ORIGINAL RESEARCH published: 01 October 2021 doi: 10.3389/fneur.2021.705407 A Phase II Study to Evaluate the Safety and Efficacy of Prasinezumab in Early Parkinson’s Disease (PASADENA): Rationale, Design, and Baseline Data Gennaro Pagano 1*, Frank G. Boess 1 , Kirsten I. Taylor 1,2 , Benedicte Ricci 3 , Brit Mollenhauer 4,5 , Werner Poewe 6 , Anne Boulay 7 , Judith Anzures-Cabrera 8 , Annamarie Vogt 1 , Maddalena Marchesi 3 , Anke Post 9 , Tania Nikolcheva 10 , Gene G. Kinney 11 , Wagner M. Zago 11 , Daniel K. Ness 11 , Hanno Svoboda 1 , Edited by: Johannes Haybaeck, Markus Britschgi 1 , Susanne Ostrowitzki 10 , Tanya Simuni 12 , Kenneth Marek 13 , Innsbruck Medical University, Austria Martin Koller 11 , Jeff Sevigny 14 , Rachelle Doody 10 , Paulo Fontoura 10 , Daniel Umbricht 1 , Azad Bonni 1 and PASADENA Investigators and Prasinezumab Study Group Reviewed by: Santiago Perez-Lloret, 1 Roche Pharma Research and Early Development (pRED), Neuroscience and Rare Diseases Discovery and Translational Consejo Nacional de Investigaciones Area, Roche Innovation Center, F. Hoffmann-La Roche Ltd., Basel, Switzerland, 2 Faculty of Psychology, University of Basel, Científicas y Técnicas Basel, Switzerland, 3 Roche Pharma Research and Early Development (pRED), Pharmaceutical Sciences, Roche Innovation (CONICET), Argentina Center, F. Hoffmann-La Roche Ltd., Basel, Switzerland, 4 Paracelsus-Elena-Klinik, Kassel, Germany, 5 Department of Camila Aquino, Neurology, University Medical Center Göttingen, Göttingen, Germany, 6 Department of Neurology, Innsbruck Medical University of Calgary, Canada University, Innsbruck, Austria, 7 Idorisa Pharmaceuticals Ltd., Allschwil, Switzerland, 8 Roche Products Ltd., Welwyn Garden *Correspondence: City, United Kingdom, 9 Feetme SAS, Paris, France, 10 F. Hoffmann-La Roche Ltd., Basel, Switzerland, 11 Prothena Gennaro Pagano Biosciences Inc., South San Francisco, CA, United States, 12 Department of Neurology, Northwestern University Feinberg gennaro.pagano@roche.com School of Medicine, Chicago, IL, United States, 13 Institute for Neurodegenerative Disorders, New Haven, CT, United States, 14 Prevail Therapeutics, New York, NY, United States Specialty section: This article was submitted to Background: Currently available treatments for Parkinson’s disease (PD) do not slow Movement Disorders, a section of the journal clinical progression nor target alpha-synuclein, a key protein associated with the disease. Frontiers in Neurology Objective: The study objective was to evaluate the efficacy and safety of prasinezumab, Received: 05 May 2021 a humanized monoclonal antibody that binds aggregated alpha-synuclein, in individuals Accepted: 01 September 2021 Published: 01 October 2021 with early PD. Citation: Methods: The PASADENA study is a multicenter, randomized, double-blind, placebo- Pagano G, Boess FG, Taylor KI, Ricci B, Mollenhauer B, Poewe W, controlled treatment study. Individuals with early PD, recruited across the US and Europe, Boulay A, Anzures-Cabrera J, Vogt A, received monthly intravenous doses of prasinezumab (1,500 or 4,500 mg) or placebo Marchesi M, Post A, Nikolcheva T, for a 52-week period (Part 1), followed by a 52-week extension (Part 2) in which all Kinney GG, Zago WM, Ness DK, Svoboda H, Britschgi M, participants received active treatment. Key inclusion criteria were: aged 40–80 years; Ostrowitzki S, Simuni T, Marek K, Hoehn & Yahr (H&Y) Stage I or II; time from diagnosis ≤2 years; having bradykinesia Koller M, Sevigny J, Doody R, Fontoura P, Umbricht D, Bonni A and plus one other cardinal sign of PD (e.g., resting tremor, rigidity); DAT-SPECT imaging PASADENA Investigators and consistent with PD; and either treatment naïve or on a stable monoamine oxidase B Prasinezumab Study Group (2021) A (MAO-B) inhibitor dose. Study design assumptions for sample size and study duration Phase II Study to Evaluate the Safety and Efficacy of Prasinezumab in Early were built using a patient cohort from the Parkinson’s Progression Marker Initiative Parkinson’s Disease (PASADENA): (PPMI). In this report, baseline characteristics are compared between the treatment- Rationale, Design, and Baseline Data. Front. Neurol. 12:705407. naïve and MAO-B inhibitor-treated PASADENA cohorts and between the PASADENA doi: 10.3389/fneur.2021.705407 and PPMI populations. Frontiers in Neurology | www.frontiersin.org 1 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design Results: Of the 443 patients screened, 316 were enrolled into the PASADENA study between June 2017 and November 2018, with an average age of 59.9 years and 67.4% being male. Mean time from diagnosis at baseline was 10.11 months, with 75.3% in H&Y Stage II. Baseline motor and non-motor symptoms (assessed using Movement Disorder Society—Unified Parkinson’s Disease Rating Scale [MDS-UPDRS]) were similar in severity between the MAO-B inhibitor-treated and treatment-naïve PASADENA cohorts (MDS-UPDRS sum of Parts I + II + III [standard deviation (SD)]; 30.21 [11.96], 32.10 [13.20], respectively). The overall PASADENA population (63.6% treatment naïve and 36.4% on MAO-B inhibitor) showed a similar severity in MDS-UPDRS scores (e.g., MDS- UPDRS sum of Parts I + II + III [SD]; 31.41 [12.78], 32.63 [13.04], respectively) to the PPMI cohort (all treatment naïve). Conclusions: The PASADENA study population is suitable to investigate the potential of prasinezumab to slow disease progression in individuals with early PD. Trial Registration: NCT03100149. Keywords: Parkinson’s disease, alpha-synuclein (α-syn), prasinezumab, monoclonal antibodies, disease progression, MDS-UPDRS = Movement Disorder Society—Unified Parkinson’s Disease Rating Scale, Phase II clinical trial, disease modification treatments INTRODUCTION Preclinical findings in cellular and animal models also support the hypothesis that certain aggregated forms of alpha-synuclein There is a high medical need to develop long-lasting therapies may be taken up by neurons and may induce the formation of that can affect the underlying cause of Parkinson’s disease intracellular alpha-synuclein inclusions in PD (20, 23–25). The (PD) and, therefore, slow disease progression (1–3). Currently appearance of intraneuronal inclusions throughout the central available treatments have powerful symptomatic effects, and peripheral nervous systems may arise upon propagation of particularly on motor symptoms, but they do not address the Lewy pathology from neuron to neuron in a concerted manner by pathological processes underlying the disease and do not prevent extracellular transfer of aggregated alpha-synuclein (21, 26–28). or slow clinical decline (2, 3). With the progressive loss of Clinical evidence also supports the hypothesis that alpha- dopaminergic and non-dopaminergic neurons and synapses, synuclein is a key driver in the etiology of PD. For instance, available therapies gradually become less effective at controlling both missense mutations (29) and increased production of alpha- PD motor symptoms (4–6). Individuals with PD will invariably synuclein due to duplication or triplication of the alpha-synuclein develop motor complications and lose their autonomy, adversely gene (SNCA) (30–32) cause early-onset autosomal dominant PD, affecting their quality of life and placing a significant burden with virtually 100% penetrance (33). Although the exact patho- on caregivers, family members and healthcare systems (7–9). physiological mechanism in these genetic causes remains unclear, Despite the high prevalence and impact of non-motor symptoms aggregation of alpha-synuclein due to missense mutation or on the quality of life of individuals with PD, treatment options overexpression is supposed to drive disease onset and progression for these symptoms are limited (10). A therapy that targets the (34). Direct transfer of aggregated alpha-synuclein from neuron underlying cause of the disease has the potential to slow motor to neuron has not been directly observed in humans. However, progression as well as address non-motor symptoms (11, 12). embryonic dopaminergic neurons transplanted into the striatum Postmortem findings suggest that the loss of dopaminergic of individuals with PD harbored inclusions reminiscent of Lewy neurons is accompanied spatially and temporally by the pathology approximately a decade after initial grafting (35, 36) progressive development of intraneuronal Lewy pathology, which, together with replicated observations in animal models which is a neuropathological hallmark of PD in distinct (23, 24), suggests the possibility of intercellular propagation of brain regions (13–17). Lewy pathology is abnormally enriched Lewy pathology. In support of a caudo-rostral propagation of in alpha-synuclein, a protein with key functions in neurons Lewy pathology, molecular imaging studies demonstrate damage (17, 18). Although the etiology of PD is yet to be elicited, or dysfunction of noradrenergic and serotonergic pathways prior the spatio-temporal association between Lewy pathology and to the dopaminergic pathways in prodromal idiopathic and neurodegeneration, together with evidence