Study design
This is a single-center, prospective, double-blind, placebo-controlled randomized trial. This clinical trial is reported according to the Standard Protocol Interventions: Recommendations for Interventional Trials (SPIRIT) guidelines [24] [study schedule (SPIRIT figure) is outlined in Figure 2, checklist see Additional file 1].
Setting and participants
Sixty high-risk IgAN participants will be followed up until 50% (30 of them) have a composite endpoint or been followed for 3 years. The trial will be conducted at Guang’anmen Hospital, Beijing, China, and was approved by the Ethics Committee of Guang’anmen Hospital (approval number: 2018-055-KY-01) in accordance with the Declaration of Helsinki and the principles outlined in the “Guidelines for Good Clinical Practice” from the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use Tripartite Guideline (January 1997). Participants will be recruited self-selected or referred from inpatients or outpatients, and hospital-based WeChat advertisement. Potential participants will be pre-screened through WeChat, further assessed by our investigator, and consented for the study. The trial will be conducted with all subjects voluntarily and signed informed consent.
Objectives
Based on optimal supportive care, the trial is aiming to assess the superiority in renal protection and reduction of severe treatment-related adverse events using YQF combined therapy, compared with immunosuppression monotherapy in high-risk IgAN.
Rationale of high-risk IgAN
Several high-quality clinical trials mentioned above [11,12] have respectively interpreted the concept of “high-risk” clinically and pathologically while not specifically. Drawn from the inclusion criteria of above-mentioned trials, this study will define "high-risk" as persistent heavy proteinuria (≥1g/d despite intensive optimal supportive care) with impaired renal function (eGFR 15-60 ml/min/1.73m2).
Inclusion and exclusion criteria
The inclusion criteria are as follows: (1) in accordance with IgAN pathological diagnosis, renal biopsy within 6 months; (2) persistent proteinuria≥1g/d despite at least 8 weeks of optimal supportive care [maximally tolerated RAS blocker which refers to no symptomatic hypotension, no hyperkalemia, and serum creatinine (SCr) increased not more than 30% of baseline, blood pressure control meeting targets (135/85 mmHg or lower), and dietary management (sodium intake less than 6g/d, protein intake of 0.6-0.8g/kg/d, and low-fat diet)]; (3) eGFR 15 to 60 ml/min/1.73 m2, calculated with the use of CKD-EPI Creatinine Equation 2009; (4) patients who maintain regular follow-up at Guang’anmen Hospital, agree to participate and obtain informed consent. (Additional file 2 shows the Informed Consent Form in more detail)
The exclusion criteria are as follows: (1) secondary IgAN; (2) comorbidity of other primary or secondary glomerular diseases; (3) comorbidity of severe primary diseases such as cardiovascular, hepatic, cerebral, hematopoietic system diseases or mental disorders; (4) allergy or intolerance to the experimental medication (e.g., RAS blockers, prednisolone, cyclophosphamide, YQF compound and its placebo compound) ; (5) contraindications of immunosuppression therapy: acute and chronic infectious diseases, malignancies, leukopenia, thrombocytopenia, gastrointestinal hemorrhage, ulcers of stomach or duodenum, post-transplantation; (6) pregnant or lactating women; (7) unwilling to participate in this study, failure to accept or tolerate Chinese medicine compound; (8) history of alcohol or drug abuse; (9) poor compliance, loss to follow-up; (10) participation in another clinical investigation.
Randomization and masking
In the case that the subjects are eligible to participate and have signed informed consent, 30 patients in each group of the YQF group and the control group will be randomly selected in 1:1 ratio after run-in phase. A central stochastic system developed by the Institute of Basic Research in Clinical Medicine, China Academy of Chinese Medical Sciences, which based on SAS 9.4.3 (SAS Institute, Cary, NC, USA), will perform the randomization. Independent experts will generate the randomization sequence which will be concealed in opaque, sealed and stapled envelopes. Participants, investigators, and other all members with clinical involvement in the trial will be masked to the treatment assignment for the duration of the trial. Relevant personnel have clear division of labor and strict permission restrictions. The masking will be removed only if the participant occurs severe side effects that need to terminate the trial.
