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Self Tolerance, Regulation and Immune imbalances (i.e. & )

Dr. Lavkush Dwivedi Dept. of Biomedical Sciences Bundelkhand University, Jhansi The immunological equilibrium: balancing3 activation and control

Activation Tolerance Effector T cells Regulatory T cells

Normal: reactions Controlled response to against pathogens pathogens Inflammatory No response to self disease, e.g. reactions against self 3 Central and peripheral tolerance to self

The principal fate of that recognize self in the generative organs is death (deletion), BUT:

Some B cells may change their specificity (called “receptor editing”)

Some T cells may differentiate into regulatory (suppressor) T lymphocytes

Abbas, Lichtman and Pillai. Cellular and Molecular , 7th edition, 2011 c Elsevier 4

Consequences of self recognition in thymus

Abbas, Lichtman and Pillai. Cellular and Molecular Immunology, 7th edition, 2011 c Elsevier 5

What self antigens are seen in the thymus?

• The thymus has a special mechanism for displaying peripheral tissue antigens in thymic medullary epithelial cells, where they signal self-reactive for death. • AIRE Genes are there which control the maturation of only non-self reactive cells. • Failure of AIRE leads to autoimmune polyendocrinopathy with candidiasis and ectodermal dysplasia (APECED), 6 Deletion of self-reactive T cells in the thymus:

Abbas, Lichtman and Pillai. Cellular and Molecular Immunology, 8th edition, 2014 AIRE (autoimmune regulator) is a regulator of gene transcription that stimulates thymic expression of many self antigens which are largely restricted to peripheral tissues 7 Peripheral tolerance

Abbas, Lichtman and Pillai. Basic Immunology, 4th edition, 2014 8 T cell anergy

Abbas, Lichtman and Pillai. Cellular and Molecular Immunology, 7th edition, 2011 c Elsevier 9 CTLA-4 competitively inhibits B7-CD28 engagement

APC APC

B7 B7 CTLA-4 CD28

T Cell T cell (activated T cell or Treg) Costimulation → T cell activation CTLA-4 blocks and removes B7 → lack of costimulation → T cell inhibition 10 Major functions of selected B7-CD28 family members • CD28-B7: initiation of immune responses • ICOS-ICOS-L: T cell help in germinal

Activation Activation center reactions ( responses)

• CTLA-4-B7: inhibits early T cell responses in lymphoid organs • PD-1:PD-L1,2: inhibits effector T cell

Inhibition responses in peripheral tissues 11 Blocking CTLA-4 promotes tumor rejection: CTLA-4 limits immune responses to tumors

Administration of antibody that blocks CTLA-4 in tumor-bearing mouse leads to tumor regression 12

The PD-1 inhibitory pathway

• PD-1 recognizes two widely expressed ligands (PD-L1, PD-L2)

• Knockout of PD-1 leads to (less severe than CTLA-4-KO)

• Role of PD-1 in T cell suppression in chronic infections, tumors? 13 T cell “exhaustion” in chronic viral infections

Memory T cells: Effector enhanced Naïve CD8+ T cells antiviral T cells Acute infection: responses clearance of virus Virus Chronic infection: persistence of virus

Exhausted T cells: inability to respond to virus (expression of inhibitory receptors, e.g. CTLA-4, PD-1) 14 Actions of PD-1

• PD-1 attenuates TCR signaling in responding T cells • Limits harmful consequences of chronic stimulation with persistent antigen (self, tumors, chronic viral infections)

• Greater role in CD8 than in CD4 T cells

• Also expressed on follicular helper T cells; function? Checkpoint blockade for cancer immunotherapy

e.g. ipilimumab Ribas A. N Engl J Med 2012;366:2517-2519. Checkpoint blockade for cancer immunotherapy

16 e.g. ipilimumab e.g. nivolumab, pembrolizumab Ribas A. N Engl J Med 2012;366:2517-2519. 17

Risks of blocking CTLA-4 or PD-1

• Blocking a mechanism of self-tolerance leads to: • Autoimmune reactions (a new cottage industry for clinicians?) – Colitis and dermatitis are common – Vitiligo, Endocrinopathies, hepatitis less common but described – Severity of adverse effects has to be balanced against potential for treating serious cancers – Less severe with anti-PD1 antibody 18 Regulatory T cells

Abbas, Lichtman and Pillai. Cellular and Molecular Immunology, 8th edition, 2014, Elsevier 19 Properties of regulatory T cells

