A Randomized, Double Blind, Placebo-controlled Study to Evaluate the Impact of Nilotinib Treatment on Safety, Tolerability, Pharmacokinetics and Biomarkers in Dementia With Lewy Bodies (DLB)
Phase: PHASE2
Conditions: Dementia With Lewy Bodies
Sponsor: Georgetown University

ClinicalTrials.gov Data Entry Cover Sheet 
Protocol ID: STUDY00000122 
Clinical Trials.gov ID: NCT04002674  
Project Title: Safety, Cognitive, and Behavioral Outcomes in Patients with Dementia with Lewy Bodies 
Treated with Nilotinib 
Principal Investigator: Fernando Pagan 
Document Date: 2/21/2019

 
 
 
 
CLINICAL TRIAL PROTOCOL 
 
 
A randomized, double blind, placebo-controlled study to evaluate the 
impact of Nilotinib treatment on safety, tolerability, pharmacokinetics 
and biomarkers in Dementia with Lewy Bodies (DLB) 
 
Author: 
 
Investigators: 
Charbel Moussa MBBS, PhD 
 
Fernando Pagan, MD 
Giuseppe Esposito, MD 
Raymond Scott Turner, MD, PhD 
Charbel Moussa, MBBS, PhD  
Yasar Torres-Yaghi, MD 
Michaeline Hebron, MS (Biomarkers and Data Manager) 
Jaeil Ahn, PhD (Biostatistician) 
Tracy Edwards, NP (Nurse Practitioner) 
Barbara Wilmarth, NP (Nurse Practitioner) 
Helen Howard, RN (Outreach Nurse Coordinator) 
 
Georgetown University Medical Center (GUMC) 
Lewy Body Dementia Research Center of Excellence 
National Parkinson's Foundation Center for Excellence 
Translational Neurotherapeutics Program (TNP) 
4000 Reservoir Rd, NW, 
Building D, Room 203-C 
Washington D.C. 20057. USA. 
Study Product: 
Nilotinib (Tasigna®, AMN107, Novartis) 
Development phase: 
II 
Draft or Version Number: 
v01 (June 23, 2020) 
 
Property of Georgetown University Medical Center may not be used, divulged, published, or 
otherwise disclosed without the consent of Georgetown University 
Clinical Trial Protocol Template Version 1.0, April 5, 2018 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
1 
 
 
TABLE OF CONTENTS 
 
List of Abbreviations 
1- PROTOCOL SUMMARY AND RATIONALE 
 
 
 
 
 
PAGE 
1.1- Study Title 
 
 
 
 
 
 
 
 
 
6 
1.2- Version Number 
 
 
 
 
 
 
 
 
6 
1.3- Study Indication 
 
 
 
 
 
 
 
 
6 
1.4- Phase of Development 
 
 
 
 
 
 
 
6 
1.5- Background 
 
 
 
 
 
 
 
 
 
6 
1.6- Objectives and Endpoints  
 
 
 
 
 
 
9 
1.7- Study Rationale 
 
 
 
 
 
 
 
 
13 
 
2-  STUDY DESIGN  
 
 
 
 
 
 
 
 
 
14 
2.1- Treatment 
 
 
 
 
 
 
 
 
 
14 
2.2- Drug Administration  
 
 
 
 
 
 
 
14 
2.3- Dosing Guidelines 
 
 
 
 
 
 
 
 
14 
2.4- Planned Exposure (e.g. Duration of the study administration) 
2.5- Number of Planned Subjects and Treatment Plan 
 
 
 
15 
2.6- Study Population 
 
 
 
 
 
 
 
 
15 
 
3- VISIT SCHEDULE AND ASSESSMENTS 
 
 
 
 
 
16 
 
4- PATIENT POPULATION  
 
 
 
 
 
 
 
29 
4.1- Number of Patients & Centers 
 
 
 
 
 
 
19 
4.2- Inclusion Criteria 
 
 
 
 
 
 
 
 
19 
4.3- Exclusion Criteria 
 
 
 
 
 
 
 
 
20 
4.4- Randomization and Registration   
 
 
 
 
 
21 
4.5- Blinding 
 
 
 
 
 
 
 
 
 
21 
4.6- Un-blinding  
 
 
 
 
 
 
 
 
 
21 
4.7- Withdrawal of subjects 
 
 
 
 
 
 
 
21 
4.8- Handling of Withdrawals 
 
 
 
 
 
 
 
22 
4.9- Termination of Study 
 
 
 
 
 
 
 
22 
4.10- Protocol Adherence 
 
 
 
 
 
 
 
22 
 
5- SAFETY MONITORING REPROTING  
 
 
 
 
 
23 
5.1- All Non-serious Adverse Events   
 
 
 
 
23 
 
 
5.2- All Serious Adverse Events 
 
 
 
 
 
23 
 
 
5.3- All Reports of Drug Exposure during Pregnancy 
 
 
24 
 
 
5.4- All reports of misuse and abuse of Nilotinib 
 
 
 
24 
 
 
5.5- Assessment and Recording of Adverse Events 
 
 
25 
5.6- Assessment of Adverse Events 
 
 
 
 
 
25 
5.7- Relatedness of Adverse Event to Investigational Product 
 
25 
 
 
5.8- Recording of Adverse Events 
 
 
 
 
 
26 
 
6-  STUDYMONTORING 
 
 
 
 
 
 
 
 
27 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
2 
 
 
 
 
6.1- Safety Monitoring 
 
 
 
 
 
 
27 
 
 
 
6.2- Data Safety and Monitoring Board (DSMB) 
 
 
27 
 
 
 
6.3- Study Monitor and auditing 
 
 
 
 
27 
 
7- Institutional Review Board (IRB) 
 
 
 
 
 
 
 
28 
7.1- Ethical Conduct of Study 
 
 
 
 
 
 
 
28 
7.2- Subject Information and Consent  
 
 
 
 
 
28 
 
8- STATISTICAL METHODS AND DATA ANALYSIS 
 
 
 
 
29 
8.1- Sample size determination  
 
 
 
 
 
 
29 
8.2- Analyses Plan  
 
 
 
 
 
 
 
 
30 
 
 
9- STUDY FEASIBILITY AND PRELIMINARY DATA 
 
 
 
 
31 
 
 
9.1- Preclinical Evidence  
 
 
 
 
 
 
31 
 
 
9.2- Human Studies 
 
 
 
 
 
 
 
37 
 
 
 
 
9.2. a- Pharmacokinetic studies  
 
 
 
39 
 
 
 
 
9.2. b- Pharmacodynamics 
 
 
 
 
39 
 
 
9.3.- Safety  
 
 
 
 
 
 
 
 
40 
 
 
 
 
9.3.1- Serious Adverse Events  
 
 
 
40 
 
 
 
 
9.3.2- Non-Serious AEs  
 
 
 
 
40 
 
 
 
 
9.3.3- QTc prolongation  
 
 
 
 
41 
 
 
 
 
9.3.4- Blood chemistry 
 
 
 
 
 
43 
 
 
9.4- Disease-related biomarker studies 
 
 
 
 
44 
 
 
9.5- Nilotinib ameliorates motor imprairment 
 
 
 
46 
 
 
9.6- Nilotinib improves cognitive symptoms  
 
 
 
47 
 
10- TREATMENTS ADMINISTERED 
 
 
 
 
 
 
 
48 
 
 
10.1- Name of the drug and all active ingredients  
 
 
48 
 
 
10.2- Pharmacological Class of the Drug 
 
 
 
 
48 
 
 
10.3- Structural Formula of the Drug  
 
 
 
 
48 
10.4- Formulation of the dosage form to be used 
 
 
 
48 
10.5- Summary of Previous Human Experience   
 
 
 
49 
10.6- Status of Drug in Other Countries 
 
 
 
 
50 
10.7- Drug Related Risks and Potential Side Effects 
 
 
50 
 
11- PRIOR AND CONCOMITANT THERAPY  
 
 
 
 
 
51 
11.1- Prohibited Medications and Contraindications 
 
 
51 
 
12- RATIONALE- NILOTINIB TO TREAT NEURODEGENERATIVE DISEASES  
52 
 
13- CLINICAL ASSESSMENTS AND OUTCOME MEASURES 
 
 
 
54 
 
 
13.1- Clinical Variables 
 
 
 
 
 
 
 
54 
13.2- Vital Signs, Height & Weight 
 
 
 
 
 
54 
13.3- Clinical Laboratory Assessments 
 
 
 
 
54 
13.4- Physical Examination 
 
 
 
 
 
 
55 
13.5- Neurological Examination  
 
 
 
 
 
55 
13.6- Adverse Events 
 
 
 
 
 
 
 
56 
13.7- MDS-UPDRS   
 
 
 
 
 
 
 
56 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
3 
 
13.8- TMT-B  
 
 
 
 
 
 
 
 
56 
13.9- MoCA   
 
 
 
 
 
 
 
 
56 
13.10- C-SSRS 
 
 
 
 
 
 
 
 
56 
13.11- TUG  
 
 
 
 
 
 
 
 
57 
13.12- NPI 
  
 
 
 
 
 
 
 
 
57 
13.13- ADAS-cog  
 
 
 
 
 
 
 
57 
13.14- ADCS-ADL  
 
 
 
 
 
 
 
57 
13.15- CAF  
 
 
 
 
 
 
 
 
57 
13.16- IAS 
 
 
 
 
 
 
 
 
 
57 
13.17- PBAs  
 
 
 
 
 
 
 
 
57 
13.16- CSF and Blood Biomarkers  
 
 
 
 
 
57 
 
14- BIOMARKERS  
 
 
 
 
 
 
 
 
 
58 
14.1- Rationale for CSF and plasma biomarkers  
 
 
 
58 
14.2- Pharmacokinetics  
 
 
 
 
 
 
59 
14.3- Amyloid PET  
 
 
60 
14.4- Cell death and exploratory biomarkers  
 
 
 
60 
14.5- Alpha-Synuclein ELISA 
 
 
 
 
 
 
60 
14.6- Homovanillic Acid and DOPAC ELISA  
 
 
 
60 
14.7- Total Tau and p-Tau181 measurement.  
 
 
 
61 
14.8- Human Neurodegenerative Disease Panels 
 
 
 
61 
 
15- DATA COLLECTION  
 
 
 
 
 
 
 
 
62 
15.1- Role of Data Management 
 
 
 
 
62 
15.2- Data Entry and Checks  
 
 
 
 
62 
15.3- Data Lock Process 
 
 
 
 
 
62 
15.4- Data handling and record keeping 
 
 
 
62 
15.5- Confidentiality 
 
 
 
 
 
 
62 
15.6- Retention of Records 
 
 
 
 
 
62 
15.7- Publications 
 
 
 
 
 
 
63 
 
16- LITERATURE REFERENCES 
 
 
 
 
 
 
 
64 
 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
4 
 
LIST OF ABBREVIATIONS 
 
AE 
Adverse Event/Adverse Experience 
AD 
Alzheimer’s Disease 
ADCS-ADL 
Alzheimer’s Disease Co-operative Study-Activity of Daily Living 
ADAS-cog 
Alzheimer’s Disease Assessment Scale-cognition 
ALT 
Alanine Aminotransferase 
AST 
Aspartate Aminotransferase 
Abl 
Abelson Tyrosine Kinase 
BBB 
Blood Brain Barrier 
BUN 
Blood Urea Nitrogen 
CIB 
Clinical Investigator’s Brochure 
CBC 
Complete Blood Count 
CNS 
Central Nervous System 
CPK 
Creatine Phosphokinase 
CRF 
Case Report Form 
CSF 
Cerebrospinal Fluid 
C-SSRS 
Columbia Suicide Severity Rating Scale 
CRU 
Clinical Research Unit 
CML 
Chronic Myeloid Leukemia 
CAF 
Clinical Assessment of Fluctuation 
DSMB 
Data Safety Monitoring Board 
DDR1 
Discoidin Domain Receptor-1  
DLB 
Dementia with Lewy bodies 
DMSO 
Dimethyl Sulfoxide 
DOPAC 
3.4-Dihydroxyphenylacetic acid 
DA 
Dopaminergic 
EDC 
Electronic Data Capture 
ELISA 
Enzyme-Linked Immunosorbent Assay 
FTLD 
Fronto-Temporal Lobar Dementia 
EKG 
Electrocardiogram 
GUMC 
Georgetown University Medical Center 
γ-GT 
Gamma-Glutamyl Transferase  
HVA 
Homovanillic Acid 
Hb 
Hemoglobin 
HCG 
Human Chorionic Gonadotropin 
HDL 
High Density Lipoprotein 
ICH 
International Conference on Harmonization 
IND 
Investigational New Drug Application 
IHC 
Immunohistochemistry 
IRB 
Institutional Review Board 
ITT 
Intent to treat 
I.P. 
Intraperitoneal 
IAS 
Irritability-Apathy Scale 
IV 
Intravenous 
LP 
Lumbar Puncture 
LDL 
Low Density Lipoprotein 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
5 
 
LDH 
Lactate Dehydrogenase 
LB 
Lewy Bodies 
MoCA 
Montreal Cognitive Assessment 
MMSE 
Mini Mental Status Exam 
MDS 
Movement Disorders Society 
N 
Number (typically refers to subjects) 
NPI 
Neuropsychiatric Inventory 
NSE 
Neuron Specific Enolase 
NIH 
National Institutes of Health 
OHRP 
Office for Human Research Protections 
OHSR 
Office of Human Subjects Research 
PHI 
Protected Health Information 
PHRC 
Partners Human Research Committee 
PDGF 
Platelet Derived Growth Factor 
PET 
Positron Emission Tomography 
PI 
Principal Investigator 
PT-INR 
Prothrombin Time-International Normalized Ratio  
PK 
Pharmacokinetics 
PDD 
Parkinson’s Disease with Dementia 
PD 
Parkinson’s disease 
PBAs 
Problem Behaviors Assessment –short form 
SAE 
Serious Adverse Event/Serious Adverse Experience 
SBP 
Systolic Blood Pressure 
SI 
Site Investigator 
SMC 
Safety Monitoring Committee 
SOP 
Standard Operating Procedure 
SN 
Substantia Nigra 
SCOPA-Cog 
Scales for Outcomes in Parkinson’s Disease-Cognition 
Src 
Protein Kinase SRC 
TDP-43 
Trans-Activation DNA Binding Protein-43 
TKI 
Tyrosine Kinase Inhibition 
TNF 
Tumor Necrosis Factor 
TREM2 
Triggering Receptors on Myeloid Cells 
TMT-B 
Trail Making Test-B 
TUG 
Timed-Up and Go 
VEGF 
Vascular Endothelial Growth Factor 
UPDRS 
Unified Parkinson’s Disease Rating Scale 
UIS 
University Information Service 
WBC 
White Blood Cells 
 
 
 
 
 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
6 
 
1- SUMMARY AND RATIONALE 
 
1.1- 
Study Title 
A randomized, double blind, placebo-controlled study to evaluate the impact of Nilotinib (Tasigna®) on 
safety, tolerability, pharmacokinetics and biomarkers in Dementia with Lewy Bodies (DLB) 
 
1.2- Version Number 
Original Protocol 
 
1.3- Study Indication 
Dementia with Lewy Bodies (DLB) 
 
1.4- Phase of Development 
II 
 
1.5- Background  
Dementia with Lewy Bodies (DLB) is an alpha-synucleinopathy (1,2) and the second most common 
form of dementia in the elderly. DLB shares striking neuropathological and clinical similarities with both 
Parkinson's disease (PD) (2) and Alzheimer's disease (AD) (3,4). The core clinical features of DLB, 
include dementia and Parkinsonism in addition to hallucinations, cognitive fluctuations and rapid eye 
movement (REM) sleep behavior disorders (RBD) (5-7). DLB and PD are characterized by death of 
dopaminergic (DA) neurons in the nigro-striatal system (8-13) and formation of inclusions known as 
Lewy bodies (LBs) that primarily contain aggregated alpha-synuclein (14-16). Misfolded alpha-
synuclein accumulates intra-neuronally within LBs and SYN (alpha-synuclein) is the highest genetic 
risk factor for PD and DLB (1,2) followed by the microtubule associated protein tau (MAPT) (17-21). 
The neuropathology of DLB overlaps with both PD and AD, and include alpha-synuclein accumulation 
in LBs, hyper-phosphorylated tau (p-tau) and beta-amyloid deposition (4,22-27). At autopsy alpha-
synuclein, p-tau and amyloid plaques may co-exist in the brains of individuals with DLB (26,28). 
Cerebrospinal fluid (CSF) biomarkers, including alpha-synuclein, dopamine metabolite homovanillic 
acid (HVA) (9,29), total tau and p-tau and amyloid beta peptides (Abeta40/42) maybe commonly shared 
in AD, PD and DLB (27,30-32). L-Dopa replacement therapies and acetylcholinesterase inhibitors may 
partially abate motor and cognitive symptoms, respectively in DLB. Selective Serotonin Re-uptake 
Inhibitors (SSRIs) and antipsychotics manage the behavioral but worsen motor symptoms in DLB. 
 
A major challenge facing DLB is to develop a therapy that can halt neuronal death and alleviate 
cognitive, motor and behavioral symptoms. Although DLB is the second most common dementia and 
a debilitating movement disorder, it stands as a unique disease with complicated pathology and 
symptomatology; however, research of DLB has been largely overshadowed by AD and PD.  No 
therapeutic approach exists to alter the levels of neurotoxic proteins such as alpha-synuclein, p-tau and 
Abeta40/42 and halt DA and other neuronal death in DLB. There is a major unmet medical need for 
further research into DLB to identify potential therapies for this disease and provide significant insights 
into the treatment of other Parkinsonian and memory disorders. One mechanism to degrade neurotoxic 
proteins is autophagy (33-37), which is a process by which the cell can degrade its own contents. There 
is evidence that autophagy is impaired in neurodegeneration (38-45), leading to failure of degradation 
of protein aggregates, including misfolded alpha-synuclein, beta amyloid and tau. Importantly, 
autophagy is exploited therapeutically in several diseases, including adult chronic myeloid leukemia 
(CML). Tyrosine kinase inhibitors (TKIs) induce autophagy (37,46,47), leading to destruction of rapidly 
dividing tumor cells in CML (46) and degradation of neurotoxic proteins, including alpha-synuclein, 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
7 
 
beta-amyloid and p-tau in PD and AD models (37,46-49). Nilotinib (Tasigna®, AMN107, Novartis, 
Switzerland) is approved by the U.S. Food and Drug Administration (FDA) and is well tolerated for CML 
treatment at oral doses of 600-800mg daily. Nilotinib penetrates the brain and promotes autophagic 
degradation of neurotoxic proteins, leading to survival of DA and other neurons and improvement of 
motor and cognitive behavior in animal models of alpha-synucleinopathy and other neurodegenerative 
diseases (37,46-50). For these studies, Nilotinib (1-10mg/kg daily) was used at significantly less than 
the clinically approved dose (up to 1200mg daily) in CML (51-53). Additionally, Nilotinib levels peak in 
the mouse brain 4hrs after intraperitoneal (I.P) injection and wash out by 8hrs (37,47). This is important 
because the lower dose and short bio-availability (up to 8hrs) will prevent Nilotinib side effects and 
induce autophagic degradation of neurotoxic proteins in post-mitotic neurons, providing a strategy to 
prevent forcing neurons into apoptosis.  
Based on strong pre-clinical evidence of the effects of Nilotinib on neurodegenerative pathologies, 
including autophagic clearance of neurotoxic proteins, neurotransmitters (dopamine and glutamate), 
immunity and behavior (49,50,54-58), we conducted an open label pilot clinical trial in individuals with 
advanced PD with dementia (PDD) and Dementia with Lewy Bodies (DLB) (59). Participants (N=12) 
were randomized 1:1 to once daily oral dose of 150mg and 300mg Nilotinib for 6 months. Our data 
suggest that Nilotinib penetrates the brain and inhibits brain Abelson (Abl) activity in  animal models 
but achieves CSF concentration that would more preferentially targets Discoidin Domain Receptor 
(DDR-1) in  humans , including PD and AD, independent of Abl activity (37,59-64). Several studies 
show that CSF alpha-synuclein, Abeta42, total tau and p-tau181 are altered in PD and DLB (65-67). 
Our data show stabilization of total CSF alpha-synuclein but a reduction in oligomeric alpha-synuclein 
levels between baseline and 6-months treatment with 150mg-300mg Nilotinib (see data). CSF HVA, 
which is an end by-product of dopamine, was also significantly increased; and CSF total tau and p-tau 
were significantly reduced (N=5, P<0.05) with 300mg Nilotinib between baseline and 6-month 
treatment. Despite the reduction of L-Dopa replacement therapies in this study, scores using the Unified 
Parkinson’s Disease Rating Scale (UPDRS) I-IV improved with 150mg (3.5 points) and 300mg (11 
points) from baseline to 6 months and worsened (13.7 points and 11.4 points) after 3-month withdrawal 
of 150mg and 300mg, respectively. Other non-motor functions e.g. constipation was resolved in all 
patients and cognition was also improved (3.5 points) using both the Mini-Mental Status Exam (MMSE) 
or the Scales for Outcomes in Parkinson’s Disease-Cognition (SCOPA-Cog) between baseline and 6 
months. MMSE scores returned to baseline after 3 months of Nilotinib withdrawal. It is important to note 
that participants who received oral daily dose of 150mg Nilotinib in this phase I open label study were 
all individuals with DLB. We conducted further phase II dose-finding random multiple dose studies that 
included placebo, 100mg, 200mg, 300mg and 400mg administered orally once 1-4 hours prior to lumbar 
puncture (LP) in PD patients. We observed that 200mg oral dose of Nilotinib results in a significant 
increase in CSF HVA and 3.4-didydroxyphenylacetic acid (DOPAC) concurrent with a decrease in CSF 
oligomeric alpha-synuclein (63). Recently, two randomized, double-blind, placebo-controlled clinical 
studies showed that Nilotinib can increase dopamine in the brain and reduce alpha-synuclein and Tau 
in Parkinson’s disease (62,63). Nilotinib can lower CSF Tau and amyloid in the CSF and reduce amyloid 
burden (plaques) via amyloid Positron Emission Tomography (Amyloid PET) scans In Alzheimer’s 
disease (64). These pathologies including dopamine loss, misfolded of alpha-synuclein and tau and 
increased amyloid burden are common in Dementia with Lewy Bodies. Taken together, data from our 
two studies indicate that 200mg Nilotinib may be an optimal dose to study in DLB patients. Our data 
are very compelling to evaluate the effects of 200mg Nilotinib in a phase II, randomized, double-blinded, 
placebo-controlled trial in patients with DLB.  
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
8 
 
 
 
We hypothesize that 200mg of Nilotinib is safe and tolerable and will alter CSF and plasma 
biomarkers in DLB patients. 
 
a- Primary outcomes: We will determine safety and tolerability using the occurrence of adverse events 
(AEs) of interest, including myelosuppression, QTc prolongation, pancreatic and hepatic disorders as 
per Nilotinib Investigator Brochure (IB). 
 
b- Secondary outcomes: We will determine 1) Nilotinib levels in CSF and plasma and 2) changes of 
DLB related CSF and plasma biomarkers, including HVA, DOPAC, Abeta40/42, total tau, ptau231/181 
and total and oligomeric alpha-synuclein.  
 
We will quantify brain amyloid burden via Florbetaben PET at baseline and 6 months at end of 
treatment. 
 
We predict that Nilotinib will stabilize CSF total alpha-synuclein and reduce oligomeric alpha-
synuclein, Abeta40/42 and p-tau.  
 
c- Clinical outcomes will include assessment of (A) cognitive and behavioral function via Montreal 
Cognitive Assessment (MoCA), the Trail Making Test (TMT)-B, AD Assessment Scale-Cognitive 
subscale (ADAS-cog), AD Cooperative Study-Activity of Daily Living (ADCS-ADL), Neuropsychiatric 
Inventory (NPI), and Clinical Assessment of Fluctuation (CAF), Irritability-Apathy Scale (IAS) and 
Problem Behaviors Assessment short (PBA-s) form. We will also determine the effects of Nilotinib on 
(B) motor function via the Unified Parkinson’s Disease Rating Scale (UPDRS)-I-III and timed-up-and-
go (TUG). 
 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
9 
 
1.6- Objectives and Endpoints 
1- Primary outcomes: we will evaluate the effects of Nilotinib on safety and tolerability  
a- Safety will be measured using the occurrence of adverse events (AEs) and serious adverse 
events (SAEs) deemed to be possibly, probably, or definitely related to the study drug. AEs 
of interest are defined as QTc prolongation, myelosuppression, hepatotoxicity and 
pancreatitis as listed in Table 1. These AEs will be tracked over the course of the trial and 
reviewed by the data and safety monitoring board (DSMB) at scheduled meetings and in 
real time. SAEs and AEs are known to be related to drug use at 800mg daily in cancer. A 
small safety trial using lower oral daily doses of 150 mg and 300 mg Nilotinib in 12 PDD 
(n=6) and DLB (n=6) patients showed one cardiac SAE over a six-month treatment period. 
Based on preliminary clinical data, investigator’s brochure (IB) and scheduled EKGs and 
lab tests, SAEs and AEs will be evaluated real-time and on case-by-case basis. 
b- Tolerability for a given participant will be defined as the ability of participants to remain on 
treatment. Overall tolerability of the drug will be defined as an acceptable number of up to 
25% discontinuations. 
Warnings and precautions: Nilotinib (Tasigna) prescribing information includes risks associated with 
arterial vascular occlusive events, including ischemic heart disease-related cardiac events, peripheral 
arterial occlusive disease and ischemic cerebrovascular events.  
Nilotinib has been shown to prolong cardiac ventricular repolarization as measured by the QT interval 
on the surface EKG in a concentration-dependent manner. Prolongation of the QT interval can result 
in a type of ventricular tachycardia called torsade de pointes, which may result in syncope, seizure, 
and/or sudden death. The baseline QTc range for inclusion into this study is 350-460 ms. If QTc 
prolongs ≥ 60 ms from baseline of individual participants AND to a value ≥ 480 ms, Nilotinib 
must be held and participants will be re-evaluated in one week on a case-by-case basis in 
consultation with a cardiologist before continuing on Nilotinib. If QTcF prolongs ≥ 500 ms then 
Nilotinib will be completely stopped and participants will be withdrawn from the study.  
Nilotinib should not be used in patients who have hypokalemia, hypomagnesemia or long QT 
syndrome. Before initiating Nilotinib electrolyte, calcium and magnesium blood levels should be tested. 
Hypokalemia or hypomagnesemia must be corrected prior to initiating Nilotinib and these electrolytes 
should be monitored periodically during drug treatment [See Table 3]. 
 
