Humoral Immune Responses PDF

Humoral Immune Responses The activation of B cells results in their proliferation and their eventual differentiation into antibody- secreting plasma cells and memory cells 3 T-dependent and T-in...

Humoral Immune Responses The activation of B cells results in their proliferation and their eventual differentiation into antibody- secreting plasma cells and memory cells 3 T-dependent and T-independent antibody responses 4  Primary and secondary humoral immune responses Primary and secondary antibody responses to protein antigens differ qualitatively and quantitatively 5 Primary and secondary humoral immune responses 6 Pathways of antigen delivery to follicular B cells 7 Activation of B Cells by Antigens and Other Signals Role of complement receptor type 2 and Toll-like receptors in B cell activation. 8  Effects of B cell antigen receptor engagement on B cells The interaction of different types of antigens (multivalent structures or proteins) with the BCR initiates B cell proliferation and differentiation in different ways. 9 Sequence of events in humoral immune responses to T cell- dependent protein antigens. 10 Migration of B cells and helper T cells and T-B interaction Antigen-activated helper T cells and B cells move toward one another in response to chemokine signals and make contact adjacent to the edge of primary follicles 11  Antigen presentation on B cells to helper T cells Protein antigens recognized by membrane Ig are endocytosed and processed, and peptide fragments are presented in association with class II MHC molecules. Helper T cells recognize MHC–peptide complexes on the B cells and then stimulate B cell responses. In responses to hapten-carrier conjugates, the hapten (the B cell epitope) is recognized by a specific B cell, the conjugate is endocytosed, the carrier protein is processed in the B cell, and peptides from the carrier (the T cell epitopes) are presented to the helper T cell. Some haptens can induce autoimmune disease. An example is hydralazine, a blood pressure-lowering drug that can produce drug- induced lupus erythematosus in certain individuals. This also appears to be the mechanism by which the anaesthetic gas halothane can cause a life-threatening hepatitis, as well as the mechanism by which penicillin-class drugs cause autoimmune hemolytic anemia. 12 Role of CD40L:CD40 Interaction in T-Dependent B Cell Activation 13  The germinal center reaction in a lymph node The characteristic events of helper T cell–dependent antibody responses, including: affinity maturation, isotype switching, and generation of long-lived plasma cells and memory B cells, occur primarily in organized structures called germinal centers that are created within lymphoid follicles during T-dependent immune responses. The complex process of genetic diversification of activated B cells and survival of the fittest that occurs in these sites is called the germinal center reaction. 14 Heavy Chain Isotype Switching 15 Affinity Maturation: Somatic Mutation of Ig Genes and Selection of High-Affinity B Cells 16 B cell selection in germinal centers 17 Production of membrane and secreted µ chains in B lymphocytes 18 Antibodies or Immunoglobulins Family of billions of similar but unique proteins produced by B lymphocytes of the immune system. Perform two functions: –Each binds to a specific antigen(molecular target, usually on an invading pathogen); –Once bound to antigens, antibodies trigger a variety of biological effector functions, including: Neutralization (interfering with the target’s function); Targeting other immune cells to dispose of the antibody-tagged pathogen, through: –Inflammation (activating immune cells for non-specific killing); –Phagocytosis, complement activation, or direct cell killing by triggering apoptosis (programmed cell death). 19  Antibodies or Immunoglobulins Antibodies, or immunoglobulins, are a family of structurally related glycoproteins produced in membrane-bound or secreted form by B lymphocytes. Membrane-bound antibodies serve as receptors that mediate the antigen- triggered activation of B cells. Secreted antibodies function as mediators of specific humoral immunity by engaging various effector mechanisms that serve to eliminate the bound antigens. The antigen-binding regions of antibody molecules are highly variable, and any one individual has the potential to produce millions of different antibodies, each with distinct antigen specificity. 