1. Uptodate to C3 Glomerulopathies
Dott. Aris Tsalouchos (Medico specializzando in Nefrologia). Università degli Studi di Firenze.

6. MPGN type III of Burkholder
(type IIIB)
MPGN type III of Strife/Anders (type IIIS/A)

7. Historical Classification of MPGN

13. Criterion 1: C3 only (%)
Criterion 2: C3 dominant and up to 1+ IgM only (%)
Criterion 3: C3 dominant and ≥ 2 orders of intensity greater than any combination of IgG, IgM, IgA, and C1q (%)

14. ‘C3 only’ is an impractical definition of C3 glomerulopathy, and we propose a definition of C3 dominant and at least two orders of magnitude more intense than any other immune reactant, which requires validation by alternative pathway evaluation.
These criteria provide a framework for identifying patients most likely to benefit from investigations of alternative complement pathway dysregulation.

16. Activation of the Alternative Pathway of Complement System
C3 “tick-over”

17. Activation of the Alternative Pathway of Complement System
“Amplification loop”

19. Late steps of complement activation and formation of the MAC.
Cell-associated C5 convertase cleaves C5 and generates C5b, which becomes bound to the convertase. C6 and
C7 bind sequentially, and the C5b,6,7 complex inserts into the plasma membrane, followed by insertion of
C8. Up to 15 C9 molecules may then polymerize around the complex to form the MAC, which creates pores
in the membrane and induces cell lysis. C5a released on proteolysis of C5 stimulates inflammation.

20. Regulation of the Alternative Pathway.
Plasma regulators of the AP of complement:
CFI
Properdin (the sole positive regulator factor)
CFH, CFHL1
Complement Factor H Related proteins (CFHR1 , CFHR2, CFHR3, CFHR4, CFHR5)
Membrane complement regulatory proteins:
Complement receptor type 1 (CR1, CD35)
Membrane cofactor protein (MCP, CD46)
Decay-accelerating factor (DAF, CD55)
Protectin (CD59)

21. Mechanisms of complement regulation by Membrane Complement Regulatory Proteins
DAF or CR1 destabilizes C3 convertases and
accelerates the dissociation of C3bBb (depicted).
Cofactor activity: MCP or CR1 binds to
C3b and serves as a cofactor for CFI-mediated cleavage and inactivation of C3b

22. Complement Factor H (CFH)
Front Immunol. 2013 Apr 23;4:93. doi: /fimmu eCollection 2013.

23. Activities of Complement Factor H (CFH)

24. Factor H-like protein 1 (CFHL1)
Factor H-like protein 1 (CFHL1) is a 43 -kDa protein that derives from an alternative
splice product of the CFH gene. It shares the seven N -terminal SCRs with factor H and has four additional amino acids. Due to the broad sequence overlapping, CFHL1 possesses the same complement regulatory activities mediated by the N-terminus of factor H.
Trends Immunol. 2008 Aug;29(8):380-7.

25. Complement factor H related proteins (CFHRs)

26. Functions of the CFHR proteins
The CFHR3 and CFHR4 proteins have been shown to enhance complement inhibitory activity of CFH, but alone lack regulatory activity
CFHR1 reduced the regulatory activity of CFH at the surface
 Because CFHR proteins also have significant differences in sequence and domain structure, they may possess so far undiscovered biological activities, which are unrelated to the complement system.
Trends Immunol. 2008 Aug;29(8):380-7.

