An Evaluation of the Safety of Intravenous (IV) Tc 99m Tilmanocept and a Comparison of Localization to Tc 99m Sulfur Colloid in Subjects With Nonalcoholic Steatohepatitis (NASH) and Healthy Controls (HC)
Phase: PHASE1
Conditions: Nonalcoholic Steatohepatitis, NASH - Nonalcoholic Steatohepatitis
Sponsor: Navidea Biopharmaceuticals

 
 
Clinical Study Protocol: NAV3-30 
Study Title: 
An Evaluation of the Safety of Intravenous (IV) Tc 99m Tilmanocept 
and a Comparison of Localization to Tc 99m Sulfur Colloid by Planar 
and 
SPECT/CT 
Imaging 
in 
Subjects 
with 
Nonalcoholic 
Steatohepatitis (NASH) and Healthy Controls (HC) 
Study Number: 
NAV3-30 
Study Phase: 
1 
Product Name: 
Technetium Tc 99m tilmanocept 
IND Number: 
132943 
No. of Subjects 
Up to 12 
Investigator(s): 
Multicenter 
Sponsor: 
Navidea Biopharmaceuticals, Inc 
4995 Bradenton Ave, Suite 240 
Dublin, OH 43017 
Sponsor Contact: 
Bonnie Abbruzzese, MS, RDN, CCRA 
Senior Director, Clinical Research 
+1 614-822-2327 
Medical Monitor: 
Michael Blue, MD, FACEP 
Senior Medical Director 
+1 614-571-4313 
 
Date 
Original Protocol: 06 October 2017  
Amendment 1 
23 April 2018 
 
Confidentiality Statement 
This document and its contents are the property of and confidential to Navidea 
Biopharmaceuticals. Any unauthorized copying or use of this document is prohibited. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
Page 2 of 71 
 
TABLE OF CONTENTS 
SYNOPSIS .................................................................................................................................6 
LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS ..........................................14 
STUDY ADMINISTRATIVE STRUCTURE .........................................................................18 
1 
INTRODUCTION ...........................................................................................................19 
1.1 
Background ............................................................................................................19 
1.2 
Liver Anatomy, Function, and Role of Kupffer cells ............................................21 
1.3 
NASH Diagnosis and Other Imaging Modalities ..................................................22 
1.4 
Preliminary Data ....................................................................................................24 
1.4.1 
99mTc-tilmanocept (Lymphoseek®) ..............................................................24 
1.4.2 
Transitioning from Subcutaneous and Intradermal Administration of 
99mTc-Tilmanocept to IV Administration in Diagnostic Imaging 
indications ....................................................................................................26 
1.4.3 
Nonclinical Evaluations – IV Administration..............................................26 
1.4.4 
Clinical Evaluations – IV Administration....................................................30 
1.4.5 
Clinical Safety using Intradermal, Subcutaneous, and Peritumoral 
Injections ......................................................................................................32 
2 
STUDY OBJECTIVES ....................................................................................................33 
2.1 
Primary Objective(s) ..............................................................................................33 
2.2 
Secondary Objective(s) ..........................................................................................33 
2.3 
Exploratory ............................................................................................................33 
3 
OVERVIEW OF METHODOLOGY AND DESIGN .....................................................34 
3.1 
Overall Study Design .............................................................................................34 
3.2 
Justification for Study Design and Population.......................................................34 
3.3 
Protocol Adherence ................................................................................................34 
3.4 
Study Duration .......................................................................................................35 
4 
STUDY POPULATION ..................................................................................................36 
4.1 
Eligibility ...............................................................................................................36 
4.1.1 
Inclusion Criteria .........................................................................................36 
4.1.2 
Exclusion Criteria ........................................................................................36 
4.2 
Recruitment ............................................................................................................37 
4.3 
Withdrawal .............................................................................................................37 
4.4 
Replacement ...........................................................................................................38 
4.5 
Subject Identification .............................................................................................38 
5 
INVESTIGATIONAL PRODUCT ..................................................................................39 
5.1 
Identification of Investigational Product ...............................................................39 
5.2 
Investigational Product Dosage and Administration .............................................39 
5.3 
Treatment Assignment ...........................................................................................40 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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5.4 
Packaging and Labeling .........................................................................................41 
5.5 
Drug Logistics and Investigational Product Accountability ..................................41 
6 
THERAPIES OTHER THAN INVESTIGATIONAL PRODUCT .................................42 
6.1 
Prior and Concomitant Therapy .............................................................................42 
6.2 
Post-Study Therapy ................................................................................................42 
7 
STUDY PROCEDURES .................................................................................................43 
7.1 
Schedule of Evaluations .........................................................................................43 
7.2 
Visit Description ....................................................................................................43 
7.2.1 
Visit 1, Screening (Day -45 to Day -1) ........................................................43 
7.2.2 
Visit 2 (Day 0) Unfiltered Tc 99m Sulfur Colloid Administration .............44 
7.2.2.1 
Before Administration of Unfiltered Tc 99m Sulfur Colloid ............44 
7.2.2.2 
Administration of Unfiltered Tc 99m Sulfur Colloid ........................44 
7.2.2.3 
After Administration of Unfiltered Tc 99m Sulfur Colloid ...............44 
7.2.3 
Visit 3 (Day 3) Tc 99m Tilmanocept Administration ..................................45 
7.2.3.1 
Before Administration of Tc 99m Tilmanocept.................................45 
7.2.3.2 
Administration of Tc 99m Tilmanocept ............................................45 
7.2.3.3 
After Administration of Tc 99m Tilmanocept ...................................45 
7.2.4 
Safety Follow Up Telephone Call (Day 8 ± 3) ............................................46 
8 
PROCEDURES AND VARIABLES ..............................................................................47 
8.1 
Population Characteristics .....................................................................................47 
8.1.1 
Demographic and Other Baseline Characteristics .......................................47 
8.1.2 
Medical and Surgical History ......................................................................47 
8.1.3 
Prior and Concomitant Medication ..............................................................47 
8.2 
SPECT/SPECT-CT Image Acquisition .................................................................47 
8.3 
Pharmacokinetics ...................................................................................................48 
8.4 
Safety .....................................................................................................................49 
8.4.1 
Adverse Events ............................................................................................49 
8.4.1.1 
Definition of Adverse Event ..............................................................49 
8.4.1.2 
Categories for Adverse Event Assessment ........................................49 
8.4.1.3 
Assessments and Documentation of Adverse Events ........................51 
8.4.1.4 
Expected Adverse Events ..................................................................52 
8.4.1.5 
Serious Adverse Events .....................................................................52 
8.4.2 
Further Safety Assessments .........................................................................54 
8.4.2.1 
Physical Exam ....................................................................................54 
8.4.2.2 
Vital Signs ..........................................................................................54 
8.4.2.3 
Electrocardiogram ..............................................................................54 
8.4.2.4 
Clinical Laboratory Parameters for Screening and Safety .................55 
9 
STATISTICAL METHODS ............................................................................................57 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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9.1 
Randomization Methods ........................................................................................57 
9.2 
Safety Variables .....................................................................................................57 
9.3 
Efficacy Variables ..................................................................................................57 
9.4 
Sample Size Justification .......................................................................................57 
9.5 
Statistical Analyses ................................................................................................58 
9.5.1 
Analysis Populations ....................................................................................58 
9.5.2 
Analysis of Baseline and Demographic Characteristics ..............................58 
9.5.3 
Analysis of Efficacy Variables ....................................................................58 
9.5.4 
Analysis of Safety Variables ........................................................................59 
9.5.5 
Handling Missing Values .............................................................................59 
9.5.6 
Interim Analysis ...........................................................................................59 
10 DATA HANDLING AND QUALITY ASSURANCE ...................................................60 
10.1 
Data Recording ......................................................................................................60 
10.1.1 
CRF Design ..................................................................................................60 
10.2 
Monitoring .............................................................................................................60 
10.3 
Data Processing ......................................................................................................60 
10.4 
Auditing .................................................................................................................60 
10.5 
Archiving ...............................................................................................................61 
10.6 
Premature Termination of the Study ......................................................................61 
10.6.1 
Study as a Whole .........................................................................................62 
10.6.2 
Center ...........................................................................................................62 
10.6.3 
Study Participant ..........................................................................................62 
11 ETHICAL AND LEGAL ASPECTS...............................................................................63 
11.1 
Ethical and Legal Conduct of the Study ................................................................63 
11.2 
Subject Information and Consent...........................................................................63 
11.3 
Financing/Financial Disclosure .............................................................................64 
11.4 
Publication Policy ..................................................................................................64 
11.5 
Subject Injury .........................................................................................................64 
12 REFERENCE LIST .........................................................................................................65 
 
LIST OF IN-TEXT TABLES 
Table 1. 
The RA Dose Matrix of 99mTc-Tilmanocept administration ............................31 
Table 2. 
Clinical Laboratory Parameters ..........................................................................55 
Table 3 
Approximate Amount of Blood Withdrawn .......................................................56 
 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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LIST OF IN-TEXT FIGURES 
Figure 1. 
Nonalcoholic steatohepatitis (NASH) progression [7] .......................................20 
Figure 2. 
Liver anatomy and lobular zoning [18] ..............................................................22 
Figure 3. 
Involvement of the macrophages in the liver damage [15] .................................24 
Figure 4. 
Structure of Tc99m-tilmanocept (Lymphoseek®) ...............................................25 
Figure 5. 
Pre-operative SPECT-CT image of a breast cancer patient who had been 
injected peritumorally with 99mTc-tilmanocept ...................................................26 
Figure 6. 
IV Administered Tc 99m tilmanocept Localization to RA Joints ......................32 
Figure 7. 
Diagrmatic representation of anticipated tessellation .........................................58 
 
LIST OF APPENDICES 
Appendix 1 Schedule of Events ..............................................................................................68 
Appendix 2 Sponsor Signatures..............................................................................................70 
Appendix 3 Investigator’s Signature ......................................................................................71 
 
 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
Page 6 of 71 
 
SYNOPSIS 
Study title 
An Evaluation of the Safety of Intravenous (IV) Tc 99m 
Tilmanocept and a Comparison of Localization to Tc 99m 
Sulfur Colloid by Planar and SPECT/CT Imaging in Subjects 
with Nonalcoholic Steatohepatitis (NASH) and Healthy 
Controls (HC) 
Study phase 
Phase 1 
Study objective(s) 
Primary  
• Determine the safety and tolerability of Tc 99m 
tilmanocept in subjects with NASH.  
Secondary  
• Determine 
the 
three 
dimensional 
tessellation 
localization of Tc 99m tilmanocept by planar and 
SPECT 
(single 
photon 
emission 
computed 
tomography)/CT (Computerized x-ray Tomography) 
imaging in subjects with NASH and asymptomatic 
controls. 
• Determine 
the 
three 
dimensional 
tessellation 
localization of unfiltered Tc 99m sulfur colloid by 
planar and SPECT/CT imaging in subjects with NASH 
and asymptomatic controls. 
• Comparison 
of 
liver 
localization 
distribution 
tessellation pattern intensity between Tc 99m 
tilmanocept and unfiltered Tc 99m sulfur colloid.  
Exploratory 
• Concordance of intrahepatic localization tessellation 
discrimination of Tc 99m tilmanocept (intrahepatic 
disease heterogeneity) by SPECT/CT imaging and 
elastography  
• Concordance of intrahepatic localization tessellation 
discrimination of Tc 99m sulfur colloid (intrahepatic 
disease heterogeneity) by SPECT/CT imaging and 
elastography 
 
 
Duration of treatment 
Up to 57 days 
Study drug 
Technetium Tc 99m tilmanocept  
Unfiltered technetium Tc 99m sulfur colloid 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Dose(s) and Route of 
administration 
Route of Administration: All administrations of Tc 99m 
tilmanocept and unfiltered Tc 99m sulfur colloid will occur 
through an intravenous (IV) route of administration. A single 
syringe will be used for each product and administered as a 
slow push into the IV. At the completion of each 
administration, a 10 cc sterile normal saline flush will be 
administered. 
The preferred site of IV placement will be the left or right 
antecubital vein. 
Doses: On Day 0, subjects will receive an on label 8.0 mCi 
dose of unfiltered Tc 99m sulfur colloid via IV administration. 
On Day 3, subjects will receive 200 µg of Tc 99m tilmanocept 
at the specified radiolabel dose of Tc 99m via IV 
administration.  
The first two enrolled subjects will be NASH subjects who 
will receive an 8.0 mCi radiolabel dose of Tc 99m tilmanocept. 
A review meeting will be held following completion of the 
first two subjects to evaluate tilmanocept safety and dose 
performance.  
At the review: 
1. If it is determined that a lower radiolabel dose of 
tilmanocept may produce images of better quality,  
three additional NASH subjects and three HCs will be 
injected and imaged with 200 µg of 2.5 mCi Tc 99m 
tilmanocept. After these six subjects have completed 
the trial, no additional subjects will be enrolled.  
 
