DOCSLIB.ORG

Impact of the Major Histocompatibility Complex Class I Peptide Repertoire on Natural Killer Cell Function

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Impact of the Major Histocompatibility Complex Class I Peptide Repertoire on Natural Killer Cell Function UNIVERSITY OF SOUTHAMPTON FACULTY OF MEDICINE Clinical and Experimental Sciences Impact of the Major Histocompatibility Complex class I peptide repertoire on Natural Killer cell function by Berenice Mbiribindi Nvunabandi Thesis for the degree of Doctor of Philosophy December 2016 UNIVERSITY OF SOUTHAMPTON ABSTRACT FACULTY OF MEDICINE Clinical and Experimental Sciences Transfer Thesis for the degree of Doctor of Philosophy IMPACT OF THE MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I PEPTIDE REPERTOIRE ON NATURAL KILLER CELL FUNCTION Berenice Mbiribindi Nvunabandi Background: Natural Killer (NK) cell activation requires the integration of inhibitory and activating signalling. Inhibitory signals are determined by members of the KIR family, which have MHC class I ligands. Peptide antagonism of MHC class I provides an alternative mechanism for loss of inhibition of KIR2DL2/3-positive NK cells. This occurs when a weak KIR binding peptide disrupts the inhibitory signalling of a strong binding peptide. Peptide antagonism has been defined only for HLA-C*0102, endogenously expressed in TAP-deficient cells, using a peptide variant of VAPWNSLSL (VAPWNSDAL and VAPWNSDYL), therefore it may be a unique property of HLA-C*0102 or more general finding. Hypothesis: We hypothesize that small changes in MHC-I peptide repertoire can impact on NK cell activation and that additional peptides can antagonise inhibitory KIR can be discovered. Results: 721.221 cells were transfected with HLA-C*0304 and ICP47 used for peptide loading and as target cells in CD107a assays. We studied previously described position 8 (P8) derivatives and also generated novel P7 derivatives of the endogenously processed peptide GAVDPLLAL. In contrast to our observations for HLA-C*0102, arginine at P7 triggered stronger inhibition than phenylalanine. L7D did not inhibit KIR2DL2/3-positive NK cells. However it did antagonise inhibition by L7R in CD107a assays. I used peptide elution and HPLC analysis to demonstrate that peptide antagonism was not related to displacement of L7R or L7F by LD7. In a study of the immunopeptidome of Hepatitis C virus expressing cells expressing HLA- C*0102 were generated to study its peptide repertoire and to identify potential NK activating or inhibitory peptides. I identified that the peptide repertoire of HLA-C was preferentially altered in comparison to HLA-A. To improve our methods for studying the influence of peptides on NK cell, I developed a Granzyme B assay based on HPLC technology. This showed that it is possible to perform a functional assay using NK lines with low backgrounds, however it was not robust enough to detect small differences in peptides. Conlcusions: Peptide antagonism is a generalizable phenomenon and it appears that HLA-C alleles has become specialised to NK cells. Contents Contents ................................................................................................................................... i List of tables ...................................................................................................................... vii List of figures ......................................................................................................................ix List of accompanying materials ............................................................................. xv DECLARATION OF AUTHORSHIP ........................................................................... xvii Acknowledgements .................................................................................................... xviii Definitions and Abbreviations ............................................................................... xix 1. Introduction................................................................................................................. 1 1.1 Natural Killer cells ....................................................................... 1 1.1.1 General description ..................................................................... 1 1.1.2 NK cells development: transcription factors and lineage commitment 1 1.1.3 NK cells development: the journey from the bone marrow to the periphery .................................................................................................. 