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Transplantation Immunopathology Lab Transplantation Centre Dpt of Medicine – CTO/IAL PD Dr Déla Golshayan, MD PhD
Tolerance in Solid Organ Transplantation
SSN Interlaken, 2013
Billingham, Brent, Medawar. Nature 1953
BL skin Tx
BL skin
no rejection
rejection
3rd party skin
Pregnant A strain
Adult A strain
Adult A strain
« Ac%vely acquired tolerance of foreign cells »
• Successful weaning from immunosuppression stable gra> func?on >1yr post Tx liver (up to 20%) >> kidney
Sustained state of immune hyporesponsiveness to donor Ags
no donor-‐specific T cells, B cells or DSA preserved gra> func?on and histology preserved immune responses against 3rd party Ags
• Biomarkers? Specific molecular and cellular signatures of immune tolerance to
iden?fy stable vs. rejectors vs. tolerant Tx recipients EU, USA consor?ums
« Opera%onal » tolerance in clinical transplanta%on
“non-self” antigens
(pathogens, tumors, Tx alloantigens)
“ self ” antigens
(self-tissues, commensal bacteria, dietary proteines …)
Immune activation Rejection
Immune regulation /suppression Tolerance
Protective Immunity
Immune Tolerance
Pathogenic effector cells Regulatory cells
Adap%ve immune responses
Transplanta%on tolerance to exploit exis%ng mechanisms to induce tolerance to allo-‐Ags
Exis?ng mechanisms to maintain tolerance to self-‐Ags
Immune tolerance
T
Immune sytem (Recipient)
Sensitized patient
Low immunological risk 1st Tx Graft
(Donor)
The immune response to an allogra?
Ag
APC cytotoxicity
complement
Inflammatory cytokines
B
mØ
anti-donor Abs (DSA) T
memory
Predictors of gra? outcome
T-‐cell-‐mediated cellular rejec%on An%body-‐mediated rejec%on
• Ideal treatment
Short-‐term administra?on
Targeted minimal immunosuppression
-‐ long-‐las?ng effect
-‐ Protec?on against cellular and humoral rejec?on
-‐ No/liWle side-‐effects
Central tolerance
Thymic educa?on of developing T cells
Selec%on
Bone marrow
immature thymocytes (cortex)
progenitor cells
mature thymocytes (medulla)
mature T cells
Delete the dangerous
high affinity poten%ally autoreac%ve
Posi%ve selec%on
Nega%ve selec%on
Thymic selec%on
Th0
THYMUS PERIPHERY
Host
Delete the dangerous
high affinity poten%ally alloreac%ve
Donor BM / HSC
Donor APC
Mixte chimerism
« Re-‐educa%on of the immune system »
HLA-Mismatched Renal Transplantation without Maintenance Immunosuppression
Kawai T, Cosimi AB, Spitzer TR, Tolkoff-Rubin N, Suthanthiran M, Saidman SL, Shaffer J, Preffer FI, Ding R, Sharma V, Fishman JA, Dey B, Ko D, Hertl M, Goes NB, Wong W, Williams WW,
Colvin RB, Sykes M, and Sachs D SUMMARY
Five patients with ESRD disease received combined bone marrow and kidney transplants from HLA single-haplotype mismatched living related donors, with the use of a non-myeloablative preparative regimen. Transient chimerism and reversible capillary leak syndrome developed in all recipients. Irreversible humoral
rejection occurred in one patient. In the other four recipients, it was possible to discontinue all immunosuppressive therapy 9 to 14 months after the transplantation, and renal function has remained stable for 2.0 to 5.3 years since transplantation. The T cells from these four recipients, tested in vitro, showed donor-specific unresponsiveness and in specimens from allograft biopsies, obtained after withdrawal of immunosuppressive therapy, there were high levels of FOXP3 but not granzyme B mRNA.