 
Gastrointestinal (GI) hemorrhage and effusions (including pleural effusion, pericardial effusion, 
ascites) or pulmonary edema were also reported in a small percentage of patients taken Nilotinib and 
GI hemorrhage and effusions should be monitored at every visit. 
 
The bioavailability of Nilotinib is increased with food, thus Nilotinib must not be taken with food. No 
food should be consumed for at least 2 hours before and for at least 1 hour after the dose is taken. 
 
Significant prolongation of the QT interval may occur when Nilotinib is inappropriately taken with food 
and/or strong CYP3A4 inhibitors and/or medicinal products with a known potential to prolong QT. 
Therefore, co-administration with food must be avoided and concomitant use with strong CYP3A4 
inhibitors and/or medicinal products with a known potential to prolong QT should be avoided [see 
exclusion criteria, section 4.3].  
 
The presence of hypokalemia and hypomagnesemia may further prolong the QT interval.  
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
10 
 
Table 1 summarizes some key laboratory tests to monitor AEs and SAEs and includes stopping rules 
and withdrawal for individual patients. 
Table 1- Monitoring laboratory tests for drug safety and stopping rules or withdrawal.  Grade 3 
classification of organ toxicity is according to standardized definitions of AEs published by the 
National Cancer Institute (NCI) of the National Institutes of Health (NIH), known as the Common 
Terminology Criteria for AEs (CTCAE, Version 4.0) or common toxicity criteria.  
Grade 1 Mild; asymptomatic or mild symptoms; clinical or diagnostic observations only; intervention 
not indicated.  
Grade 2 Moderate; minimal, local or noninvasive intervention indicated; limiting age-appropriate 
instrumental activities of daily living (ADL).  
Grade 3 Severe or medically significant but not immediately life-threatening; hospitalization or 
prolongation of hospitalization indicated; disabling; limiting self-care ADL.  
Grade 4 Life-threatening consequences; urgent intervention indicated.  
Grade 5 Death related to AE.  
Assessments 
Toxicity Criteria 
Endpoints 
Myelosuppression 
≥Grade 3 
 
 
 
 
Thrombocytopenia 
and Neutropenia  
Perform complete blood counts (CBC) every month or 
as needed. Withhold Nilotinib for hematological toxicities 
if absolute neutrophil count (ANC) <1.0 x 109 /L and/or 
platelet count <50 x 109 /L.  
Monitor blood counts every 7 days and if problem 
resolves resume treatment within 2 weeks. Patient’s 
blood counts must be monitored every 7 days. If 
Myelosuppression recurs patients must be withdrawn. 
QT Prolongation 
≥Grade 3 
 
EKGs with a QTc 
≥480 ms 
and/or an increase 
of QTc ≥ 60 ms 
from baseline 
Withhold Nilotinib and perform an analysis of serum 
potassium and magnesium, and if below lower limit of 
normal, correct with supplements to within normal limits. 
Concomitant medication usage must be reviewed. 
Patients with prolonged QTc ≥60 ms change from 
baseline or QTc ≥480 ms should be withdrawn from the 
study unless there is a causative electrolyte abnormality 
that has been corrected or CYP inhibitor that may be 
withdrawn. 
Resume within 2 weeks if QTcF returns to <450 ms and 
to within 20 ms of baseline. Repeat EKG 7 days after 
dose adjustment and if QTcF returns to ≥480 ms 
following withholding, Nilotinib must be discontinued. 
Hepatotoxicity 
≥Grade 3 
 
Elevated 
hepatic 
transaminases  
 
Withhold Nilotinib and monitor hepatic transaminases, 
including alanine aminotransferase (ALT), aspartate 
aminotransferase (AST) and alkaline phosphatase 
every 7 days. If transaminase level returns to baseline, 
resume treatment and monitor patients every 7 days. If 
hepatotoxicity recurs patients must be withdrawn from 
the study. 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
11 
 
Bilirubin  
≥Grade 3 
 
Elevated Bilirubin 
 
Withhold Nilotinib and monitor bilirubin every 7 days. If 
bilirubin level returns to baseline, resume treatment and 
monitor patients every 7 days. If bilirubin elevation 
recurs patients must be withdrawn from the study. 
Pancreatitis 
≥Grade 3 
 
Elevated serum 
lipase and 
amylase  
Withhold Nilotinib and monitor lipase and amylase 
every 7 days. If lipase and amylase levels return to 
baseline, resume treatment and monitor patients every 
7 days. If pancreatic toxicity recurs patients must be 
withdrawn from the study. 
 
 
2-Secondary outcomes:  
a) Pharmacokinetics: We will measure the CSF concentration of Nilotinib. 
b) Pharmacodynamics: We will determine the effects of Nilotinib on primary biomarkers, including  
changes of CSF levels of HVA between baseline and 6 months. Based on the data collected from the 
proof of concept study in PD we predict an increase in CSF HVA and DOPAC levels following Nilotinib 
treatment.  
 
We will also measure the CSF concentration of surrogate/exploratory biomarkers: including: 
1) Abeta40/42, total tau, ptau231/181 and total and oligomeric alpha-synuclein 
2) Quantify brain amyloid burden via Florbetaben PET at baseline and 6 months at end of 
treatment. 
- 
We 
will 
use 
ELISA 
to 
measure 
exploratory 
CSF 
and 
plasma 
neuro-
inflammatory/modulatory markers, including Triggering Receptors on Myeloid Cells 
(TREM)-2 interleukins (IL)- 1α&β, 2, 3, 4, 5, 6, 7, 8 (CXCL8), 9, 10, 12, 13, 15, 17 α, and 
chemokines (C-C) including, CXCL10, CCL2, CL7, CCL22, CCL3, CCL4, platelet-derived 
growth factor (PDGF)-AA, PDGF-AB/BB, CCL5, CX3CL1 (fractalkine), Tumor necrosis 
growth factor (TNF)-α, transforming growth factor (TGF)-α, and vascular endothelial growth 
factor (VEGF). We will also measure NSE and S100B as markers of cell death. 
 
3-Clinical outcomes will include assessment of:  
a) cognitive and behavioral function via Montreal Cognitive Assessment (MoCA), the Trail 
Making Test (TMT)-B, AD Assessment Scale-Cognitive subscale (ADAS-cog), AD 
Cooperative Study-Activity of Daily Living (ADCS-ADL), Neuropsychiatric Inventory (NPI), 
and Clinical Assessment of Fluctuation (CAF), Irritability-Apathy Scale (IAS) and Problem 
Behaviors Assessment short (PBA-s) form. 
b) We will also determine the effects of Nilotinib on motor function via the Unified Parkinson’s 
Disease Rating Scale (UPDRS)-I-III and timed-up-and-go (TUG). 
 
Based on preclinical and clinical studies (see below), we hypothesize that Nilotinib will be safe and 
tolerated in individuals with DLB. We also predict that Nilotinib will be detected in the CSF. Nilotinib 
will also increase CSF HVA and DOPAC levels and improve cognitive and motor symptoms in DLB 
patients. Nilotinib may also stabilize CSF levels of total alpha-synuclein, and reduce oligomeric alpha-
synuclein and p-tau levels and reduce CSF amyloid and brain amyloid burden. Nilotinib may affect cell 
death markers, including NSE and S100B and phosphorylated neurofilaments. We further hypothesize 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
12 
 
that we may see evidence of change in surrogate and biomarkers of pathophysiology as outlined in our 
secondary outcomes that will help us to build a better clinical development program going forward. 
 
 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
13 
 
 
1.7- Rationale to advance to Phase II Nilotinib trial in DLB (Study Rationale) 
We performed an open label phase I clinical trial using two commercially available doses of Nilotinib 
(150 and 300mg capsules) in patients with advanced PDD and DLB. These indications have some 
overlapping pathologies and clinical symptoms and share common plasma and CSF biomarkers, 
including loss of dopamine and changes in alpha-synuclein, Abeta42/40, total tau and p-tau (181, 231). 
We obtained preliminary data showing that Nilotinib crosses the blood-brain-barrier (BBB) and is 
detected in the CSF, suggesting Abl inhibition and downstream target engagement (alpha-synuclein, 
tau and Abeta42) in the CNS (pharmacodynamics). Nilotinib increased CSF HVA and DOPAC levels 
as a downstream biomarker of dopamine metabolism. These data provide strong feasibility to test 
Nilotinib in a larger placebo-controlled, phase II clinical trial to demonstrate safety, tolerability, 
pharmacokinetics/pharmacodynamics and changes in disease biomarkers and clinical outcomes in 
patients with DLB. Recently, two randomized, double-blind, placebo-controlled clinical studies showed 
that Nilotinib can increase dopamine in the brain and reduce alpha-synuclein and Tau in moderately 
advanced PD patients (62,63). Nilotinib can lower CSF Tau and amyloid in the CSF and reduce amyloid 
burden (plaques) via amyloid Positron Emission Tomography (Amyloid PET) scans in participnats with 
dementia due to mild-to-moderate AD (64). Pharmacokinetics studies showed that alteration of these 
toxic proteins was independent of Abl Inhibition as Nilotinib reached a CSF concentration sufficient to 
inhibit DDR1 receptors but not Abl. These pathologies including dopamine loss, misfolded of alpha-
synuclein and tau and increased amyloid burden are common in Dementia with Lewy Bodies. Taken 
together, data from our studies indicate that 200mg Nilotinib may be an optimal dose to study in DLB 
patients. Our data are very compelling to evaluate the effects of 200mg Nilotinib in a phase II, 
randomized, double-blinded, placebo-controlled trial in patients with DLB.  
 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
14 
 
3) Study Design 
We propose to perform a phase II randomized, double-blinded, placebo-controlled study to 
evaluate the impact of an oral daily dose of 200mg Nilotinib on safety, tolerability, 
pharmacokinetics, pharmacodynamics and clinical outcomes in patients with DLB.  
 
2.1- Treatment: Sixty (60) participants will be recruited and randomly assigned 1:1 to placebo (group 
1) or 200 mg Nilotinib (group 2). Participants will be treated for 6 months and monitored every month 
(4 weeks) in a total of up to 11 visits that include screening, baseline, 1, 2, 3, 4, 5, 6 months follow up 
and 7-month washout. Blood and cerebrospinal fluid (CSF) will be collected at baseline and at 6 months 
to determine Nilotinib effects on CSF biomarkers. Amyloid PET will also be conducted at baseline and 
end of treatment.  
 
2.2- Drug Administration: Nilotinib will be taken orally once daily (taken without a meal) for 6 calendar 
months and 1 month wash out. Study partners must provide written informed consent prior to screening. 
Based on data obtained in our proof of concept phase I study, a phase II study will primarily examine 
the effects of Nilotinib on safety and tolerability and clinical outcomes.  
 
 
2.3- Dosing Guidelines 
One 200 mg capsule of Tasigna or matching placebo will be administered orally once a day. Key 
elements will be detailed in the educational brochure including special warnings and pre-cautions: 
- 
Brief background on Nilotinib, its authorized indication and posology 
- 
Information on the cardiac risks associated with the use of Nilotinib and that Nilotinib can cause 
prolongation of the QT interval and that patients at risk of arrhythmia, especially torsade de 
pointes, should not take Nilotinib. 
The need to avoid co-prescription with any other medicines that might prolong the QT interval 
- 
Caution in prescribing to patients with a history of or risk factors for coronary heart disease 
- 
That Nilotinib may cause fluid retention, cardiac failure and pulmonary edema 
- 
That Nilotinib is metabolized by CYP3A4 and that strong inhibitors or inducers of this enzyme 
may significantly affect exposure to Nilotinib. 
- 
That inhibitors may increase the potential for adverse drug reactions in particular QT interval 
prolongation 
- 
To warn participants about over the counter (OTC) medicines in particular St John’s Wort 
- 
The need to inform patients about the effects of food on Nilotinib 
- 
 Not to eat within two hours before and one hour after taking Nilotinib 
- 
The need to avoid foods such as grapefruit juice which inhibit CYP3A4 enzymes 
 
2.4- Planned Exposure (e.g. Duration of the study administration)  
We anticipate this project to be completed in 4 years (48 months); including 3 years (36 months) for 
pre-screening and enrolment, 36 months for treatment, neurological and safety examinations and 6 
months for post treatment evaluation as well as data analysis. 
 
 
 
 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
15 
 
Table 2- Summary of primary, secondary and exploratory objectives with the associated 
endpoints and evaluation criteria:   
Outcomes 
Objective 
Endpoint 
Primary 
Safety and 
tolerability  
1- Safety will be measured using the occurrence of AEs) 
and SAEs deemed to be possibly, probably, or 
definitely related to the study drug. AEs of interest are 
defined as QTc prolongation, myelosuppression, 
hepatotoxicity and pancreatitis as listed in Table 1 
and Novartis IB, on which we base our power 
consideration. These AEs will be tracked over the 
course of the trial and reviewed by the data and safety 
monitoring board (DSMB) at scheduled meetings and 
in real time. SAEs and AEs are known to be related 
to drug use at 800mg daily in cancer. A small safety 
trial using lower oral daily doses of 150mg and 300mg 
Nilotinib in 12 PD patients showed one cardiac SAE 
over a six-month treatment period. Based on 
preliminary clinical data, IB and scheduled EKGs and 
lab tests, SAEs and AEs will be evaluated real-time 
on case-by-case basis. 
2- Tolerability for a given participant will be defined as 
the ability of participants to remain on treatment. 
Overall tolerability of the drug will be defined as an 
acceptable number of up to 25% discontinuations. 
Secondary  
Pharmacokinetics 
and biomarkers 
1- Nilotinib level in CSF 
2- CSF levels of HVA and DOPAC between baseline 
and 6 months.  
3- CSF alpha-synuclein (total and oligomeric), total tau 
and 
p-tau, 
Abeta40/42, 
NSE, 
S100B 
and 
phosphorylated neurofilaments. 
4- Amyloid burden between Baseline and 6-months 
using Florbetaben PET 
5- CSF 
ELISA 
and 
plasma 
neuro-
inflammatory/modulatory markers, including TREM2. 
clinical 
outcomes 
Cognition 
and 
behavior 
MoCA, ADAS-cog, ADCS-ADL, CAF, TMT-B, NPI, IAS, 
PBAs. 
 
Motor 
MDS-UPDRS I-III and TUG 
 
2.5- Number of Planned Subjects and Treatment Plan 
Up to 100 subjects will be screened for the study with the goal of enrolling 60 for treatment, including 
30 participants with DLB on placebo (group 1) and 30 participants on 200 mg Nilotinib (group 2). A sub-
group of 50 participants will be enrolled in the amyloid PET study. 
 
2.6- Study Population 
This study will be conducted in DLB patients with 2.5≥Hoehn & Yahr≤3 and UPDRS I-III ≤50 and 
15≥UPDRS III (motor) ≥40 and MoCA≥18. Eligible participants must not be on MAO-B inhibitors 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
16 
 
(Rasagiline or Selegiline) for 4 weeks and must not be on ≥800mg Levodopa daily. Participants must 
be stable on acetylcholinesterase inhibitors and other medications for at least 6 weeks. 
 
4) VISIT SCHEDULE AND ASSESSMENTS 
  
Screening. This visit (Table 3) will determine study eligibility. Potential participants and their 
study partners must review and sign an informed consent form (ICF) prior to any study-related 
procedures. Information regarding demographics, concurrent medications, and medical history will be 
gathered from the participant and study partner at the screening visit. Prior to any study-related 
activities, subjects and their legally authorized representatives (LAR) will be thoroughly informed on all 
aspects of the study and will be requested to sign an ICF. Prior to obtaining written informed consent, 
information will be given at a complexity level that is understandable by the subject in both oral and 
written form by staff. Participants will also be informed about the drug safety and side effects. 
 
Visits/month (m) 
Screening       Baseline 
1m 
2m 
3m 
4m 
5m 
6m 
7m (washout) 
 
Consent 
  
X 
 
 
 
 
 
 
 
 
Demographics 
X 
Vitals, H&P 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Neuro Exam  
X 
X 
X 
X 
X 
X 
X 
X 
X 
EKG 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
Blood draws*  
X 
X 
X 
X 
X 
X 
X 
X 
X 
CSSR-S 
 
X 
X 
X 
X 
X 
X 
X 
X 
X 
TMT-B 
 
X 
X 
 
 
X 
 
 
X 
X 
MDS-UPDRS  
X 
X 
 
 
X 
 
 
X 
X 
MoCA 
 
X 
X 
 
 
X 
 
 
X 
X 
Hoehn & Yahr  
X 
X 
 
 
X 
 
 
X 
X 
CAF  
 
 
X 
X 
X 
X 
X 
X 
X 
X 
PBAs 
 
 
X 
 
 
X 
 
 
X 
X 
IAS  
 
 
X 
 
 
X 
 
 
X 
X 
ADAS-cog 
 
 
X 
 
 
X 
 
 
X 
X 
ADCS-ADL 
 
 
X 
 
 
X 
 
 
X 
X 
NPI  
 
 
X 
 
 
X 
 
 
X 
X 
TUG  
 
 
X 
 
 
X 
 
 
X 
X 
LP  
 
 
X 
 
 
 
 
 
X 
PET  
 
 
X 
 
 
 
 
 
X 
 
Table 3- A detailed schedule of 9 visits (±2 visits) and the assessments done at each visit. History and Physical (H&P), 
Electrocardiograms (EKG), Columbia Suicide Severity Rating Scale (CSSRS), Montreal Cognitive Assessment (MoCA), Trail 
Making Test (TMT)-B, AD Assessment Scale-Cognitive subscale (ADAS-cog), AD Cooperative Study-Activity of Daily Living 
(ADCS-ADL), Neuropsychiatric Inventory (NPI), Irritability-Apathy Scale (IAS), Problems Behavior Assessment Short (PBAs) 
form and Clinical Assessment of Fluctuation (CAF). Movement Disorders Society-Unified Parkinson’s Disease Rating Score 
(MDS-UPDRS)-I-III, timed-up-and-go (TUG) and Lumbar Puncture (LP). 
*Blood draw will be performed for standard blood chemistry, including Cholesterol, low density lipoprotein (LDL), high 
density lipoprotein (HDL), albumin, total protein, alkaline phosphatase, total bilirubin, creatinine, calcium, chloride, sodium, 
potassium, magnesium, inorganic phosphorus, bicarbonate, creatine phosphokinase (CPK), gamma-glutamyl transferase (γ-
GT), lactate dehydrogenase (LDH), lipase, α-amylase, alanine aminotransferase (ALT), aspartate aminotransferase (AST), 
blood urea nitrogen (BUN), thyroid stimulating hormone (TSH), vitamin B12, and glucose. A standard hematology panel 
including, complete blood count (CBC) with differential counts will be performed. Hemoglobin (Hb), hematocrit, red blood cell 
count, platelet count, and white blood cell (WBC) with differential count will be measured. In addition, prothrombin time-
international normalized ratio (PT-INR) will be measured for assessment of coagulation.  
Additional blood draws will be performed 30 minutes before LPs at baseline and 6 months. LPs are mandatory at 
baseline line and end of study visit. LP must be performed within 2 hours from the last Levodopa dose or in ON-STATE.  
Study drug will be dispensed at the end of baseline visit, as well as every visit. 
Visits should be scheduled on a monthly basis or every 30 days (± 3 days). 
Women of child bearing potential must be willing to take a pregnancy test at every visit.  
Unscheduled or miscellaneous visits are allowed if necessary. 
Amyloid PET imaging can be done as a standalone visit between Screening and Baseline visits or during Baseline 
upon confirmation of diagnosis and eligibility. PET should be performed before dispensation of study drug at Baseline and within 
1 week from last dose at end of study ( ± 7 days). 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
17 
 
 
Demographics, vital signs, history and physical (H&P) and neurological exams will be performed 
at the screening visit. The MoCA and TMT-B, Hoehn and Yahr and MDS-UPDRS I-III will be performed 
at screening to confirm diagnosis of DLB and determine eligibility criteria. Electrocardiograms (EKGs) 
will be performed with standard blood chemistry, including cholesterol, low density lipoprotein (LDL), 
high density lipoprotein (HDL), albumin, total protein, alkaline phosphatase, total bilirubin, creatinine, 
calcium, chloride, sodium, potassium, magnesium, inorganic phosphorus, bicarbonate, creatine 
phosphokinase (CPK), gamma-glutamyl transferase (γ-GT), lactate dehydrogenase (LDH), lipase, α-
amylase, alanine aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen 
(BUN), thyroid stimulating hormone (TSH), vitamin B12, and glucose. A standard hematology panel 
including, complete blood count (CBC) with differential counts will be performed. Hemoglobin (Hb), 
hematocrit, red blood cell count, platelet count, and white blood cell (WBC) with differential count will 
be measured. In addition, prothrombin time-international normalized ratio (PT-INR) will be measured 
for assessment of coagulation. Should treatment with any HMG-CoA (or 3-hydroxy-3-methylglutaryl-
coenzyme A) reductase inhibitor (a lipid lowering agent) be needed to treat lipid elevations, evaluate 
the potential for a drug-drug interaction before initiating therapy as certain HMG-CoA reductase 
inhibitors are metabolized by the CYP3A4 pathway. If test results warrant therapy, local standards of 
practice and treatment guidelines will be followed.  
 
The Columbia Suicide Severity Rating Scale (C-SSRS) will be performed at the screening visit.  
 
Baseline visit will be scheduled 2-4 weeks after screening and results from all screening procedures 
will be reviewed and all inclusion/exclusion criteria must be met prior to baseline assessments. The 
Columbia Suicide Severity Rating Scale (C-SSRS) will be performed at baseline visit and at every visit 
as indicated in Table 3. Vital signs, history and physical (H&P), EKG, neurological exams and blood 
draws will be performed at baseline and very other visit as indicated in Table 3. 
 
Cognitive assessments will be performed via MoCA (global cognition), and TMT-B (executive 
function). The MoCA and TMT-B will be done at both screening and baseline to ensure subjects can 
perform these tests and to minimize practice effects on baseline score. ADCS-ADL will measure 
activities of daily living and ADAS-cog will measure global cognitive change. Cognitive fluctuations will 
be measured via CAF. Motor assessments will be performed via UPDRS I-III and Timed-Up and Go 
(TUG).  Psychiatric assessments will be performed using NPI, PBAs and IAS.  
A mandatory blood draw and LP will be performed at baseline. LP must be performed within 2 hours 
from the last Levodopa dose. CSF collection via LP and blood draw will be performed for biomarker 
pharmacokinetics, pharmacodynamics and biomarkers measurement (CSF alpha-synuclein (total and 
oligomeric), CSF total tau/p-tau18, HVA and DOPAC and inflammatory/neurotrophic markers). Blood 
will also be collected for monitoring laboratory tests as indicated above. A mandatory Amyloid PET 
must also be performed upon confirmation of diagnosis and eligibility at or before Baseline.  
The baseline procedures may be completed over several visits. After recruitment, participants will be 
randomized into 2 groups (1:1) receiving a single daily dose of 200 mg Nilotinib or matching placebo 
(group 2).  
Drugs will be dispensed at the end of the baseline visit. 
 
At 1, 2, 4 and 5 months visits, vital signs, history and physical (H&P), EKG, neurological exams and 
blood draws and CSSR-S and CAF will be performed at very other visit as indicated in Table 3. 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
18 
 
  
At 3 and 6 months follow up and 7 months wash out visits, vital signs, history and physical (H&P), 
EKG, neurological exams and blood draws and CSSR-S will be performed. Cognitive assessments 
will be performed via MoCA, ADCS-ADL, and ADAS-cog. Cognitive fluctuations will be measured via 
CAF.  Motor assessments will be performed via UPDRS I-III and Timed-Up and Go (TUG).  
Psychiatric assessments will be performed using NPI, PBAs and IAS. A mandatory blood draw and 
LP will be performed at 6-month end of treatment visit. LP must be performed within 2 hours from the 
last Levodopa dose and within 1-4 hours since last Nilotinib dose, because we detected Nilotinib in 
human CSF up to 4 hours after oral administration (59). 
 
Medication Compliance Check. Site staff will remind subjects to bring unused study drug with 
them at each visit as well as the completed drug compliance diary. Site staff will count the number of 
capsules remaining at each in-person clinic visit for which the subject brings the unused drug with 
him/her. This information will be recorded in the subject specific drug accountability form. The total 
number of returned medication will be reviewed against the drug compliancy diary. If drug compliance 
is determined to be problematic site staff will review with the subject the importance of using the study 
drug as per protocol. Any dose adjustments will be documented on the drug exposure form (indicating 
start and stop date).  Additionally, site staff will check the subject’s overall compliance with the study 
requirements. This will also include checks of protocol compliance, and concomitant medication use, 
in addition to the proper use of study drug in order to assess the reliability of subject generated data.  
 