3 1.2 NK cell education ........................................................................ 4 1.2.1 Chronic exposure of NK cells to activating ligands make them tolerant. 4 1.2.2 Inhibitory receptor repertoire selectivity of NK cells receptors ..... 6 1.2.3 Self-MHC Class I and Inhibitory receptors recognition is involved in NK Cell Education ........................................................................................... 8 1.2.4 Mechanisms leading to hyporesponsive NK cells: licensing or disarming? 9 1.3 NK cells in health and disease ................................................... 11 1.3.1 NK cells in cancer ..................................................................... 11 1.3.2 NK cells in pregnancy ............................................................... 12 1.3.3 NK cells in infection .................................................................. 13 1.3.4 NK cells in autoimmune diseases .............................................. 15 1.4 Therapeutic applications of NK cells ......................................... 17 1.4.1 NK cells in Liver disease ............................................................ 19 1.4.2 NK cells as therapeutic targets for the treatment of liver disease20 1.5 The molecular specificity of NK cell receptors ........................... 21 1.5.1 Receptor-mediated NK cell responses ....................................... 22 1.5.2 NK cell recognition and response receptors .............................. 23 1.5.3 Activating receptors .................................................................. 24 1.5.4 Inhibitory receptors .................................................................. 27 i 1.6 Killer Immunoglobulin like Receptors: KIR ................................. 28 1.6.1 Structural and diversity of KIR receptors ................................... 28 1.6.2 Structural basis of KIR-HLA recognition ..................................... 30 1.6.3 HLA-C Recognition by inhibitory HLA-C specific KIR ................... 32 1.6.4 The role of the peptide: peptide presentation ........................... 33 1.6.5 The role of the peptide: KIR recognition .................................... 36 1.6.6 NK cells inhibitory receptors and peptide selectivity .................. 37 1.6.7 Antagonism of NK cells inhibition through inhibitory receptors . 38 1.6.8 Considerations for viral infections............................................. 40 1.7 Specificity of Activating KIR2D receptors ................................... 41 1.8 NK cell activation ...................................................................... 42 1.9 Hypothesis, aims and Experimental plan ................................... 43 1.9.1 Aim 1: Demonstrate that peptide antagonism is not restricted to only one HLA-C allele ...................................................................................... 44 1.9.2 Aim 2: Design of a novel method for peptide high throughput screen …………………………………………………………………………………44 1.9.3 Aim 3: Identifying viral induced changes in peptide repertoire and assessing how this may affect NK cell reactivity ....................................... 45 2. Materials and Methods ...................................................................................... 46 2.1 Peptides ................................................................................... 46 2.2 Cells lines, PBMC and cell culture .............................................. 46 2.3 ICP47 gene transfection in 721.221:C*0304 cells ..................... 47 2.4 Peptide stabilization assay ........................................................ 48 2.5 Measurement of the decay of cell surface HLA-C molecules ....... 49 2.6 Degranulation Assays ............................................................... 49 2.7 Peptide elution from MHC class I and HPLC analysis .................. 50 2.8 PBMC DNA extraction ................................................................ 51 2.9 KIR genotyping ......................................................................... 52 2.10 Confocal microscopy ................................................................ 53 2.11 KIR binding assay ..................................................................... 53 2.11.1 Affinity of P7 peptide variants for KIR ....................................... 53 2.11.2 KIR binding and peptide titration .............................................. 54 2.11.3 KIR binding, HLA-C and Degranulation Assay ............................ 