NEJM 2008;358:353-‐361
Mixed-‐chimerism induc%on In solid organ Tx
CsA (day -1 to d+60–90)
Cyclophosphamide (2x60 mg/kg d-5, d-4) ATG (3–4 x15–30 mg/kg) TI (7 Gy)
Donor BM Tx
HLA-identical
Multiple myeloma d0
Kidney Tx
Buhler et al., Transplanta/on 2002
Cyclophosphamide (2x60 mg/kg d-5, d-4) Rituximab (2x375 mg/m2 d-7, d-2) anti-CD2 mAb (MEDI507 d-1, d0, d+1) TI (7 Gy d-1)
Prednisone 2 mg/kg d0 tapering until d+10
HLA-mismatched
No malignancy
CsA (day -1 to 9 mths)
Kawai et al., NEJM 2008
Massachusetts General Hospital & the ITN / NIH
20 studies with search: “transplantation tolerance” and “chimerism” Rank Status Study
1 Active, not recruiting Reducing the Risk of Transplant Rejection: Simultaneous Kidney and Bone Marrow Transplant
2 Active, not recruiting Bone Marrow Transplant to Induce Tolerance in Kidney Transplant Recipients From Living Donors /deceased donors
4 Recruiting Combined Kidney and Bone Marrow Transplantation to Prevent Kidney Transplant Rejection
8 Recruiting HLA-Identical Sibling Renal Transplant Tolerance With Donor Hematopoietic Stem Cells and Campath-1H
11 Completed Combined Bone Marrow and Kidney Transplant for Multiple Myeloma With Kidney Failure
14 Active, not recruiting Islet Cell Transplantation Alone and CD34+ Enriched Donor Bone Marrow Cell Infusion in Patients With Type 1 Diabetes Mellitus; Steroid Free Regimen
Recruiting Autoimmune diseases (Rheumatoid arthritis, multiple sclerosis), hemopathies
Ongoing clinical trials
DC
Graft
Teff
Treg
The alloresponse – Rejec%on/tolerance
Muller/Golshayan. 2011
« Control » of the alloreac?ve Teff cells a>er Tx Regulatory T cells Regulatory B cells Tolerogenic DC Mesenchymal stem cells Cytokines
Expansion and func?on of Teff pool
Treg frequency and func?on
Teff Treg
Balance between dele%on and regula%on
Peripheral tolerance
THYMUS
CD4+CD25-Foxp3-
PERIPHERY
DC Pathogenic T cells
Foxp3-
IL-4
IL-12
TGF-β+IL-6/IL-21 (danger signals)
IL-23
Th1
Th2
Th17
Th0
ROR-γt
GATA-3
T-bet
Teff
CD4+CD25+Foxp3+
Regulatory T cells
Foxp3+
TGF-β
RA (CD103+DC) - IL-2 Foxp3 Treg
Golshayan and Pascual ,Trends Transplant 2009
Differen%a%on of T cells
Preven?on of acute and chronic rejec?on Induc?on of donor-‐specific Tx tolerance in lymphopenic hosts
Golshayan et al., Blood 2007. Joffre et al., Nat Med 2008. Tsang et al., J Clin Invest 2008 Preven?on of Tx vasculopathy/rejec?on by ex vivo-‐expanded human nTreg
Nadig et al., Nat Med 2010
Transfer of ex-‐vivo expanded Treg
Objec%ves: Prevent immunological rejec?on of Tx organs without the need for long-‐term use of pharmacological immunosuppression Clinical trials in living donor renal Tx recipients -‐ Source: peripheral vs. cord blood -‐ GMP facili?es -‐ purity, numbers (expansion methods) -‐ know/select donor Ag in advance for specificty
Treg in the clinic
Inflamma%on and stability of Treg
Ischemia/reperfusion Gra? injury Systemic infec%ons (virus)
Jaeger and Kuchroo. Scand J Immunol 2010
Wekerle and Sykes, 2004
Combining applicable strategies for the clinic
transient
Combining mixed chimerism and Treg transfer
BL/6
Balb/c full thickness tail skin
BL/6 d0
2x107 Balb/c BM cells i.v
6 weeks later Rapa (0.1 mg d-1, d0, d+2)
Anti-CD154 mAb (1mg d0)
CTLA-4 Ig (0.5 mg d+2)
3-4x106 recipient-derived Treg
with Tregs
no Tregs
% chimerism
with Tregs
no Tregs
3rd party Tx +Tregs
3rd party Tx
% gra? survival
Tolerant recipients exhibit unique expression pa\erns compared with stable recipient under chronic immunosuppression
ITN and Indices of Tolerance/RISET study groups
To combine biomarkers and clinical data to decide on IS weaning -‐ Transcriptome -‐ miRNA -‐ flow cytometry of peripheral blood -‐ func?onal cell-‐based assays
RT-PCR gene expression analyses of urine sedimentary cells
Newell et al., and Sagoo et al., J Clin Invest 2010
Iden%fica%on of a B cell signature associated with tolerance in renal Tx recipients
FACS analyses of PBMC
Thaunat et al., Blood 2010
Intragra? ter%ary germinal centers CD20+CD27+ cellular rejec%on
Effector and regulatory B cells
Am J Transplant.2010;10(9):1970-‐80
CD20+
B cell « help » to maintain optimal memory T
B cells help alloreac?ve T-‐cell differen?a?on, prolifera?on and survival to generate op?mal numbers of func?onal TM
Thaunat et al., Blood 2010
Intragra? ter%ary germinal centers CD20+CD27+ cellular rejec%on
Effector and regulatory B cells
Mesenchymal stem cells (MSC)