 
 
Drug Related Risks and Potential Side Effects 
Nilotinib has a number of side effects  as stated in the IB, including gastrointestinal complications, 
vomiting and nausea, and in some rare cases dizziness (51). Abl has an essential role in mammalian 
heart growth and development and adult complications with Abl inhibition, include edema, 
nausea/vomiting, muscle cramps, neutropenia, thrombocytopenia, fever, liver toxicity, arthralgia, and 
exanthema/rash (68). Side effects are identified for Abl in homozygous Abl mutant mice, which display 
dramatically enlarged hearts due to abnormally increased cardiomyocyte proliferation during later 
stages of embryogenesis (69). Disruption of Abl in mice results in neonatal lethality accompanied by 
pleiotropic developmental defects with variable penetrance, including ranting, splenic and thymus 
atrophy, B cell lymphopenia, dysfunctional osteoblasts, and foreshortened crania (70-72). However, 
clinical use of Nilotinib (600-800mg daily) is fairly tolerated in CML patients, but no clinical and 
longitudinal data exist about the long term side effects of this drug since its approval in 2007. Therefore, 
clinical use of Abl inhibition may have dose-limiting toxicity but we will use much lower doses in the 
current studies (see study design). Importantly, Nilotinib is washed out of the brain within several hours 
(37,47), so lower dose and prolonged period of administration may decrease misfolded protein levels, 
leading to slowing of motor, non-motor and cognitive decline. Furthermore, to evaluate adverse drug 
effects, all patients will be monitored every month for specific warnings, including prolongation of QT 
interval. Prior to administration and periodically, patients will be monitored for hypokalemia or 
hypomagnesaemia. EKGs will also be obtained to monitor the QTc at baseline and periodically 
thereafter as detailed in Table 3. Nilotinib will not be administered to patients with hypokalemia, 
hypomagnesaemia, or long QT syndrome. Concomitant drugs known to prolong the QT interval and 
strong CYP3A4 inhibitors will also be avoided.  
Nilotinib should be taken on an empty stomach 2hrs after eating to ensure appropriate absorption and 
no food should be consumed one hour after Nilotinib. The following SAEs can occur with Nilotinib: 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
19 
 
1- Myelosuppression  
2- QT Prolongation  
1- Sudden Deaths  
2- Cardiac and Arterial Vascular Occlusive Events  
3- Pancreatitis and Elevated Serum Lipase   
4-  Hepatotoxicity  
5-  Electrolyte Abnormalities   
6- Hemorrhage  
7- Fluid Retention  
 
4- PATIENT POPULATION 
4.1- Number of Patients & Centers 
This is a single center study at Georgetown University Medical Center (GUMC) Translational 
Neurotherapeutics Program (TNP) with total duration of 4 years and open enrollment for 3 years. We 
currently have no competing trials for DLB and we anticipate efficient recruitment to this study. We will 
recruit DLB patients from our Movement Disorders and Memory Disorders clinics at MedStar 
Georgetown University Hospital (MGUH). We have a registry of over 400 potential participants 
diagnosed with mild to moderate DLB and their study partners. Study procedures will be conducted at 
GUMC Clinical Research Unit (CRU) of Georgetown-Howard Universities Center for Clinical and 
Translational Science (GHUCCTS). GHUCCTS encompasses MedStar Health Research Institute (10 
hospitals), including Georgetown University Hospital (GUH), Howard University, the Washington DC 
VA Medical Center (with its hospital and five clinics) and the Oak Ridge National Laboratory. CRU 
GHUCCTS is NIH-funded and combines the five institutions into one research powerhouse, creating 
one of the largest clinical trials networks in the country 
A total of 60 participants with DLB will be enrolled. We anticipate screening 100 individuals to enroll a 
total of 60 participants over 4 years. There are no competing clinical trials for DLB in the greater 
Washington-Baltimore area. We will advertise to community support groups and websites 
(Clinicaltrials.gov). Only patients that meet all inclusion/exclusion criteria and are willing to make all 
visits will be included in the study. Our program has had significant success with recruitment and 
retention in clinical trials because of the active clinical involvement at GUH and in the satellite centers, 
including McLean Medical Center, Montgomery Hospital and Washington Hospital Center. Most 
patients will be local to study center, but any patient who qualifies and can commit to all study visits will 
be allowed enrollment into the study regardless of residence. Overall we do not anticipate any issues 
in recruiting or maintaining 60 patients in this clinical trial for 6-month treatment. We will also advertise 
in our newsletters and websites. 
 
4.2- Inclusion Criteria  
1- Written informed consent 
2- Capable of providing informed consent and complying with study procedures. Subjects who 
are unable to provide consent may use a Legally Authorized Representative (LAR). 
3- Clinical diagnosis of DLB according to McKeith et al (32) with both dementia MoCA≥18 and 
Parkinsonian defined as bradykinesia in combination with rest tremor, rigidity or both 
UPDRS I-III is less than 50 and/or UPDRS-III between 15 -40 on-state. Dementia and 
Parkinsonism must be present with at least one other symptom such as fluctuation, visual 
hallucinations or REM sleep behavioral disorder (RBD) 
4- 2.5 ≥Hoehn and Yahr stage ≤3 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
20 
 
5- : MDS-UPDRS-III 15-40 on-state (or up to 70 on the off-state) 
6- Abnormal DaTScan  
7- Stable concomitant medical and/or psychiatric illnesses in the judgement of the PI 
8- Patients between the age of 25-90 years, medically stable 
9- Must NOT be stable on mono-amine oxidase (MAO)-B inhibitors (Selegeline or rasagiline) 
for at least 4 weeks before enrollment and during Nilotinib treatment. 
10- Must be medically stable on 800mg Levodopa daily for at least 4 weeks 
11- QTc interval 350-460 ms, inclusive 
12- Participants must be willing to undergo LP at baseline and 6 months after treatment 
13- 13- Participants must be willing to do an LP and PET scan. 
 
4.3- Exclusion Criteria  
1. Patients with hypokalemia, hypomagnesaemia, or long QT syndrome- QTc≥461 ms 
2. Concomitant drugs known to prolong the QTc interval and history of any cardiovascular 
disease, including myocardial infraction or cardiac failure, angina, arrhythmia 
3. History or presence of cardiac conditions including:  
a. Cardiovascular or cerebrovascular event (e.g. myocardial infarction, unstable 
angina, or stroke) 
b. Congestive heart failure 
c. First, second- or third-degree atrioventricular block, sick sinus syndrome, or other 
serious cardiac rhythm disturbances 
d. Any history of Torsade de Pointes 
4. Treatment with any of the following drugs at the time of screening or the preceding 30 days, 
and/or planned use over the course of the trial: 
a. Treatment with Class IA or III antiarrhythmic drugs (e.g. quinidine) 
b. Treatment with QT prolonging drugs (www.crediblemeds.org)- excluding 
Selective Serotonin Reuptake Inhibitors (SSRIs) (e.g. Citalopram, Paxil, Zoloft, 
Cymbalta, Sertraline, etc...) 
c. Strong CYP3A4 inhibitors (including grapefruit juice). The concomitant use of 
strong CYP3A4 inhibitors (e.g., ketoconazole, itraconazole, clarithromycin, 
atazanavir, indinavir, nefazodone, nelfinavir, ritonavir, saquinavir, telithromycin, 
voriconazole) must be avoided. Grapefruit products may also increase serum 
concentrations of Nilotinib. Should treatment with any of these agents be 
required, therapy with Nilotinib should be interrupted. 
d. Anticoagulants, including Coumadin (warfarin), heparin, enoxaparin, daltiparin, 
xarelto, etc. 
e. St. John’s Wort and the concomitant use of strong other CYP3A4 inducers (e.g., 
dexamethasone, phenytoin, carbamazepine, rifampin, rifabutin, rifapentine, 
phenobarbital) must be avoided since these agents may reduce the concentration 
of Nilotinib. 
5. Abnormal liver function defined as AST and/or ALT > 100% the upper limit of the normal 
6. Renal insufficiency as defined by a serum creatinine > 1.5 times the upper limit of normal 
7. History of HIV, clinically significant chronic hepatitis, or other active infection 
8. Females must not be lactating, pregnant or with possible pregnancy 
9.  Medical history of liver or pancreatic disease  
10. Clinical signs indicating syndromes other than DLB, including, PD, PD with Dementia 
(PDD), corticobasal degeneration, supranuclear gaze palsy, multiple system atrophy, 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
21 
 
chronic traumatic encephalopathy, signs of frontal dementia, history of stroke, head injury 
or encephalitis, cerebellar signs, early severe autonomic involvement, Babinski sign 
11. Current evidence or history in past two years of epilepsy, focal brain lesion, head injury with 
loss of consciousness or DSM-IV criteria for any major psychiatric disorder including 
psychosis, major depression, bipolar disorder, alcohol or substance abuse 
12. Evidence of any significant clinical disorder or laboratory finding that renders the participant 
unsuitable for receiving an investigational drug including clinically significant or unstable 
hematologic, hepatic, cardiovascular, pulmonary, gastrointestinal, endocrine, metabolic, 
renal or other systemic disease or laboratory abnormality 
13. Active neoplastic disease, history of cancer five years prior to screening, including breast 
cancer (history of skin melanoma or stable prostate cancer are not exclusionary) 
14. Contraindications to LP: prior lumbosacral spine surgery, severe degenerative joint disease 
or deformity of the spine, platelets < 100,000, use of Coumadin/warfarin, or history of a 
bleeding disorder 
15. Must not be on any immunosuppressant medications or IVIG 
16. Must not be enrolled as an active participant in another clinical study 
 
4.4- Randomization and Registration will be performed by an internet based randomization module. 
Randomization of the subjects to the 2 treatment groups will be performed in a stratified manner. The 
chance for randomization to the groups is 1:1 for placebo: 200 mg Nilotinib.  
 
4.5- Blinding. The investigators will be blinded to the dosage. Medications for any patient will be 
labeled by the CRU with a package medical identification number (Med. Id). A patient specific patient 
identification number (Pat. Id.) will be assigned to each patient. The investigator will have to note the 
Pat.Id on the designated medication package number after randomization. Drug dispensation will take 
place once a month during the monthly visit. 
 
4.6- Un-blinding may occur for emergency purposes in case an AE or SAE makes it necessary for the 
treating physician to unblind the study treatment– if possible prior contact will be made with the clinical 
trials co-coordinator (CTC) or project manager. If this is not feasible, the CTC will be contacted within 
24 hours after un-blinding. The CTC should not be made aware of what the treatment assignment was. 
If un-blinding occurs the subject is automatically withdrawn and the procedure for withdrawal will be 
followed.  
 
4.7- Withdrawal of subjects. In accordance with the Declaration of Helsinki, each subject is free to 
withdraw from the study at any time without giving reasons for her/his decision. The investigator may 
also withdraw the subject at any time in the interest of the subject’s safety, including severe study-
related toxicity as detailed in Table 1 or in the case of un-blinding as described above. The primary 
reason for withdrawal (e.g. subject wish, safety, withdrawal of consent, etc.) must be recorded in the 
subject’s medical record and on the withdrawal form in the electronic Case Report Form (eCRF). Should 
a subject decide to withdraw after administration of study drug, or should the investigator decide to 
withdraw the subject, all efforts will be made to complete and report the observations up to the time of 
withdrawal as thoroughly as possible.  
A study subject will be discontinued from participation in the study if: 
 Any clinical AE, laboratory abnormality, concurrent illness, or other medical condition or situation 
occurs such that continued participation in the study would not be in the best interest of the 
subject. 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
22 
 
 The participant meets any exclusion criteria (either newly developed or not previously 
recognized). 
 
Subjects are free to withdraw from participation in the study at any time upon request. 
 
4.8- Handling of Withdrawals 
A subject may choose to discontinue participation in the study at any time. Subjects who permanently 
discontinue study drug should complete early study drug termination procedures per protocol. The 
subject should then return any unused study drug and will be asked to return to the study site for a final 
safety visit.  
 
4.9- Termination of Study  
This study may be prematurely terminated if, in the opinion of the principal investigator(s) (PIs), there 
is sufficient reasonable cause.   
 
Circumstances that may warrant termination include, but are not limited to: 
 Determination of unexpected, significant, or unacceptable risk to subjects. 
 Enrollment is unsatisfactory. 
 Insufficient adherence to protocol requirements. 
 Data are not sufficiently complete and/or evaluable.  
 
4.10- Protocol Adherence 
The Principal Investigators (PIs) agree to adhere to the protocol detailed in this document and agrees 
that any changes to the protocol must be approved by the site Institutional Review Board (IRB). The 
PIs will be responsible for enrolling only those study subjects who have met protocol eligibility criteria. 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
23 
 
5- SAFETY MONITORING AND REPORTING 
The AE definitions and reporting procedures provided in this protocol comply with all applicable 
regulations and ICH guidelines. The PI(s) will carefully monitor each subject throughout the study for 
possible AEs. All AEs will be documented on CRFs designed specifically for this purpose. It is also 
important to report all AEs, especially those that result in permanent discontinuation of the 
investigational product being studied, whether serious or non-serious. 
 
5.1- All Non-serious Adverse Events 
An AE is any unfavorable and unintended sign (including a clinically significant abnormal laboratory 
finding, for example), symptom, or disease temporally associated with a study, use of a drug product 
or device whether or not considered related to the drug product or device.  
 
Adverse drug reactions (ADR) are all noxious and unintended responses to a medicinal product related 
to any dose. The phrase “responses to a medicinal product” means that a causal relationship between 
a medicinal product and an adverse event is at least a reasonable possibility, i.e., the relationship 
cannot be ruled out. Therefore, a subset of AEs can be classified as suspected ADRs, if there is a 
causal relationship to the medicinal product. 
 
Examples of AEs include: new conditions, worsening of pre-existing conditions, clinically significant 
abnormal physical examination signs (i.e. skin rash, peripheral edema, etc.), or clinically significant 
abnormal test results (i.e. lab values or vital signs), with the exception of outcome measure results, 
which are not being recorded as AEs in this trial (they are being collected, but analyzed separately). 
Stable chronic conditions (i.e., diabetes, arthritis) that are present prior to the start of the study and do 
not worsen during the trial are NOT considered AEs.  Chronic conditions that occur more frequently 
(for intermittent conditions) or with greater severity, would be considered as worsened and therefore 
would be recorded as AEs. 
 
AEs are generally detected in two ways: 
 
 
Clinical  symptoms reported by the subject or signs detected on examination. 
 
Ancillary Tests  abnormalities of vital signs, laboratory tests, and other diagnostic procedures 
(other than the outcome measures, the results of which are not being captured as AEs). 
 
If discernible at the time of completing the AE log, a specific disease or syndrome rather than individual 
associated signs and symptoms should be identified by the PI and recorded on the AE log.  However, 
if an observed or reported sign, symptom, or clinically significant laboratory anomaly is not considered 
by the Site Investigator to be a component of a specific disease or syndrome, then it should be recorded 
as a separate AE on the AE log.  Clinically significant laboratory abnormalities, such as those that 
require intervention, are those that are identified as such by the PI. 
 
Subjects will be monitored for AEs from the time they sign consent until completion of their participation 
in the study (defined as death, consent withdrawal, loss to follow up, and early study termination for 
other reasons or following completion of the entire study).   
 
5.2- All Serious Adverse Events 
An SAE is defined as an adverse event that meets any of the following criteria: 
1. Results in death. 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
24 
 
2. Is life threatening: that is, poses an immediate risk of death as the event occurred. 
a. This serious criterion applies if the study subject, in the view of the PI or Sponsor, is at 
immediate risk of death from the AE as it occurs.  It does not apply if an AE hypothetically 
might have caused death if it were more severe. 
3. Requires inpatient hospitalization or prolongation of existing hospitalization. 
a. Hospitalization for an elective procedure (including elective PEG tube/g-tube/feeding tube 
placement) or a routinely scheduled treatment is not an SAE by this criterion because an 
elective or scheduled “procedure” or a “treatment” is not an untoward medical occurrence. 
4. Results in persistent or significant disability or incapacity. 
a. This serious criterion applies if the “disability” caused by the reported AE results in a 
substantial disruption of the subject’s ability to carry out normal life functions. 
5. Results in congenital anomaly or birth defect in the offspring of the subject (whether the subject 
is male or female).   
6. Necessitates medical or surgical intervention to preclude permanent impairment of a body 
function or permanent damage to a body structure. 
7. Important medical events that may not result in death, are not life-threatening, or do not require 
hospitalization may also be considered SAEs when, based upon appropriate medical judgment, 
they may jeopardize the subject and may require medical or surgical intervention to prevent one 
of the outcomes listed in this definition. Examples of such medical events include blood 
dyscrasias or convulsions that do not result in inpatient hospitalization, or the development of 
drug dependency or drug abuse. 
 
An inpatient hospital admission in the absence of a precipitating, treatment-emergent, clinical AE may 
meet criteria for "seriousness" but is not an adverse experience, and will therefore, not be considered 
an SAE. An example of this would include a social admission (subject admitted for other reasons than 
medical, e.g., lives far from the hospital, has no place to sleep). 
 
The PI is responsible for classifying AEs as serious or non-serious. 
 
5.3- All Reports of Drug Exposure during Pregnancy. 
Females, who are lactating/breast feeding, pregnant or with possible pregnancy must not participate in 
this study. Nilotinib and other drugs and procedures used in this study may be unsafe for a fetus, an 
infant, sperm and eggs. Women of child bearing potential must agree to avoid pregnancy during 
participation in this study and for 3 months after the completion of this study. A man must agree not to 
conceive a child during participation in this study and for 3 months after the completion of this study. If 
a woman becomes pregnant during the study or if a man fathers a child during the study, they should 
immediately notify the principal investigator and study coordinator(s) and must be immediately 
withdrawn from the study.  
5.4- All reports of misuse and abuse of Nilotinib and other medication. 
Site staff will check the subject’s overall compliance with the study requirements. This will also include 
checks of protocol compliance, Nilotinib and concomitant medication use, in addition to the proper use 
of study drug in order to assess potential risks of drug abuse or misuse. The PI shall report within 24 
hours of discovery any drug misuse experience to the IRB, FDA, Novartis and study monitor.  
 
5.5- Assessment and Recording of Adverse Events 
The PIs will carefully monitor each subject throughout the study for possible AEs. All AEs will be 
documented on CRFs designed specifically for this purpose.  All AEs will be collected and reported in 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
25 
 
the electronic data capture (EDC) system. The PIs shall promptly review all information relevant to the 
safety of the investigational product, including all SAEs.  Special attention will be paid to those that 
result in permanent discontinuation of the investigational product being studied, whether serious or 
non-serious. 
 
5.6- Assessment of Adverse Events 
At each visit (including telephone interviews), the subject will be asked if they have had any problems 
or symptoms since their last visit in order to determine the occurrence of AEs.   If the subject reports 
an AE, the Investigator will probe further to determine: 
1. Type of event 
2. Date of onset and resolution (duration) 
3. Severity (mild, moderate, severe) 
4. Seriousness (does the event meet the above definition for an SAE) 
5. Causality, relation to investigational product and disease 
6. Action taken regarding investigational product 
7. Outcome 
 
5.7- Relatedness of Adverse Event to Investigational Product 
The relationship of the AE to the investigational product should be specified by the PIs, using the 
following definitions: 
 
1.  Not Related:          
 
Concomitant illness, accident or event with no reasonable 
association with treatment. 
 
2.  Unlikely: 
 
 
The reaction has little or no temporal sequence from 
administration of the investigational product, and/or a more likely alternative 
etiology exists. 
 
3.  Possibly Related: 
 
The reaction follows a reasonably temporal sequence from 
administration of the investigational product and follows a known response 
pattern to the suspected investigational product; the reaction could have 
been produced by the investigational product or could have been produced 
by the subject’s clinical state or by other modes of therapy administered to 
the subject. (Suspected ADR) 
 
4.  Probably Related: 
 
The reaction follows a reasonably temporal sequence from 
administration of investigational product; is confirmed by discontinuation of 
the investigational product or by re-challenge; and cannot be reasonably 
explained by the known characteristics of the subject’s clinical state. 
(Suspected ADR) 
 
5.  Definitely Related: 
 
The reaction follows a reasonable temporal sequence from 
administration of investigational product; that follows a known or expected 
response pattern to the investigational product; and that is confirmed by 
improvement on stopping or reducing the dosage of the investigational 
product, and reappearance of the reaction on repeated exposure. 
(Suspected ADR) 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
26 
 
5.8- Recording of Adverse Events 
All clinical AEs are recorded in the AE Log in the subject’s study binder. Study staff should fill out the 
AE Log and enter the AE information into the EDC system within 48 hours of the site learning of a new 
AE or receiving an update on an existing AE. Entries on GUMC RedCAP database and the AE Log 
(and into the EDC) will include the following: name and severity of the event, the date of onset, the date 
of resolution, relationship to investigational product, action taken, and primary outcome of event.  The 
PI shall report within 24 hours of discovery any serious adverse experience whether or not considered 
related to the study to the IRB, FDA and study monitor who will contact the DSMB. Serious adverse 
experiences should also be reported immediately by telephone and subsequently in writing within 48 
hours of the occurrence to the following: 
Robert Schiff, Ph.D., RAC, CQA, FRAPS 
Medical Monitor 
1120 Bloomfield Avenue 
West Caldwell, NJ 07006 
Tel:  973-227-1830 
Fax:  973-227-5330 
Cell: 973-568-3361 
Email: Rschiff13@aol.com 
The Institutional Review Board will be notified of such adverse experiences 
 
The minimum necessary information to be provided at the time of the initial report will include the 
following: 
 
Study identifier 
A description of the event  Whether study was discontinued 
Study Center 
Date of onset  
The reason why the event is 
classified as serious 
Subject number 
Current status  
Investigator assessment 
of 
the 
association between the event and 
study treatment 
 
Copies of each report and documentation of IRB notification and receipt will be kept in the Clinical 
Investigator’s study file/site binder. 
 
Adverse Event Reporting Period 
The study period during which adverse events must be reported is normally defined as the period from 
the initiation of any study procedures to the end of the study treatment follow-up 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
27 
 
6- STUDY MONITORING  
6.1- Safety Monitoring  
The study PIs will review safety data throughout the trial and may stop the trial for safety if they 
determine that there is a significant difference in the rate of a particular AE that would indicate a risk 
that is greater than the possible benefit of the study drug.  
 
Unanticipated problems involving risks to subjects or others including adverse events will be reported 
to the Partners Human Research Committee (PHRC) in accordance with PHRC unanticipated problems 
including AEs reporting guidelines.  
 
6.2- Data Safety Monitoring Board (DSMB) will be formed at the time of the initiation of the study and 
will include medical experts on Nilotinib effects on cardiac toxicity (TBD), neurologist (TBD), a clinical 
pharmacologist (TBD) and a biostatistician (TBD). No investigator involved in the trial will be a member 
of the DSMB. The DSMB will review the protocol to identify any necessary modifications. If 
modifications are necessary, revisions will be reviewed by the DSMB prior to its recommendation on 
initiation of the project.  The DSMB, based on its review of the protocol, will identify the data parameters 
and format of the information to be regularly reported. The DSMB will be informed of the occurrence of 
any SAEs and immediately notified of fatal or life-threatening events. The DSMB may at any time 
request additional information from the PIs. The DSMB will be provided with data blinded to treatment 
status, but they may request un-blinded data if there is a safety concern. Based on the review of safety 
data, the DSMB will make recommendations regarding the conduct of the study. These may include 
amending safety monitoring procedures, modifying the protocol or consent, terminating the study or 
continuing the study as designed. The discussions and decisions of the DSMB will be summarized in 
written reports and provided to the PIs. The DSMB will meet in person or by conference call every 6 
months or as necessary when requested by the PI or the monitor. 
The DSMB will also provide boundaries for when to terminate the study or end participation of an 
individual patient according to the guidelines of DMSB charters adopted by the NIH. The stopping rules 
should include stopping rules as in Table 1 and other criteria related to: 
Pancreatitis, QT prolongation (select a method for QT correction (typically Frederica’s method) and 
include in the protocol), develop individual stopping rules for withdrawal from the study for QT 
prolongation, and create algorithms for the follow-up of abnormal blood tests that potentially signal the 
start of a drug induced event (e.g. neutropenia or thrombocytopenia). The DSMB will monitor each 
patient in real-time and on a case-by-case basis and unblind any case when necessary.  
 
6.3- Study monitor and auditing will ascertain adherence to the study protocol by investigators and 
ensure proper documentation and investigator blinding of biomarker data processing and deposition. 
This study will be monitored by a clinical monitor. The monitors will maintain liaison with the 
investigators by telephone, letter, email and personal visits in order to assure the sponsors that the 
clinical study is completed according to the protocol requirements and that Good Clinical Practices are 
being followed according to 21 CFR parts 50, 56, 812, FDA Guidelines and the ICH E6 Guideline.  The 
PI will allocate adequate time for such monitoring activities. The PI will also ensure that the monitor or 
other compliance or quality assurance reviewer is given access to all the above noted study-related 
documents and study related facilities (device testing location), and has adequate space to conduct the 
monitoring visit.  
 
Auditing inspections will serve to verify strict adherence to the protocol and the accuracy of the data 
management, in accordance with the federal regulations. The PI will permit study-related monitoring 
and audits by the IRB, the sponsor, and government regulatory bodies, of all study related documents 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
28 
 
(e.g. source documents, regulatory documents, data collection instruments, study data etc) at regular 
intervals throughout the study. The PI will ensure the capability for inspections of applicable study-
related facilities (device testing location). Participation as a PI in this study implies acceptance of 
potential inspection by government regulatory authorities and applicable University compliance and 
quality assurance offices.  
The investigator should be aware that representatives of the FDA might also inspect the study site and 
subject records. If contacted for an audit of this study by any regulatory agency, the investigator should 
notify the sponsor immediately.  
Study monitor will ascertain adherence to the study protocol by investigators and ensure proper 
documentation and investigator blinding of biomarker data processing and deposition. 
 