from in vitro and in SNCA genetic PD (37, 38). vivo models, suggests that pathologically aggregated forms of The growing understanding of the role of alpha-synuclein alpha-synuclein may contribute to axonal and neuronal damage, in the development of Lewy pathology and the pathogenesis formation of Lewy pathology and consequent neuronal loss and of PD support the rationale that targeting alpha-synuclein may disease progression (18–22). have therapeutic potential (39). Preclinical in vivo models of Frontiers in Neurology | www.frontiersin.org 2 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design alpha-synucleinopathy, such as transgenic mice overexpressing study duration were built, in part, using a patient cohort from the wild-type human alpha-synuclein or that develop pathology Parkinson’s Progression Markers Initiative (PPMI) study group. upon intracerebral injection of aggregated recombinant alpha- The PPMI is a landmark, global, observational clinical study synuclein, are valuable when studying drug mechanisms of individuals with PD designed to comprehensively evaluate targeting alpha-synuclein. These models may help identify the cohorts of significant interest using advanced imaging, biological downstream mode of action of therapeutic compounds. Indeed, sampling and clinical and behavioral assessments to identify neuropathological and behavioral deterioration in various biomarkers of PD progression (46). We compared the baseline mouse models of alpha-synuclein pathology was shown to be characteristics of the treatment-naïve and the monoamine ameliorated by treatment with monoclonal antibodies binding to oxidase B (MAO-B) inhibitor-treated cohorts of the PASADENA alpha-synuclein (12, 40–44). population. We also compared the baseline characteristics of the Prasinezumab (previously known as RO7046015/PRX002) is total PASADENA population with the characteristics of a subset an investigational, humanized monoclonal immunoglobulin G1 of individuals with early PD enrolled in the PPMI study, which antibody directed against an epitope in the carboxyl terminus was selected using similar inclusion criteria to the PASADENA of human alpha-synuclein (11, 40, 41, 45). It binds to human study (47). aggregated alpha-synuclein with a high affinity and avidity (11, 40, 41). Preclinical pharmacologic studies to evaluate efficacy and potency of the murine form of prasinezumab METHODS (9E4) were performed in two transgenic mouse lines featuring alpha-synuclein aggregation disorders: Line D and Line 61 Study Design mice. The mice were treated with weekly intraperitoneal The Phase II study of Anti alpha-Synuclein AntiboDy in administration of 9E4 or certain other alpha-synuclein antibodies Early ParkiNson’s diseAse (PASADENA) to evaluate the safety over 5–6 months and showed reduced neuronal and synaptic and efficacy of prasinezumab (NCT03100149) is a multicenter, loss and a reduction in intraneuronal build-up of alpha- randomized, double-blind, placebo-controlled study across ∼60 synuclein pathology (measured as alpha-synuclein inclusions in sites in the United States, France, Austria, Germany, and cortical and subcortical regions), reduction of gliosis, and an Spain. The study was designed to evaluate the efficacy and improvement in both cognitive and motor behaviors (12, 40–42). safety of intravenous prasinezumab (received every 4 weeks) in Although blockade of cell-to-cell transmission of alpha- participants with early-stage PD (Hoehn and Yahr [H&Y] Stages synuclein by extracellular neutralization of pathogenic I–II, time since diagnosis ≤2 years). species has been proposed as the main mechanism of action The study consists of two parts: a 52-week, double-blind, of prasinezumab, preclinical experiments also suggest a placebo-controlled treatment period (Part 1), followed by a 52- potential engagement of the lysosomal pathway (12). Together, week extension period during which all participants received these preclinical data support the therapeutic potential of active treatment but remained blinded to original dose allocation prasinezumab in slowing the progression of PD. (Part 2) (Figure 1). A 12-week safety follow-up was mandatory In a Phase I single-ascending-dose study in healthy volunteers for all participants, regardless of whether cessation of treatment and a Phase I multiple-ascending-dose study in individuals occurred after Part 1 or Part 2. with PD, prasinezumab was safe, able to penetrate the blood– In Part 1, participants were randomized with a 1:1:1 allocation brain barrier (measured in the cerebrospinal fluid) and showed ratio to either placebo, a high dose (4,500 mg for body weight robust peripheral binding to alpha-synuclein (11, 45). Peripheral ≥65 kg; 3,500 mg for body weight <65 kg) or a low dose binding, measured as the lowering of circulating, free (unbound) (1,500 mg for all body weights) of prasinezumab. Body weight serum alpha-synuclein, occurred within 1 h of administration influences clearance (and volume of distribution) such that of prasinezumab and was maintained for longer durations with exposure increases in patients with lower body weight. Therefore, higher doses of prasinezumab (11). Results also demonstrated a the use of an ∼25% lower dose in participants with lower dose-dependent increase of prasinezumab in cerebrospinal fluid body weight (<65 kg) is implemented at high doses where there concentration, which was ∼0.3% relative to serum across all is an increased risk for infusion-related reactions. To reduce dose groups (11). Prasinezumab has a high apparent binding the risk of infusion-related reactions, participants in the high- affinity to aggregated alpha-synuclein, which makes it bind more dose group received a 2,000 mg intravenous infusion on Day 1 selectively to aggregated alpha-synuclein over monomeric alpha- followed by an up-titration to the full dose on Day 28 which they synuclein (11). Together with the observed cerebrospinal fluid then received every 4 weeks. In addition, the first three study concentrations achieved, it is predicted that >90% of aggregated treatment infusions (irrespective of treatment allocation) were alpha-synuclein will be engaged by prasinezumab in the brain prolonged to 2 h and were preceded by pre-medication with non- of individuals with PD at doses ≥1,500 mg (11). The 1,500 and sedating antihistamine and acetaminophen (11). Randomization 4,500 mg doses were selected as both were expected to saturate was stratified by sex, age group (<60 vs. ≥60 years) and use of the target in a Phase II study in individuals with early PD. MAO-B inhibitor at baseline (yes vs. no). Here we report the study rationale, design, and baseline Participants from Part 1 were eligible to continue to patient characteristics of PASADENA, a Phase II clinical trial Part 2 provided dopamine transporter with single-photon testing efficacy and safety of prasinezumab in individuals with emission computerized tomography (DaT-SPECT) and magnetic early PD. The study design and assumptions for sample size and resonance imaging (MRI) scans had been completed at screening Frontiers in Neurology | www.frontiersin.org 3 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design FIGURE 1 | PASADENA study design schematic. *Low dose = 1,500 mg, † High dose = 4,500 mg for ≥65 kg; 3,500 mg for <65 kg. DaT-SPECT, dopamine transporter imaging with single-photon emission computerized tomography; IV, intravenous; Q4W, every month. and Week 52, and participants had received at least 10 doses of Key inclusion criteria included: idiopathic PD with study treatment (placebo or prasinezumab) during Part 1. bradykinesia and one of the other cardinal signs of PD In Part 2, participants randomized to treatment with (resting tremor, rigidity) and no other known or suspected cause prasinezumab in Part 1 remained on the same dose for the of PD; aged 40–80 years; a DaT-SPECT consistent with PD; duration of Part 2. Those participants initially randomized body weight range of ≥45 to ≤110 kg and a body mass index of to placebo were re-randomized to either 1,500 or 4,500 mg 18–34 kg/m2 ; and either treatment naïve or on a stable dose of a prasinezumab using a 1:1 allocation ratio. Randomization was MAO-B inhibitor for at least 90 days. stratified by dopaminergic therapy since start of study (yes vs. Key exclusion criteria included: medical history indicating no), sex, age group at start of study (<60 vs. ≥60 years) and use a Parkinson syndrome other than idiopathic PD; known of MAO-B inhibitor at baseline (yes vs. no). carriers of certain familial PD genes (Parkin, PINK1, DJ1); If symptomatic PD treatment was initiated during Part 1, Mini Mental State Examination (MMSE) ≤25; use of any investigators were required to record the reason(s) and the type of the following: catechol-O-methyl transferase inhibitors and dose of symptomatic PD treatment prescribed. Participants (entacapone, tolcapone), amantadine or anticholinergics, or who initiated symptomatic PD treatment could then continue dopaminergic medication (levodopa and both ergot and non- in the study, as per their regular scheduled