Donepezil

Patients who met the consensus diagnostic criteria² for probable DLB were recruited from 48 psychiatric or neurological specialty centers throughout Japan from October 2007 to February 2010. Diagnosis of each patient was validated after discussion by the central committee. Outpatients (≥50 years old) with mild to moderate-severe dementia (10-26 on the Mini-Mental State Examination [MMSE]¹⁸ and Clinical Dementia Rating¹⁹ ≥0.5) and with behavioral symptoms (Neuropsychiatric Inventory-plus [NPI-plus] ≥8) were eligible. NPI-plus is a 12-item version of the NPI, with the original 10 items supplemented by 2 DLB-relevant domains of sleep and cognitive fluctuation.¹¹ , ²⁰ , ²¹ Patients had caregivers who routinely stayed with them at least 3 days per week and 4 hours per day, provided information for this study, assisted compliance with treatment, and escorted patients to required visits.
Exclusion criteria included Parkinson disease diagnosed at least 1 year prior to the onset of dementia; focal vascular lesions on magnetic resonance imaging or computed tomography that might cause cognitive impairment; other neurological or psychiatric diseases; clinically significant systemic disease; complications or history of severe gastrointestinal ulcer, severe asthma, or obstructive pulmonary disease; systolic hypotension (<90mmHg); bradycardia (<50m⁻¹); sick sinus syndrome; atrial or atrioventricular conduction block; QT interval prolongation (≥450 milliseconds); hypersensitivity to donepezil or piperidine derivatives; severe parkinsonism (Hoehn and Yahr score ≥ IV)²² ; and treatment with ChEIs or any investigational drug within 3 months prior to screening. ChEIs, antipsychotic agents, and antiparkinson drugs other than L-dopa or dopamine agonists were not allowed during the study.
Written informed consent was obtained from the patient (if at all possible) and his/her caregiver before initiating the study procedures. The study was conducted in accordance with the principles of the Declaration of Helsinki. The protocol was approved by the institutional review board at each center.

The AChE inhibitory activity was measured using the spectrophotometric method developed by Ellman, et al. [61 (link)], with slight modifications as suggested by Rhee, et al. [62 (link)]. Acetylthiocholine was used as the substrate to detect the inhibition of AChE. The reaction mixture contained 40 µL of Buffer C (Tris–HCl, 50 mM, pH 8, containing 0.1 M of sodium chloride and 0.02 M of magnesium chloride hexahydrate), 20 μL of the tested sample solution, 20 µL of AcSCh (15 mM, PBS pH 7.4), and 100 µL of DTNB (3 Mm, dissolved in Buffer C). The mixture was pre-incubated for 3 min at 25 °C. Finally, the enzymatic reaction was started with the addition of 20 µL of 0.5 U/mL AChE (137 U/mg solid) and then incubated at 25 °C for 30 min. The amount of product released was monitored in an EPOCH 2 (BIOTEK ^®) microplate reader every 1 min at 405 nm. All reactions were performed in triplicate in a 96-well microplate. Tested sample solutions from the essential oil were made by dissolving 10 mg in 1 mL MeOH. Two more dilutions were included in MEOH (10× dilution) to obtain final concentrations (1000, 100, and 10 ug/mL) of essential oil in MeOH. The IC₅₀ value was calculated by curve fitting of data (non-linear regression analysis, PRISM 8.0.1, GraphPad, San Diego, CA, USA). Donepezil was used as positive control. Any increase in absorbance because of spontaneous hydrolysis of the substrate was corrected by subtracting the absorbance at the end of pre-incubation from the absorbance measured after the addition of the enzyme. The IC₅₀ value was measured from the corresponding rate of the reaction curve with Graph Pad v8.0.1.5.