Robert Schiff, Ph.D., RAC, CQA, FRAPS 
Medical Monitor 
1120 Bloomfield Avenue 
West Caldwell, NJ 07006 
Tel:  973-227-1830 
Fax:  973-227-5330 
Cell: 973-568-3361 
Email: Rschiff13@aol.com 
 
7) Institutional Review Board (IRB) 
This study will be conducted in compliance with current Good Clinical Practices (GCP) and Title 21Part 
56 of the United States of America Code of Federal Regulations (CFR) relating to IRBs.  
7.1- Ethical Conduct of Study 
The study will be conducted in accordance with GCP defined by the International Conference on 
Harmonization (ICH) and the ethical principles of the Declaration of Helsinki. 
7.2- Subject Information and Consent 
This study will be conducted in compliance with Title 21 Part 50 of the United States of America Code 
of Federal Regulations (CFR), Federal Regulations and ICH Guidance Documents pertaining to 
informed consent. At the first visit, prior to initiation of any study-related procedures, subjects will be 
informed about the nature and purpose of the study, participation/termination conditions, and risks and 
benefits. Subjects will be given adequate time to ask questions and become familiar with the study prior 
to providing consent to participate. Subjects will give their written consent to participate in the study 
and will be provided with a copy of the fully executed consent form for their records.  
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
29 
 
 
8- Statistical Methods and Data Analysis 
 
8.1- Sample size determination 
Safety: No formal sample size calculation is conducted. Sample size of 30 patients in each arm is 
based on feasibility. Safety of the treatment arm will be judged by the absence of events of major 
concern, including QTc prolongation and myelosuppression. According to Novartis Investigator 
Brochure, the frequency of AEs of all grades leading to discontinuation in the Nilotinib 300mg treatment 
was 14.0% and the incidence of SAEs in the Nilotinib 300 mg treatment recorded 9%. With the Nilotinib 
200 mg treatment, the treatment arm is expected to have less than 98.9% probability of observing at 
least one instance of any AE with an expected incidence of at least 14% in the treatment arm when the 
binomial distribution is assumed. For SAEs, this probability of at least one event will decrease to at 
most 94.1% with an expected incidence of at least 9% in the treatment arm. 
 
Tolerability: If we define a dose as tolerable when we observe no more than 25% intolerance or the 
discontinuation ratio is greater than 25%, the probability of early discontinuation of this trial (>25%), 
i.e., observing 8 or more AEs out of 30 is 5% (1-95.0%) given 30 participants in the Nilotinib 200mg 
arm. If we assume 9% of probability of SAEs occurrence, the probability of discontinuation due to SAEs 
of this trial, i.e., observing 8 or more severe AEs out of 30 is 0.4% (1-99.6%).    
 
Efficacy endpoints. Given sample size of 30 per arm, we calculate power to detect the efficacy of 
200mg Nilotinib treatment in terms of primary biomarkers and clinical endpoint between baseline and 
6 months. We will test the null hypothesis that the endpoint change in the control group over 6 months 
equals the endpoint change in the treatment group using a one-sided significance level 0.025.  
It is assumed that 10% complete losses and additional 20% partial loss of information due to 
symptomatic treatment leads to ``drop-out’’ rate of 30%. Considering 30% drop-out, a total of 42 
patients (21 patients in each arm) will be considered. Given observed statistics on changes of efficacy 
endpoints in the pilot study, we anticipate powers to detect changes in each endpoint as shown in Table 
4.  
 
Table 4- Estimated power based on two endpoints 
Endpoint 
   Placebo 
Treatment  Difference 
             SD 
   Power (N=30)     Power (N=21) 
HVA 
0% 
50% 
50% 
50 
97% 
90% 
UPDRS I-IV 
3 
-7 
10 
12 
90% 
77% 
We will have 90% power to test our null hypotheses for HVA biomarkers based on comparing means 
using a normal distribution with 21 patients in each arm. For testing clinical outcomes (MDS- UPDRS), 
a number of 21 or more patients will achieve at least 77% power to detect the change in UPDRS I-IV. 
Due to normality given a small sample size, a simulation based non-parametric Mann-Whitney-
Wilcoxon test (MWW) power is also computed given the mean change rate and SD. For HVA, MWW 
power is approximately 87% with 21 patients in each arm and MWW power for UPDRS is approximately 
73%. 
Amyloid PET power analysis: These imaging studies will significantly add to our understanding of the 
potential change of CNS amyloid via PET versus CSF in DLB. We anticipate to continue to enroll 
participants until 2023. We estimate that around 50 participants should be enrolled within the currently 
approved project period for the existing parent award and we therefore propose to add amyloid 
imaging to data collection.  This is a pilot or roof-of-concept study, but based on the data obtained in 
the AD study with n=12 patients per group (placebo versus nilotinib), we performed power analysis 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
30 
 
considering a similar effect on amyloid PET. Therefore, considering the changes of amyloid in the 
frontal lobe over 6 months as co-primary endpoints, the study overall type I error of 5% is split into 
2.5% for evaluating each endpoint. In a two-sided t-test for two-sample mean, at the significance 
level of 0.025 and 80% power, 25 subjects for 200mg Nilotinib group and 25 subjects for placebo 
group will have a minimal detectable effect size of 0.9 SD or 0.9 Cohen’s D between 200mg nilotinib 
and placebo. Considering 20% dropout rate, 
20 subjects for 200mg Nilotinib group 
and 20 subjects for placebo group will have a minimal detectable effect size of 1.03 Cohen’s D 
between 200mg nilotinib and placebo. The results of this proof-of-concept study will guide the 
development of a larger future clinical trial.  
 
8.2- Analyses plan:  
 
Safety: The frequency of AEs classified by MedDRA system organ class and preferred term and 
clinically significant changes in EKG and laboratory parameters will be summarized as simple 
proportions with Clopper-pearson exact 95% confidence bounds. The proportion of participants 
experiencing each type of event will be compared by Fisher’s exact test.  
Tolerance: The proportion tolerant of Nilotinib 200mg dose will be estimated with exact confidence 
intervals.  
 
Target Engagement: The measurements of all endpoints at baseline and the change from baseline to 
6 months will be summarized in terms of mean, median, range, and standard deviation. To compare 
efficacy of Nilotinib, 200mg, a shared-baseline repeated-measures ANOVA will be used with estimates 
of treatment-specific changes over 6 months and differences between treatments obtained from linear 
contrasts. This model will include fixed effects of visit (baseline and 6 months) and a dose by post-
baseline visit interaction and unstructured covariance for the repeated measurements. One additional 
baseline covariate may be included if a large difference in a potentially prognostic measure results by 
chance. A significant dose effect will be judged based on one-tailed testing at a p-value of 0.025. Other 
CSF and plasma exploratory biomarkers and clinical outcomes will be explored by the same model. 
The relationship between 6-month changes in tau, alpha-synuclein, dopamine metabolism and amyloid 
or other biomarkers and 6-month changes in clinical outcomes will be summarized as simple 
correlations with a visual verification that correlations within each dose roughly match correlations 
across doses, i.e., absence of Simpson’s paradox. 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
31 
 
9- STUDY FEASIBILITY AND PRELIMINARY DATA 
9.1- Preclinical Evidence 
We previously demonstrated 
that Nilotinib penetrates the brain, 
inhibits Abl activity and promotes 
autophagic degradation of amyloid 
proteins 
in 
transgenic 
mice 
(37,47,48,50,73). These findings 
were also reproduced by several 
other laboratories (37,60,61). To 
determine whether Nilotinib enters 
the brain, male and female 2 months 
old wild type (WT) C57BL/6 mice 
were intraperitoneally (I.P.) injected 
with an escalating dose of 1, 10, 20, 
40 and 80 mg/kg Nilotinib dissolved 
in 30µL DMSO (n=3 per group) and 
sacrificed at 2, 4, 6, 8 and 12hr post-
injection (n=3 per time point). The 
concentration 
of 
Nilotinib 
was 
measured by mass spectrometry 
using 13C labeled Nilotinib as a 
standard control in the plasma (Fig. 
1A) and total brain homogenates 
(Fig. 1B). At 10 mg/kg Nilotinib 
reached the highest brain/blood 
ratio (5.08%) at 305nM peak (Tmax) 
4hr post-injection; and 1 mg/Kg 
brain/blood was 3.06% at 205nM 
(Fig. 1C). Higher doses of Nilotinib 
(20, 40 and 80mg/kg) injection did 
not result in higher brain levels, 
perhaps due to the role of the 
ATP-binding 
cassette 
(ABC) 
transporters, 
which 
facilitate 
Nilotinib efflux at the BBB (73). 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
32 
 
Abl activation is associated with accumulation of alpha-Synuclein. To examine the 
relationship between Abl and alpha-Synuclein, male C57BL/6 mice were stereotaxically injected with 
1x104 multiplicity of infection (m.o.i) lentiviral clones driving the expression of Abl, or alpha-Synuclein 
(or LacZ) bilaterally 
into the substantia 
nigra (SN). Lentiviral 
injection significantly 
increased 
alpha-
Synuclein (42%) over 
LacZ 
level 
(Fig. 
2A&B, 1st blot, N=9 
animals) 
6 
weeks 
post-injection. Alpha-
Synuclein expression 
led to an increase in 
total 
Abl 
(110%) 
relative to actin and 
tyrosine 412 (T412) 
phosphorylation 
(289%) 
relative 
to 
total Abl (Fig. 2A&B, 
p<0.05, 
N=9) 
compared to LacZ 
expressing 
mice, 
indicating 
Abl 
activation. 
Conversely, lentiviral 
expression of Abl in 
the mouse SN led to 
an increase (204%) in 
total Abl relative to 
actin 
(Fig. 
2D&E, 
p<0.05, 
N=9) 
and 
T412 phosphorylation 
(231%) relative to total 
Abl and resulted in 
increased 
levels 
of 
monomeric (51%) and 
high molecular weight 
alpha-Synuclein 
(relative to actin) 6 
weeks 
post-injection 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
33 
 
compared to Lac-Z. To verify that both lentiviral LacZ and Abl were expressed, a V5 tag of the 
lentiviruses were probed with IHC. Staining of 20μm thick SN sections showed V5 (Fig. 2E) and Abl 
(Fig. 2F) co-localized (Fig. 2G) in lentiviral-Abl injected brains, while staining of Lac-Z injected SN 
showed V5 (Fig. 2H) and endogenous Abl (Fig. 2I) without co-localization (Fig. 2J), indicating lentiviral 
expression.  
Western Blot (WB) was performed on homogenized frozen striatal (caudate) brain tissues 
(described in (36)) from 9 sporadic PD patients and 7 age-matched control subjects. Human post-
mortem PD striatal extracts showed an increase in total Abl (220%) relative to actin and T412 (267%) 
relative to total Abl (Fig. 2K&L) compared to control subjects (p<0.02, two-tailed t-test), suggesting a 
relationship between Abl 
activation 
and 
alpha-
Synuclein in PD.  
Intraperitoneal 
(I.P.) injection of 10-20 
mg/kg Nilotinib into WT 
mice (N=5 animals/time 
point), led to detection of 
up to 30ng Nilotinib 
(310nM) per mg brain 
tissue 
3-4hr 
after 
injection (Fig. 2M) and 
Nilotinib 
was 
still 
detectable at 3.4ng/mg 
(35nM) 
7-8 
hr., 
indicating that Nilotinib 
enters the brain and is 
washed out after several 
hours. Two months old 
C57BL6 mice were I.P. 
injected once a day with 
10mg/kg 
Nilotinib 
or 
DMSO 
(30μL) for 
3 
consecutive weeks, and 
total brain extracts were 
analyzed with WB. 
Nilotinib led to a slight 
(11%) 
decrease 
in 
total Abl relative to 
tubulin but T412 Abl 
was 
significantly 
decreased (52%) relative to total Abl compared to DMSO (Fig. 2N&O, p<0.05, N=10 animals). No 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
34 
 
changes were detected in T245 Abl relative to actin or total Abl (Fig. 2N, 3rd blot). These data were 
confirmed by other groups (37,60,61).  
Nilotinib decreases brain and blood alpha-Synuclein levels. Nilotinib effects on alpha-
Synuclein 
levels 
were tested in 7-8 
months 
old 
transgenic 
alpha-
Synuclein mice that 
harbor 
the 
A53T 
mutation of alpha-
Synuclein (74). Total 
brain lysates showed 
accumulation 
(ELISA) of up to 
885ng/ml 
human 
alpha-Synuclein (Fig. 
3A) in 7-8 months 
mice 
treated 
with 
DMSO compared to 
wild 
type 
control 
(p<0.05, N=10), but 
daily IP injection of 
10mg/kg Nilotinib for 
3 weeks significantly 
decreased 
alpha-
Synuclein 
to 
467ng/ml 
(p<0.05, 
N=10), indicating that 
once a day treatment 
with 
Nilotinib 
decreases 
brain 
alpha-Synuclein 
levels. A significant 
increase in caspase-
3 activity (Fig. 3B, 
670%, 
p<0.05, 
N=64), 
indicating 
apoptotic death, was 
observed 
in 
A53T 
mice 
brains 
and 
Nilotinib reversed this 
increase to 201% compared to wild type age-matched controls with and without Nilotinib (N=64). The 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
35 
 
effects of lower dose and longer periods of treatment were evaluated in the blood and brain of 5-6 
months old A53T mice, which were injected every other day with 1mg/kg or 5mg/kg Nilotinib for 6 weeks 
(compared to 3 weeks). Whole blood was collected via cardiac puncture and brain tissues were 
extracted in lysis buffer and analyzed by ELISA. A total of 665ng/ml human alpha-Synuclein was 
observed in the brain of A53T treated with DMSO compared to C57BL/6 age-matched control (Fig. 3C, 
p<0.05, N=10), but alpha-Synuclein levels decreased to 503ng/ml with 5mg/kg and 344ng/ml with 
1mg/kg Nilotinib for 6 weeks. A higher concentration of human alpha-Synuclein (1635ng/ml) was 
detected in the whole blood of A53T mice treated with DMSO compared to C57BL/6 age-matched 
control (Fig. 3D, p<0.05, N=10), and, again, alpha-Synuclein levels decreased to 1439ng/ml with 
5mg/kg and 888ng/ml with 1mg/kg Nilotinib for 6 weeks, suggesting that the decrease in brain levels 
may also decrease blood alpha-Synuclein. Daily IP injection of 10mg/kg Nilotinib for 3 weeks into 7-8 
months A53T mice again decreased monomeric (41%) and high molecular weight human alpha-
Synuclein (37) relative to actin compared to A53T mice treated with DMSO.  
To ascertain that autophagy is involved in Nilotinib-mediated alpha-Synuclein clearance in A53T 
mice in vivo, autophagic vacuoles (AVs) were isolated via subcellular fractionation using a 
discontinuous Metrizamide gradient (75) and the levels of alpha-Synuclein and p-Tau were measured 
via ELISA. Subcellular fractionation was performed in an age-dependent manner to determine whether 
higher levels of protein accumulation alter autophagic flux through the AV10 or AV20 Metrizamide 
gradients, which contain LC3B (Fig. 3E, inset), indicating phagophore/autophagosome presence and 
the lysosomal fraction containing lysosomal associated membrane protein (LAMP)-2a. Human alpha-
Synuclein was detected in AV10 (340ng/ml) and AV20 (401ng/ml) in 1 month old A53T brains (Fig. 3E, 
p<0.05, N=5) but 10mg/kg Nilotinib significantly decreased alpha-Synuclein levels (56ng/ml) in AV10 
(Fig. 3E, p<0.05, N=5). However, Nilotinib significantly increased alpha-Synuclein levels in the 
lysosomes (268ng/ml) compared to DMSO (59ng/ml). Alpha-Synuclein was even higher in AV10 
(940ng/ml) and AV20 (410ng/ml) in 5 months old A53T mice treated with DMSO (Fig. 3G, p<0.05, N=5) 
but alpha-Synuclein levels were decreased by Nilotinib in AV10 (245ng/ml) and increased in AV20 
(642ng/ml) compared to DMSO. Nilotinib also significantly increased alpha-Synuclein levels in the 
lysosomes (333ng/ml) compared to DMSO (109ng/ml) in the same age group. P-Tau was also used as 
another protein marker that can potentially be degraded by autophagy. P-Tau was detected in AV10 
(710ng/ml), AV20 (590ng/ml) and lysosomes (129ng/ml) in 1 month old A53T mouse brain (Fig. 3F, 
p<0.05, N=5), indicating Tau hyper-phosphorylation. However, 10mg/kg Nilotinib increased p-Tau in 
AV10 (1001ng/ml), AV20 (890ng/ml) and lysosomes (321ng/ml) compared to DMSO within the same 
age group (Fig. 3F, p<0.05, N=5). p-Tau was higher in AV10 (1540ng/ml) and AV20 (2055ng/ml) in 5 
months old A53T mice treated with DMSO (Fig. 3F, p<0.05, N=5) but Nilotinib decreased p-Tau in AV10 
(610ng/ml) and increased it in AV20 (1133ng/ml) compared to DMSO. Nilotinib also decreased p-Tau 
in the lysosomes (345ng/ml) compared to DMSO (530ng/ml). 
 These data suggest that Nilotinib inhibition of Abl triggers autophagic protein clearance 
in vivo. 
Nilotinib protects SN tyrosine hydroxylase (TH) neurons from alpha-Synuclein toxicity. 
Staining of 20µm thick brain sections showed human alpha-Synuclein expression in mice injected with 
lentiviral alpha-Synuclein into the SN and treated with DMSO (Fig. 4B) compared to LacZ treated with 
Nilotinib (or DMSO data not shown) mice (Fig. 4A, N=12); but Nilotinib led to 84% (by stereology) 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
36 
 
decrease of human alpha-Synuclein (Fig. 4C, p<0.05, N=12) in SN neurons. A significant decrease in 
TH+ neurons (89% by stereology) was observed in lentiviral alpha-Synuclein treated with DMSO (Fig. 
4E&H) compared to LacZ treated with Nilotinib (Fig. 4D&G), and Nilotinib treatment of alpha-Synuclein 
expressing mice reversed TH+ neuron loss back to 82% (Fig. 4F&I, by stereology) of LacZ level (p<0.05, 
N=12). To evaluate alpha-Synuclein and Nilotinib effects on dopamine (DA) metabolism, DA and its 
metabolite Homovanilic acid (HVA) in SN brain extracts were measured using ELISA. A significant 
decrease (p<0.05, N=8) in DA (62%) and HVA (36%) were observed in SN extracts of lentiviral alpha-
Synuclein treated with DMSO compared to LacZ with and without Nilotinib (Fig. 4J). However, Nilotinib 
significantly (P<0.05, N=8) reversed DA and HVA loss back to control (Fig. 4J, N=8). Alpha-Synuclein 
expression in SN decreased rotarod performance 39% (Fig. 4K, p<0.05, N=14) of LacZ with and without 
Nilotinib, but Nilotinib reversed motor performance to 86% of LacZ (Fig. 4K, p<0.05, N=14).  
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
37 
 
9.2- Human Studies 
To examine the effects of lower doses of Nilotinib on safety and efficacy in PDD and PD with MCI and 
DLB, twelve late stage (Hoehn and Yahr 3-5) PDD and DLB participants were randomized into 150mg 
(N=5) or 300mg (N=7) groups and received oral daily doses of Nilotinib for 6 months with 3-month 
follow up (Chart 3 and Table 6). The primary objective was to determine the safety and tolerability of 
Nilotinib for 6 months throughout a total of 11 visits. Electrocardiograms (EKG), physical and 
neurological exams and blood draws for laboratory chemistry were performed every 2 weeks for the 
first 2 months, and every month thereafter. Participants were on stable medical conditions on L-Dopa 
treatment and other PD medications for 4 weeks prior to study enrollment as indicated in Table 6. 
Patients with cardiovascular complications, infarcts, QTc interval >450ms and those who are on drugs 
that prolong QTc or with potential drug contraindications were excluded as per package insert. Thirty 
patients were screened and 12 enrolled. The secondary objectives determined whether Nilotinib 
crosses the BBB. Nilotinib concentration was measured in plasma and CSF at baseline, 2 months 
(interim) and 6 months. Pharmacodynamics analysis was performed to determine target engagement 
via measurement of CSF phosphorylated Abl (activity) in addition to neurodegeneration biomarkers, 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
38 
 
including alpha-Synuclein, dopamine metabolite HVA, total Tau and p-Tau181/231, Aβ42 and Aβ40. 
Cell death markers, including CSF NSE and glial and neuronal marker S100B were measured. 
Tertiary/exploratory outcomes included measurement of motor and non-motor symptoms using UPDRS 
I-IV. Montreal Cognitive Assessment (MoCA) in the range of 18-26 was used at screening to determine 
mild cognitive impairment (MCI), moderate cognitive impairment (17-10) and severe cognitive 
impairment (<10) in PD and LBD patients. Nilotinib effects on progression of dementia were evaluated 
using the MMSE and SCOPA-Cog.  
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
39 
 
9.2.a- Pharmacokinetic studies  
Blood and CSF were collected from twelve patients at baseline prior to treatment (N=12, control) 
and 2 months into treatment (N=12). Blood and CSF were collected from nine patients at six months 
(N=9) of Nilotinib treatment (total N=33 samples) and collection time via lumbar puncture (LP) was 
staggered at 0, 1, 2, 3, 4 or 5hrs after oral administration of Nilotinib. Blood was collected half an hour 
prior to LP. Two CSF samples were eliminated because they had >25ng/ml hemoglobin. 
Pharmacokinetic analysis demonstrates that Nilotinib (Fig. 5A&E) peaks in the plasma 2hrs (Tmax) after 
dosing at 55.27nM and 86.75nM (Cmax) in the 150mg and 300mg groups, respectively. Nilotinib (Fig. 
5B&E) also peaks in the CSF 2hrs (Tmax) after dosing at 5.09nM and 11.1nM (Cmax) in the 150mg and 
300mg groups, respectively. Area under the curve (AUC) is 157.27 and 232.8 ng/ml*h for plasma and 
7.98 and 14.13 ng/ml*h for CSF in 150mg and 300mg groups, respectively. Nilotinib was quantified in 
reference to 13C Nilotinib standard and samples were precipitated and dialyzed to obtain unbound 
Nilotinib. Nilotinib was not detected in the CSF (t1/2 ~1.8) 3hrs after dosing suggesting short 
bioavailability of unbound or free Nilotinib compared to plasma t1/2=12.4 (Fig. 5E). The CSF:plasma 
ratio of Nilotinib (Fig. 5C) is 12% and 5% with 300mg and 150mg Nilotinib, respectively.  
9.2.b- Pharmacodynamics. To demonstrate whether Nilotinib inhibits Abl via de-
phosphorylation, we measured pan-tyrosine CSF Abl relative to total Abl via ELISA.  
Phosphorylated Abl was reduced with 150mg and 300mg Nilotinib (Fig. 5B), suggesting that Nilotinib 
(~12% -22%) decreases CSF Abl (>30%) phosphorylation (inhibition). These data support the pre-
clinical results that Nilotinib penetrates the brain. The differences in Tmax and Cmax between human and 
mice can be attributed to several factors, including route of administration (i.e. I.P-mouse versus oral-

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
40 
 
human), methods of dissolving the drug (DMSO in mice), integrity of BBB in advanced PDD and DLB, 
differences in drug metabolism between mice and human, concomitant medication and diet. 
Additionally, Nilotinib was quantified in whole brain extracts in mice versus human CSF, where drug-
protein binding is likely to be different. Collectively these data support further investigation to determine 
Nilotinib concentration in CSF in larger trials with early PD.  
 
9.3- Safety 
 
9.3.1- Serious Adverse Events (SAEs). Adverse Events (AEs) that required or resulted in 
hospitalization were classified as SAEs and were listed in Table 7. One patient in the 300 mg Nilotinib 
group (NIL-9) was withdrawn at week 4 due to moderate cardiac ischemia and myocardial infarct and 
was double-stented for left bundle block. One participant in the 300mg Nilotinib group was hospitalized 
with urinary tract infection (UTI) and one participant in the 150mg group was hospitalized with 
pneumonia and UTI (Table 7).  
 
9.3.2- Non-serious AEs. Non-serious AEs, which did not require hospitalization were classified 
per system and/or preferred term and summarized in Table 8 as number of events and number of 
affected participants in each group. One participant in 150mg and one in 300mg group had slight QTc 
prolongation>450ms (detailed in Table 9). Participants in the 150mg group reported three UTIs, two 
cases of pneumonia, one cold virus, one mild back pain, one mild headache, one mild dysgraphia, one 
mild left foot drag, one mild confusion, one mild hallucination, one mild paranoia, one mild agitation, 
one moderate anxiety, one mild incontinence, one moderate itching and one skin irritation. Participants 
in the 300mg group reported one incident of blurry vision, one diarrhea, one nausea, one mild fatigue, 
and two generalized weakness, three UTIs, one pneumonia, one weight loss, one tooth extraction, one 
dizziness, two mild hallucination, one mild paranoia, one mild crying episode, one mild urinary urgency, 
one mild cough and one eczematous lesion.  