study visits. For ergot [pramipexole, ropinirole, rotigotine] dopamine agonists) participants who started dopaminergic treatment (levodopa or for more than a total of 60 days or within 60 days of baseline and dopamine agonist), the Movement Disorder Society—Unified prior participation in any prasinezumab study. Parkinson’s Disease Rating Scale (MDS-UPDRS) including Part A full list of inclusion and exclusion criteria can be found in IV (motor assessment while on dopaminergic treatment) and Supplementary Tables 1, 2. digital biomarker in-clinic assessments at subsequent visits were performed in an “Off ” state, i.e., patients had not received Objectives and Endpoints levodopa since the previous evening (>8 h prior). The MDS- Clinical assessments performed at baseline and at different UPDRS Part III (motor assessment) was repeated at least 1 h after study visits are summarized in Figures 2, 3. A full list of receiving levodopa in the clinic (while patients are in an “On” endpoints is available online in the PASADENA study protocol state), along with digital biomarker in-clinic assessments. (NCT03100149) (48). Study Population Primary Endpoint The inclusion and exclusion criteria were developed to select an The primary objective of the study was to assess the efficacy early PD population with a measurable and predictable rate of of prasinezumab 1,500 and 4,500 mg vs. placebo at Week 52 in progression over a 1-year period. enrolled participants. The primary endpoint was the change from Frontiers in Neurology | www.frontiersin.org 4 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design FIGURE 2 | Schedule of activities in PASADENA Part 1. *Parkinson’s Disease Questionnaire−39 (PDQ39) is at baseline, Week 20, and Week 48; † SCOPA-AUT (Scales for Outcomes in Parkinson’s Disease—Autonomic Dysfunction) is at baseline, Weeks 16, 28, 40 and 52. ‡ Digital includes PASADENA Digital Motor Score, Patient Global Impression of Severity, Daily diary, Patient Assessment of Constipation Symptoms (PAC-SYM), EuroQol-5D (EQ-5D) and Hospital Anxiety and Depression Scale (HADS). § Magnetic resonance imaging (MRI) includes safety, diffusion tensor imaging (DTI), resting state and arterial spin labeling (ASL). CGI-C, Clinical Global Impression of Change; CGI-I, Clinical Global Impression of Improvement; DaT-SPECT, dopamine transporter imaging with single-photon emission computerized tomography; H&Y, Hoehn & Yahr; MDS-UPDRS, Movement Disorder Society—Unified Parkinson’s Disease Rating Scale; MMSE, Mini Mental State Examination; MoCA, Montreal Cognitive Assessment; PDSS-2, Parkinson’s Disease Sleep Scale Revised Version 2; PGI-C, Patient Global Impression of Change; RBDSQ, Rapid Eye Movement Sleep Behavior Disorder Screening Questionnaire; SE-ADL, Schwab and England Activities of Daily Living. baseline at Week 52 in MDS-UPDRS sum of Parts I + II + III (SE-ADL) score; time to worsening in motor or non-motor vs. placebo. symptoms (increase of ≥3 points in MDS-UPDRS Part I or MDS-UPDRS Part II); time to start of dopaminergic Secondary Endpoints PD treatment (levodopa or dopamine agonists); and safety The effects of prasinezumab 1,500 and 4,500 mg vs. placebo at (Supplementary Table 3), tolerability, immunogenicity, and Week 52 on MDS-UPDRS Part IA, Part IB, Part I total, Part pharmacokinetics (PK) of prasinezumab. Safety and tolerability II total, Part III total, and Part III subscores (bradykinesia, were assessed for up to 104 weeks, with or without dopaminergic rigidity, resting tremor, and axial symptoms) were included as treatment. PK of prasinezumab was assessed using population secondary endpoints. Part I assessed non-motor experiences PK modeling. of daily living, with Part IA focused on complex behaviors (cognitive impairment, hallucinations, and psychosis, etc.) and Exploratory Endpoints Part IB focused on non-motor experiences (sleep and urinary Exploratory endpoints of this study included: MDS-UPDRS problems, constipation, pain, etc.). Part II assessed motor Part III subscores determined by independent central raters experiences of daily living (eating, dressing, handwriting, getting (using video recordings to address consistency and accuracy in out of bed, etc.) and Part III assessed motor signs of PD the trained site raters), imaging analysis of striatum, caudate, (speech, finger tapping, bradykinesia, gait, and freezing, etc.) and putamen (average, ipsilateral, and contralateral) for DaT- (49). Part IA and Part III were administered by the study SPECT binding ratio values, and the change from baseline investigator, and Part IB and Part II were completed by on a sensor-based measure derived from Roche PD Mobile the participant. Application v2 digital biomarkers (50) (smartphone and wrist- Other secondary endpoints included; Montreal Cognitive worn wearable) assessments (see Figure 4 for an overview of Assessment (MoCA) Total score; Clinical Global Impression the remote monitoring tests). The analyses of primary and of Improvement (CGI-I); Patient Global Impression of Change secondary endpoints were also repeated, with the results for the (PGI-C); DaT-SPECT in the ipsilateral (to the clinically dominant two prasinezumab 1,500 and 4,500 mg pooled doses vs. placebo, side) putamen; Schwab and England Activities of Daily Living as a pre-specified exploratory analysis. Frontiers in Neurology | www.frontiersin.org 5 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design FIGURE 3 | Schedule of activities in PASADENA Part 2. *Digital includes PASADENA Digital Motor Score, Patient Global Impression of Severity, Daily diary, Patient Assessment of Constipation Symptoms (PAC-SYM), EuroQol-5D (EQ-5D), and Hospital Anxiety and Depression Scale (HADS). † Magnetic resonance imaging (MRI) includes safety, diffusion tensor imaging (DTI), resting state and arterial spin labeling (ASL). CGI-I, Clinical Global Impression of Improvement; DaT-SPECT, dopamine transporter imaging with single-photon emission computerized tomography; MDS-UPDRS, Movement Disorder Society—Unified Parkinson’s Disease Rating Scale; MoCA, Montreal Cognitive Assessment; PDQ39, Parkinson’s Disease Questionnaire−39; PDSS-2, Parkinson’s Disease Sleep Scale Revised Version 2; PGI-C, Patient Global Impression of Change; SCOPA-AUT, Scales for Outcomes in Parkinson’s Disease—Autonomic Dysfunction; SE-ADL, Schwab and England Activities of Daily Living. FIGURE 4 | Table of digital measures included in the Roche Parkinson’s Disease Mobile Application v2. SDMT, Symbol Digit Modalities Test. A full list of exploratory endpoints is included in global motor function was developed. Data from a PASADENA- Supplementary Table 4. like PPMI cohort were used to inform sensor feature selection for the PASADENA Digital Motor Score, as follows: MDS-UPDRS Parts II and III item-level scores which significantly declined PASADENA Digital Motor Score in Year 1 of PPMI were identified [n = 21; whereby one item Summary scores of sensor data from digital health technology (posture) was not tested with the digital biomarker; final set tools should be developed independent of existing clinical data to of MDS-UPDRS items = 20]. A blinded subset of PASADENA ensure that the digital score does not inherit the shortcomings of data (n = 157) was used to map sensor feature data (aggregated the clinical measure (e.g., restriction of range, reduced resolution over 2-week periods) onto each MDS-UPDRS item with the of scale). However, this requires independent longitudinal sensor blinded PASADENA population. The set of sensor features was datasets to build and validate such a digital score (50). Since such used to predict the sum of the 20 identified MDS-UPDRS items. data are not yet available, a provisional single summary sensor- PASADENA Digital Motor Scores were generated for every 2 based measure, the “PASADENA Digital Motor Score”, reflecting weeks of the study and submitted to linear random coefficient Frontiers in Neurology | www.frontiersin.org 6 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design (i.e., slope) models testing for differences in slopes between DaT-SPECT signal loss at Week 52 and the pairwise comparison censored patients in each treatment group over the 26 2-week of each active dose arm with placebo (47, 52). periods of the PASADENA study. A second set of exploratory digital biomarker analyses Covariate Adjustment comprised 17 individual pre-specified sensor features, which Analyses of efficacy endpoints (primary, secondary, and were selected based on a previous communication that reported exploratory) included the following covariates in the model: cross-sectional correlations between sensor features and MDS- background therapy at baseline (MAO-B inhibitor treatment [yes UPDRS scores (51) and available literature. These individual vs. no]), age group (<60 vs. ≥60 years), sex (male vs. female), sensor features were also analyzed with linear random coefficient DaT-SPECT binding ratio in the contralateral (to the clinically models testing for differences in slopes across treatment groups most affected side) putamen at baseline, and treatment group