New-Onset Complement-Mediated Thrombotic Microangiopathy during the COVID-19 Pandemic

The coronavirus disease-19 (COVID-19) is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The virus is alleged to enable a proinflammatory state that leads to the activation of the coagulation and the complement cascade [1]. The complement system is part of the innate immune system that can be activated by different agents, such as immune complexes or microorganisms (classical and lectin pathway), or that is continuously active (alternative pathway) and needs to be actively controlled by regulatory proteins in a state in which activation is not needed. The alternative complement pathway is involved in renal diseases and is a central component of the pathomechanism in complement-mediated thrombotic microangiopathy or atypical haemolytic uremic syndrome (cTMA/aHUS) and in complement 3 (C3) nephropathy.

Both COVID-19 itself and vaccinations against SARS-CoV-2 have been linked to relapses and to first manifestations of cTMA/aHUS, respectively [2, 3]. In a previous study, we could show that COVID-19 was associated with a higher risk of relapse of TMA than the SARS-CoV-2 vaccinations in a cohort of patients with an established diagnosis of cTMA/aHUS [4]. In this study, we investigated the impact of the COVID-19 pandemic on patients with a new onset of cTMA/aHUS within the Vienna TMA cohort and whether new-onset case numbers would increase during the pandemic (until end of May 5, 2023, when the World Health Organization declared the end of the public health emergency of international concern).

We examined patients with new-onset aHUS/cTMA during the COVID-19 pandemic. The Institutional Review Board (IRB) of the Medical University of Vienna approved the study (unique IRB identifier: 1368/2014). All patients provided written informed consent including consent to the publication of details of their medical case, and the study was conducted in accordance with the principles of the Declaration of Helsinki. The data that support the findings of this study are not publicly available due to their containing information that could compromise the privacy of research participants but are available from the corresponding author (C.A.) upon reasonable request.

The data were derived from the Vienna TMA cohort which was established in 2014 at the Division of Nephrology and Dialysis, Department of Medicine III, of the Medical University of Vienna. It continuously includes all patients that present with complement-mediated TMAs, secondary TMAs to other disorders as well as patients with other complement-mediated kidney diseases (such as C3 glomerulonephritis) that are followed at our institution [5, 6]. The study period extended from March 2020 following the patients until May 5, 2023, thus covering the whole period of the public health emergency of international concern as declared by the World Health Organization.

We included all patients that presented with new-onset aHUS/cTMA during the follow-up (FU) period. We took special interest in aHUS/cTMA episodes that were related to SARS-CoV-2 infections or SARS-CoV-2 vaccinations.

We recorded SARS-CoV-2 infections in case of a positive result of a real-time quantitative polymerase chain reaction (RT-qPCR) test, and a RT-qPCR result with a cycle threshold of less than 30 was deemed positive for the presence of SARS-CoV-2. Both paper-based results and reports of positive results of patients were considered. SARS-CoV-2 vaccinations, free of charge in Austria since the second quarter of 2021 with specific recommendations for individuals at high risk for severe COVID-19, are entered in a central electronic database with details on the date of application and type of vaccine used and could thus be analysed in relation to TMA onset or flares. A TMA episode was defined either by the presence of specific laboratory and clinical signs (mechanical haemolysis, thrombocytopenia, acute kidney injury) and/or of TMA on a kidney biopsy (thrombi in glomerular capillaries or arterioles, mesangiolysis, endothelial swelling, double contouring of basement membranes, multilayering of arterioles). An association with COVID-19 or SARS-CoV-2 vaccination was considered if any signs or symptoms of TMA occurred within 8 weeks of the respective event and no other triggering factor could be identified. All standard laboratory work-up was conducted at the Department of Laboratory Medicine, Medical University of Vienna, and complement-specific analyses were performed at the Research Laboratory, Department of Internal Medicine and Haematology 3rd Department of Internal Medicine and Haematology, and Research Group for Immunology and Haematology, Semmelweis University – Eötvös Loránd Research Network (Office for Supported Research Groups), Budapest, Hungary [7]. Genetic analyses were performed by the same institutions [6]. No additional genetic or laboratory test was performed for the purpose of this study.

