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Mechanisms of Immune Tolerance The Liver as Immunoprivileged Organ
Amy S Rosenberg MD Supervisory Medical Officer Office of Biotechnology
Products CDER FDA
Immunological tolerance bull Definition
ndash unresponsiveness to an antigen induced by exposure of lymphocytes to that antigen antigen-specific (unlike ldquoimmunosuppressionrdquo)
bull Significance ndash All individuals are tolerant of their own antigens to a greater or
lesser extent (self-tolerance) breakdown of self-tolerance results in autoimmunity
ndash Therapeutic potential Inducing tolerance may be exploited to prevent graft rejection treat autoimmune and allergic diseases and prevent immune responses in gene therapy and stem cell transplantation
2
How Self is Self
Self proteins can be immunogenic and tolerance to them broken (eg by administration of a therapeutic homolog) TolerogenicityImmunogenicity of self proteins depends largely on
ndash Abundance determines the degree to which developing potentially autoreactive T and B cells are tolerized
ndash Alteration in chemicalphysical structure aggregation post-translational modifications (PTMs) chemical degradation
ndash Adjuvants bull Extrinsic innate immune response modifiers
bull Intrinsic immunomodulatory properties of the protein
Expect Immunogenicity No tolerance
Neutralize Product Hypersensitivity
Rare Immunogenicity Autoimmunity
Foreign proteins Self Proteins
FOREIGN SELF
Expect Immunogenicity
No tolerance Neutralize Product
Hypersensitivity
Potential Immunogenicity
Incomplete tolerance Altered structure Antigen Present
Epitope spreading
Rare Immunogenicity
Robust tolerance Novel Route of Administration Adjuvants HLA Haplotype Specific
bull Low abundance self-protein bull Aggregates of self proteins bull Chemical degradationmodification of self proteins bull Adjuvants
Antibody Response to Proteins The Mediators
BCR
Short lived
Plasma cell
Antigen
DC
Peptide
Cytokines
MHCII TCR
Memory B cell
Antibodies Helper T cell Tfh
B cell
Antibodies
Long lived
Plasma cell
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Immunological tolerance bull Definition
ndash unresponsiveness to an antigen induced by exposure of lymphocytes to that antigen antigen-specific (unlike ldquoimmunosuppressionrdquo)
bull Significance ndash All individuals are tolerant of their own antigens to a greater or
lesser extent (self-tolerance) breakdown of self-tolerance results in autoimmunity
ndash Therapeutic potential Inducing tolerance may be exploited to prevent graft rejection treat autoimmune and allergic diseases and prevent immune responses in gene therapy and stem cell transplantation
2
How Self is Self
Self proteins can be immunogenic and tolerance to them broken (eg by administration of a therapeutic homolog) TolerogenicityImmunogenicity of self proteins depends largely on
ndash Abundance determines the degree to which developing potentially autoreactive T and B cells are tolerized
ndash Alteration in chemicalphysical structure aggregation post-translational modifications (PTMs) chemical degradation
ndash Adjuvants bull Extrinsic innate immune response modifiers
bull Intrinsic immunomodulatory properties of the protein
Expect Immunogenicity No tolerance
Neutralize Product Hypersensitivity
Rare Immunogenicity Autoimmunity
Foreign proteins Self Proteins
FOREIGN SELF
Expect Immunogenicity
No tolerance Neutralize Product
Hypersensitivity
Potential Immunogenicity
Incomplete tolerance Altered structure Antigen Present
Epitope spreading
Rare Immunogenicity
Robust tolerance Novel Route of Administration Adjuvants HLA Haplotype Specific
bull Low abundance self-protein bull Aggregates of self proteins bull Chemical degradationmodification of self proteins bull Adjuvants
Antibody Response to Proteins The Mediators
BCR
Short lived
Plasma cell
Antigen
DC
Peptide
Cytokines
MHCII TCR
