Immunology Block 3

T cell Deletion (periphery): Activation induced cell death

(AICD) • Repeated stimulation of T cells with self antigen induces co-expression of Fas and FasL (Both ligand and receptor) it will self induce cell death or (in the case of APC presenting self antigens) two lymphocytes in close proximity can meet and bind to each other's Fas and FasL (self recognition of self antigen and two FasL-Fas binding) and they kill each other. Self antigens recognized wihtout strong costimulation do not stimulate production of anti • Binding of FasL to Fas induces the apoptosis of Fas-bearing cells by activation of the caspase signaling cascade leading to cleavage of nuclear DNA and proteolysis of vital cellular proteins. • FasL is expressed predominantly on T cells activated by the interaction between T cell receptor (TCR) and antigen-presenting cells (APCs). • Microbe-infected cells are recognized by FasL expressing cytotoxic T lymphocytes (CTLs) Such cells are killed by CTLs through Fas-FasL-mediated apoptosis Mice with mutation in Fas and FasL genes and children with mutations in these all develop autoimmune diseases with lymphocyte accumulation.

Principle factors in development of autoimmunity

*- inheritance of susceptibility genes* - susceptibility genes may interfere with pathways of self-tolerance & lead to the persistence of selfreactive T and B lymphocyte *- environmental triggers such as infections* - environmental stimuli may cause cell and tissue injury and inflammation and activate these selfreactive lymphocytes - results in the generation of effector T cells and autoantibodies that are responsible for the autoimmune disease - promotes the influx of lymphocytes into tissues & activation of antigen-presenting cells (APCs) and subsequently of self-reactive T cells, resulting in tissue injury

Mature B lumphocytes that enctounter self-antigens in the peripheral lymphoid tissues can become incapable of responding to that antigen by:

*1. B cells recognize self antigen & don't receive T cell help (helper T cells have been eliminated or are tolerant) & B cells become anergic because of this block in signaling* • anergic B cells may leave lymphoid follicles and are subsequently excluded from the follicles • these excluded B cells may die because they do not receive necessary survival stimuli (anti-apoptosis stimuli) *2. B cells that recognize self antigens in the periphery may also undergo apoptosis* *3. Inhibitory receptors on the B cells may be engaged and prevent activation & Regulatory T cells may also contribute to peripheral B cell tolerance*

Active vs Passive immunotherapy

*Active: * Engages immune system Durable *Passive* enhances pre-existing immune response short lived.

Two best defined mechanisms responsible for induction of anergy

1) Loss of TCR mediated activation signalling due to loss of costimulation 2) Increased delivery of inhibitory signals.

Passive immunotherapy

1. Antibody therapy 2. Adoptive cell therapy 3. Chimeric antigen receptor engineered T cell therapy (CAR-T)

The Role of Innate and Adaptive Immunity in Promoting Tumor Growth

1. Chronic inflammation, infection (e.g., gastric cancer—Helicobacter pylori; hepatocellular carcinoma—HBV, HCV) 2. Cells of the innate immune system are considered the most direct tumorpromoting culprits among immune cells. Tumor-associated macrophages of the alternatively activated (M2) phenotype as well as other cells are sources of VEGF and TGFβ, which promotes angiogenesis, and matrix metalloproteinases. 3. The adaptive immune system can promote chronic activation of innate immune cells, thereby promoting cancer progression.

Tumor evasion mechanisms

1. Lack of T cell recognition of tumor (loses the antigen that CTLs are activated for) aka failure to produce tumor antigen 2. Lack of T cell recognition of tumor (class I MHC deficient tumor) 3. Inhibition of T cell 4. Tumor treated as self antigen (Tolerance) 5. Antibody modulation - antibodies against tumor antigen is endocytosed and lost 6. Tumor makes a matrix around itself so CTLs can't get to it

Peripheral T cell tolerance: Anergy

1. T Cell-intrinsic anergy: Refers to long lived functional unresponsibeness that is induced when these cells recognize self antigens. self antigens are usually displayed with low levels of (APC) costimulators (B7) T cell-engagement of self antigens may cause preferential T cell binding of inhibitory receptors to B7 that is expressed on APCs anergic cells may express increased inhibitory receptors CTLA-4, PD-1 (NOT CD28) lack of optimal TCR complex signaling leads to increased expression of ubiquitin ligase enzymes eventual proteasome activation (and removal of that T cell). Anergic cells survive but are incapable of responding to antigen so they die.

Another way of describing T cell death in periphery

1. Uptake & processing of self antigen 2. Presentation of processed peptide to CD4+ T cell 3. IL-2 production & expression of the IL-2R occurs followed by their autocrine binding 4-6. Activated autoreactive T cells express Fas and FasL on their surfaces or as soluble s-FasL after cleavage of the membrane-associated FasL 7-9. FasL-Fas R interactions lead to apoptosis by activationinduced cell death

CTL response induction to tumor cells.