54 2.12 Minigenes for endogenous peptide presentation ....................... 55 2.12.1 Minigene preparation ................................................................ 55 2.12.2 Minigenes transformation into competent cells ......................... 56 2.12.3 Extraction of plasmids containing minigenes ............................ 56 ii 2.13 Transfection of minigenes constructs into 721.221:C*0304 cells by electroporation ......................................................................... 57 2.14 Cytotoxicity Assay using Propidium iodide (PI) .......................... 58 2.15 Granzyme B Assay optimisation ................................................ 58 2.15.1 Determining the optimal wavelength for Caspase 3 Peptide (C3P) detection 58 2.15.2 Determining the optimal conditions for
Load more

Recommended publications
  • ENSG Gene Encodes Effector TCR Pathway Costimulation Inhibitory/Exhaustion Synapse/Adhesion Chemokines/Receptors
    ENSG Gene Encodes Effector TCR pathway Costimulation Inhibitory/exhaustion Synapse/adhesion Chemokines/receptors ENSG00000111537 IFNG IFNg x ENSG00000109471 IL2 IL-2 x ENSG00000232810 TNF TNFa x ENSG00000271503 CCL5 CCL5 x x ENSG00000139187 KLRG1 Klrg1 x ENSG00000117560 FASLG Fas ligand x ENSG00000121858 TNFSF10 TRAIL x ENSG00000134545 KLRC1 Klrc1 / NKG2A x ENSG00000213809 KLRK1 Klrk1 / NKG2D x ENSG00000188389 PDCD1 PD-1 x x ENSG00000117281 CD160 CD160 x x ENSG00000134460 IL2RA IL-2 receptor x subunit alpha ENSG00000110324 IL10RA IL-10 receptor x subunit alpha ENSG00000115604 IL18R1 IL-18 receptor 1 x ENSG00000115607 IL18RAP IL-18 receptor x accessory protein ENSG00000081985 IL12RB2 IL-12 receptor x beta 2 ENSG00000186810 CXCR3 CXCR3 x x ENSG00000005844 ITGAL CD11a x ENSG00000160255 ITGB2 CD18; Integrin x x beta-2 ENSG00000156886 ITGAD CD11d x ENSG00000140678 ITGAX; CD11c x x Integrin alpha-X ENSG00000115232 ITGA4 CD49d; Integrin x x alpha-4 ENSG00000169896 ITGAM CD11b; Integrin x x alpha-M ENSG00000138378 STAT4 Stat4 x ENSG00000115415 STAT1 Stat1 x ENSG00000170581 STAT2 Stat2 x ENSG00000126561 STAT5a Stat5a x ENSG00000162434 JAK1 Jak1 x ENSG00000100453 GZMB Granzyme B x ENSG00000145649 GZMA Granzyme A x ENSG00000180644 PRF1 Perforin 1 x ENSG00000115523 GNLY Granulysin x ENSG00000100450 GZMH Granzyme H x ENSG00000113088 GZMK Granzyme K x ENSG00000057657 PRDM1 Blimp-1 x ENSG00000073861 TBX21 T-bet x ENSG00000115738 ID2 ID2 x ENSG00000176083 ZNF683 Hobit x ENSG00000137265 IRF4 Interferon x regulatory factor 4 ENSG00000140968 IRF8 Interferon
    [Show full text]
  • Human and Mouse CD Marker Handbook Human and Mouse CD Marker Key Markers - Human Key Markers - Mouse
    Welcome to More Choice CD Marker Handbook For more information, please visit: Human bdbiosciences.com/eu/go/humancdmarkers Mouse bdbiosciences.com/eu/go/mousecdmarkers Human and Mouse CD Marker Handbook Human and Mouse CD Marker Key Markers - Human Key Markers - Mouse CD3 CD3 CD (cluster of differentiation) molecules are cell surface markers T Cell CD4 CD4 useful for the identification and characterization of leukocytes. The CD CD8 CD8 nomenclature was developed and is maintained through the HLDA (Human Leukocyte Differentiation Antigens) workshop started in 1982. CD45R/B220 CD19 CD19 The goal is to provide standardization of monoclonal antibodies to B Cell CD20 CD22 (B cell activation marker) human antigens across laboratories. To characterize or “workshop” the antibodies, multiple laboratories carry out blind analyses of antibodies. These results independently validate antibody specificity. CD11c CD11c Dendritic Cell CD123 CD123 While the CD nomenclature has been developed for use with human antigens, it is applied to corresponding mouse antigens as well as antigens from other species. However, the mouse and other species NK Cell CD56 CD335 (NKp46) antibodies are not tested by HLDA. Human CD markers were reviewed by the HLDA. New CD markers Stem Cell/ CD34 CD34 were established at the HLDA9 meeting held in Barcelona in 2010. For Precursor hematopoetic stem cell only hematopoetic stem cell only additional information and CD markers please visit www.hcdm.org. Macrophage/ CD14 CD11b/ Mac-1 Monocyte CD33 Ly-71 (F4/80) CD66b Granulocyte CD66b Gr-1/Ly6G Ly6C CD41 CD41 CD61 (Integrin b3) CD61 Platelet CD9 CD62 CD62P (activated platelets) CD235a CD235a Erythrocyte Ter-119 CD146 MECA-32 CD106 CD146 Endothelial Cell CD31 CD62E (activated endothelial cells) Epithelial Cell CD236 CD326 (EPCAM1) For Research Use Only.
