Harnessing tolerance mechanisms with monoclonal antibodies Prof. Herman Waldmann

Transcription

1 Harnessing tolerance mechanisms Herman Waldmann Sir William Dunn School of Pathology, Oxford University, UK 1 1. Transplantation 2 Lymphocyte depletion strategies to minimise drug immunosuppression in transplantation Alemtuzumab The first humanised therapeutic antibody 3 The screen versions of these slides have full details of copyright and acknowledgements 1

2 Distribution of the CD52 antigen within the hemopoietic system Hemopoietic stem cells CD52+ Lymphocytes Basophil Neutrophil Monocyte Dendritic cells Eosinophil Platelets Erythroid Hale G, et al., The CAMPATH-1 antigen (CDw52), Tissue Antigens 1990; 35(3): % freedom from rejection Incidence of acute rejection on low dose CyA monotherapy 1 maintenance drug 80.7% 77.3% 69.5% 69.5% 3 maintenance drugs Years post-transplant Control: 69.5% Alemtuzumab: 66.4% Watson CJ et al., 2005, Alemtuzumab (CAMPATH 1H) induction therapy in cadaveric kidney transplantation -- efficacy and safety at five years, Am J Transplant, 5(6): Routes to transplantation tolerance Bone marrow Lymphocyte Monocyte Eosonophil Basophil Neutrophil Establishing mixed blood chimerism Tolerisation with an expendable cell source (e.g., blood transf usions) Tolerisation to the tissue itself 6 The screen versions of these slides have full details of copyright and acknowledgements 2

3 Achieving mixed chimerism Monoclonal antibodies to ablate T-cells from host and donor blood systems Additional drugs or irradiation to provide some myelosuppression and hemopoietic space Has proven straightforward to achieve in rodents Translation to humans harder, where even across minor H barriers sustained mixed chimerism is hard to achieve Buhler LH, et al., Transplantation 2002; 74(10): It has been common in tolerance research to achieve tolerance empirically, and to then work out mechanisms Such mechanisms are usually relevant to self-tolerance Short courses of Rx which blindfold T-cells of the immune system generate immunological tolerance to allogeneic skin transplants Waldmann, H., T. C. Chen, et al., (2006), Regulatory T cells in transplantation, Semin Immunol 18(2): Blockade of co-receptors and co-stimulatory molecules w i th monoclonal antibodies T-cell CD40L TCR CD4 CD40 MHCII Dendritic cell 9 The screen versions of these slides have full details of copyright and acknowledgements 3

4 Non depleting CD4 + CD8 antibodies promote transplant tolerance GRAFT B RECIPIE T A GRAFT C 10 Graft survival and tolerance Graft rejection Cells that regulate 11 Cells that can damage Features of dominant transplantation tolerance Resistance to breakdown by infusion of naive T-cells Tolerant T-cells can adoptively transfer of suppression to a naive mouse in an antigen-specific way Linked suppression 12 The screen versions of these slides have full details of copyright and acknowledgements 4

5 Tolerised mice show linked-suppression 100 % graft survival 50 C (+ B) (B X C) F Time (days) 120 CBA hosts tolerised to B10 skin Third party graft Balb/C 13 Infectious tolerance Dominant tolerance which is passed on from one cohort of T-cells to another as a result of cohabitation with antigen Qin, S., Cobbold, S.P. Pope, H., Elliott, J., Kioussis, D. & Waldmann, H. 1993, Infectious transplantation tolerance, Science 12; 259: Summary points 1. Regulatory T-cells can be found in tolerated grafts 2. TGFβ is involved in therapeutic tolerance 3. TGFβ can convert naive T-cells to regulatory cells that suppress graft rejection 4. Persistent innocuous low level signalling by antigen also induces regulatory T-cells 5. Regulatory T-cells generate privileged microenvironments 6. Transplantation tolerance may be possible if we can exploit this T reg -tissue interplay; Monoclonal antibodies are a useful tool for this purpose 15 The screen versions of these slides have full details of copyright and acknowledgements 5

