Academic Trainees Meeting – 5 th May, 2011

Academic Trainees Meeting – 5th May, 2011

Interesting aspects of complement regulation……

Matthew Pickering

Wellcome Trust Senior Fellow in Clinical ScienceConsultant Rheumatologist

Complement activation protein deficiency

Terminal pathway

Recurrent Neisseria infections

C3

Infection

Classical pathway

SLE-like illness

Recurrent infection with encapsulated bacteria e.g. pneumococci, Haemophilus influenzae

Vasculitis, glomerulonephritis

Complement dysregulation

Terminal pathway dysregulation

Paroxysmal nocturnal haemoglobinuria

C1 inhibitor deficiency

[classical pathway dysregulation]

Hereditary angioedema

Alternative pathway dysregulation

Dense deposit disease,

Atypical haemolytic uraemic syndrome

renal thrombotic microangiopathy

Disorders of complement

Activation protein deficiency

‘too little’ complement

Tell us what might happen if we

therapeutically inhibit complement

Regulatory protein deficiency

‘too much’ complement’

Provide diseases in which complement inhibiting therapies

ought to be effective

C3b

C3

C3b C3b C3b

FOREIGN SURFACE

C3b

‘C3b amplification loop’

Complement activation

lectin pathway

Bacterial Carbohydrate, ficolins

C4b2a

classical pathway

immune complexes

C3bBb

alternative pathway

‘always on’

C5 activation

MAC

C5a

MAC = membrane attack complex

C3b

C3

Complement regulation

lectin pathway

C4b2a

classical pathway

C3bBb

alternative pathway

MAC = membrane attack complex

iC3b C3b

CR1CD46CD59

MAC

Factor I

DAF(CD55)

C1 inhibitor

C4bp C1 inhibitor Factor H

Factor I

iC3b

Factor H

Factor H

Physiological control of complement activation

REGULATORS

Complement dysregulation and disease:

ACTIVATORS

Loss of function Gain of function

The balance is influenced by mutations (extreme) and and/or polymorphisms (‘fine tuning’)

What does factor H do? Critical negative regulator of the alternative pathway and C3b

amplification loop

What happens to C3 levels in individuals with complete genetic deficiency of CFH?

Uncontrolled spontaneous activation of the alternative pathway and secondary consumption of C3

Why is factor H important? It is associated with human disease:

Dense deposit disease

mutations

rare

‘protective’ and ‘at risk’polymorphisms

common

Dense deposit disease Electron-dense transformation of the glomerular basement

membrane

Glomerular C3 staining in DDD

DDD retinopathy

Dense deposit disease

C3b

C3

C3bBb

Factor H

B, D

C3 nephritic factor

Anti-factor H

Associated with plasma C3 activation:

Dense deposit disease Animal models:

Spontaneous porcine factor H deficiency and gene-targeted factor H-deficient mice

Profound plasma C3 depletion – 5% of normal C3 levels

Spontaneous renal disease – ‘murine/porcine DDD’

Factor H deficiency

Wild-type

C3 staining wild-type

0

200

400

600

Cfh-/-

Pla

sma

C3

- m

g/l

Dense deposit disease What have the animal models taught us?

The renal disease does not develop if activation of C3 is blocked

The renal disease does develop if C5 activation is blocked Dense deposits still develop Glomerular inflammation reduced but not absent

Murine dense deposit disease is dependent on the ability to activate C3 but not C5

Glomerular basement membrane deposits in mice with combined

deficiency of factor H and C5

Pickering MC, et al. PNAS 2006 103(25):9649-54.

Human complement deficiency

DeficiencyState:

Plasma C3:

Factor I

low absent

C3

Associations: Recurrent infection

immune complex-mediated renal diseasee.g. MPGN type I

Factor H

low

Dense depositdisease

C3b iC3b, C3d

Pickering MC, Cook HT. Clin Exp Immunol. 2008 51(2):210-30.

Plasma C3 regulation Continuous activation of C3 occurs in plasma through

the C3 ‘tick-over’pathway

C3b

C3

C3bBb

Factor H

C3b

Factor I

iC3b

C3dC3c

Factor BFactor D

Dense deposit disease Administration of factor I to mice with combined deficiency

of H and I restores GBM C3 staining

0 24 48 72

0

50

100

150

200

hours

Pla

sma

C3

leve

ls (

mg

/l)

injections

Rose KL et al. J Clin Invest. 2008 118(2):608-18.

Why is factor H important? It is associated with human disease:

Dense deposit diseaseAtypical haemolytic uraemic syndrome

mutations

rare

‘protective’ and ‘at risk’polymorphisms

common

Atypical haemolytic uraemic syndrome

Atypical Haemolytic uraemic syndrome

renal thrombotic microangiopathy

Alternative pathway dysregulation

Associated with:

COMPLEMENT MUTATIONS

Loss of function mutations in regulators

• Factor H • Mutations• Hybrid gene (copy number variation)

• Factor I• MCP (CD46)

