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

Impact of C3 on transplant

• utcome

Steven Sacks MRC Centre for Transplantation Kings College London

Ancient origin of C3

Fujita, Nature Rev. Immunol. 2002

• 450m
• 350m

..well conserved thiol group

C3 C3

C3a C3a

C3b C3b C = O S C = O NH SH

Activating surface Activating surface

APC APC

Antimicrobial effects

C3b

Bacterium Bacterium C3

C3a

MACRO

OPSONISATION OPSONISATION

C3b

Bacterium Bacterium C3

C3a

PMNL

INFLAMMATION & IMMUNITY INFLAMMATION & IMMUNITY

C3b

Bacterium Bacterium C3

C5b-9

C5,6,7,8,9

C3a

LYSIS LYSIS

C5a

C3b C5b iC3b

C3d

Protection of host cells

C3b

Host cell Host cell INACTIVATION OF C3 INACTIVATION OF C3

Factor I

Host cell Host cell

C5 C5b-9

C6,7,8,9

FAILURE TO INACTIVATE C3 FAILURE TO INACTIVATE C3

C5a

PMNL

Regulator of complement

Enhanced activity of C3 at times around transplantation

Donor Transplanted organ Donor organ 300x 1000x

Tliney - rat Naesens 2009 Dammans 2010

Ischaemia reperfusion injury

C3 mRNA Complement Tubule necrosis

5 6 7 8 9 1 0 11 12 50 10 0 15 0 W T - W T (n=6 ) C 3d e f - W T (n =6)

Native nephrectomy ** p < 0 .0 5 * p < 0 .0 5 Day post-transplantation

Creatinine ( µmol/L)

Farrar, FASEB J 2006

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

Acute allograft rejection

7 50 100 C3+/+ donor (n=10) 8 10 12 14 C3-/- donor (n=10) 20 40 60 80 100 Days after transplantation % Animal Survival

B6 donor kidney into B10.BR recipients* or BALB/c recipients+

*Nat Med 2002; +Lin 2006

T cell priming

2 4 6 8 10 *** 3H TdR (cpm x 10

4)

5 10 15 20 25 WT C5aR-/- ** Donor strain DC IFN-γ (ng/ml)

Dendritic cell with C3 and MHC II T cell proliferation and cytokine release

J Immunol 2006 Blood 2006 & 2009 Mol Immunol 2011

C3a and C5a as immune stimulants

C3

C3a, C5a Antigen-presenting cell of the donor

C3aR/C5aR

Mouse: J Immunol 2006 Blood 2006, 2009 *Lalli, Blood 2008 Human: Mol Immunol 2011

• 1. Secretion of

complement components by activated APC

• 1. Secretion of

complement components by activated APC

• 2. Complement

activation

• 2. Complement

activation

• 3. C3aR and C5aR

signalling increases IL-12, MHC II, CD40, CD80

• 3. C3aR and C5aR

signalling increases IL-12, MHC II, CD40, CD80

• 4. Th1 activation/

expansion/survival and cytokine release*

• 4. Th1 activation/

expansion/survival and cytokine release*

C3 is essential for normal T cell priming through the action of C3a and C5a on antigen presentation C3 is essential for normal T cell priming through the action of 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 tissue and lowers the threshold for B cell stimulation Opsonisation of antigen with C3b leads to retention in lymphoid 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 Recipient

DAF C3 or receptor CR1 analogue C3a or C5a Anti-C5

Donor organ

Gene silencing

Reducing the expression of C3 mRNA in renal tissue reduces

• rgan susceptibility to warm

ischaemia reperfusion injury and offers a new therapeutic

• ption

Zheng, Transplantation 2006

Organ painting

Epithelial sheet

Native CR1 Cloned fragment Tailed Membrane- inserted

C3b

Effect of organ painting in rat

Reduces post-transplant acute renal failure Reduces chronic vascular damage to 20wks Donor organs are less immunogenic at 2wks Increases organ viability after prolonged ischaemia

Patel JASN 2006

10 20 30 20 40 60 80 100 A PT 07 0 (n= 22) C

• ntrol agent (n=

19) p = .0 3 63.6% 26.3%

D ays post tx

% Graft survival

Days post transplant

Recovery after 16h cold ischaemia

Pilot study in man

Donor organ treatment with Mirococept (APT070) or control No safety issues (10mg dose) No antibodies against APT070 Small amounts of serum APT070 No systemic inhibition of complement (C3, C4, CH50)

n=2 n=2 n=12 n=4

Baseline-adjusted creatinine AUC (d1-14) Molecular Immunology 2007

Next Step: EMPIRIKAL trial

Efficacy of Mirococept for Preventing Ischaemia-Reperfusion Injury in Kidney Allografts 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: What triggers C3 at the time of transplantation? Looking for new targets: What triggers C3 at the time of transplantation?

Recognition molecules

C1q C1r C1s

Antibody

Cell surface

MBL/ficolin MASP1 MASP2

Mannose

Cell surface

C4 C4 C2 C2 C3 C3

Classical pathway Lectin pathway

Antigen

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 at the time of kidney transplantation Lectin molecules may be one of the first triggers to activate C3 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 W Zhou A Farrar R Smith N Sheerin Q Peng W Shwaeble E Asgari K Brown V Binda Clinical trial G Koffman K Smith C Watson M Drage I Rebollo-Mesa Funders Wellcome Trust Medical Research Council Kidney Patients Association