2. If the review shows no safety signals and acceptable 
quality of imaging, one additional NASH subject as 
well as three HCs will be injected and imaged with 
200 µg of 8.0 mCi Tc 99m tilmanocept. Once a total 
of three NASH subjects and three HCs have 
completed the trial, a second review meeting will be 
held to compare results from NASH subjects to those 
of the HCs. If it is determined at the second review 
meeting that a lower radiolabel dose of tilmanocept 
may produce additional clinically meaningful results, 
3 NASH subjects and 3 HCs will be dosed with 200 
µg of 2.5 mCi Tc 99m tilmanocept. If it is determined 
that additional doses will not provide clinically 
meaningful results, the trial will be concluded. 
 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Inclusion criteria 
ALL SUBJECTS: 
1. The subject has provided written informed consent with 
HIPAA 
(Health 
Information 
Portability 
and 
Accountability Act) authorization before the initiation 
of any study-related procedures. 
2. The subject is ≥18 years of age at the time of consent.  
3. The subject has a body mass index (BMI) between 18 
and 45. 
 
CONTROL SUBJECTS: 
4. The subject is deemed to be clinically free of any 
infectious/inflammatory disease(s) for at least 4 weeks 
prior to the consent date. 
5. The subject has not taken any antibiotics for at least 4 
weeks prior to the consent date.  
 
NASH SUBJECTS: 
4. The subject has biopsy-confirmed NASH within 12 
months prior to enrollment.  
5. The subject has a Nonalcoholic Fatty Liver Disease 
(NAFLD) Activity Score (NAS) of ≥ 4, with a score of 
at least 1 for each steatosis, lobular inflammation, and 
hepatocyte ballooning.  
6. The subject has fibrosis staging of F3-F4.  
Exclusion criteria 
ALL SUBJECTS: 
1. The subject is pregnant or lactating. 
2. The subject size or weight is not compatible with 
imaging per the investigator. 
3. The subject has received radiation therapy or 
chemotherapy or has a previous diagnosis of cancer 
other than basal cell carcinoma.  
4. The subject has renal insufficiency as demonstrated by 
a glomerular filtration rate (GFR) < 60 mL/min. 
5. The subject has a chronic or persistent infection or has 
any condition that would, in the opinion of the 
examining physician, preclude their participation. 
6. The subject has a known allergy to or has had an 
adverse reaction to dextran exposure. 
7. The subject has received an investigational product 
within 30 days prior to the Tc 99m sulfur colloid 
administration. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
Page 9 of 71 
 
8. The subject has received any radiopharmaceutical 
within 7 days prior to the administration of Tc 99m 
sulfur colloid. 
9. The subject is HIV positive.  
10. The subject has a history of alcohol abuse or currently 
consumes alcohol in excess of 3 drinks/day for men or 
2 drinks/day for women.  
11. The subject has hepatitis B or C. 
 
CONTROL SUBJECTS:  
12. The subject has hepatic insufficiency as demonstrated 
by ALT (alanine aminotransferase [SGPT]) or AST 
(aspartate aminotransferase [SGOT]) greater than two 
times the upper limit of normal (ULN). 
13. The subject has been diagnosed with NASH, NAFLD, 
or other chronic liver disease.  
14. The subject has been diagnosed with metabolic 
syndrome or Type I or II diabetes.  
 
NASH SUBJECTS: 
12. The subject has any chronic liver disease aside from 
NASH/NAFLD. 
13. The subject has uncontrolled diabetes as indicated by an 
A1c >9% within the 3 months prior to enrollment.  
Study design 
Prospective, open-label, multicenter, safety, comparative 
study of injected Tc 99m tilmanocept and unfiltered Tc 99m 
sulfur colloid in the detection of and assessment of three 
dimensional tessellation localization to the liver in subjects 
with and without moderate to severe NASH by planar and 
SPECT/CT imaging.  
On Day 0, subjects will receive IV administration of 8.0 mCi 
of unfiltered Tc 99m sulfur colloid. Subjects will have planar 
imaging of the liver followed by SPECT/CT scanning of the 
abdomen, liver, and spleen at 20 minutes ± 5 minutes post-
administration. 
On Day 3, subjects will receive IV administration of 200 µg 
of tilmanocept at the specified radiolabeled dose of Tc 99m. 
Subjects will have planar imaging of the liver followed by 
SPECT/CT scanning of the abdomen, liver, and spleen at three 
specified time points post-administration: 20 minutes ± 5 
minutes, 75 minutes ± 15 minutes and 180 minutes ± 15 
minutes.  

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Methodology 
The study includes: screening, enrollment, pre- and post-
injection assessments, administration of unfiltered Tc 99m 
sulfur colloid, administration of Tc 99m tilmanocept, imaging 
and follow-up. 
Visit 1, Screening (Day -45 to Day -1): The screening visit 
will include informed consent, review of study eligibility, 
demography, collection of medical history including 
medications, vital signs, ECG (electrocardiogram), physical 
exam including height and weight, clinical labs, urinalysis, a 
serum pregnancy test for subjects of childbearing potential, 
and a transient elastography. Adverse event monitoring will 
begin from Visit 1 Screening and continue through subject 
completion of the trial. 
Visit 2, Day 0: 
Pre-Tc 99m Sulfur Colloid Administration: 
• Before Administration (Day 0) 
o Urine pregnancy test will be performed for 
subjects of childbearing potential 
o Assessment of adverse events 
o Concomitant medication review 
o Vital Signs - within 15 minutes prior to 
administration 
 
Tc 99m Sulfur Colloid Administration : Subjects will 
receive the on label 8.0 mCi dose.  
Post-Tc 99m Sulfur Colloid Administration:  
• 20 ± 5 Minutes Post-Administration: 
o Assessment of adverse events 
o Vital Signs 
o Planar imaging of the liver/spleen followed by 
SPECT/CT of the liver/spleen and abdomen  
 
Visit 3, Day 3: 
Pre-Tc 99m Tilmanocept Administration:  
• Before administration (Day 3) 
o Urine pregnancy test will be performed for 
subjects of childbearing potential 
o Assessment of adverse events 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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o Concomitant medication review 
o Vital Signs - within 15 minutes prior to 
administration 
 
Tc 99m Tilmanocept Administration: Subjects will receive 
the open label 200 µg at the specified radiolabeled dose.   
Post-Tc 99m Tilmanocept Administration:  
• 20 ± 5 Minutes Post-Administration: 
o Assessment of adverse events 
o ECG (completed before vital signs) 
o Vital Signs 
o Planar imaging of the liver/spleen followed by 
SPECT/CT of the liver/spleen and abdomen  
• 75 ± 15 Minutes Post-Administration: 
o Assessment of adverse events 
o Planar imaging of the liver/spleen followed by 
SPECT/CT of the liver/spleen and abdomen  
• 180 ± 15 Minutes Post-Administration: 
o Assessment of adverse events 
o Planar imaging of the liver/spleen followed by 
SPECT/CT of the liver/spleen and abdomen  
• After Completion of All Imaging: 
o Blood draw for hematology and chemistry 
testing 
o Urine collection for urinalysis  
 
Day 8 ± 3: 
Safety Follow-Up Telephone Call 5 ± 3 Days Post-Injection 
of Tc 99m Tilmanocept:  
• Concomitant medication review 
• Assessment of adverse events 
Planned study dates 
Start of study 
recruitment / November 
2017 
End of Recruitment/June 
2018 
End of Study/September 2018 
Planned number of 
study centers 
Approximately 1-3 centers in the United States  
Number of subjects 
Up to 12 evaluable subjects may be enrolled.  

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Primary Endpoint 
• Proportion of subjects experiencing a noxious 
pharmacologic activity/adverse drug reaction (ADR)  
Secondary Endpoints 
• Determine the three dimensional tessellation 
localization of Tc 99m tilmanocept by planar and 
SPECT/CT imaging in subjects with NASH and 
asymptomatic controls. 
• Determine 
the 
three 
dimensional 
tessellation 
localization of unfiltered Tc 99m sulfur colloid by 
planar and SPECT/CT imaging in subjects with NASH 
and asymptomatic controls. 
• Comparison of liver localization distribution 
tessellation pattern intensities between Tc 99m 
tilmanocept and unfiltered Tc 99m sulfur colloid 
Exploratory Endoints 
• Concordance of intrahepatic localization tessellation 
discrimination of Tc 99m tilmanocept (intrahepatic 
disease heterogeneity) by SPECT/CT imaging and 
elastography  
• Concordance of intrahepatic localization tessellation 
discrimination of Tc 99m sulfur colloid (intrahepatic 
disease heterogeneity) by SPECT/CT imaging and 
elastography  
Data Analyses  
The following analysis population will be defined for the 
study: 
• Intent-to-Diagnose (ITD) Population - Subjects who 
are enrolled in the study, administered Tc 99m 
tilmanocept and unfiltered Tc 99m sulfur colloid, and 
are imaged will be included in the ITD analysis 
population. 
• Safety Population – All patients who are enrolled and 
administered Tc 99m tilmanocept and unfiltered Tc 
99m sulfur colloid in the study will be included in the 
safety population. 
 
All safety data analyses will be conducted on the safety 
population.  All efficacy data analyses will be conducted on 
the ITD population. 
The number and percentage of subjects experiencing noxious 
pharmacologic activity or ADRs will be tabulated by subject 
type, Tc 99m tilmanocept dose group (if applicable), and 
overall. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Per subject localization rates and percentage will be calculated 
by time point, by subject type, Tc 99m tilmanocept dose group 
(if applicable), and overall.   
Localization intensities (tessellation) will be summarized by 
descriptive statistics (mean, median, standard deviation, 
minimum, maximum and range) at each time point.  All results 
will be compared to statistics computed on the control group 
at each time point. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS 
ADR 
adverse drug reaction 
AE 
adverse event 
ALT 
alanine aminotransferase (SGPT) 
AST 
aspartate aminotransferase (SGOT) 
BMI 
body mass index 
BUN 
blood urea nitrogen 
CCL2 
CC-chemokine ligand 2 
CCR2 
CC-chemokine receptor 2 
CD206 
Mannose-binding receptor (Ca2+-binding lectin) 
CO2 
carbon dioxide 
CRA 
clinical research associate 
CRF 
case report form 
CRO 
contract research organization 
CSF1 
Macrophage colony-stimulating factor 1 
CSF1R 
CSF1 receptor 
CT 
X-ray computed tomography 
CX3CR1 
CX3C-chemokine receptor 1 
CXCL 
CXC-chemokine ligand 
CXCR 
CXC-chemokine receptor 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Cy3 
Cy3 cyanine dyes fluoresce orange (~550 nm excitation, ~570 nm 
emission) 
  
DAMPs 
Damage-associated molecular patterns 
DICOM 
Digital Imaging Communications in Medicine 
DTPA 
diethylene triamine pentaacetic acid  
 
ECG 
electrocardiogram 
EU 
European Union 
FASL 
FAS ligand 
FDA 
Food and Drug Administration 
FNR 
false negative rate 
GCP 
Good Clinical Practice 
GFR 
Glomerular Filtration Rate 
HC 
Healthy controls 
HCT 
hematocrit 
Hgb 
hemoglobin 
HIPAA 
Health Information Portability and Accountability Act 
ICF 
informed consent form 
ICH 
International Conference on Harmonization  
%IDSN 
percentage of Injected Dose in the Sentinel Node 
IL-10 
Interleukin-10 
IL-1β 
Interleukin-1β 
IL-6 
Interleukin-6 
ILM 
intraoperative lymphatic mapping 
IRB 
Institutional Review Board 
ISF 
investigator site file 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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ITD 
intent-to-diagnose population 
IV 
intravenous 
KD 
Dissociation constant 
kDa 
kiloDalton (molecular weight designation) 
LN 
lymph node 
LSEC 
Liver sinusoidal endothelial cell 
LY6C  
Lymphocyte antigen 6 complex 
M1 
Macrophage 1 
M2 
Macrophage 2 
mCi 
milliCurie (37x106 becquerels; 37megabecquerels) 
MedDRA 
Medical Dictionary for Regulatory Activities 
MoMFs 
Monocyte-derived macrophages 
MRI 
Magnetic Resonance Imaging  
MRS 
Magnetic Resonance Spectroscopy  
MSR1 
Macrophage scavenger receptor 1 
MTD 
maximum tolerated dose 
NAFLD 
nonalcoholic fatty liver disease 
NAS 
nonalcoholic fatty liver disease activity score 
NASH 
nonalcoholic steatohepatitis 
NHANES 
National Health and Nutrition Examination Survey 
NO 
Nitric oxide 
NOAEL 
no-observed adverse-effect level 
PGE2 
Prostaglandin E2 
PI 
principal investigator 
RA 
Rheumatoid Arthritis  
RBC 
red blood cells 
ROS 
Reactive oxygen species 
SAE 
serious adverse event 
SAP 
statistical analysis plan 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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SC 
Subcutaneous 
SCC 
squamous cell carcinoma 
SLN 
sentinel lymph node 
SLNB 
sentinel lymph node biopsy 
SPECT 
single photon emission computed tomography 
SUSARs 
Suspected, Unexpected, Serious Adverse Reactions 
Tc 99m 
technetium-99m metastable isotope; γ-emitting (t½ = 6.02 h) 
TcSC 
Tc 99m Sulfur Colloid 
Tessellation 
Three dimensional diffential localization or heterogeneity of intrahepatic 
signal 
 
TGF-β 
Transforming Growth Factor-β 
Tilmanocept 
DTPA Mannosyl Dextran (the US Adopted Name for the drug substance 
of Lymphoseek®) 
TMF 
trial master file 
TNF 
Tumour necrosis factor 
ULN 
upper limit of normal 
US 
United States 
VEGFA 
Vascular endothelial growth factor A 
VBD 
vital blue dye 
WBC  
white blood cell 
wt 
weight 

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Date: 23 April 2018 
 
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STUDY ADMINISTRATIVE STRUCTURE 
The principal investigator (PI) must sign the protocol signature sheet before study participant 
recruitment may start.  Likewise, all protocol amendments must be signed and dated by the PI 
before coming into effect. 
The name and address of the participating center, the investigators, and all required signature 
documents will be maintained in the trial master file (TMF). 
In addition to the PI, there are additional onsite roles that may be performed by other sub-
investigators:   
• 
Subject referral to the study 
• 
Review of subject eligibility and medical records  
• 
Clinical evaluations 
• 
Safety assessments 
• 
Injection and imaging  
• 
On-site image analysis 
 
Study personnel not listed in this section are identified in a separate personnel list.  This list 
will be updated as needed.  The list of personnel will be available in the center’s investigator 
site file (ISF). 