APCs ingest tumor cells as antigens and present antigens to T cells. Often the tumor cells don't express costimulators or class II MHC. CTLs require recognition of antigen or costimulation from class I restricted CD4 cells (tumor cells stop both). Dendritic cells therefore ingest tumor cells or their proteins and antigens and processess and display on class I MHC via cross-presentation (aka cross priming). These dendritic cells will express the costimulator that CD8 T cells need to become CTLs. It is not known how they do this because bacteria induce the costimulators (possible dead cancer cells induce it).

Tumor therapy: Vaccination

Active: One way is to vaccinate patients with their own tumor cells or antigens from these cells. Vaccines can be administed as recombinant proteins with adjuvants. Another way of doing is is dendritic cells expanded in vitro from blood precursors and exposed to tumor cells or defined tumor antigen and these tumor antigen pulsed dendritic cells are used as vaccines. It is hoped that these APCs with tumor antigens will mimic the normal pathway of cross presentation and generate CTLs against the tumor cells. Tumor vaccines are only moderately successful. (maybe tumors developed mechanisms to suppress immune system) Also, if you are sick, you will have weakened immune system. Might be better for early stage when you have healthy immune system. Tumors caused by oncogenic viruses can be prevented by vaccinating against these viruses. (Hep-B and HPV)

Two models for how negative selection vs Treg generation are done

Affinity model T-cell tolerance mechanism (checkpoints) (key word "affinity") Differential signaling hypothesis

Syngeneic - Allogenic - Xenogenic -

Animals that are identical to one another (and the grafts exchanged among these animals) Animals (and grafts) of one species that differ from other animals of the same species. animals (and grafts) of different species

Some MHC alleles lead to increased risk of

Ankylosing spondylitis RA type 1 DM Pemphiguys vulgaris

T cell deletion (Periphery): normal immune response

Antigen recognition induces production of pro-apoptotic proteins in T cells that induce cell death. In normal immune responses, the activity of these pro-apoptotic proteins is counteracted by anti-apoptotic proteins that are induced by costimulation and by growth factors produced during the responses. However, self antigens, recognized without strong costimulation, do not stimulate production of anti-apoptotic proteins and the relative deficiency of survival signals induces death of the cells that recognize these antigens.

T cell deletion - Activation induced cell death (periphery)

Apoptosis of Mature Lymphocytes: Recognition of self antigens may trigger pathways of apoptosis that result in elimination (deletion) of the self-reactive lymphocytes. Two likely mechanisms: FasL--Fas Binding. Activation-induced cell death (AICD). Normal immune response

Peripheral tolerance

Autoreactive mature B and T cells leave central lymphoid organs and enter peripheral lymphoid tissue where they are exposed to self antigens where: a) B and T cells can undergo functional inactivation (anergy) b) B and T cells can undergo apoptosis (Negative selection) c) T cell activation can be blocked by regulatory T cells

Receptor editing

B cell central tolerance: changing receptor specificity ▪ Some immature B cells that recognize self antigens in the bone marrow may: re-express RAG genes, resume immunoglobulin (Ig) light-chain gene recombination, or express a new Ig light chain ▪ new light chain associates with the previously expressed Ig heavy chain ▪ new antigen receptor may no longer be specific for self antigen ▪ reduces the chance that potentially harmful self-reactive B cells will leave the marrow ▪ estimated that 25% to 50% of mature B cells in a normal individual may have undergone receptor editing during their maturation

Central tolerance in general:

BM and Thymus immature B and T cells exposed to self antigens can undergo any of the following mechanisms a) B&T celsl may undergo apoptosis and are deleted (negative selection) b) B cells can change their receptor specificity (Receptor editing) c) Some T cells can develop into regulatory T cells Tolerance of self antigens not present in these organs can be induced and maintained in peripheral organs

Tumor therapy: Checkpoint blockade

Boost host immune response against tumors by blocking normal inhibitory signals for lymphocytes thus removing brakes (checkpoints) on the immune response. An antibody against CTLA-4 was approved for melanoma. Antibodies for PD-1 PD-L1 have sown impressive efficacy. This targets the PD ligand ON THE TUMOR. This immune response is largely specific for peptides produced by mutated genes in the tumors. Sometimes this leads to autoimmunity though.

DM type 1 RA

CLA-4 allele CD25 allele MHC allele Type 4 CTLA4 Type 4

How does T cell choose which receptor will bind APC-ligand and B7?

CTLA-4 has a higher affinity for B7 molecules than does CD28 CTLA-4 may have higher expression levels in T cells specific for autoantigens When APCs present self antigens B7 levels are low, T cell CTLA-4 receptor will preferentially bind B7 When APCs present microbial antigens during infection B7 levels are high, the low-affinity activating receptor CD28 is engaged to B7 ligands to a greater extent than CTLA-4 (Remember: b/c microbial antigen CTLA-4 expression levels are presumably lower)

Autoreactive lymphocyte

Capable of recognizing self self antigens. When the mechanisms fail, the immune system attacks the individuals own cells and tissues which is known as autoimmunity. Autoimmune disease result from autoimmunity.