if the model’s residuals were normally distributed; if the residuals (4,500 mg prasinezumab, 1,500 mg prasinezumab or placebo). were not normally distributed, mixed-effect models for repeated For each continuous endpoint the baseline of the endpoint measures (MMRM) were applied. variable was included in the model. Statistical Analyses of Primary Efficacy Endpoint Standard Protocol Approvals, Registrations, and Change in MDS-UPDRS sum of Parts I + II + III from baseline Patient Consents vs. placebo was analyzed using an MMRM, with covariates This study was conducted in full conformance with the described in the Covariate adjustment section as fixed effects. The International Conference on Harmonization E6 guideline for model also included baseline MDS-UPDRS sum of Parts I + II + Good Clinical Practice and the principles of the Declaration of III, week of treatment (as a categorical factor), a treatment-by- Helsinki, or the laws and regulations of the country in which week interaction term and an interaction term between baseline the research was conducted, whichever afforded the greater MDS-UPDRS by week. An unstructured variance-covariance protection to the individual. The study protocol, Informed matrix was used to model the random error. The model tested the Consent Forms, and any information given to the participant, null hypothesis of no treatment difference at a two-sided α-level were approved by the Institutional Review Board or Ethics of 20% for the following comparisons: Committee (NCT03100149). • 4,500 or 3,500 mg (high dose) prasinezumab vs. placebo. • 1,500 mg (low dose) prasinezumab vs. placebo. Sample Size Justification A sample size of ∼100 randomized participants per group (300 All patients randomized in the study were included in participants in total for the three groups) was estimated, which the analyses. All the assessments flagged on and after the allowed for a power of ∼80% at two-sided α-level of 20% to “first symptomatic PD treatment date” (either initiation of detect a three-point difference in MDS-UPDRS sum of Parts symptomatic PD treatment or a change to MAO-B inhibitor I + II + III between groups from baseline at Week 52. The dose) were not included in the analysis per the hypothetical 3-point difference was selected based on clinical judgement of strategy described in the ICH E9(R1) Addendum on Estimands expert consultants in movement disorders and modeling on and Sensitivity Analyses in Clinical Trials by the European PPMI data, which were used to model the disease progression of Medicines Agency. The MMRM model does not perform an the placebo arm in the PASADENA study. The power calculation imputation of missing data; however, high correlations between was based on simulations of the MMRM analysis planned for the successive observations of a subject allow data from subjects primary efficacy variable. Assessments performed while on any who started dopaminergic PD treatment to contribute to the dopaminergic PD treatment started after randomization were not estimation at their final time-point. The primary endpoint of included in the analysis of the primary endpoint. MDS-UPDRS sum of Parts I + II + III was re-analyzed with data The assumptions on progression, variability, dropout rate, and from the two treatment arms pooled and compared with placebo likelihood to start dopaminergic PD treatment within the first as an exploratory endpoint. 52 weeks of treatment, with or without a MAO-B inhibitor as background therapy, were derived from analyses based on the Statistical Analyses of Secondary and Exploratory PPMI database and various sources of information from the Efficacy Endpoints literature (47, 52). The percentage of patients defined as non- For the endpoints utilizing MDS-UPDRS Part IA, Part IB, Part evaluable at Week 52 was predicted to be 25% in the placebo I total, Part II total, Part III total, Part III subscores, CGI- group and 20% in the treatment groups. It was expected that I, and PGI-C, the information collected after symptomatic PD patients treated with prasinezumab would be less likely to start treatment was handled as for the primary analysis. The analysis dopaminergic PD treatment during the study period, compared of all other endpoints included all the data available regardless of with those treated with placebo. This estimate included non- start of symptomatic PD treatment. evaluable data from patients who prematurely dropped out An analysis of covariance (ANCOVA) was used to analyze and/or started dopaminergic PD treatment after randomization. the change from baseline at Week 52 in MoCA Total The sample size of 100 patients per arm also provided 76% score and SE-ADL score with covariates described in the power (α = 20%, two sided) to reject the null hypothesis, Covariate adjustment section. The change (between baseline assuming a 37.5% reduction for the key secondary endpoint, the and Week 52) in DaT-SPECT striatal binding ratio in the Frontiers in Neurology | www.frontiersin.org 7 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design ipsilateral putamen to the clinically most affected side, was Subgroups included in analyses were: analyzed using ANCOVA. Arterial spin labeling (ASL) MRI • MAO-B inhibitors at baseline (yes vs. no) was performed at sites with the technical capability. Effects • H&Y Stage at baseline (I vs. II) are tested for in pre-specified regions of interest; striatum, • Rapid Eye Movement Sleep Behavior Disorder Screening caudate, and putamen (ipsilateral, contralateral, and average Questionnaire (RBDSQ) at baseline (RBDSQ ≥5 vs. <5) were assessed for each region). Between-group changes in • Data-driven subphenotypes (Diffuse malignant vs. mild motor ASL MRI over 1 year in regions of interest were also tested predominant vs. intermediate) at baseline using ANCOVAs. • Alpha-synuclein skin (positive vs. negative) (staining by The CGI-I was intended as a measure of change in health immunohistochemistry on skin biopsy sections at baseline) status from baseline CGI—Severity of illness (CGI-S). For the • DaT-SPECT striatal binding ratio in the ipsilateral putamen to CGI-I, patients were divided into one of two groups: the clinically most affected side (very abnormal vs. abnormal). • “Responders”: Score of 1–4 (i.e., rated as “no change,” For the derivations of the data-driven subphenotypes, scales were “minimally improved,” “much improved,” or “very classified into: motor scales (MDS-UPDRS Part II and MDS- much improved”). UPDRS Part III) and non-motor scales (Scales for Outcomes • “Progressors”: Score of 5–7 (i.e., rated as “minimally worse,” in Parkinson’s Disease—Autonomic Dysfunction [SCOPA-AUT], “much worse,” or “very much worse”). RBDSQ, and MoCA). After each one of the scales had been The proportion of patients rated by CGI-I grouping at Week 24 divided into percentiles, the data-driven subphenotypes were and Week 52 was analyzed using a logistic regression model. defined as follows: The estimated odds ratio for “responders” and “progressors” • Diffuse malignant: Score on motor scales being greater than at Week 24 and Week 52 for treated patients compared with the 75th percentile; and at least one score on a non-motor scale placebo were calculated with 80% confidence interval. Analysis greater than the 75th percentile; or all three non-motor scores of the PGI-C followed the same methodology outlined for the greater than the 75th percentile CGI-I above. • Mild motor predominant: Motor and all non-motor scores less Time to worsening of motor or non-motor symptoms (of than the 75th percentile ≥3-point change from baseline in MDS-UPDRS Part I or Part • Intermediate: All those individuals not meeting criteria for II) and time to start of dopaminergic (levodopa or dopamine other subtypes. agonist) treatment were plotted using a Kaplan-Meier survival plot and analyzed using a Cox proportional hazards model to obtain a treatment difference between each of the prasinezumab Inclusion Criteria for the PPMI Cohort dose levels against placebo. A 3-point minimum change in MDS- The design of this study and the assumptions for progression, UPDRS Part II was chosen as this has been previously identified variability, dropout rate, and likelihood to start symptomatic as the smallest change of score that is clinically meaningful to treatment within the first 52 weeks of treatment, with or without patients (53). a MAO-B inhibitor, were derived from analyses of data collected in the PPMI observational clinical study. Details regarding the PPMI study have been previously published and are available Subgroup Analyses at ppmi-info.org (52, 54). Patients from the PPMI population Subgroups with ≥20% of patients from the modified intent-to- were selected for comparison with the PASADENA cohort treat population at baseline were analyzed. The model used for using the following criteria, which align with the PASADENA the primary endpoint was run in each subgroup, excluding the study criteria: to have at least two of the following: resting subgroup being analyzed if that was a covariate (e.g., MAO- tremor, bradykinesia, rigidity (must have