    [Show full text]
  • Tools for Cell Therapy and Immunoregulation
    RnDSy-lu-2945 Tools for Cell Therapy and Immunoregulation Target Cell TIM-4 SLAM/CD150 BTNL8 PD-L2/B7-DC B7-H1/PD-L1 (Human) Unknown PD-1 B7-1/CD80 TIM-1 SLAM/CD150 Receptor TIM Family SLAM Family Butyrophilins B7/CD28 Families T Cell Multiple Co-Signaling Molecules Co-stimulatory Co-inhibitory Ig Superfamily Regulate T Cell Activation Target Cell T Cell Target Cell T Cell B7-1/CD80 B7-H1/PD-L1 T cell activation requires two signals: 1) recognition of the antigenic peptide/ B7-1/CD80 B7-2/CD86 CTLA-4 major histocompatibility complex (MHC) by the T cell receptor (TCR) and 2) CD28 antigen-independent co-stimulation induced by interactions between B7-2/CD86 B7-H1/PD-L1 B7-1/CD80 co-signaling molecules expressed on target cells, such as antigen-presenting PD-L2/B7-DC PD-1 ICOS cells (APCs), and their T cell-expressed receptors. Engagement of the TCR in B7-H2/ICOS L 2Ig B7-H3 (Mouse) the absence of this second co-stimulatory signal typically results in T cell B7-H1/PD-L1 B7/CD28 Families 4Ig B7-H3 (Human) anergy or apoptosis. In addition, T cell activation can be negatively regulated Unknown Receptors by co-inhibitory molecules present on APCs. Therefore, integration of the 2Ig B7-H3 Unknown B7-H4 (Mouse) Receptors signals transduced by co-stimulatory and co-inhibitory molecules following TCR B7-H5 4Ig B7-H3 engagement directs the outcome and magnitude of a T cell response Unknown Ligand (Human) B7-H5 including the enhancement or suppression of T cell proliferation, B7-H7 Unknown Receptor differentiation, and/or cytokine secretion.
    [Show full text]
  • Supplementary Table 1: Adhesion Genes Data Set
    Supplementary Table 1: Adhesion genes data set PROBE Entrez Gene ID Celera Gene ID Gene_Symbol Gene_Name 160832 1 hCG201364.3 A1BG alpha-1-B glycoprotein 223658 1 hCG201364.3 A1BG alpha-1-B glycoprotein 212988 102 hCG40040.3 ADAM10 ADAM metallopeptidase domain 10 133411 4185 hCG28232.2 ADAM11 ADAM metallopeptidase domain 11 110695 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 195222 8038 hCG40937.4 ADAM12 ADAM metallopeptidase domain 12 (meltrin alpha) 165344 8751 hCG20021.3 ADAM15 ADAM metallopeptidase domain 15 (metargidin) 189065 6868 null ADAM17 ADAM metallopeptidase domain 17 (tumor necrosis factor, alpha, converting enzyme) 108119 8728 hCG15398.4 ADAM19 ADAM metallopeptidase domain 19 (meltrin beta) 117763 8748 hCG20675.3 ADAM20 ADAM metallopeptidase domain 20 126448 8747 hCG1785634.2 ADAM21 ADAM metallopeptidase domain 21 208981 8747 hCG1785634.2|hCG2042897 ADAM21 ADAM metallopeptidase domain 21 180903 53616 hCG17212.4 ADAM22 ADAM metallopeptidase domain 22 177272 8745 hCG1811623.1 ADAM23 ADAM metallopeptidase domain 23 102384 10863 hCG1818505.1 ADAM28 ADAM metallopeptidase domain 28 119968 11086 hCG1786734.2 ADAM29 ADAM metallopeptidase domain 29 205542 11085 hCG1997196.1 ADAM30 ADAM metallopeptidase domain 30 148417 80332 hCG39255.4 ADAM33 ADAM metallopeptidase domain 33 140492 8756 hCG1789002.2 ADAM7 ADAM metallopeptidase domain 7 122603 101 hCG1816947.1 ADAM8 ADAM metallopeptidase domain 8 183965 8754 hCG1996391 ADAM9 ADAM metallopeptidase domain 9 (meltrin gamma) 129974 27299 hCG15447.3 ADAMDEC1 ADAM-like,
    [Show full text]
  • CD Markers Are Routinely Used for the Immunophenotyping of Cells