Memory B cell
Antibodies Helper T cell Tfh
B cell
Antibodies
Long lived
Plasma cell
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
How Self is Self
Self proteins can be immunogenic and tolerance to them broken (eg by administration of a therapeutic homolog) TolerogenicityImmunogenicity of self proteins depends largely on
ndash Abundance determines the degree to which developing potentially autoreactive T and B cells are tolerized
ndash Alteration in chemicalphysical structure aggregation post-translational modifications (PTMs) chemical degradation
ndash Adjuvants bull Extrinsic innate immune response modifiers
bull Intrinsic immunomodulatory properties of the protein
Expect Immunogenicity No tolerance
Neutralize Product Hypersensitivity
Rare Immunogenicity Autoimmunity
Foreign proteins Self Proteins
FOREIGN SELF
Expect Immunogenicity
No tolerance Neutralize Product
Hypersensitivity
Potential Immunogenicity
Incomplete tolerance Altered structure Antigen Present
Epitope spreading
Rare Immunogenicity
Robust tolerance Novel Route of Administration Adjuvants HLA Haplotype Specific
bull Low abundance self-protein bull Aggregates of self proteins bull Chemical degradationmodification of self proteins bull Adjuvants
Antibody Response to Proteins The Mediators
BCR
Short lived
Plasma cell
Antigen
DC
Peptide
Cytokines
MHCII TCR
Memory B cell
Antibodies Helper T cell Tfh
B cell
Antibodies
Long lived
Plasma cell
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Expect Immunogenicity No tolerance
Neutralize Product Hypersensitivity
Rare Immunogenicity Autoimmunity
Foreign proteins Self Proteins
FOREIGN SELF
Expect Immunogenicity
No tolerance Neutralize Product
Hypersensitivity
Potential Immunogenicity
Incomplete tolerance Altered structure Antigen Present
Epitope spreading
Rare Immunogenicity
Robust tolerance Novel Route of Administration Adjuvants HLA Haplotype Specific
bull Low abundance self-protein bull Aggregates of self proteins bull Chemical degradationmodification of self proteins bull Adjuvants
Antibody Response to Proteins The Mediators
BCR
Short lived
Plasma cell
Antigen
DC
Peptide
Cytokines
MHCII TCR
Memory B cell
Antibodies Helper T cell Tfh
B cell
Antibodies
Long lived
Plasma cell
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
FOREIGN SELF
Expect Immunogenicity
No tolerance Neutralize Product
Hypersensitivity
Potential Immunogenicity
Incomplete tolerance Altered structure Antigen Present
Epitope spreading
Rare Immunogenicity
Robust tolerance Novel Route of Administration Adjuvants HLA Haplotype Specific
bull Low abundance self-protein bull Aggregates of self proteins bull Chemical degradationmodification of self proteins bull Adjuvants
Antibody Response to Proteins The Mediators
BCR
Short lived
Plasma cell
Antigen
DC
Peptide
Cytokines
MHCII TCR
Memory B cell
Antibodies Helper T cell Tfh
B cell
Antibodies
Long lived
Plasma cell
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Antibody Response to Proteins The Mediators
BCR
Short lived
Plasma cell
Antigen
DC
Peptide
Cytokines
MHCII TCR
Memory B cell
Antibodies Helper T cell Tfh
B cell
Antibodies
Long lived
Plasma cell
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Cellular Interactions in Generation of T cell Mediated Responses
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
CD4+ T cell
TCR TCR TCR
APC
Uptake by APC
Necrotic and apoptotic
cell material
MHC class-II
Cell T cell help
MHC class-II MHC class-I
APC
CD4+ T cell CD8+ T cell
A B
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
T Cells More Robustly Tolerant than B Cells to Self Proteins
(Weigle 1980)
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
T Cells More Robustly Tolerant to Self-ProteinsThymic Mechanisms
bull Negative selection ndash T cells with high affinity for thymicperipheral tissue antigens that
access thymus in sufficient quantities ndash T cells with high affinity for peripheral tissue antigens whose