Tumor antigens: Viral antigens

Caused by oncogenic viruses, the tumor antigens may be products of the viruses. (HPV E6, E7 proteins in Cervical carinoma, EBNA proteins in EBV induced lymphomas)

Differential signaling hypothesis

Central T cell tolerance mechanism ® Qualitatively and quantitatively different signals are delivered during positive and negative selection ® Nature of the signalling downstream of the TCR differs greatly between positive and negative selection due to difference in strenght of signal received. ® Strong binding is associated with ◊ robust calcium influx ◊ migratory arrest ◊ lag period (gene changes) ◊ programmed cell death & phagocytosis Treg (FoxP3+/CD4+/CD25+) cells that are autoantigen-specific are generated by intermediate degrees of binding and subsequent "intermediate strength" of signal received by developing CD4 T cells express FoxP3 become Tregs!

Two types of antigens that are not self but are produced by cells or tissues that have to be tolerated

Commensal microbes in intestine/skin Tolerance of fetal antigens

Tumor antigen: Products of diverse mutated genes

Common human tumors harbour a large numcer of mutations in diverse genes that play no role (passenger mutations). Many of these may stimulate adaptive immune response. Virutally any gene may be mutagenized randomly in different tumors

Immune surveillance

Control and elimination of malignant cells by immune system

T cell central tolerance principle mechanisms

Death of immature T cell (negative selection Generation of CD4+ regulatory T cells (Treg ( (FoxP3+/CD4+/CD25+))

Types of allorecognition

Direct and indirect

Infections in autoimmunity

Environmental stimuli; infection: *Normal Tolerance *- Infections may activate self-reactive lymphocytes & trigger the development of autoimmune diseases *Microbe-activated APCs * ❖Microbes may activate the APCs to express costimulators ❖if then these APCs present self antigens, the specific T cells are activated and not rendered tolerant ❖autoimmunity *Molecular mimicry* ❖ Some microbes may produce peptide antigens that are similar to, & cross-react with, self antigens ❖ Immune responses to these microbial peptides may result in an immune attack against self antigens ❖immune responses initiated by the microbes may become directed at self cells and tissues lead to autoimmunity ❖Best example: rheumatic fever Streptococci attempt to evade eradication by expressing host self antigens M antigen (M protein): molecular mimicry Host immune response - production of antibodies Unfortunately same antibodies react against cardiac myosin.. Autoimmunity! Actually autoreactive to heart and joints

First set and second set rejection

First one takes longer Second set has cells ready to reject tumor so it occurs much faster. (also occurs if you give effector cells to different mouse)

Mutations in Tregs

FoxP3 + regulatory T cells are required for the maintenance of self-tolerance (mutations result in systemic multi-organ autoimmune diseases) Examples of diseases: IPEX (Immune dysregulation Polyendocrinopathy Enteropathy Xlinked) syndrome: Rare inflammatory disease caused by mutations of Foxp3, destroys the immunoregulatory environment of affected male infants Presentation is most commonly the clinical triad of watery diarrhea, eczematous dermatitis, and endocrinopathy (most commonly insulindependent diabetes mellitus) Systemic, multi-organ autoimmune disease

Indirect allorecognition

If graft cells (or alloantigens) are injested by recipient dendritic cells, donor antigens are processed and prsented by the self MHC molecules on recipient APCs. It is similar to cross-presentation. Alloreactive CTLs are induced by indirect pathway, these CTLs are specific for donor alloantigens displayed by the recipient's self MHC molecules on the recipient's APCs, so they cannot recognize and kill cellls in the grapft (which, of course, express donor MHC molecules). When graft alloantigens are recognied y the indirect pathway, the subsequent rejection of graft is likely mediated mainly by alloreactive CD4+ T cells. These T cells may enter the graft together with with host APCs, recognize graft antigens that are picked up and displayed by these APCs, and secrete cytokines that injure the graft by inflammatory reaction.

T cell central tolerance: negative selection

If lymphocyte that has not completed its maturation (immature T cell) interacts strongly with a self antigen displayed as a peptide bound to a self MHC (both CD8 and CD4 via MHC1 and MHC II respectively), that lymphocyte receives signals that trigger apoptosis (causing negative selection). This way, cells die before they are competent.

Tumor immunoediting

Immune responses to tumor cells impart selective pressures that result in the survival and outgrowth of variant tumor cells with reduced immunogenicity Could induce a different tumor. This is why if you get cancer again, this is not good. This is a problem with immunotherapy

Tumor antigens: Aberrantly expressed proteins

In several human tumors, antigens that elicit immune responses appear to be normal (unmnated) proteins whose expression is dysregulated in the tumors. They are completely normal but for some reason, abnormal expression may make it immunogenic. One example is a protein that is expressed only in embryos. Shouldn't be expressed in adults. (Tyrosinase, gp100, cancer/testis antigens in various tumors.

