The study was conducted at 10 accredited medical centers in Russia and included 32 patients from 23 to 71 years of age, with PNH confirmed by flow cytometry to assess the size of the PNH clone among red blood cells (RBC), granulocytes, and monocytes, with intravascular hemolysis and concomitant clinical symptoms at present or in past medical history with or without eculizumab administration. PNH clone testing was performed according to the previously described protocol of high-sensitivity flow cytometry . All patients were vaccinated against meningococcal infection with a tetravalent vaccine (Neisseria meningitidis serotypes A, C, Y, and W135). For naive patients, it was obligatory to exceed the LDH upper limit of normal by 1.5 times. The key exclusion criteria were the presence of diseases associated with bone marrow failure with the PNH clone (aplastic anemia, myelodysplastic syndrome, idiopathic myelofibrosis), previous infectious diseases caused by Neisseria meningitidis, active nonspecific infectious diseases, and completion of eculizumab treatment less than 70 days before the enrollment. Informed consent was obtained from all patients for being included in the study prior to any procedure.
A randomized, open-label, comparative, multicenter study was approved by the Ministry of Health of Russia (No. 546 dated October 17, 2017) and the Ethics Committee of MoH (No. 153 eff. date August 22, 2017) and was conducted in accordance with the ethical standards of the responsible committees on human experimentation (national and local) and with the Helsinki Declaration of 1975, as revised in 2013. The study consisted of the following periods: Screening up to 4 weeks, Treatment (26 weeks), and Follow-up (2 weeks).
Patients were allocated by variable block size (4 and 6 block random allocation sequence was generated by biostatistician in Stata14) randomization using the IVRS system with a 1:1 ratio into two groups: Group A, in which patients received the Biosimilar (Elizaria), and Group B, in which patients received the Originator (Soliris). Preliminary stratification was performed depending on the status of previous treatment with eculizumab (patients not receiving eculizumab/patients receiving a maintenance dose of eculizumab before enrollment in the study). Eculizumab-naive patients in both groups received the initial cycle of therapy consisting of four weekly intravenous administrations of the medicinal product at a dose of 600 mg, with subsequent maintenance therapy at a dose of 900 mg every 2 weeks. Patients previously treated with eculizumab started the treatment at the maintenance dose.
Evaluation of the pharmacokinetic (PK) parameters of the medicinal product was performed based on the determination of total (free and bound) eculizumab by biolayer interferometry (Octet® QKe System (Pall ForteBio)) with using Octet® Software, v.10.0 (Pall ForteBio). The following basic pharmacokinetic (PK) parameters were calculated: minimum product concentration (Cmin), maximum product concentration (Cmax), minimum product concentration at the end of the dosing interval after establishing a stationary distribution (Ctrough), area under the concentration–time curve throughout the dosing interval after establishing a stationary distribution (AUCt,ss). As additional PK parameters, the time to reach the maximum product concentration (Tmax), elimination constant (Kel), medicinal product elimination half-life (T1/2), mean retention time of medicinal product in the systemic circulation (MRT), clearance (Cl), and stationary volume of distribution (Vss) were calculated.
The analysis of pharmacodynamic (PD) parameters included evaluation of the membrane attack complex concentration in the blood serum (MAC, C5b-9) by the enzyme-linked immunosorbent assay (MicroVue Complement SC5b-9 Plus EIA MicroVue Complement SC5b-9 Plus EIA (Quidel, USA)) at the same time periods as for the PK assessment.
The safety of therapy was assessed by the incidence and severity of adverse drug reactions (ADRs) according to symptoms, physical examination, assessment of vital signs, electrocardiography (ECG), laboratory and instrumental studies, and patient diaries.
Immunogenicity was determined by the level of anti-drug antibodies, including the neutralizing activity of antibodies to eculizumab, by the method based on the bridging enzyme-linked immunosorbent assay. The evaluation was performed at 1, 5, 13, 21 weeks of treatment and after the end of the treatment.
All data was collected using the platform EDC system CSOnline version 7.5.501.1 by Ennov clinical®.
Comparative evaluation of hemolysis activity based on the area under the LDH concentration–time curve (LDH AUC) during the maintenance therapy with the investigational or reference product was the primary endpoint of the study. The secondary points included the area under the LDH concentration–time curve (LDH AUC) during 26 weeks of treatment, the number of packed red blood cells (pRBC) transfusions performed, the change in the values of the Functional Assessment of Chronic Illness Therapy-Fatigue Scale (FACIT-Fatigue) and the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire (EORTC QLQ-C30), the number/proportion of patients with various thrombotic complications, patients requiring pRBC transfusions, and patients with breakthrough hemolysis.
The standard methods available in Stata (StataCorp LLC), version 14, were used to analyze the data. The PK and PD data analysis was performed using the PkSolver software. LDH AUC during the period of maintenance therapy with the investigational or reference product was performed to assess the primary efficacy endpoint in terms of chronic hemolysis in a PP population. To demonstrate the difference in LDH AUC between the groups, a 95% confidence interval was built, which should not have crossed the noninferiority boundary, which is set in this study as 50% of the previously identified placebo-controlled effect at a value of 150,635 U/L*days. The χ2 criterion or Fisher’s exact test was used to analyze the secondary efficacy parameters, which are categorical variables. Descriptive statistics was used to analyze the secondary efficacy parameters, which are quantitative variables. For the intergroup analysis, Student’s t-test or Mann–Whitney test was used, depending on the type of data distribution. A normality test was performed using the Shapiro–Wilk test.