Study procedures
As a means of evaluating activities of daily living in this cognitively impaired patient population, the Amsterdam Instrumental of Activities of Daily Living questionnaire (A-IADL-Q) (22) (32) will be employed. This instrument is completed by the study partner, hence study participants are required to have a study partner able to accompany them on visits to clinic. The screening for the trial consists of several elements in order to ensure adherence to the inclusion and exclusion criteria (see table 1). A medical history, physical and neurological examination, vital signs, ECG, and cognitive measures including the WAIS-IV Coding test, to test executive function and working memory, the MMSE and a Neuropsychological Test Battery (NTB) (23–25)(33–35) composed of CogState tests of episodic verbal memory (the International Shopping List Test), episodic visual memory (the One Card Learning test), working memory (the One Back test) and two measures of attention (‘Detection’ and ‘Identification’). A composite score of perfomance on these latter three tests is the primary efficacy measure for this study.
To assess AD-specific speech changes the Winterlight speech assessment (WLA) will be employed regularly to assess any change from baseline in the study participants (36). Also, blood will be drawn for hematology, blood biochemistry and assessment of blood biomarkers. Also, a lumbar puncture will be performed to assess inclusion criteria in the form of AB42 and p-Tau-levels, and have an extra matrix to assess longitudinal pharmacodynamic effects of varoglutamstat by means of investigational biomarkers. A cerebral MRI will be performed to rule out relevant cerebral vascular disease, or other pathological changes.
Upon passing screening, the first 90 participants will be randomized 1:1:1 by a computerized allocation system to receive either PQ912 300mg BID, 600mg BID or placebo BID following a 12-week escalation period. Escalation occurs in up to five steps; weeks 1–2: 50 mg/placebo once daily, weeks 3–4: 50 mg/placebo BID, weeks 5–8: 150 mg/placebo BID, weeks 9–12: 300 mg/placebo BID, weeks 13–24: 300 mg/placebo or 600 mg/placebo BID, in order to minimize possible side-effects. At the baseline visit, study participants will be assessed again on the NTB, WAIS-IV coding test and the WLA. A full physical examination will also be performed, as well as en EEG and blood draws for hematology, biochemistry and apolipoprotein E (ApoE), Human Leukocyte Antigen (HLA) and CYP2c19 genotyping.
As illustrated in Fig. 1, the participants will be seen in the clinics 4 weeks after baseline and assessed for AE’s, drug accountability and vital signs. In addition, blood will be sampled for hematology and biochemistry and a physical exam will be conducted. At week 12, the above procedures will be repeated as well as the NTB, WLA and WAIS-IV Coding test. These procedures will be repeated every 12 weeks.
The participants attend clinic visits once every 12 weeks until study termination, where several key measurements are made at timepoints, including physical and neurological examination, WAIS-IV Coding test, WLA, NTB, Amsterdam IADL-Q. Every 24 weeks blood will be analyzed for biomarkers. Blood levels of PQ912 will be measured at week 12, 24 and 48 as well as at EOT, for study participants enrolled early in the trial. In addition, an EEG will be performed at week 24 and 48 and an additional cerebral MRI will be performed at EOT. CSF will be collected for biomarker measurement at week 48.
Should the patients experience significant intolerable side effects (starting from a maximal dose of 300 mg BID/placebo or above), the investigators may decrease drug dosage by 50% for a period or temporarily suspend treatment with IMP. Should treatment-naïve study participants progress significantly in their symptoms during the study, initiation of standard AD medication is allowed.
Endpoints
The safety analysis will be based on significant changes on i) physical and neurological examinations, ii) significant changes on blood samples, iii) amyloid-related imaging abnormalities (ARIA) and iv) reported adverse events.
The primary efficacy analysis consists of the pooled Z-score of the CogState ‘Detection’, ‘One Back’ and ‘Identification’ tests (see ‘Statistical Evaluation’ for method of evaluation).