Hypersensitivity

Injurous or pathologic immune reaction - Dysregulated /uncontrolled Response to foreign antigen resulting in tissue injury - Immune Responses that may be directed against self (autologous) antigens, as result of the faulty recognition or breakdown of tolerance.

Chronic rejection

Is an indolent form of graft damage that occurs over months or years leading to progressive loss of graft function. It may be manifested as fibrosis of the graft and gradual narrowing of graft blood vessels (called arteriosclerosis). In both lesions, the culprites are believed to be T cells that react against graft alloantigens and secrete cytokines which stimulate the proliferation and activities of fibroblasts and vascular smooth muscle cells in the graft. Alloantibodies also contribute to chronic rejection. Although treatments to prevent or curtail acute rejection have steadily improved, leading to better 1-year survival transplants, chronic rejection is refractory to most of these therapies and is becoming the principle cause of graft failure. Chronic inflammatory reaction in vessel wall intimal smooth muscle proliveration, vessel occulsion Fibrosis - graft arteriosclerosis

Normal T cell response in peripheral lymphoid organ

It requires antigen recognition and costimulation. Those that are not stimulated (and are not memory cells) will die. Dendritic cells in normal uninfected tissues and peripheral lymphoid organs are in a resting state and express little B7 and are constantly processing and displaying self antigens. If an autoreactive T cell came along and TCR binds to self antigen (MCH==(self antigen) }==(autoreactive TCR complex), that is signal one. But it doesn't not receive strong costimulation so it is not activated. The presence or absence of costimulation is therefore a major determining factor if T cells are activated or tolerized. Antigen recognition without adequate costimulation results in T cell anergy or death, or makes T cells sensitive to suppression by regulatory T cells.

Principle immune mechanism of tumor eradication

Killing of tumor cells by CTLs specific for tumor antigens Majority of tumor antigens that elicit response are endogeniously synthesized cytosolic or nuclear proteins that are displayed as class I MHC associated proteins. Therefore recognized by CD8+ CTLs. They kill the cells producing these antigens.

Tumor antigens: Common antigens

Malignant tumors express various types of molecules that may be recognized by the immune system as foreign antigens. The tumor must express antigens if the immune system is to react against the tumor that is seen as "non-self" Products of diverse mutated genes. (mutated self protein that does not contribute to tumorgenesis) Products of ongogenes or mutated tumor supressor genes Overexpressed or Aberrantly expressed proteins Viral antigens

Treg Features

Most regulatory T cells are CD4+ and express high levels of CD25, the α chain of the interleukin-2 (IL-2) receptor (allows optimal recognition), and transcription factor FoxP3, which is required for the development and function of Tregs (without this, they will not become Tregs)

Recognition of allogenic MHC complex

Normally, Self MHC molecule presents foreign peptide to T cell selected to recognize self MHC weakly, but may recognize self MHC+foreign peptide complexes well. Allogenic MHC: The self MHC-restricted T cell recognizes the allogenic MHC moelcule whose structure resembples a self MHC+foreign complex. So it doesn't matter what peptides is in it because the MHC itself fits in the TCR (there is no mechanism for selectively eliminating T cells with TCRs have high affinity for allogenic MHC molecules because they aren't present in the thymus. ) Or The allogenic MHC with a self-peptide or allopeptide together will set off the TCR. (This MHC needs a little help).

Acute rejection

Occurs within days or weeks after transplantation and is the principle cause of early graft failure. It is mediated by T cells and *antibodies specific for alloantigens* in the graft. The T cells may be CD8+ CTLs that directly destroy graft cells or CD4+ cells that secrete cytokines and induce inflammation which destroys the graft. T cells may also react against cells in graft vessels, leading to vascular damage. Antibodies contribute especially to the vascular component of acute rejection. Antibody-mediated injury to graft vessels is caused mainly by complement activation by the clinical pathway. Current immunosuppressive therapy is designed mainly to prevent and reduce acute rejection by blocking the activated of alloreactive T cells. Endothelialitis or Parenchymal cell damage, interstitial inflammation

Hyperacute rejection

Occurs within minutes of transplantation and is characterized by thrombosis of graft vessels and ischemic necrosis of the graft. It is mediated by circulating antibodies that are specific for antigens on graft endothelial cells and that are present before transplantation. Almost immediately after transplantation, antibodies bind to antigens on graft vascular endothelium and thrombus formation. These pre-formed antibodies may be natural IgM antibodies specific for blood group antigens or specific for allogenic MHC molecules (which is IgG) that are induced by exposure to allogenic cells due to previous blood transfucions, pregnancy, or organ transplantation. This is not a common problem in clinical transplantation because we know that donor and recipients are blood matches before and the recipients are tested for antibodies against the cells of the prospective donor (this test is known as cross-match). This is still a problem in xenotransplantation though.

