation of NLRP3 inflammasome in vitro and in vivo. Further investigation revealed that echinatin exerts its inhibitory effect on NLRP3 inflammasome by binding to heat-shock protein 90 (HSP90), inhibiting its ATPase activity, and disrupting the association between the cochaperone SGT1 and HSP90-NLRP3. Importantly, in vivo experiments demonstrated that administration of echinatin obviously inhibits NLRP3 inflammasome activation and ameliorates LPS-induced septic shock and DSS-induced colitis in mice. Moreover, echinatin exerted favorable pharmacological effects on liver inflammation and fibrosis in mouse model of non-alcoholic steatohepatitis (NASH). Collectively, our study identified echinatin as a novel inhibitor of NLRP3 inflammasome and may be developed as a potentially therapeutic approach for the treatment of NLRP3-driven diseases.
Authors
Guang Xu, Shubin Fu, Xiaoyan Zhan, Zhilei Wang, Ping Zhang, Wei Shi, Nan Qin, Yuanyuan Chen, Chunyu Wang, Ming Niu, Yuming Guo, Jia-bo Wang, Zhaofang Bai, Xiaohe Xiao
Abstract
Tumor antigen-specific CD4 T cells accumulate at tumor sites evoking their involvement in antitumor effector functions in situ. Contrarily to CD8 cytotoxic T-lymphocyte exhaustion, that of CD4 T cells remains poorly appreciated. Here, using phenotypic, transcriptomic and functional approaches, we characterized CD4 T-cell exhaustion in head and neck, cervical and ovarian cancer patients. We identified a CD4 tumor-infiltrating lymphocyte (TIL) population, defined by high PD-1 and CD39 expression, which contained high proportions of cytokine-producing cells, although the quantity of cytokines produced by these cells was low evoking an exhausted state. Terminal exhaustion of CD4 TILs was instated regardless of TIM-3 expression suggesting divergence with CD8 T-cell exhaustion. ScRNA-Seq and further phenotypic analyses uncovered, however, similarities with the CD8 T-cell exhaustion program. In particular, PD-1hiCD39+ CD4 TILs expressed the exhaustion transcription factor TOX and the chemokine CXCL13 and were tumor antigen-specific. In vitro, PD-1 blockade enhanced CD4 TIL activation, as evidenced by increased CD154 expression and cytokine secretion, leading to improved dendritic cell maturation and consequently to higher tumor-specific CD8 T-cell proliferation. Our data identify CD4 TIL exhaustion as a player of responsiveness to immune checkpoint blockade.
Authors
Camille-Charlotte Balança, Anna Salvioni, Clara-Maria Scarlata, Marie Michelas, Carlos Martinez-Gomez, Carlos Gomez-Roca, Victor Sarradin, Marie Tosolini, Carine Valle, Frédéric Pont, Gwénaël Ferron, Laurence Gladieff, Sébastien Vergez, Agnès Dupret-Bories, Eliane Mery, Philippe Rochaix, Jean-Jacques Fournié, Jean-Pierre Delord, Christel Devaud, Alejandra Martinez, Maha Ayyoub
Abstract
CIS43 is a potent neutralizing human monoclonal antibody (mAb) that targets a highly conserved ‘junctional’ epitope in the Plasmodium falciparum (Pf) circumsporozoite protein (PfCSP). Enhancing the durability of CIS43 in vivo will be important for clinical translation. Here, two approaches were used to improve the durability of CIS43 in vivo while maintaining potent neutralization. First, the Fc domain was modified with the “LS” mutations (CIS43LS) to increase CIS43 binding affinity for the neonatal Fc receptor (FcRn). CIS43LS and CIS43 showed comparable in vivo protective efficacy. CIS43LS had nine to thirteen-fold increased binding affinity for human (6.2 nM vs. 54.2 nM) and rhesus (25.1 nM vs. 325.8 nM) FcRn at endosomal pH 6.0 compared to CIS43. Importantly, the half-life of CIS43LS in macaques increased from 22 days to 39 days compared to CIS43. The second approach for sustaining antibody levels of CIS43 in vivo is through adeno-associated virus (AAV) expression. Mice administered once with AAV expressing CIS43 had sustained antibody levels of ~ 300 μg/mL and mediated protection against sequential malaria challenges up to 36 weeks. Based on these data, CIS43LS has advanced to Phase I clinical trials and AAV delivery provides a potential next generation approach.