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Date: 23 April 2018 
 
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1 
INTRODUCTION 
1.1 
Background 
This clinical study provides a preliminary evaluation of Navidea’s radiopharmaceutical Tc 
99m tilmanocept as an imaging agent for the detection of CD206 (Cluster of Differentiation 
206, the macrophage mannose receptor) expressing macrophages in the liver as a possible 
indicator of pathological processes related to nonalcoholic steatohepatitis (NASH). NASH is 
a chronic inflammatory disorder. Tc 99m tilmanocept contains mannose groups attached to a 
dextran backbone, enabling Tc 99m tilmanocept to bind strongly to CD206 expressed on the 
surface of several classes of activated macrophages. Elevated levels of activated macrophages 
are associated with liver inflammation raising the possibility that imaging Tc 99m tilmanocept 
localizations by single-photon emission computed tomography (SPECT) could be used alone 
or in combination with other diagnostic tests to identify persons with NASH. 
NASH is an emerging public health concern affecting millions of persons. There is an urgent 
unmet need to develop noninvasive methods to augment and improve the accuracy of 
diagnostic evaluations of patients suspected of having NASH. Currently, liver biopsies in 
combination with various clinical findings are the most accurate means for diagnosing NASH 
[2]. However, such procedures are invasive and potentially challenging to deliver to a patient 
population as large as the one comprised of those suspected of having NASH. The primary 
objective of this study is to determine diagnostic feasibility and to explore a variety of imaging 
protocol parameters to provide a preliminary determination of an optimized imaging protocol 
for Tc 99m tilmanocept in NASH patients. Developing a non-invasive NASH diagnostic would 
greatly facilitate the delivery of new NASH therapeutics, many of which are currently in 
clinical trials [3-6]. 
Nonalcoholic fatty liver disease (NAFLD) is characterized by an accumulation of the fat in the 
liver cells. Histological evaluation of a liver biopsy revealing accumulation of fat without 
inflammation and associated damage is diagnosed as NAFLD.  NAFLD is considered to be a 
hepatic manifestation of metabolic syndrome and develops in patients with non-significant 
alcohol usage. NAFLD progression leads to further accumulation of fat in the hepatocytes, 
inflammation, and damage to liver cells that is diagnosed NASH (see Figure 1).  NASH is a 
silent disease. Further progression of NASH leads to NASH-associated liver fibrosis, cirrhosis 
and hepatocellular carcinoma. These NASH derived pathologies are the fastest growing 
indications for liver transplantation, while at the same time liver transplantation shortage is a 
major issue [8-10]. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
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Figure 1. Nonalcoholic steatohepatitis (NASH) progression [7] 
Changing diets and activity levels are causing a growing global obesity epidemic with the 
result that metabolic syndrome and its associated NAFLD are becoming more prevalent. Many 
of these individuals with NAFLD are progressing to NASH, exasperating the growing need for 
better and more assessable means to diagnose and manage this emerging patient population. 
Recent analysis of this epidemic showed global prevalence of NAFLD to be approximately 
25% - 37% with the highest risk in the Middle East, South America and lowest in Africa [11, 
12].  It is estimated that 30% of the US population has developed NAFLD with the prevalence 
of NASH currently at 5.7% [12].  Moreover, in 2010, 417,524 patients had NASH-associated 
cirrhosis, and 4,104,871 had NASH-associated advanced fibrosis based on National Health 
and Nutrition Examination Survey (NHANES) data [13].  
NASH is an inflammatory syndrome in which immune cells infiltrate into and accumulate in 
liver tissue [14, 15]. Numerous among these immune cells are macrophages. There are two 
types of macrophages in the liver. The first type consists of Kupffer cells (aka stellate 
macrophages) that reside within the endothelial lining of blood vessels in the liver. The second 
type of macrophages in the liver consists of macrophages differentiated from bone marrow 
derived monocytes that have infiltrated into the liver and developed into activated 
macrophages in response to inflammatory signals originating in the liver [14]. It is 
hypothesized that Tc 99m tilmanocept, through its affinity to CD206, will bind to and 
accumulate in both types of liver macrophages permitting the degree and anatomical 
distribution of Tc 99m tilmanocept localization to be used as an indicator of the presence and 
intensity of NASH related hepatic inflammation. In addition, NASH results in the development 
of fibrotic lesions in the liver, basically scars resulting from resolved inflammation. These 
fibrotic lesions are depleted of macrophages, thus are expected to be regions of much reduced 
Tc 99m tilmanocept localization relative to regions where NASH related inflammation is 
ongoing [1, 16, 17]. In both cases of either increased Tc 99m tilmanocept localization due to 
active NASH mediated inflammation or decreased Tc 99m tilmanocept localization due to 
macrophage depletion due to fibrotic resolution of NASH mediated inflammation, the degree 
and distribution of Tc 99m tilmanocept localization is expected to significantly differ in NASH 
involved livers relative to the degree and distribution of Tc 99m tilmanocept localization 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
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observed in livers which are not involved in NASH inflammation. This clinical study will 
provide a preliminary evaluation of the degree and distribution of Tc 99m tilmanocept 
localization in healthy livers and those involved in NASH inflammation. If this and follow on 
studies are successful in establishing that Tc 99m tilmanocept enabled imaging can identify 
areas of NASH involvement or resolution, then it may be possible to develop Tc 99m 
tilmanocept as a NASH imaging agent that can assist physicians to non-invasively diagnose 
and monitor NASH, thus satisfying the need for a non-invasive means to diagnose and assess 
NASH.   
1.2 
Liver Anatomy, Function, and Role of Kupffer cells 
The liver is the largest internal organ, responsible for filtration of the blood coming from a 
digestive tract before passing it to the rest of the body (Figure 2) [19].  Liver is made up of 
microscopic lobules, including acinus. Acinus is constructed of interconnected hexagonal units 
packed with spokes-like internal canals transporting blood and bile ducts across tightly packed 
hepatocytes. Each acinus unit has three functional zones [20, 21].  Zone 1 (periportal) performs 
a majority of metabolic functions and contains the most oxygenated blood. Zone 2 (midzone) 
is situated between zones 1 and 3.  Zone 3 (centrilobular) borders the central vein although it 
has the poorest oxygenation, and is the most susceptible to a restriction in blood supply to 
tissues resulting in ischemic damage, abnormal retention of lipids within the cell (steatosis), 
hepatocellular ballooning and hepatocyte death resulting in scar tissues (fibrosis) leading to 
possible progression to cirrhosis. Zone 3 is the anatomical location where NASH pathology 
and its consequences are most pronounced. In addition, activated Kupffer cells produce TGFβ 
and other cytokines that promote liver fibrosis [22].  Interestingly, during the process of liver 
inflammation and hepatocyte injury (ballooning), Kupffer cells have an ability to differentiate 
into activated macrophages, many of which highly express CD206 (see Figure 3) [23].  This 
observation further supports the hypothesis that Tc 99m tilmanocept binding to CD206 can 
facilitate imaging of NASH associated inflammation.  

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Date: 23 April 2018 
 
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Figure 2. Liver anatomy and lobular zoning [18] 
1.3 
NASH Diagnosis and Other Imaging Modalities 
Since NASH manifests as a chronic and gradually progressive inflammatory syndrome of the 
liver and since NASH-related liver inflammation is associated with large numbers of liver 
infiltrating CD206 expressing activated macrophages, Tc 99m tilmanocept enabled imaging of 
NASH inflamed livers could facilitate a diagnosis of NASH through observation of Tc 99m 
tilmanocept localization to areas of CD206-expressing macrophage aggregations associated 
with NASH inflammation.  
Today, physicians use a variety of imaging modalities to inform a diagnosis of NASH [24-26]. 
These alternatives to Tc 99m tilmanocept based imaging for NASH include ultrasound (US), 
computerized tomography (CT), magnetic resonance spectroscopy (MRS) and magnetic 
resonance imaging (MRI).  All of these imaging modalities have their pros and cons, but most 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
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importantly and unlike Tc 99m tilmanocept, none of these other imaging modalities directly 
image inflammatory processes [25].  Instead they measure either liver fat deposition or 
evidence of liver fibrosis [16, 24, 27]. NASH, especially in its earlier stages and before 
extensive tissue damage has occurred and when one would most likely want to initiate therapy, 
may not be associated with significant fibrosis. Thus, evaluating livers only for fibrosis 
introduces a diagnostic sensitivity issue into the diagnostic work up of patients suspected of 
having NASH. Conversely, evaluating a patient’s liver for fat content can introduce a 
diagnostic specificity issue related to discriminating between NAFLD and NASH. Since, 
NAFLD is a very common condition with limited clinical consequences, confusing NAFLD 
and NASH is problematic. By contrast, Tc 99m tilmanocept may be capable of imaging NASH 
inflammation directly. As inflammation is central to the pathology of NASH [24], Tc 99m 
tilmanocept may localize conspicuously and specifically in the livers of patients with NASH.  
In addition to the imaging modalities discussed in the previous paragraph, many investigators 
have evaluated the utility of using various 99mtechnetium colloids as imaging liver agents [1, 
16, 17, 27]. In fact, Tc 99m sulfur colloid will be evaluated as a comparator to Tc 99m 
tilmanocept for liver imaging in this protocol. Colloids are particles that are actively 
phagocytized by Kupffer cells. When injected into the blood circulation, Tc 99m sulfur colloid 
is rapidly cleared from the circulation by the Kupffer cells [26]. Tc 99m sulfur colloid 
localization to Kupffer cells enables SPECT imaging of livers. Tc 99m tilmanocept localization 
is expected to differ from Tc 99m sulfur colloid localization in two ways. First, colloidal 
particles are expected to penetrate poorly into the liver. Thus, Tc 99m sulfur colloid will be 
unable to localize to aggregates of activated macrophages that have penetrated into liver tissue. 
Second, Tc 99m sulfur colloid localization to Kupffer cells is independent of their cellular 
activation state and CD206 expression phenotype. Conversely, Tc 99m tilmanocept 
localization is CD206 dependent and the extent of localization is expected to vary relative to 
the level of CD206 expression and the density of CD206 expressing cells. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
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Figure 3. Involvement of the macrophages in the liver damage [15] 
Abbreviations: CCL2, CC-chemokine ligand 2; CCR2, CC-chemokine receptor 2; CSF1, macrophage colony-
stimulating factor 1; CSF1R, CSF1 receptor; CX3CR1, CX3C-chemokine receptor 1; CXCR, CXC-chemokine 
receptor; CXCL, CXC-chemokine ligand; DAMPs, damage-associated molecular patterns; FASL, FAS ligand; 
IL-1β, interleukin-1β; IL-6, interleukin-6; IL-10, interleukin-10; LSEC, liver sinusoidal endothelial cell; LY6C, 
lymphocyte antigen 6 complex; MoMFs, monocyte-derived macrophages; MSR1, macrophage scavenger 
receptor 1; NO, nitric oxide; PGE2, prostaglandin E2; ROS, reactive oxygen species; TNF, tumour necrosis 
factor; VEGFA, vascular endothelial growth factor A. 
1.4 
Preliminary Data 
1.4.1 
99mTc-tilmanocept (Lymphoseek®) 
Tc 99m tilmanocept is a radiopharmaceutical imaging agent that was originally designed to 
bind specifically and with high affinity to CD206 on macrophages that reside in lymph nodes 
(Figure 4). Tilmanocept is a macromolecule consisting of multiple units of DTPA 
(diethylenetriaminepentaacetic acid) and mannose, each synthetically attached to a 10 kDa 
dextran backbone via amine terminated molecular leashes. The mannose acts as a ligand for 
CD206 while the DTPA moieties serve as chelating agents for labeling with 99mtechnetium 
(99mTc).  