27. Pathogenesis of C3 glomerulopathy

28. Genetic causes of C3 glomerulopathy.
A unique Spanish pedigree was identified that associates C3G-DDD with a
heterozygous mutation in the C3 causing the deletion of two amino acid residues (C3-923delDG)
Generates an active C3-convertase that is:
- normally regulated by DAF on the surface of endothelial cells
- resistant to decay by factor H in the plasma
Conclusion: fluid phase-restricted dysregulation of the AP

29. Genetic causes of C3 glomerulopathy.
CFH/CFHL1

30. Copy number variations in the CFHR gene cluster in patients with C3 glomerulopathy.

31. Autoimmune forms of C3 glomerulopathy C3 nephritic factor (C3 Nef)
C3Nefs are IgG or IgM antibodies that bind to the C3bBb convertase .
Stabilization of the C3bBb convertase by C3Nef results in higher and prolonged activity, C3 and C5 consumption and activation of the terminal pathway.
C3Nef has been detected in ∼50% in the cases of MPGN I and MPGN
III, and in up to 80% of MPGN II/DDD patients.
Also identified in patients with antiphospholipid syndrome and even in healthy individuals.
Immunology Letters (2014) 163–171

32. reported a frequency of ∼20% for C4Nef alone or together with C3Nef.
The only study analyzing a cohort of 100 hypocomplementemic MPGN patients
reported a frequency of ∼20% for C4Nef alone or together with C3Nef.
Screening for C4Nef is currently not part of the routine diagnostics.
It is unclear at present if C4Nefs are common and if they are relevant to disease. Therefore, it would be worthwhile to clarify the frequency and relevance of C4Nef.

33. Autoimmune forms of C3 glomerulopathy Factor B and C3b autoantibodies
Autoantibodies targeting the individual components of the C3bBb convertase, C3b and factor B were also described
At present, one patient with factor B autoantibody and two patients with combined factor B and C3b autoantibodies have been described in detail.
Anti-factor C3b
Thorough functional characterization of the factor B autoantibody revealed stabilization of the C3bBb convertase against both intrinsic and factor H mediated decay. The C3b antibodies were not further characterized.
Mol Immunol 2010;47:1476–83
N Engl J Med 2011;365:2340–2

34. Autoimmune forms of C3 glomerulopathy Factor H autoantibodies
The first factor H autoantibody described was a “mini-autoantibody” consisting of lambda light chain dimers the binding site of this antibody was
shown to be in domain 3 of factor H
One patient diagnosed with DDD had monoclonal gammopathy and autoantibodies against factor H and CFHL1
In a recent study, 10 of 32 patients with C3 glomerulonephritis had
evidence of monoclonal gammopathy associated with complement activation and the authors suggested that monoclonal immunoglobulins acted as autoantibodies to complement-regulatory proteins. This potential association should be investigated in the future.

35. EPIDEMIOLOGY OF C3 GLOMERULOPATHIES.
INCIDENCE/PREVALENCE
Estimated incidence at 1 to 2 per million population per year with a reported prevalence of 2 to 3 per 1 million population
In one large series from France, the ratio of C3GN to DDD was about 2 : 1.
AGE OF ONSET
DDD is primarily a disease of children and young adults.
Patients with C3GN are significantly older, with a mean age at diagnosis of 30 years.
GENDER
Affects males and females equally in many cohorts, although some studies have shown a female predominance.
GEOGRAPHIC VARIATION
Mutation in CFHR5 in which almost all the cases have been found in Cyprus.
Clin J Am So c Nephrol. 2009;4:22-32
Kidney Int. 2012;82:
Clin J Am Soc Nephrol. 2014;9:46-53
Proc Natl Acad Sci USA. 2013;110:

36. Clinical manifestations
Dense Deposit Disease
At presentation, almost all patients with DDD have proteinuria, usually with hematuria.
Nephrotic-range proteinuria is present in two thirds of patients, and frank nephrotic syndrome in 12% to 65% in different series.
A number of patients have initial signs and symptoms of acute nephritic syndrome and may have episodes of acute kidney injury that show complete clinical resolution.
In about half of patients, clinical onset of DDD is preceded by acute infection, often an upper respiratory tract infection.
COMPREHENSIVE CLINICAL NEPHROLOGY Copyright © 2015