Authors
Neville Kielau Kisalu, Lais Da Silva Pereira, Keenan J. Ernste, Yevel Flores-Garcia, Azza H. Idris, Mangaiarkarasi Asokan, Marlon Dillon, Scott MacDonald, Wei Shi, Xuejun Chen, Amarendra Pegu, Arne Schön, Fidel Zavala, Alejandro B. Balazs, Joseph R. Francica, Robert A. Seder
Abstract
Subjects with obesity frequently have elevated serum vasopressin levels, noted by the stable analog, copeptin. Vasopressin acts primarily to reabsorb water via urinary concentration. However, fat is also a source of metabolic water, raising the possibility that vasopressin might have a role in fat accumulation. Fructose has also been reported to stimulate vasopressin. Here we tested the hypothesis that fructose induced metabolic syndrome is mediated by vasopressin. Orally administered fructose, glucose or high fructose corn syrup increased vasopressin (copeptin) concentrations and was mediated by fructokinase, an enzyme specific for fructose metabolism. Suppressing vasopressin with hydration both prevented and ameliorated fructose-induced metabolic syndrome. The vasopressin effects were mediated by the Vasopressin 1b receptor, as Vasopressin 1b receptor knockout mice were completely protected while V1a knockout paradoxically showed worse metabolic syndrome. The mechanism is likely mediated in part by de novo expression of V1b in the liver that amplifies fructokinase expression in response to fructose. Thus, our studies document a new role for vasopressin in water conservation via the accumulation of fat as a source of metabolic water. Clinically, it also suggests that increased water intake may be a beneficial way to both prevent or treat metabolic syndrome.
Authors
Ana Andres-Hernando, Thomas J. Jensen, Masanari Kuwabara, David J. Orlicky, Christina Cicerchi, Nanxing Li, Carlos A. Roncal-Jimenez, Gabriela E. Garcia, Takuji Ishimoto, Paul S. Maclean, Petter Bjornstad, Laura Gabriela Sanchez-Lozada, Mehmet Kanbay, Takahiko Nakagawa, Richard Johnson, Miguel Lanaspa
Abstract
The G/T transversion, rs35705950, located approximately 3 kb upstream of the MUC5B start site, is the cardinal risk factor for idiopathic pulmonary fibrosis (IPF). Here, we investigate the function and chromatin structure of this -3 kb region and provide evidence that it functions as a classically defined enhancer subject to epigenetic programming. We use nascent transcript analysis to show that RNA polymerase II loads within 10 bp of the G/T transversion site, definitively establishing enhancer function for the region. By integrating Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) analysis of fresh and cultured human airway epithelial cells with nuclease sensitivity data, we demonstrate that this region is in accessible chromatin that affects the expression of MUC5B. Through applying paired single nucleus RNA- and ATAC-seq to frozen tissue from IPF lungs, we extend these findings directly to disease, with results indicating that epigenetic programming of the -3 kb enhancer in IPF occurs in both MUC5B-expressing and non-expressing lineages. In aggregate, our results indicate that the MUC5B-associated variant, rs35705950, resides within an enhancer that is subject to epigenetic remodeling and contributes to pathologic misexpression in IPF.