    ptglab.com 1 CD MARKER ANTIBODIES www.ptglab.com Introduction The cluster of differentiation (abbreviated as CD) is a protocol used for the identification and investigation of cell surface molecules. So-called CD markers are routinely used for the immunophenotyping of cells. Despite this use, they are not limited to roles in the immune system and perform a variety of roles in cell differentiation, adhesion, migration, blood clotting, gamete fertilization, amino acid transport and apoptosis, among many others. As such, Proteintech’s mini catalog featuring its antibodies targeting CD markers is applicable to a wide range of research disciplines. PRODUCT FOCUS PECAM1 Platelet endothelial cell adhesion of blood vessels – making up a large portion molecule-1 (PECAM1), also known as cluster of its intracellular junctions. PECAM-1 is also CD Number of differentiation 31 (CD31), is a member of present on the surface of hematopoietic the immunoglobulin gene superfamily of cell cells and immune cells including platelets, CD31 adhesion molecules. It is highly expressed monocytes, neutrophils, natural killer cells, on the surface of the endothelium – the thin megakaryocytes and some types of T-cell. Catalog Number layer of endothelial cells lining the interior 11256-1-AP Type Rabbit Polyclonal Applications ELISA, FC, IF, IHC, IP, WB 16 Publications Immunohistochemical of paraffin-embedded Figure 1: Immunofluorescence staining human hepatocirrhosis using PECAM1, CD31 of PECAM1 (11256-1-AP), Alexa 488 goat antibody (11265-1-AP) at a dilution of 1:50 anti-rabbit (green), and smooth muscle KD/KO Validated (40x objective). alpha-actin (red), courtesy of Nicola Smart. PECAM1: Customer Testimonial Nicola Smart, a cardiovascular researcher “As you can see [the immunostaining] is and a group leader at the University of extremely clean and specific [and] displays Oxford, has said of the PECAM1 antibody strong intercellular junction expression, (11265-1-AP) that it “worked beautifully as expected for a cell adhesion molecule.” on every occasion I’ve tried it.” Proteintech thanks Dr.
    [Show full text]
  • Immune Evasion by Herpes Simplex Viruses
    Chapter 5 Immune Evasion by Herpes Simplex Viruses Angello R. Retamal-Díaz, Eduardo Tognarelli, Alexis M. Kalergis, Susan M. Bueno and Pablo A. González Additional information is available at the end of the chapter http://dx.doi.org/10.5772/64128 Abstract Infection with herpes simplex viruses type 1 (HSV-1) and type 2 (HSV-2) is extremely frequent in the human population, as well as recurrent reactivations due to lifelong infection. Infection and persistence of HSVs within healthy individuals likely results as a consequence of numerous molecular determinants evolved by these pathogens to escape both immediate and long-term host antiviral mechanisms. Indeed, HSVs harbor an arsenal of proteins that confer them stealth by negatively modulating immune function. Consequently, these viruses perpetuate within the host, altogether silently shedding onto other individuals. In this chapter, we discuss HSV determinants that interfere with cellular antiviral factors, as well as viral determinants that hamper innate and adaptive immune components intended to control such microbes. The identifica‐ tion of HSV evasion molecules that modulate the immune system, as well as the understanding of their mechanisms of action, should facilitate the design of novel prophylactic and therapeutic strategies to overcome infection and disease elicited by these viruses. This chapter is intended to provide an overview of the evasion mecha‐ nisms evolved by herpes simplex viruses to escape numerous host antiviral mediators. Keywords: immune evasion, innate immunity, adaptive immunity, antiviral response 1. Introduction Herpes simplex viruses (HSVs, HSV-1 and HSV-2) are extremely prevalent in the human population with virtually half of the world inhabitants infected with HSV-1 [1] and nearly 500 million with HSV-2 [2].