expression in thymus is mediated by transcription factor AIRE bull Natural (thymically generated) regulatory T cells (Tregs)
ndash Lineage deviation of T cells with high affinity to proteins expressed in thymus to immune suppressive regulatory T cells (Tregs) distinguished by FOXP3 transcription factor neuropilin and helios
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Consequences of Self Antigen Recognition in Thymus
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier 10
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Mutations in Tregulatory cell Transcription Factor FoxP3 Confer Autoimmunity by Deficiency of Tregs
(Sakaguchi et al 2008)
Mother of IPEX patient
IPEX patient
Normal
Autoimmune disease
Inflammatory bowel disease
Allergy
X
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Confidential 7172016 12
167
Figure described in Palomares O Martiacuten-Fontecha M Lauener R Traidl-Hoffmann C Cavkaytar O Akdis M Akdis CA Regulatory T cells and immune regulation of allergic diseases roles of IL-10 and TGF- β Genes Immun 2014 Dec15(8)511-20
Natural tTreg nTreg originate in the thymus positively selected Inducedperipheral (ip)Tregs generated in the periphery originate from CD4+ T cells in response to environmental antigens
Tregs Arise in the Periphery (ip Tregs) as well as in the Thymus (tnTregs)
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
13
What Self Antigens are Seen in the Thymus
bull Ubiquitous cell-associated and circulating proteins bull Peripheral tissue antigens expression mediated by
Autoimmune Regulator (AIRE) transcription factor in thymic medullary epithelial cells and extrathymic bone marrow derived APC (eTACs) bull signal self-reactive thymocytes for death or lineage
deviation to Tregs
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
AIRE Promotes Expression of Peripheral Tissue Antigens in the Thymus DeletingDeviating High Affinity Autoreactive T cells
(Mathis and Benoist 2007)
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
AIRE Mutations are Associated with Autoimmune Polyendocrine Syndrome Type 1
(Kampe et al 2008)
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
AIRE Mediated Expression of of Liver Proteins in the Thymus
Cyp1a2 Prg4 Apoa1 Fgg Tdo2 Ambp Gys2 Fgb Alb Itih4 Apoa2 Apoc2 Mat1a Orm1 Itih3 Hp Igfbp1 Reln Hal Cyp26a1 Hamp
Cytochrome P450 family 1 subfamily A polypeptide 2
Proteoglycan 4 Apolipoprotein A-I Fibrinogen gamma chain Tryptophan 23-dioxygenase Alpha-1-microglobulinbikunin precursor Glycogen synthase 2 (liver) Fibrinogen beta chain Albumin Inter-alpha-trypsin inhibitor heavy chain
family member 4 Apolipoprotein A-II Apolipoprotein C-II Methionine adenosyltransferase I alpha Orosomucoid 1 Inter-alpha-trypsin inhibitor heavy chain 3 Haptoglobin insulin-like growth factor binding protein 1 reelin Histidine ammonia-lyase cytochrome P450 family 26 subfamily a
polypeptide 1 Hepcidin antimicrobial peptide
2272 2631 6268 1749 1384 1392 922 1535 1465 191 3341 3456 995 2983 1448 1083 152 120 103 273 337
206 317 959 27 225 266 195 367 439 58 1118 1216 378 116 58 496 83 223 22786 714 9461
Gene ID Gene Annotation WT Aire KO
11 8 6 6 6 5 4 4 3 3 2 2 2 2 2 2 1 05 05 04 04
Fold Change
223 94 8043 5906 279 3641 52 6643 73775 681 11107 249 424 5651 300 15456 416 15 28 14 163
Liver Expression Protein atlas
The list is the combination of SI from PNAS 2012 by Matthieu Giraud and 02670 Table2 (Microarray data from Affymetrix MgU74Av2 gene chip by using 20040429 Affymetrix annotation FPKM values or number of Fragments Per Kilobase gene model and Million reads were calculated as the sum of all its protein-coding transcripts and the average FPKM value for replicate samples were used as abundance scores) from RNA-seq analysis
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
