mod 1 slide 3 (B-cells & antibodies)

antibodies (Ab) or immunoglobulins (Ig)

glycoproteins that appear in circulation after immunization and they have the ability to react specifically with the antigen used to immunize

antigen

substance — protein from a vaccine component, pathogen, tumor cell, allergen, that causes an antibody response

epitope

the part of the antigen that interacts with antibody — usually a small peptide sequence

hapten

a small molecule that can elicit an immune response only when it reacts with an endogenous self-protein; examples (antibiotic drugs)

monoclonal antibody (mAb)

• identical immunoglobulines, generated from a single B-cell clone

• these antibodies recognize unique epitopes, or binding site, on a single antigen

• produced by recombinant biotechnology protocols and are used as therapies

clonal expansion

• process of rapid cell division resulting in the multiplication of genetically identical cell clones from a single parent cell

  • for example: a single activated B-cells can, within 7 days, give rise to approx. 4000 antibody-secreting plasma cells — over 2000 antibody molecules can be produced per plasma cell per second for typically up to 4-5 days

humoral

relating to the body fluids, especially with regard to immune responses involving antibodies in body fluids as distinct from cells

antibody response — adaptive

• compared to the primary antibody response, the secondary antibody response occurs ___ ___ and produces antibody levels that are ____ and more ____

• primary response involves ____ (first antibody to appear on B-cells)

• secondary response mostly involves ____

• more quickly; higher; sustained

• IgM

• IgG

why are all FDA approved mAbs based on IgG class of antibodies rather than the other 4 classes?

• crosses placenta

• molecular weight not that high, prevalent found in body, HALF LIFE

vaccination: activation of B and T-cells, production of Ig’s and memory cells

• upon SC or IM administration, the vaccine components are taken up by ____ ____ such as macrophages and dendritic cells (DCs) that reside in the peripheral tissue

  • the APCs that have taken up antigens become _____ and start ___ ___ nearby lymph nodes

• inside the lymph nodes, the antigen is process by the ___ and presented to ___ and ___ ___, which, when recognizing the antigen and receiving the appropriate co-stimulatory signals, become activated

• these antigen-specific B and T-cells clonally expand to produce multiple cells recognizing the ___ antigen

  • in addition, memory B and T-cells are formed that provide ___-___ protection against infection with the pathogen

• phagocytic APCs

  • activated; migrating towards

• APCs; T and B-cells

• same

  • long-term (sometimes lifelong)

antibody response: cells involved

• example: human __ cell responses to influenza vaccination (containing non-pathogenic viral protein antigens) in humans

• the B-cell that contain a surface antibody (B cell receptor; naive) matching to the influenza antigen is ___ ___ and becomes activated

• this ultimately leads to production of ___ ___ ___ and ____ ____ that secrete neutralizing antibodies to the initial antigen

• B cell

• clonally selected

• memory B cells and plasma cells

B cell development and differentiation

• B cell development occurs in both the ___ ____ and ____ ____

• in the bone marrow, B cells mature from ___ cells

  • during this differentiation, gene rearrangements of the antibody result in what?

  • BCR is fully functional antibody (___) bound to a B cell that is capable of binding antigen

  • following an interaction with antigen, specific B cells differentiate into ____ (antibody secreting) cells and ____ ___ cells

  • each B cell carries a ____ antibody

• bone marrow and lymphoid organs (spleen and lymph nodes)

• stem

  • the generation and surface expression of a mature B cell receptor (BCR)

  • IgM

  • plasma cells and memory B cells

  • distinct antibody — no 2 B cells are alike!

antibodies are bound to the surface of B cells via their Fc receptor

• antibodies on B cells (called B cell receptors) look a lot like ___ ___ receptors on ___ ___

  • remember: TCR binds ____ antigen that APC carries via ____

• what do both the cells do?