Activated CTL reaction to tumor antigen

Once naïve CD8 T cellls have differentiated into effector CTLs, they are able to kill tumor cells expressing the relevant antigens without a requirement for costimulation or T cell help. Thus, CTL differentiation may be induced by cross presentation of tumor antigens by host dendritic cells but CTLs are effective against the tumor itself. Antitumor CD4+ T cell responses and antibodies are also detected in patients but whether they actually protect individuals against tumor growth is not established. Macrophages and NK cells can kill tumor cells in vitro but protective role of these are unkonwn as well.

Autoimmune disease may be specific to what

Organ (affecting one or a few organs) Systemic (widespread) - tissue injury in autoimmune diseases may be caused by antibodies against self antigens or by T cells reactive with self antigens - cautionary note: in many cases, diseases associated with uncontrolled immune responses are called autoimmune without formal evidence that the responses are directed against self antigens

Tumor antibody therapy

Passive Monoclonal antibodies against various tumor antigens have been used in many cancers. The antibodies bind to tumor antigens and activate host effector mechansims such as phagocytes or complement system. Antibody for CD20 (expressed in B cells) is used for B cell tumors. Other monoclonal antibodies may be used by blocking growth factor signaling (anti-Her2/Neu for brest cancer and anti-EGF receptor for various) or inhibiting angiogenesis (antibody against vascular endothelial growth factor for colon cancer and other tumors)

Tumor therapy: Chimeric antigen receptors

Passive CAR-T - improvement on adoptive cellular therapy Modification of T cell therapy, a chimeric antigen receptor that recognizes a tumor antigen and coupled to intracellular signalling domains is genetically introducted to a patient's T cells and then expanded ex vivo and transferred back into patient. This is effective against leukemias. Can change T cell receptor a bit. Make it more effective. If you know tumor has specific antigen on the surface. You can change the T cell receptor. You are engineering TCR. The new one will be more potent. This is because the T cell generate a recombined engineering protein. It looks like antibody (VH+VL) so it is like high affinity for tumor antigen. Then, on cytoplasmic portion, cell signalling molecule can be used that is stronger than usual and the response overall is much stronger. Expanding quantitiy + quality

Tumor: Adoptive cellular therapy

Passive T lymphs may be isolated from the blood or tumor infiltrates of a patient, expanded by culture with growth factors and injected back into the same patient. The T cells presumably contain tumor-specific CTLs which find the tumor and destroy it. Just expanding quanitity

Peripheral T lymphocyte Tolerance

Peripheral T lymphocyte tolerance is Important for preventing T cell responses to self antigens that are not present in thymus. It may also provide backup mechanisms for preventing autoimmunity in situations where central tolerance is incomplete (autoreactive central tolerance escapees)

HLA linked genes

Relative risk: incidence is higher than general population. ❖ incidence of a particular autoimmune disease often is greater among individuals who inherit a particular HLA allele(s) than in the general population. ❖ increased incidence is called the odds ratio or relative risk of an HLAdisease association ❖ although an HLA allele may increase the risk of developing a particular autoimmune disease, the HLA allele is not, by itself, the cause of the disease ❖ the disease never develops in the vast majority of people who inherit an HLA allele that confers increased risk of the disease

Self antigens differ from foreign microbial antigens by:

Self antigens are present in thymus, Inducing deletion and generate regulatory T cells, whereas microbial antigens are generally excluded from thymus Self antigens are displayed by resting APCs in absence of innate immunity and second signals. Thus favoring induction of T cell anergy or death or suppression by regulatory T cells. Microbes elicit innate immune reactions leading to expression of costimulators and cytokiens that promote T cell proliferation and differentiation into effector cells. Self antigens are present throughout life and may therefore cause prolonged repeated TCR engagement again promoting anergy apoptosis and development of regulatory T cells.

How do HLA linked alleles confer susceptibility?

Some believe: ❖particular MHC alleles may be especially effective at presenting pathogenic self peptides to autoreactive T cells ❖the alleles are inefficient at displaying certain self antigens in the thymus, leading to defective negative selection of T cells

T cell central tolerance: Generation of Tregs

Some immature autoreactive CD4 T cells in thymus with high affinity develop into regulatory T cells instead of negative selection and enter Peripheral tissues.