Gain of function mutations in activation proteins

• C3• Factor B

ACQUIRED COMPLEMENT DYSREGULATION

Anti-factor H autoantibodies

Atypical Haemolytic uraemic syndrome – factor H mutations

Factor I

iC3b

C3b

C3

B, D

C3bBb

HOST SURFACERENAL ENDOTHELIUM

C3b C3b C3b C3b C3b

C5 activation

MAC C5a

CD46

C3 regulation

Surface recognition

Murine model of factor H-associated atypical haemolytic uraemic syndrome

Gene-targeted factor H-deficient mice transgenically expressing a mutant mouse factor H protein (FH16-20)

Cfh-/-FH16-20Cfh-/-

0

25

50

75

100

Pla

sma

C3

- m

g/l

wild-type mouse CFH Mutated mouse FH16-20

Renal histology in Cfh-/-.FH16-20

Murine model of factor H-associated atypical haemolytic uraemic syndrome

Use this model to determine contribution of C5 activation to renal injury

Spontaneous renal disease does not occur in C5-deficient Cfh-/-FH16-20 animals

Murine model of factor H-associated atypical haemolytic uraemic syndrome

Cfh-/-FH16-20 animals are hypersensitive to experimentally triggered renal injury – this injurious response is C5 dependent

C3

C9

Atypical haemolytic uraemic syndrome - therapy

C5 inhibition successful in case reports – examples: Eculizumab for aHUS – N. Engl. J. Med. 2009 360:5 pp542-543 Eculizumab for congenital aHUS – N. Engl. J. Med. 2009 360:5 pp544-6

Open Label Controlled Trial of Eculizumab in Adult Patients With Plasma Therapy-sensitive / -resistant Atypical Hemolytic Uremic Syndrome (aHUS)

Successful outcomes announced in ASN 2010 meeting http://clinicaltrials.gov/ct2/results?term=eculizumab

Why is factor H important? It is associated with human disease:

Dense deposit diseaseAtypical haemolytic uraemic syndrome

mutations

rare

‘protective’ and ‘at risk’polymorphisms

common

Factor H and Age-related macular degeneration

Factor H and AMD – the ‘Y402H’ polymorphism

From Sofat et al., Atherosclerosis 213 (2010) 184-90

Alternative pathway dysregulation

Age-related macular degeneration

Factor H and Age-related macular degeneration

Ocular drusen

Associated with:

Polymorphic variants in:Regulators• Factor H Y402H ‘at risk’

V62I ‘protective’

activation proteins• C3 C3FF ‘at risk’

• Factor BBf32Q ‘protective’

Age-related macular degeneration

Functional differences inthe Valine62Isoleucine CFHpolymorphism

62Isoleucine more efficient at preventing red cell lysis

14nM vs. 22.6nM at 50% lysis

62Valine

62Isoleucine

Factor H and Age-related macular degeneration

Age-related macular degeneration

Complement dysregulation and eye disease – age-related macular degeneration

Dense deposit disease

mutations

alternative pathway activation

Ocular drusen

‘at risk’polymorphisms

‘protective’polymorphisms

DDD retinopathy

Factor H 402H*Factor H 62VFactor B 32R

C3F

Factor H 402Y*Factor H 62IFactor B 32Q

C3SCFHR1/3 deletion*

*functional consequences not understood

Factor H null allelesC3 3923∆DG

Why is factor H important? It is associated with human disease:

Dense deposit diseaseAtypical haemolytic uraemic syndrome

mutations

rare

‘protective’ and ‘at risk’polymorphisms

common

Age-related macular degeneration Meningococcal sepsis

Factor H and susceptibility to meningococcal infection

Meningococcal sepsis

The factor H family

Why are the factor H-related proteins important? They are associated with human disease:

mutations

rare

‘protective’ and ‘at risk’polymorphisms

common

The factor H family: copy number variation

CFH CFHR3 CFHR1 CFHR4 CFHR2 CFHR5

Most frequent CFH-CFHR allele

Deletion homozygotes: African American 16%Hageman et al, Ann. Medicine 2006 European Americans 4.7%

CFH CFHR4 CFHR2 CFHR5

CFHR1-3 deletion allele polymorphism (common)

Others (uncommon - <1%)

CFH CFHR1 CFHR4 CFHR2 CFHR5

CFH CFHR3 CFHR4 CFHR2 CFHR5

CFH CFHR3 CFHR2 CFHR5

CFH CFHR3 CFHR1 CFHR3 CFHR1 CFHR4 CFHR2 CFHR5

CFH CFHR3 CFHR1 CFHR1 CFHR4 CFHR2 CFHR5

Why are the factor H-related proteins important? They are associated with human disease:

‘protective’ and ‘at risk’polymorphisms

commonAge-related macular

degeneration

CFHR1-3 deletion allele polymorphism

associated with protection against

AMD

Mol Immunology 44 (2007):3921.

Complement therapeuticsPathologies in which

complement is activated

Complement therapeuticsExamples of the many complement inhibitors in development

Eric Wagner and Michael Frank Nature Reviews 2010, vol. 9, 43-56.

Thanks

Elena Goicoechea de Jorge Katherine Vernon Mitali Patel Kirsten Rose Talat Malik Sharmal Narayan Marieta Ruseva Tamara Montes Lola Sanchez-Nino

Danielle Paixao-Cavalcante Fadi Fakhouri Terence Cook Marina Botto Santiago Rodriguez de Cordoba Veronique Fremeaux -Bacchi Patrick Maxwell Danny Gale