Authors
Fabienne Gally, Sarah K. Sasse, Jonathan Kurche, Margaret A. Gruca, Jonathan H. Cardwell, Tsukasa Okamoto, Hong Wei Chu, Xiaomeng Hou, Olivier Poirion, Justin Buchanan, Sebastian Preissl, Bing Ren, Sean P. Colgan, Robin D. Dowell, Ivana V. Yang, David A. Schwartz, Anthony N. Gerber
Abstract
Limited experimental evidence bridges nutrition and cancer immunosurveillance. Here, we show that ketogenic diet (KD) or its principal ketone body, 3-hydroxybutyrate (3HB), most specifically in an intermittent scheduling, induced T cell-dependent tumor growth retardation of aggressive tumor models. In conditions in which anti-PD-1, alone or in combination with anti-CTLA-4, failed to reduce tumor growth in mice receiving a standard diet, KD or oral supplementation of 3HB reestablished therapeutic responses. Supplementation of KD with sucrose (which breaks ketogenesis, abolishing 3HB production) or with a pharmacological antagonist of the 3HB receptor GPR109A abolished the antitumor effects. Mechanistically, 3HB prevented the ICB-linked upregulation of PD-L1 on myeloid cells while favoring the expansion of CXCR3+ T cells. KD induced compositional changes of the gut microbiota with distinct species such as Eisenbergiella massiliensis commonly emerging in mice and humans subjected to carbohydrate low diet interventions and highly correlating with serum concentrations of 3HB. Altogether, these results demonstrate that KD induces a 3HB-mediated antineoplastic effect that relies on T-cell mediated cancer immunosurveillance.
Authors
Gladys Ferrere, Maryam Tidjani Alou, Peng Liu, Anne-Gaëlle Goubet, Marine Fidelle, Oliver Kepp, Sylvère Durand, Valerio Iebba, Aurélie Fluckiger, Romain Daillère, Cassandra Thelemaque, Claudia Grajeda-Iglesias, Carolina Alves Costa Silva, Fanny Aprahamian, Deborah Lefevre, Liwei Zhao, Bernhard Ryffel, Emeline Colomba, Monica Arnedos, Damien Drubay, Conrad Rauber, Didier Raoult, Francesco Asnicar, Tim Spector, Nicola Segata, Lisa Derosa, Guido Kroemer, Laurence Zitvogel.
A roadmap from single-cell transcriptome to patient classification for the immune response to trauma
Abstract
Immune dysfunction is an important factor driving mortality and adverse outcomes after trauma but remains poorly understood, especially at cellular level. To deconvolute trauma-induced immune response, we applied single-cell RNA sequencing to circulating and bone marrow mononuclear cells in injured mice and circulating mononuclear cells in trauma patients. In mice, the greatest changes in gene expression were seen in monocytes across both compartments. After systemic injury, the gene expression pattern of monocytes markedly deviated from steady state with corresponding changes in critical transcription factors (TFs), which can be traced back to myeloid progenitors. These changes were largely recapitulated in human single-cell analysis. We generalized the major changes in human CD14+ monocytes into six signatures, which further defined two trauma patient subtypes (SG1 vs. SG2) identified in the whole blood leukocyte transcriptome in the initial 12h after injury. Compared with SG2, SG1 patients exhibited delayed recovery, more severe organ dysfunction and a higher incidence of infection and non-infectious complications. The two patient subtypes were also recapitulated in burn and sepsis patients, revealing a shared pattern of immune response across critical illness. Our data will be broadly useful to further explore the immune response to inflammatory diseases and critical illness.
Authors
Tianmeng Chen, Matthew J. Delano, Kong Chen, Jason L. Sperry, Rami A. Namas, Ashley J. Lamparello, Meihong Deng, Julia Conroy, Lyle L. Moldawer, Philip A. Efron, Patricia A. Loughran, Christopher W. Seymour, Derek C. Angus, Yoram Vodovotz, Wei Chen, Timothy R. Billiar
Abstract
Management of Gastrointestinal stromal tumor (GIST) has been revolutionized by the identification of activating mutations in KIT and PDGFRA and clinical application of receptor tyrosine kinase (RTK) inhibitors in advanced disease. Stratification of GIST into molecularly defined subsets provides insight into clinical behavior and response to approved targeted therapies. Although these RTK inhibitors are effective in most GIST, resistance remains a significant clinical problem. Development of effective treatment strategies for refractory GIST requires identification of novel targets to provide additional therapeutic options. Global kinome profiling has potential to identify critical signaling networks and reveal protein kinases essential in GIST. Using Multiplexed Inhibitor Beads and Mass Spectrometry, we explored the majority of the kinome in GIST specimens from the three most common molecular subtypes (KIT-mutant, PDGFRA-mutant, Succinate dehydrogenase (SDH)-deficient) to identify novel kinase targets. Kinome profiling with loss-of-function assays identified an important role for G2-M tyrosine kinase, Wee1, in GIST cell survival. In vitro and in vivo studies revealed significant efficacy of MK-1775 (Wee1 inhibitor) in combination with avapritinib in KIT and PDGFRA-mutant GIST cell lines, and notable efficacy of MK-1775 as a monotherapy in the PDGFRA-mutant line. These studies provide strong preclinical justification for the use of MK-1775 in GIST.