37. Extra-renal manifestations
Dense Deposit Disease
Patients with DDD may develop ocular drusen, lipoproteinaceous deposits of complement-containing debris within the Bruch membrane beneath the retinal pigment epithelium
This pathology is similar to age-related macular degeneration (AMD) but, in contrast to AMD, drusen in DDD may be found as early as the second decade of life.
Visual loss may occur (typically over two to three decades) usually as a consequence of retinal atrophy and sometimes associated with subretinal neovascular membrane formation.
COMPREHENSIVE CLINICAL NEPHROLOGY Copyright © 2015

38. Like AMD, MPGNII is a disease of the ECM, where defects in the regulation of the alternative pathway of complement, either by genetic alterations in genes including CFH, or the presence of auto-antibodies against FH, lead to deposits in the basement membranes of the glomeruli.
Y402H polymorphism of complement factor H and FHL-1
Exchange of tyrosine to histidine at aminoacid position 402

39. Extra-renal manifestations
Dense Deposit Disease
A small minority of patients with DDD have acquired partial lipodystrophy (APL), a condition with symmetric destruction of adipose tissue from the face, arms, and upper portions of the trunk
Acquired partial lipodystrophy has not been reported in humans with DDD resulting from genetic deficiency of CFH, implying a direct effect of the C3NeF itself in causing the adipocyte damage, rather than this being a manifestation of systemic complement alternative pathway dysregulation per se
Partial lipodystrophy can precede renal disease by some years and is
sometimes noticed acutely following an otherwise minor infection, suggesting that cytokine release or other manifestations of immunological activation can trigger adipocyte damage in the disease.
COMPREHENSIVE CLINICAL NEPHROLOGY Copyright © 2015

40. Clinical manifestations
C3 Glomerulonephritis
Because C3GN has only recently been recognized, clinical manifestations are less well defined.
In the French series, 27% of patients with C3GN had nephrotic syndrome at presentation.
Two thirds of patients had microhematuria at presentation.
One third had elevated blood pressure
CFHR5 nephropathy
The commonest cause of C3GN is complement factor H-related 5 (CFHR5) nephropathy, which is endemic in Greek Cypriots (1:6.000 of the population of the country).
CFHR5 nephropathy is characterised clinically by microscopic haematuria with episodes of macroscopic haematuria and acute renal dysfunction occurring at times of upper respiratory tract or other infections.
Mild to moderate proteinuria is seen late in the disease and the nephrotic syndrome is not a feature.
Circulating complement C3 and C4 levels are normal
Practical Nephrology 2014

41. Summary of clinical and laboratory features of C3 glomerulopathies
Practical Nephrology 2014

47. Evaluation of alternative complement pathway

50. Recurrence after Renal Transplantation
The risk for recurrence of C3 glomerulopathy is derived from small data sets:
In a study that included 18 transplants in dense deposit disease, recurrence was reported in 11 kidneys (61%), and there was a trend toward greater likelihood of transplant recurrence in dense deposit disease compared with either MPGN type 1 or MPGN type 3 groups
Kidney Int2006;69: 504–511
In a recent study, recurrence in C3 glomerulonephritis (6 out of 10 (60%)) and dense deposit disease (6 out of 11 (54.5%)) was similar.
Notably, some patients have developed thrombotic microangiopathy after renal transplantation
Kidney Int. 2012;82:

53. Treatment for C3 glomerulopathy
Cellular immunosuppression
Despite the recent designation of G3 glomerulopathy, the assumption must be that it has always existed (either in the form of dense deposit disease or C3 glomerulonephritis).