Non HLA associated genes

Some polymorphisms in non-HLA genes are associated with different autoimmune diseases & may contribute to failure of self-tolerance or abnormal activation of lymphocytes Polymorphisms in the gene encoding tyrosine phosphatase PTPN22 may lead to uncrontrolled activation of both B and T cells and are assoicated with numerous autoimmune diseases Variants of innate immune cytoplasmic microbial sensor NOD-2 can cause reduced resistance to intestinal microbes and this is associated with Crohns's disease (inflammatory bowel disease) CTLA4 - Type 1 DM and RA CD25 - MS, Type 1 DM C2, C4 (complement) - Systemic lypus erythematosus

Mendelian inheritance

Some rare autoimmune disorders are Mendelian in origin, caused by mutations in single genes that have high penetrance and lead to autoimmunity in most or all individuals who inherit these mutations AIRE CTLA4 FOXP3 FAS

Tumor antigens: Products of oncogenes or mutated tumor suppressor genes

Some tumor antigens are products of mutated or translocated oncogenes or tumor supressor genes that presumably are involved in the process of malignant transformation called driver mutations. These mutations may be seen as forein (as proteins). (Oncogenic products: Mutated Ras, Bcr/Abl fusion proteins; Tumor supressor gene products: Mutated p53)

T cell repressing tumor cells

Some tumors express ligands for T cell inhibitory receptors such as PD-1. Tumors may also induce only low levels of B7 costumulators on APCs resuling in preferential engagement of inhibitory receptor CTLA-4 on T cells rather than the stimulatory CD28. Resulting in reduced T cell activation upon recognition of tumor antigens. Some tumors may induce Tregs supressing antitumor immune response. Making Tregs suppress the CTLs Some tumors secrete immunosuppressive cytokines such as TGF-Beta or induce regulatory T cells that suppress immune responses.

Affinity model

T cell central tolerance mechanism The key factor in determining positive and negative selection is the *strength of the antigen recognition by the maturing T cell * ® high-avidity recognition induces negative selection - apoptosis ® low-avidity recognition leads to positive selection - anergy ® Treg (CD4+CD25+) cells that are autoantigen-specific are generated by intermediate degrees of binding *this is the one in the middle.*

Transplant antigens

The antigens of allograpfts that serve as the principle targets of rejection are proteins encoded in the MHC. Homologous MHC genes and molecules are present in all mammals; human MHC is called human Leukocyte antigen (HLA) complex. There are six class I MHC alleles (HLA-A, -B, and -C from each parent) Usually more than eight class I MHC alles (one allele of HLA-DQ and -DP and one or two of DR from each parent and some combinations of these). MHCs are highly polymorphic, over 13,000 HLA alleles among humans The response to MHC antigens on another individuals' cells is one of the strongest immune responses known They are the main antigens that cause graft rejection Other's that contribute are *minor* histocompatibility antigens

Cancer immunotherapy

The main strategies for cancer immunotherapy aim to provide antitumor effectors (antibodies and T cells) to patients, actively immunize patients against their tumors and stimulate the patients' own antitumore immune responses. Most treatment for tumors that can't be removed surgically rely on chemotherapy and irradiation. Immunotherapy remains a major goal of immunologists.

Preventing rejection

The mainstay of preventing and treating the rejection of organ transplants is immunosuppression, designed mainly to inhibit T cell activation and effector functions. The development of immunosuppresive drugs made it feasible to transplant organs from donors that were not HLA-matched with recipients especially in cases of heart lung and liver. Side effects: infection and cancers Long-term goal of transplant immunology: ↑ immunological tolerance specific for the graft alloantigens; ↓ side effects on host adaptive immunity

Autoimmune regulator (AIRE)

There are self-proteins normally present only in the thymus or peripheral tissues. The autoimmune regulator (AIRE) is responsible for the thymic expression of those antigens. Mutations in AIRE leads to autoimmune problems.

Tumor cells that stop expressing class I MHC

They cannot display antigens for CD8+ T cells. However, NK cells recognize molecules expressed on tumor cells but not normal cells and can be effective against class I MHC negative tumors

Tumor therapy: Cytokine therapy

Treating patients with cyokines that promote lymphocyte activation. The first cytokin used this way is IL-2. Its clinicall use is limited by serious toxic effects at high doses that are needed to stimulate antitumor T cell responses. IL-2 also enhances the numbers and function sof Tregs which may interfere with antitumor immunity.

Therapeutic applications of CTLA-4 and PD-1

Treatment of cancer patients with antibodies that block these receptors. This can lead to enhanced antitumor immune responses. This treatment therapy is known as a checkpoint blockated, because the inhibitory receptors impose checkpoints in immune responses and the treatment blocks these checkpoints. Patients treated with this have some degree of autoimmunity, which is understantible because the inhibitory receptors are constatntly functioning to keep autoreactive T cells in check. Animals with CTLA-4 or PD-1 blocked or KO, are autoimune to their own tissues

