Study Design
There was a potential of continuous improvement in the 900 mg group at endpoint of phase 2 trial, so the 24-week treatment duration may not be long enough to witness the effectiveness of GV-971. Therefore, it is necessary to increase the treatment duration in phase 3 trials of GV-971[16]. This phase 3 study was a 36-week multicenter, randomized, double-blind, placebo-controlled parallel-group clinical trial. Participants with mild-to-moderate AD were assigned randomly (1:1 ratio), to receive GV-971 (450 mg, b.i.d.) or placebo.
The trial protocol was approved by the Ethics Review Board of Shanghai Mental Health Center (Shanghai, China) and is registered on https://clinicaltrials.gov/NCT02293915. The protocol was also approved by the Institutional Review Boards of all participating sites. The Clinical New Drug Research and Development Team of the Department of Psychogeriatrics of Shanghai Mental Health Center and the sponsor designed the study in consultation with academic advisors. All participants or their representatives provided written informed consent before participation in the trial.
Data were gathered by the study investigators through the Merge system (www.merge.com/eClinical), analyzed by IQVIA (Durham, NC, USA) after the data were locked, and interpreted by the academic main investigators in collaboration with the sponsor. The academic authors had attest to the accuracy and integrity of the data and the fidelity of this report to the study protocols, which are available with the full protocol (protocol number: 971-III; version 7.1) and statistical analysis plan (supplementary appendix).
There were 8 protocol amendments and 4 protocol versions in the course of the trial. A planned 24-week interim analysis was deleted, and the study continued to its primary outcome at 36 weeks. The other amendments included deletion of the age-adjustments on the interpretation of the Fazekas scale of allowable white matter pathology on magnetic resonance imaging (MRI) and adjustment in how routine laboratory studies were interpreted. The subject whose Mini-Mental State Examination (MMSE)[17] fluctuation was more than 2 score from screening visit to randomize visit should be double checked according to the inclusion/exclusion criteria in order to accurately enroll the subjects participant. Allowable heart rates were broadened (to > 55 beats per minute) and apolipoprotein E (APOE) genotyping was made mandatory. The intended sample size of 788 was somewhat over-recruited resulting in a final sample size of 818 randomized subjects.
Participants
Participants were aged 50–85 years of age and met the diagnostic criteria for probable AD according to the National Institute of Neurological and Communicative Disorders and Stroke and the Alzheimer’s Disease and Related Disorders Association[18]. Participants had mild-to-moderate AD, with a MMSE score from 11 to 22 inclusive for participants with only a primary-school education, and from 11 to 26 inclusive for those with an education beyond primary school, in line with representative China national sample of age-, gender-, education- reference norms [17, 19]. A total Hachinski Ischemia Scale[20] score of ≤ 4 was required, and the Hamilton Hamilton Depression Scale 17[21] score had to be ≤ 10.
In phase 2 trial, more strict MRI markers associated with AD were not included as a part of the inclusion/exclusion criteria for AD diagnosis at the protocol design. The lack of these markers may lead to some bias in AD diagnosis and lead to the result of no significant difference on the primary endpoint of ADAS-Cog changes between 900 mg group and placebo group[16]. So we strengthened brain MRI assessment in the phase 3 trial. During screening, brain magnetic resonance imaging (MRI) with oblique coronal hippocampus images was undertaken; a medial temporal lobe atrophy visual rating scale score ≥ 2 was required[22]. MRI data were sent to an Imaging Advisory Team via email in digital imaging and communications in medicine format along with a brief medical history. A Fazekas scale[23] for white-matter lesions of grade ≥ 3 (moderate-to-severe) or > 2 lacunar-infarction lesions of diameter 1.0–2.0 cm or infarction lesions in vital brain areas (thalamus, hippocampus, or entorhinal cortex) excluded participation. The brain MRI results of all participants were presented for diagnostic review by the Imaging Advisory Team to minimize diagnostic error. The investigators reviewed the laboratory-examination results and comments from the Imaging Advisory Team on the brain MRI examination to ascertain if participants met the enrollment criteria.
Female participants were postmenopausal (last menopause at least 24 weeks prior to study entry), surgically sterilized, or of childbearing age who agreed to use contraceptive measures during the trial. Women of childbearing age and women < 24 weeks from the start of menopause underwent a urine pregnancy test during screening, and the result was required to be negative for study entry.
Participants were required to have completed education of at least primary-school level or higher and be able to complete the protocol-specified cognitive tests. Progressive impairment of memory must have been present for ≥ 12 months. Care partners for the participants had frequent contact (≥ 4 days every week for ≥ 2 h on each of these days). Caregivers had to be willing to provide critical trial data on caregiver-based scales: CIBIC+[24, 25], Alzheimer’s Disease Cooperative Study-Activities of Daily Living (ADCS-ADL) scale[26] and Neuropsychiatric Inventory (NPI)[27, 28]. Before implementation of a protocol-related procedure or examination, participants and caregivers provided written informed consent. If participants could not sign due to limited cognition, their legal guardians signed on their behalf.
Participants were excluded if they had taken part in another clinical trial < 30 days before initiation of this trial or had dementia due to non-AD causes. Participants had a normal neurologic examination. They were excluded if they had abnormal laboratory values, including: liver-function tests (alanine transaminase [ALT] or aspartate transaminase [AST]) > 1.5-times the upper limit of normal (× ULN); creatinine > 1.5 × ULN; white blood cell counts, platelet counts, or hemoglobin levels below the lower limit of normal; or blood glucose > 1.5 × ULN. Participants with systolic blood pressure ≥ 160 mmHg or < 90 mmHg or diastolic blood pressure ≥ 100 mmHg or < 60 mmHg during screening were excluded from the trial. They were also excluded if they had unstable or severe cardiac, pulmonary, hepatic, renal or hematopoietic disease (including unstable angina, uncontrolled asthma, active gastric bleeding or cancer), a resting heart rate < 55 bpm after 10 min of rest, a visual/hearing disorder that prevented completion of neuropsychologic tests and scale evaluations, or a history of alcohol abuse, drug abuse or severe psychopathology (including major depression). Participants could not be on cholinesterase inhibitors or memantine while enrolled in this trial and were required to be off these agents for ≥ 4 weeks before randomization. The investigators excluded participants who they thought could not complete this trial, who participated in the Phase 2 GV-971 trial, who were on AD therapies that could not be stopped, or were relatives of staff of IQVIA or Shanghai Green Valley Pharmaceuticals.