20 Structure of an antibody Molecule 21 Hypervariable Region of Immunoglobulins Complementarity-determining regions The N-terminal domains of heavy and light chains form the V regions of antibody molecules, which differ among antibodies of different specificities. The V regions of heavy and light chains each contain three separate hypervariable regions of about 10 amino acids that are spatially assembled to form the antigen- combining site of the antibody molecule. 22  Antigen – Antibody Binding Complementarity-determining regions 23  General features of Ab structure All antibodies have a common symmetric core structure of two identical covalently linked heavy chains and two identical light chains, each linked to one of the heavy chains. Each chain consists of two or more independently folded Ig domains of about 110 amino acids containing conserved sequences and intrachain disulfide bonds. 24 General features of Ab structure Shown here are a membrane-bound IgG molecule; the T cell receptor; a major histocompatibility complex (MHC) class I molecule; the CD4 coreceptor of T cells; CD28, a costimulatory receptor on T cells; and the adhesion molecule ICAM-1. N and C at the ends of polypeptide chains refer to the amino and carboxy termini, respectively. 25 Antibodies Characterization Non-reducing kDa gel 150 kDa 50 25 Reducing gel 26 Features of Immunoglobulin Binding Antigen 27 Human Immunoglobulin Repertoire Antibodies are classified into different isotypes and subtypes on the basis of differences in the heavy chain C regions, which consist of three or four Ig C domains, and these classes and subclasses have different functional properties. The antibody classes are called IgM, IgD, IgG, IgE, and IgA. Both light chains of a single Ig molecule are of the same light chain isotype, either κ or λ, which differ in their single C domains. 28  Human Antibodies Isotypes  Any variable region can use one of five different heavy chain constant regions.  This choice defines five different isotypes, or classes of human antibodies with distinct biological functions: IgG: γ(gamma chain); IgM: µ (mu); IgD: δ (delta); IgA: α (alpha); IgE: ε (epsilon).  B cells can switch the isotypes of antibodies they produce by exchanging heavy chain constant regions, but the switch is irreversible. Most of the effector functions of antibodies are mediated by the C regions of the heavy chains, but these functions are triggered by binding of antigens to the combining site in the V region. 29 Human Antibodies Isotypes: GAMDE 31 Clonal Expansion  Lymphocytes specific for an antigen undergo considerable proliferation after exposure to that antigen.  The term clonal expansion refers to an increase in the number of cells that express identical receptors for the antigen and thus belong to a clone.  The increase in antigen- specific cells enables the adaptive immune response to keep pace with rapidly dividing infectious pathogens. 32 Memory  Exposure of the immune system to a foreign antigen enhances its ability to respond again to that antigen.  Responses to second and subsequent exposures to the same antigen, are usually more rapid, larger, and often qualitatively different from the first, or primary, immune response to that antigen.  Each exposure to an antigen generates long-lived memory cells specific for the antigen. These memory cells are more efficient at responding to and eliminating the antigen than are naïve lymphocytes.  Memory B lymphocytes produce antibodies that bind antigens with higher affinities than antibodies produced in the first immune response.  Memory T cells also react much more rapidly and vigorously to antigen challenge than do naïve T cells 33 Changes in antibody structure during humoral immune response 34  Monoclonal antibodies  Monoclonal antibodies are produced from a single clone of B cells and recognize a single antigenic determinant. Monoclonal antibodies can be generated in the laboratory and are widely used in research, diagnosis, and therapy. 35  Monoclonal antibodies applications: research, medical diagnosis and therapy  Identification of phenotypic markers unique to particular cell types. These specific monoclonal antibodies have been used to define clusters of differentiation (CD) markers for classification of lymphocytes and other leukocytes and various cell types.  Immuno-diagnosis. The diagnosis of many infectious and systemic diseases relies on the detection of particular antigens or antibodies in the blood, urine, or tissues by use of monoclonal antibodies in immunoassays.  Tumor identification. Labeled monoclonal antibodies specific for various cell proteins are used to determine the tissue source of tumors by staining histological tumor sections.  