CD4+ T cells Specific for Tissue Restricted Antigens do not Undergo Extensive Thymic Deletion Expanded Tregs Mediate Tolerance
Thymus Periphery
Ubiquitous self-antigen
Tissue-restricted self-antigen (no thymic expression)
Tconv
Treg
No self-antigen
Deletion Pancreatic self antigen
Treg expansion Lung or
intestinal self antigen
R
R R
R
R
X Ignorance
Suppression
Legoux et al 2015 Immunity 43 896ndash908
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
CD4+ T cell Fate Depends on Cytokine Milieu in Site of Activation
Immunosuppression Engraftment
IL-23 IL-6 IL-21
TGFβ
IL-4
IL-12
Naiumlve CD4+ T cell
Th2 (IL-4 IL-5
IL-13)
Treg (IL-10 TGFβ)
TH17 (IL-17)
TH1 (IFNγ)
Liver graft rejection
Stat4 T-bet
Stat6 Gata3
Stat3 RORγ
Stat5 Foxp3
(Sanchez-Fueyo A and TB Strom Gastroenterology 2011)
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Autoimmunity is Suppressed by both Thymic and Peripheral Tregs
Local immune suppression Oral tolerance Fetal tolerance Mucosal tolerance
Nrp-1
APC
Thymus tTreg
Tnaive
Teff
pTreg
Teff Site of Inflammation
specialized APC
Immune homeostasis Autoimmune responses
Yadav M et al Frontiers in Immunology 2013
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Mechanisms by which Tregs Suppress Immune Responses
20
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Tolerance Mechanisms in the Liver bull 80 of the blood that passes through the liver sinusoids comes from the GI tract
carrying harmless dietary and commensal organism antigens the default immune response must be tolerancehellipmaintained by the myriad of tolerogenic APC in the liverrdquo hellipcontinuous exposure of liver cells to these entities leads to ldquoendotoxin tolerancerdquo ldquo
bull Antigen presentation within the liver mediated by a variety of cell types including plasmacytoid and myeloid dendritic cells (pmDC) Kupffer Cells sinusoidal endothelial cells hepatic stellate cells and hepatocytes generally leads to T cell tolerance and not immunity
ndash pDC-IL-27 secretion mediates expression of PDL-1 on pDC promoting activation and expansion of Tregs
ndash mDC express PDL-1 induce IDO
ndash KC- expression of Fas-L kills CD8T cells IDO IL-10 expression of PDL-1
ndash Hepatocytes induce apoptosis of CD4+ and CD8+ Tcells
ndash LSEC functional inactivation of CD8+ T cells and bias CD4+ T cells to Treg-dependent on a cell surface expressed lectin (LSEC lectin)
bull CD8+T cells transiently activated then undergo apoptosis or exhaustion bull Generating robust immune responses appears to hinge on full activation of CD4+ T
cells which provide help to CD8+ T cells
21
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Immunosuppressive Circuits in Liver Mediated Mainly Through Liver Resident Myeloid Cells
IL-10
TGF-β1
IDO
Cox2-gtPGE2
FasL
HEP
Dead HEP
ldquoMDSCrdquo
LSEC
pDC
mDC
HSC Kupffer
PD-L1
LSECtin
IL-10
IL-27
PD-L1
IDO PD-L1
TGF-β1 + retinol
PD-L1
kynurenine
IL-10 TGFβ1 other
IFN-γ
HGF
T reg
T reg
8+
4+
Crispe IN 2014 Hepatology
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Generating Immune Responses in the Liver
bull NK NKT γδ and mucosal associated innate lT cell (MAIT) activation and responses may be critical in response to infectious organisms ndash Promote DC maturation into APCs activate CD4+ CD8+ T cells Bcells
PMNs and macrophages via contact and cytokine dependent interactions
bull However none of these populations uses HLA the single greatest risk factor for autoimmune disease as restriction element for immune response suggesting criticality for mediating response to infection but potentially only indirect mechanism of action in triggering AIH spectrum
23
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Innate T Cells Present at High Levels in the Human Liver Recognize Microbial or Stress Induced Antigens Indicative of
Infection but not in the Context of HLA (Doherty D 2016 J Autoimmunity)