• T cells

  • peptide; MHC

• both bind antigens (usually peptides)

B cells need helper T cells (CD4+)

• ___ ____ are also classified as professional antigen presenting cells (APC) and carry a unique antibody that can bind antigen

• B cells talk to ____ helper T cells via ____ (3 signal model)

• B cell undergoes ____ ___ and ____ in lymphoid tissue to plasma cells and memory B cells

• B cell

• CD4+; MCH II

• clonal expansion and differentiate

antibody structure - general

• ______ (mirror image)

  • 2 Fab (binds antigen)

  • Fc (binds Fc receptors)

• _____

  • 2 identical heavy (red)

  • 2 identical light (blue)

• _____

  • variable (VH and VL)

    • variable (V) regions (VH and VL)

      • no 2 B cells will do what?

  • constant (CH and CL)

• regions

• chains

• domains

  • will secrete antibodies with the same variable region

structure of an antibody (Ig) — specifics

• what does the antibody consist of? (5)

• heavy chain constant Fc region determines what?

  • the Fc portion of the Ig and binds to Fc receptors on ____ cells

  • what do disulfide bonds do?

• antibody repertoire (total # of unique Ig’s available to a human) is ___

• only ______ BP DNA in human genome

1. 2 identical heavy chains (H

2. 2 identical light chains (L)

3. 2 identical antigen binding sites

4. variable regions in both H and L chains

5. constant region in both H and L chains

• Ig isotype (IgG, IgM, IgE, IgA, IgD)

  • immune cells

  • hold constant chains together

• 10¹⁸

• 3 × 10⁹

antigens (peptides) bind to Fab portion of antibody

• each B cell containing a specific antibody on its surface can interact with what?

• what type of epitope specificity is determined by strength of interaction between epitope and Fab portion of antibody?

• are they covalent or non covalent?

  • what binding is associated with it? (3)

• an antigen epitope

• lock and key epitope

• non covalent

  • H bonding

  • van der waals

  • electrostatic

chemical manipulation of antibodies

• treatment with the enzyme ____: produces 1 F(ab’)₂ piece (the Fc portion is hydrolyzed [degrade] into tiny peptides)

• treatment with the enzyme ____: produces 2 x Fab pieces and an Fc piece

• pepsin

• papain

antigen-antibody response

• ______: reaction of soluble antigen with soluble Ig

• ______: reaction of antigen expressed on cell-surface with Ig (visible clumps)

• ______: reaction of Ig with viruses and soluble toxins released by bacteria (result in loss of infectivity)

• ______: coating of pathogens and foreign particles, prior to recognition of phagocytic cells

• ______ ______: the complement is composed of approx. 20 serum proteins (24-550 kD), which promote ingestion of microorganisms by phagocytic cells (phagocytosis) or by coating bacteria (opsonization)

• precipitation

• agglutination

• neutralization

• opsonization

• complement activation

how do other Ig isotypes arise, if IgM is the only Ig isotype bound on developing B cells?

• class (isotype) ___

• remember; IgM is the ____ isotype found on _____ B cells

  • antigen matching with BCR provides clonal expansion

• but switching from an IgM secreting plasma cell to an IgE or IgG secreting plasma cells requires ____

• dependent upon antigen ____ AND ____ ___ ___ ___

• involves ____ ____ ____ ____ at DNA/RA level

• switching

• major; naïve

• help

• stimulation and interaction with T cells

• heavy chain constant rearrangement (Fc)

class switching example

• to produce IgG antibodies, the variable regions must be ____ and the constant regions _____

• in order to change the function of the molecule but keep the specificity, one needs to ____ the light chain and the heavy chain variable regions and ____ the heavy chain constant region

• preserved; exchanged

• conserve; replace

therapeutic antibodies - monoclonal antibodies

• since we know antibodies are specific to an epitope of an antigen; can we take advantage in a therapeutic setting?

• create ___ ___ of an antibody specific to an antigen — perhaps a self or non-self protein antigen that is implicated in a disease

• monoclonal antibodies (mAbs)

• single clones

what does the mAb STELARA do?