The ADCS-ADL-MCI is a functional evaluation scale that assesses the ability of patients to perform ADLs (with recall "In the past 4 weeks") through a structured questionnaire administered to the informant/carer by a physician or qualified rater [21 (link)]. The ADCS-ADL-MCI was derived from its parent measure, the ADCS-ADL [12 (link)], and adapted to be suitable for MCI patients. Two forms of the ADCS-ADL-MCI are the 18-item and the 24-item version. While the 24-item version was administered as part of the ADC-008 trial, the 18-item version is explored in the present study due to practical considerations (i.e., its use in current clinical trials such as the phase 3 aducanumab trials EMERGE [NCT 02484547] and ENGAGE [NCT 02477800]). Items in the measure predominantly include IADLs, such as balancing a checkbook, navigating outside the home, shopping, using household appliances, or finding personal belongings. Physically getting dressed and the ability to be left alone are also being assessed. The 18-item ADCS-ADL-MCI is scored from 0–53 based on the subject's degree of independence in performing specific tasks (i.e., independent, partially independent, fully dependent). Except for item 3 (i.e., 'Regarding physically getting dressed, which best describes his/her usual performance in the past 4 weeks?', with response options ranging from '0 = Someone else dressed him/her' to '4 = Dressed completely without supervision or physical help'), all other items follow a gating response format [22 (link)]. That is, the first step of the item asks about whether the patient performed the specific task – and in case of endorsement – the second step usually follows the response format ranging from '1 = With physical help' to '3 = Without supervision or help' or a similar wording probing the details and manner of performing the specific task. Some items include four to five response options that describe how the IADL was performed (rather than three response options as in the more standardized response format for items 1, 2, 4, 5, and 7). Further, the second step of some items includes subquestions that are more descriptive regarding a particular activity (such as watching television), including only 'yes' and 'no' response options. Lower scores on the measure denote more substantial impairment [12 (link), 16 (link)]. In this study, the ADCS-ADL-MCI was assessed at baseline and months 6, 12, 18, 24 and 36 for patients with MCI. Per the ADC-008 trial design, patients who progressed to mild AD dementia were moved into an open-label study with donepezil, and where the ADCS-ADL-MCI was not included in that study design. Consequently, these patients are no longer included in the analysis after having progressed to mild AD dementia and, therefore, this study does not provide any information on the measurement properties of the ADCS-ADL-MCI as applicable to patients with mild AD dementia.

Chi-square for categorical variables and Mann–Whitney U test for continuous variables due to non-normality were used to compare the baseline characteristics between patients with RLS and RLS-free controls. Cox proportional hazards regression models were applied to explore the association between RLS and the risk of dementia after adjusting for age, sex, income, residence, CCI, and history of other comorbidities. Among the Cox regression models, we used the Fine–Gray subdistribution hazard model with mortality as a competing risk given the old age of the study population. The proportional hazard assumption was satisfied in our Cox model (Schoenfeld individual test p-value > 0.05).
Sensitivity analyses were performed using four different models. In model 1, dementia was defined as the prescription of anti-dementia medications (donepezil, rivastigmine, galantamine, and memantine) at least twice and a diagnosis of the ICD-code of dementia. Although these medications were approved for only AD (rivastigmine additionally for Parkinson’s disease dementia), they can be used for cognitive symptoms in other types of dementia based on recommendations from multiple guidelines [31 (link)–33 (link)]. The previous study revealed that the definition of all-cause dementia by ICD-10 code plus anti-dementia medications had a positive predictive value of 94.7% when reviewing the medical records of 972 patients in two hospitals [34 (link)]. In model 2, medication history was added to the ICD code to define RLS. Patients with RLS ICD-code (G25.8) who had taken dopamine agonists (ropinirole or pramipexole) twice or more were regarded as patients with RLS (n = 1458). In this sensitivity model, we excluded patients with Parkinson’s disease because they could also take dopamine agonists. In model 3, patients taking antipsychotic agents were excluded because the antidopaminergic property of antipsychotic agents could lead to a misdiagnosis of RLS (n = 2482). The following antipsychotic agents approved in South Korea were used in this study: haloperidol, sulpiride, chlorpromazine, perphenazine, pimozide, risperidone, olanzapine, quetiapine, paliperidone, amisulpride, aripiprazole, ziprasidone, clozapine, blonanserin, and zotepine. In model 4, patients with RLS only diagnosed by psychiatrists or neurologists were included (n = 1154) to preclude the possible misdiagnosis by non-expert physicians.
To evaluate the effect of dopamine agonists (pramipexole and ropinirole) on the development of dementia, the risk of dementia was compared after dividing RLS patients by dopamine agonist use. Patients with RLS who were prescribed pramipexole or ropinirole at least once were considered dopamine agonist users. All missing data were addressed using listwise deletion. Data processing and statistical analyses were performed using SAS version 9.4 (SAS Institute, Cary, NC, USA). Statistical significance was set at a two-tailed p-value of < 0.05.