Checkpoint blockade therapy

Treatment of cancer with antibodies that block receptors: • the inhibitory receptors impose checkpoints to prevent autoimmunity (CTLA4, PD-1 tolerance mechanisms, impose checkpoints.) • Tolerance mechanisms = tolerance checkpoints • the cancer chemotherapeutic treatment blocks these checkpoints ("removes the brakes" on immune responses) • patients treated with checkpoint blockade often develop autoimmune reactions consistent with the idea that the inhibitory receptors are constantly functioning to keep autoreactive T cells in check Immunosupressive Cancer turned Cancer suppressive-T cells off Checkpoint blockade turned (results in)Cancer suppressive -T cells back on! • In experimental animals: if CTLA-4 or PD-1 molecules are blocked (by treatment with antibodies) or eliminated (by gene knockout), the animals develop autoimmune reactions against their own tissues • Polymorphisms in the CTLA4 gene have been associated with some autoimmune diseases in humans • Group 7: Rare patients with mutations in one of their two copies of the CTLA4 gene also develop multi-organ inflammation

Three antibody approaches for killing tumors (Target therapy)

Tumor specific antibody - antibody binds to tumor and NK cells with Fc receptors (CD16) are activated and kill tumor Tumor specific antibody or antibody fragment conjugated to toxin - Antibody-toxin conjugates bind to the tumor cell, internalized and kills the cell Tumor specific antibody (or antibody fragment) conjugated to radionucleotide - radioactive antigody bind to the tumor cell and radiation kills tumor and nearby cells.

Immunological tolerance

Unresponsiveness to self antigens. *Lack of immune response to antigens* It is maintained despite the fact that the moleculear mechanisms by which lymphocyte receptor specificities are generated are not biased to exclude receptors for self antigens.

CD4 cells in tumor rejection

We mainly only talk about CD8+ recognition of tumor antigens in these lectures. Tumor antigens also may be recognized by CD4 + T cells, but less is known about the role that CD4 + T cells play in tumor immunity

Peripheral T cell tolerance: Loss of TCR mediated activation

When T cells recognize antigens without cosimulation, the TCR complex may lose its ability to transmit activating signals. In other cases, this is related to the activation of enzymes (ubiquitin ligases) which modify signaling proteins and target them for intracellular destruction by proteases.

Immunogenic

When antigens elicit a response that activates lymphocytes to become effector and memory cells which is a productive immune response, the response is said to be immunogenic.

Central B cell tolerance

When immature B lymphocytes interact strongly with self antigens in BM, the B cells either change their receptor specificity (receptor editing) or are killed (deletion). They can also become anergic.

Tolerogenic

When lymphocytes are functionally inativated or killed, resulting in tolerance. Antigens that induce tolerance are tolerogenic. This happens to allow coexistence of microbiota with human host Assist immune system of pregnant females to tolerate foreign antigens derived from the father.

Peripheral T cell Tolerance: Increased delivery of inhibitory signals

a) On recognition of self antigens, T cells may also prefientially engage one of the inhibitory receptors of the CD28 family, CDLA-4 or PD-1. Anergic T cells may express higher levels of these inhibitory receptors which will inhibit responses to subsequent antigen recognition. b) Engagement of inhibitory receptors = The inhibitory receptors regulate T cell responses. They expression increases so they cant bind to small amount of B7 by accident.

Forced high levels of B7 costimulators results in

autoimmune reactiosn against antigens in that tissue. Providing second signals may break anergy and activate autoreactive T cells.

Tolerance to fetal antigens

i. Some foreign paternal antigens are expressed by the fetus which creates immunological dilemma of the mother. They need to be tolerated. ii. One mechanism for tolerance is generation of peripheral FoxP3+ regulatory T cells specific for these paternal antigens. iii. Placentation is strongly correlated with the ability to generate stable peripheral regulatory T cells. Maybe mothers who don't have this suffer from recurrent pregnancy loss (unclear) iv. Other mechansisms include 1) the exclusion of inflammatory cells from pregnant uterus. 2) Poor antigen presentation in placenta 3) Inability to generate harmful Th1 responses in a healthy pregnant uterus

Commensal microbe tolerance

i. We need some microbes because they serve essential functions, such as in the gut for digestion and skin. There's a lot of them ii. Mature lymphocytes in these tissues are cabable of recognizing the organisms but do not react against them so they mcirobes are not eliinated and inflammation is not triggered. iii. This is done by the presence of abundant IL-10 producing regulatory T cells, some dendritic cells signaling from TLRs that lead to inhibition rather than activation. Also the separation of some bacteria from intestinal immune system by the epithelium helps. iv. Crosstalk between an organism and its gut commensal microbiota has both potentiating and detrimental effects on the immune response.

Peripheral tolerance is induced when

mature T cells recognize self antigens in peripheral tissues and leads to: a) Functional inactivation (anergy) of mature peripheral autoreactive T cells b) Self reactive lymphocytes are suppressed by regulatory T cells c) Cell death of mature, peripheral, autoreactive T cells.

Direct allorecognition

most Tissues contain dendritic cells and when the tissues are transplanted, the dendritic cells are carried to the graft. When T cells in the recipient recognize donor allogenic MHC molecules on graft dendritic cells, the T cells are activated; This is direct recognition (aka direct presentation) of alloantigens. This stimulates the development of alloactive T cells (CTLs)

Allografts and xenograpfts (allogenic and xenogenic grafts) are always...

rejected by a recipient with a normal immune system. The antigens are alloantigens and xenoantigens. Antibodies and T cells receting against these are alloreactive and xenoreactive.

