Treating C3 glomerulopathy with eculizumab

The report includes five patients (all white Caucasians) with C3GN (two males and three females) and two patients with DDD (both female, Table 1). Three patients had recurring allograft C3G. One patient was diagnosed with de novo allograft C3GN after kidney transplantation with ESRD due to focal segmental glomerulosclerosis (FSGS). Four patients received immunosuppressive agents prior to eculizumab with varying success. The mean age at diagnosis was 29 years (range = 14–59 years). At diagnosis all subjects had either impaired renal function (mean serum creatinine 2 mg/dl; range = 0.8–3.3 mg/dl) and/or elevated levels of urinary protein (urinary protein to creatinine ratio [UPCR] 3.6 g/g; range = 0.3–5.4 g/g). All patients presented with varying levels of hematuria.

Patient C3GN1 was diagnosed with C3GN in his/her late thirties, 4 years after receiving an allogenic kidney transplant (baseline creatinine 1.6 mg/dl post transplantation; immunosuppression with tacrolimus, MMF [mycophenolate mofetil], and prednisone) due to ESRD of MPGN (initial diagnosis 11 years earlier with classification as MPGN type I). The patient presented with elevated levels of serum creatinine (1.9 mg/dl), urinary protein (5–6 g/g), and hematuria. The kidney biopsy revealed discrete glomerular fibrosis, mesangial hypercellularity, lymphocyte infiltration, and pronounced C3 staining in accordance with C3GN (Table 2). Complement assays showed complement activation with low C3 levels (Additional file 1: Table S2). Additionally, C3 convertase autoantibodies were detected (C3 nephritic factors, C3NeF; Table 2). Immunosuppression was maintained with prednisone, tacrolimus, and MMF. One-time administration of rituximab did neither affect creatinine levels nor urinary protein. Treatment with eculizumab over 27 months maintained stable kidney transplant function (serum creatinine levels of 1.6–2.2 mg/dl), greatly improved urinary protein (0.2–0.4 g/g), and led to stable hematuria (Fig. 1a). Complement testing at week 17 and 43 of treatment suggested effective complement blockade (CH50 < 10 IE/ml, ref. range 20–50 IE/ml; Additional file 1: Table S2).

Patient C3GN2 with a history of HIV infection (HAART had been successfully applied to control HIV infection) presented with serum creatinine levels of 2.3 mg/dl, urinary protein levels of 0.3 g/g, and hematuria in his/her early forties. Kidney biopsy revealed global mesangial and endocapillary hypercellularity with discrete glomerular sclerosis. Immunofluorescence and electron microscopy revealed distinct C3 deposition with little to no immunoglobulin deposition (Table 2) leading to the diagnosis of C3GN. Complement analysis showed increased C3 turnover and sMAC formation (Additional file 1: Table S3). Antibody and genetic testing showed no pathologic mutations, but identified antibodies against C3NeF, C3b, and CFH (Table 2). Eculizumab treatment resulted in an improvement of renal function with slightly improved serum creatinine levels at 1.2–1.7 mg/dl and low range urinary protein (0.1–0.2 g/g; Fig. 1b). Notably, a short interruption of eculizumab treatment for 12 weeks led to a dramatic increase of urinary protein (maximum urinary protein levels 1.8 g/g), worsened renal function (maximum serum creatinine levels 2.2 mg/dl), and enhanced sMAC formation (maximum sMAC levels 914 ng/dl). Re-initiation of eculizumab promptly reduced urinary protein (0.1–0.3 g/g), restored kidney function (serum creatinine levels 1.4–1.8 mg/dl), and improved hematuria.

Patient C3GN3 presented with relapsing C3GN in his/her mid-thirties, 3 months after receiving allogenic kidney transplantation (post transplantation creatinine 1.5 mg/dl; immunosuppression with tacrolimus, MMF, and prednisone) due to ESRD of C3GN (initial diagnosis 7 years earlier). Previous treatment approaches included prednisone, MMF, and azathioprine. The patient presented with elevated serum creatinine (2.1 mg/dl), discrete urinary protein (0.3 g/g), and hematuria (Additional file 1: Table S4). Transplant kidney biopsy revealed no signs of graft rejection, but showed mesangial proliferation, lymphocyte infiltration and predominant C3 staining without immunoglobulin deposition (Table 2). Electron microscopy confirmed relapsing C3GN. Antibody and genetic testing revealed neither autoantibodies nor pathophysiological relevant mutations in any of the known disease genes (Table 2). Notably a heterozygous variant change of questionable pathophysiological relevance in exon 17 of C3 was detected (detailed discussion Additional file 1: Text S1). Treatment with eculizumab stabilized kidney function (serum creatinine 1.9–2.2 mg/dl), urinary protein (0.25–0.35 g/g), and hematuria (Fig. 1c). Immunosuppression was maintained with prednisone, tacrolimus, and MMF. Of note, eculizumab had to be paused for 4 weeks due to immunosuppression-induced agranulocytosis.