either resting tremor B inhibitors at baseline [yes vs. no]). The subgroup analyses or bradykinesia), and confirmation from imaging core that DaT- were performed in the primary endpoint and the following SPECT screening was consistent with dopamine transporter secondary/exploratory endpoints: deficit. Data were downloaded in May 2020 and, from the • MDS-UPDRS Part I total 423 individuals in the PPMI population, a cohort of • MDS-UPDRS Part II 336 participants were selected based on the above criteria. No • MDS-UPDRS Part III Total score and subscores individuals in the PPMI cohort received MAO-B inhibitors • MDS-UPDRS sums of Part II and III at baseline. • DaT-SPECT striatal binding ratio in the ipsilateral putamen to the clinically most affected side Statistical Comparisons Between PASADENA and • PASADENA Digital Motor Score PPMI Cohorts • MoCA score The standardized mean difference (SMD) in prognostic scores • Time to worsening in motor signs (increase of ≥3 points in was used to assess differences between baseline data for the MDS-UPDRS Part III) PASADENA and PPMI cohorts, and for the PASADENA MAO- • Time to worsening in motor or non-motor symptoms B inhibitor-treated vs. treatment-naïve patient groups. The SMD (increase of ≥3 points in MDS-UPDRS Part I or MDS-UPDRS was calculated as the absolute value in the difference in means Part II) of a covariate across the treatment groups, divided by the Frontiers in Neurology | www.frontiersin.org 8 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design pooled standard deviation (SD). SMDs larger than 0.25 indicate compared with the PPMI cohort. The PASADENA population that the groups were too different from one another for a had longer average time from diagnosis (10.11 [6.50] months reliable comparison of change from baseline in that variable (55). vs. 6.44 [6.31] months, respectively; SMD: −0.573) compared Overall, 20 covariates were selected for the analysis, including with the PPMI cohort. DaT-SPECT striatal binding ratios for demographic, imaging, and clinical assessment (MDS-UPDRS the PASADENA population for the contralateral putamen (0.80 Parts I + II + III) data. [0.25] vs. 0.68 [0.27], respectively; SMD: −0.445) and ipsilateral putamen (1.06 [0.32] vs. 0.96 [0.39], respectively; SMD: −0.285) to the clinically most affected side were both higher compared RESULTS OF BASELINE DATA ANALYSIS with the PPMI cohort. On average, PASADENA patients scored Baseline PASADENA Demographics higher in MoCA (27.87 [2.03] vs. 27.24 [2.29], respectively; A total of 443 patients were screened between June 2017 SMD: −0.291) and lower in SCOPA-AUT (8.05 [5.71] vs. 9.75 and November 2018 at 60 sites. Overall, 127 patients failed [6.23], respectively; SMD: 0.284) scores compared with the PPMI screening due to not meeting certain inclusion and exclusion cohort. All other baseline characteristics were balanced between criteria, such as brain DaT-SPECT screening consistent with the PASADENA population and the PPMI cohort (Table 1 and PD, concomitant disease, or condition within 6 months of Supplementary Figure 2). screening and MMSE ≤25. Overall, 316 patients were enrolled at 57 centers across the following five countries: United States (160 patients [50.6%]), Spain (50 patients [15.8%]), France (65 DISCUSSION patients [20.6%]), Germany (35 patients [11.1%]), and Austria (6 patients [1.9%]). Of those patients enrolled into the PASADENA PASADENA is the first Phase II study to test the efficacy study, 115 (36%) had received MAO-B inhibitor treatment at of a monoclonal antibody that selectively binds aggregated enrollment and 201 (64%) were treatment naïve. The mean alpha-synuclein to slow disease progression in early PD. The (SD) age of PASADENA patients was 59.9 (9.10) years and the PASADENA study enrolled individuals diagnosed with early- population included 213 (67.4%) men and 103 (32.6%) women, stage PD, requiring that they be, inter alia, either treatment naïve with a mean time from diagnosis of 10.11 (6.50) months and 238 or on stable treatment with MAO-B inhibitors and have a DaT- (75.3%) individuals being H&Y Stage II. The mean (SD) MDS- SPECT-confirmed dopaminergic deficit. In order for this Phase UPDRS sum of Parts I + II + III at baseline was 31.41 (12.78) and II proof-of-concept study to measure the disease-modifying the mean total scores for the individual parts were: Part I, 4.61 potential of prasinezumab within the study period of 1 year, (3.83); Part II, 5.33 (4.04); and Part III, 21.47 (9.00). The mean it was essential to define a population with a measurable rate baseline DaT-SPECT striatal binding ratios for the PASADENA of progression. The progression rate may depend on baseline population were 0.80 (0.25) for the contralateral putamen and disease severity and other factors (56). Several previous studies 1.06 (0.32) for the ipsilateral putamen. have reported that the progression rate in individuals with early PD is generally faster shortly after diagnosis of PD and MAO-B Inhibitor-Treated vs. before the start of levodopa or dopamine agonist therapy Treatment-Naïve Patients in the usage (47, 57). Individuals at this early stage of disease still have vulnerable dopaminergic neurons and protecting them PASADENA Population against the development of further alpha-synuclein pathology Within the PASADENA study population, patients treated with may potentially slow motor disease progression. A 1-year MAO-B inhibitors at baseline were on average younger (58.2 treatment duration, if well-powered, is expected to be sufficient to [9.00] years vs. 60.8 [9.00] years, respectively; SMD:−0.290) demonstrate relevant between-group differences, resulting from and had a longer time from diagnosis (11.96 [6.10] months vs. an effect of treatment on disease progression in individuals with 9.06 [6.50] months, respectively; SMD: 0.461) vs. the treatment- early PD. naïve group. MAO-B inhibitor-treated patients also had a Current treatments for PD, such as levodopa, improve motor higher MoCA score vs. the treatment-naïve group (28.27 [1.96] symptoms and aim to increase dopamine levels, compensating vs. 27.65 [2.04], respectively; SMD: 0.309). All other baseline for the dopaminergic cell and synaptic loss (58). It is possible characteristics were balanced between patients who received a that the potential effects of treatment with a disease-modifying MAO-B inhibitor (n = 115) and those who were treatment naïve therapy on motor symptoms in individuals with early PD (n = 201; Table 1 and Supplementary Figure 1). might be masked by these powerful symptomatic therapies (47, 52). People with early PD treated with MAO-B inhibitors PASADENA Population vs. PPMI Cohort, have a reduced likelihood of starting levodopa or dopamine Selected Using PASADENA Eligibility agonists therapy compared with placebo, while maintaining a Criteria relatively high progression rate (56, 57). Individuals with early When comparing the PASADENA population baseline PD who were either treatment naïve or treated with a MAO-B characteristics with those of the PPMI cohort, MDS-UPDRS inhibitor at baseline were, therefore, included in Part 1 of the mean scores were lower for the PASADENA population for PASADENA study. The recruited PASADENA study population Part I (4.61 [3.83] vs. 5.60 [3.93], respectively; SMD: 0.255) and was also similar to other early PD therapeutic trial populations Part IB (3.45 [2.78] vs. 4.33 [3.10], respectively; SMD: 0.297) (56, 59, 60). Frontiers in Neurology | www.frontiersin.org 9 October 2021 | Volume 12 | Article 705407 Frontiers in Neurology | www.frontiersin.org Pagano et al. TABLE 1 | PASADENA; MAO-B inhibitor-treated vs. treatment-naïve patients, and PASADENA (all patients) vs. PPMI. PASADENA PASADENA SMD (CI) PASADENA PPMI SMD (CI) MAO-B Treatment-naïve All patients (n = PD patients (n = inhibitor-treated patients 316) 336) patients (n = 201) (n = 115) Age, years mean (SD) 58.2 (9.00) 60.8 (9.00) −0.290* (−0.520, −0.059) 59.90 (9.10) 61.30 (9.69) 0.166 (0.012, 0.320) Gender Male, n (%)† 74 (64.3) 139 (69.2) −0.048 (−0.156, 0.060) 213 (67.4) 220 (65.5) −0.019 (−0.092, 0.053) Years of education, mean (SD) 16.39 (5.20) 15.22 (4.80) 0.234 (0.006, 0.467) 15.65 (4.99) 15.53 (3.03) −0.029 (−0.183, 0.125) Time from diagnosis, months mean (SD) 11.96 (6.10) 9.06 (6.50) 0.461* (0.225, 0.689) 10.11 (6.50) 6.44 (6.30) −0.573* (−0.730, −0.417) H&Y Stage II, n (%)† 83 (72.2) 155 (77.1) −0.049 (−0.150, 0.051) 238 (75.3) 197 (59.0) −0.167 (−0.238, −0.096) RBDSQ (SD) 3.51 (2.65) 3.43 (2.75) 0.031 (−0.199, 0.260) 3.46 (2.71) 4.14 (2.69) 0.250 (0.095, 0.405) DaT-SPECT contralateral putamen (SD) 0.81 (0.24) 0.78 (0.25) −0.102 (−0.331, 0.128) 0.80 (0.25) 0.68 (0.27) −0.445* (−0.601, −0.289) DaT-SPECT ipsilateral putamen (SD) 1.02 (0.30) 1.09 (0.33) −0.194 (−0.421, 0.038) 1.06 (0.32) 0.96 (0.39) −0.285* (−0.439, −0.129) MoCA (SD) 28.27 (1.96) 27.65 (2.04) 0.309* (0.076, 0.539) 27.87 (2.03) 27.24 (2.29) −0.291* (−0.446, −0.136) SCOPA-AUT (SD) 7.71 (4.82) 8.25 (6.16) −0.096 (−0.323, 0.136) 8.05 (5.71) 9.75 (6.23) 0.284* (0.128, 0.439) MDS-UPDRS sum of Parts I + II + III, mean (SD) 