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
41 
 
 
 
9.3.3- QTc prolongation. One participant (NIL-9) in the 300 mg was withdrawn due to moderate 
cardiac ischemia and myocardial infarct and was double-stented for left bundle block (Table 7&9). NIL-
8 had slight QTc prolongation>450ms at week 8, and NIL-15 had two episodes of slight QTc 
prolongation>450ms at week 2 and week 8 but QTc returned to normal range in subsequent visits. 
NIL8 withdrew from the study at week 24 due to caregiver burden, but returned for safety follow up visit 
(week 36) and another patient (NIL-12) did not return at week 36.  
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
42 
 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
43 
 
 
9.3.4- Blood chemistry. Laboratory tests (Table 10) were performed throughout eleven visits 
according to study protocol. Laboratory results of some participants’ were outside the normal range at 
baseline due to comorbidities that were not exclusionary. A total number of three AEs below normal 
range of ALT level (<100%) were observed in the 150mg group Nilotinib. Two AEs above normal range 
(>100%), including one ALT and one bilirubin were observed in the 150mg group Nilotinib. Two AEs 
with an increase of ALT and one AST above normal range (>100%) were observed in the 300mg group 
Nilotinib. 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
44 
 
9.4- Disease related biomarker studies. CSF alpha-Synuclein concentration was significantly 
reduced at six months compared to baseline in the 150mg group (Table 11, N=4), but alpha-Synuclein 
did not change in the 300mg group (N=5). CSF HVA also significantly increased in the 300mg group at 
six-month compared to baseline (Table 11, N=5), but no significant differences were observed in the 
150mg group between baseline and six months treatment. CSF total Tau was significantly reduced at 
six months compared to baseline (Table 11, N=4) in 150mg group and no changes in total Tau were 
observed in 300mg groups, and these results were confirmed by Invitrogen and Millipore ELISA. CSF 
pTau181 concentration was increased at six months compared to baseline with 300mg Nilotinib (Table 
11, N=5) and the ratio of pTau181:total Tau was increased at six months in 300mg groups (p=0.0225). 
However, CSF pTau231 was reduced at six months compared to baseline with 150mg (Table, N=4) 
and 300 mg (Table 11, N=5) Nilotinib and the ratio of pTau23:total Tau did not change.  
CSF Aβ40 concentration was reduced at six months compared to baseline with 150mg (Table 
11, N=4), but Aβ40 was stable with 300mg Nilotinib. There was no significant change in CSF Aβ42 
(Table 11) and the ratio of CSF Aβ42:Aβ40 was not different. CSF concentration of NSE was reduced 
in 150mg group (Table 11, N=4) and the glial and neuronal cell death marker, S100B was reduced in 
300mg group (Table 11 N=5) at six months compared to baseline.  

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
45 
 
Plasma concentration of alpha-Synuclein was not significantly different at six months compared 
to baseline in the 150mg and 300mg group (Table 12). Plasma HVA level trended towards an increase 
after Nilotinib treatment in 300mg and 150mg groups at six months compared to baseline. The plasma 
level of Aβ40 did not change after Nilotinib treatment in both 150mg and 300mg groups.  
We performed further CSF analysis of oligomeric α-synuclein in the same subjects and 
normalized oligomeric to monomeric α-synuclein. The level of oligomeric α-synuclein was increased in 
the 150mg group (Fig. 6A) but stable in the 300mg group (Fig. 6B), consistent with the changes in 
monomeric α-synuclein. However the ratio of oligomeric/monomeric α-synuclein was significantly 
higher in the 150mg group at 6 months compared to baseline (Fig. 6C, p<0.05) but did not change in 
the 300mg group (Fig. 6D). Additionally, the change in CSF oligomeric/monomeric α-synuclein (Fig. 
6E) between baseline and 6 months was 40% in the 150mg group but this increase was attenuated 
(8%) in the 300mg group, suggesting that Nilotinib reduces CSF oligomeric α-synuclein levels. These 
results are consistent with our preclinical data (37,50,54) and previous evidence that the ratio of 
oligomeric/monomeric α-synuclein longitudinally increases in the CSF of PD patients (76,77). These 
data further suggest that Nilotinib reduces α-synuclein levels in PD but a placebo group is needed to 
better understand the effects of Nilotinib on α-synuclein species. 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
46 
 
 
9.5- 
Nilotinib 
ameliorates 
motor 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
47 
 
impairment. Participants’ motor symptoms were monitored with UPDRS throughout all visits (Table 
13). An average decrease of 3.4 points and 3.6 points in UPDRS-III (motor) was observed at six months 
compared to baseline with 150mg and 300mg Nilotinib, respectively. An average decrease of 7 points 
and 11.1 points in UPRDS I-IV was observed at six months in 150mg and 300mg groups, respectively. 
An average decrease of 7 points and 11.1 points in UPRDS I-IV were observed at 6 months in 150mg 
and 300mg groups, respectively. In the 300mg Nilotinib group, UPDRSI-IV significantly improved 
(p=0.0098) at six months compared to baseline (Table 14). The improvement in UPDRS was 
progressive throughout the six months (24 weeks) trial, and this improvement was reversed in 
participants (who returned) in the follow up visit off drug at thirty six weeks.  
 
9.6- Nilotinib improves cognitive symptoms. Participants’ progression of cognitive decline 
was monitored with MMSE throughout all visits (Table 15). An average increase of 3.85 (out of 30) and 
3.5 points in MMSE was observed at six months compared to baseline with 150mg and 300mg Nilotinib, 
respectively. In the 300mg group MMSE significantly increased (p=0.013) at six months compared to 
baseline (Table 14). The increase in MMSE scores was incremental throughout the twenty four-week 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
48 
 
treatment, and it declined in the thirty weeks-week follow up visit. Improvement in cognitive symptoms 
was also observed using Scales for Outcomes in Parkinson’s Disease-Cognition (SCOPA-Cog) with 
an average increase of 1.85 (out of 43) and 2 points at six months compared to baseline with 150mg 
and 300mg Nilotinib, respectively.  
 
 
10- TREATMENT ADMINSTERED 
10.1- Name of the drug and all active ingredients 
One hard white to yellowish powder in red opaque hard gelatin (Tasigna) capsule contains 200 mg 
Nilotinib (as hydrochloride monohydrate) and 117.08 mg lactose (as monohydrate). The Investigator 
Brochure (IB) indicating the pharmacological, safety and kinetics of the drug is attached.  
In brief, Nilotinib has been clinically prescribed since FDA approval in 2007, and substantial 
pharmacokinetics data are published (51-53,78). TKIs such as Imatinib (Glevec) are effective in many 
patients with CML in chronic phase (79,80). Nilotinib (AMN107) is a second generation selective Bcr-
Abl inhibitor, which is effective following Imatinib resistance and intolerance (79). Nilotinib was 
approved by the FDA in 2007 for CML treatment (up to 1200 mg orally daily) (51-53).  
 
10.2- Pharmacological Class of the Drug 
Nilotinib belongs to the pharmacotherapeutic group: Antineoplastic agents, protein kinase inhibitors, 
ATC code: L01XE08 
Nilotinib is a potent inhibitor of the Abl tyrosine kinase activity of the BCR-ABL oncoprotein both in cell 
lines and in primary Philadelphia-chromosome positive leukemia cells. The substance binds with high 
affinity to the ATP-binding site in such a manner that it is a potent inhibitor of wild-type BCR-ABL and 
maintains activity against 32/33 imatinib-resistant mutant forms of BCR-ABL. As a consequence of this 
biochemical activity, Nilotinib selectively inhibits the proliferation and induces apoptosis in cell lines and 
in primary Philadelphia-chromosome positive leukemia cells from CML patients. In murine models of 
CML, as a single agent Nilotinib reduces tumor burden and prolongs survival following oral 
administration. 
 
Nilotinib has little or no effect against the majority of other protein kinases examined, including Src, 
except for the platelet derived growth factor (PDGF), Kinase tyrosine kinase (KIT) and Ephrin receptor 
kinases, which it inhibits at concentrations within the range achieved following oral administration at 
therapeutic doses recommended for the treatment of CML.  
 
10.3- Structural Formula of the Drug 
Nilotinib has the molecular formula C28H22F3N7O and a molecular weight 
of 529.52g/mol. The structural formula of Nilotinib is as follows: 
 
10.4- Formulation of the dosage form to be used 
One hard white to yellowish powder in red opaque hard gelatin (Tasigna) capsule contains 200 mg 
Nilotinib (as hydrochloride monohydrate) and 117.08 mg lactose (as monohydrate). No excipients have 
any known effects and the full list of excipients in hard capsule includes:  
Lactose monohydrate 
Crospovidone 
Poloxamer 188 
Silica, colloidal anhydrous 
Magnesium stearate 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
49 
 
Hard capsule shell 
Gelatin 
Titanium dioxide  
Red iron oxide  
Yellow iron oxide  
Printing ink 
Shellac 
Black iron oxide.  
Placebo: Matching placebo that contains all ingredients except Nilotinib will be used. The drug will be 
procured in 200mg capsules and matching placebo will be manufactured by Sharp Clinical Sevices. 
 
10.5- Summary of Previous Human Experience   
Several oncology clinical trials data were reported for Tasigna. Exposure to Tasigna in a total of 717 
patients from a randomized Phase III study in patients with newly diagnosed Ph+ CML in chronic phase 
treated at the recommended dose of 300 mg twice daily (n=279) and from an open-label multicenter 
Phase II study in patients with imatinib-resistant or intolerant CML in chronic phase (n=321) and 
accelerated phase (n=137) treated at the recommended dose of 400 mg twice daily. No data were 
reported on CNS clinical trials. The following safety data were reported: 
a- In patients with newly diagnosed CML in chronic phase 
The median duration of exposure was 48.0 months (range 0.1-58.7 months). The most frequent 
(≥10%) non-hematological adverse reactions were rash, pruritus, headache, nausea, fatigue, alopecia 
and myalgia. Most of these adverse reactions were mild to moderate in severity. Upper abdominal pain, 
constipation, diarrhea, asthenia, dry skin, muscle spasms, arthralgia, vomiting, abdominal pain and 
peripheral edema were observed less commonly (<10% and ≥5%) were of mild to moderate severity, 
manageable and generally did not require dose reduction. 
Discontinuation due to adverse drug reactions was observed in 9% of patients. 
Treatment-emergent hematological toxicities include myelosuppression: thrombocytopenia (18%), 
neutropenia (15%) and anemia (7%). Pleural and pericardial effusions, regardless of causality, 
occurred in 1% and <1% of patients, respectively, receiving Tasigna 300mg twice daily. Gastrointestinal 
hemorrhage, regardless of causality, was reported in 3% of these patients.  
The change from baseline in mean time-averaged QTcF interval at steady state was 6 msec. No 
patient had an absolute QTcF >500 msec while on the study medicinal product. QTcF increase from 
baseline exceeding 60 msec was observed in <1% of patients while on the study medicinal product. 
No sudden deaths or episodes of torsade de pointes (transient or sustained) were observed. No 
decrease from baseline in mean left ventricular ejection fraction (LVEF) was observed at any time 
during treatment. No patient had a LVEF of <45% during treatment nor an absolute reduction in LVEF 
of more than 15%. 
 
b- In patients with imatinib-resistant or intolerant CML in chronic phase and accelerated 
phase 
The data described below reflect exposure to Tasigna in 458 patients in an open-label 
multicenter Phase II study in patients with imatinib-resistant or intolerant CML in chronic phase (n=321) 
and accelerated phase (n=137) treated at the recommended dose of 400 mg twice daily. The most 
frequent (≥10%) non hematological drug-related adverse events were rash, pruritus, nausea, fatigue, 
headache, vomiting, myalgia, constipation and diarrhea. Most of these adverse events were mild to 
moderate in severity. Alopecia, muscle spasms, decreased appetite, arthralgia, abdominal pain, bone 
pain, peripheral edema, asthenia, upper abdominal pain, dry skin, erythema and pain in extremity were 
observed less commonly (<10% and ≥5%) and have been of mild to moderate severity (Grade 1 or 2). 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
50 
 
Discontinuation due to adverse drug reactions was observed in 16% of chronic phase and 10% of 
accelerated phase patients. 
Treatment-emergent hematological toxicities include myelosuppression: thrombocytopenia (31%), 
neutropenia (17%) and anemia (14%). Pleural and pericardial effusions as well as complications of fluid 
retention occurred in <1% of patients receiving Tasigna. Cardiac failure was observed in <1% of 
patients. Gastrointestinal and CNS hemorrhage were reported in 1% and <1% of patients, respectively. 
QTcF exceeding 500 msec was observed in <1% of patients. No episodes of torsade de pointes 
(transient or sustained) were observed. 
 
Most frequently reported adverse reactions in Tasigna clinical studies for cancer: 
 
Non-hematological adverse reactions (excluding laboratory abnormalities) are reported in at least 5% 
of the patients in Tasigna clinical studies. These are ranked under heading of frequency using one 
decimal precision for percentages and the following convention: very common (≥1/10) or common 
(≥1/100 to <1/10). Within each frequency grouping, adverse reactions are presented in order of 
decreasing seriousness. 
 
10.6- Status of Drug in Other Countries 
Tasigna is available in the United States, Europe and other countries of the world and requires 
a prescription. Nilotinib is subject to restricted medical use and the marketing authorization holder shall 
submit periodic safety update reports for this product in accordance with the requirements set out in 
the list of Union reference dates (EURD list) provided for under Article 107c(7) of Directive 2001/83/EC 
and published on the European medicines web-portal determine compliance.  
 
10.7- Drug Related Risks and Potential Side Effects 
Nilotinib has a number of side effects  as stated in the IB, including gastrointestinal complications, 
vomiting and nausea, and in some rare cases dizziness (51). Abl has an essential role in mammalian 
heart growth and development and adult complications with Abl inhibition, include edema, 
nausea/vomiting, muscle cramps, neutropenia, thrombocytopenia, fever, liver toxicity, arthralgia, and 
exanthema/rash (68). Side effects are identified for Abl in homozygous Abl mutant mice, which display 
dramatically enlarged hearts due to abnormally increased cardiomyocyte proliferation during later 
stages of embryogenesis (69). Disruption of Abl in mice results in neonatal lethality accompanied by 
pleiotropic developmental defects with variable penetrance, including ranting, splenic and thymus 
atrophy, B cell lymphopenia, dysfunctional osteoblasts, and foreshortened crania (70-72). However, 
clinical use of Nilotinib (600-800mg daily) is fairly tolerated in CML patients, but no clinical and 
longitudinal data exist about the long term side effects of this drug since its approval in 2007. Therefore, 
clinical use of Abl inhibition may have dose-limiting toxicity but we will use much lower doses in the 
current studies (see study design). Importantly, Nilotinib is washed out of the brain within several hours 
(37,47), so lower dose and prolonged period of administration may decrease misfolded protein levels, 
leading to slowing of motor, non-motor and cognitive decline. Furthermore, to evaluate adverse drug 
effects, all patients will be regularly (every two week for the first two months) monitored for specific 
warnings, including prolongation of QT interval. Prior to administration and periodically, patients will be 
monitored for hypokalemia or hypomagnesaemia. EKGs will also be obtained to monitor the QTc at 
baseline, 7 days after initiation, and periodically thereafter as detailed in Table 2. Nilotinib will not be 
administered to patients with hypokalemia, hypomagnesaemia, or long QT syndrome. Concomitant 
drugs known to prolong the QT interval and strong CYP3A4 inhibitors will also be avoided.  
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
51 
 
Nilotinib should be taken on an empty stomach.  It should be taken one hour before eating, or 2hrs 
after eating to ensure appropriate absorption.   
 
11-PRIOR AND CONCOMITANT THERAPY 
Throughout the study, the subject may be prescribed concomitant medications or treatments deemed 
necessary to provide adequate supportive care, provided that the medications are licensed in the United 
States. All concomitant medications and/or treatments received by a subject should be recorded on the 
appropriate source document and eCRF.   
Prohibited and contra-indicated medications in those receiving Nilotinib are listed in the 
inclusion/exclusion criteria and detailed below. 
 
11.1- Prohibited Medications and Contraindications 
Prohibited Medications 
Prohibited medications for study subjects are as follows: 
- 
Anti-coagulant medications, including Coumadin, heparin, enoxaparin, fondiparinaux, etc 
- 
CYP3A4 enzyme inhibitors, including grapefruit juice 
- 
 St John’s Wort 
 
Pregnancy & Nursing Mothers 
There are no adequate and well-controlled studies in pregnant women. Subjects or partners of male 
subjects must not become pregnant during the study or 3 months after stopping study drug. If a female 
subject becomes pregnant, study treatment must be discontinued immediately. Caution should be 
exercised; therefore, no subject should nurse their infant while participating in this study. 
 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
52 
 
 
12-RATIONALE 
TO 
USE 
NILOTINIB 
TO 
TREAT 
NEURODEGENERATIVE 
DISEASES. 
Neurodegenerative diseases, including PD, are a group of genetic and sporadic disorders associated 
with neuronal death and progressive nervous system dysfunction. Cancer is also a collection of related 
genetic diseases, in which cells begin to divide without stopping and spread into surrounding tissues. 
Unlike neurodegeneration, in which no regeneration happens when damaged or aging post-mitotic 
neurons die, damaged cells survive when they should die in cancer, resulting in uncontrolled mitotic 
cell division to form tumors. Cancerous tumors are malignant as they spread or invade nearby tissues 
by cellular contiguity or metastasize via blood and/or humoral transport. In neurodegeneration, the 
spread of disease by contiguity is supported by the hypotheses that toxic or “prion-like” proteins 
propagate along neuroanatomical pathways (81), leading to progressive spread of disease and cell 
death. In neurodegeneration, failure of cellular quality control mechanisms leads to inadequate protein 
degradation via the proteasome or autophagy (82), resulting in intracellular accumulation of neurotoxic 
proteins. Consequently, these proteins are secreted from a pre-synaptic neuron and can traverse the 
synaptic cleft and enter a contiguous post-synaptic neuron. Secreted proteins may not penetrate an 
adjacent cell via the synapse but they may be re-routed into the cell and recycled via the endosomal 
system to fuse with autophagic vacuoles like the autophagosome or the lysosome (83-85). Microglia, 
the brain resident immune cells may also phagocytose and destroy toxic proteins (86). 
  
Accumulation of neurotoxic proteins, including alpha-Synuclein (Lewy bodies), beta-amyloid 
plaques, Tau tangles, Huntington, prions and TDP-43 are major culprits in neurodegeneration. These 
toxic proteins trigger progressive apoptotic cell death leading to loss of many central nervous system 
(CNS) functions, including mentation, cognition, language, movement, gastrointestinal motility, sleep 
and many others. The discoveries of toxic protein propagation from cell to cell (81), leading to 
progression of neurodegeneration triggered a series of pre-clinical and clinical studies to limit protein 
propagation via antibodies (active and passive immune therapies) that can capture the protein and 
destroy it en route to healthy neurons. This approach is fraught with difficulties, including failure to arrest 
neurocognitive decline and brain edema/inflammation. Manipulation of autophagy is a novel therapeutic 
approach that focuses on degradation of neurotoxic proteins at the manufacturing site in order to 
prevent their secretion and propagation. This novel strategy leads to unclogging the cell’s disposal 
machine and degradation of toxic proteins, thus preserving neuronal survival via bulk digestion. 
Preservation of neuronal survival maintains the level of neurotransmitters that are necessary for 
cognitive, motor and other CNS functions, leading to alleviation of symptoms as well as arrest of 
neurodegeneration. As neurons are post-mitotic cells, pulsatile autophagy may promote protein 
degradation and provide an effective disease-modifying therapy for neurodegenerative diseases.  
 
Autophagy is a double-edged sword in cancer, either preventing accumulation of damaged 
proteins and organelles to suppress tumors, or promoting cell survival mechanisms that lead to tumor 
growth and proliferation (87). Leukemia and many other cancer treatments have been revolutionized 
by manipulation of autophagy, which leads to bulk degradation of unwanted or toxic molecules (88). 
For example in leukemia, genetic mutations and DNA damage can lead to large numbers of abnormal 
white blood cells (leukemia cells and leukemic blast cells) to accumulate in the blood and bone marrow, 
crowding out normal blood cells. Autophagy can lead to the degradation of the products of cancer-
causing genes (oncogenes), tumor suppressor genes, damaged DNA and essential components of the 
cytosol, thereby controlling abnormal mitotic division and limiting tumor growth. Autophagy can also 
lead to self-cannibalization via promotion of programmed cell death, or apoptosis (89,90). Activation of 
the tumor suppressor p53 in response to DNA damage leads the cell to arrest proliferation, initiate DNA 
repair, and promote survival. However, if the DNA damage cannot be resolved by p53, it can trigger 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
53 
 
apoptotic death (89,90). Cell division and apoptosis are mediated by signaling mechanisms via the 
endosomal (early and recycling) system (83). Tyrosine kinases are activated via auto phosphorylation, 
triggering various signaling mechanisms that mediate cell division and/or apoptosis (91,92). Tyrosine 
kinase inhibition via de-phosphorylation leads to signaling via the late endosomal-lysosomal pathway, 
thus increasing autophagic degradation (83-85) and tumor growth. 
 
TKIs have significantly improved the life quality and expectancies in many cancers, including 
CML (93,94). CML is characterized by the translocation of chromosomes 9 and 22 to form the 
“Philadelphia” chromosome resulting in the expression of a constitutively active Breakpoint Cluster 
Region-Abelson (BCR-Abl) tyrosine kinase. This oncogenic protein activates intracellular signaling 
pathways and induces cell proliferation. Our laboratory investigated TKIs that activate autophagy and 
are FDA-approved for CML, thus significantly reducing research and development efforts and cost by 
re-purposing (reviewed in (57)) for new indications. Abl is activated in neurodegeneration (48,56,89,90). 
A fraction of Nilotinib crosses the blood-brain-barrier (BBB), inhibits Abl and facilitates autophagic 
amyloid clearance, leading to neuroprotection and improved cognition and motor behavior (49,50,54-
58). Mice treated with a much lower dose of these drugs (<25% of the typical CML dose) show 
significant motor and cognitive improvement and degradation of alpha-Synuclein, beta-amyloid, Tau 
and TDP-43 without evidence of increased inflammation (49,50,54-58). There was also significant 
reversal of neurotransmitter alterations, including dopamine and glutamate in several models of 
neurodegeneration. As a modulator of myeloid cells (95), Nilotinib may also positively regulate neuronal 
death and produce neuro-restorative effects via increased production of necessary growth factors and 
proliferation of myeloid-derived glia. Autophagic toxic protein clearance and production of growth 
factors may restore loss of neurotransmitters, leading to improved motor and cognitive functions. 
Nilotinib provides a double-edge sword via manipulation of autophagy to inhibit cell division and tumor 
growth in CML on one hand, and promote toxic protein degradation and neuronal survival in 
neurodegeneration on the other hand.  

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
54 
 
13-CLINICAL ASSESSMENTS AND OUTCOME MEASURES 
13-1. Clinical Variables 
Assessments will be performed at visits as noted above throughout the study for clinical evaluation. In 
addition to the assessments evaluated below, subjects will provide information on their demographics, 
past medical history, including PD, as well as concomitant medication usage. 
 
13.2- Vital Signs, Height & Weight 
Vital signs, including systolic and diastolic blood pressure, pulse rate (radial artery)/minute, respiratory 
rate/minute, temperature and weight will be assessed at specified visits. Height will be measured and 
recorded at the Screening Visit only.  
 
Medical history will be assessed as follows: 
 History of PD (inquiring on date of diagnosis and first symptoms) 
 PD treatment history 
 Smoking history 
 Important medical information for inclusion/exclusion  
 Significant 
medical 
and 
surgical 
history 
(e.g. 
Allergy/Immunologic, 
Cardiovascular, 
Dermatological, 
ENT, 
Gastrointestinal, 
Gynecologic/Urologic, 
Hepatobiliary, 
Hermato/Lymphatic, Metabolic/Endocrine, Musculoskeletal, Neurologic, Ophthalmologic, 
Psychiatric, Pulmonary, renal, and Other) 
Questioning of Comorbidities will be done using the Charlston Comorbidity Index and should be 
done at every visit after Baseline: 
Metastatic solid tumor:   
 
 
 
 
 
no/yes 
AIDS:  
 
 
 
 
 
 
 
no/yes 
Moderate-to-severe liver disease:  
 
 
 
no/yes 
Hemiplegia:   
 
 
 
 
 
 
no/yes 
Moderate-to-severe renal failure:   
 
 
 
no/yes 
Moderate-Diabetes with endorgan damage:  
 
 
no/yes 
Neoplasia:   
 
 
 
 
 
 
no/yes 
Leukemia:  
 
 
 
 
 
 
 
no/yes 
Lymphoma:   
 
 
 
 
 
 
no/yes 
Myocardial infarct:   
 
 
 
 
 
no/yes 
Congestive heart failure:   
 
 
 
 
no/yes 
Peripheral vascular disease:  
 
 
 
 
no/yes 
Cerebrovascular disease:   
 
 
 
 
no/yes 
Dementia:  
 
 
 
 
 
 
 
no/yes 
Chronic pulmonary disease:  
 
 
 
 
no/yes 
Connective tissue disease: 
 
 
 
 
no/yes 
Ulcer disease:  
 
 
 
 
 
 
no/yes 
Mild liver disease:   
 
 
 
 
 
no/yes 
Diabetes:  
 
 
 
 
 
 
 
no/yes 
 Concomitant medications will be recorded at every visit after baseline.   
13.3- Clinical Laboratory Assessments 
The following safety laboratory tests will be performed during the study: 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
55 
 
Standard blood chemistry, including Cholesterol, low density lipoprotein (LDL), high density lipoprotein 
(HDL), albumin, total protein, alkaline phosphatase, total bilirubin, creatinine, calcium, chloride, sodium, 
potassium, magnesium, inorganic phosphorus, bicarbonate, creatine phosphokinase (CPK), gamma-
glutamyl 
transferase 
(γ-GT), 
lactate 
dehydrogenase 
(LDH), 
lipase, 
α-amylase, 
alanine 
aminotransferase (ALT), aspartate aminotransferase (AST), blood urea nitrogen (BUN), thyroid 
stimulating hormone (TSH), vitamin B12, and glucose. A standard hematology panel including, 
complete blood count (CBC) with differential counts will be performed. Hemoglobin (Hb), hematocrit, 
red blood cell count, platelet count, and white blood cell count with differential count will be measured. 
In addition, prothrombin time-international normalized ratio (PT-INR) will be measured for assessment 
of coagulation.  
 
o Serum hCG for women of childbearing potential (WOCBP)  
 
Additional testing may be ordered if needed, to further assess an AE, or if there is any suspicion that a 
subject may be pregnant, throughout the course of the study. 
 