Patient DDD1 presented in his/her early twenties with impaired renal function (serum creatinine 1.5 mg/dl), nephrotic range urinary protein (5.7 g/g), and hematuria. Complement analysis showed activation of the alternative pathway and high sMAC levels (Additional file 1: Table S5). A kidney biopsy at the department of pediatric nephrology 10 years before had shown mesangial proliferation, and prominent C3 staining with little immunoglobulin staining. Electron microscopy confirmed DDD (Table 2). Antibody analysis and genetic testing identified neither autoantibodies against C3NeF, nor pathophysiological relevant mutations in CFH (Table 2). Treatment with eculizumab stabilized kidney function (serum creatinine 1.1–1.3 mg/dl) and initially improved urinary protein (2.2–2.7 g/g). Hematuria levels significantly decreased. However, urinary protein deteriorated after 87 weeks of therapy (3.0–3.7 g/g) (Fig. 1d). Blood pressure was carefully controlled and within normal range.

Patient C3GN4 presented with de novo C3GN at the in his/her early twenties. He had received a kidney transplant due to FSGS with ESRD 10 years earlier. In line with the initial diagnosis of FSGS an immunosuppressive therapy with prednisone, cyclosporine, and cyclophosphamide had been administered. One month following kidney transplantation, the patient had been successfully treated with cyclophosphamide and plasmapheresis due to relapsing FSGS [41]. 11 years after transplantation, laboratory testing showed a gradual loss of transplant function with elevated serum creatinine (3.3 mg/dl), prominent urinary protein (3.4 g/g), and hematuria. Transplant kidney biopsy revealed glomerular sclerosis, mesangial and endocapillary proliferation. Immunofluorescence and electron microscopy confirmed the diagnosis of de novo C3GN (Table 2; C3GN4, 1^st biopsy). Further analysis showed activation of the alternative pathway and a weakly positive C3NeF (Additional file 1: Table S6). Genetic testing revealed heterozygous changes of unknown clinical significance in CFI and COL4A3 (Table 2, detailed discussion Additional file 1: Text S2).

Immunosuppression after kidney transplantation was maintained with prednisone, tacrolimus, and MMF. After administration of eculizumab, kidney transplant function and urinary protein greatly improved. Serum creatinine levels stabilized at 1.6–1.8 mg/dl and urinary protein levels at 1 g/g. However after 3 months of treatment, eculizumab was stopped due to lack of compliance. 1.5 years later the patient presented again with relapsing disease, serum creatinine levels of 3 mg/dl, and urinary protein of 4.3 g/g. A second kidney biopsy showed progressing glomerular sclerosis, and immunofluorescence revealed complement deposition but no IgG. (Table 2; C3GN4, 2^nd biopsy). Treatment with eculizumab was reapplied. Hematuria levels remained stable, however serum creatinine and urinary protein levels further increased, and treatment with eculizumab was discontinued after 3 months (Fig. 2a).

Patient DDD2 presented with elevated serum creatinine levels (2.1 mg/dl), nephrotic range urinary protein (5.4 g/g), and hematuria at the in his/her late fifties (Additional file 1: Table S7). Kidney biopsy displayed mesangial proliferation, lymphocyte infiltration, and discrete glomerular sclerosis. Immunofluorescence showed distinct C3 staining with little immunoglobulin staining. Electron microscopy found dense deposits in accordance with DDD (Table 2). Further diagnostic workup revealed alternative pathway complement activation but no autoantibodies against C3NeF and CFH. Genetic analysis detected a heterozygous missense variant in CFHR5 (Table 2, detailed discussion in Additional file 1: Text S3). Treatment with eculizumab was initiated stabilizing kidney function at serum creatinine levels of 2 mg/dl and urinary protein levels of 2 g/g. However, in the following months kidney function and urinary protein deteriorated. Thus, treatment with eculizumab was terminated and dialysis was initiated (Fig. 2b).

Patient C3GN5 presented in his/her mid-teens with nephrotic range urinary protein (5.6 g/g), normal renal function (serum creatinine levels 0.8 mg/dl), and hematuria. Kidney biopsy showed pronounced glomerular sclerosis, mesangial proliferation, and distinct C3 staining with little immunoglobulin deposition. Electron microscopy confirmed the diagnosis of C3GN (Table 2, C3GN5, 1^st and 2^nd biopsy). Complement analysis revealed extensive activation of the alternative pathway with low C3 levels (Additional file 1: Table S8). Antibody and genetic testing detected C3NeF, but no mutation in known disease genes (Table 2). Treatment with plasmapheresis, plasma infusion, prednisone, and MMF had initially induced reduction of urinary protein (details published in [25]). Due to deterioration of urinary protein (5.6 g/g) eculizumab therapy was initiated. With the initiation of eculizumab, MMF was stopped but was readministered after 2.5 months since urinary protein increased. A follow-up biopsy after 1 year of eculizumab treatment revealed glomerular IgG, and κ light chain deposition with identical staining pattern for mouse antibodies (Table 2, C3GN5, 2^nd biopsy). Since eculizumab is a humanized monoclonal antibody with human IgG2/IgG4 heavy and κ light chain, and mouse variable regions [42], this glomerular antibody deposition may reflect eculizumab. A break of 17 weeks due to insurance payment negotiations led to a dramatic increase of urinary protein (> 15 g/g) and decline of renal function. Reinitiation of eculizumab could not halt disease progression (Fig. 2c).