Immunological ignorance

when antigen specific lymphocytes don't react in any way. (antigen presence is ignored (self antigens - tolerogenic).

Similarities between CTLA-4 and PD-1

• B7 receptor family members • Expressed by activated T cells • Level of expression affected by the strength and/or duration of TCR signaling • Regulate an overlapping set of intracellular T-cell signaling proteins • Reduce T-cell proliferation, glucose metabolism, cytokine production, and survival

Treg Development

• CD4+ T cells that recognize self antigens may differentiate into regulatory T cells (Tregs) develop in the thymus or peripheral tissues

Regulation of T cell responses by CTLA-4

• CTLA-4 is expressed transiently on activated CD4+ T cells • expressed always on Tregs • in general: functions to terminate activation of responding T cells (shutting off T cell responses) Recognizes some B7 costimulators on APCs that bind to CD28 • also mediates the suppressive function of regulatory T cells • CTLA-4 works by: blocking and removing B7 molecules from the surface of APCs • reduces costimulation and preventing the activation of T cells Polymorphisms in the CTLA4 gene have been associated with some autoimmune diseases in humans.

Differences between CTLA4 and PD-1

• CTLA-4 limits T-cell responses early in an immune response, primarily in lymphoid tissues • PD-1 limits T-cell responses later in an immune response, primarily in peripheral tissues • CTLA-4 expressed by T cells; PD-1 expressed by T cells and other immune cells • CTLA-4 ligands expressed by professional antigen-presenting cells; PD-1 ligands expressed by antigen-presenting cells and other immune cells, and can be inducibly expressed on nonimmune cells, including tumor cells • PD-1 engagement interferes with more T-cell signaling pathways than does CTLA-4 engagement10 • CTLA-4 affects Treg functioning; the role of PD-1 on Tregs is unclear

IL-2 role in T cells.

• IL-2 is an example of a cytokine that serves two opposite roles: • promotes immune responses by stimulating T cell proliferation • inhibits immune responses by maintaining functional regulatory T cells

Regulation of T cell responses by PD-1

• PD-1 is expressed on CD4+ and CD8+ T cells after antigen stimulation • has an immunoreceptor tyrosinebased inhibitory motif (ITIM) typical of receptors that deliver inhibitory signals • PD-1 terminates responses of T cells to: • self antigens • chronic infections, & virus infections

Features of an Effective Immune Response

• Specificity • Trafficking • Adaptability • Target elimination • Durability (immune memory)

T cell tolerance in periphery: Suppression: Treg Function

• Tregs function to suppress (or inhibit) the activation of autoreactive naïve T cells and their differentiation by: *• contact-dependent mechanisms* • Regulatory T cells express CTLA-4, may block or remove B7 molecules expressed by APCs; make these APCs incapable of providing costimulation via CD28 & activating T cells *• secreting cytokines that inhibit T cell responses* • Some Tregs produce cytokines (e.g., IL-10, TGF-β) that inhibit the activation of lymphocytes, dendritic cells, & macrophages

TGF-beta in T cells

• cytokine transforming growth factor β (TGF-β) also plays a role in the generation of regulatory T cells (Tregs) • stimulate expression of the FoxP3 transcription factor (generating Tregs) • many cell types can produce TGF-β, but the source of TGF-β for inducing regulatory T cells in the thymus or peripheral tissues is not defined (also inhibits lymphocytes, dendritic cells and macrophages)

Diseases associated with autoantibody production are caused by

• defective tolerance in B lymphocytes • defective tolerance and diminished Tfh cell help (T Follicular Helper)

T-independent (self) antigens not recognized by T cells

• self polysaccharides • self lipids • self nucleic acids To prevent autoantibody production against these, we need to induce B cell tolerance.

Maintenance of Tregs

• survival and function of regulatory T cells are depend on cytokine IL-2 • Tregs have a high level of expression of the IL-2 receptor, and may bind & consume the IL-2 T cell growth factor, & reduce its availability for responding T cells • This role of IL-2 accounts for the severe autoimmune disease that develops in mice in which IL-2 or IL-2 receptor genes are deleted

B cell central tolerance: Deletion

▪ immature B cells that strongly recognize self antigens receive death signals and die by apoptosis ▪ similar to negative selection of immature T cells ▪ negative selection of B cells eliminates lymphocytes with high-affinity receptors for abundant, & widely expressed, cell membrane & soluble self antigens (same as in T cell compartment)

B cell central tolerance: Anergy

▪ self antigens, such as soluble proteins, may be recognized in the bone marrow with low avidity ▪ B cells specific for these antigens survive, but antigen receptor expression is reduced, and the cells become functionally unresponsive (anergic)