This clinical trial was conducted at 34 participating sites in the psychiatry, neurology, and geriatric departments of hospitals in several regions of China. [18F]-FDG-PET was carried out at two sites (Beijing and Shanghai) where appropriate technology and expertise were routinely available.
Interventions
One capsule provides 150 mg of GV-971. The placebo is identical to the GV-971 capsule with regard to taste, odor, appearance and design, and is acceptable based on the quality-standard inspection approved by the National Medical Products Administration (NMPA). GV-971 capsules are manufactured according to Good Manufacturing Practice (GMP).
Two groups were intervened by GV-971 and placebo, respectively. The study comprised a 2-week screening period, 4-week run-in period, and 36-week treatment period. During the run-in period, each participant took three placebo capsules b.i.d. During the 36-week treatment period, each participant took three GV-971 capsules (150 mg/capsule) or three placebo capsules b.i.d.
Drug distribution was undertaken according to the drug number generated by an interactive web response system (IWRS). After participants were determined to be eligible for trial participation, investigators logged into the IWRS system to obtain a randomization number and drug number for each participant. During the 36-week treatment period, participants were randomized, (1:1 ratio), to receive GV-971 (450 mg, b.i.d.) or the matching placebo. During each on-site visit (Figure S1), investigators recorded the distribution and return of trial drugs. Participants, caregivers, and trial staff remained blinded to treatment assignment throughout the trial.
Outcomes
The primary efficacy endpoint was the drug–placebo difference in the change from baseline on the ADAS-Cog12 (score range, 0-75, with higher scores indicating greater cognitive impairment) at week36[29]. Secondary endpoints were intergroup differences in changes from baseline in the CIBIC+[24, 25], ADCS-ADL scale (score range, 0-78, with lower scores indicating worse functioning)[26] and NPI[27, 28]. Intergroup differences in change from baseline in a global index defined for relative cerebral metabolic rate for glucose (CMRglu) on [18F]-FDG PET[30] after 36 weeks of double-blind treatment was measured in a subgroup of participants. There was no change in the choice of outcome measures in the course of the trial.
All adverse events (AEs) that occurred during drug treatment were captured on case report forms. AE evaluation comprised classification of the organ system, grade, relationship to drug exposure, treatment measures required, and the outcome.
Ten on-site visits and four telephone interviews were conducted with each participant. The on-site visits occurred at weeks −6 and −4, day 0; and weeks 4, 8, 12, 16, 24, 36, and 40. Telephone interviews were undertaken at weeks 2, 20, 28, and 32.
Sample Size
Sample size was calculated with nQuery Advisor 7.0. According to the results of the GV-971 Phase 2 trial and other anti-AD clinical trials, a minimum ADAS-Cog drug-placebo difference of ≥ 1.4 was assumed. Completion of the trial by 315 participants in each arm would provide 80% power to detect an effect size of ≥ 0.2 between GV-971 and placebo groups with a two-sided alpha level of 0.05. Therefore, we planned to enroll 788 participants, allowing for an anticipated 20% dropout rate. In total, 818 participants were randomized.
Data Statistics and Study Parameters
The primary endpoint (drug-placebo difference in change from baseline of the ADAS-Cog12 score at week-36) was analyzed using analysis of covariance (ANCOVA) with the treatment group, education level, pooled center, and age group as fixed effects and the baseline MMSE score and baseline ADAS-Cog12 score as covariates. Control-based pattern imputation was used to supply the missing data in the primary analysis. In general, this method assumed that, after trial withdrawal, participants from the treatment arm would exhibit the same future evolution of the disease as participants receiving control treatment. Participants who discontinued the trial from the control arm were assumed to evolve in the same way as control participants who remained in the trial. In addition, a mixed-model repeated-measures model (MMRM) was used for sensitivity analyses. The primary efficacy analysis was based on the full analysis set (FAS), which included all randomly assigned participants who received at least one dose of the double-blind study drug and had a baseline value and at least one post-baseline efficacy assessment.
For secondary endpoints, CIBIC + was analyzed by a stratified Cochran-Mantel-Haenszel test for present of grade change from baseline, including the stratification factors of MMSE at baseline, education level and age group. ANCOVA was used to assess the drug-placebo difference in the change from baseline on ADCS-ADL and NPI scores.
Safety analyses were based on the safety analysis set, which included all randomly assigned participants who received at least one dose of a study agent. Safety analyses were based on a summary of AEs, laboratory assessments, electrocardiograms, vital signs and physical examinations.
Participants in the cerebral [18F]-FDG-PET sub-cohort underwent PET at baseline and week-36. We used the statistical region of interest (sROI) approach to compute the relative global CMRglu global index[30]. sROI was introduced based cross-validation and ADNI data, and was used in a recent clinical trial[31]. The sROI defined global CMRglu index is actually the standard uptake value ratio between a set of regions affected by AD and a set of regions spared by AD in terms of glucose uptake decline over time[30].
SAS v9.4 (SAS Institute, Cary, NC, USA) was used for statistical analyses. P < 0.05 (two-sided) indicated significance.