30.21 (11.96) 32.10 (13.20) −0.150 (−0.378, 0.081) 31.41 (12.78) 32.63 (13.04) 0.094 (−0.059, 0.248) MDS-UPDRS Part I, mean (SD) 4.49 (3.40) 4.68 (4.06) −0.051 (−0.279, 0.180) 4.61 (3.83) 5.60 (3.93) 0.255* (0.100, 0.409) MDS-UPDRS Part IA, mean (SD) 0.96 (1.54) 1.27 (1.67) −0.195 (−0.422, 0.037) 1.16 (1.62) 1.27 (1.57) 0.071 (−0.083, 0.224) 10 MDS-UPDRS Part IB, mean (SD) 3.53 (2.50) 3.14 (2.93) 0.045 (−0.185, 0.273) 3.45 (2.78) 4.33 (3.10) 0.297* (0.142, 0.451) MDS-UPDRS Part II, mean (SD) 5.19 (3.90) 5.41 (4.13) −0.055 (−0.284, 0.174) 5.33 (4.04) 6.12 (4.20) 0.190 (0.036, 0.344) MDS-UPDRS Part III, mean (SD) 20.53 (8.81) 22.01 (9.09) −0.165 (−0.394, 0.065) 21.47 (9.00) 20.92 (8.88) −0.062 (−0.216, 0.092) MDS-UPDRS Part III Axial Symptoms, mean (SD)† 0.77 (0.50) 0.78 (0.57) −0.004 (−0.233, 0.225) 0.78 (0.54) 0.68 (0.71) −0.153 (−0.307, 0.001) MDS-UPDRS Part III Bradykinesia, mean (SD)‡ 9.91 (5.60) 10.48 (5.49) −0.102 (−0.331, 0.127) 10.27 (5.53) 10.60 (5.60) 0.059 (−0.095, 0.212) MDS-UPDRS Part III Rigidity, mean (SD)‡ 3.90 (2.58) 4.17 (2.83) −0.103 (−0.331, 0.128) 4.07 (2.74) 3.86 (2.61) −0.078 (−0.232, 0.075) MDS-UPDRS Part III Resting Tremors, mean (SD)‡ 2.71 (2.77) 3.12 (2.74) −0.147 (−0.377, 0.082) 2.97 (2.75) 2.58 (2.42) −0.151 (−0.305, 0.003) Demographics and baseline disease characteristics compared using the SMD. *Indicates not balanced covariates (>0.25 SMD). † For binary variables the table shows the difference in proportions. ‡ Part III subscores are defined as: Bradykinesia (sum of item 3.4, finger tapping; item 3.5, hand movements; item 3.6, pronation-supination movements of hands; item 3.7, toe tapping; item 3.8, leg agility; item 3.9, arising from chair; item 3.13, posture; and item 3.14, body bradykinesia); Rigidity (sum of item 3.3. [Neck, Upper Limbs and Lower Limbs]); Resting tremors (sum of item 3.17, rest tremor amplitude [Lip/Jaw, Upper Limbs and Lower Limbs] and Item 3.18, constancy of tremor); and axial symptoms (sum of item 3.10, gait; item 3.11, freezing of gait; and item 3.12, postural stability). CI, confidence interval; DaT-SPECT, dopamine transporter with single-photon emission computerized tomography; H&Y, Hoehn and Yahr; October 2021 | Volume 12 | Article 705407 MAO-B, monoamine oxidase B; MDS-UPDRS, Movement Disorders Society—Unified Parkinson’s Disease Rating Scale; MoCA, Montreal Cognitive Assessment; PD, Parkinson’s disease; PPMI, Parkinson’s Progression Markers Initiative; RBDSQ, Rapid Eye Movement Sleep Behavior Disorder Screening Questionnaire; SCOPA-AUT, Scales for Outcomes in Parkinson’s Disease—Autonomic Dysfunction; SD, standard deviation; SMD, standardized mean difference. PASADENA Phase II Study Design Pagano et al. PASADENA Phase II Study Design In this study, a comparison of the baseline characteristics of supported with a panel of exploratory biomarkers assessing the PASADENA study participants showed that those who received potential effects on PD pathology and progression of neuronal treatment with MAO-B inhibitors were younger with a longer damage. The MDS-UPDRS is comprised of four parts: Part time from diagnosis compared with the treatment-naïve group; I, Mentation, Behavior, and Mood; Part II, Activities of Daily however, the two groups had similar overall symptom severity at Living; Part III, Motor Examination; and Part IV, Complications baseline. This observed similarity may be due to improvement in of Therapy (49, 64). Each parkinsonian sign or symptom is MDS-UPDRS scores as a result of MAO-B inhibitor treatment, rated on a 5-point scale (ranging from 0 to 4), with higher which provides symptomatic relief to individuals with early PD scores indicating more severe impairment (64). The MDS- by prolonging the action of dopamine in the brain (61, 62). UPDRS demonstrates good reliability, validity and sensitivity to The younger age may be due to the fact that individuals with change over a range of measures of time from diagnosis and older onset of PD usually receive levodopa rather than MAO-B severity (64). Previous studies in individuals with early PD have inhibitors (63). demonstrated a linear increase in MDS-UPDRS of ∼6–12 points The design and assumptions of this study were informed per year following diagnosis and prior to initiating symptomatic from analyses of data collected in the PPMI observational treatment (47, 52, 56, 57). A positive effect on MDS-UPDRS clinical study. Therefore, it was important to compare the scores may, therefore, indicate a potential effect of prasinezumab baseline characteristics of the PASADENA study population on global PD progression. It is important to note that the increase with the PPMI cohort to ensure the assumptions made were in MDS-UPDRS scores in the treatment-naïve population with also valid for the PASADENA population. The PASADENA early PD is derived from the Part III motor examination scores; population and PPMI cohort showed different distributions for this population exhibits decline to a far lesser extent in activities some demographic measures; for example, more PASADENA of daily living (MDS-UPDRS Part I) and motor problems in daily participants were in H&Y Stage II (75 vs. 59%) and had on life (MDS-UPDRS Part II) (64). average a 3.7-month-longer time from diagnosis than the PPMI A deficit of striatal dopamine transporters, measured by DaT- cohort. However, the DaT-SPECT striatal binding ratios for SPECT imaging, is used to confirm that neurodegeneration both the ipsilateral and contralateral putamen suggested that of striatal dopaminergic terminals is indicative of PD (65). the PASADENA population had slightly less advanced disease in Individuals at the earliest stage of PD show the fastest terms of severity and potentially of progression. inverse exponential decline of DaT-SPECT signal (i.e., loss of Dose selection for the PASADENA trial was based on data dopaminergic terminals) in the ipsilateral putamen to the most from previous studies. In a Phase I multiple-ascending-dose clinically affected side (66, 67). Therefore, DaT-SPECT in the study (NCT02157714), individuals with mild-to-moderate PD ipsilateral putamen was selected as a secondary outcome measure who received prasinezumab up to 60 mg/kg intravenously every in the PASADENA study to determine the disease-modifying 4 weeks reported no serious or severe adverse events (11). potential of prasinezumab on the neurodegeneration of the Rapid-dose and time-dependent mean reductions from baseline nigrostriatal terminals. vs. placebo in free serum alpha-synuclein levels of up to 97% Smartphones and smartwatches are built with high-quality were reported in trial participants after a single infusion at the sensors that, together with novel software technologies, enable highest dose (P = 0.002), with similar reductions after two the remote, non-invasive, frequent, and sensitive measurement additional infusions. Mean cerebrospinal fluid concentration of and analysis of motor symptoms in PD (68–70). Digital prasinezumab also increased with dose, to ∼0.3% relative to monitoring of motor symptoms using smartphones has been the concentration in serum across all dose cohorts. Currently, previously used in the Phase I study of prasinezumab in assays to quantify engagement of prasinezumab with aggregated individuals with PD (NCT02157714), in which preliminary forms of alpha-synuclein in vivo are not available. Thus, the reliability and clinical validity were established (71). Study dose selection for the Phase II study was primarily based on participants completed a daily battery of tests and carried the human serum and cerebrospinal fluid PK data extrapolation, phone with them throughout the day for passive monitoring. and the relationship of these data with histopathological and The study revealed high adherence and a strong correlation functional endpoints. The doses used in the PASADENA trial between smartphone sensor data and clinical measures of motor were selected to fall in the therapeutic exposure range predicted signs and, notably, the detection of clinical manifestations that from preclinical efficacy models; the high prasinezumab dose were not apparent at site visits (71). A second version of (4,500 mg for body weight ≥65 kg; 3,500 mg for body weight this digital biomarker approach was therefore implemented <65 kg) was selected to match exposure at the 60 mg/kg in the PASADENA study to maximize the probability of dose in the multiple-ascending-dose study, and the 1,500 mg detecting a potential therapeutic effect of prasinezumab and prasinezumab dose to yield exposure levels above those effective potentially provide new insights into the functioning and on alpha-synuclein pathology in the mouse model, with sufficient behavior of individuals with PD. The test-retest reliability separation between the two to enable exposure response analyses. and preliminary clinical validity of the sensor-based outcome Both doses selected were expected to bind >90% of pathological measures were demonstrated with high intraclass correlation aggregated alpha-synuclein in the central nervous system, thus coefficients and significant correlations