In addition to the primary analysis of efficacy and safety evaluation, several secondary and exploratory endpoints have been defined for this study. Secondary endpoints have been defined as linear change in overall cognition based on the complete NTB and in the CogState battery by itself, using the same statistical model as the primary efficacy analysis. At week 48, changes in global relative theta power (4–8 Hz) in the EEG-data as well as changes from baseline score of A-IADL-Q will also be analyzed.
As an exploratory endpoint neuronal activity will be evaluated in the same manner as described by Briels et al. 2020 (26) (36), Poil et al. 2013 (27) and Scheltens et al. 2018 (19) in regards to global relative power in the delta (0.5–4 Hz), alpha (8–13 Hz) and beta (13–30Hz) frequency bands, looking into global posterior dominant peak frequency, Amplitude Envelope Correlation (AEC) in the 4–13 Hz band and functional network topology measures such as centrality, modularity, minimum spanning tree in an attempt to acquire new EEG AD-specific biomarkers in this modality.
Besides collection at screening and baseline, blood for biomarker measurement will be collected at week 24, 48 and the EOT-visit, whereas CSF will be collected at screening and at EOT. There will be a broad assessment of biomarkers with potential use for quantifying neuroinflammation, synaptic toxicity and neurodegeneration. The panel of biomarkers assessed will include levels of: YKL-40 (inflammatory marker), Neurogranin (synaptic marker), Beta secretase 1 protein [BACE-1], Tau and pTau, Protein fragments of Tau, GFAP and extracellular matrix (ECM) molecules (neurocan, brevican and Tenascin-R). In addition to the earlier mentioned well-established biomarkers, exploratory biomarkers in both plasma and CSF will be investigated e.g. Neurofilament light chain (NFL); protein fragments of Tau, GFAP and extracellular matrix (ECM) molecules (neurocan, brevican and Tenascin-R); pGlu-peptide substrates of QC (e.g. pGlu- and total C-C motif chemokine ligand 2 [CCL2], Orexin A) and Aβ peptides (including full length and truncated A peptides) will be assessed.
Statistical evaluation
The main statistical evaluation will be performed in the Full Analysis Set population, such that all participants who are randomized and received at least one dose of study medication will be included in the main statistical analysis.
An interim analysis is planned to be performed shortly following the 90th participant reach 24 weeks post-baseline. At this point a data safety monitoring board (DSMB) consisting of experts within relevant medical and scientific fields will conduct an unblinded safety analysis, in order to decide on study continuation and secondly which dose of investigational medicinal product (IMP) should be used for the remainder of the study.
As the study is planned to have differences in participation duration and dosage, a linear model of the primary efficacy parameter will be employed to assess effectiveness. The primary efficacy analysis will consist of the pooled Z-score based on the cognitive scores from the CogState ‘One Back’, ‘Identification’ and ‘Detection’ tests (see Fig. 2). The two different dose groups will be pooled and the slope coefficient of the Z-score over time will be compared to that of the placebo group. This approach enables incorporation of all data on participating individuals, irrespective of their follow-up time, thus ensuring a maximum amount of information obtained from study participants over the course of the trial.
Based on extrapolation of the results in the SAPHIR study, evaluation of other empirical data, and expert opinion, it was determined that at 48 weeks, an effect size of approximately 0.35 is a reasonable expectation. The effect size, Cohen’s D, of 0.35 corresponds to an abolishment of the decline in the combined Z-score for cognition in the PQ912 treated study participants. Based on the assumptions above, and an allocation ratio of 11:14 between active and placebo, by comparison via a two-sample T-test, a total sample size of around 250 should provide a power close to or exceeding 80%.
The sample size is not adjusted for the rate of drop-out as it is anticipated that all participating individuals will have at least two measurements of the combined Z-score, irrespective of their follow-up time, and therefore contribute to the analysis of the primary efficacy endpoint.