Interventions
Run-in period (pre-trial, 4 weeks)
Eligible participants will enter a run-in period for 4 weeks, during which they will receive an optimized basic treatment, including living behavior management (smoking cessation, alcohol restriction, weight control, low-salt and proper protein diet), maximum tolerated dose of ACEI or ARB, blood pressure control to a target below 135/85 mmHg, controlling glycated hemoglobin≤7% by using insulin or oral hypoglycemic agents in diabetics, reaching targets in serum uric acid (UA) (<420umol/L in male, <360umol/L in female) by uric acid-lowering medications in hyperuricemia, and stopping other Chinese medicine treatments.
Treatment period (in-trail, 48 weeks)
At the end of the run-in period, participants who fulfill all eligible criteria and no exclusion criteria, will be randomized to either the YQF combined immunosuppression therapy (YQF group) or matching placebo combined with immunosuppression therapy (control group) in a double-blind fashion with a total treatment period of 48 weeks. Both groups will continue their basic treatment as mentioned above in the pre-trial phase.
Immunosuppression therapy: oral prednisolone (0.5-0.8 mg/kg/d, exact dose decided by the investigator, maximum dose not exceeding 60 mg/d) for 8 weeks, then tapered by 5~10 mg/d every 4 weeks, with a total treatment period of 24-32 weeks. Participants with persistent proteinuria≥1g/d after 8-weeks of corticosteroid monotherapy, will receive 0.8-1.0g of intravenous cyclophosphamide (CTX) every 4 weeks, total dose of not exceeding 8g (exact dose decided by the site Investigator). If severe CTX-related adverse events occur, such as alanine transaminase (ALT) exceeding the upper limit of 2 times, infections requiring hospitalization, granulocytes<3.0×109/L and platelets < 50.0×109/L, stop using CTX, symptomatically treated and record adverse events. The frequency of detection is increased to once every 2 weeks, and the trial is terminated if persistent infection or myelosuppression occurs.
YQF Formula Granule: obtained from Sichuan Xinlvyao Co. (Chengdu, Sichuan, China), manufacturing process complying with Chinese GMP. The compounds are blends of individual herbal extract from YQF formula (consist of: astragalus membranaceus, saposhnikovia divaricata(turcz.) schischk, Flos Lonicerae, Angelica Sinensis, Dioscorea Nipponica, hedyotis diffusa willd, rhubarb, Spatholobus Suberectus), dissolved in 150ml boiled water and taken orally twice a day.
YQF placebo: obtained from Sichuan Xinlvyao Co. (Chengdu, Sichuan, China), manufacturing process complying with Chinese GMP. Major component: malt dextrine, with similar appearance and the package, dissolved in 150ml boiled water and taken orally twice a day.
Follow-up period (post-trial)
All participants will continue their previous basic treatment agents.
Follow-up assessments
Participants will be visited at regular intervals, for a planned mean of at least 3 years until the end point occurs. In run-in period and treatment period, study visits occur every 4 weeks until week 24, and every 12 weeks face-to-face or by telephone in consideration of choice of the participants and investigators, till the end of the trial. The measurements will be performed at qualified laboratories during the follow-up period.
Laboratory measurements including: (1) urine tests: 24h urine total protein (UTP) measured using biuret method, albumin/creatinine ratio (ACR) ; (2) blood tests: serum creatinine, albumin (ALB), blood urea nitrogen (BUN), uric acid (UA), blood glucose (GLU), total cholesterol (CHO), electrolytes (K, Na, Cl), triglyceride (TG), alanine transaminase (ALT), aspartate aminotransferase (AST), hemoglobin (HGB), white blood cell count (WBC), and platelets (PLT).
General conditions inspection includes body weight, appetite, excretory functions, stamina, mobility, sleep, etc. Vital signs inspection includes temperature, respiration, pulse and blood pressure. Rational symptoms and chronic inflammation status will be recorded by investigators on case report forms (CRFs) as syndrome scores for effectiveness assessment. Each symptom is scored on a 4-point scale ranging from 0 (absent) to 3 (severe), and a total symptom score is calculated by adding together the values for all 3 symptoms. Similarly, we get a total chronic inflammation status score (see Additional file 5).
The final follow-up duration will be continued until the occurrence of primary end point or keeping visit up to 3 years (Figure 3).
Outcome measures definition
The primary composite end point is defined as the first occurrence of 40% decrease in eGFR from baseline eGFR, progression to continuous renal replacement, or death due to renal disease.