Tc 99m tilmanocept 
 
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Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
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Figure 4. Structure of Tc99m-tilmanocept (Lymphoseek®) 
Showing attachment of multiple units of DTPA and mannose to a backbone of dextran. The chelating agent, 
DTPA, is shown holding an atom of 99mTechnetium. 
The final construct has an average molecular weight of 18.7 kDa, is highly water soluble and 
has an apparent particle diameter of 7 nm. Tilmanocept’s multiple mannose moieties permit it 
to bind multivalently with very high affinity (KD = 2.7 x 10-11M) to CD206 through CD206’s 
8 mannose binding domains. Multivalent binding is required for this high affinity interaction 
[28]. After binding to CD206, tilmanocept is internalized and accumulates in endosomes. 
Tilmanocept is protected by the U.S. patent #6,409,990. 
Following various preclinical studies in both cells based assay systems and in various animal 
species and following numerous clinical studies (including three phase 3 trials) in human 
subjects with breast cancers, melanomas, and head and neck cancers [29-34],  Lymphoseek® 
was approved by the United States Food and Drug Administration (FDA) in 2013 [35] for use 
as a lymphatic mapping agent in patients undergoing surgeries to remove melanomas and 
breast tumors without presurgical evidence of metastatic disease. Since then, the indications 
for which Lymphoseek® has been approved have been expanded to all lymphatic mapping to 
assist in the localization of lymph nodes draining a primary tumor site in patients with solid 
tumors for which this procedure is a component of intraoperative management and for guiding 
sentinel lymph node biopsy (SLNB). Additionally, Lymphoseek® is approved in the European 
Union (EU) for use in imaging and intraoperative detection of sentinel lymph nodes (SLNs) 
draining a primary tumor in adult patients with breast cancer, melanoma, or localized 
squamous cell carcinoma of the oral cavity. Lymphoseek® has an extraordinary safety record. 
Since its first regulatory approval in 2013, Lymphoseek® has been injected into over 170,000 
cancer patients for various SLN related indications. To date, there have been no serious adverse 
events associated with these procedures that have been attributed to Lymphoseek®. Typical of 
how Lymphoseek® is used, Figure 5 shows a SPECT/CT image of a breast cancer patient 
injected peritumorally with Lymphoseek®. Localization to a SLN is clearly shown. 
DTPA 
Molecular 
Leash 
    Mannose 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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Figure 5. Pre-operative SPECT-CT image of a breast cancer patient who had 
been injected peritumorally with 99mTc-tilmanocept 
Lymphatic transport and accumulation in the first encountered lymph node, the sentinel lymph node (SLN), is 
observed. 
1.4.2 
Transitioning from Subcutaneous and Intradermal Administration of 99mTc-
Tilmanocept to IV Administration in Diagnostic Imaging indications 
For all previously approved SLN and lymphatic mapping indications, the approved routes of 
administration were subcutaneous, intradermal or peritumoral injection. Intravenous (IV) 
injection, as is proposed in this protocol has not been approved as part of the label for 
Lymphoseek®. It is well known that CD206 expressing macrophages aggregate at sites of 
inflammation associated with a number of serious illnesses. Navidea wishes to expand the 
approved diagnostic indications for which 99mTc-Tilmanocept is approved to include several 
new diagnostic imaging indications that exploit the pathological aggregations of CD206 
expressing macrophages associated with these illnesses. Most if not all of these alternative 
indications, including the one proposed in this protocol for imaging NASH inflammation, will 
require IV administration.  
1.4.3 
Nonclinical Evaluations – IV Administration 
Dosing In Rats: In preparation to initiate the IV route of administration, eleven (11) preclinical 
tests were conducted to assess safety, toxicity, and interaction potential at doses hundreds to 
thousands of times the expected maximum human dose, as summarized below: 
Type of Study / 
Description 
Test System 
Method of 
Administration 
Dosing 
Central nervous system 
safety pharmacology  
Rat 
Intravenous 
37, 190, and 380 µg/animal 
or equivalent 490X and 61X 
the anticipated study doses 
of 50 µg and 400 µg in 
humans 

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Date: 23 April 2018 
 
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Type of Study / 
Description 
Test System 
Method of 
Administration 
Dosing 
Expanded single-dose 
toxicology (including 
toxicokinetics and local 
tolerance) 
Rat 
Intravenous 
37, 190, and 380 µg/animal 
or equivalent 490X and 61X 
the anticipated study doses 
of 50 µg and 400 µg in 
humans 
Respiratory Safety 
Pharmacology 
Evaluation Using Head-
Out Plethysmography of 
Tilmanocept following 
Intravenous Bolus 
Injection in Male Rats 
Rat 
Intravenous 
60, 120, and 300 µg/animal 
or equivalent 320X and 41X 
the anticipated study doses 
of 50 µg and 400 µg in 
humans 
In Vitro Evaluation of 
Tilmanocept as an 
Inhibitor of Cytochrome 
P450 (CYP) Enzymes in 
Human Liver 
Microsomes 
Human Liver 
Samples 
In vitro 
0.6 to 600 nM 
In Vitro Evaluation of 
Tilmanocept as an 
Inhibitor of Human ABC 
and SLC Transporters 
Human Liver 
Samples 
In vitro 
0.04, 0.4 µM 
Pharmacokinetics, 
Excretion, and 
Distribution by 
Quantitative Whole-
Body Autoradiography 
Following Intravenous 
Administration of 
99mTc-Tilmanocept in 
Rats 
Rat 
Intravenous 
25 µg in 0.5 mL with 
collection of  Blood, Urine, 
Feces, and Carcasses for 
QWBA 
Hemolysis and protein 
flocculation 
Human blood 
samples 
In vitro 
2.5, 25, and 250 µg/mL 
whole human blood 
Target profiling screen 
Ion Channel 
In vitro 
See Individual Tests Below 

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Date: 23 April 2018 
 
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Type of Study / 
Description 
Test System 
Method of 
Administration 
Dosing 
K Ion Channel 
Ion Channel 
The cardiac potassium 
channel, hERG, is 
responsible for a rapid 
delayed rectifier current 
(IKr) in human ventricles. 
This channel has been 
selected for evaluation 
because inhibition of IKr 
is the most common cause 
of cardiac action potential 
prolongation by non-
cardiac drugs. In this 
assay, hERG potassium 
channels are expressed in 
a human embryonic 
kidney (HEK293) cell line 
that lacks endogenous 
IKr. 
0.025, 
0.05, 0.25, 
0.5 mg/mL 
Na Ion Channel 
Ion Channel 
Cloned hNav1.5 sodium 
channels (SCN5A gene 
expressed in CHO cells) 
0.025, 
0.05, 0.25, 
0.5 mg/mL 
Ca Ion Channel 
 
 
Ion Channel 
1. Cloned L-type calcium 
channels (hCav1.2, 
encoded by the human 
CACNA1C gene and 
coexpressed with the β2 
subunit, encoded by the 
human CACNB2 gene 
and the α2δ1 subunit 
encoded by the human 
CACNA2D1 gene in 
CHO cells), responsible 
for ICa,L, high threshold 
calcium current. 
2. Cloned hNav1.5 
sodium channels (SCN5A 
gene expressed in CHO 
cells). 
0.025-0.5 mg/mL 
 
Conclusions from These Tests: 
CNS: In conclusion, a single IV administration of tilmanocept was well tolerated in rats at 
levels of 0.15, 0.75, and 1.50 mg/kg. Brief sedation shortly after dosing was observed at 0.75 
and 1.50 mg/kg, which resolved by the time of the first functional observational battery 
assessments and was attributed to the mannosyl-dextran content of tilmanocept. There were no 
tilmanocept-related effects on functional observational battery parameters. 

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Date: 23 April 2018 
 
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Single Bolus Toxicity: Tilmanocept-related clinical pathology changes were limited to 
minimally greater, dose-related, aspartate aminotransferase (AST) values for 0.75 and 1.5 
mg/kg males and females that were likely caused by muscle or erythrocyte release as changes 
did not occur in other hepatobiliary-related clinical pathology parameters. This change had 
resolved at the end of the recovery phase. 
At the Day 2 necropsy, dark focus in the glandular stomach was considered to be a potential 
test article-related gross pathology finding in males at ≥ 0.75 mg/kg. The gross finding of dark 
focus in the stomach correlated microscopically with focal erosion or minimal hemorrhage. 
Focal erosion in the stomach was test article related in males at ≥0.75 mg/kg, but was 
considered to be of little toxicological significance. No test article-related organ weight 
changes were noted.  At the end of the recovery phase (Day 15), there were no test article-
related findings in gross pathology, organ weights, or histopathology. 
Respiratory: In conclusion, respiratory function was assessed in male Crl:CD(SD) rats given 
a single intravenous injection dose of vehicle control article or 0.150, 0.300, or 0.750 mg/kg 
of tilmanocept at a dose volume of 3 mL/kg.  Administration of tilmanocept had no effect on 
mortality, but it was associated with severe abnormal clinical observations of hypoactivity, 
ataxia, labored or irregular respiration, and pale skin of entire body for animals given 0.750 
mg/kg.  Administration of >0.150 mg/kg tilmanocept had no direct effect on respiration rate, 
but it was associated with higher tidal volume (up to +26, +14, and +50% in animals given 
0.150, 0.300, or 0.750 mg/kg, respectively) and higher minute volume (up to +18, +5, and 
+40% 0.5 through 1 hour post-dose in animals given 0.150, 0.300, or 0.750 mg/kg, 
respectively). 
CYP:  This study was designed to independently evaluate the in vitro inhibitory effect of 
tilmanocept on major CYP enzymes in human liver microsomes. Specifically, the inhibition of 
CYP1A2, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6 and CYP3A4/5 (utilizing two 
different substrates) by tilmanocept was assessed with the aim of ascertaining the potential of 
tilmanocept to inhibit the metabolism of concomitantly administered drugs. 
To evaluate tilmanocept as a direct, time-dependent and metabolism-dependent inhibitor of 
CYP activity, human liver microsomes from a pool of 200 human donor sources were 
incubated with marker substrates in the presence or absence of tilmanocept. To distinguish 
between timedependent and metabolism-dependent inhibition, tilmanocept was preincubated 
with human liver microsomes for 30 min without and with an NADPH-generating system, 
respectively, prior to the incubation with the marker substrate. Known direct and metabolism-
dependent inhibitors of CYP enzymes were included as positive controls in all experiments. 
Under these experimental conditions, there was little or no evidence of direct, time- or 
metabolismdependent inhibition of any CYP enzyme by tilmanocept. 
ABC and SLC transporters: It was found that tilmanocept was not an inhibitor of  human 
transporters (namely P-gp, BRCP, OAT1B1, OATP1B3, OAT1, OAT3, and OCT2) under the 
conditions tested.  

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Drug Distribution: Drug-related radioactivity was widely distributed throughout the whole 
body. The highest mean concentrations of radioactivity were observed in urine > kidney cortex 
> liver > kidney > kidney medulla > bile > spleen > spleen red pulp > spleen white pulp > 
urinary bladder between 0.25 and 36 hours post-dose. The concentration of radioactivity in 
other tissues was low (<500 ng equivalents 99mTc-Tilmanocept/g) and maximum levels were 
generally observed between 0.25 and 2 hours post-dose. Concentrations declined over time 
and were not detectable in the majority of the tissue by 36 hours post-dose. The exceptions 
were the kidney, liver, spleen, and urine, in which concentrations were still detectable at 48 
hours post-dose. By 72 hours post-dose levels of radioactivity in all tissues were not detectable. 
The main route of elimination of drug-related radioactivity following an IV administration of 
99mTc-Tilmanocept was via urine, with a mean 38.9% of the dose administered detected in 
urine over 0 to 120 hours post-dose. A mean of 10.0% of the administered dose was eliminated 
via feces. The total recoveries were a mean of 50.8 ± 4.87% over the measurable time period 
for technetium Tc99m detection (t0.5 = 6.02 hr). 
Hemolysis: No hemolysis and no flocculation were observed following in vitro treatment of 
human whole blood with tilmanocept at final whole blood concentrations of 2.5, 25 or 250 
μg/mL. 
Ion Channels (Na+, K+, Ca2+): Although there is a small dose-dependent effect that is 
fractional to the positive control, variability within concentration renders the median value 
observations not significantly different (Kruskal-Wallis). 
1.4.4 
Clinical Evaluations – IV Administration  
The first questions that needed to be addressed when initiating studies evaluating 99mTc-
Tilmanocept against new diagnostic targets using a new route of administration are: Can 99mTc-
Tilmanocept specifically and sufficiently localize to aggregations of pathology related CD206 
expressing macrophages to enable diagnostic imaging of disease associated inflammation, and 
what is the optimal imaging protocol to facilitate this desired imaging capability. In February 
of 2017, Navidea initiated the first such study evaluating these questions in which IV injected 
99mTc-Tilmanocept was evaluated for localization in skeletal joints of subjects with skeletal 
joint inflammation due to rheumatoid arthritis (RA). As of this writing, this study in patients 
with active RA is still ongoing but has already provided positive affirmation that IV injected 
99mTc-Tilmanocept localizes specifically and sufficiently to CD206 macrophages in RA 
inflamed joints with enough intensity to enable diagnostic imaging.  
The study (Clincaltrials.gov number NCT02865434) was designed to determine an optimal 
imaging protocol utilizing IV injected 99mTc-Tilmanocept to visualize aggregations of CD206 
expressing macrophages in skeletal joints inflamed due to involvement of rheumatoid arthritis 
(RA) and to evaluate safety variables. The study is expected to have 39 participants: 33 with 
active RA and 6 (healthy controls) without joint inflammation. Twenty-seven of the 
participants will be divided into 9 cohorts comprised of three participants each. The 
participants in each cohort will receive a different combination of tilmanocept mass dose labels 
with a different specific activity of 99mtechnetium (99mTc) as described in Table 1. 