All patients that were hospitalized with a TMA episode were routinely tested for SARS-CoV-2 by RT-qPCR. Furthermore, hospitalized patients in our institution and other hospitals were routinely tested at least once per week for the presence of SARS-CoV-2. SARS-CoV-2 RT-qPCR testing for the general population – including asymptomatic individuals – was widely available, free of charge, and routinely performed and documented as measure to maintain work, allow social contact, and contain spread of the disease. This allowed for conclusive assessment of SARS-CoV-2 infections in the patient group investigated. As we, in addition, performed personal interviews with the participants during the observation period, we do not believe that we have missed mild to severe COVID-19 episodes or hospitalizations for TMA in other hospitals among study participants. Of course, we could not rule out subclinical TMA episodes related or unrelated to COVID-19 or SARS-CoV-2 vaccination.

The data are presented as count and frequency, as mean and standard deviation or as median and interquartile range, as appropriate. Fisher’s exact test was used to calculate differences in the proportions of categorical variables and odds ratios. FU time ended at May 5, 2023. We used a 95% confidence interval, and a two-sided p value of <0.05 was considered significant. We used MS Excel and GraphPad Prism 9 for data management and analysis and GraphPad Prism 9 for the generation of figures.

By the end of February 2020, the Vienna TMA cohort consisted of 219 patients with a TMA including 55 patients with a diagnosis of aHUS/cTMA. During FU until May 2023, we enrolled 7 further patients with a new diagnosis of aHUS/cTMA into the Vienna TMA cohort. The total FU time for these new patients was 1.76 ± 0.24 years (total amount: 12.32 years) from first TMA to the end of the FU. The patient characteristics are specified in Figure 1. Demographic data, SARS-CoV-2 vaccinations, and episodes of COVID-19 are summarized in Table 1.

All patients received at least one dose of a SARS-CoV-2 vaccine. Those were widely available in Austria to the general population during the second quarter of 2021. A total of 19 doses were administered in 7 patients. One patient received one dose, 2 patients received two doses, 3 patients received 3 doses, and one patient received a total of five doses. The most commonly used vaccine was COMIRNATY (Pfizer, New York City, NY; n = 14, 74%) followed by Spikevax (Moderna, Cambridge, MA; n = 3, 16%) and Jcovden (Janssen, New Brunswick, NJ; n = 2, 10%).

Of the 7 patients with new-onset aHUS/cTMA during the COVID-19 pandemic, five (71%) were tested positive for SARS-CoV-2 by RT-qPCR during the 3-year FU. All infections were mild to moderate, with one requiring hospitalization, because of a concomitant acute kidney failure.

Between March 2020 and May 2023, a total of 7 patients experienced their first aHUS/cTMA episode. No patient experienced a TMA relapse or more than one episode after the initial diagnosis during the FU period. Figure 2 shows the individual course of all 7 patients including SARS-CoV-2 vaccinations, COVID-19 episodes, and complement therapies since their first TMA episode. One patient died after informed and consensual discontinuation of haemodialysis.

Three TMA episodes were in close temporal connection and therefore attributable to either COVID-19 (n = 1; 33%) or SARS-CoV-2 vaccination (n = 2; 66%), respectively (Table 2). All 3 patients had systemic signs of TMA, and TMA was confirmed by kidney biopsy in all cases. The other four TMA episodes not related to a COVID-19 infection or SARS-CoV-2 vaccination included three pregnancy-associated aHUS/cTMA cases and one aHUS/cTMA patient with a family history of aHUS/cTMA. Three of those patients had the first TMA episode before any recorded exposure to either virus or vaccination, and 1 patient had received three doses of a SARS-CoV-2 vaccine and had mild COVID-19 3 months before an episode of pregnancy-associated aHUS/cTMA.

The complement profile of all patients included in this report is presented in Table 3. We found known disease-associated genetic variants in complement factor H in 2 patients (in one associated with familial aHUS/cTMA and in another with aHUS/cTMA after a SARS-CoV-2 vaccine).

One patient that suffered a TMA episode after a SARS-CoV-2 vaccination received an additional immunization with an mRNA vaccine under complement inhibitory therapy and showed no further signs of TMA thereafter.

