KDIGO-CMKD-CC-Speakers-Guide-v4-July-17-FINAL.pptx

Pathology

1. Atypical Hemolytic Uremic Syndrome and C3 Glomerulopathy KDIGO 2016 Controversies Conference This presentation is based on: Goodship T. et al., Kidney Intl (2017) 91:539-551.

2. Supported by The content of this presentation reflects the position of the authors of the original paper. It does not reflect the official opinion of Alexion or Achillion and should not be viewed as a position endorsed by them. Responsibility for the information and views expressed in this presentation lies entirely with the authors. Disclaimer : Eculizumab is not currently approved for the treatment of C3G.

3. Part 1: Introduction

4. The Roles of Complement Ricklin D, et al. Nat Rev Nephrol 12: 383-401, 2016.

5. Atypical Hemolytic Uremic Syndrome ( aHUS ) Ultra-rare disease characterized by acute kidney injury (AKI), thrombocytopenia, and microangiopathic hemolytic anemia. At least 50% of aHUS patients have an underlying inherited and/or acquired complement abnormality. Eculizumab, a humanized mAb against C5, makes it possible to control aHUS and prevent development of ESRD in the majority of patients.

6. C3 Glomerulopathy ( C3G) C3G comprises a group of kidney diseases driven by uncontrolled activation of the complement cascade that leads to C3 deposition within the glomerulus. The dysregulation of C3 convertase is driven by genetic and/or acquired defects. A biopsy is required to make the diagnosis. Two major subtypes are recognized: dense deposit disease (DDD) and C3 glomerulonephritis (C3GN) .

7. Complement Dysregulation in C3G & a HUS Mastellos DC, et al. Trends in Immunology 38: 383-394, 2017

8. SECTION A Atypical Hemolytic Uremic Syndrome ( aHUS )

9. Part 2: Renal Pathology: a HUS

10. aHUS Pathology aHUS is a thrombotic microangiopathy (TMA). Pathology resembles tissue responses to endothelial injury. The presence of C5b-9 staining is not a reliable indicator of aHUS. In general, it is not possible to determine etiology from morphology.

11. Morphological Features in Microangiopathy Active Lesions Chronic Lesions Glomeruli Thrombi Endothelial swelling or denudation Fragmented red blood cells Subendothelial flocculent material by EM Mesangiolysis Microaneurysms Glomeruli Double contours of peripheral capillary walls by LM, with variable mesangial interposition New subendothelial basement membrane by EM Widening of the subendothelial zone by EM   Arterioles Thrombi Endothelial swelling or denudation Intramural fibrin Fragmented red blood cells Intimal swelling Myocyte necrosis Arterioles Hyaline deposits   Arteries Thrombi Myxoid intimal swelling Intramural fibrin Fragmented red blood cells   Arteries Fibrous intimal thickening with concentric lamination (onion skin)  

12. a HUS Case Study 30-y.o. male, 2-week history of general malaise, presented with AKI ( plasma creatinine (Cr) 250 µ mol / L) and microangiopathic hemolytic anemia ( hemoglobin ( Hb ) 9.5 g/ dL and fragmented cells on a peripheral blood film ). Platelet count 130 x 10 9 / L. Renal biopsy showed endothelial swelling and denudation, mesangiolysis, double contours of the glomerular basement membrane and subendothelial accumulation of electron-lucent, flocculent material. No evidence of intraluminal thrombus. Patient’s maternal grandmother had developed kidney failure after the delivery of her second child.

13. a HUS Case Study Which of the following are correct? The renal biopsy appearances are not compatible with a diagnosis of aHUS . Screening for secondary causes of a renal thrombotic microangiopathy should be undertaken. Genetic screening for inherited complement abnormalities should be undertaken. The family history is not relevant. Treatment with high-dose steroids should be commenced . Both B and C. Both D and E.

14. Part 3 : Clinical Phenotype and Assessment: a HUS

15. a HUS : Presentation Current classifications of aHUS reflect a better understanding of disease mechanisms, including the impact of genetic background and etiologic triggers. Precipitating factors include autoimmune conditions, transplants, pregnancy, infections, drugs, and metabolic conditions. The time course and persistence of an aHUS episode are not well understood. Many patients appear to be at life-long risk for the recurrent acute presentation of aHUS . Disease penetrance for an acute episode of aHUS in carriers of known pathogenic mutations increases with age.