54. Treatment for C3 glomerulopathy
Cellular immunosuppression
There are no controlled trials to support the use of anti-cellular immune therapy in C3 glomerulopathy.
Strategies to reduce C3Nef with either mycophenolate mofetil or rituximab have not been studied formally, and there are reports of both response and non response in the published
Literature.
The most recent reports suggest that the anticellular immunosuppressive approach remains wholly unsatisfying:
McCaughan et al. reported a failure to respond to glucocorticoid, mycophenolate mofetil, and rituximab therapy
Bomback et al. reveal the failure of both prednisone and mycophenolate mofetil treatment despite appropriate escalation of both agents.
Am J Transplant2012;12: 1046–1051
Clin J Am Soc Nephrol2012;7: 748–756

55. Treatment for C3 glomerulopathy
Eculizumab

56. Treatment for C3 glomerulopathy
Eculizumab

57. Treatment for C3 glomerulopathy
Eculizumab
Treatment protocol with eculizumab was standard and homogenous in all reported patients: loading dose of 900 mg i.v. weekly for 5 weeks, followed by 1,200 mg i.v. every other week.
Most patients (8 out of 11 ) were treated for 12 months, while 2 patients were treated only for 2 months both showing a rapid response to treatment (one case with native kidneys, the other one with disease recurrence post-renal transplant).

58. Treatment for C3 glomerulopathy
Eculizumab

59. Treatment for C3 glomerulopathy
Eculizumab
Taken together, these results highlight three important points:
Treatment with eculizumab for C3G should probably be initiated early, before major sclerotic modifications have occurred in the renal glomeruli.
Proof of terminal pathway activation, indicated by elevated C5b-9 levels, may be an indicator of patients who can respond to treatment.
Treatment with eculizumab appears to be beneficial also in patients with advanced renal damage,and appears to be able not only to reduce proteinuria but also to improve renal function.
The effect of C5 inhibition on renal pathology is still debated. The limited available pathological data indicate that eculizumab decreased endocapillary proliferation and inflammatory cell infiltration in four out of six patients with repeated kidney biopsies.

60. Treatment for C3 glomerulopathy
Plasma therapy

61. Treatment for C3 glomerulopathy
Plasma therapy

62. Treatment for C3 glomerulopathy
Plasma therapy

63. Treatment for C3 glomerulopathy
Plasma therapy
Deletion of Lys224 in regulatory domain 4 of Factor H
Chronic treatment with periodical FFP-infusion was successful in preventing disease progression in two patients with C3 deposition glomerulopathy (C3DG) caused by alternative complement pathway dysregulation because of dysfunctional CFH and C3NeF.
On the basis of a CFH half-life of 6 days, FFP infusions (10–15 ml/kg body weight/infusion) were given every 14 days for a total of 36 months.

65. Treatment for C3 glomerulopathy.
Future therapeutic approaches
Recombinant proteins is the feature

66. In vitro studies using sera from patients with DDD showed that soluble CR1 prevents dysregulation of the alternative pathway C3 convertase, even in the presence of C3 nephritic factors.
In mice deficient in complement factor H and transgenic for human CR1, soluble CR1 therapy stopped alternative pathway activation, resulting in normalization of serum C3 levels and clearance of iC3b from glomerular basement membranes.
Short-term use of soluble CR1 in a pediatric patient with end stage renal failure demonstrated its safety and ability to normalize activity of the terminal complement pathway

68. Cp40, a second-generation compstatin analog in clinical development, is a 14 aminoacid
cyclic peptide that selectively inhibits complement activation in humans and non -human
primates by binding to C3 and C3b.
in vitro using multiple assays of complement activity, was demonstrated that:
Cp40 prevents complement -mediated lysis of sheep erythrocytes in sera from C3G patients, prevents complement dysregulation in the presence of patient-derived autoantibodies to the C3 and C5 convertases, and prevents complement dysregulation associated with disease-causing genetic mutations

70. Tripterygium wilfordii
ln 2012, a report was published on the effectiveness of Tripterygium wimnrdii (triptolide), a herbal extract with in vitro immunomodulatory effects and the ability to reduce renal complement expression
A reduction in proteinuria at different degrees was achieved in 8 out of 10 patients with DDD
The broader therapeutic use of this agent, that has been employed for rheumatoid arthritis and other diseases, is limited by reports of severe side effects in about half of the treated patients
Kidney int 2002;62(4):i
Clin Nephrol 2012:78(3):207—215