The secondary composite end points are defined as (1) main outcome measurement: the mean annual reduction in eGFR based on SCr (eGFR-slope); (2) proteinuria remission (prescribed as proteinuria <0.5g/d) at week 24, 36, 48 in treatment period, and month 6, 12, 24, or 36 if possible. The remission rate of symptoms and inflammation status will be evaluated at week 48, respectively.
Adverse events and safety
Adverse events and SAEs will be inspected at each follow up, including infections requiring hospitalization, thromboembolic events, hepatic dysfunction, hematopoietic disorders, fracture or osteonecrosis and new onset diabetes. SAEs are defined according to the definitions of Good Clinical Practice (GCP) by the China Food and Drug Administration (CFDA). The researcher will report the events to the principal investigator and the Ethics Committee within 24 hours of occurrence of the SAE, who will give a final decision on whether or not to continue the study treatment and identify appropriate support for the patients involved. Furthermore, treatment interruption due to any events and relevant measures will be recorded on the CRFs. The influence of all adverse events will be analyzed at the end of the trial.
Termination and withdrawal
Participants will be withdrawn from the trial in any of the following situations: (1) SAEs related with immunosuppression: severe infections (e.g., pulmonary infection requiring ventilatory support), serious granulocytopenia/thrombocytopenia (e.g., WBC<2.0×109/L, PLT<20×109/L); (2) participants or investigators fail to obey the study protocol; (3) participants behave poor compliance, experimental medications are taken less than 30%; (4) termination requested by data and safety monitoring board, ethics committee or study principal investigator: in the best interest of the participants, the investigator can request for termination of individual participation if the study treatment is of insignificant clinical benefit.
Sample size calculation
On basis of our recent trial from above [22], eGFR-Slope after 12-month treatment as the primary endpoint, we define δ=(μ1-μ2)/σ, σ as the pooled standard deviation (sample size calculation, see Additional file 4), where μ is the treated mean, respectively. We calculate δ=0.994, approximately equivalent as 1.0, according to the look-up table of counts (Medical Statistics Method, PH Jin, Shanghai Medical College Press, see Additional file 3), using α=0.05 and β=0.1, we require a sample size of 46 participants. Assuming 25% dropout adaptation and in view of n=30 as a rule of thumb for a small size clinical trial, we plan to recruit 60 patients for this study (30 patients per group).
Statistical analysis
Investigators will fill the CRFs as making follow-up observations by interviewing the participants. Paper CRFs will be interpreted into database by those investigators in order to ensure data validity. An independent statistician will unblind and extract clean data from final database for analysis after locking. Data analysis of treatment effect and safety follows the intention-to-treat principle. Investigators can access the final database only when both data analysis and study are complete. The endpoint events will be described by Kaplan-Meier method and the between-group difference will be compared using log-rank test. T-test will be used for scale variables, chi-square test or variance analysis will be used for binomial and nominal variables, nonparametric test will be used for ordinal variables. The eGFR-slope will be calculated as the individual slopes obtained from individual linear regressions of eGFR during the follow-up period. All analyses will be performed using SPSS software version 23.0 (IBM Analytics). Differences are considered to indicate statistically significant for 2-sided P < 0.05.
Quality control and monitoring
A Data and Safety Monitoring Board (DSMB) will be established to monitor the performance of the overall study, ensure safety and welfare of participants, and review quality of the data on regular meetings. The DSMB, which is independent of the present trial, consists of three members, including a clinical nephrologist, a statistician and an ethical specialist (see Additional file 6). The DSMB will assess that participants receive good clinical care and that safety concerns are interpreted and addressed appropriately. Moreover, the DSMB will make recommendations and decisions on any modification of the protocol, even termination of the study based on the interim analysis of efficacy and safety. In order to improve compliance with the protocol until completion, we will intensify patient management through one-to-one WeChat follow-up by designate researchers. Participants’ medical records (including CRFs, consent forms, etc.) will be archived securely and used only within the validity period.
Intention-to-treat analysis will be used for subjects who dropout or withdraw from the study, which includes every subject who is randomized according to randomized treatment assignment regardless of whether he/she receives the treatment of this group or not, should finally be included in the assigned group for statistical analysis of efficacy.
Last observation carried forward method will be applied in missing data of longitudinal observations.