6 1. Regulatory T-cells T are present in tolerated skin grafts Expand T-cells Tolerant Empty Deplete T-cells Tolerant Empty Expand T-cells % graft survival 100 Expanded T-cells 50 Depleted T-cells Expanded syngeneic skin Time (days) Tolerant Empty Naive cells Graca, L. et al., (2002), "Identification of regulatory T cells in tolerated allografts" J Exp Med, 195(12): T-cell receptor transgenic mice Mic e carrying just one type of TCR to just one transplantation antigen 17 Tolerance induction in a TCR transgenic mouse Mice transgenic f or a TCR directed towards a male peptide; Crossed onto a RAG -/- background Antigen is H-Y (male) as presented by H2-E k Antigen (Dby ) peptide is REEALHQFRSGRKPI Male mice hav e NO T-cells (thy mic deletion) Female T-cells are ALL TCR + CD4 + (monospecif ic) No f oxp3 expression in thy mus or periphery of normal f emale mice Female mice rapidly REJECT male skin graf ts CD4 or CD40L mabs can tolerise f emale mice to male skin (coreceptor or costimulation blockade) 18 The screen versions of these slides have full details of copyright and acknowledgements 6

7 FoxP3 +, CD4 + CD25 +, GITR + regulatory T-cells are found in tolerated skin grafts FoxP3/HPRT Tolerated 1st skin Tolerated 1st skin lym phs enriched T olerated 1st skin CD4+ T-cell s Tolerated 2nd skin Tol erated 2nd skin lym phs enriched Tolerated 2nd skin CD4+ T-cells 20% 49% CD4 Cobbold S et al., The Journal of Immunology, 2004, 172: CD25 19 Anti-CD4 blockade of TCR-transgenic CD4 + T-cells responding to Dby peptide induces foxp3 expression in vitro in a TGFβ dependent manner Proliferation Cobbold S et al., The Journal of Immunology, 2004, 172: RT-PCR TGFβ is needed in this form of therapeutic transplantation tolerance 21 The screen versions of these slides have full details of copyright and acknowledgements 7

8 The induction of skin graft tolerance by anti-cd CD4 is TGFβ but not IL-10 dependent Cobbold S et al., The Journal of Immunology, 2004, 172: Naive T-cells T exposed to antigen in i n the presence of TGFβ can be induced to become foxp3 + regulatory T-cells T These cells can be expanded in vitro Expanded TGFβ-induced Tregs are able to prevent graft rejection This provides a clue to one role for TGFβ in inducing regulatory 23 O v erriding tolerance checkpoints for rejection T-cells T reg Induction Expansion Homing and recirculation Tissue entry Damage 24 The screen versions of these slides have full details of copyright and acknowledgements 8

9 Disabled i T-cells T and checkpoints in tolerance T-cells T reg TGFβ Induction Expansion Homing and recirculation Tissue entry Damage 25 Successful override of tolerance checkpoints when TGFβ signalling is unavailable T-cells T reg Induction Expansion Homing and recirculation Tissue entry Damage Persistent exposure to antigen under incomplete signalling conditions favours tolerance and induction of regulatory T- T cells 27 The screen versions of these slides have full details of copyright and acknowledgements 9

10 An altered peptide ligand of the Dby-E k mh peptide Induces tolerance to male skin graf ts in f emale monospecif ic TCR transgenic mice Polarises CD4 + CD25 - T-cells to IL-10 producing Tr1 cells in vitro Induces CD4 + CD25 + FoxP3 + T-cells which are easily demonstrable in the skin graf t H Weak agonist Chen, T-C et al., J. Clin. Invest. 131: 1754 (2004) 28 Altered peptide ligands can induce foxp3 expression in peripheral T-cells T A1(M) x RAG-1 -/- thymus A1(M) x RAG-1 -/- spleen Skin elsewhere Tolerated skin Tolerant thymus Tolerant spleen Whole CBA/Ca spleen cells CD4 + CD25 - CD4 + CD Normalised foxp3 mrna expressi on Spleen 3% CD4 + CD25 + T-cells 29 Graft 15% CD4 + CD25 + T-cells Chen, T-C et al., J. Clin. Invest. 131: 1754 (2004) Natural privilege against immune damage is an established feature of a number of tissues such as the placenta and the anterior chamber of the eye 30 The screen versions of these slides have full details of copyright and acknowledgements 10