16. a HUS : Extrarenal Manifestations Extrarenal manifestations are reported in up to 20% of patients. It is unclear whether these manifestations are a direct consequence of complement activation, TMA, or other factors such as severe hypertension and uremia . ( Suppl Table 1)

17. a HUS : Extrarenal Manifestations Digital gangrene, skin Cerebral artery thrombosis/ stenosis Extracerebral artery stenosis Cardiac involvement/myocardial infarction Ocular involvement Neurologic involvement Pancreatic , gastrointestinal involvement Pulmonary involvement Intestinal involvement

18. a HUS : Laboratory Analysis Investigations should focus on determining the underlying etiology and excluding other diagnoses. Measure ADAMTS13 activity to diagnose or exclude thrombotic thrombocytopenic purpura (TTP) . Because the incidence of TTP is much lower in children than in adults, expert opinion recommends that in children, treatment with eculizumab should not be delayed while ADAMTS13 activity is being determined ; however , signs of nonresponse should be carefully monitored . Investigation for STEC-HUS should be routine in all patients with presumed aHUS.

19. a HUS : Laboratory Analysis Serum /plasma levels of complement proteins should be measured in all patients with primary aHUS prior to plasma therapy . C3 levels will be low in 30-50% of aHUS cases. Low C3 levels are also noted in acute STEC- aHUS and pneumococcal aHUS . Complement split products and assays of complement function can also be obtained, although the significance of some of these assays requires further study.

20. a HUS Case Study 24-y.o. female presented with: AKI (Cr 500 µ mol /L) thrombocytopenia (platelet count 50 x 10 9 /L ); and a microangiopathic haemolytic anaemia, following a 2-week history of intermittent diarrhea .

21. a HUS Case Study Which of the following investigations should be undertaken to elucidate the cause of illness? Measure serum levels of C3 and C4. Measure ADAMTS13 activity. Screen for STEC infection. Screen for factor H autoantibodies. Screen for anticardiolipin antibodies. All of the above. None of the above.

22. Part 4 : G enetic and Acquired Drivers of Disease: a HUS

23. a HUS : Genetic Drivers of Disease Studies of hundreds of aHUS patients have provided an excellent understanding of genetic drivers of disease, leading to the development of individualized care. Genetic screening and molecular diagnostics, with expert interpretation of the results, should inform therapeutic decisions .

24. a HUS : Genetic Testing The minimum set of genes that should be screened includes CFH , CD46 , CFI , C3 , CFB , THBD , CFHR1 , CFHR5 , and DGKE . Genetic testing should also include the risk haplotypes CFH-CFHR3 and MCP ggaac as they modify disease penetrance and severity . Delays in obtaining results from genetic or molecular diagnostic studies should not prevent a clinical diagnosis or postpone treatment, as early anticomplement treatment is crucial to preserve renal function and avoid irreversible sequelae . Technologies to detect copy number variation, hybrid genes and other complex genomic rearrangements in the CFH/CFHRs genomic region must be included in the genetic testing .

25. a HUS : Genetic Testing Genetic analysis is essential in living-related kidney donor transplantation. Transplantation from living-related kidney donors should only be considered if causative genetic or acquired factors are clearly identified in the recipient and the related donor is free of these factors . Genetic testing is recommended for patients in whom discontinuation of eculizumab is being considered.

26. Understanding Genetic Variants Genetic variants should be classified as “benign,” “likely benign,” “variant of uncertain significance (VUS),” “likely pathogenic,” or “pathogenic,” following international guidelines. Pathogenic variants compromise protection of host endothelial cells and platelets from complement damage/activation .

27. a HUS : Acquired Drivers of Disease Acquired drivers of disease are autoantibodies to complement proteins or protein complexes that impair normal function. The best-studied acquired drivers are FH autoantibodies, which are usually seen in association with deletion of the CFHR3 and CFHR1 genes. The deletion of CFHR3 and CFHR1 is a common copy number variation that can be identified on genetic testing. The finding of FH autoantibodies should be confirmed in a second sample at least 4 weeks after the initial sample.

28. a HUS : Acquired Drivers of Disease Testing should also be performed in the pre-renal transplant period. In pediatric patients, FH autoantibody assays should be performed following consensus guidelines: at diagnosis and if positive, at day 7, 14, 28, monthly and one year. Relapses of anti-FH associated HUS occur in about 20-25% patients .

29. a HUS Case Study 35-y.o. female presented with AKI, thrombocytopenia, and a microangiopathic hemolytic anemia . Treated with eculizumab and made a full recovery. On genetic screening, she was found to carry a heterozygous CFH/CFHR1 hybrid gene . Family history: Both parents alive. Two brothers and 3 sisters. Two children aged 6 and 2.