Origin bone marrow niche adherant fibroblast-‐like cells (CD45-‐CD31-‐CD105+CD44+) Self-‐renewal & plas%city home to injured ?ssues and favour regenera?ve processes preven?on of ischemia/reperfusion injury Immunoregulatory
Immunogenicity ? MSC express MHC I
Galipeau J et al. MiSOT consor?um
MSC in Tx
Treatment of GVHD post BM Tx
- universal donor strategy (Osiris therapeutics)
no benefit in phase 3 clinical trial
- allogeneic/autologous MSC beneficial in an acute setting (severe GVHD, tissue repair post infarct) or for long-term
Treatment of severe acute GVHD with 3rd-party haploidentical MSC
Le Blanc et al., Lancet 2004
The immune response throughout the life of a Tx
Balance Teff/Treg in vivo Most experimental studies done in lymphopenic hosts Strength of the alloresponse (early direct pathway) Innate immune ac?va?on (local inflammatory events / infec?ons) Cross-‐reac?ve memory T cells
Limita%ons in clinical solid organ Tx
APC
MHC:Ag
T cell
B7
TCR
CD28
Cytokines TNF-, IL-12 proIL-1β
NF-κB
TLR
MyD88
TRAF6 IKK β
IκB NF-κB
IRAK
MAPK NLR Inflammasomes
TLR
Endosome
Innate immune receptors: sensing « danger »
Gra? injury Infec%ons
Difficul?es to translate robust tolerance inducing protocols from rodents to large animals and humans -‐ Memory T cells account for a bigger propor?on of the T-‐cell repertoire in larger animals and humans
Exposure to environmental an?gens and pathogens Decreased func?on of the thymus with age, change in the propor?ons of naïve vs. memory T-‐cell pools
-‐ Cross-‐reac?vity between alloreac?ve and pathogen-‐specific T cells (heterologous immunity)
-‐ Differen?al suscep?bility to immunosuppressive drugs (cos?mula?on) than naïve T cells
- TM are more resistant to Treg-mediated regulation -‐ Homeosta?c expansion in lymphopenic hosts
Wu et al. Nat Med 2004. Adams et al. J Clin Invest 2003
Memory T cells : a barrier to Tx tolerance
Turka LA and lechler RI. Nat Rev Immunol 2009
Perspec%ves for clinical Tx
Billingham, Brent, Medawar. Nature 1953
Fondation Medi-CAL Futur
Tx Immunopathology Lab & Co
Lerisa Govender Dr Jean-‐Christophe Wyss Rajesh Kumar Pr Manuel Pascual
Many thanks to: Pr L Buhler, Pr JD Seebach, HUG Pr M Thome, UNIL Pr T Wekerle, Austria Pr RI Lechler, UK
… of Mice and Men …
Rationale of using an induction therapy
T-cell depletion/modulation at the time of Tx and antigen presentation
- High precursor frequency of alloreactive T cells
- I/R injury potentiates alloantigen presentation and immune activation
promotion of tolerance
Donor APC
Parenchymal cells
Donor-reactive T cells
t after Tx
v v
v
v
v
v
v
danger signals
Maintenance of tolerance and inflamma%on
IL-‐2c+MR1 LPS 1x s.c
>d60
- A large fraction of peripheral T cells in an individual can react with an allo-MHC (cross-reactive clones) - Higher affinity for allo-MHC than for the positive-selecting self-MHC
High precursor frequency of alloreactive T cells
Peripheral T cells 1% alloreactive <0.0001% specific for a nominal Ag presented by self-MHC
Reiser et al. Nat Immunol 2000
donor-reactive T cells
t after Tx
v
v
v
v
v
v
v
Strength of the direct pathway early immune response
20 studies with search: “transplanta?on tolerance” and “chimerism”
Etudes cliniues en cours
• Completed Reducing the Risk of Transplant Rejection: Simultaneous Kidney and BM Transplant
• Recruiting Renal Allograft Tolerance Through Mixed Chimerism
• Active, not recruiting Induction of Donor Specific Tolerance in Recipients of Live Donor Kidney Allografts by Donor Stem Cell Infusion • Active, not recruiting Combined Kidney and BM Transplantation to Prevent Kidney Transplant Rejection • Recruiting Delayed Tolerance in Recipients of Living Kidney Allografts by Donor Stem Cell Infusion • Completed Induction of Donor Specific Tolerance in Recipients of Cardiac Allografts by Donor Stem Cell Infusion • Recruiting HLA-Identical Sibling Renal Transplant Tolerance • Completed Combined BM and Kidney Transplant for Multiple Myeloma With Kidney Failure • Completed Islet cell transplantation alone and CD34+ enriched donor BM cell infusion in patients with type I
diabetes, steroids free
APC
Naïve T
Treg
IL-‐10 TGF-‐β IL-‐35
IL-‐2 IL-‐35
CTLA-‐4 Galec%n-‐1
Perforin Granzyme
Inhibi%on of clonal expansion
modula%on of APC
Inhibi%on of effector func%on of adap%ve and innate cells
B mø
TCR
Cos%mul
IDO
Foxp3
Peripheral tolerance
-‐ Naturally occuring Treg (nTreg) -‐ Induced Treg (iTreg) (by in vitro or in vivo manipula?on) Tr1 IL-‐10+ Foxp3+ iTreg CD8+CD28+ CD3+DN