13.4- Physical Examination 
A physical examination will be performed and recorded. A full physical examination (including vital 
signs) will be performed by a trained physician as indicated in the schedule of assessments (Table 2).  
The Physical examination will include: 
 General appearance 
 Abdomen 
 Cardiovascular systems 
 Lungs 
 Lymph nodes 
 Musculoskeletal systems 
 Skin (with special attention to dermatological tolerability) 
 Extremities 
 Head, ears, eyes, nose, throat, and mouth 
 Thyroid gland 
 Vital signs: 
 
 
Body weight (Kg or pound) 
 
 
Height (cm or inch) (at the Screening Visit only) 
 
 
Blood pressure systolic and diastolic (mmHg) 
 
 
Heart rate (bpm) 
 
13.5- Neurological Examination 
A neurological examination will be performed and recorded as detailed in Table 2. Neurological 
examination will include: 
 Cranial nerves 
 Motor functions 
 Sensation 
 Coordination 
 Speech 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
56 
 
13.6- Adverse Events 
AEs will be documented at each study visit, including the Screening Visit once the ICF has been signed 
by the subject. Information on AEs of study medication and on inter-current events will be determined 
at each visit by direct questioning of the subjects’ review of concomitant medications, and vital sign 
results. AEs are incidents or complaints that do not require hospitalization, SAEs require hospitalization 
(see section 13). 
13.7- MDS-UPDRS (includes Hoehn & Yahn staging) is used to follow the longitudinal course of PD 
and it is the most commonly used scale in the clinical study of PD. UPDRS motor section in particular 
is used to follow the progression of a person's PD. The UPDRS is made up of these sections: Part I: 
evaluation of mentation, behavior, and mood. Part II: self-evaluation of the activities of daily life (ADLs) 
including speech, swallowing, handwriting, dressing, hygiene, falling, salivating, turning in bed, walking, 
and cutting food. Part III: clinician-scored monitored motor evaluation. Part IV: complications of therapy. 
Part V: Hoehn and Yahr staging of severity of Parkinson's disease. Part VI: Schwab and England ADL 
scale 
13.8- TMT-B. The Trail Making Test (TMT) is a neuropsychological test of visual attention and task 
switching (Reitan, 1958). It consists of two parts in which the subject is instructed to connect a set of 
25 dots as quickly as possible while still maintaining accuracy. The test can provide information about 
visual search speed, scanning, speed of processing, mental flexibility, as well as executive functioning. 
The Trail Making Test B (TMT-B) has been shown to be sensitive to changes in cognitive ability in HD, 
and has been used in an HD clinical trial (Huntington Study Group Reach2HD Investigators, 2015).  
 
13.9- MOCA is designed as a rapid screening instrument for mild cognitive dysfunction.  It assesses 
different cognitive domains, including attention and concentration, executive functions, memory, 
language, visuo-constructional skills, conceptual thinking, calculations and orientation.  It has excellent 
psychometric properties and has become a widely used screening instrument for mild cognitive 
impairment (Smith, Gildeh & Holmes, 2007). It is administered by a nurse or clinician and takes 10 - 20 
minutes.  
  
13.10- C-SSRS 
The US FDA recommends the use of a suicidality assessment instrument that maps to the Columbia 
Classification Algorithm for Suicide Assessment (C-CASA). The C-CASA was developed to assist the 
FDA in coding suicidality data accumulated during the conduct of clinical trials of antidepressant drugs. 
One such assessment instrument is the Columbia Suicide Severity Rating Scale (C-SSRS). The C-
SSRS involves a series of probing questions to inquire about possible suicidal thinking and behavior. 
The US FDA also recommends assessment of depression during every visit, so the Geriatric 
Depression Scale (GDS) will also be performed. 
Only investigators who have been fully trained in the administration of the C-SSRS and GDS will assess 
subject suicidality and depression. As part of training, investigators are prepared to respond to and 
manage instances in which patients express suicidal ideation or exhibit suicidal behavior. 
At the Baseline Visit, the C-SSRS Baseline version will be administered. This version is used to assess 
suicidality over the subject’s lifetime and specifically for the previous 6-month time period. 
At each visit and the Final Safety visit, as applicable, the Since Last Visit version of the C-SSRS will be 
administered. This version of the scale assesses suicidality since the subject’s last visit. 
Information obtained from: http://www.cssrs.columbia.edu/ 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
57 
 
13.11- TUG- Timed Up and Go (TUG) is an assessment of mobility, balance, walking ability, and fall 
risk (Podsiadlo et al, 1991).  It uses the time that a person takes to rise from a chair, walk three meters, 
turn around, walk back to the chair, and sit down.  It requires both static and dynamic balance. 
13.12- NPI. The behavioral outcome measure for this trial is the NPI. The NPI is a well-validated, 
reliable, multi-item instrument to assess psychopathology in AD based on interview with the study 
partner. The NPI evaluates both the frequency and severity of 10 neuropsychiatric disturbances. 
Frequency assessments range from 1 (occasionally, less than once per week) to 4 (very frequently, 
once or more per day or continuously) as well as severity (1=mild, 2=moderate, 3=severe).  The overall 
score and the score for each subscale are the product of severity and frequency.  
13.13- ADAS-Cog aims to evaluate cognitive impairment in the assessment of AD. Recommended for 
second stage or more detailed assessments and/or for particular research evaluations rather than for 
applications in routine care settings. It takes 30-45 min and administered by an interviewer. Requires 
additional training. ADAS-cog was included in this LBD study to better capture potential changes in 
ADL and non-ADLs and severity of cognitive impairment. 
13.14- ADCS-ADL is an activity of daily living inventory developed by the ADCS to assess functional 
performance in participants with AD. Using a structured interview format, study partners are queried as 
to whether participants attempted each item in the inventory during the prior 4 weeks and their level of 
performance.  The ADCS-ADL scale discriminates well between normal participants and those with 
mild AD and it has good test-retest reliability. The ADCS-ADL includes some items from traditional 
basic ADL tests (e.g., grooming, dressing, walking, bathing, feeding, toileting) as well as instrumental 
(complex) activities of daily living (e.g., shopping, preparing meals, using household appliances, 
keeping appointments, reading).  
13.15- CAF consists of seven items of confusional behaviour (falls, fluctuation, drowsiness, attention, 
disorganised thinking, altered level of consciousness, communication), scores for which are summed 
to provide a severity score for fluctuating confusion ranging from 0 to 21. 
13.16- IAS was developed to measure apathy and irritability in patients with dementia, including AD 
and HD (Chatterjee 2005). The IAS is a 28-item self-administered questionnaire collecting information 
about different aspects of irritability and apathy utilizing a 0-3 scale for each item to indicate severity. 
Both a patient and a study partner version can be administered. The IAS will be completed separately 
by Subjects and Study Partners (Chatterjee 2005). 
 
13.17- PBA-s is a structured interview in which a trained interviewer rates the frequency and severity 
of neuropsychiatric symptoms through observation and the reporting of the Subject and Study Partner. 
Symptoms rated include depressed mood, suicidal ideation, anxiety, irritability, angry or aggressive 
behavior, apathy, perseverative thinking or behavior, obsessive-compulsive behaviors, delusional or 
paranoid thinking, hallucinations, and disoriented behavior. Each behavioral problem is rated for both 
severity and frequency on a 0-4- point scale; severity and frequency ratings are then multiplied to 
provide an overall score for each symptom. 
 
13.18- CSF and Blood Biomarkers  
Subjects will also be asked to provide blood samples for biomarker analysis per Schedule of Activities. 
Blood samples will be stored in a sample repository at Georgetown University Medical Center (GUMC) 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
58 
 
laboratory for Dementia and Parkinsonism, where bio fluids will be indefinitely stored and may be used 
for further biomarker validation studies. All samples will be labeled with a code. The code will not include 
any identifiable information. Any analysis performed on these samples is for research purposes only. 
Unused samples will remain in the biorepository for future PD-related research. There is no scheduled 
date on which the samples will be destroyed. Samples may be stored for research until they are used, 
damaged, decayed or otherwise unfit for analysis. Subjects have the option of declining participation in 
this portion of the study at any time by withdrawing their consent to have their sample used. However, 
it will not be possible to destroy samples that may have already been used.  
 
14- BIOMARKERS  
 
14-1- Rationale for CSF and plasma biomarkers. The biomarkers proposed include CSF levels of 
alpha-Synuclein, total Tau and p-Tau181/231, which yielded useful results in our open label pilot trial and 
are identified as potential CSF biomarkers for PD and DLB pathology.  
Several studies indicate that reduction of CSF alpha-Synuclein is associated with alpha-Synuclein 
pathology in the CNS (96-98). CSF alpha-Synuclein, which probably comes from a neuronal source, 
has a lower level than the highly abundant presence of peripheral alpha-Synuclein (99). However, only 
a few studies have investigated CSF alpha-Synuclein levels in patients with neuropathologically 
confirmed alpha-Synucleinopathies (65,100). CSF alpha-Synuclein is reduced in de novo PD patients 
compared with healthy individuals (101), and it is lower in patients with advanced alpha- 
Synucleinopathies compared to other neurological disorders (100). Our pilot study showed that alpha-
Synuclein is significantly decreased with 150mg Nilotinib between baseline and 6 months, perhaps 
reflecting the natural progression of the disease. However, 300mg Nilotinib attenuates the loss of CSF 
alpha-Synuclein between baseline and 6 months treatment. Therefore, we expect Nilotinib to reduce 
the decrease in CSF alpha-Synuclein level indicating reduced cell death, which will also be compared 
with direct markers of cell death, including NSE and S100B. 
 
Tau pathology is also frequently found in the CNS of PD patients and have been associated with the 
development of cognitive impairment and PD dementia (102). Several studies also demonstrate 
increased levels of CSF total Tau and p-Tau181 levels (65-67) in PD patients, but another study report 
slightly decreased or normal levels of CSF total Tau and p-Tau (103). The CSF variably as reported in 
the literature in CSF Tau levels may be due to disease stage and/or differences in diagnostic criteria. 
Therefore, based on the preliminary data we obtained from our pilot trial showing changes in CSF Tau 
with Nilotinib treatment, we will measure CSF Tau in this study as another marker of cell death and 
compare Tau with NSE and S100B at baseline and 6 months in a placebo-controlled study. 
 
The R47H and other variants of triggering receptors on myeloid cells (TREM)-2, which result in 
loss of TREM2 function, are strong risk factors for PD and DLB (104-106). Activated microglia in the 
SN proliferate and produce reactive oxygen species and pro-inflammatory cytokines, resulting in 
progressive degeneration of DA neurons in PD  (107,108). TREM2 may regulate microglial response 
and phagocytosis. TREM2 inhibits inflammatory responses in microglia via suppression of NF-kB 
pathways and activation of innate immunity (109), while TREM2 loss of function results in reduced 
microglial phagocytosis (110-112). Therefore, measuring TREM2 levels in the CSF may provide 
another important pharmacodynamics effect indicating neuro-inflammation and the phagocytic activity 
of microglia to potentially reduce alpha-synuclein levels after Bosutinib treatment of DLB patients. The 
biomarkers proposed include CSF levels of total and oligomeric alpha-Synuclein, total tau and p-
tau181/231, which are identified as potential CSF biomarkers for PD and DLB pathology.  
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
59 
 
Other candidate biomarkers for PD include catecholamines, such as dopamine and noradrenaline, and 
their metabolites. Levels of CSF HVA, which is the end byproduct of the neuronal metabolite of 
dopamine is reduced in PD (113,114), reflecting diminution of stores of central dopamine. We will 
measure CSF HVA and quantify DaTscan to measure dopamine transporters activity in collaboration 
with GE Healthcare at baseline, 3 months (interim) and 6-months to correlate imaging and CSF 
biomarkers of dopamine metabolism. DaTscan is clinically useful in early stages of PD and since we 
observe an increase in HVA level with Nilotinib treatment, we expect Nilotinib to increase HVA levels, 
reflecting an increase in central dopamine stores. We also expect Nilotinib to increase dopamine 
transporters activity, which will be quantified at baseline and compared to 3 and 6 months Nilotinib 
treatment. We will also measure changes in exploratory biomarkers of pathophysiology, which may 
show Nilotinib effects on biomarker levels that will help us to build a better clinical development program 
going forward. 
 
CSF, plasma DLB biomarkers: Blood draw (15ml) and lumbar puncture (LPs) to obtain CSF (~15ml) 
will be performed on all patients at baseline and 12-months after treatment. Blood will be drawn 0.5hr 
before LP s outlined above. Plasma will be isolated immediately after blood draw and will be aliquoted 
and stored at -80oC. CSF will be aliquoted and stored at -80oC. Freeze and thaw cycles will be avoided. 
To avoid CSF contamination with blood, the first 1 mL of CSF collection will be discarded and all 
samples will be centrifuged at 1000g for 15 minutes. Samples that contain a detectable level of 
Hemoglobin will be eliminated from alpha-Synuclein and HVA evaluation. 
Plasma and CSF sample preparation for Mass Spec to determine Nilotinib pharmacokinetics  
Plasma and CSF samples (20 µl) will be thawed initially on ice at room temperature and transfused to 
Eppendorf tubes containing 100µl of water. 500µl extraction solvent, Acetonitrile/Methanol (50:50) 
containing the internal standard (5ng/mL of Nilotinib_13C_2H3) will be added to the sample. The 
mixture will be vortexed and incubated for 20min on ice to accelerate protein precipitation. After 
incubation, the samples will be vortexed and centrifuged at 13,000 rpm for 20 min at 4OC. The 
supernatant is freeze- dried using speed vacuum and reconstituted in 200µL of Methanol: Water (50:50) 
and processed by Mass Spectrometry. 
 
14.2- Pharmacokinetics. Plasma and CSF will be collected 2hrs (Tmax Figure 5) after oral 
administration of Nilotinib to determine the pharmacokinetic parameters of Nilotinib after dosing with 
150 mg and 300 mg. Note that no Nilotinib was detected in CSF after 4 hrs. in our prior study (Figure 
5 A). Quantitation of Nilotinib will be performed using multiple reactions monitoring mass spectrometry. 
The samples will be resolved on an Acquity UPLC BEH C18 1.7µm, 2.1 x 50 mm column online with a 
triple quadrupole mass spectrometer (Xevo-TQ-S, Waters Corporation, USA) operating in the multiple 
reaction monitoring (MRM) mode (The sample cone voltage and collision energies will optimized for 
both analytes to obtain maximum ion intensity for parent and daughter ions using “IntelliStart” feature 
of MassLynx software (Waters Corporation, USA). The instrument parameters will be optimized to gain 
maximum specificity and sensitivity of ionization for the parent [m/z = 530.27 (Nilotinib), 438.25 and 
daughter ions [m/z = 289.01 (Nilotinib). Signal intensities from all MRM Q1/Q3 ion pairs for both 
analytes are ranked to ensure selection of the most intense precursor and fragment ion pair for MRM-
based quantitation. The metabolite ratios are calculated by normalizing the peak area of endogenous 
metabolites within tissue samples normalized to the internal standard Nilotinib_13C_2H3.  
 
14.3- Amyloid PET. Abeta positron emission tomography (PET) imaging Florbetaben F18 
(Neuraceq™; Piramal Imaging) is a radioactive tracer to help visualize beta-amyloid plaque deposits in 
AD. Florbetaben F18 PET amyloid imaging offers a unique opportunity to detect the presence of Abeta 
in the human body during life. This technology is FDA-approved to detect Abeta neuritic plaque density 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
60 
 
in adult patients with cognitive impairment who are being evaluated for AD and/or other causes of 
cognitive decline. The objective of Abeta image interpretation “is to estimate beta-amyloid neuritic 
plaque density in brain gray matter. Florbetaben is not intended for use in predicting development of 
dementia or other neurological condition, its use in the current studies is intended to monitor responses 
to Nilotinib treatment.  
 
14.4- Cell death and exploratory biomarkers. NSE and S100B (glia and neurons), total Tau and 
phosphorylated Tau will be measured using Millipore ELISA at baseline and after 12 months Nilotinib 
treatment. Nilotinib may have a modulatory effect on myeloid cells (95), which can either proliferate and 
differentiate into peripheral macrophages or become myeloid-derived glia that cross the BBB and 
produce neurotrophic and/or inflammatory markers. Nilotinib may affect CNS glial progenitor cells. Our 
preliminary preclinical and clinical data show Nilotinib-induced alterations of peripheral and CNS 
inflammatory makers. Therefore, we will perform unbiased multiplex ELISA (Millipore) to profile a panel 
of 44 plasma and CSF markers, including TREM2, interleukins (IL)- 1α&β, 2, 3, 4, 5, 6, 7, 8 (CXCL8), 
9, 10, 12, 13, 15, 17 α, and chemokines (C-C) including, CXCL10, CCL2, CL7, CCL22, CCL3, CCL4, 
platelet-derived growth factor (PDGF)-AA, PDGF-AB/BB, CCL5, CX3CL1 (fractalkine), Tumor necrosis 
growth factor (TNF)-α, transforming growth factor (TGF)-α, and vascular endothelial growth factor 
(VEGF), GFAP, neurofilaments, glial fibrillary acidic protein (GFAP) and TDP-43.  
 
14.5- Alpha-Synuclein ELISA. Solid phase alpha-Synuclein sandwich ELISA (Cat#SIG38974, 
Biolegend) will be performed on CSF and plasma. To avoid repetitive freeze and thaw cycles, 
immediately after LP and blood draws, 15 mL CSF and 5mL plasma will be aliquoted on ice into 0.5mL 
tubes and stored at -80oC. Fresh aliquots were used to perform or repeat ELISA. Total alpha- Synuclein 
rabbit monoclonal antibody (amino acids 118-123) will be coated on the microwells and 200µl CSF or 
plasma will be added to designated wells. CSF samples will be diluted 1:10 while plasma samples will 
be diluted 1:50. After overnight sample incubation at 2-8oC, alpha-Synuclein will be captured by the 
coated antibody. After washing, a biotinylated mouse monoclonal alpha-Synuclein (amino acids 103-
107) detection antibody will be added to each well to detect the captured alpha-Synuclein (amino acids 
118-123). Samples will be incubated with 50µl of detection antibody for 2hrs at room temperature. After 
washing, 200µl of streptavidin HRP will be added and incubated for 1hr at room temperature to 
recognize the bound biotinylated detection antibody. Samples will then washed and incubated with 
100µl of chemiluminescent substrates. Plates will be shaken for 10-15 seconds and read immediately 
by a luminometer. The magnitude of the luminescence is proportional to the quantity of alpha-Synuclein 
in the samples. 
 
14.6- Homovanillic Acid and DOAPC ELISA. A 100µl CSF or plasma samples will be incubated with 
100µl HRP-conjugate reagent and incubated for 1hr at 37oC using solid phase sandwich ELISA 
(MyBioSource, Cat# MBS064661). All samples at baseline and 6 months will be analyzed side-by-side 
using same reagents. After washing, 50µl of chromogen solution A and 50µl of chromogen solution B 
will be added to the solution and incubated for 15min at 37oC. The reaction will be stopped with 50µl 
stop solution and the optical density will be read at 450nM. The magnitude of the absorbance is 
proportional to the quantity of CSF and plasma HVA.HVA and DOPAC levels will be confirmed by LC-
MS. 
 
14.7- Total Tau and p-Tau181 measurement. Solid phase human total Tau sandwich ELISA 
(Invitrogen, Cat# KHB0042) and p-Tau181 (Invitrogen, Cat# KH0061) will be performed on CSF 
samples. All samples at baseline and 6 months will be analyzed side-by-side using same reagents. A 
monoclonal Tau or p-Tau181 capture antibodies will be coated onto micro-wells. 50µl of CSF will be 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
61 
 
added to each well, allowing human Tau or p-Tau181 antigen to bind to the immobilized capture 
antibody, and incubated for 2hrs at room temperature. After 2hr incubation, samples will be washed 
and incubated with 100µl of total Tau or p-Tau181 detection antibody and incubated for 1hr at room 
temperature. After washing, 100µl of HRP labeled anti-rabbit IgG will be added to each well and 
incubated for 30min at room temperature. Samples will be washed to remove all unbound enzyme 
and100µl TMB, a HRP substrate, will be added to develop color. The magnitude of the absorbance for 
this developed color is proportional to the quantity of CSF total Tau or p-Tau181.  
 
14.8- Human Neurodegenerative Disease Panels. We will use a multiplex Xmap technology that 
uses magnetic microspheres internally coded with two fluorescent dyes to measure markers of 
neurodegeneration. All samples at baseline and 6 months will be analyzed side-by-side using same 
reagents. Through precise combinations of these two dyes, multiple proteins are measured within the 
sample. Each of these spheres is coated with a specific capture antibody. The capture antibody binds 
to the detection antibody and a reporter molecule, completing the reaction on the surface of the bead. 
25µL of CSF or plasma will be incubated overnight at 4oC with 25µL of a mixed bead solution, containing 
human total Tau, pTau231 and NSE (Millipore, Cat#: HND1MAG-39K) or S100B, Aβ42, and Aβ40 (CSF 
Aβ40 is diluted 1:10). After washing, samples will be incubated with 25µL detection antibody solution 
for 1.5hrs at room temperature (Millipore, CAT#: HND4MAG-36K), 25µL of Streptavidin-Phycoerythrin 
will be added to each well containing the 25µl of detection antibody solution. Samples were then 
washed and suspended in 100µl of sheath fluid. Samples will be then run on MAGPIX with Xponent 
software. The Median Fluorescent Intensity (MFI) data will be analyzed using a 5-parameter logistic or 
spline curve-fitting method for calculating analyte concentrations in samples.  

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
62 
 
15- DATA COLLECTION 
All data collected as part of this study will be entered into a secure data management site maintained 
by Georgetown University Information Service (UIS). All data will be stored on a UIS and IRB approved 
databases, including long-term storage on Amazon Web Service (WAS) and will be deposited on 
RedCAP for an external and independent data analysis and monitoring. Data will be stored in EDC 
maintained by GUMC. This platform facilitates: 
1. Capture of clinical and research data from neurologic patients for individual projects in a 
structured and secure system; 
2. Aggregating and sharing uniform, de-identified and/or anonymized datasets for secondary 
analyses. 
 
15.1- Role of Data Management 
Data Management (DM) is responsible for the development, execution and supervision of plans, 
policies, programs, and practices that control, protect, deliver, and enhance the value of data and 
information assets. 
 
All data will be managed in compliance with applicable Sponsor and regulatory requirements. Site 
personnel will collect, transcribe, correct, and transmit the data onto source documents, Case Report 
Forms (CRFs), and/or other forms used to report, track and record clinical research data. DM is 
responsible for developing, testing, and managing clinical data management activities. 
 
15.2- Data Entry and Checks 
The site personnel are instructed to enter information into the EDC. Data capture is the responsibility 
of the staff at the site under the supervision of the PIs.  During the study, the PIs must maintain complete 
and accurate documentation for the study. The EDC provides password protection. An edit checking 
and data clarification process will be put in place to ensure accuracy of the data. Logic and range 
checks as well as more sophisticated rules may be built into the eCRFs to provide immediate error 
checking of the data entered.  The system has the capability to automatically create electronic queries 
for forms that contain data that are out of range, out of window, missing or not calculated correctly.   
 
15.3- Data Lock Process 
The platform will have the ability to lock the project-specific visits to prevent any modification of data 
once the project is closed.  Once this option is activated, every user will have Read-Only access to the 
data. 
 
15.4- Data handling and record keeping 
The PIs are responsible to ensure the accuracy, completeness, legibility, and timeliness of the data 
reported.  Data reported in the eCRF derived from source documents should be consistent with the 
source documents and discrepancies should be explained. 
 
15.5- Confidentiality 
The EDC software and RedCAP databases and patient data reside on Georgetown University IRB-
approved servers. Physical and software access to the servers and security is provided by GUMC 
investigators. 
 