Among the 7 patients, we recorded five infections that triggered one TMA episode (20%) and 19 vaccinations triggered two TMA episodes (10%; p = 0.52, odds ratio 0.47; 95% CI: 0.04–8.39). In the 3 years before the outbreak of the pandemic, a total of 11 patients were diagnosed with aHUS in our centre, while during the pandemic a total of 7 patients were newly diagnosed with aHUS (p = 0.13).

This 65-year-old man came to the emergency department with nausea and severe hypertension 12 days after he had received a SARS-CoV-2 vaccination (Jcovden, Janssen, New Brunswick, NJ). The laboratory work-up showed acute kidney injury without haemolysis. A renal biopsy 9 days after admission demonstrated severe scarring, chronic and active TMA, and subtle C3 deposits by indirect immunohistochemistry. Electron microscopy showed intramembranous and subendothelial deposits, compatible with C3 nephropathy. Serological evaluation of the complement system showed a decreased concentration of C3 and of CFH, decreased activity of the alternate complement pathway, as well as a highly elevated sC5b-9 concentration. Thus, these results support the presence of complement activation, with signs of alternative complement pathway dysregulation. Genetic testing revealed a pathogenic, heterozygous frameshift duplication in exon 5 of CFH (c.617_618dupTG, p.Glu207Trpfs*11). Despite treatment with complement blockade, he remained dependent on dialysis. The patient received one dose of COMIRNATY (Pfizer, New York City, NY) while on ravulizumab therapy and showed no further signs of TMA. The patient died during FU, because he refused to further undergo haemodialysis.

This 60-year-old woman presented with general malaise to a rural hospital 6 months after mild COVID-19 and 2 weeks after her first SARS-CoV-2 vaccination (COMIRNATY, Pfizer, New York City, NY). She was referred to our centre because of laboratory signs indicating TMA. A kidney biopsy confirmed the clinical diagnosis, showing arteriolar TMA. Additionally, serum C5b-9 was elevated. She received a total of 5 sessions of plasma exchange, to which she responded well. The patient currently has impaired kidney function. Genetic testing revealed the CFH-H3 and the CD46ggaac risk-haplotypes, both heterozygous [8].

This 61-year-old female patient with a history of kidney transplantation due to anti-glomerular basement membrane disease presented with acute kidney injury, microangiopathic haemolysis, and biopsy-proven glomerular and arteriolar microangiopathy during an episode of moderate COVID-19. She responded well after 2 doses of eculizumab with a return of serum creatinine to baseline values and resolution of microangiopathic haemolysis. She had received a total of 5 SARS-CoV-2 vaccines and had measurable antibody levels. No systemic complement dysregulation was seen in this patient. Genetic analysis revealed a heterozygous variant of unknown significance in the complement factor B gene (p.Glu255Gly).

This single-centre cohort study covers patients with newly diagnosed aHUS/cTMA during 3 years of the COVID-19 pandemic. We included all patients with incident aHUS/cTMA and looked for the aetiology of the disease with a special emphasis on COVID-19 and SARS-CoV-2 vaccinations.

Both viral infections and vaccinations have been linked to the development of aHUS/cTMA in (genetically) vulnerable patients in the past [9‒11]. The molecular mechanism for the development of complement-mediated diseases with regard towards viral infections and vaccinations includes an augmented proinflammatory state that leads to disturbances in the complement and coagulation cascade [12]. Additionally, in vitro studies have shown a possible competitive blocking of cell surface binding of complement factor H by the SARS-CoV-2 spike protein and serum from COVID-19 patients was shown to be able to induce complement-mediated cell death and increased C5b-9 deposition on cell surfaces [13].

COVID-19 was associated with thrombotic microangiopathies in several case reports and series. Both the thrombotic thrombocytopenic purpura and the haemolytic uremic syndrome have been described. Treatment consisted of supportive measures, plasma therapy, and anti-complement agents depending on the clinical course of the disease. Outcomes were variable with some patients returning to baseline kidney function and others reaching end-stage kidney disease. Interestingly, the severity of kidney disease does not seem to correlate with the severity of COVID-19 in any reported cases or case series [2, 14‒16]. In the past, we described that relapses of aHUS/cTMA in patients with a previously established diagnosis of the disease were more frequently associated with COVID-19 than the SARS-CoV-2 vaccination [4]. Limitations include the small case numbers and the retrospective nature of this study.

