Response to “A Caution Against the Use of C5B-9 Endothelial Assay to Support Eculizumab Therapy”

We would like to thank Dr. Duineveld and colleagues for showing interest in our article titled "Eculizumab first" in the management of posttransplant thrombotic microangiopathy [1Federica Maritati Corradetti Valeria Bini Claudia et al."Eculizumab First" in the Management of Posttransplant Thrombotic Microangiopathy.Kidney Int Rep. 2024; https://doi.org/10.1016/j.ekir.2024.01.013Abstract Full Text Full Text PDF Scopus (0) Google Scholar], by highlighting the role of the ex-vivo C5b-9 assay in the evaluation of disease activity. Indeed, the diagnosis of posttransplant thrombotic microangiopathy (TMA) is based on clinical course and laboratory data as described in our paper (Methods Section). We underly how a rapid diagnosis and a prompt treatment with eculizumab leads to a good outcome, as demonstrated by the trend of hemoglobin, platelets, LDH, and eGFR. The test of complement activation on endothelial cells was performed, together with genetic analysis, in another Center (Mario Negri Institute in Bergamo) before eculizumab starting and the results were available some weeks after diagnosis and treatment. Thus, this test was not used for disease management. However, the retrospective revision of the ex-vivo C5b-9 formation assay showed that, among 25 patients tested, 84% and 92% resulted positive respectively on unstimulated and activated endothelial cells. This finding reinforces, in our opinion, the hypothesis of a complement hyperactivation secondary to a significant trigger (ischemia/reperfusion injury, rejection) in patients with some genetic predisposition to atypical hemolytic-uremic syndrome (15% of tested patients had rare complement gene variants and almost 90% carried at least one risk haplotype/polymorphism). Our findings are consistent with other previous studies reported in literature. The capability of the ex-vivo C5b-9 formation assay to evaluate the presence of factors in serum that augment complement activation on endothelial cells, and to identify patients with TMA caused by complement dysregulation is in fact supported by several publications from different independent groups, including Giuseppe Remuzzi in Bergamo [2Gastoldi Sara Aiello Sistiana Miriam Galbusera et al An ex vivo test to investigate genetic factors conferring susceptibility to atypical haemolytic uremic syndrome.Front Immunol. 2023 Feb 9; 141112257Crossref PubMed Scopus (3) Google Scholar,3Galbusera Miriam Noris Marina Gastoldi Sara et al.An Ex Vivo Test of Complement Activation on Endothelium for Individualized Eculizumab Therapy in Hemolytic Uremic Syndrome.Am J Kidney Dis. 2019 Jul; 74: 56-72Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar], S.A.M.E.G. Timmermans in Maastricht [4Werion Alexis Storms Pauline Zizi Ysaline et al.Epidemiology, Outcomes, and Complement Gene Variants in Secondary Thrombotic Microangiopathies.Clin J Am Soc Nephrol. 2023 Apr 21; 18: 881-891Crossref PubMed Google Scholar,5Sjoerd A.M.E.G. Timmermans, Myrurgia A. Abdul-Hamid, Joris Vanderlocht et al Patients with hypertension-associated thrombotic microangiopathy may present with complement abnormalities. Kidney Int. 2017 Jun;91(6):1420-1425Google Scholar] and Marta Palomo in Barcelona [6Marta Palomo, Miquel Blasco, Patricia Molina et al Complement Activation and Thrombotic Microangiopathies. Clin J Am Soc Nephrol. 2019 Dec 6; 14(12): 1719–1732.Google Scholar,7Blasco Miquel Guillen-Olmos Elena Diaz-Ricart Maribel et al.Complement Mediated Endothelial Damage in Thrombotic Microangiopathies.Front Med (Lausanne). 2022 Apr 25; 9811504PubMed Google Scholar]. In addition, several published findings showed that C5b-9 formation assay is positive in patients with secondary forms of acute TMA [3Galbusera Miriam Noris Marina Gastoldi Sara et al.An Ex Vivo Test of Complement Activation on Endothelium for Individualized Eculizumab Therapy in Hemolytic Uremic Syndrome.Am J Kidney Dis. 2019 Jul; 74: 56-72Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar, 4Werion Alexis Storms Pauline Zizi Ysaline et al.Epidemiology, Outcomes, and Complement Gene Variants in Secondary Thrombotic Microangiopathies.Clin J Am Soc Nephrol. 2023 Apr 21; 18: 881-891Crossref PubMed Google Scholar, 5Sjoerd A.M.E.G. Timmermans, Myrurgia A. Abdul-Hamid, Joris Vanderlocht et al Patients with hypertension-associated thrombotic microangiopathy may present with complement abnormalities. Kidney Int. 2017 Jun;91(6):1420-1425Google Scholar]. The above studies document that endothelial complement dysregulation occurs also in TMA forms in which the environmental trigger or the underlying disease play a stronger role in inducing endothelial injury and complement activations than the genetic background. By the way, rare complement gene variants have been reported in 10-40% of patients with secondary TMA. In addition, risk variants and haplotypes may predispose to develop secondary forms of TMA. Finally, previous studies showed that a positive C5b-9 test on endothelium predicts the response to eculizumab and may support treatment monitoring [3Galbusera Miriam Noris Marina Gastoldi Sara et al.An Ex Vivo Test of Complement Activation on Endothelium for Individualized Eculizumab Therapy in Hemolytic Uremic Syndrome.Am J Kidney Dis. 2019 Jul; 74: 56-72Abstract Full Text Full Text PDF PubMed Scopus (60) Google Scholar,8Sjoerd A.M.E.G. Timmermans, Alexis Werion, Jan GMC Damoiseaux et al Diagnostic and Risk Factors for Complement Defects in Hypertensive Emergency and Thrombotic Microangiopathy. Hypertension. 2020 Feb;75(2):422-430Google Scholar] Thus, after literature revision, we can affirm that our results are consistent with previous data and confirm that the test is effective to identify patients with active TMA associated with complement activation. A CAUTION AGAINST THE USE OF C5B-9 ENDOTHELIAL ASSAY TO SUPPORT ECULIZUMAB THERAPYKidney International ReportsPreviewIn their recent study Maritati and colleagues reported 45 patients with early onset TMA after kidney transplantation.[1] Using serum of these patients an increased C5b-9 deposition on endothelial cells was observed in 84%. The authors suggest that these positive test results support the effective use of eculizumab. We caution against such conclusion. Initially, it was hoped that the ex-vivo complement assay would allow to evaluate the presence of factors in serum that augment complement activation on endothelial cells, and to identify patients with (risk of) TMA caused by complement dysregulation. Full-Text PDF Open Access