15.6- Retention of Records 
Research records will be retained in accordance with site IRB policies.  
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
63 
 
15.7- Publications 
The PIs will be responsible for publications of results from this trial. Responsibilities will include the 
following: 
 Analyze and interpret data gathered in this study, and write publications from these data. 
 Submit manuscripts to selected journals and address peer reviewers’ comments. 
 Submit abstracts to selected meetings and present data at the meetings. 
 Determine authorship on the basis of the Uniform Requirements for Manuscripts. 
 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
64 
 
16 
LITERATURE REFERENCES 
1. 
Guerreiro, R., Ross, O. A., Kun-Rodrigues, C., Hernandez, D. G., Orme, T., Eicher, J. D., 
Shepherd, C. E., Parkkinen, L., Darwent, L., Heckman, M. G., Scholz, S. W., Troncoso, J. C., 
Pletnikova, O., Ansorge, O., Clarimon, J., Lleo, A., Morenas-Rodriguez, E., Clark, L., Honig, L. 
S., Marder, K., Lemstra, A., Rogaeva, E., St George-Hyslop, P., Londos, E., Zetterberg, H., 
Barber, I., Braae, A., Brown, K., Morgan, K., Troakes, C., Al-Sarraj, S., Lashley, T., Holton, J., 
Compta, Y., Van Deerlin, V., Serrano, G. E., Beach, T. G., Lesage, S., Galasko, D., Masliah, E., 
Santana, I., Pastor, P., Diez-Fairen, M., Aguilar, M., Tienari, P. J., Myllykangas, L., Oinas, M., 
Revesz, T., Lees, A., Boeve, B. F., Petersen, R. C., Ferman, T. J., Escott-Price, V., Graff-
Radford, N., Cairns, N. J., Morris, J. C., Pickering-Brown, S., Mann, D., Halliday, G. M., Hardy, 
J., Trojanowski, J. Q., Dickson, D. W., Singleton, A., Stone, D. J., and Bras, J. (2018) 
Investigating the genetic architecture of dementia with Lewy bodies: a two-stage genome-wide 
association study. Lancet Neurol 17, 64-74 
2. 
Dickson, D. W. (2018) Neuropathology of Parkinson disease. Parkinsonism Relat Disord 46 
Suppl 1, S30-S33 
3. 
Bharadwaj, R., Cimino, P. J., Flanagan, M. E., Latimer, C. S., Gonzalez-Cuyar, L. F., Juric-
Sekhar, G., Montine, T. J., Marshall, D. A., and Keene, C. D. (2018) Application of the condensed 
protocol for the NIA-AA guidelines for the neuropathological assessment of Alzheimer's disease 
in an academic clinical practice. Histopathology 72, 433-440 
4. 
Brenowitz, W. D., Keene, C. D., Hawes, S. E., Hubbard, R. A., Longstreth, W. T., Jr., Woltjer, R. 
L., Crane, P. K., Larson, E. B., and Kukull, W. A. (2017) Alzheimer's disease neuropathologic 
change, Lewy body disease, and vascular brain injury in clinic- and community-based samples. 
Neurobiology of aging 53, 83-92 
5. 
Montine, T. J., Koroshetz, W. J., Babcock, D., Dickson, D. W., Galpern, W. R., Glymour, M. M., 
Greenberg, S. M., Hutton, M. L., Knopman, D. S., Kuzmichev, A. N., Manly, J. J., Marder, K. S., 
Miller, B. L., Phelps, C. H., Seeley, W. W., Sieber, B. A., Silverberg, N. B., Sutherland, M., 
Torborg, C. L., Waddy, S. P., Zlokovic, B. V., Corriveau, R. A., and Committee, A. C. O. (2014) 
Recommendations of the Alzheimer's disease-related dementias conference. Neurology 83, 
851-860 
6. 
Corriveau, R. A., Koroshetz, W. J., Gladman, J. T., Jeon, S., Babcock, D., Bennett, D. A., 
Carmichael, S. T., Dickinson, S. L., Dickson, D. W., Emr, M., Fillit, H., Greenberg, S. M., Hutton, 
M. L., Knopman, D. S., Manly, J. J., Marder, K. S., Moy, C. S., Phelps, C. H., Scott, P. A., Seeley, 
W. W., Sieber, B. A., Silverberg, N. B., Sutherland, M. L., Taylor, A., Torborg, C. L., Waddy, S. 
P., Gubitz, A. K., and Holtzman, D. M. (2017) Alzheimer's Disease-Related Dementias Summit 
2016: National research priorities. Neurology 89, 2381-2391 
7. 
McKeith, I. G., Boeve, B. F., Dickson, D. W., Halliday, G., Taylor, J. P., Weintraub, D., Aarsland, 
D., Galvin, J., Attems, J., Ballard, C. G., Bayston, A., Beach, T. G., Blanc, F., Bohnen, N., 
Bonanni, L., Bras, J., Brundin, P., Burn, D., Chen-Plotkin, A., Duda, J. E., El-Agnaf, O., Feldman, 
H., Ferman, T. J., Ffytche, D., Fujishiro, H., Galasko, D., Goldman, J. G., Gomperts, S. N., Graff-
Radford, N. R., Honig, L. S., Iranzo, A., Kantarci, K., Kaufer, D., Kukull, W., Lee, V. M. Y., 
Leverenz, J. B., Lewis, S., Lippa, C., Lunde, A., Masellis, M., Masliah, E., McLean, P., 
Mollenhauer, B., Montine, T. J., Moreno, E., Mori, E., Murray, M., O'Brien, J. T., Orimo, S., 
Postuma, R. B., Ramaswamy, S., Ross, O. A., Salmon, D. P., Singleton, A., Taylor, A., Thomas, 
A., Tiraboschi, P., Toledo, J. B., Trojanowski, J. Q., Tsuang, D., Walker, Z., Yamada, M., and 
Kosaka, K. (2017) Diagnosis and management of dementia with Lewy bodies: Fourth consensus 
report of the DLB Consortium. Neurology 89, 88-100 
8. 
Postupna, N., Latimer, C. S., Larson, E. B., Sherfield, E., Paladin, J., Shively, C. A., Jorgensen, 
M. J., Andrews, R. N., Kaplan, J. R., Crane, P. K., Montine, K. S., Craft, S., Keene, C. D., and 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
65 
 
Montine, T. J. (2017) Human Striatal Dopaminergic and Regional Serotonergic Synaptic 
Degeneration with Lewy Body Disease and Inheritance of APOE epsilon4. Am J Pathol 187, 
884-895 
9. 
Morimoto, S., Takao, M., Hatsuta, H., Nishina, Y., Komiya, T., Sengoku, R., Nakano, Y., Uchino, 
A., Sumikura, H., Saito, Y., Kanemaru, K., and Murayama, S. (2017) Homovanillic acid and 5-
hydroxyindole acetic acid as biomarkers for dementia with Lewy bodies and coincident 
Alzheimer's disease: An autopsy-confirmed study. PloS one 12, e0171524 
10. 
Thomas, A. J., Attems, J., Colloby, S. J., O'Brien, J. T., McKeith, I., Walker, R., Lee, L., Burn, 
D., Lett, D. J., and Walker, Z. (2017) Autopsy validation of 123I-FP-CIT dopaminergic 
neuroimaging for the diagnosis of DLB. Neurology 88, 276-283 
11. 
Walker, Z., Moreno, E., Thomas, A., Inglis, F., Tabet, N., Stevens, T., Whitfield, T., Aarsland, D., 
Rainer, M., Padovani, A., and Da, T. D. L. B. P. S. G. (2016) Evolution of clinical features in 
possible DLB depending on FP-CIT SPECT result. Neurology 87, 1045-1051 
12. 
Walker, Z., Moreno, E., Thomas, A., Inglis, F., Tabet, N., Rainer, M., Pizzolato, G., Padovani, 
A., and Da, T. D. L. B. P. S. G. (2015) Clinical usefulness of dopamine transporter SPECT 
imaging with 123I-FP-CIT in patients with possible dementia with Lewy bodies: randomised 
study. Br J Psychiatry 206, 145-152 
13. 
Papathanasiou, N. D., Boutsiadis, A., Dickson, J., and Bomanji, J. B. (2012) Diagnostic accuracy 
of (1)(2)(3)I-FP-CIT (DaTSCAN) in dementia with Lewy bodies: a meta-analysis of published 
studies. Parkinsonism Relat Disord 18, 225-229 
14. 
Spillantini, M. G., Schmidt, M. L., Lee, V. M., Trojanowski, J. Q., Jakes, R., and Goedert, M. 
(1997) Alpha-synuclein in Lewy bodies. Nature 388, 839-840 
15. 
Spillantini, M. G., Crowther, R. A., Jakes, R., Hasegawa, M., and Goedert, M. (1998) alpha-
Synuclein in filamentous inclusions of Lewy bodies from Parkinson's disease and dementia with 
lewy bodies. Proceedings of the National Academy of Sciences of the United States of America 
95, 6469-6473 
16. 
Goedert, M. (2001) Alpha-synuclein and neurodegenerative diseases. Nature reviews. 
Neuroscience 2, 492-501 
17. 
Martin, E. R., Scott, W. K., Nance, M. A., Watts, R. L., Hubble, J. P., Koller, W. C., Lyons, K., 
Pahwa, R., Stern, M. B., Colcher, A., Hiner, B. C., Jankovic, J., Ondo, W. G., Allen, F. H., Jr., 
Goetz, C. G., Small, G. W., Masterman, D., Mastaglia, F., Laing, N. G., Stajich, J. M., Ribble, R. 
C., Booze, M. W., Rogala, A., Hauser, M. A., Zhang, F., Gibson, R. A., Middleton, L. T., Roses, 
A. D., Haines, J. L., Scott, B. L., Pericak-Vance, M. A., and Vance, J. M. (2001) Association of 
single-nucleotide polymorphisms of the tau gene with late-onset Parkinson disease. JAMA 286, 
2245-2250 
18. 
Zabetian, C. P., Hutter, C. M., Factor, S. A., Nutt, J. G., Higgins, D. S., Griffith, A., Roberts, J. 
W., Leis, B. C., Kay, D. M., Yearout, D., Montimurro, J. S., Edwards, K. L., Samii, A., and Payami, 
H. (2007) Association analysis of MAPT H1 haplotype and subhaplotypes in Parkinson's 
disease. Ann Neurol 62, 137-144 
19. 
Healy, D. G., Abou-Sleiman, P. M., Lees, A. J., Casas, J. P., Quinn, N., Bhatia, K., Hingorani, A. 
D., and Wood, N. W. (2004) Tau gene and Parkinson's disease: a case-control study and meta-
analysis. J Neurol Neurosurg Psychiatry 75, 962-965 
20. 
Maraganore, D. M., Hernandez, D. G., Singleton, A. B., Farrer, M. J., McDonnell, S. K., Hutton, 
M. L., Hardy, J. A., and Rocca, W. A. (2001) Case-Control study of the extended tau gene 
haplotype in Parkinson's disease. Ann Neurol 50, 658-661 
21. 
Wood, N. W. (1998) Genetic risk factors in Parkinson's disease. Ann Neurol 44, S58-62 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
66 
 
22. 
Apaydin, H., Ahlskog, J. E., Parisi, J. E., Boeve, B. F., and Dickson, D. W. (2002) Parkinson 
disease neuropathology: later-developing dementia and loss of the levodopa response. Arch 
Neurol 59, 102-112 
23. 
Ferman, T. J., Smith, G. E., Kantarci, K., Boeve, B. F., Pankratz, V. S., Dickson, D. W., Graff-
Radford, N. R., Wszolek, Z., Van Gerpen, J., Uitti, R., Pedraza, O., Murray, M. E., Aakre, J., 
Parisi, J., Knopman, D. S., and Petersen, R. C. (2013) Nonamnestic mild cognitive impairment 
progresses to dementia with Lewy bodies. Neurology 81, 2032-2038 
24. 
Graff-Radford, J., Murray, M. E., Lowe, V. J., Boeve, B. F., Ferman, T. J., Przybelski, S. A., 
Lesnick, T. G., Senjem, M. L., Gunter, J. L., Smith, G. E., Knopman, D. S., Jack, C. R., Jr., 
Dickson, D. W., Petersen, R. C., and Kantarci, K. (2014) Dementia with Lewy bodies: basis of 
cingulate island sign. Neurology 83, 801-809 
25. 
Labbe, C., Ogaki, K., Lorenzo-Betancor, O., Soto-Ortolaza, A. I., Walton, R. L., Rayaprolu, S., 
Fujioka, S., Murray, M. E., Heckman, M. G., Puschmann, A., McCarthy, A., Lynch, T., Siuda, J., 
Opala, G., Rudzinska, M., Krygowska-Wajs, A., Barcikowska, M., Czyzewski, K., Sanotsky, Y., 
Rektorova, I., McLean, P. J., Rademakers, R., Ertekin-Taner, N., Hassan, A., Ahlskog, J. E., 
Boeve, B. F., Petersen, R. C., Maraganore, D. M., Adler, C. H., Ferman, T. J., Parisi, J. E., Graff-
Radford, N. R., Uitti, R. J., Wszolek, Z. K., Dickson, D. W., and Ross, O. A. (2015) Role for the 
microtubule-associated protein tau variant p.A152T in risk of alpha-synucleinopathies. 
Neurology 85, 1680-1686 
26. 
Irwin, D. J., Xie, S. X., Coughlin, D., Nevler, N., Akhtar, R. S., McMillan, C. T., Lee, E. B., Wolk, 
D. A., Weintraub, D., Chen-Plotkin, A., Duda, J. E., Spindler, M., Siderowf, A., Hurtig, H. I., Shaw, 
L. M., Grossman, M., and Trojanowski, J. Q. (2018) CSF tau and amyloid-beta predict cerebral 
synucleinopathy in autopsied Lewy body disorders. Neurology  
27. 
Armstrong, R. A., Cairns, N. J., and Lantos, P. L. (1998) The spatial patterns of Lewy bodies, 
senile plaques, and neurofibrillary tangles in dementia with Lewy bodies. Experimental 
neurology 150, 122-127 
28. 
Koga, S., Aoki, N., Uitti, R. J., van Gerpen, J. A., Cheshire, W. P., Josephs, K. A., Wszolek, Z. 
K., Langston, J. W., and Dickson, D. W. (2015) When DLB, PD, and PSP masquerade as MSA: 
an autopsy study of 134 patients. Neurology 85, 404-412 
29. 
Vermeiren, Y., Van Dam, D., Aerts, T., Engelborghs, S., Martin, J. J., and De Deyn, P. P. (2015) 
The monoaminergic footprint of depression and psychosis in dementia with Lewy bodies 
compared to Alzheimer's disease. Alzheimers Res Ther 7, 7 
30. 
Bousiges, O., Bombois, S., Schraen, S., Wallon, D., Quillard, M. M., Gabelle, A., Lehmann, S., 
Paquet, C., Amar-Bouaziz, E., Magnin, E., Miguet-Alfonsi, C., Delbeuck, X., Lavaux, T., Anthony, 
P., Philippi, N., Blanc, F., e, P. L. M. n., and collaborators. (2018) Cerebrospinal fluid Alzheimer 
biomarkers can be useful for discriminating dementia with Lewy bodies from Alzheimer's disease 
at the prodromal stage. J Neurol Neurosurg Psychiatry  
31. 
Arai, H., Morikawa, Y., Higuchi, M., Matsui, T., Clark, C. M., Miura, M., Machida, N., Lee, V. M., 
Trojanowski, J. Q., and Sasaki, H. (1997) Cerebrospinal fluid tau levels in neurodegenerative 
diseases with distinct tau-related pathology. Biochem Biophys Res Commun 236, 262-264 
32. 
Aerts, M. B., Esselink, R. A., Claassen, J. A., Abdo, W. F., Bloem, B. R., and Verbeek, M. M. 
(2011) CSF tau, Abeta42, and MHPG differentiate dementia with Lewy bodies from Alzheimer's 
disease. Journal of Alzheimer's disease : JAD 27, 377-384 
33. 
Gan-Or, Z., Dion, P. A., and Rouleau, G. A. (2015) Genetic perspective on the role of the 
Autophagy-Lysosome Pathway in Parkinson disease. Autophagy, 0 
34. 
Winslow, A. R., and Rubinsztein, D. C. (2011) The Parkinson disease protein alpha-synuclein 
inhibits autophagy. Autophagy 7, 429-431 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
67 
 
35. 
Winslow, A. R., Chen, C. W., Corrochano, S., Acevedo-Arozena, A., Gordon, D. E., Peden, A. 
A., Lichtenberg, M., Menzies, F. M., Ravikumar, B., Imarisio, S., Brown, S., O'Kane, C. J., and 
Rubinsztein, D. C. (2010) alpha-Synuclein impairs macroautophagy: implications for Parkinson's 
disease. J Cell Biol 190, 1023-1037 
36. 
Lonskaya, I., Hebron, M. L., Algarzae, N. K., Desforges, N., and Moussa, C. E. (2012) Decreased 
parkin solubility is associated with impairment of autophagy in the nigrostriatum of sporadic 
Parkinson's disease. Neuroscience  
37. 
Hebron, M. L., Lonskaya, I., and Moussa, C. E. (2013) Nilotinib reverses loss of dopamine 
neurons and improves motor behavior via autophagic degradation of alpha-synuclein in 
Parkinson's disease models. Human molecular genetics 22, 3315-3328 
38. 
Boland, B., Kumar, A., Lee, S., Platt, F. M., Wegiel, J., Yu, W. H., and Nixon, R. A. (2008) 
Autophagy induction and autophagosome clearance in neurons: relationship to autophagic 
pathology in Alzheimer's disease. J Neurosci 28, 6926-6937 
39. 
Kegel, K. B., Kim, M., Sapp, E., McIntyre, C., Castano, J. G., Aronin, N., and DiFiglia, M. (2000) 
Huntingtin expression stimulates endosomal-lysosomal activity, endosome tubulation, and 
autophagy. J Neurosci 20, 7268-7278 
40. 
Nixon, R. A., Wegiel, J., Kumar, A., Yu, W. H., Peterhoff, C., Cataldo, A., and Cuervo, A. M. 
(2005) Extensive involvement of autophagy in Alzheimer disease: an immuno-electron 
microscopy study. J Neuropathol Exp Neurol 64, 113-122 
41. 
Ravikumar, B., Duden, R., and Rubinsztein, D. C. (2002) Aggregate-prone proteins with 
polyglutamine and polyalanine expansions are degraded by autophagy. Hum Mol Genet 11, 
1107-1117 
42. 
Sabatini, D. M. (2006) mTOR and cancer: insights into a complex relationship. Nat Rev Cancer 
6, 729-734 
43. 
Stefanis, L., Larsen, K. E., Rideout, H. J., Sulzer, D., and Greene, L. A. (2001) Expression of 
A53T mutant but not wild-type alpha-synuclein in PC12 cells induces alterations of the ubiquitin-
dependent degradation system, loss of dopamine release, and autophagic cell death. J Neurosci 
21, 9549-9560 
44. 
Webb, J. L., Ravikumar, B., Atkins, J., Skepper, J. N., and Rubinsztein, D. C. (2003) Alpha-
Synuclein is degraded by both autophagy and the proteasome. J Biol Chem 278, 25009-25013 
45. 
Nixon, R. A. (2013) The role of autophagy in neurodegenerative disease. Nature medicine 19, 
983-997 
46. 
Salomoni, P., and Calabretta, B. (2009) Targeted therapies and autophagy: new insights from 
chronic myeloid leukemia. Autophagy 5, 1050-1051 
47. 
Lonskaya, I., Hebron, M. L., Desforges, N. M., Franjie, A., and Moussa, C. E. (2013) Tyrosine 
kinase inhibition increases functional parkin-Beclin-1 interaction and enhances amyloid 
clearance and cognitive performance. EMBO Mol Med 5, 1247-1262 
48. 
Lonskaya, I., Hebron, M. L., Desforges, N. M., Schachter, J. B., and Moussa, C. E. (2014) 
Nilotinib-induced autophagic changes increase endogenous parkin level and ubiquitination, 
leading to amyloid clearance. J Mol Med (Berl) 92, 373-386 
49. 
Lonskaya, I., Hebron, M., Chen, W., Schachter, J., and Moussa, C. (2014) Tau deletion impairs 
intracellular beta-amyloid-42 clearance and leads to more extracellular plaque deposition in 
gene transfer models. Molecular neurodegeneration 9, 46 
50. 
Lonskaya, I., Desforges, N. M., Hebron, M. L., and Moussa, C. E. (2013) Ubiquitination increases 
parkin activity to promote autophagic alpha-synuclein clearance. PLoS One 8, e83914 
51. 
Deremer, D. L., Ustun, C., and Natarajan, K. (2008) Nilotinib: a second-generation tyrosine 
kinase inhibitor for the treatment of chronic myelogenous leukemia. Clin Ther 30, 1956-1975 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
68 
 
52. 
Skorski, T. (2011) BCR-ABL1 kinase: hunting an elusive target with new weapons. Chem Biol 
18, 1352-1353 
53. 
Mahon, F. X., Hayette, S., Lagarde, V., Belloc, F., Turcq, B., Nicolini, F., Belanger, C., Manley, 
P. W., Leroy, C., Etienne, G., Roche, S., and Pasquet, J. M. (2008) Evidence that resistance to 
nilotinib may be due to BCR-ABL, Pgp, or Src kinase overexpression. Cancer Res 68, 9809-
9816 
54. 
Hebron, M. L., Lonskaya, I., Olopade, P., Selby, S. T., Pagan, F., and Moussa, C. E. (2014) 
Tyrosine Kinase Inhibition Regulates Early Systemic Immune Changes and Modulates the 
Neuroimmune Response in alpha-Synucleinopathy. J Clin Cell Immunol 5, 259 
55. 
Wenqiang, C., Lonskaya, I., Hebron, M. L., Ibrahim, Z., Olszewski, R. T., Neale, J. H., and 
Moussa, C. E. (2014) Parkin-mediated reduction of nuclear and soluble TDP-43 reverses 
behavioral decline in symptomatic mice. Hum Mol Genet 23, 4960-4969 
56. 
Hebron, M., Lonskaya, I., and Moussa, C. E. (2013) Nilotinib reverses loss of dopamine neurons 
and improves motor behavior via autophagic degradation of α-synuclein in Parkinson’s disease 
models. Hum. Mol. Genet. 22, 3315-3328 
57. 
Moussa, C. E. (2015) Parkin Is Dispensable for Mitochondrial Function, but Its Ubiquitin Ligase 
Activity Is Critical for Macroautophagy and Neurotransmitters: Therapeutic Potential beyond 
Parkinson's Disease. Neuro-degenerative diseases  
58. 
Lonskaya, I., Hebron, M. L., Selby, S. T., Turner, R. S., and Moussa, C. E. (2015) Nilotinib and 
bosutinib modulate pre-plaque alterations of blood immune markers and neuro-inflammation in 
Alzheimer's disease models. Neuroscience 304, 316-327 
59. 
Pagan, F., Hebron, M., Valadez, E. H., Torres-Yaghi, Y., Huang, X., Mills, R. R., Wilmarth, B. 
M., Howard, H., Dunn, C., Carlson, A., Lawler, A., Rogers, S. L., Falconer, R. A., Ahn, J., Li, Z., 
and Moussa, C. (2016) Nilotinib Effects in Parkinson's disease and Dementia with Lewy bodies. 
J Parkinsons Dis 6, 503-517 
60. 
Karuppagounder, S. S., Brahmachari, S., Lee, Y., Dawson, V. L., Dawson, T. M., and Ko, H. S. 
(2014) The c-Abl inhibitor, nilotinib, protects dopaminergic neurons in a preclinical animal model 
of Parkinson's disease. Scientific reports 4, 4874 
61. 
Mahul-Mellier, A. L., Fauvet, B., Gysbers, A., Dikiy, I., Oueslati, A., Georgeon, S., Lamontanara, 
A. J., Bisquertt, A., Eliezer, D., Masliah, E., Halliday, G., Hantschel, O., and Lashuel, H. A. (2014) 
c-Abl phosphorylates alpha-synuclein and regulates its degradation: implication for alpha-
synuclein clearance and contribution to the pathogenesis of Parkinson's disease. Human 
molecular genetics 23, 2858-2879 
62. 
Pagan, F. L., Hebron, M. L., Wilmarth, B., Torres-Yaghi, Y., Lawler, A., Mundel, E. E., Yusuf, N., 
Starr, N. J., Anjum, M., Arellano, J., Howard, H. H., Shi, W., Mulki, S., Kurd-Misto, T., Matar, S., 
Liu, X., Ahn, J., and Moussa, C. (2019) Nilotinib Effects on Safety, Tolerability, and Potential 
Biomarkers in Parkinson Disease: A Phase 2 Randomized Clinical Trial. JAMA Neurol  
63. 
Pagan, F. L., Hebron, M. L., Wilmarth, B., Torres-Yaghi, Y., Lawler, A., Mundel, E. E., Yusuf, N., 
Starr, N. J., Arellano, J., Howard, H. H., Peyton, M., Matar, S., Liu, X., Fowler, A. J., Schwartz, 
S. L., Ahn, J., and Moussa, C. (2019) Pharmacokinetics and pharmacodynamics of a single dose 
Nilotinib in individuals with Parkinson's disease. Pharmacol Res Perspect 7, e00470 
64. 
Turner, R. S., Hebron, M. L., Lawler, A., Mundel, E. E., Yusuf, N., Starr, J. N., Anjum, M., Pagan, 
F., Torres-Yaghi, Y., Shi, W., Mulki, S., Ferrante, D., Matar, S., Liu, X., Esposito, G., Berkowitz, 
F., Jiang, X., Ahn, J., and Moussa, C. (2020) Nilotinib Effects on Safety, Tolerability, and 
Biomarkers in Alzheimer's Disease. Ann Neurol  
65. 
Mollenhauer, B., Locascio, J. J., Schulz-Schaeffer, W., Sixel-Doring, F., Trenkwalder, C., and 
Schlossmacher, M. G. (2011) alpha-Synuclein and tau concentrations in cerebrospinal fluid of 
patients presenting with parkinsonism: a cohort study. Lancet Neurol 10, 230-240 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
69 
 