Authors
Shuai Ye, Dinara Sharipova, Marya Kozinova, Lillian R. Klug, Jimson W. D'Souza, Martin G. Belinsky, Katherine J. Johnson, Margret B. Einarson, Karthik Devarajan, Yan Zhou, Samuel Litwin, Michael C. Heinrich, Ronald P. DeMatteo, Margaret von Mehren, James S. Duncan, Lori Rink
Abstract
COPD is a chronic respiratory disease characterized by small airway remodeling and alveolar emphysema due to environmental stresses such as cigarette smoking (CS). Oxidative stress is commonly implicated in COPD pathology, but recent findings suggest that one oxidant-producing NADPH oxidase homolog, dual oxidase 1 (DUOX1), is downregulated in the airways of COPD patients. We evaluated lung tissue sections from COPD patients for small airway epithelial DUOX1 protein expression, in association with measures of lung function and small airway and alveolar remodeling. We also addressed the impact of DUOX1 for lung tissue remodeling in mouse models of COPD. Small airway DUOX1 levels were decreased in advanced COPD, and correlated with loss of lung function and markers of emphysema and remodeling. Similarly, DUOX1 downregulation in correlation with extracellular matrix remodeling was observed in a genetic model of COPD, transgenic SPC-TNF-α mice. Finally, development of subepithelial airway fibrosis in mice due to exposure to the CS-component acrolein, or alveolar emphysema induced by administration of elastase, were in both cases exacerbated in Duox1-deficient mice. Collectively, our studies highlight that downregulation of DUOX1 may be a contributing feature of COPD pathogenesis, likely related to impaired DUOX1-mediated innate injury responses involved in epithelial homeostasis.
Authors
Caspar Schiffers, Cheryl van de Wetering, Robert A. Bauer, Aida Habibovic, Milena Hristova, Christopher M. Dustin, Sara Lambrichts, Pamela M. Vacek, Emiel F.M. Wouters, Niki L. Reynaert, Albert van der Vliet
Abstract
Transient partial remission, a period of low insulin requirement experienced by most patients soon after diagnosis has been associated with mechanisms of immune regulation. A better understanding of such natural mechanisms of immune regulation might identify new targets for immunotherapies that reverse T1D. In this study, using Cox model multivariate analysis we validate our previous findings that patients (n = 84) with the highest frequency of CD4+ CD25+CD127hi (127-hi) cells at diagnosis experience the longest partial remission and we show that the 127-hi cell population is a mix of Th1- and Th2-type cells with a significant bias towards anti-inflammatory Th2-type cells. In addition, we extend these findings to show that patients with the highest frequency of 127-hi cells at diagnosis are significantly more likely to maintain beta-cell function. Moreover, in patients treated with Alefacept in the TIDAL clinical trial, the probability of responding favorably to the anti-inflammatory drug was significantly higher in those with a higher frequency of 127-hi cells at diagnosis than those with a lower 127-hi cell frequency. These data are consistent with the hypothesis that 127-hi cells maintain an anti-inflammatory environment that is permissive for partial remission, beta-cell survival and response to anti-inflammatory immunotherapy.
Authors
Aditi Narsale, Breanna Lam, Rosita Moya, TingTing Lu, Alessandra Mandelli, Irene Gotuzzo, Benedetta Pessina, Gian Maria Giamporcaro, Rhonda Geoffrey, Kerry Buchanan, Mark Harris, Anne-Sophie Bergot, Ranjeny Thomas, Martin J. Hessner, Manuela Battaglia, Elisavet Serti, Joanna D. Davies