Here, we report patients with a new diagnosis of aHUS/cTMA during the COVID-19 pandemic. Of those 7 incident patients, three cases were associated with either COVID-19 (n = 1; 14%) or a SARS-CoV-2 vaccine (n = 2; 28%). We could not detect a statistically significant difference for the risk of new TMA episodes after either COVID-19 or SARS-CoV-2 vaccinations (p = 0.52, odds ratio 0.47; 95% CI: 0.04–8.39). Of course, our case numbers are low, which is partly due to the rare nature of the disease. In our previous publication however, we were able to show that COVID-19 episodes are more frequently associated with aHUS/cTMA relapses, than SARS-CoV-2 vaccines [4]. We cannot make a statement if a specific type of vaccine is more prone to trigger TMA episodes than another. This is both due to the low case number and to the fact that TMA episodes were attributed to both mRNA and vector-based vaccines in this publication and also our previous one [4]. However, literature suggests that mRNA vaccines are more prone to cause TTP than HUS and vector-based vaccines are more prone to cause HUS. This may be caused by vector-specific properties resulting in complement activation. In addition, it has been shown that adenoviral-based vaccines can directly target CD46 and thus impair complement regulation on cell surfaces [17].

In general, SARS-CoV-2 vaccination-associated cases of aHUS were milder and showed a better outcome than COVID-19-associated cases in the literature [18]. We speculate that both SARS-CoV-2 vaccinations and infections can represent triggering factor for aHUS/cTMA episodes in vulnerable individuals. In patients with COVID-19-associated aHUS, genetic complement variants were reported in about 50% of patients, which is a similar number as compared to the previously reported general aHUS population. COVID-19 therefore seems to be a viral trigger factor for aHUS in a vulnerable population. In our 3 patients with COVID-19 or SARS-CoV-2 vaccination-associated aHUS, we could not detect any disease-causing genetic variants. However, in our previously reported population of aHUS patients we recorded four COVID-19 or SARS-CoV-2 vaccination-associated aHUS flares among 27 incident aHUS patients [4].

TMA is a rare but potentially life-threatening complication of COVID-19 and, probably to a lesser extent, SARS-CoV-2 vaccinations. Clinicians need to be aware of this complication in patients newly presenting with TMA episodes and always thoroughly record patient’s disease history. The incidence of new aHUS cases did not differ from the pre-pandemic era in a large tertiary care centre cohort.

We gratefully acknowledge the help of Dorottya Csuka, Ágnes Szilágyi, Katalin Marossy, György Sinkovits, Márta Kókai, Éva Zsuzsanna Szendrei, Lászlóné Kertész, Edina Szabó, and Beáta Takács with complement and genetic analyses.

The Ethics Committee of the Medical University of Vienna approved the study (unique identifier: 1368/2014). All patients provided written informed consent, and the study was conducted in accordance with the principles of the Declaration of Helsinki.

Dr. Aigner has received lecture fees from Novartis. Dr. Gaggl has received lecture fees from Alexion. Dr. Kain has received personal fees from Takeda, Jansen, and Vifor and is a council member of the Renal Pathology Society and the European Society of Pathology. Dr. Kozakowski has received research funding from Astellas, Chiesi, and the Medical Scientific Fund of the Mayor of Vienna and personal fees from Sanofi. Dr. Gere Sunder-Plassmann has received funding from Amicus, Chiesi and Sanofi. None of the aforementioned funding was in relationship with the presented manuscript.

This project was funded by the “Medical Scientific Fund of the Mayor of the City of Vienna” (project number: 17001, principal investigator: Christof Aigner).

C.A., M.G., A.S., and G.S.-P. designed the study and analysed the data; C.A. and S.S. collected the data; C.A. designed the figures; C.A., M.G., S.S., A.S., R.K., N.K., A.O., Z.P., R.S.-P., and G.S.-P. drafted and revised the paper. All authors approved the final version of the manuscript.

The data that support the findings of this study are not publicly available due to their containing information that could compromise the privacy of research participants but are available from the corresponding author (C.A) upon reasonable request.