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Table 1. 
The RA Dose Matrix of 99mTc-Tilmanocept administration 
Dose Matrix 
50 µg Tilmanocept 
200 µg Tilmanocept 400 µg Tilmanocept 
10 mCi 99mTc 
Cohort 1 
Cohort 2 
Cohort 3 
5 mCi 99mTc 
Cohort 4 
Cohort 5 
Cohort 6 
1.0 mCi 99mTc 
Cohort 7 
Cohort 8 
Cohort 9 
 
 
Two additional cohorts of 6 participants each will be recruited and imaged using the selected 
optimal dose. One cohort (Cohort 10) will be comprised of individuals with active RA, while 
the other cohort (Cohort 11) will consist of healthy individuals without joint disease. In 
addition to imaging of skeletal joints that may be involved in RA, participants in Cohorts 10 
and 11 with be evaluated for 99mTc-Tilmanocept biodistribution, dosimetry and blood plasma 
half-life.  
Three more subjects remain to be injected and imaged in each Cohort 10 and Cohort 11. To 
date, no serious adverse events have been observed and no adverse events have been attributed 
to tilmanocept in this study. 
Figure 6A and Figure 6B show examples of the kinds of imaging results that are being obtained 
in this study. Figure 6A shows that hands of a participant with active RA, while Figure 6B 
shows an image of an RA involved ankle of a different individual. In both figures, localization 
of 99mTc-Tilmanocept to RA inflamed joints is clearly visible.  

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Date: 23 April 2018 
 
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(A) 
(B) 
Figure 6. IV Administered Tc 99m tilmanocept Localization to RA Joints 
(A) 99mTc-Tilmanocept localization in hands (small joints’ detection area after 200 µg/10 mCi 
dose of 99mTc-tilmanocept; picture on the left) and (B) in ankles and feet (50 µg/10 mCi dose 
of 99mTc-tilmanocept; picture on the right) after injections of 99mTc- tilmanocept administrated 
intravenously. 
1.4.5 
Clinical Safety using Intradermal, Subcutaneous, and Peritumoral Injections 
Over 200,000 patients in clinical trials and commercial use in the U.S. for ILM with SLNB 
have been exposed to Tc 99m tilmanocept (Lymphoseek®), and there have been no safety 
signals, no deaths due to drug, and no Serious Adverse Events (SAEs) due to Lymphoseek®. 
There are no known drug interactions leading to contraindications with the use of 
Lymphoseek®. Post-marketing reports have shown less than 0.12% of subjects experiencing 
AEs, with the most common one being lack of node localization.  

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2 
STUDY OBJECTIVES 
2.1 
Primary Objective(s)  
• Determine the safety and tolerability of Tc 99m tilmanocept in subjects with NASH.  
 
2.2 
Secondary Objective(s) 
• Determine the three dimensional tessellation localization of Tc 99m tilmanocept by 
planar and SPECT/CT imaging in subjects with NASH and asymptomatic controls. 
• Determine the three dimensional tessellation localization of unfiltered Tc 99m sulfur 
colloid by planar and SPECT/CT imaging in subjects with NASH and asymptomatic 
controls. 
• Comparison of liver localization distribution tessellation pattern intensity between Tc 
99m tilmanocept and unfiltered Tc 99m sulfur colloid 
 
2.3 
Exploratory 
• Concordance of intrahepatic localization tessellation discrimination of Tc 99m 
tilmanocept (intrahepatic disease heterogeneity) by SPECT/CT imaging and 
elastography  
• Concordance of intrahepatic localization tessellation discrimination of Tc 99m sulfur 
colloid (intrahepatic disease heterogeneity) by SPECT/CT imaging and elastography 
 

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3 
OVERVIEW OF METHODOLOGY AND DESIGN 
3.1 
Overall Study Design 
This is a prospective, open-label, multicenter, safety, comparative study of IV administered Tc 
99m tilmanocept and unfiltered Tc 99m sulfur colloid in the detection of and assessment of 
three dimensional tessellation localization to the liver in subjects with and without moderate 
to severe NASH by planar and SPECT/CT imaging.   
On Day 0, subjects will receive IV administration of 8.0 mCi of unfiltered Tc 99m sulfur 
colloid. Subjects will have planar imaging of the liver followed by SPECT/CT scanning of the 
abdomen, liver, and spleen at 20 minutes ± 5 minutes post-administration.   
On Day 3, subjects will receive IV administration of 200 µg of tilmanocept at the specified 
radiolabeled dose of Tc 99m. Subjects will have planar imaging of the liver followed by 
SPECT/CT scanning of the abdomen, liver, and spleen at three specified time points post-
administration: 20 minutes ± 5 minutes, 75 minutes ± 15 minutes, and 180 minutes ± 15 
minutes. 
The proposed study includes 3 visits and a safety follow-up telephone call: a screening visit 
for initial determination of eligibility and evaluation of clinical status (“Visit 1”), Day 0 (“Visit 
2”) the day of unfiltered Tc 99m sulfur colloid administration and imaging, Day 3 (“Visit 3”) 
the day of Tc 99m tilmanocept administration and imaging, and a safety follow-up telephone 
call 5 ± 3 days following Tc 99m tilmanocept administration.  
The Schedule of Events (Appendix 1) contains a list of all study procedures and time points. 
Study activities are described in detail in Section 7.  
Following completion of the first two enrolled NASH subjects, a review meeting will be held 
to evaluate tilmanocept safety and dose performance.  
3.2 
Justification for Study Design and Population 
This feasibility study is designed to evaluate the safety of Tc 99m tilmanocept and to compare 
Tc 99m tilmanocept tessellation localization in the livers of subjects with moderate to severe 
NASH and healthy controls (HCs) to unfiltered Tc 99m sulfur colloid localization. Planar 
imaging of the liver using a SPECT camera will be combined with SPECT/CT imaging of the 
abdomen to provide greater resolution of areas of intrahepatic localization. 
This study is designed to evaluate the use of Tc 99m tilmanocept as an imaging agent in 
subjects with biopsy-confirmed moderate to severe NASH by evaluating intrahepatic 
localization tessellation. 
3.3 
Protocol Adherence 
Strict adherence to all specifications outlined in this protocol is required for all aspects of the 
study conduct; the investigator may not modify or alter the procedures described in this 
protocol. If protocol modifications are necessary, all alterations that are not solely of an 

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administrative nature require a formal protocol amendment for the involvement of Institutional 
Review Board(s) (IRB(s)). 
If an investigator has deviated from the protocol in order to eliminate an immediate hazard to 
subjects or for other inevitable medical reasons, the investigator shall document all such 
deviations, including the reasons thereof, and submit the document to the sponsor and the IRB 
as applicable. 
3.4 
Study Duration 
Subjects will be “on study” for up to 57 days depending on the duration of the screening 
window (up to 45 days). 

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4 
STUDY POPULATION 
4.1 
Eligibility 
Subjects who fulfill all respective inclusion and none of the respective exclusion criteria will 
be eligible for enrollment into the study.  All inclusion/exclusion criteria must be verified 
before a subject may be considered eligible for administration of unfiltered Tc 99m sulfur 
colloid and imaging (Day 0 procedures).  A subject will be considered enrolled in the study on 
the morning of study Day 0 when they arrive at the study site and receive Tc 99m sulfur colloid 
via IV administration.  Written and dated (with time noted) informed consent will be obtained 
from all subjects. A subject who withdraws consent prior to arrival at the study site on Day 0 
will be considered a screen failure. 
4.1.1 
Inclusion Criteria 
ALL SUBJECTS: 
1. The subject has provided written informed consent with HIPAA (Health Information 
Portability and Accountability Act) authorization before the initiation of any study-
related procedures. 
2. The subject is ≥18 years of age at the time of consent.  
3. The subject has a body mass index (BMI) between 18 and 45. 
 
CONTROL SUBJECTS: 
4. The subject is deemed to be clinically free of any infectious/inflammatory disease(s) for 
at least 4 weeks prior to the consent date. 
5. The subject has not taken any antibiotics for at least 4 weeks prior to the consent date. 
 
NASH SUBJECTS: 
4. The subject has biopsy-confirmed NASH within 12 months prior to enrollment.  
5.  The subject has a NAFLD Activity Score (NAS) of ≥ 4, with a score of at least 1 for 
each steatosis, lobular inflammation, and hepatocyte ballooning.  
6.  The subject has fibrosis staging of F3-F4.   
 
4.1.2 
Exclusion Criteria 
ALL SUBJECTS: 
1. The subject is pregnant or lactating. 
2. The subject size or weight is not compatible with imaging per the investigator. 
3. The subject has received radiation therapy or chemotherapy or has a previous diagnosis 
of cancer other than basal cell carcinoma.  
4. The subject has renal insufficiency as demonstrated by a GFR of < 60 mL/min. 

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5. The subject has a chronic or persistent infection or has any condition that would, in the 
opinion of the examining physician, preclude their participation. 
6. The subject has a known allergy to or has had an adverse reaction to dextran exposure. 
7. The subject has received an investigational product within 30 days prior to the Tc 99m 
sulfur colloid administration. 
8. The subject has received any radiopharmaceutical within 7 days prior to the 
administration of Tc 99m sulfur colloid. 
9. The subject is HIV positive.  
10. The subject has a history of alcohol abuse or currently consumes alcohol in excess of 3 
drinks/day for men or 2 drinks/day for women.  
11. The subject has hepatitis B or C. 
 
CONTROL SUBJECTS:  
12. The subject has hepatic insufficiency as demonstrated by ALT (alanine 
aminotransferase [SGPT]) or AST (aspartate aminotransferase [SGOT]) greater than 
two times the upper limit of normal (ULN). 
13. The subject has been diagnosed with NASH, NAFLD, or other chronic liver disease.  
14. The subject has been diagnosed with metabolic syndrome or Type I or II diabetes.  
 
NASH SUBJECTS: 
12. The subject has any chronic liver disease aside from NASH/NAFLD. 
13. The subject has uncontrolled diabetes as indicated by an A1c >9% within the 3 months 
prior to enrollment.  
 
4.2 
Recruitment 
Subjects will be recruited from gastroenterology practices in accordance with the inclusion and 
exclusion criteria listed above.  Potentially suitable subjects will be asked by their treating 
physician about their willingness to participate in this study.  Healthy, “NASH-free” subjects 
(controls) may be recruited via IRB approved advertisements and clinically assessed for the 
“absence” of liver and inflammatory diseases. 
4.3 
Withdrawal 
In accordance with the Declaration of Helsinki, each subject is free to withdraw from the study 
at any time and without providing a reason. 
A subject who withdraws consent prior to administration of Tc 99m sulfur colloid at the study 
site on Day 0 will be considered a withdrawn subject. 
Should a subject withdraw after administration of the investigational product, all efforts will 
be made to complete and report the observations up to the time of withdrawal as thoroughly as 

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possible. An explanation should be given of why the subject is withdrawing or being 
withdrawn from the study.  
The investigator may withdraw a subject from the study at any time at the discretion of the 
investigator for any of the following reasons:  
• 
A protocol violation occurs  
• 
A serious or intolerable AE occurs  
• 
A clinically significant change in a laboratory parameter occurs  
• 
At the investigator’s/sponsor’s discretion as long as it is in the best interest of the 
subject  
• 
The sponsor or investigator terminates the study  
• 
The subject requests to be discontinued from the study 
 
4.4 
Replacement 
Subjects will be replaced under the following conditions: 
• 
Subjects who did not receive both unfiltered Tc 99m sulfur colloid and Tc 99m 
tilmanocept  administration or did not proceed to imaging following either 
administration.  
 
4.5 
Subject Identification 
After the subject provides written informed consent, the site will assign the subject a 7-digit 
subject number. Subject numbers are to be assigned in a sequential manner using the following 
format:  
Digits 1 to 2: Study number “30”  
Digits 3 to 4: Site number (e.g., “01”)  
Digits 5 to 7: Sequential subject number (e.g., “001”, “002”, “003”) 
 
 
For example, the first subject consented at Site 01 is subject number “30-01-001.”  
 
Subjects will maintain the same number given at screening for the entire study. If a subject is 
a screen failure, the number will not be used for any other subject. 

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5 
INVESTIGATIONAL PRODUCT 
5.1 
Identification of Investigational Product 
Tc 99m tilmanocept is a radiopharmaceutical that binds to mannose binding receptors (CD206) 
that reside on the surfaces of dendritic cells and macrophages. 
5.2 
Investigational Product Dosage and Administration 
On Day 0, subjects will receive an on label 8.0 mCi dose of unfiltered Tc 99m sulfur colloid 
via IV administration. On Day 3, subjects will receive 200 µg tilmanocept via IV 
administration at the specified radiolabeled dose of Tc 99m. A single syringe will be used for 
each product and injected as a slow push into the IV. At the completion of each injection, a 10 
cc sterile normal saline flush will be administered. The preferred site of IV placement will be 
the left or right antecubital vein. The final administered tilmanocept dose will be ± 20% of the 
mass dose and radiolabel mCi dose assignment.  
The first two enrolled subjects will be NASH subjects who will receive an 8.0 mCi radiolabel 
dose of Tc 99m tilmanocept. A review meeting will be held following completion of the first 
two subjects to evaluate tilmanocept safety and dose performance.  
At the review (as diagrammed below): 
1. If it is determined that a lower radiolabel dose of tilmanocept may produce images of 
better quality, three additional NASH subjects and three HCs will be injected and 
imaged with 200 µg of 2.5 mCi Tc 99m tilmanocept. After these six subjects have 
completed the trial, no additional subjects will be enrolled.  
 
2. If the review shows no safety signals and acceptable quality of imaging, one additional 
NASH subject as well as three HCs will be injected and imaged with 200 µg of 8.0 
mCi Tc 99m tilmanocept. Once a total of three NASH subjects and three HCs have 
completed the trial, a second review meeting will be held to compare results from 
NASH subjects to those of the HCs. If it is determined at the second review meeting 
that a lower radiolabel dose of tilmanocept may produce additional clinically 
meaningful results, 3 NASH subjects and 3 HCs will be dosed with 200 µg of 2.5 mCi 
Tc 99m tilmanocept. If it is determined that additional doses will not provide clinically 
meaningful results, the trial will be concluded. 
 