11 5.. Regulatory T-cells T and the concept of acquired privilege 31 S uppression by a regulatory T-cell T clone is dependent on IDO activity in certain types of dendritic cell CD11c+ DCs B220+ B220+ B220- CD8 α+ CD8 α+ CD8α- Treg A IDO+ DCs/Total DCs (%) IDO + DCs IDO - DCs D 250 IDO-WT CD11c+ IDO-KO IDO-WT CD11c- IDO-KO Dby K b Mellor et al., International Immunology, 16, , 2004 CPM (x10-3) % 50 x5 0 Treg Medium 1mT 10xTrp 32 T he notion of acquired immunological privilege Chemokines Regulatory Cytokines Cell death and damage Waldmann et al., Seminars in Immunology 16 (2004) The screen versions of these slides have full details of copyright and acknowledgements 11

12 Stages in the evolution of t ransplantation tolerance 1. Antibodies act as blindfolds to prevent rejection 2. Selection from natural T reg pool, and de novo polarisation of naive T-cells to T reg activity 3. T reg attracted to site of antigen and there take over the job of preventing rejection 4. Grafts heal (TGFβ) and antigens continue to be released and processed by host APC 34 Why is tolerance long term and infectious? 5. Those antigen-presenting-cells are quiescent 6. Many T-cells seeing antigen associated with quiescent APC are inactivated 7. Others are induced to become able to regulate (persistent danger-free antigen presentation) 8. New cohorts of T-cells exposed to these antigens are recruited to become regulatory (infectious tolerance) Autoimmune diseases 36 The screen versions of these slides have full details of copyright and acknowledgements 12

13 Blockade of TCR complex with CD3 m onoclonal antibodies T-cell CD40L CD40 MHCII TCR CD4 Dendritic cell 37 Mutation of a humanised antibody to diminish a m ajor side effect of cytokine release Mutation at Asp (297) prevents glycosylation Bolt et al., 1993, The generation of a humanised, non-mitogenic antibody that retains immunosuppressive activity in vitro, Eur. J. Immunol. 23; C hanges in residual beta-cell function C-peptide release after glucose challenge 0.15 P = P = 0.01 P = nm/min m 12 m 18 m ChAglyCD3 Placebo Keymeulen B, et al., Insulin needs after CD3-antibody therapy in new-onset type 1 diabetes N Engl J Med 2005; 352(25): The screen versions of these slides have full details of copyright and acknowledgements 13

14 Insulin needs IU/kg/day 0.70 P = P = P = ChAglyCD3 Placebo Baseline 6 m 12 m 18 m Keymeulen B, et al., Insulin needs after CD3-antibody therapy in new-onset type 1 diabetes, N Engl J Med 2005; 352(25): Lymphocyte depletion strategies in autoimmune disease Alemtuzumab The first humanised therapeutic antibody From Coles et al., J. Neurol (1): Epub 2005 Jul The screen versions of these slides have full details of copyright and acknowledgements 14

15 Speculations 43 Why regulation at all? Tissues should be permissive of appropriate immune responses to microbes without risk of irreversable tissue damage Regulatory mechanisms may have co-evolved with adaptive immunity to cooperate with tissues to ensure this 44 Mechanisms of regulation Immunity Suppression/regulation T-cell T-cell Cells of the innate immune system Cells of the innate immune system 45 The screen versions of these slides have full details of copyright and acknowledgements 15

16 46 The screen versions of these slides have full details of copyright and acknowledgements 16