Therapy. Advances in medical research have led to the identification of cells and molecules that are involved in the pathogenesis of many diseases. Monoclonal antibodies, because of their exquisite specificity, provide a means of targeting these cells and molecules. Many monoclonal antibodies are used therapeutically today. Some examples include: Antibodies against the cytokine tumor necrosis factor (TNF) used to treat rheumatoid arthritis and other inflammatory diseases. Antibodies against CD20 for the treatment of B cell–derived tumors and for depleting B cells in certain autoimmune disorders. Antibodies against epidermal growth factor receptors to target cancer cells. antibodies against vascular endothelial growth factor in patients with macular degeneration. 36 Therapeutic Antibodies 37 Therapeutic Antibodies 38 Monoclonal Antibodies in Clinical Use 39  International Nonproprietary Names (INNs) for Monoclonal Antibodies  INN system begun in 1950 by the World Health Organization (WHO) to provide a unique (generic) name to identify each pharmaceutical substance  INN system has important goals and benefits: Clear identification, safe prescription and dispensing of medicines to patients. Communication and exchange of information among health professionals and scientists worldwide. 40 Revised monoclonal antibody (mAb) nomenclature scheme Geneva, 26 May 2017 Programme on International Nonproprietary Names (INN) © World Health Organization 2017 41 Limitations of monoclonal antibodies Not orally bioavailable: delivery by injection usually needed Optimal for extracellular targets; difficult intracellular delivery. Cannot penetrate the blood-brain barrier. Chemistry, Manufacturing and Controls (CMC) Issues: – Complex molecules produced by living cells; hard to characterize and to control batch-to-batch variation and stability Potential for immunogenicity: If recognized as foreign by the immune system, will trigger the formation of anti-drug antibodies (ADA). – A mouse antibody injected into a human will elicit a Human Anti-Mouse Antibody (HAMA) response; – Even human antibodies can trigger antibody responses in humans; HAHA responses. 43 Anti-Drug Antibodies (ADA) in Patients 44 Recombinant Antibodies Re-engineered to reduce immunogenicity in humans Paul Carter Nat. Rev. Cancer, Nov, 2001. 118-129. 45  Recombinant Antibodies: Anti-CD20  Rituximab was developed using cloning and recombinant DNA technology from human and murine (mice or rat) genes. It was originally FDA approved in 1997 for treatment of non-Hodgkin Lymphoma that was resistant to chemotherapy.  Ofatumumab is the first fully humanized monoclonal antibody that targets the CD20 molecule and is approved for the treatment of previously untreated patients with CLL.  Obinutuzumab is a glycoengineered antibody that demonstrated significantly higher efficacy over rituximab in B-cell malignancies such as CLL. The effect of the glycoengineering improves the binding of monoclonal antibodies with immune cells. 46  Second- and third-generation anti-CD20 mAbs  The second-generation anti-CD20 mAbs include ofatumumab, veltuzumab, and ocrelizumab. These are humanized to reduce immunogenicity.  The third-generation humanized CD-20 mAbs have an engineered Fc region to increase their binding affinity for the FcγRIIIa receptor. Three third-generation mAbs, AME-133v, PRO131921 and GA101, are undergoing active clinical development. 48 Shundong Cang, et al., Journal of Hematology & Oncology 2012  49 Shundong Cang, et al., Journal of Hematology & Oncology 2012  Antigens Metabolites DNA RNA Protein ✹ Antigens are substances specifically bound by antibodies or T lymphocyte antigen receptors. Antigens that bind to antibodies include a wide variety of biologic molecules, including sugars, lipids, carbohydrates, proteins, and nucleic acids. This is in contrast to most T cell antigen receptors, which recognize only peptide antigens. 50 Antigenic Determinants Macromolecular antigens contain multiple epitopes, or determinants, each of which may be recognized by an antibody. Linear epitopes of protein antigens consist of a sequence of adjacent amino acids, and conformational determinants are formed by folding of a polypeptide chain. 51 Antigen-Antibody Complexes  The relative concentrations of polyvalent antigens and antibodies may favor the formation of immune complexes that may deposit in tissues and cause damage.  Antibody binding to antigen can be highly specific, distinguishing small differences in chemical structures, but cross-reactions may also occur. 52