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5.3 
Treatment Assignment 
In this prospective, open-label, non-randomized, multicenter, safety, comparative study, all 
subjects will receive IV administration of both 8.0 mCi unfiltered Tc 99m sulfur colloid and 
200 µg tilmanocept at the specified radiolabeled Tc 99m dose . After the completion of two 

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NASH subjects at the 8.0 mCi radiolabel dose of Tc 99m tilmanocept, the tilmanocept dose 
will reviewed, as detailed in Section 5.2.  
5.4 
Packaging and Labeling 
Unfiltered Tc 99m sulfur colloid will be prepared at an on label 8.0 mCi dose according to 
standard clinical practice by Nuclear Medicine technologists at the clinical site [36].  
The dose of Tc 99m tilmanocept should be ordered from Cardinal Health once the subject has 
been scheduled for IV administration and imaging. 
Tilmanocept cartons ready for radiolabeling will be shipped and stored at the region-specific 
Cardinal Health radiopharmacy.  Tilmanocept is provided in a vial. Vials are packaged as a kit. 
A carton contains five vials of tilmanocept. One kit, which contains one tilmanocept vial, 
should be used for no more than one subject. The carton also contains five shield labels and 25 
syringe labels. This package has been designed specifically for tilmanocept and protects the 
vials during shipment, handling, and storage.  Navidea will provide a radiolabeling protocol 
and Quality Control worksheets. The Cardinal Health radiopharmacy will radiolabel the 
specified tilmanocept dose at the assigned millicurie dose Tc 99m in 1.0 mL and deliver one 
syringe to the clinical site radiopharmacy that is ready for administration. 
5.5 
Drug Logistics and Investigational Product Accountability 
The investigator (or designated personnel) will confirm receipt of the investigational product 
in writing and will use the investigational product only within the framework of this clinical 
study and in accordance with this study protocol.  For each subject, he/she will keep a record 
of the investigational product dispensed and store all other forms that accompanied the delivery 
of the radiolabeled product to the clinical site. These documents are to be filed in the 
investigator site file. Overall drug accountability and reconciliation will be completed by the 
sponsor or its representative.  A list of investigational product vials and other materials that 
were returned or destroyed must be recorded and signed by the PI or an appropriately qualified 
designee as documented in the study site responsibility sheet. An overall accountability and 
reconciliation form of the investigational product will be prepared and completed. If there are 
any discrepancies, they must be investigated and their resolution documented. All unused study 
kits will be destroyed in accordance with institutional destruction procedures. 

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6 
THERAPIES OTHER THAN INVESTIGATIONAL PRODUCT 
6.1 
Prior and Concomitant Therapy 
All medications taken 30 days prior to the first screening visit through the 5 ± 3 days post-
tilmanocept injection (Day 8 ± 3) safety follow-up telephone call will be documented. Subjects 
receiving radiation therapy or chemotherapy within 12 months of tilmanocept injection or who 
have been diagnosed with cancer other than basal cell carcinoma are not eligible for 
participation in the trial. The subject’s history of treatments for NASH will also be collected. 
6.2 
Post-Study Therapy  
There are no post-study therapy restrictions. 

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7 
STUDY PROCEDURES 
7.1 
Schedule of Evaluations 
A schedule of evaluations is provided in the Schedule of Study Events (see Appendix 1). 
7.2 
Visit Description 
7.2.1 
Visit 1, Screening (Day -45 to Day -1) 
• 
Preliminary review of inclusion and exclusion criteria 
• 
Obtain signed informed consent for study participation 
• 
Allocation of unique subject number; this number will be used to document the 
subject data in the case report forms (CRFs) and enrollment log 
• 
Demography – date of birth, gender, race 
• 
Medical/surgical history- all relevant prior medical and surgical conditions will be 
recorded in the CRF. Documented medical conditions will also note the month and 
year of onset if the condition is still active. The subject’s history of NASH treatment 
will also be collected.  
• 
Concomitant Medications (within 30 prior to screening visit). 
• 
Vital signs (body temperature, heart rate, blood pressure, and respiratory rate after at 
least 1 minute in a resting position) 
• 
ECG – The electrocardiogram (ECG) reading will be assessed for clinical 
significance by the investigator, and any clinically significant abnormal findings will 
be noted on the subject’s medical history. 
• 
Physical examinations will include an assessment of height, weight and calculation of 
BMI and an examination of general appearance, skin, eyes, ears, nose, throat, head 
and neck (including thyroid), lungs, heart, abdomen, musculoskeletal, and nervous 
system. Any clinically relevant finding is to be documented as a baseline finding. 
Physical exams that are conducted as standard of care prior to signing informed 
consent may be used if they are performed within 30 days of injection. 
• 
Blood draw for routine hematology, chemistry, and serum pregnancy test for women 
of child-bearing potential. Females of child bearing potential are defined as women 
that are not surgically sterile (hysterectomy, tubal ligation, or bilateral oophorectomy) 
nor postmenopausal for at least 1 year prior to screening. Women who are not of 
childbearing potential will not require a pregnancy test.  
• 
Urine collection for routine urinalysis  
• 
Transient elastography will be performed to assess liver fibrosis  
• 
Final confirmatory review of inclusion and exclusion criteria.  
AE monitoring will begin from Visit 1 Screening and continue through subject completion of 
the trial. 
 

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7.2.2 
Visit 2 (Day 0) Unfiltered Tc 99m Sulfur Colloid Administration 
7.2.2.1 
Before Administration of Unfiltered Tc 99m Sulfur Colloid 
The following will be completed on the day of administration prior to the administration of 
unfiltered Tc 99m sulfur colloid: 
• A urine pregnancy test for women of child-bearing potential. Females of child 
bearing potential are defined as women that are not surgically sterile (hysterectomy or 
bilateral oophorectomy) nor postmenopausal for at least 1 year prior to screening. 
Women who are not of childbearing potential will not require a pregnancy test. 
• Assessment of AEs 
• Concomitant medication review 
• Vital signs after at least 1 minute in a resting position (body temperature, heart rate, 
blood pressure, and respiratory rate) within 15 minutes prior to administration of 
unfiltered Tc 99m sulfur colloid 
 
7.2.2.2 
Administration of Unfiltered Tc 99m Sulfur Colloid 
IV administration of unfiltered Tc 99m sulfur colloid will be at study time Day 0 Time 00:00. 
The preferred site of IV placement will be the left or right antecubital vein. 
All subjects will receive an on label 8.0 mCi dose of unfiltered Tc 99m sulfur colloid. The 
filled syringe will be connected to the IV for a slow push injection. At the completion of the 
injection, a 10 cc sterile normal saline flush will be administered.  
The IV administration will be performed in the nuclear medicine department by an onsite 
Certified Nuclear Medicine Technologist or Nuclear Medicine Physician. Subjects will be 
continuously monitored for AEs. 
7.2.2.3 
After Administration of Unfiltered Tc 99m Sulfur Colloid 
The following procedures will be completed at the specified times: 
• 20 ± 5 Minutes After Administration 
o Assessment of AEs  
o Vital signs after at least 1 minute in a resting position (body temperature, heart 
rate, blood pressure, and respiratory rate) 
o Planar imaging of the liver/spleen followed by SPECT/CT of the liver/spleen 
and abdomen 
 

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7.2.3 
Visit 3 (Day 3) Tc 99m Tilmanocept Administration 
7.2.3.1 
Before Administration of Tc 99m Tilmanocept 
The following procedures will be completed on the day of injection prior to the administration 
of Tc 99m tilmanocept: 
• 
A urine pregnancy test for women of child-bearing potential. Females of child 
bearing potential are defined as women that are not surgically sterile (hysterectomy or 
bilateral oophorectomy) nor postmenopausal for at least 1 year prior to screening. 
Women who are not of childbearing potential will not require a pregnancy test. 
• 
Assessment of AEs 
• 
Concomitant medication review 
• 
Vital signs after at least 1 minute in a resting position (body temperature, heart rate, 
blood pressure, and respiratory rate) within 15 minutes prior to administration of Tc 
99m tilmanocept 
 
7.2.3.2 
Administration of Tc 99m Tilmanocept 
IV administration of Tc 99m tilmanocept will be at study time Day 3 Time 00:00. The preferred 
site of IV placement will be the left or right antecubital vein. 
All subjects will receive an open label 200 µg dose of Tc 99m tilmanocept at the specified 
radiolabel dose. The filled syringe will be connected to the IV for a slow push injection. At the 
completion of the injection, a 10 cc sterile normal saline flush will be administered.  
The IV administration will be performed in the nuclear medicine department by an onsite 
Certified Nuclear Medicine Technologist or Nuclear Medicine Physician. Subjects will be 
continuously monitored for AEs. 
7.2.3.3 
After Administration of Tc 99m Tilmanocept 
The following procedures will be completed at the specified times:  
• 
20 ± 5 Minutes After Administration: 
o Assessment of AEs  
o ECG (completed before vital signs) 
o Vital signs after at least 1 minute in a resting position (body temperature, heart rate, 
blood pressure, and respiratory rate)  
o Planar imaging of the liver/spleen followed by SPECT/CT of the liver/spleen and 
abdomen 
 
• 
75 ± 15 Minutes After Administration: 
o Assessment of AEs  

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o Planar imaging of the liver/spleen followed by SPECT/CT of the liver/spleen and 
abdomen 
 
• 
180 ±15 Minutes After Administration: 
o Assessment of AEs  
o Planar imaging of the liver/spleen followed by SPECT/CT of the liver/spleen and 
abdomen 
 
• 
After Completion of All Imaging: 
o Blood draw for hematology and chemistry testing 
o Urine collection for urinalysis  
 
7.2.4 
Safety Follow Up Telephone Call (Day 8 ± 3) 
A safety follow-up telephone call will be performed 5 ± 3 days after administration of Tc 
99m tilmanocept (Day 8 ± 3): 
o Concomitant medication review 
o Assessment and review of AEs 
 

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8 
PROCEDURES AND VARIABLES 
8.1 
Population Characteristics 
8.1.1 
Demographic and Other Baseline Characteristics 
Up to six (6) female or male subjects with biopsy-confirmed moderate to severe NASH will 
be enrolled.  Up to six (6) female or male subjects without evidence of any liver or 
inflammatory diseases will be enrolled. There are no minimum or maximum numbers for 
gender in the groups.  
8.1.2 
Medical and Surgical History 
Relevant medical and surgical histories will be obtained on all study subjects. For NASH 
subjects, this will include a history of all NASH treatment.   
As part of the medical history, the date of the last spontaneous menstruation will be recorded, 
if childbearing potential is not excluded by surgical sterilization. 
8.1.3 
Prior and Concomitant Medication 
All prior medications used within the last 30 days before the first screening examination 
through the 5 ± 3 day post-tilmanocept injection (Day 8 ± 3) follow-up safety telephone call 
will be documented.  
8.2 
SPECT/SPECT-CT Image Acquisition  
The camera used to obtain the images should be a dual-headed SPECT or SPECT/CT camera 
equipped with a low-energy, high-resolution collimator with a 15% window (20% can be used 
if 15% setting not available) centered over a 140keV peak.   
The camera must have passed the daily SPECT QC tests as per the manufacturer’s 
recommendation for that day’s scan schedule. 
Whenever possible, subjects should be asked to void after injection and prior to the first 
imaging session and again between imaging sessions.  
Subjects should be positioned supine with arms at their side. The subject’s head, legs, hands, 
and hips should be in the true anterior position. The subject should be instructed to remain as 
motionless as possible during the scan, and asked to breathe as shallowly as comfortable when 
the camera is passing over the chest. To help minimize leg motion, a light taping around the 
ankles can be used to keep the legs together. A pillow under the knees is often very helpful for 
comfort. 
A planar scan of the liver followed by SPECT/CT should be obtained at the following time 
points following administration of the specified product: 

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Unfiltered Tc 99m Sulfur Colloid: 
• 
20 ± 5 minutes post administration 
 
Tc 99m Tilmanocept: 
• 
20 ± 5 minutes post administration 
• 
75 ± 15 minutes post administration 
• 
180 ± 15 minutes post administration 
 
It is anticipated that the planar scan would be acquired for 25-30 minutes. Using state-of-the-
art 2-headed cameras (nominal 20” x 15” FOV), a target of 5-7 million counts should be 
obtained for the summed view of both heads. For higher activity injections, it may be possible 
to attain this target in a shorter time, allowing the subject more time off the bed between scans. 
A simple measure of the subject’s “thickness” (i.e. from a lateral visual view, the height of the 
subject’s anterior-most chest/abdomen above the scan bed) should be documented. This will 
be used for the attenuation correction to be applied to the anterior/posterior views obtained 
from the planar scan. 
Following completion of the the planar imaging session at the designated time points, a 3D 
SPECT/CT should be obtained on the liver, spleen, and abdomen, followed by a patient break.  
It is anticipated that each planar or SPECT/CT scan would be acquired for 25-30 minutes (30 
minutes preferred). A break between scans should address the comfort of the patient, 
encouraging the subject to void, consume fluids or food and to move or rest freely.  It is very 
important that the subject remain motionless for the duration of each scan. 
The SPECT transverse slices will be reconstructed using the manufacturer’s recommended 
reconstruction algorithms including the recommended attenuation correction. 
It is strongly recommended that each site should perform all imaging acquisitions per their 
camera manufacturer and model parameters and in accordance with their institutional practices 
for both the planar imaging as well as the SPECT/CT imaging acquisitions. 
De-identified DICOM (Digital Imaging Communications in Medicine) copies of all acquired 
images will be transmitted to Navidea Biopharmaceuticals. The areas of localization will be 
identified, documented and captured in the CRFs. 
8.3 
Pharmacokinetics 
No pharmacokinetic investigation will be performed in this study.  