    [Show full text]
  • Supplemental Figures 032819.Pptx
    Summary of Supplemental Figures and Tables 1) Supplemental Figure 1. Frequency of phenotypically defined endothelial cells is unchanged in aged mice. 2) Supplemental Figure 2. Aged LSKs have increased myeloid/megakaryocytic bias 3) Supplemental Figure 3. RBCs are decreased and Platelets are increased in peripheral blood of aged mice 4) Supplemental Figure 4. Aged BMME cultures have increased MSC populations. 5) Supplemental Figure 5. Aged BMME cells increase young LSK cell engraftment following competitive transplantation 6) Supplemental Figure 6. Flow cytometry gating strategy for sorting of young and aged murine Mφs 7) Supplemental Figure 7. Flow cytometry gating strategy for sorting of human Mφs 8) Supplemental Figure 8. Axl-/- LT-HSCs have increased cell engraftment following competitive transplantation 9) Supplemental Figure 9. Schematic representation of mechanisms by which age-dependent defects in marrow Mφs induce megakaryocytic bias in HSC. 10) Table S1. List of antibodies used in the flow cytometric and cell sorting analyses described 11) Table S2. List of cell populations analyzed and their respective immunophenotypes 12) Table S3. List of top GO-BP categories enriched for significantly upregulated genes from murine aged vs young marrow macrophages 13) Table S4. List of significant KEGG Pathways enriched for significantly upregulated genes from murine aged vs young marrow macrophages 1 A Sinusoidal Arteriolar BV786 – CD31 Sca1 – PE-Cy7 B C Arteriolar EC Sinusoidal ECs (Lin-, CD45-, CD31+ Sca1+) (CD45-Lin-CD31+Sca1-) 0.08 0.15 0.06 0.10 0.04 0.05 0.02 PercentLiveof Cells PercentLiveof Cells 0.00 0.00 Young Aged Young Aged Supplemental Figure 1.
    [Show full text]
  • The Impact of HSV-1 Infection, SOCS1 Peptide, And
    Wright State University CORE Scholar Browse all Theses and Dissertations Theses and Dissertations 2016 The Impact of HSV-1 Infection, SOCS1 Peptide, and SOCS3 Peptide Mimetic on Cell Viability, Morphology, and Cytoskeleton Proteins of Unpolarized and Cytokine-Polarized M1 RAW 264.7 Murine Macrophages Mubarak Almutairi Wright State University Follow this and additional works at: https://corescholar.libraries.wright.edu/etd_all Part of the Immunology and Infectious Disease Commons, and the Microbiology Commons Repository Citation Almutairi, Mubarak, "The Impact of HSV-1 Infection, SOCS1 Peptide, and SOCS3 Peptide Mimetic on Cell Viability, Morphology, and Cytoskeleton Proteins of Unpolarized and Cytokine-Polarized M1 RAW 264.7 Murine Macrophages" (2016). Browse all Theses and Dissertations. 1513. https://corescholar.libraries.wright.edu/etd_all/1513 This Thesis is brought to you for free and open access by the Theses and Dissertations at CORE Scholar. It has been accepted for inclusion in Browse all Theses and Dissertations by an authorized administrator of CORE Scholar. For more information, please contact [email protected]. The Impact of HSV-1 Infection, SOCS1 peptide, and SOCS3 peptide mimetic on Cell Viability, Morphology, and Cytoskeleton Proteins of Unpolarized and Cytokine-Polarized M1 RAW 264.7 Murine Macrophages A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science. By Mubarak Huraysan Almutairi B.S., King Saud University, 2010 2016 Wright State University WRIGHT STATE UNIVERSITY THE GRADUATE SCHOOL April 25, 2016 I HEREBY RECOMMEND THAT THE THESIS PREPARED UNDER MY SUPERVISION BY Mubarak Huraysan Almutairi ENTITLED The Impact of HSV-1 Infection, SOCS1 peptide, and SOCS3 peptide mimetic on Cell Viability, Morphology, and Cytoskeleton Proteins of Unpolarized and Cytokine- Polarized M1 RAW 264.7 Murine Macrophages BE ACCEPTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF Master of Science.