30. a HUS Case Study Which of the following are correct? Other members of the family are not at risk of developing aHUS . There is no need to offer screening to other family members for the CFH/CFHR1 hybrid gene. The finding of the CFH/CFHR1 hybrid gene in an unaffected family member has no implications for that individual. Family members should be offered genetic counselling. aHUS can present at any age in unaffected carriers of the CFH/CFHR1 hybrid gene . Both A and C. Both D and E.

31. Part 5 : Treatment Strategies: a HUS

32. a HUS : Treatment All patients with a clinical diagnosis of primary aHUS are eligible for treatment with eculizumab. The dosing schedule reported in the trials that led to the approval of eculizumab should be followed. Treatment duration is controversial as there is no evidence to support life-long therapy in all aHUS patients .

33. a HUS : Treatment Two options for long-term dosing have been considered: The minimal dose required to achieve complement blockade. A discontinuation dosing schedule . No data exist to support either option, and both require monitoring of complement activity . There are no data to inform the frequency of testing Dose reduction or discontinuation require ongoing monitoring of complement activity. In patients who have undergone transplant, especially patients who have lost previous allografts, discontinuation is not recommended.

34. a HUS : Treatment If access to eculizumab is unavailable, plasma therapy can be used. The use of plasma exchange when eculizumab is available may be associated with some improvement, but delaying use of eculizumab may lead to a suboptimal therapeutic outcome. Eculizumab increases the risk of meningococcal infection. Patients should receive vaccination against meningococcus (including Type B); however, vaccination should not delay the start of eculizumab therapy. Antibiotic prophylaxis is mandated during the first 2 weeks.

35. a HUS Treatment : Transplant Kidney transplantation should be delayed for at least 6 months after the start of dialysis as limited renal recovery is possible several months after starting eculizumab. Living-related kidney donation carries a risk for recurrence in the recipient and a risk of de novo disease in the donor should the donor carry an at-risk genetic variant. Liver transplant remains an option in patients with liver-derived complement protein abnormalities, in particular for renal transplant recipients with uncontrolled disease activity despite eculizumab therapy.

36. a HUS Case Study 44-y.o. male on hemodialysis for 15 years. Primary renal disease is aHUS . Received two cadaver renal transplants, both of which were lost within 3 months of transplantation to recurrent disease despite treatment with plasma exchange. Genetic screening showed a heterozygous mutation in factor H, which has been reported previously in six other patients with aHUS . Being considered for a third transplant.

37. a HUS Case Study Which of the following are correct? Prophylactic use of eculizumab immediately before and then long-term after the transplant is unlikely to prevent disease recurrence in the transplanted kidney. Prophylactic plasma exchange immediately before and then long-term after the transplant is highly likely to prevent disease recurrence after the transplant. A living-related transplant should not be considered under any circumstances. If he receives prophylactic eculizumab, he should be vaccinated against meningococcus.

38. a HUS Research Recommendations: Summary A comparative study of biopsies from patients with well-documented malignant hypertension and patients with well-documented alternative complement pathway disease. A longitudinal study of patients with features of chronic microangiopathy on biopsy but without a history of acute presentation. Clinical studies Define how complement biomarkers correlate with current or impending aHUS relapse and/or renal involvement. Identify risk factors for relapse upon cessation of anti-complement therapy. Identify alternative anti-complement therapeutics.

39. SECTION B C3 Glomerulopathy (C3G)

40. Part 2: Renal Pathology: C3G

41. C3G Pathology The C3G disease spectrum is caused by abnormal control of complement activation, deposition or degradation that results in predominant glomerular C3 fragment deposition. A renal biopsy using immunofluorescence (IF) is required to diagnose C3G. Electron microscopy (EM) is used to sub-classify C3G as DDD or C3GN .

42. Morphological Features Light Microscopy Active lesions Mesangial expansion with or without hypercellularity Endocapillary hypercellularity including monocytes and/or neutrophils Capillary wall thickening with double contours (the combination of capillary wall thickening and mesangial increase is referred to as a membranoproliferative pattern) Necrosis Cellular/ fibrocellular crescents   Chronic lesions Segmental or global glomerulosclerosis Fibrous crescents Immunofluorescence Microscopy Typically dominant C3 staining Electron Microscopy DDD: Dense osmiophilic mesangial and intramembranous electron dense deposits C3GN: Amorphous mesangial with or without capillary wall deposits including subendothelial, intramembranous and subepithelial electron dense deposits Sub-epithelial ‘humps’ may be seen in both DDD and C3GN

43. C3G Pathology : Controversies Correlations between renal biopsy appearances, etiology, and clinical outcome are ill-defined. IF staining is subjective and semiquantitative. Well-defined for dense deposit disease (DDD). Not clear if characteristic for C3 glomerulonephritis (C3GN). Distinguishing DDD and C3GN by EM can be difficult.