66. 
Parnetti, L., Chiasserini, D., Bellomo, G., Giannandrea, D., De Carlo, C., Qureshi, M. M., Ardah, 
M. T., Varghese, S., Bonanni, L., Borroni, B., Tambasco, N., Eusebi, P., Rossi, A., Onofrj, M., 
Padovani, A., Calabresi, P., and El-Agnaf, O. (2011) Cerebrospinal fluid Tau/alpha-synuclein 
ratio in Parkinson's disease and degenerative dementias. Mov Disord 26, 1428-1435 
67. 
Tateno, F., Sakakibara, R., Kawai, T., Kishi, M., and Murano, T. (2012) Alpha-synuclein in the 
cerebrospinal fluid differentiates synucleinopathies (Parkinson Disease, dementia with Lewy 
bodies, multiple system atrophy) from Alzheimer disease. Alzheimer Dis Assoc Disord 26, 213-
216 
68. 
Kantarjian, H., Sawyers, C., Hochhaus, A., Guilhot, F., Schiffer, C., Gambacorti-Passerini, C., 
Niederwieser, D., Resta, D., Capdeville, R., Zoellner, U., Talpaz, M., Druker, B., Goldman, J., 
O'Brien, S. G., Russell, N., Fischer, T., Ottmann, O., Cony-Makhoul, P., Facon, T., Stone, R., 
Miller, C., Tallman, M., Brown, R., Schuster, M., Loughran, T., Gratwohl, A., Mandelli, F., Saglio, 
G., Lazzarino, M., Russo, D., Baccarani, M., and Morra, E. (2002) Hematologic and cytogenetic 
responses to imatinib mesylate in chronic myelogenous leukemia. N Engl J Med 346, 645-652 
69. 
Qiu, Z., Cang, Y., and Goff, S. P. (2010) c-Abl tyrosine kinase regulates cardiac growth and 
development. Proc Natl Acad Sci U S A 107, 1136-1141 
70. 
Li, B., Boast, S., de los Santos, K., Schieren, I., Quiroz, M., Teitelbaum, S. L., Tondravi, M. M., 
and Goff, S. P. (2000) Mice deficient in Abl are osteoporotic and have defects in osteoblast 
maturation. Nat Genet 24, 304-308 
71. 
Schwartzberg, P. L., Stall, A. M., Hardin, J. D., Bowdish, K. S., Humaran, T., Boast, S., Harbison, 
M. L., Robertson, E. J., and Goff, S. P. (1991) Mice homozygous for the ablm1 mutation show 
poor viability and depletion of selected B and T cell populations. Cell 65, 1165-1175 
72. 
Tybulewicz, V. L., Crawford, C. E., Jackson, P. K., Bronson, R. T., and Mulligan, R. C. (1991) 
Neonatal lethality and lymphopenia in mice with a homozygous disruption of the c-abl proto-
oncogene. Cell 65, 1153-1163 
73. 
Bakhsheshian, J., Wei, B. R., Chang, K. E., Shukla, S., Ambudkar, S. V., Simpson, R. M., 
Gottesman, M. M., and Hall, M. D. (2013) Bioluminescent imaging of drug efflux at the blood-
brain barrier mediated by the transporter ABCG2. Proceedings of the National Academy of 
Sciences of the United States of America 110, 20801-20806 
74. 
Giasson, B. I., Duda, J. E., Quinn, S. M., Zhang, B., Trojanowski, J. Q., and Lee, V. M. (2002) 
Neuronal alpha-synucleinopathy with severe movement disorder in mice expressing A53T 
human alpha-synuclein. Neuron 34, 521-533 
75. 
Lonskaya, I., Shekoyan, A. R., Hebron, M. L., Desforges, N., Algarzae, N. K., and Moussa, C. 
E. (2012) Diminished Parkin Solubility and Co-Localization with Intraneuronal Amyloid-beta are 
Associated with Autophagic Defects in Alzheimer's Disease. J Alzheimers Dis  
76. 
Majbour, N. K., Vaikath, N. N., Eusebi, P., Chiasserini, D., Ardah, M., Varghese, S., Haque, M. 
E., Tokuda, T., Auinger, P., Calabresi, P., Parnetti, L., and El-Agnaf, O. M. (2016) Longitudinal 
changes in CSF alpha-synuclein species reflect Parkinson's disease progression. Mov Disord  
77. 
Majbour, N. K., Vaikath, N. N., van Dijk, K. D., Ardah, M. T., Varghese, S., Vesterager, L. B., 
Montezinho, L. P., Poole, S., Safieh-Garabedian, B., Tokuda, T., Teunissen, C. E., Berendse, 
H. W., van de Berg, W. D., and El-Agnaf, O. M. (2016) Oligomeric and phosphorylated alpha-
synuclein as potential CSF biomarkers for Parkinson's disease. Mol Neurodegener 11, 7 
78. 
van Erp, N. P., Gelderblom, H., and Guchelaar, H. J. (2009) Clinical pharmacokinetics of tyrosine 
kinase inhibitors. Cancer Treat Rev 35, 692-706 
79. 
Kantarjian, H. M., Giles, F., Gattermann, N., Bhalla, K., Alimena, G., Palandri, F., Ossenkoppele, 
G. J., Nicolini, F. E., O'Brien, S. G., Litzow, M., Bhatia, R., Cervantes, F., Haque, A., Shou, Y., 
Resta, D. J., Weitzman, A., Hochhaus, A., and le Coutre, P. (2007) Nilotinib (formerly AMN107), 
a highly selective BCR-ABL tyrosine kinase inhibitor, is effective in patients with Philadelphia 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
70 
 
chromosome-positive chronic myelogenous leukemia in chronic phase following imatinib 
resistance and intolerance. Blood 110, 3540-3546 
80. 
de Lavallade, H., Apperley, J. F., Khorashad, J. S., Milojkovic, D., Reid, A. G., Bua, M., Szydlo, 
R., Olavarria, E., Kaeda, J., Goldman, J. M., and Marin, D. (2008) Imatinib for newly diagnosed 
patients with chronic myeloid leukemia: incidence of sustained responses in an intention-to-treat 
analysis. J Clin Oncol 26, 3358-3363 
81. 
Polymenidou, M., and Cleveland, D. W. (2012) Prion-like spread of protein aggregates in 
neurodegeneration. The Journal of experimental medicine 209, 889-893 
82. 
Ciechanover, A., and Kwon, Y. T. (2015) Degradation of misfolded proteins in 
neurodegenerative diseases: therapeutic targets and strategies. Experimental & molecular 
medicine 47, e147 
83. 
Mellman, I. (1996) Endocytosis and molecular sorting. Annual review of cell and developmental 
biology 12, 575-625 
84. 
Jerram, A. H., Smith, P. F., and Darlington, C. L. (1996) A dose-response analysis of the 
behavioral effects of (+)MK-801 in guinea pig: comparison with CPP. Pharmacology, 
biochemistry, and behavior 53, 799-807 
85. 
Luzio, J. P., Rous, B. A., Bright, N. A., Pryor, P. R., Mullock, B. M., and Piper, R. C. (2000) 
Lysosome-endosome fusion and lysosome biogenesis. Journal of cell science 113 ( Pt 9), 1515-
1524 
86. 
Kettenmann, H., Hanisch, U. K., Noda, M., and Verkhratsky, A. (2011) Physiology of microglia. 
Physiol Rev 91, 461-553 
87. 
Yang, Z. J., Chee, C. E., Huang, S., and Sinicrope, F. A. (2011) The role of autophagy in cancer: 
therapeutic implications. Mol Cancer Ther 10, 1533-1541 
88. 
Macintosh, R. L., and Ryan, K. M. (2013) Autophagy in tumour cell death. Seminars in cancer 
biology 23, 344-351 
89. 
Crighton, D., Wilkinson, S., O'Prey, J., Syed, N., Smith, P., Harrison, P. R., Gasco, M., Garrone, 
O., Crook, T., and Ryan, K. M. (2006) DRAM, a p53-induced modulator of autophagy, is critical 
for apoptosis. Cell 126, 121-134 
90. 
Yee, K. S., Wilkinson, S., James, J., Ryan, K. M., and Vousden, K. H. (2009) PUMA- and Bax-
induced autophagy contributes to apoptosis. Cell death and differentiation 16, 1135-1145 
91. 
Nakada, M., Kita, D., Teng, L., Pyko, I. V., Watanabe, T., Hayashi, Y., and Hamada, J. (2013) 
Receptor tyrosine kinases: principles and functions in glioma invasion. Advances in experimental 
medicine and biology 986, 143-170 
92. 
Cadena, D. L., and Gill, G. N. (1992) Receptor tyrosine kinases. FASEB J 6, 2332-2337 
93. 
Jabbour, E., Kantarjian, H., and Cortes, J. (2015) Use of second- and third-generation tyrosine 
kinase inhibitors in the treatment of chronic myeloid leukemia: an evolving treatment paradigm. 
Clin Lymphoma Myeloma Leuk 15, 323-334 
94. 
Holyoake, T. L., and Helgason, G. V. (2015) Do we need more drugs for chronic myeloid 
leukemia? Immunol Rev 263, 106-123 
95. 
Zhang, H., and Li, S. (2013) Molecular mechanisms for survival regulation of chronic myeloid 
leukemia stem cells. Protein Cell 4, 186-196 
96. 
Hong, Z., Shi, M., Chung, K. A., Quinn, J. F., Peskind, E. R., Galasko, D., Jankovic, J., Zabetian, 
C. P., Leverenz, J. B., Baird, G., Montine, T. J., Hancock, A. M., Hwang, H., Pan, C., Bradner, 
J., Kang, U. J., Jensen, P. H., and Zhang, J. (2010) DJ-1 and alpha-synuclein in human 
cerebrospinal fluid as biomarkers of Parkinson's disease. Brain 133, 713-726 
97. 
Ohrfelt, A., Grognet, P., Andreasen, N., Wallin, A., Vanmechelen, E., Blennow, K., and 
Zetterberg, H. (2009) Cerebrospinal fluid alpha-synuclein in neurodegenerative disorders-a 
marker of synapse loss? Neurosci Lett 450, 332-335 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
71 
 
98. 
Kasuga, K., Tokutake, T., Ishikawa, A., Uchiyama, T., Tokuda, T., Onodera, O., Nishizawa, M., 
and Ikeuchi, T. (2010) Differential levels of alpha-synuclein, beta-amyloid42 and tau in CSF 
between patients with dementia with Lewy bodies and Alzheimer's disease. J Neurol Neurosurg 
Psychiatry 81, 608-610 
99. 
Mollenhauer, B., Trautmann, E., Otte, B., Ng, J., Spreer, A., Lange, P., Sixel-Doring, F., Hakimi, 
M., Vonsattel, J. P., Nussbaum, R., Trenkwalder, C., and Schlossmacher, M. G. (2012) alpha-
Synuclein in human cerebrospinal fluid is principally derived from neurons of the central nervous 
system. Journal of neural transmission 119, 739-746 
100. Mollenhauer, B., Cullen, V., Kahn, I., Krastins, B., Outeiro, T. F., Pepivani, I., Ng, J., Schulz-
Schaeffer, W., Kretzschmar, H. A., McLean, P. J., Trenkwalder, C., Sarracino, D. A., Vonsattel, 
J. P., Locascio, J. J., El-Agnaf, O. M., and Schlossmacher, M. G. (2008) Direct quantification of 
CSF 
alpha-synuclein 
by 
ELISA 
and first 
cross-sectional study 
in 
patients with 
neurodegeneration. Experimental neurology 213, 315-325 
101. Mollenhauer, B., Trautmann, E., Taylor, P., Manninger, P., Sixel-Doring, F., Ebentheuer, J., 
Trenkwalder, C., and Schlossmacher, M. G. (2013) Total CSF alpha-synuclein is lower in de 
novo Parkinson patients than in healthy subjects. Neurosci Lett 532, 44-48 
102. Lleo, A., Cavedo, E., Parnetti, L., Vanderstichele, H., Herukka, S. K., Andreasen, N., Ghidoni, 
R., Lewczuk, P., Jeromin, A., Winblad, B., Tsolaki, M., Mroczko, B., Visser, P. J., Santana, I., 
Svenningsson, P., Blennow, K., Aarsland, D., Molinuevo, J. L., Zetterberg, H., and Mollenhauer, 
B. (2015) Cerebrospinal fluid biomarkers in trials for Alzheimer and Parkinson diseases. Nature 
reviews. Neurology 11, 41-55 
103. Kang, J. H., Irwin, D. J., Chen-Plotkin, A. S., Siderowf, A., Caspell, C., Coffey, C. S., Waligorska, 
T., Taylor, P., Pan, S., Frasier, M., Marek, K., Kieburtz, K., Jennings, D., Simuni, T., Tanner, C. 
M., Singleton, A., Toga, A. W., Chowdhury, S., Mollenhauer, B., Trojanowski, J. Q., Shaw, L. M., 
and Parkinson's Progression Markers, I. (2013) Association of cerebrospinal fluid beta-amyloid 
1-42, T-tau, P-tau181, and alpha-synuclein levels with clinical features of drug-naive patients 
with early Parkinson disease. JAMA Neurol 70, 1277-1287 
104. Benitez, B. A., Cruchaga, C., and United States-Spain Parkinson's Disease Research, G. (2013) 
TREM2 and neurodegenerative disease. N Engl J Med 369, 1567-1568 
105. Jonsson, T., and Stefansson, K. (2013) TREM2 and neurodegenerative disease. N Engl J Med 
369, 1568-1569 
106. Rayaprolu, S., Mullen, B., Baker, M., Lynch, T., Finger, E., Seeley, W. W., Hatanpaa, K. J., 
Lomen-Hoerth, C., Kertesz, A., Bigio, E. H., Lippa, C., Josephs, K. A., Knopman, D. S., White, 
C. L., 3rd, Caselli, R., Mackenzie, I. R., Miller, B. L., Boczarska-Jedynak, M., Opala, G., 
Krygowska-Wajs, A., Barcikowska, M., Younkin, S. G., Petersen, R. C., Ertekin-Taner, N., Uitti, 
R. J., Meschia, J. F., Boylan, K. B., Boeve, B. F., Graff-Radford, N. R., Wszolek, Z. K., Dickson, 
D. W., Rademakers, R., and Ross, O. A. (2013) TREM2 in neurodegeneration: evidence for 
association of the p.R47H variant with frontotemporal dementia and Parkinson's disease. 
Molecular neurodegeneration 8, 19 
107. Wu, D. C., Teismann, P., Tieu, K., Vila, M., Jackson-Lewis, V., Ischiropoulos, H., and 
Przedborski, S. (2003) NADPH oxidase mediates oxidative stress in the 1-methyl-4-phenyl-
1,2,3,6-tetrahydropyridine model of Parkinson's disease. Proceedings of the National Academy 
of Sciences of the United States of America 100, 6145-6150 
108. Hu, X., Zhang, D., Pang, H., Caudle, W. M., Li, Y., Gao, H., Liu, Y., Qian, L., Wilson, B., Di 
Monte, D. A., Ali, S. F., Zhang, J., Block, M. L., and Hong, J. S. (2008) Macrophage antigen 
complex-1 mediates reactive microgliosis and progressive dopaminergic neurodegeneration in 
the MPTP model of Parkinson's disease. J Immunol 181, 7194-7204 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
72 
 
109. Sims, R., van der Lee, S. J., Naj, A. C., Bellenguez, C., Badarinarayan, N., Jakobsdottir, J., 
Kunkle, B. W., Boland, A., Raybould, R., Bis, J. C., Martin, E. R., Grenier-Boley, B., Heilmann-
Heimbach, S., Chouraki, V., Kuzma, A. B., Sleegers, K., Vronskaya, M., Ruiz, A., Graham, R. 
R., Olaso, R., Hoffmann, P., Grove, M. L., Vardarajan, B. N., Hiltunen, M., Nothen, M. M., White, 
C. C., Hamilton-Nelson, K. L., Epelbaum, J., Maier, W., Choi, S. H., Beecham, G. W., Dulary, 
C., Herms, S., Smith, A. V., Funk, C. C., Derbois, C., Forstner, A. J., Ahmad, S., Li, H., Bacq, 
D., Harold, D., Satizabal, C. L., Valladares, O., Squassina, A., Thomas, R., Brody, J. A., Qu, L., 
Sanchez-Juan, P., Morgan, T., Wolters, F. J., Zhao, Y., Garcia, F. S., Denning, N., Fornage, M., 
Malamon, J., Naranjo, M. C. D., Majounie, E., Mosley, T. H., Dombroski, B., Wallon, D., Lupton, 
M. K., Dupuis, J., Whitehead, P., Fratiglioni, L., Medway, C., Jian, X., Mukherjee, S., Keller, L., 
Brown, K., Lin, H., Cantwell, L. B., Panza, F., McGuinness, B., Moreno-Grau, S., Burgess, J. D., 
Solfrizzi, V., Proitsi, P., Adams, H. H., Allen, M., Seripa, D., Pastor, P., Cupples, L. A., Price, N. 
D., Hannequin, D., Frank-Garcia, A., Levy, D., Chakrabarty, P., Caffarra, P., Giegling, I., Beiser, 
A. S., Giedraitis, V., Hampel, H., Garcia, M. E., Wang, X., Lannfelt, L., Mecocci, P., Eiriksdottir, 
G., Crane, P. K., Pasquier, F., Boccardi, V., Henandez, I., Barber, R. C., Scherer, M., Tarraga, 
L., Adams, P. M., Leber, M., Chen, Y., Albert, M. S., Riedel-Heller, S., Emilsson, V., Beekly, D., 
Braae, A., Schmidt, R., Blacker, D., Masullo, C., Schmidt, H., Doody, R. S., Spalletta, G., 
Longstreth, W. T., Jr., Fairchild, T. J., Bossu, P., Lopez, O. L., Frosch, M. P., Sacchinelli, E., 
Ghetti, B., Yang, Q., Huebinger, R. M., Jessen, F., Li, S., Kamboh, M. I., Morris, J., Sotolongo-
Grau, O., Katz, M. J., Corcoran, C., Dunstan, M., Braddel, A., Thomas, C., Meggy, A., Marshall, 
R., Gerrish, A., Chapman, J., Aguilar, M., Taylor, S., Hill, M., Fairen, M. D., Hodges, A., Vellas, 
B., Soininen, H., Kloszewska, I., Daniilidou, M., Uphill, J., Patel, Y., Hughes, J. T., Lord, J., 
Turton, J., Hartmann, A. M., Cecchetti, R., Fenoglio, C., Serpente, M., Arcaro, M., Caltagirone, 
C., Orfei, M. D., Ciaramella, A., Pichler, S., Mayhaus, M., Gu, W., Lleo, A., Fortea, J., Blesa, R., 
Barber, I. S., Brookes, K., Cupidi, C., Maletta, R. G., Carrell, D., Sorbi, S., Moebus, S., Urbano, 
M., Pilotto, A., Kornhuber, J., Bosco, P., Todd, S., Craig, D., Johnston, J., Gill, M., Lawlor, B., 
Lynch, A., Fox, N. C., Hardy, J., Consortium, A., Albin, R. L., Apostolova, L. G., Arnold, S. E., 
Asthana, S., Atwood, C. S., Baldwin, C. T., Barnes, L. L., Barral, S., Beach, T. G., Becker, J. T., 
Bigio, E. H., Bird, T. D., Boeve, B. F., Bowen, J. D., Boxer, A., Burke, J. R., Burns, J. M., 
Buxbaum, J. D., Cairns, N. J., Cao, C., Carlson, C. S., Carlsson, C. M., Carney, R. M., 
Carrasquillo, M. M., Carroll, S. L., Diaz, C. C., Chui, H. C., Clark, D. G., Cribbs, D. H., Crocco, 
E. A., DeCarli, C., Dick, M., Duara, R., Evans, D. A., Faber, K. M., Fallon, K. B., Fardo, D. W., 
Farlow, M. R., Ferris, S., Foroud, T. M., Galasko, D. R., Gearing, M., Geschwind, D. H., Gilbert, 
J. R., Graff-Radford, N. R., Green, R. C., Growdon, J. H., Hamilton, R. L., Harrell, L. E., Honig, 
L. S., Huentelman, M. J., Hulette, C. M., Hyman, B. T., Jarvik, G. P., Abner, E., Jin, L. W., Jun, 
G., Karydas, A., Kaye, J. A., Kim, R., Kowall, N. W., Kramer, J. H., LaFerla, F. M., Lah, J. J., 
Leverenz, J. B., Levey, A. I., Li, G., Lieberman, A. P., Lunetta, K. L., Lyketsos, C. G., Marson, 
D. C., Martiniuk, F., Mash, D. C., Masliah, E., McCormick, W. C., McCurry, S. M., McDavid, A. 
N., McKee, A. C., Mesulam, M., Miller, B. L., Miller, C. A., Miller, J. W., Morris, J. C., Murrell, J. 
R., Myers, A. J., O'Bryant, S., Olichney, J. M., Pankratz, V. S., Parisi, J. E., Paulson, H. L., Perry, 
W., Peskind, E., Pierce, A., Poon, W. W., Potter, H., Quinn, J. F., Raj, A., Raskind, M., Reisberg, 
B., Reitz, C., Ringman, J. M., Roberson, E. D., Rogaeva, E., Rosen, H. J., Rosenberg, R. N., 
Sager, M. A., Saykin, A. J., Schneider, J. A., Schneider, L. S., Seeley, W. W., Smith, A. G., 
Sonnen, J. A., Spina, S., Stern, R. A., Swerdlow, R. H., Tanzi, R. E., Thornton-Wells, T. A., 
Trojanowski, J. Q., Troncoso, J. C., Van Deerlin, V. M., Van Eldik, L. J., Vinters, H. V., Vonsattel, 
J. P., Weintraub, S., Welsh-Bohmer, K. A., Wilhelmsen, K. C., Williamson, J., Wingo, T. S., 
Woltjer, R. L., Wright, C. B., Yu, C. E., Yu, L., Garzia, F., Golamaully, F., Septier, G., 
Engelborghs, S., Vandenberghe, R., De Deyn, P. P., Fernadez, C. M., Benito, Y. A., Thonberg, 

 Georgetown University 
Medical Center  
[NILOTINIB IN DEMENTIA WITH LEWY BODIES (DLB) –
CLINICAL TRIAL PROTOCOL] 
 
73 
 
H., Forsell, C., Lilius, L., Kinhult-Stahlbom, A., Kilander, L., Brundin, R., Concari, L., Helisalmi, 
S., Koivisto, A. M., Haapasalo, A., Dermecourt, V., Fievet, N., Hanon, O., Dufouil, C., Brice, A., 
Ritchie, K., Dubois, B., Himali, J. J., Keene, C. D., Tschanz, J., Fitzpatrick, A. L., Kukull, W. A., 
Norton, M., Aspelund, T., Larson, E. B., Munger, R., Rotter, J. I., Lipton, R. B., Bullido, M. J., 
Hofman, A., Montine, T. J., Coto, E., Boerwinkle, E., Petersen, R. C., Alvarez, V., Rivadeneira, 
F., Reiman, E. M., Gallo, M., O'Donnell, C. J., Reisch, J. S., Bruni, A. C., Royall, D. R., Dichgans, 
M., Sano, M., Galimberti, D., St George-Hyslop, P., Scarpini, E., Tsuang, D. W., Mancuso, M., 
Bonuccelli, U., Winslow, A. R., Daniele, A., Wu, C. K., Gerad/Perades, C. A. E., Peters, O., 
Nacmias, B., Riemenschneider, M., Heun, R., Brayne, C., Rubinsztein, D. C., Bras, J., Guerreiro, 
R., Al-Chalabi, A., Shaw, C. E., Collinge, J., Mann, D., Tsolaki, M., Clarimon, J., Sussams, R., 
Lovestone, S., O'Donovan, M. C., Owen, M. J., Behrens, T. W., Mead, S., Goate, A. M., 
Uitterlinden, A. G., Holmes, C., Cruchaga, C., Ingelsson, M., Bennett, D. A., Powell, J., Golde, 
T. E., Graff, C., De Jager, P. L., Morgan, K., Ertekin-Taner, N., Combarros, O., Psaty, B. M., 
Passmore, P., Younkin, S. G., Berr, C., Gudnason, V., Rujescu, D., Dickson, D. W., Dartigues, 
J. F., DeStefano, A. L., Ortega-Cubero, S., Hakonarson, H., Campion, D., Boada, M., Kauwe, J. 
K., Farrer, L. A., Van Broeckhoven, C., Ikram, M. A., Jones, L., Haines, J. L., Tzourio, C., Launer, 
L. J., Escott-Price, V., Mayeux, R., Deleuze, J. F., Amin, N., Holmans, P. A., Pericak-Vance, M. 
A., Amouyel, P., van Duijn, C. M., Ramirez, A., Wang, L. S., Lambert, J. C., Seshadri, S., 
Williams, J., and Schellenberg, G. D. (2017) Rare coding variants in PLCG2, ABI3, and TREM2 
implicate microglial-mediated innate immunity in Alzheimer's disease. Nature genetics 49, 1373-
1384 
110. Garcia-Reitboeck, P., Phillips, A., Piers, T. M., Villegas-Llerena, C., Butler, M., Mallach, A., 
Rodrigues, C., Arber, C. E., Heslegrave, A., Zetterberg, H., Neumann, H., Neame, S., Houlden, 
H., Hardy, J., and Pocock, J. M. (2018) Human Induced Pluripotent Stem Cell-Derived Microglia-
Like Cells Harboring TREM2 Missense Mutations Show Specific Deficits in Phagocytosis. Cell 
reports 24, 2300-2311 
111. Hong, S., and Stevens, B. (2017) TREM2: Keeping Microglia Fit during Good Times and Bad. 
Cell Metab 26, 590-591 
112. Sasaki, A. (2017) Microglia and brain macrophages: An update. Neuropathology : official journal 
of the Japanese Society of Neuropathology 37, 452-464 
113. Zubenko, G. S., Marquis, J. K., Volicer, L., Direnfeld, L. K., Langlais, P. J., and Nixon, R. A. 
(1986) Cerebrospinal fluid levels of angiotensin-converting enzyme, acetylcholinesterase, and 
dopamine metabolites in dementia associated with Alzheimer's disease and Parkinson's 
disease: a correlative study. Biol Psychiatry 21, 1365-1381 
114. Goldstein, D. S., Holmes, C., and Sharabi, Y. (2012) Cerebrospinal fluid biomarkers of central 
catecholamine deficiency in Parkinson's disease and other synucleinopathies. Brain 135, 1900-
1913