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8.4 
Safety  
8.4.1 
Adverse Events 
8.4.1.1 
Definition of Adverse Event 
The definitions below follow International Conference on Harmonization (ICH) – Good 
Clinical Practice (GCP) (see also ICH Guideline for Clinical Safety Data Management:  
Definitions and Standards for Expedited Reporting). 
Adverse Event (AE) 
An AE is defined as any untoward medical occurrence in a subject administered a 
pharmaceutical product and which does not necessarily have a causal relationship with this 
treatment.  An AE can therefore be any unfavorable and unintended sign (including an 
abnormal laboratory finding), symptom, or disease temporally associated with the use of a 
medicinal (investigational) product, whether or not considered related to the medicinal 
(investigational) product.  
Any clinically significant change in a condition (worsening) from screening that results in a 
change in subject management will be considered an AE and will be recorded on the AE page 
of the CRF. 
By definition for this study, all untoward medical occurrences beginning on the day of Visit 1 
Screening (Days -45 to -1) through the assessment of the Safety Follow-up Telephone Call 
(Day 8 ± 3) are to be reported as AEs.  AEs continuing after study completion will be followed 
to normalization or stabilization.  SAEs will be reported from the time of consent through the 
end of participation. 
8.4.1.2 
Categories for Adverse Event Assessment 
All AEs will be assessed and documented by the investigator according to the categories 
detailed below. 
Seriousness 
For each AE, the seriousness must be determined according to the criteria given in 
Section 8.4.1.5. 

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Severity 
The severity of an AE is classified according to the following categories, taking into account 
the possible range of the intensity of the event: 
• 
Mild - 
The AE is transient and easily tolerated by the subject. 
• 
Moderate -  The AE causes the subject discomfort and interrupts the subject’s usual 
activities 
• 
Severe -  
The AE causes considerable interference with the subject’s usual 
activities and may be incapacitating or life-threatening. 
 
Specific drug treatment 
Any specific drug treatment will be documented.  
Causal relationship to investigational product 
The investigator will use the following definitions to assess the relationship of the AE to the 
use of investigational product:  
Definitely related: 
Event can be fully explained by administration of the investigational 
product. 
Probably related: 
Event is most likely to be explained by administration of the 
investigational product rather than the subject’s clinical state or 
other agents/therapies. 
Possibly related: 
Event may be explained by administration of the investigational 
product or by the subject’s clinical state or other agents/therapies. 
Probably not related: 
Event is most likely to be explained by the subject’s clinical state or 
other agents/therapies, rather than the investigational product. 
Definitely not related: 
Event can be fully explained by the subject’s clinical state or other 
agents/therapies. 
For causality assessments, events meeting the categories of definitely, probably, or possibly 
related will be considered to be related to investigational product. 
Causal relationship to study procedure 
The investigator will use the following definitions to assess the relationship of the AE to the 
study procedure:  

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Definitely related: 
Event can be fully explained by the study procedure. 
Probably related: 
Event is most likely to be explained by the study rather than the 
subject’s clinical state or other agents/therapies. 
Possibly related: 
Event may be explained by the study procedure or by the subject’s 
clinical state or other agents/therapies. 
Probably not related: 
Event is most likely to be explained by the subject’s clinical state or 
other agents/therapies, rather than the study procedure. 
Definitely not related: 
Event can be fully explained by the subject’s clinical state or other 
agents/therapies. 
For causality assessments, events meeting the categories of definitely, probably, or possibly 
related will be considered to be related to study. 
8.4.1.3 
Assessments and Documentation of Adverse Events 
Attention shall be paid to the occurrence of AEs for the duration of subject participation. 
Untoward medical events beginning on Visit 1 Screening (Days -45 to -1) through the 
completion of the Follow-up Telephone Call will be reported as AEs.  Thus, subjects should 
be closely observed by the investigator both during and after the evaluation. 
Any AE (observed, volunteered, or elicited) should be recorded in detail in the source 
documentation. 
The following information is required: 
• 
The date and time of onset of any AE. 
• 
The duration (the entire duration of an event or symptom, calculated from date of 
onset to date of end, if not recorded directly). 
• 
The seriousness of the AE will be assessed by the investigator.  If the investigator 
deems that an AE qualifies as an SAE, a special form provided by the sponsor should 
be completed and the event must be immediately reported to the sponsor.  A 
definition of SAEs is provided in Section 8.4.1.5.   
• 
The maximum intensity (mild, moderate, or severe). 
• 
Whether drug treatment was administered for the event, any specific drug treatment 
must be documented. 
• 
The relationship of the AE to the investigational product and to study conduct (for 
definitions, see above). 
The outcome of the AE (resolved, resolved with sequelae, not resolved, unknown, 
death). 
 
AEs will be coded according to an internationally recognized dictionary (Medical Dictionary 
for Regulatory Activities [MedDRA]). 

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8.4.1.4 
Expected Adverse Events 
Investigational Product-Related Risks 
In all completed studies of Tc 99m tilmanocept, involving 575 subjects, only three events 
(breast pain and injection site pain reported by subjects with breast cancer and injection site 
irritation reported by a subject with head and neck squamous cell cancer) were deemed 
definitely related to the administration of Tc 99m tilmanocept by the investigator. The most 
commonly reported adverse reactions have been lack of effect (<0.067%), injection site pain 
(<0.02%) and rash (<0.02%). AEs from the radioactive dose are not expected, since the applied 
radiation doses are far below doses that can cause acute effects in human tissues.  
Post-marketing surveillance shows that Tc 99m tilmanocept (Lymphoseek®) has been 
administered to more than 200,000 patients with not a single drug-related SAE.  Routes of 
administration have included: subcutaneous, intradermal, and peritumoral. The intended route 
of administration in this study is IV.   
Precautionary Measures 
Special precautionary measures are not considered necessary for this study.  In case of 
emergency, standard emergency procedures will be employed. 
Unexpected Adverse Events 
An unexpected AE is defined as an adverse reaction that in nature and severity is not consistent 
with the applicable product information (e.g., Investigator’s Brochure).  Any adverse 
experience that is not listed in the current Investigator’s Brochure or which is, with regard to 
the specificity or severity, not consistent with the risk information shall be regarded as 
unexpected. 
 
Examples would be (a) acute renal failure listed in the Investigator’s Brochure with a 
subsequent new report of interstitial nephritis and (b) hepatitis with a first report of fulminant 
hepatitis.  “Unexpected” as used in this definition refers to an adverse drug experience that has 
not been previously observed and included in the Investigator’s Brochure, rather than from the 
perspective of such experience not being anticipated from the pharmacological properties of 
the investigational product. 
8.4.1.5 
Serious Adverse Events 
Definition of Serious Adverse Events 
Definition 
The following SAE definition is based on ICH guidelines and the final rule issued by the FDA 
and effective 06 Apr 1998. 

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An SAE is classified as any untoward medical occurrence that at any dose: 
• 
results in death, or 
• 
is life threatening, or 
• 
requires inpatient hospitalization or prolongation of existing hospitalization, or 
• 
results in persistent or significant disability/incapacity, or 
• 
is a congenital anomaly/birth defect, or 
• 
is an important medical event (see paragraphs below). 
 
The term ‘life threatening’ in the definition refers to an event in which the subject was at risk 
of death at the time of the event; it does not refer to an event that hypothetically might have 
caused death if it were more severe. 
Medical and scientific judgment should be exercised in deciding whether it is appropriate to 
report an AE as serious also in other situations, such as important medical events that may not 
be immediately life threatening or result in death or hospitalization but may jeopardize the 
subject or may require intervention to prevent one of the other outcomes listed in the definition 
above.  These should also usually be considered serious.  Examples of such events are intensive 
treatment in an emergency room or at home for allergic bronchospasm or blood dyscrasias or 
convulsions that do not result in subject hospitalization. 
Actions and reporting obligations in case of serious adverse events 
The investigator should take appropriate diagnostic and therapeutic measures to minimize the 
risk to the subject.   
If any SAE occurs over the course of the study, investigators or other site personnel will inform 
Navidea Biopharmaceutical representatives within one day (i.e., within 24 hours) of becoming 
aware of the SAE. Written notification of the SAE will be emailed to Navidea 
Biopharmaceuticals Pharmacovigilance at safety@navidea.com. For fatal or life-threatening 
AEs where important or relevant information is missing, active follow-up is undertaken 
immediately. 
Pregnancy will have the same time reporting obligations to the sponsor as SAEs. Upon 
notification, Navidea will provide a form for collection of pregnancy information. 
All SAEs must also be recorded on the AE CRFs. 
Notification of the IRB(s) 
The sponsor and/or the investigator will notify the IRB(s) about all relevant events (e.g., SAEs 
and Suspected, Unexpected, Serious Adverse Reactions [SUSARs]) according to all applicable 
regulations. 

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Notification of the authorities 
The sponsor will process and report all relevant events (e.g., SAEs, SUSARs) to the authorities 
according to all applicable regulations. 
Sponsor’s notification of the investigators 
The sponsor will inform all investigators about reported relevant events (e.g., SAEs, SUSARs) 
according to all applicable regulations. 
8.4.2 
Further Safety Assessments 
8.4.2.1 
Physical Exam  
Complete physical examinations will be conducted according to the Schedule of Study Events 
(see Appendix 1). Height and body weight will be collected at Visit 1 (Screening).  
Physical examinations will be performed for the following body systems: 
• 
General Appearance 
• 
Skin/dermatological 
• 
Eyes, ears, nose, throat 
• 
Head and neck (including thyroid) 
• 
Lungs 
• 
Heart 
• 
Abdomen (liver, kidney, spleen, gastrointestinal) 
• 
Musculoskeletal 
• 
Nervous system 
 
8.4.2.2 
Vital Signs 
Vital signs comprise the measurement of body temperature, heart rate, and respiration, systolic 
and diastolic blood pressure. All measurements will be taken after the subject has been in a 
resting position for at least 1 minute.  
Vital signs will be measured at screening, within 15 minutes before each product 
administration, and 20 ± 5 minutes after each product administration.   
Any clinically significant change from screening (worsening) that results in a change in subject 
management will be considered an AE and will be recorded on the AE page of the CRF. 
8.4.2.3 
Electrocardiogram 
A standard 12-lead ECG will be obtained at screening and 20 ± 5 minutes following Tc 99m 
tilmanocept administration. The ECG will be measured with the subject in a resting position 
for at least 1 minute. No continuous ECG monitoring will be required.  At a minimum the heart 

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rate, QRS, PR and QT intervals will be collected.  QTc will be calculated using both the 
Fridericia and Bazett formulas. 
On-site investigator’s responsibilities 
The immediate cardiac safety of the subject will be ensured by the on-site qualified physician.  
Any 12-lead ECG intervals, waveform abnormalities, and rhythm changes that are clinically 
significant in that they result in a change in subject management will be considered an AE.  In 
the case of an SAE, once SAE notification is decided upon, investigators are required to follow 
the procedure described for SAE notification and document abnormal ECG findings (intervals 
and waveforms).  Any interval data or abnormal waveform finding that resulted in an AE (i.e., 
change of patient management) must be followed to normalization or stabilization.  
Each 12-lead ECG tracing must be signed and dated and stored in the subject’s source 
documentation.   
8.4.2.4 
Clinical Laboratory Parameters for Screening and Safety 
Clinical laboratory tests to be evaluated in this study include hematology, serum chemistry, 
and urinalysis.  Clinical laboratory tests will include the following as defined in Table 2. 
Table 2. 
Clinical Laboratory Parameters 
Hematology 
Hemoglobin (Hgb), hematocrit (HCT), platelets, neutrophils, 
basophils, lymphocytes, monocytes, red blood cells (RBC), white 
blood cells (WBC) 
Serum chemistry 
Aspartate aminotransferase (AST), alanine aminotransferase 
(ALT), alkaline phosphatase, total bilirubin, creatinine, chloride, 
hemoglobin A1c (screening only), potassium, sodium, total 
protein, albumin, carbon dioxide (CO2)/bicarbonate, blood urea 
nitrogen (BUN), glucose  
Urinalysis 
pH, specific gravity 
 
All laboratory reports must be promptly reviewed for clinical significance by the investigator, 
and upon review, initialed and dated by the investigator. 
Good clinical practice would suggest that a copy of the laboratory results also be provided to 
the subject’s referring physician. 
Any change in a laboratory value, which results in a change in subject management (additional 
controls or treatment required), will be reported as a clinically significant change. Clinically 
significant changes in laboratory parameters, which are not the result of laboratory error, are 
to be recorded as AEs. 