with MDS-UPDRS Part both could show signal of efficacy on disease progression (11). III subscale items, respectively (72). As mentioned, summary The effect of treatment with prasinezumab on clinical digital scores should be developed independent of clinical progression rate was determined using the MDS-UPDRS and was measures. However, the independent longitudinal sensor datasets Frontiers in Neurology | www.frontiersin.org 11 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design required to build and validate such a digital score are not we would recruit untreated individuals in the prodromal stage yet available (50). Therefore, the “PASADENA Digital Motor of PD, however, there are currently no available biomarkers that Score” was developed as an exploratory outcome measure for give a definitive diagnosis of prodromal PD (79). Therefore, we this study to reflect global motor function. We note that this included participants with a recent clinical diagnosis of PD (<2 exploratory outcome measure would require validation in an years), as this is currently the earliest stage at which PD can be independent cohort before elevating this measure higher in the definitively identified. endpoint hierarchy. Finally, proof-of-concept studies can present statistical A definitive diagnosis of PD can only be made post-mortem as challenges, such as smaller sample sizes, which can reduce biomarker tools that detect alpha-synuclein in the brain in vivo statistical power. In this study, the Sponsor decided to use an are not currently available (73). However, pathological forms of 80% confidence interval (α = 0.20) rather than a more common alpha-synuclein have been detected in peripheral neurons present 95% (α = 0.05) to ensure completion of the trial in an acceptable in skin biopsy samples from individuals with PD; the degree of timeframe. Although reducing the sample size may lead to less peripheral nerve pathology detected in samples was found to precision, point estimates reported together with 80% confidence correlate with disease severity (74, 75). Longitudinal skin biopsy intervals still provide useful information about the plausible sampling has been implemented in the PASADENA study for magnitude of the effect between prasinezumab and placebo, as the direct and in vivo assessment of alpha-synuclein pathology, lower confidence levels decrease the width of the confidence and its progression in response to treatment with prasinezumab. interval. The trade-off between sample size and confidence level Clinical data and samples, including skin biopsies, will be confirmed that the endpoints with a confidence interval that does collected from DaT-SPECT negative screen failure participants not contain the value of null effect should be investigated in to determine whether the detection of alpha-synuclein skin future clinical trials (80). pathology may be used as a sensitive, specific, and less invasive tool to diagnose PD. While this study was designed to assess the safety and efficacy CONCLUSIONS of prasinezumab in early PD, there were some limitations to The PASADENA Phase II study was designed to assess the safety the study. In order to assess the disease-modifying effect of a and tolerability and clinical effect of prasinezumab on disease drug in PD, the disease must progress by a measurable amount progression in patients with early PD. This study will focus during the study period. Over a 1-year period, individuals with on the effect of treatment in early PD, as disease progression early PD progress <2 points on both MDS-UPDRS Parts I and is measurable and predictable in this cohort and will not be II (47). This is below the threshold for clinical meaningfulness masked by treatment with dopaminergic therapy. The primary (>2 points) when a patient’s quality of life is negatively affected outcome measure will be supported by clinical measures and (76). However, the MDS-UPDRS Part III progresses more quickly imaging to investigate the potential physiological impact of in early PD and can often pass the threshold for clinical treatment. In addition, novel digital biomarkers will be used to meaningfulness; 4.63 points (47, 76). This may in part be due to assess potentially subtle effects of treatment on motor function in the original UPDRS being developed when PD was thought to individuals with PD. be a predominantly motor disease, thus, non-motor symptoms such as constipation, fatigue and sleep problems are inadequately represented on the scale (77). DATA AVAILABILITY STATEMENT This study included patient-reported outcome measures of quality of life (e.g., PGI-C, SE-ADL, CGI-I); however, the change The datasets presented in this study can be found in online in these outcome measures is usually minimal in early PD. repositories. Qualified researchers may request access to Therefore, there is a need for a sensitive outcome measure that individual patient-level data through the clinical study data captures both early and non-motor PD symptoms, addresses request platform (https://vivli.org/). Further details on Roche’s symptom diversity and is correlated with clinically meaningful criteria for eligible studies are available here (https://vivli.org/ impacts on quality of life (53, 77). Another limitation of members/ourmembers/). For further details on Roche’s Global studies assessing disease progression in PD is the confounding Policy on the Sharing of Clinical Information and how to request effect of dopaminergic PD treatment. We accounted for this access to related clinical study documents, see here (https://www. by analyzing the change in MDS-UPDRS using a hypothetical roche.com/research_and_development/who_we_are_how_we_ estimand strategy, in which the participants are included in work/clinical_trials/our_commitment_to_data_sharing.htm). the model until the visit prior to starting dopaminergic PD Data used in the preparation of this article were obtained from treatment. In addition, we performed a sensitivity analysis using the Parkinson’s Progression Markers Initiative (PPMI) database the treatment policy estimand strategy, in which all participants (ppmi-info.org/data). For up-to-date information on the study, are included regardless of starting dopaminergic PD treatment. visit ppmi-info.org. In this analysis, we have tried to account for the confounding effect of dopaminergic PD treatment by including an 8-h off ETHICS STATEMENT medication time for the “Off ” evaluation in this study, the long duration response to levodopa may begin to have a PK impact The studies involving human participants were reviewed on patients within their first years of levodopa use (78). Ideally, and approved by Ethikkommission der Medizinischen Frontiers in Neurology | www.frontiersin.org 12 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design Universität Innsbruck, Innsbruck, Austria; Comité de Carbone, Claire Carroll, Laura Casado Fernandez, Catherine Protection des Personnes (CPP) Ouest IV, Nantes, France; Cassé-Perrot, Anna Castrioto, Helene Catala, Justine Chan, Ethikkommission der Universität Leipzig and Geschäftsstelle Samia Cheriet, Anthony Ciabarra, Joseph Classen, Juliana der Ethikkommission an der medizinischen Fakultät der Coleman, Robert Coleman, Yaroslau Compta, Anne-Gaëlle Universität Leipzig, Leipzig, Germany; Ethikkommission der Corbillé, Jean-Christophe Corvol, Mariana Cosgaya, Nabila Fakultät für Medizin der Technischen Universität München, Dahodwala, Philippe Damier, Elodie David, Thomas Davis, München, Germany; Ethikkommission der Universität Ulm Marissa Dean, Berengere Debilly, Janell DeGiorgio, Andres (Oberer Eselsberg), Ulm, Germany; Landesamt für Gesundheit Deik, Laure Delaby, Marie-Helene Delfini, Pascal Derkinderen, und Soziales Berlin and Geschäftsstelle der Ethik-Kommission Philipp Derost, Maria de Toledo, Lisa Deuel, Ann Marie des Landes Berlin, Berlin, Germany; Ethikkommission des Diaz-Hernandez, Cameron Dietiker, Karina Dimenshteyn, Julio FB Medizin der Philipps-Universität Marburg, Marburg, Dotor, Franck Durif, Jens Ebentheuer, Karla Maria Eggert, Germany; Ethikkommission an der Medizinischen Fakultät Sara Eichau Madueño, Claudia Eickhoff, Aaron Ellenbogen, der Eberhard-Karls-Universität und am Universitätsklinikum Philipp Ellmerer, Ines Esparragosa Vazquez, Alexandre Eusebio, Tübingen, Tübingen, Germany; Ethikkommission an der Siobhan Ewert, John Fang, Danielle Feigenbaum, Frederique Med. Fakultät der HHU Düsseldorf, Düsseldorf, Germany; Fluchere, Alexandra Foubert-Samier, Marie Fournier, Anne Ethikkommission der LÄK Hessen, Frankfurt, Germany; Fradet, Valerie Fraix, Samuel Frank, Franca Fries, Monique CEIm Hospital Universitari Vall d’Hebron, Barcelona, Spain; Galitzky, Marisol Gallardó Pérez, Jose Manuel García Moreno, Copernicus Group Independent Review Board, Puyallup, Carmen Gasca, Thomas Gasser, Joyce Gibbons, Caroline Washington, USA; Western Institutional Review Board, Giordana, Alicia Gonzalez Martinez, Ira Goodman, Arantza Puyallup, Washington, USA; The University of Kansas Medical Gorospe, Marie Goubeaud, David Grabli, Mangone Graziella, Center Human Research