Type 1 NKT cell Type 2 NKT cell MAIT cell γδ T cell γδ T cell γδ T cell γδ T cell
Vα24Jα18 Diverse Vα72Jα33 Vδ1 Vδ1 Vγ9Vδ2 Vδ3
CD1d CD1d MR1 MICA MICB Rae1 CD1c CD1d Butyrophilin 3A1 CD1d
Glycolipids Glycolipids Riboflavin metabolites Stress-inducible proteins Glycolipids Pyrophosphates Glycolipids
Cell Type T Cell Receptor Antigen-presenting
Molecule Stimulatory
Ligands
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Genetic Basis of ToleranceAutoimmunity
bull Genetic predisposition of autoimmune diseases bull HLA genes
ndash Major genetic association with autoimmune diseases ndash Disease-associated alleles are present in normal individuals
bull Non-HLA genes ndash Many loci identified by genomic methods (eg genome wide
association studies [GWAS]) ndash Examples include FoxP3 AIRE CD25 (IL2R-α) NOD (sensor of
microbes) PTPN22 (tyrosine phosphatase) bull Multiple genes are associated with autoimmunity
ndash No single mutation causes common autoimmune diseases
25
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
HLA is the Strongest Genetic Factor for Susceptibility to Autoimmune Disease
26
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Non-HLA Genes in Autoimmunity
27
Lymphadenopathy enteropathy eczema infection
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
HLA Association with AIH
DQB DQA DRB1 DRB1 DRB3 DRB3 Previous data DRB1 DRB1 DRB1
Locus Allele OR p Value n
603 103
1301 1302
101 202
13
13 + 3 3
218 455 680 016 149 055 69 106 53
ns 0013 000247 000845 ns ns 100E-06 100E-07 100E-04
66 33 39 39 84 95
111
111 33
Goldberg AC et al 2001 Human Immunology
Previous data refer to odds ratio (OR) observed when a larger cohort of patients was analyzed by low resolution PCR-SSP for the HLA-DRB1 locus
DRB11301 appears the major susceptibility factor its only amino acid different from DRB11302 is in position 86 corresponding to pocket 1 in the peptide-presenting groove-glycine for valine
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Polymorphisms in TNF-α Gene Associated with AIH
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
TNFA2 + + ndash ndash
Phenotype Patients Comparison OR P
HLA-A1 + ndash + ndash
HLA-B8 + ndash + ndash
HLA-DRB10301 + ndash + ndash
(n = 83) 35 12 4 32
(n = 83) 43 4 0 36
(n = 85) 40 8 4 71
(n = 98) 15 11 9 63
(n = 98) 17 9 3 69
(n = 102) 15 12 3 4
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
+ + vs ndash ndash 36 + ndash vs ndash ndash NS ndash + vs ndash ndash NS
lt00004
lt00004
lt00003
Controls
Cookson S et al 1999 Hepatology
Of note is high degree of linkage disequilibrium between the TNFA locus and both HLA B8 and HLA DRB10301 within the extended haplotype B8-TNFA2- DRB10301
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Pathogenesis of Organ-Specific Autoimmunity
Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011 c Elsevier
Failure of control mechanisms is the underlying cause of most autoimmune diseases
30 Abbas Lichtman and Pillai Cellular and Molecular Immunology 7th edition 2011
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Deliberately Breaking Self Tolerance
bull Monoclonal antibodies to checkpoint inhibitors (PD-1 PDL-1)
bull Monoclonal antibodies to suppressive molecules (eg CTLA-4 IL-2Ra)
bull Chimeric Antigen Receptor T cells (CAR-T) bull Inhibitors of Indoleamine Diooxygenase (IDO)
31
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
The Immune System on a Knifersquos Edge Tipping the Balance for Therapy of Serious Diseases
Cancer Chronic Infection
Autoimmunity Graft Rejection
Checkpoint Inhibitor
Antagonists
Checkpoint Inhibitor Agonists
Treg
Teff
Treg
Teff
Treg
Teff
Autoimmunity CancerChronic Infection
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
Acknowledgements
bull Mark Avigan and John Senior OSE FDA bull Michael Norcross OBP FDA bull Jeff Bluestone UCSF bull Abul Abbas UCSF bull Daniela Verthelyi OBP FDA bull Steven Kozlowski OBP FDA
33
34
34