    [Show full text]
  • A Novel Ebola Virus VP40 Matrix Protein-Based Screening for Identification of Novel Candidate Medical Countermeasures
    viruses Communication A Novel Ebola Virus VP40 Matrix Protein-Based Screening for Identification of Novel Candidate Medical Countermeasures Ryan P. Bennett 1,† , Courtney L. Finch 2,† , Elena N. Postnikova 2 , Ryan A. Stewart 1, Yingyun Cai 2 , Shuiqing Yu 2 , Janie Liang 2, Julie Dyall 2 , Jason D. Salter 1 , Harold C. Smith 1,* and Jens H. Kuhn 2,* 1 OyaGen, Inc., 77 Ridgeland Road, Rochester, NY 14623, USA; [email protected] (R.P.B.); [email protected] (R.A.S.); [email protected] (J.D.S.) 2 NIH/NIAID/DCR/Integrated Research Facility at Fort Detrick (IRF-Frederick), Frederick, MD 21702, USA; courtney.fi[email protected] (C.L.F.); [email protected] (E.N.P.); [email protected] (Y.C.); [email protected] (S.Y.); [email protected] (J.L.); [email protected] (J.D.) * Correspondence: [email protected] (H.C.S.); [email protected] (J.H.K.); Tel.: +1-585-697-4351 (H.C.S.); +1-301-631-7245 (J.H.K.) † These authors contributed equally to this work. Abstract: Filoviruses, such as Ebola virus and Marburg virus, are of significant human health concern. From 2013 to 2016, Ebola virus caused 11,323 fatalities in Western Africa. Since 2018, two Ebola virus disease outbreaks in the Democratic Republic of the Congo resulted in 2354 fatalities. Although there is progress in medical countermeasure (MCM) development (in particular, vaccines and antibody- based therapeutics), the need for efficacious small-molecule therapeutics remains unmet. Here we describe a novel high-throughput screening assay to identify inhibitors of Ebola virus VP40 matrix protein association with viral particle assembly sites on the interior of the host cell plasma membrane.
    [Show full text]
  • Download Special Issue
    Advances in Virology Virus Immune Evasion: New Mechanism and Implications in Disease Outcome Guest Editors: Rika Draenert, John Frater, and Julia G. Prado Virus Immune Evasion: New Mechanism and Implications in Disease Outcome Advances in Virology Virus Immune Evasion: New Mechanism and Implications in Disease Outcome Guest Editors: Rika Draenert, John Frater, and Julia G. Prado Copyright © 2012 Hindawi Publishing Corporation. All rights reserved. This is a special issue published in “Advances in Virology.” All articles are open access articles distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is prop- erly cited. Editorial Board AmiyaK.Banerjee,USA Alain Kohl, UK Jean Rommelaere, Germany Jay C. Brown, USA Young-Min Lee, USA Peter Rottier, The Netherlands Michael Bukrinsky, USA Blanca Lupiani, USA Dominique Schols, Germany Bruno Canard, France Shinji Makino, USA Robert Snoeck, Belgium Jaquelin Dudley, USA Masao Matsuoka, Japan Edward Stephens, USA Eric O. Freed, USA Trudy Morrison, USA H. J. Thiel, Germany Robert C. Gallo, USA A. Osterhaus, The Netherlands Subhash Verma, USA Gary S. Hayward, USA George N. Pavlakis, USA WilliamS.M.Wold,USA William Irving, UK Finn S. Pedersen, Denmark Frank Kirchhoff,Germany Stefan Pohlmann,¨ Germany Contents Virus Immune Evasion: New Mechanism and Implications in Disease Outcome, Rika Draenert, John Frater, and Julia G. Prado Volume 2012, Article ID 490549, 1 page Viral Infection: An Evolving
    [Show full text]
  • 2B4 (CD244) Signaling by Recombinant Antigen-Specific Chimeric