Protection Program, Kansas City, Stephan Grimaldi, Jeffrey Gross, Raquel Guimaraes-Costa, Kansas, USA; Oregon Health & Science University Independent Andreas Hartmann, Christian Hartmann, Travis Hassell, Robert Review Board, Portland, Oregon, USA; Northwestern University Hauser, Antonio Hernandez, Jorge Hernandez-Vara, Günter Institutional Review Board, Chicago, Illinois, USA; Spectrum Höglinger, Christian Homedes, Andrea Horta-Barba, Jean-Luc Health Human Research Protection Program, Grand Rapids, Houeto, Julius Huebl, Jennifer Hui, Stuart Isaacson, Joseph Michigan, USA; The University of Vermont Committees Jankovic, Annette Janzen, Junior Jauregui, Jocelyne Jiao, Maria on Human Subjects, Burlington, Vermont, USA; Beth Jose Marti Domenech, Xavier Joseph, Srinath Kadimi, Pat Israel Deaconess Medical Center Committee on Clinical Kaminski, Silja Kannenberg, Jan Kassubek, Maya Katz, Kevin Investigations, New Procedures and New Forms of Therapy, Klos, Shannon Klos, Christopher Kobet, Jennifer Koebert, Boston, Massachusetts, USA; Vanderbilt Human Research Patricia Krause, Andrea Kuehn, Jaime Kulisevsky Bojarsky, Protection Program Health, Boston, Massachusetts, USA; Rajeev Kumar, Martin Kunz, Lille Kurvits, Kimberly Kwei, Vanderbilt Human Research Protection Program Health, Simon Laganiere, Brice Laurens, Johannes Levin, Oren Levy, Nashville, Tennessee, USA; University of Maryland, Baltimore Peter LeWitt, Gurutz Linazasoro Cristóbal, Irene Litvan, Karlo Institutional Review Board, Baltimore, Maryland, USA; Lizarraga, Katherine Longardner, Rocio Lopez, Lydia Lopez University of Southern California Institutional Review Board, Manzanares, Sara Lucas del Pozo, Maria Rosario Luquin Puido, Los Angeles, California, USA; Columbia University Medical Nijee Luthra, Kelly Lyons, Sylvia Maass, Gerrit Machetanz, Center Institutional Review Board, New York, New York, USA; Yolanda Macias, David Maltete, Jorge Uriel Manez Miro, Louise- University of Southern California San Francisco Institutional Laure Mariani, Juan Marin, Kathrin Marini, Ana Marques, Review Board, San Francisco, California, USA; University Gloria Marti, Maria Jose Marti Domenech, Saul Martinez, of Pennsylvania Institutional Review Board, Philadelphia, Wassilios Meissner, Sara Meoni, Brit Mollenhauer, Dunia Philadelphia, USA; HCA—HealthOne Institutional Review Mon Martinez, Johnson Moon, Elena Moro, Peter Morrison, Board, Denver, Colorado, USA. The patients/participants Christoph Muehlberg, Manpreet Multani, Christine Murphy, provided their written informed consent to participate in Anthony Nicholas, Rajesh Pahwa, Antonio Palasi, Heidi Pape, this study. Neepa Patel, Prity Patel, Marina Peball, Elizabeth Peckham, Terry Peery, Rafael Perez, Jesus Perez, Alisa Petit, Elmar Pinkhardt, Werner Poewe, Elsa Pomies, Cecile Preterre, Joseph PASADENA INVESTIGATORS Quinn, Olivier Rascol, Philippe Remy, Irene Richard, Benjamin Roeben, Emily Ruether, Jost-Julian Rumpf, David Russell, Hayet Claudia Altendorf, Chareyna Anandan, Giulia Andrews, Salhi, Daniela Samaniego-Toro, Alexandra Samier-Foubert, Solène Ansquer, Raphaele Arrouasse, Sana Aslam, Jean- Antonio Sanchez, Emmanuelle Schmitt, Alfons Schnitzler, Oliver Philippe Azulay, Jeanette Baker, Ernest Balaguer Martinez, Schorr, Julie Schwartzbard, Kerstin Schweyer, Klaus Seppi, Shadi Barbu, Kara Bardram, Danny Bega, Helena Bejr-Kasem Victoria Sergo, Holly Shill, Andrew Siderowf, Tanya Simuni, Marco, Isabelle Benatru, Eve Benchetrit, Felix Bernhard, Amir Umberto Spampinato, Ashok Sriram, Natividad Stover, Caroline Besharat, Sagari Bette, Amelie Bichon, Andrew Billnitzer, Tanner, Arjun Tarakad, Carolyn Taylor, Claire Thalamus, Sophie Blondeau, Thomas Boraud, Freiderike Borngräber, James Thomas Toothaker, Nadege Van Blercom, Nora Vanegas- Boyd, Kathrin Brockmann, Matthew Brodsky, Ethan Brown, Arrogave, Lydia Vela, Sylvian Vergnet, Tiphaine Vidal, Jonathan Christof Bruecke, Fabienne Calvas, Monica Canelo, Federico Vöglein, Ryan Walsh, Cheryl Waters, Mirko Wegscheider, Endy Frontiers in Neurology | www.frontiersin.org 13 October 2021 | Volume 12 | Article 705407 Pagano et al. PASADENA Phase II Study Design Weidinger, Caroline Weill, Gregor Wenzel, Tatiana Witjas, critically revised important intellectual content. GP and ABon Isabel Wurster, Brenton Wright, Milan Zimmermann, and provided final approval of the manuscript. All authors were Rafael Zuzuarregui. involved in data interpretation and revised and gave input on the article. PRASINEZUMAB STUDY GROUP FUNDING Markus Abt, Atieh Bamdadian, Teresa Barata, Nicholas Barbet, Sara Belli, Frank Boess, Azad Bonni, Edilio Borroni, The authors declare that F. Hoffmann-La Roche Ltd was the Anne Boulay, Markus Britschgi, Jerome Chague, Valerie sponsor and sole funder of the study. Cosson, Christian Czech, Dennis Deptula, Cheikh Diack, Rachelle Doody, Juergen Dukart, Giulia D’Urso, Sebastian Dziadek, Hannah Eddleston, Chris Edgar, Laurent Essioux, ACKNOWLEDGMENTS Morgan Farell, Rebecca Finch, Paulo Fontoura, Waltraud The authors thank all the subjects who participated in Gruenbauer, Andrea Hahn, Stefan Holiga, Michael Honer, this study. PPMI was a public-private partnership sponsored Shirin Jadidi, Kelly Johnson-Wood, Markus Keller, Timothy by the Michael J. Fox Foundation (MJFF) for Parkinson’s Kilchenmann, Martin Koller, Thomas Kremer, Thomas Research and is co-funded by MJFF, AbbVie, Allergan, Amathus Kustermann, Claire Landsdall, Michael Lindemann, Florian Therapeutics, Avid Radiopharmaceuticals, Bial Biotech, Biogen Lipsmeier, Cecile Luzy, Marianne Manchester, Maddalena Idec, BioLegend, Bristol-Myers Squibb, Calico, Celgene, Denali Marchesi, Ferenc Martenyi, Meret Martin-Facklam, Katerina Therapeutics Inc., 4D Pharma Plc, Eli Lilly and Company, Mironova, Annabelle Monnet, Emma Moore, Daniel K Ness, F. Hoffmann-La Roche Ltd., GE Healthcare, Genentech Inc., Markus Niggli, Tania Nikolcheva, Susanne Ostrowitzki, GlaxoSmithKline, Golub Capital BDC, Handl Therapeutics, Gennaro Pagano, Benedicte Passmard, Agnes Poirier, Insitro, Janssen Neuroscience, Lundbeck, Merck, Meso Scale, Anke Post, Megana Prasad, Nathalie Pross, Tiffany Quock, Neurocrine Biosciences, Pfizer, Piramal, Prevail Therapeutics, Benedicte Ricci, Ellen Rose, Christoph Sarry, Christine Sanofi Genzyme, Servier, Takeda, Teva Pharmaceutical Industries Schubert, Dennis Selkoe, Jeff Sevigny, Kaycee Sink, Hannah Ltd., UCB, Verily Life Sciences, and Voyager Therapeutics. Staunton, Tim Steven, Alexander Strasak, Hanno Svoboda, Industry partners contribute to PPMI through financial and Kirsten Taylor, Radhika Tripuraneni, Dylan Trundell, Daniel in-kind donations and have a lead role in providing feedback Umbricht, Lynne Verselis, Annamarie Vogt, Ekaterina on study parameters through the Partners Scientific Advisory Volkova-Volkmar, Cornelia Weber, Silke Weber, and Board (PSAB). Through close interaction with the study, the Wagner Zago. PSAB was positioned to inform the selection and review of potential progression markers that could be used in AUTHOR CONTRIBUTIONS clinical testing. GP, FB, KT, BM, WP, GK, WZ, DN, SO, KM, MK, and JS designed the study. GP, FB, KT, BM, WP, ABou, AV, MM, SUPPLEMENTARY MATERIAL AP, SO, and TS were involved in data collection. GP, KT, BR, JA-C, AV, and MM analyzed the data. GP and ABon The Supplementary Material for this article can be found drafted the work. 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Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or Conflict of Interest: GP, FB, KT, BR, JA-C, AV, MM, TN, HS, MB, SO, RD, DU, endorsed by the publisher. and ABon are employees of F. Hoffmann-La Roche Ltd. ABou is an employee of Idorsia Pharmaceuticals Ltd. AP is an employee of Feetme SAS. JS is an employee of Prevail Therapeutics. GK, WZ, DN, and MK are employees of Copyright © 2021 Pagano, Boess, Taylor, Ricci, Mollenhauer, Poewe, Boulay, Prothena Biosciences Inc. The authors declare that F. Hoffmann-La Roche Anzures-Cabrera, Vogt, Marchesi, Post, Nikolcheva, Kinney, Zago, Ness, Ltd was the sponsor of the study. F. Hoffmann-La Roche Ltd and Prothena Svoboda, Britschgi, Ostrowitzki, Simuni, Marek, Koller, Sevigny, Doody, Corporation plc were the funders of the study. F. Hoffmann-La Roche Ltd and Fontoura, Umbricht, Bonni and PASADENA Investigators and Prasinezumab Prothena Corporation plc were involved in the study design, collection, analysis, Study Group. This is an open-access article distributed under the terms of interpretation of data, the writing of this article and the decision to submit it for the Creative Commons Attribution License (CC BY). The use, distribution publication. or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in The remaining authors declare that the research was conducted in the absence of this journal is cited, in accordance with accepted academic practice. No use, any commercial or financial relationships that could be construed as a potential distribution or reproduction is permitted which does not comply with these conflict of interest. terms. Frontiers in Neurology | www.frontiersin.org 17 October 2021 | Volume 12 | Article 705407