Impact of C3 on transplant outcome Steven Sacks MRC Centre for - PowerPoint PPT Presentation

Impact of C3 on transplant outcome Steven Sacks MRC Centre for Transplantation King � s College London

Ancient origin of C3 -450m -350m Fujita, Nature Rev. Immunol. 2002

..well conserved thiol group C3a C3a C3 C3 S C = O C3b C3b SH C = O NH Activating surface Activating surface

Antimicrobial effects OPSONISATION LYSIS INFLAMMATION & IMMUNITY OPSONISATION LYSIS INFLAMMATION & IMMUNITY PMNL APC APC C3a C3a C3a C5a MACRO C3 C3 C3 C5,6,7,8,9 C3b C3b C3b C5b-9 Bacterium Bacterium Bacterium Bacterium Bacterium Bacterium

Protection of host cells INACTIVATION OF C3 FAILURE TO INACTIVATE C3 INACTIVATION OF C3 FAILURE TO INACTIVATE C3 PMNL Regulator of C5a complement C5 Factor I C6,7,8,9 C3b C3b C5b iC3b C3d C5b-9 Host cell Host cell Host cell Host cell

Enhanced activity of C3 at times around transplantation Donor Donor organ Transplanted organ 300x 1000x Tliney - rat Naesens 2009 Dammans 2010

Ischaemia reperfusion injury C3 mRNA Complement Tubule necrosis W T - W T (n=6 ) 15 0 C 3d e f - W T (n =6) * p < 0 .0 5 µ mol/L) ** p < 0 .0 5 10 0 Creatinine ( 50 0 5 6 7 8 9 1 0 11 12 Day post-transplantation Native nephrectomy Farrar, FASEB J 2006

Acute tubular injury is dependent on C5a and C5b-9, whose Acute tubular injury is dependent on C5a and C5b-9, whose production is fuelled by local synthesis of C3 mainly by tubule production is fuelled by local synthesis of C3 mainly by tubule epithelium epithelium

Acute allograft rejection 100 C3+/+ donor (n=10) % Animal Survival 50 C3-/- donor (n=10) 0 0 7 8 10 12 14 20 40 60 80 100 Days after transplantation B6 donor kidney into B10.BR recipients * or BALB/c recipients + *Nat Med 2002; + Lin 2006

T cell priming Dendritic cell with T cell proliferation and C3 and MHC II cytokine release 10 25 Donor strain DC 4 ) WT 8 3H TdR (cpm x 10 20 C5aR-/- IFN- γ (ng/ml) 6 15 ** 4 10 *** 2 5 0 0 J Immunol 2006 Blood 2006 & 2009 Mol Immunol 2011

C3a and C5a as immune stimulants 4. Th1 activation/ 4. Th1 activation/ Antigen-presenting expansion/survival and expansion/survival and cytokine release* cytokine release* cell of the donor 3. C3aR and C5aR 3. C3aR and C5aR signalling increases signalling increases 1. Secretion of 1. Secretion of C3aR/C5aR IL-12, MHC II, CD40, IL-12, MHC II, CD40, complement complement CD80 components by CD80 components by activated APC activated APC C3a, C5a Mouse: J Immunol 2006 C3 Blood 2006, 2009 2. Complement 2. Complement *Lalli, Blood 2008 activation activation Human: Mol Immunol 2011

C3 is essential for normal T cell priming through the action of C3 is essential for normal T cell priming through the action of C3a and C5a on antigen presentation C3a and C5a on antigen presentation

B cell priming H-2b mouse anti H-2k IgG response Marsh, Transplantation 2001

Opsonisation of antigen with C3b leads to retention in lymphoid Opsonisation of antigen with C3b leads to retention in lymphoid tissue and lowers the threshold for B cell stimulation tissue and lowers the threshold for B cell stimulation Fang 1998 Dempsey 1996

Evolution of late graft failure Contribution of C3

Long term effects of C3 in man � Peritubular C5b-9 correlates with chronic kidney disease � C3F/S polymorphism associates with chronic allograft nephropathy � Capillary C4d associates with chronic antibody mediated rejection

Chronic kidney injury Adriamycin nephropathy in BALB/c at 6 wks Wild type (C3) Wild type (HE) C3 deficient (HE) Sheerin, FASEBJ 2008

New opportunities for therapy created by understanding C3 Donor Donor organ Recipient DAF C3 or receptor CR1 analogue C3a or C5a Anti-C5

Gene silencing Reducing the expression of C3 mRNA in renal tissue reduces organ susceptibility to warm ischaemia reperfusion injury and offers a new therapeutic option Zheng, Transplantation 2006

Organ painting Native CR1 Cloned fragment Tailed Membrane- inserted C3b Epithelial sheet

Effect of organ painting in rat � Reduces post-transplant acute renal failure Recovery after 16h cold ischaemia 100 � Reduces chronic vascular A PT 07 0 (n= 22) C ontrol agent (n= 19) 80 damage to 20wks 63.6% p = 0 .0 3 % Graft survival 60 � Donor organs are less 40 26.3% immunogenic at 2wks 20 0 � Increases organ viability after 0 10 20 30 D ays post tx Days post transplant prolonged ischaemia Patel JASN 2006

Pilot study in man � Donor organ treatment with Mirococept (APT070) or control Baseline-adjusted creatinine AUC (d1-14) � No safety issues (10mg dose) � No antibodies against APT070 � Small amounts of serum APT070 n=2 n=2 n=12 n=4 � No systemic inhibition of complement (C3, C4, CH50) Molecular Immunology 2007

Next Step: EMPIRIKAL trial E fficacy of M irococept for P reventing I schaemia- R eperfusion I njury in K idney Al lografts � MRC funded 2011 � Double-blind randomized Phase II study of Mirococept v. perfusion control � 280 patients in each of treatment and control arms � Main endpoint: incidence of Delayed Graft Function � Adaptive design to explore dose-response over a range of 5 to 25mg � Multi-centre study in the UK � Likely to recruit first patients in 2012

Looking for new targets: Looking for new targets: What triggers C3 at the time of transplantation? What triggers C3 at the time of transplantation?

Recognition molecules Classical pathway Lectin pathway C1q MBL/ficolin C1r MASP1 MASP2 C1s C4 C4 Mannose C2 C2 Antibody Antigen C3 C3 Cell surface Cell surface

Impact of lectin pathway on post-ischaemic damage (Heart and bowel: PNAS 2011)

Lectin molecules may be one of the first triggers to activate C3 Lectin molecules may be one of the first triggers to activate C3 at the time of kidney transplantation at the time of kidney transplantation

Conclusions � C3 is a key target in preventing graft injury at the time of transplantation, with potential to increase graft longevity and widen the range of donor organs suitable for transplantation � Other targets such as the lectin pathway may become more relevant as we grow to understand what triggers complement activation following metabolic and immunological insult to the donor organ � Therapeutic targeting requires detailed knowledge of which complement pool contributes to a specific graft injury, in particular whether the pool is intravascular or extravascular

Thanks Lab studies Clinical trial � W Zhou � G Koffman � A Farrar � K Smith � R Smith � C Watson � N Sheerin � M Drage � Q Peng � I Rebollo-Mesa � W Shwaeble � E Asgari Funders � K Brown � Wellcome Trust � V Binda � Medical Research Council � Kidney Patients Association