    Published OnlineFirst July 28, 2009; DOI: 10.1158/1078-0432.CCR-08-2810 Cancer Therapy: Preclinical 2B4 (CD244) Signaling by Recombinant Antigen-specific Chimeric Receptors Costimulates Natural Killer Cell Activation to Leukemia and Neuroblastoma Cells Bianca Altvater,1 Silke Landmeier,1 Sibylle Pscherer,1 Jaane Temme,1 Katharina Schweer,1 Sareetha Kailayangiri,1 Dario Campana,3 Heribert Juergens,1 Martin Pule,2 and Claudia Rossig1 Abstract Purpose: Novel natural killer (NK) cell–directed strategies in cancer immunotherapy aim at specifically modulating the balance between NK cell receptor signals toward tu- mor-specific activation. The signaling lymphocyte activation molecule–related receptor 2B4 (CD244) is an important regulator of NK cell activation. We investigated whether 2B4-enhanced activation signals can redirect the cytolytic function of human NK cells to NK cell–resistant and autologous leukemia and tumor targets. Experimental Design: In vitro–stimulated NK cells from healthy donors and pediatric leukemia patients were gene modified with CD19 or GD2-specific chimeric receptors containing either the T-cell receptor ζ or 2B4 endodomain alone or combined. Results: Chimeric 2B4 signaling alone failed to induce interleukin-2 receptor up-regula- tion and cytokine secretion but triggered a specific degranulation response. Integration of the 2B4 endodomain into T-cell receptor ζ chimeric receptors significantly enhanced all aspects of the NK cell activation response to antigen-expressing leukemia or neuro- blastoma cells, including CD25 up-regulation, secretion of IFN-γ and tumor necrosis factor-α, release of cytolytic granules, and growth inhibition, and overcame NK cell re- sistance of autologous leukemia cells while maintaining antigen specificity.
    [Show full text]
  • Impaired Activating Receptor Expression Pattern in Natural Killer Cells from Patients with Multiple Myeloma
    Letters to the Editor 732 specificity for the CD20 molecule for potential use in adoptive by IMF SC 110212, University of Muenster to CS. MP and CS immunotherapy of B-cell malignancies. We show that cTCR þ contributed equally. CD8 þ NKT cells can be expanded in large quantities and are M Pieper1, C Scheffold1, S Duwe2, C Rossig2, G Bisping1, highly effective in lysing CD20 molecule expressing Daudi and 1 3 2 1 1 Raji tumor cell lines, as well as freshly isolated malignant B M Stelljes , TF Tedder , H Jurgens , WE Berdel and J Kienast 1Department of Medicine/Hematology and Oncology, lymphocytes from B-CLL patients. In competitive targeting University of Muenster, Muenster, Germany; studies, we were able to dissect the major pathways of tumor 2 þ Department of Pediatric Hematology and Oncology, cell recognition and found that endowing CD8 NKT cells with University of Muenster, Muenster, Germany and CD20z chimeric receptors resulted in a significant increase of 3Department of Immunology, Duke University, cytotoxic activity against CD20 þ Daudi targets as compared to Durham, NC, USA nontransduced effectors. E-mail: [email protected] We also demonstrate that chimeric receptor mediated cytotoxicity is dependent on the expression level of the target antigen. Tumor targets expressing high levels of CD20 molecule (Daudi) were lysed more efficiently than low level expressing References targets (Raji). Of importance, cytotoxicity of ex vivo expanded cTCR þ CD8 þ NKT cells was more potent as compared to 1 Eshhar Z, Waks T, Gross G, Schindler DG. Specific activation and ex vivo expanded cTCR þ CD8 þ T cells.
    [Show full text]