• Phenotype: CD4+, high IL-2 receptor (CD25), low IL-7 receptor, Foxp3 transcription factor; other markers • Essential features of stable Tregs: – Foxp3 expression: Foxp3 mutations --> autoimmune disease (IPEX); in mice, disease can be corrected by providing normal Foxp3+ cells – CD25 (IL-2Ra) expression: IL-2 is a necessary survival factor – CTLA-4 expression: required for suppressive function of most Tregs 20 Mechanisms of action of Foxp3+ Treg cell • CTLA-4 on Tregs removes B7 on APCs, reduces CD28 engagement and T cell activation – Genetic deletion of CTLA-4 in Foxp3+ cells results in severe systemic autoimmunity and lymphoproliferation • Inhibitory produced by Tregs (TGF-b, IL-10) suppress immune responses (DCs, MACs, T cells) – IL-10 deletion in Foxp3+ cells results in colitis – IL-10 is also produced by Foxp3- cells • Consumption of IL-2 Functions of Interleukin-2: the dogma 21 22 Dual roles of IL-2 in T cell responses

Surprising conclusion from knockout mice: the non-redundant function of IL-2 is in controlling immune responses Pathogenesis of autoimmunity 23 Therapy of immune disorders: rational approaches target 24 lymphocyte activation and subsequent 25 The landscape of T cell activating and inhibitory receptors

TIGIT 26

Autoimmunity • Autoimmunity is a self- damaging immune effector response manifested in various autoimmune diseases (AID). • Any disease that results from such an aberrant immune response is termed an "autoimmune disease

Immunodeficiency • Immunodeficiency is a state in which the 's ability to fight infectious disease and cancer is compromised or entirely absent. • Most cases of immunodeficiency are acquired ("secondary") due to extrinsic factors that affect the patient's immune system. 27 Immunodeficiency

• Humoral – innate - complement, MBL acquired immunity – immunoglobulins (B lymphocytes)

• Cell mediated immunity – innate immunity – - acquired immunity – T lymphocytes

• Primary – congenital, genetically defined, symptoms predominantly at early age

• Secondary – the onset of symptoms at any age chronic diseases effect of irradiation immunosuppression surgical intervention, injuries stress 28

Immunodeficiencies – major clinical features

• - microbial infections (encapsulated bacteria) respiratory - pneumonia, sinusitis, otitis GIT – diarrhea • – microbial infections (pyogenic), sepsis edema (HAE) – C1-INH deficiency • T lymphocytes - bacterial, fungal, viral GIT – diarrhoea respiratory – pneumonia, sinusitis • Phagocytes - abscesses, recurrent purulent skin infections granulomatous inflammation 29 I. Primary – phagocytic cell defects Qualitative – phagocytes functional disorders, various enzyme deficits, inability of phagocytes to degrade the ingested material

Chronic Granulomatous Disease (CGD) • Approximately in 60% X-linked

• Enzymatic inability to generate toxic oxygen metabolites (H2O2) during oxygen consumption) - defect in neutrophilic cytochrome b (part of complex containing NADPH oxidase) • Inability to kill catalase producing bacteria such as Staph.aureus, Pseud.aeruginosa • Clinical features: granulomas of skin, organs • Treatment: long-term ATB prophylaxis, in more severe cases BMT 30 II. Primary immunodeficiencies – disorders X-linked hypogamaglobulinemia (XLA, Bruton’s agammaglobulinemia) • Most common X-linked form • Block of maturation of pre-B lymphocytes into B lymphocytes (Bruton’s tyrosine kinase defect) • Undetectable or very low serum levels of Ig, absence of B cells • Symptoms: pneumonias, pyogenic otitis, increased occurrence of pulmonary fibrosis • Treatment: life-long Ig replacement

CVID – Common Variable Immunodeficiency • Heterogenous group of B cell functional defects • Low levels of IgG and IgA, B cell counts normal • Symptoms: onset mostly between 2nd and 3rd decade recurrent respiratory tract infections (pneumonia, sinusitis) • Treatment: life-long Ig replacement 31 II. Primary immunodeficiencies – B cell disorders Selective IgA deficiency • Disorder of B cell function: absence of IgA, levels of the other Ig normal • Recurrent mild/moderate infections or asymptomatic • Risk of reaction to live attenuated vaccines or generation of anti-IgA antibodies after blood transfusion

Selective IgG subclasses deficiency, specific IgG deficiency • B cell functional disorder • Onset of symptoms in childhood, mostly respiratory tract infections caused by encapsulated bacteria (H.influenzae, Pneumococci)

Transitory hypogammaglobulinemia of infancy • Milder and transitory decrease of IgG and IgA 32