• ____ 2 inflammatory cytokines IL12 and IL23

• it uses Fab portion to bind to a ____ ___ on IL12 and IL23

• _____ these cytokines from acting on their cell receptors - blocks signaling

• work _____ of cell

• blocks

• shared epitope

• prevents

• outside

I just discovered a new humanized Mab that works on a bone osteoclast protein target.  In keeping with the rules, I should name it:

1. Rimtumumab

2. Rimsozumab

3. Rimvizumab

4. Rimciximab

2. Rimsozumab

Which of the following biologics is classified as an immunomodulating human Mab according to nomenclature rules?

1. Certolizumab  (CIMZIA)

2. Golimumab (SIMPONI)

3. Vedolizumab (ENTYVIO)

4. Secukinumab (COSENTYX)

2. Golimumab (SIMPONI)

nomenclature can be confusing with new approved mAbs

• break it down

  • substem A: ne (neural)

  • substem B: zu (humanized)

• what is the 4 letter nonsensical extension (-jjmr)

  • 4 letter random extension is mandated by the FDA for new _____ ____ including originator and biosimilar biologics

  • these suffixes are intended to make it easier for health care providers to distinguish between biologic medications made by different manufacturers

• biologic medications

Alhemo (concizumab-mtci)

for routine prophylaxis of bleeding in hemophilia A and B

Bizengri (zenocutuzumab-zbco)

to treat non-small cell lung cancer and pancreatic adenocarcinoma

Ebglyss (lebrikizumab-lbkz)

to treat moderate to severe atopic dermatitis

Kisunla (donanemab-azbt)

to treat Alzheimer’s disease

germinal center

a specialized structure within secondary lymphoid organs in which responding B cells undergo somatic hypermutation and selection for increased antigen affinity (affinity maturation)

somatic hypermutation

• process that allows B cells to mutate the genes that they use to produce antibodies

• this enables the B cells to produce antibodies that are better able to bind to antigens

• these mutations occur in the Fab portion of the anibody

affinity maturation

process by which antibodies gain increased affinity and anti pathogen activity and is the result of somatic hypermutation (SHM) of immunoglobulin genes in B cells, couples to selection for antigen binding

class switch recombination

process by which proliferating B cells rearrange the constant region genes (Fc) in the heavy chain to switch from expressing one class of immunoglobulin (such as IgM) to another (such as IgG)

affinity maturation

• drives B cells to evolve toward expression of ____ affinity B cell antigen receptor (BCRs)

• GC-B cells that produce high affinity antibodies via ____ ____ are selected to differentiate into plasma cells and memory B cells through specific gene expression changes that regulate proliferation and differentiation

• higher

• affinity maturation

how and where does this (affinity maturation) occur?

• the ___ ___ of lymphoid organs are specialized compartments within peripheral lymphoid tissues (eg. lymph nodes, spleen) and the main structure where antigen activated B cells diversify by _____ _____ to generate high affinity antibodies

• the GC is divided into 2 anatomical compartments

  • ___ ____ site of antigen driven selection and affinity maturation

  • ___ ____ site of B cell proliferation and somatic hypermutation (SHM)

• germinal center (GC); somatic hypermutation (SHM)

• light zone (LZ)

• dark zone (DZ)

germinal center dynamics

• the processes leading to the formation of high affinity B cells that target a specific antigen and the formation of antibody secreting plasma cells and memory B cells is based on their relative ____ — much akin to Darwinian selection — in an individual

• ___ ____ helper cells play a huge role in affinity maturation

• the intensity of T cell help received in the LZ determines the number of ___ ___ a B ell will undergo in the DZ

• a B cell that receives the _____ amount of help necessary for survival would undergo only one cell cycle per LZ/DZ cycle, whereas a B cell that receives a _____ T cell signal can undergo multiple cell cycles in the DZ without returning to the LZ

• fitness

• T-follicular

• division cycles

• minimum; strong