44. C3G Case Study 7-y.o.male with 3-day h/o macroscopic hematuria 4-wk after being culture-positive for Group A β -hemolytic Strep pharyngitis. Normal blood pressure and urine output, no edema. Urinalysis: numerous RBCs; RBC casts; 4+ protein; negative leukocytes and nitrites. Serum Cr 1.0 mg/ dL (normal 0.2 -0.6 mg/dL); normal electrolytes and albumin. ASO elevated (1240 IM/ mL [normal < 150 IU/ mL]) ; ANA negative; C3 39 mg/ dL (normal 87 -181 mg/ dL ). FH: negative for renal disease.

45. C3G Case Study Diagnosed with APSGN and monitored as an outpatient. One month later, creatinine has improved to 0.7 mg/dL, UPC ratio is 0.7 (normal < 0.2 ), and C3 is 22 mg/dL. What would you do now? Get a renal biopsy. Continue to monitor and get a renal biopsy 3 months after onset if C3 has not improved. Start an oral course of prednisone at 2 mg/kg/ day. Get genetic testing. Get comprehensive complement studies.

46. Part 3 : Clinical Phenotype and Assessment: C3G

47. C3G: Presentation C3G generally follows a chronic, indolent course with persistent AP activation resulting in a 10-year renal survival of approximately 50%. There are, however, cases of C3G that present as a rapidly progressive GN.

48. C3G: Extrarenal Manifestations Acquired partial lipodystrophy (APL) and retinal drusen are reported and appear to be direct consequences of complement activation .

49. C3G: Laboratory Analysis Serum /plasma levels of complement proteins should be measured in all patients with C3G prior to plasma therapy . C3 levels will be low in 30-50% of aHUS cases and up to 75% of C3G cases. Complement split products and assays of complement function can also be obtained, although the significance of some of these assays requires further study.

50. C3G Case Study 33-y.o.female with a diagnosis of DDD. Ophthalmologic examination showed drusen. Complement labs showed: C3, < 0.15 g /L (normal 0.9 -1.8g/ L); C3c, 3.3mg/L (normal <2.0 mg/L) B, 23.9 mg/L (normal 22-50 mg/ dL ); Bb, 2.3 mg/L (normal <2.2mg/L) C5, 12 mg/ dL (normal 10-21 mg/ dL ); soluble C5b-9, 0.25mg/L (normal <0.3mg/L) CH50, <5 U/ml (normal 30-90 U/ml); alternative pathway functional assay (APFA) 0% (normal 50-130%)

51. C3G Case Study What do these results mean? There is no ongoing complement activity. The ongoing complement activity is primarily at the level of the alternative pathway. The ongoing complement activity is primarily at the level of the classical pathway. The ongoing complement activity is primarily at the level of the terminal pathway.

52. Part 4 : G enetic and Acquired Drivers of Disease: C3G

53. C3G: Genetic Drivers of Disease Understanding of the genetics of C3G is not yet comparable to that of aHUS. There is no clear benefit to performing genetic analysis in all cases of C3G.

54. C3G: Genetic Testing The minimum set of genes that should be screened includes CFH , CD46 , CFI , C3 , CFB , THBD , CFHR1 , CFHR5 , and DGKE . Technologies to detect copy-number variation, hybrid genes, and other complex genomic rearrangements in the CFH/CFHRs genomic region must be included in the genetic testing .

55. C3G: Genetic Testing Genetic analysis is essential in living-related kidney donor transplantation . All planned recipients of a living-related kidney should be screened. If a genetic abnormality is found, the donor should be tested to exclude that genetic abnormality .

56. Understanding Genetic Variants Genetic variants should be classified as “benign,” “likely benign,” “variant of uncertain significance (VUS),” “likely pathogenic,” or “pathogenic,” following international guidelines. C3G appears mechanistically more complex than aHUS. There is limited information about genotype/phenotype correlations to distinguish different C3G subtypes, inform prognosis and/or recommend treatment.

57. C3G: Acquired Drivers of Disease Acquired drivers of disease are autoantibodies to complement proteins or protein complexes that impair normal function . In C3G, the most common autoantibodies are to C3 convertase , a serine protease formed from C3b and Bb. These autoantibodies are called C3Nefs. They stabilize C3 convertase and prolong its half-life. Other antibodies in C3G include FH autoantibodies, C4Nefs and C5Nefs. In older adults, serum free light chains (FLC) should be assayed.