III. Primary immunodeficiencies – T cell disorders diGeorge syndrome • Disorder of development of 3rd and 4th branchial pouch → congenital heart disease + absence of thymus + absence of parathyroid glands • Complete or partial • Symptoms: symptoms of cong.heart dis. – prominent immunodeficency – variable (mild functional → absence of T cells) hypocalcemic spasms – possible mental deficit • Treatment: symptomatic, in complete form BMT 33 IV. Primary immunodeficiencies – combined defects of T and B cells SCID – Severe Combined ImmunoDeficiency • X-linked recessive disease, combined disorder of humoral and cell mediated immunity • Severe disorder (patients often die during until 2nd year of age), onset of symptoms soon after birth (severe diarrhoea, pneumonia, meningitis, BCGitis) • Immunological features: typically lymphopenia, absence of T cells, hypogammaglobulinemia • Forms: ✓ X-linked form – the most common, T-B+NK-, defect of common gamma chain shared by receptors of various important cytokines → absence of T cells ✓ AR form – T-B-, enzymatic defect (ADA, PNP) → accumulation of metabolites toxic for DNA synthesis → lymphopenia ✓ Ommen sy. - defect of recombinases → defect of VDJ recombination → proliferation of one or more clones of autoreactive T cells • Treatment: BMT is of critical significance, in ADA deficiency gene therapy ATB, IVIG 34 V. Primary immunodeficiencies – complement system disorders

Hereditary angioedema (HAE) • Absence or functional defect of C1-inhibitor • Recurrent swellings of skin or mucosa (oropharynx, gut) triggered by intercurrent infect, trauma or surgery • Uncontroled activation of kinin system • Laryngeal edemas could be life-threatening, immediate treatment is necessary 35 Secondary immunodeficiency

• Acute and chronic viral infections – infectious mononucleosis, influenza • Metabolic disorders- uremia • Autoimmune diseases – against immunocompetent cells (, lymphocytes); autoimmune phenomena also after administration of certain drugs (e.g. oxacilin, quinidine) • Chronic GIT diseases • Malignant diseases (leukemia) • Hypersplenism/asplenia • Burn, postoperative status, injuries • Severe nutritional disorders • Chronic infections • Ionizing radiation • Drug induced immunodeficiencies (chemotherapy) • Immunosupressive therapy • Chronic stress • Chronic exposure to harmful chemical substances 36 Acquired ImmunoDeficiency Syndrome (A.I.D.S.)

• Caused by retrovirus HIV 1 or HIV 2 • Virus has a tropism for cells bearing CD4 surface marker (Th CD4+ lymphocytes); also infects and CNS cells • Viral genome transcribes into human DNA and infected cell provides viral replication • Diagnosis: serological (3 to 6 weeks after primoinfection, PCR • Transmission: sexual contact contact with blood or blood products mother-to-child – prenatally, delivery, breast feeding • Phases: acute (flu-like symptoms) ✓asymptomatic – months to years, viral replication, loss of Th cells ✓symptomatic – infections, autoimmune disorders, malignancies, ✓final – systemic breakdown, opportune infections (Pneumocystis ✓jirovecii, Cryptococcus neoformans, Toxoplasma gondii, ✓Candida albicans, CMV etc.) ✓- Kaposi’s sarcoma • Treatment – ✓Reverse transcriptase inhibitors (e.g.zidovudine) ✓Protease inhibitors - block the viral protease enzyme ✓Combined drug therapy ✓Antimicrobial agents 37 Autoimmune diseases

• Experimental models are revealing pathways of immune regulation • But experimental animals are often inadequate models of human diseases • Improving technologies for human genetic and phenotypic analyses are enabling studies of patients • Challenges: – Defining which mechanisms of fail in different autoimmune diseases – Using this knowledge to develop therapies 38

AUTOIMMUNE PATOLOGICAL RESPONSE- ETIOLOGY • the diseases are chronic and usually irreversible • incidence: 5%-7% of population, higher frequencies in women, increases with age • factors contribute to autoimmunity:

✓ internal (HLA association, polymorphism of genes, defect in genes regulating apoptosis, polymorphism in genes for TCR a H immunoglobulin chains, association with immunodeficiency, hormonal factors) ✓ external (infection, stress by activation of neuroendocrine axis and hormonal dysbalance, drug and ionization through modification of autoantigens) 39 CLINICAL CATEGORIES • systemic - affect many organs and tissue

• organoleptic - affect predominantly one organ accompanied by affection of other organs (inflammatory bowel diseases, celiac disease, AI hepatitis, pulmonary fibrosis)

• organ specific - affect one organ or group of organs connected with development or function 40

SYSTEMIC AUTOIMMUNE DISEASES

• Systemic lupus erythematosus • Rheumathoid arthritis • Sjögren‘s syndrome • Dermatopolymyositis • Systemic sclerosis • Mixed connective tissue disease • Vasculitis 41 SYSTEMIC LUPUS ERYTHEMATOSUS • chronic, inflammatory, multiorgan disorder

• autoantibodies react with nuclear material and attack cell function, immune complexes with dsDNA deposit in the tissue