58. C3G Case Study 24-y.o.male with a diagnosis of DDD on hemodialysis. Planning living-related donor transplant from sibling. Family history: Cousin also on dialysis. Genetic testing identified: An ultra-rare variant in the complement factor H gene, CFH c.3229T>C, which substitutes Cys at position 1077 for Arg (p.Cys1077Arg); this is classified as a VUS (variant of uncertain significance ). A common copy-number variant (CNV) in which one copy of the complement factor H–related 3 and –related 1 genes are deleted (del CFHR3-CFHR1 ).

59. C3G Case Study What do these results mean and what would you do? These results can be dismissed. He should not receive a kidney transplant because factor H is synthesized by the liver, and his disease will recur. The donor sibling should be screened for the complement factor H gene variant and, if found , he should be excluded from donating. The donor sibling should be screened for the complement factor H gene variant and, if found , he should be cleared to donate.

60. Part 5 : Treatment Strategies: C3G

61. C3G Treatment : All Patients  All Patients   Optimal blood pressure control (suggested blood pressure below the 90% in children and ≤120/80 in adults) Priority agents include angiotensin converting enzyme inhibitors and angiotensin receptor blockers Optimal nutrition for both normal growth in children, healthy weight in adults Lipid control  Moderate Disease Description Urine protein over 500 mg/24 hours despite supportive therapy OR Moderate inflammation on renal biopsy OR Recent increase in serum creatinine suggesting risk for progressive disease   Recommendation Prednisone Mycophenolate mofetil  Severe Disease  Description Urine protein over 2000 mg/24 hours despite immunosuppression and supportive therapy OR Severe inflammation represented by marked endo - or extracapillary proliferation with or without crescent formation despite immunosuppression and supportive therapy OR Increased serum creatinine suggesting risk for progressive disease at onset despite immunosuppression and supportive therapy   Recommendation Methylprednisolone pulse dosing as well as other anti-cellular immune suppressants have had limited success in rapidly progressive disease Data are insufficient to recommend eculizumab as a first-line agent for the treatment of rapidly progressive disease

62. C3G: Treatment A retrospective study supports the effectiveness of mycophenolate mofetil in C3GN patients. No specific recommendation can be made for plasma therapy or rituximab (an anti-CD20 antibody). Since the pathogenesis of C3G is due to dysregulation and hyperactivity of the alternative pathway of complement, eculizumab has been tried in a limited number of patients with varied results.

63. C3G Treatment : Transplant No specific data are available to inform decisions surrounding transplantation in C3G. Recommendations reflect expert opinion and limited case reports. C3G recurs in allografts at a high rate, leading to graft loss in ~50% of patients.

64. C3G Case Study 32-y.o . male with C3GN diagnosed 2 years ago. On ACEs and ARBs with good control of HTN but increasing proteinuria and decreasing renal function. CKD G3 with eGFR 45 mL/min/ 1.73 m 2 Complement studies: No rare genetic variants identified in CFH , CD46 , CFI , C3 , CFB , THBD , CFHR1 , CFHR5 , and DGKE C3, <0.55g/L (normal 0.9 - 1.8 g /L; C3c, 2.2mg/L (normal <2.0 mg/L); C5, 8.5 mg/ dL (normal 10-21 mg/ dL ); soluble C5b-9, 5.45 mg /L (normal < 0.3 mg /L); CH50, <26 U/mL (normal 30-90 U/ mL) ; APFA (alternative pathway functional assay 3% (normal 50-130%) C3Nef +2; C5Nef +4

65. C3G Case Study What would you do next? Add rituximab. Begin plasma exchange. Add mycophenolate mofetil . Add eculizumab.

66. C3G Research Recommendations: Summary A multicenter study analyzing biopsies to define the relationship of morphology to etiology, clinical course, and response to therapy. Comprehensive genetic testing to fill the knowledge gap in establishing robust phenotype-genotype correlations. Clinical studies Assess the value of proximal (at the level of the AP) anti-complement therapy. Development and trial of novel complement inhibitors. Determine value of complement biomarkers to inform clinical outcome in C3G patients and stratify them into targeted treatment groups.

67. Conclusions While there are knowledge gaps in both aHUS and C3G, the evidence base for the management of patients with C3G lags behind that of aHUS; addressing this disparity should be a priority. Although these two diseases are presented as distinct entities, there is substantial overlap in their pathogenesis and clinical presentation.

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