• general symptoms: include malaise, fever, weight loss • multiple tissue are involved including the skin, mucosa, kidney, joints, brain and cardiovascular system

• characteristic features: butterfly rash, renal involvement, CNS manifestation, pulmonary fibrosis 42 RHEUMATOID ARTHRITIS

• chronic, inflammatory disease with systemic involvement • characterized by an inflammatory joint lesion in the synovial membrane, destruction of the cartilage and bone, results in the joint deformation • clinical features: arthritis, fever, fatigue, weakness, weight loss • systemic features: vasculitis, pericarditis, uveitis, nodules under skin, intersticial pulmonary fibrosis • diagnostic tests: elevated C- reactive protein and ESR, elevated serum gammaglobulin levels - autoantibodies against IgG = rheumatoid factor (RF), a-CCP (cyclic citrulline peptid), ANA - X-rays of hands and legs- show a periarticular porosis, marginal erosion 43 Vasculitis

• characterized by inflammatory destruction of vessels leading to thrombosis and aneurysms • proliferation of the intimal part of blood-vessel wall and fibrinoid necrosis • affect mostly lung, kidneys, skin

• diagnostic tests: elevated ESR, CRP, leucocytosis, biopsy of affected organ (necrosis, granulomas), angiography 44

ORGANOLEPTIC AUTOIMMUNE DISEASES • Ulcerative Colitis • Ulcerative colitis • chronic inflammation of the • Crohn‘s disease large intestine mucosa and submucosa • Autoimmune hepatitis • features: diarrhea, bloody • Primary biliary cirhosis and mucus stools • Pulmonary fibrosis • extraintestinal features (arthritis, uveitis) • Autoantibodies: pANCA, a- large intestine 45 Crohn‘s disease

• The granulomatous inflammation of whole intestinal wall with ulceration and scarring that can result in abscess and fistula formation • The inflammation of Crohn's disease the most commonly affects the terminal ileum, presents with diarrhea and is accompanied by extraintestinal features - arthritis, spondylitis • antibodies against Saccharomyces cerevisiae (ASCA), a- pancreas 46 AUTOIMMUNE ENDOCRINOPATHY • Hashimoto‘s thyroiditis ✓ Result to hypothyroidism ✓ Treated with autoantibodies against thyroidal peroxidase (a-TPO) and thyroglobuline (a-TG) • Graves-Basedow disease ✓ Autoantibodies against thyrotropin receptor cause thyroid cells proliferation. ✓ Thyrotoxicosis due to overproduction of thyroid hormone (patient exhibit fatigue, nervousness, increased sweating, palpitations, weight loss, exophtalmus) • Diabetes mellitus I. type • Addison‘s disease • Autoimmune polyglandular syndrome 47 AUTOIMMUNE ENDOCRINOPATHY

Diabetes mellitus I type ✓ Characterized by an inability to process sugars in the diet, due to a decrease in or total absence of insulin production ✓ Results from immunologic destruction of the insuline- producing β-cells of the islets of Langerhans in the pancreas ✓ Anti-GAD (glutamic acid decarboxylase (primary antigen), anti- islet cell, anti- insulin autoantibodies formed due to infilteration of islets with B & T Cells. 48 AUTOIMMUNE NEUROPATHY

• Myasthenia gravis ✓ Caused due to overexcitation of muscles, induced by binding of auto-antibodies on Ach receptors. Also autoantibodies bind with ACH Esterase enzyme (Responsible for break down of extra Acetylcholine) • Multiple sclerosis ✓ Chronic demyelinizing disease with abnormal reaction of T cells to myeline protein because of mimicry between a virus and myeline protein. ✓ Autoantibodies against MOG (myelin-oligodendrocyte glycoprotein) 49 AUTOIMMUNE CYTOPENIA

• AI hemolytic disease- autoantibodies against membrane erythrocyte antigens

• AI trombocytopenia - autoantibodies against trombocyte antigens (GPIIb/IIIa)

• AI neutropenia - autoantibodies against membrane neutrofil antigens 50

Treatment of autoimmune diseases

❑ Systemic AI – non-specific immunosuppression ✓ Alkylating agents - Cyclophosphamide ✓ Purine analogs - Azathioprine, mycophenolate, ✓ Anti-metabolites - Methotrexate ✓ Anti-biotics - cyclosporin A, tacrolimus ✓ monoclonal antibodies ❑ Organ specific AI - Non-specific immunosuppression ❑ Endocrinopathies-Substitution of lacking product of endocrinal gland destroyed by AI process ✓ Insulin, thyroid gland hormones 51

“Nothing can be more auto toxicus than your negative attitude because life is 10% what you face and 90%, the way you deal with that.”

Thanks