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Any clinically significant changes in laboratory values are to be followed up with repeated 
tests at appropriate intervals (as determined by the investigator) until the values return to 
baseline level or until the abnormality is explained by the investigator.  
The expected amount of blood to be withdrawn is shown in Table 3. 
Table 3 
Approximate Amount of Blood Withdrawn 
Time point of examination 
 
Blood Volume 
Screening: Laboratory examination  
Chemistry    5 mLs 
Hematology 4 mLs 
 
 
9 mL (~2 tsp) 
Post Tilmanocept Injection and 
Imaging: Laboratory examination 
Chemistry    5 mLs 
Hematology 4 mLs 
 
 
9 mL (~2 tsp) 
TOTAL: 
 
18 mL (~4 tsp) 
 

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9 
STATISTICAL METHODS 
This is a prospective, open-label, multicenter, safety, comparative study of injected Tc 99m 
tilmanocept and unfiltered Tc 99m sulfur colloid in the detection of and assessment of 
localization (tessellation discrimination) to the liver in subjects with and without moderate to 
severe NASH by planar and SPECT/CT imaging.  The statistical objective of the study is to 
determine the safety of Tc 99m tilmanocept and to compare Tc 99m tilmanocept localization 
(tessellation characteristics) to unfiltered Tc 99m sulfur colloid between subjects with and 
without NASH. 
Exploratory contrasts with available elastography and Tc99m tilmanocept images may be  
completed. 
A study center is defined as a treatment administration site or group of treatment administration 
sites under the control and supervision the same PI.   
9.1 
Randomization Methods 
The study is not randomized. 
9.2 
Safety Variables 
The primary variable for this study is whether a subject observes any noxious pharmacologic 
activity/adverse drug reaction (ADR) during their enrollment in the trial.  This is a safety 
endpoint.  The pharmacologic activity or ADR must be observed after the dose of Tc99m 
tilmanocept is administered. 
NOTE: Adverse Drug Reaction 
An ADR includes all noxious and unintended responses to a medicinal product related to any 
dose or dose regimen. The phrase “responses to a medicinal product” means that a casual 
relationship between a medicinal product and an AE is at least a reasonable possibility, i.e. 
the relationship cannot be ruled out. 
9.3 
Efficacy Variables 
The efficacy variables for this study are as follows: 
• Liver uptake of unfiltered Tc99m-labeled sulfur colloid and tilmanocept; and, 
• Intrahepatic distribution (tessellation) of Tc99m-labeled sulfur colloid and tilmanocept 
9.4 
Sample Size Justification 
The sample size of N=12 subjects is deemed sufficient to accomplish the Sponsor’s 
objectives of determining the safety of IV tilmanocept and localization patterns of 
tilmanocept and sulfur colloid in NASH and asymptomatic subjects. 

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9.5 
Statistical Analyses 
9.5.1 
Analysis Populations 
The following analysis population will be defined for the study: 
• 
Intent-to-Diagnose (ITD) Population - Subjects who are enrolled in the study, 
administered Tc 99m tilmanocept and unfiltered Tc 99m sulfur colloid, and are 
imaged will be included in the ITD analysis population. 
• 
Safety Population – All patients who are enrolled and administered Tc 99m 
tilmanocept and unfiltered Tc 99m sulfur colloid in the study will be included in the 
safety population. 
 
All safety data analyses will be conducted on the safety population.  All efficacy data analyses 
will be conducted on the ITD population. 
9.5.2 
Analysis of Baseline and Demographic Characteristics 
The distribution of each baseline and demographic variable of interest will be summarized by 
dose group. Continuous variables will be summarized via mean, median, standard deviation, 
and range. Categorical variables will be summarized via counts and percentages. 
9.5.3 
Analysis of Efficacy Variables 
Co-location of tilmanocept and sulfur colloid intrahepatic tessellation localization (Figure 7) 
will be assessed through frequency tables derived by creating virtual slices of the liver.. These 
scans will be false-colored and overlaid, matching landmarks (e.g. large blood vessels) for 
image registration. The resulting voxel colors will be classified as binding tilamanocept, 
binding sulfur colloid, or binding both. No formal hypothesis tests are planned. Frequency 
tables will created for individual subjects and overall. 
The voxel radiation intensities will be used to create non-parametric density estimates of 99mTc 
gamma radiation intensities for both Tc 99m tilmanocept and Tc 99m sulfur colloid. Density 
estimates will be created for both individual subjects and for NASH subjects and healthy 
controls pooled separately. 
 
Figure 7. Diagrmatic representation of anticipated tessellation 

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Complete analyses and details will be described in the Statistical Analysis Plan (SAP) for the 
study. 
9.5.4 
Analysis of Safety Variables 
The number and percentage of subjects with any noxious pharmacologic activity or ADRs will 
be tabulated by dose group and overall. 
All AEs will be observed for each subject from the time of signing of informed consent until 
exit from the study.  A treatment emergent AE is defined as an AE whose start date is on or 
after the procedure date.  If the procedure date and/or the AE start date are missing, the AE 
will be considered treatment emergent.  
Prior to analysis, all AEs will be coded using the MedDRA coding dictionary. Based on these 
coded terms, treatment emergent AEs will be summarized by dose group and overall as 
follows: 
• 
by system organ class and preferred terms 
• 
by system organ class and preferred terms and relation to study drug 
• 
by system organ class and preferred terms and severity 
 
Observed and change from baseline vital sign parameters, ECG parameters and hematology, 
clinical chemistry and urinalysis parameters will be summarized using descriptive statistics 
(mean, median, standard deviation, minimum, maximum and range) at each time point by dose 
group and overall.  
Other safety analyses will be described in the SAP for the study. 
9.5.5 
Handling Missing Values 
The analysis of the primary efficacy variables will be carried out on the observed data, i.e., a 
complete case analysis.     
9.5.6 
Interim Analysis 
There are no interim analyses planned for this study. 

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10 DATA HANDLING AND QUALITY ASSURANCE 
10.1 Data Recording 
Data required according to this protocol is captured in the subject’s source documentation and 
are to be recorded in the CRFs (provided by the sponsor) as soon as possible. 
10.1.1 
CRF Design 
Paper CRFs will be used for collecting all data generated during the trial. CRF completion 
details will be documented in a separate document that will be provided by the sponsor and 
maintained in the TMF. 
10.2 Monitoring 
This study will be monitored regularly by a clinical research associate (CRA) from the sponsor 
or a contract research organization (CRO).  Monitoring procedures include one or more visits 
designed to clarify all prerequisites before the study starts.  Interim monitoring visits will take 
place on a regular basis according to a schedule fixed by mutual agreement.  During these 
visits, the CRA will check for completion of the entries on the CRFs, their compliance with 
the protocol and with GCP, and will compare the CRF entries with the source data. 
All data recorded in the CRF will be captured in the source documentation. 
The CRA will verify the correct use of the investigational product.  The investigational product 
will not be supplied to the investigator site prior to a favorable opinion from the IRB and the 
regulatory authority and, if appropriate, from the radiation protection authorities. In 
addition, the CRA will determine whether all AEs and SAEs have been appropriately reported 
(including adherence to the time periods required for SAEs). 
10.3 Data Processing 
Study data documentation will be maintained specifying all relevant aspects of data processing 
for the study (including data validation, cleaning, correcting, releasing).  This documentation 
will be stored in the TMF. 
For data coding (e.g., AEs, medication, medical/surgical history), internationally recognized 
and accepted dictionaries will be used.  These and the processes used for coding will be 
specified in the data management plan. 
10.4 Auditing 
A member of the sponsor’s (or a designated CRO) quality assurance unit may arrange to visit 
the investigator in order to audit the performance of the study at the study site and the study 
documents originating there.  The auditor(s) will usually be accompanied by a CRA or the 
study team leader.  The investigator will be informed about the outcome of the audit. 

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In addition, inspections by health authority representatives and IRB(s) are possible at any time.  
The investigator is to notify the sponsor of any such inspection immediately. 
10.5 Archiving 
Essential documents shall be archived safely and securely in such a way that ensures that they 
are readily available upon authorities’ request.  Patient (hospital) files will be archived 
according to local regulations and in accordance with the maximum period of time permitted 
by the hospital, institution, or private practice.  Where the archiving procedures do not meet 
the minimum timelines required by the sponsor, alternative arrangements must be made to 
ensure the availability of the source documents for the required period. 
The investigator/institution notifies the sponsor if the archival arrangements change 
(e.g., relocation or transfer of ownership). 
The ISF is not to be destroyed without the sponsor’s approval. 
The investigator’s contract will contain all regulations relevant for the study center. 
10.6 Premature Termination of the Study 
Termination by the Sponsor 
The Sponsor may terminate the study at any time for any of the following reasons: 
1. Failure to enroll subjects 
2. Protocol violations 
3. Inaccurate or incomplete data 
4. Unsafe or unethical practices 
5. Questionable safety of the investigational product 
6. Suspected lack of efficacy of the investigational product 
7. Administrative decision 
 
Termination by the Investigator 
If the Investigator terminates the study prematurely, the Investigator must do the following: 
• 
Return all unused investigational products and related study materials to the Sponsor. 
• 
Provide the IRB(s) and the sponsor with a written statement describing why the study 
was terminated prematurely.  Prompt compliance with this requirement is essential so 
that the sponsor may comply with its regulatory obligations. 
 

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10.6.1 
Study as a Whole 
The sponsor retains the right to prematurely terminate the study as a whole at any time.   
At the discretion of the sponsor, the entire study may be canceled for medical reasons.  In 
addition, the sponsor retains the right to end the study at any time if the study cannot be carried 
out as agreed upon in the protocol. In case of early termination or suspension of the study, the 
principal investigator/sponsor will promptly inform the investigator/institutions, regulatory 
authorities, and IRB of the termination or suspension and the reason for that. 
10.6.2 
Center 
At any time, the study may be terminated at an individual center if:  
• 
The center cannot comply with the requirements of the protocol. 
• 
It is not possible for the center to comply with GCP standards. 
 
10.6.3 
Study Participant 
Individual subjects may be withdrawn from the study according to the criteria specified in 
Section 4.3. 
 

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11 ETHICAL AND LEGAL ASPECTS 
11.1 Ethical and Legal Conduct of the Study 
The planning and conduct of this clinical study are subject to national laws. Only when all of 
the requirements of the appropriate regulatory authority have been fulfilled will the study 
begin. The study will be conducted in accordance with the ethical principles that have their 
origin in the Declaration of Helsinki and the ICH-GCP Guidelines of 17 Jan 1997. At the 
discretion of the investigator, the entire study may be canceled for medical reasons. In addition, 
the sponsor retains the right to end the study for medical-scientific or GCP-relevant reasons. 
In case of premature termination, the investigators, IRB(s) and Regulatory Authorities will be 
informed by the Study Manager. As required by local law, current safety-relevant information 
will be provided to the IRB(s) and the regulatory authorities by the sponsor. The sponsor will 
also inform all investigators about relevant safety events according to the applicable 
regulations. 
11.2 Subject Information and Consent 
All relevant information on the study will be summarized in the subject consent form and 
additionally as required by the investigator’s institution in an integrated subject information 
and consent sheet. A sample informed consent form (ICF) is provided as a document separate 
to this protocol. 
Based on this subject ICF, the investigator will explain all relevant aspects of the study to each 
subject, before entry into the study (i.e., before examinations and procedures associated with 
selection for the study are performed).  
The investigator will also mention that written approval of the IRB has been obtained.  Each 
subject will have ample time and opportunity to ask questions and will be informed about the 
right to withdraw from the study at any time without any disadvantage and without having to 
provide reasons for this decision.  Following this informative discussion, the subject will be 
asked if he/she is willing to sign and personally date a statement of informed consent.  Only if 
the subject voluntarily agrees to sign the ICF and has done so, may he/she enter the study.  
Additionally, the investigator or his/her designee will personally sign and date the form.  The 
subject will receive a duplicate of the signed and dated form.  
The investigator will record in the source documentation the consent process including the 
time and date of obtaining informed consent. In the event that informed consent is obtained on 
the date that baseline study procedures are performed, the study record or subject's clinical 
record must clearly show that informed consent was obtained prior to these procedures.  
The ICF and any other written information provided to subjects will be revised whenever 
important new information becomes available that may be relevant to the subject’s consent, or 
there is an amendment to the protocol which necessitates a change to the content of the subject 
information and/or the written ICF. The investigator will inform the subject of changes in a 
timely manner and will ask the subject to confirm his/her participation in the study by signing 

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the revised ICF. Any revised written ICF and written information must receive the IRB’s 
approval/favorable opinion in advance of use. 
11.3 Financing/Financial Disclosure 
Each investigator (including principal and/or any subinvestigators; as well as their spouses and 
dependent children) who is directly involved in the treatment or evaluation of research subjects 
has to provide a financial disclosure according to all applicable legal requirements. All relevant 
documentation will be filed in the sponsor TMF and the ISF, as appropriate. 
11.4 Publication Policy 
The sponsor will be responsible for determining when any trial results should be published.  
The sponsor will work jointly with the investigator(s) to publish information in a timely 
manner.  The investigator(s) shall not submit any information gleaned under the direct support 
or sponsorship of the sponsor to journals or professional societies without the prior written 
approval of the sponsor.  A “publication” is meant to include any abstract, letter, manuscript 
or public announcement in any form or length that contains information gleaned under the 
direct support or sponsorship of the sponsor. 
11.5 Subject Injury 
In general, if a subject is injured as a direct result of the investigational product but not due to 
medical negligence on the part of the principal investigator or study staff, the sponsor will pay 
for reasonable and necessary medical treatment for the injury, to the extent the expenses are 
not covered by the subject’s medical insurance, a government program, or other responsible 
third party.  If laws or regulations of the locality in which the study is taking place require 
additional payment of expenses, the sponsor shall comply with such law or regulation.  Where 
applicable, the sponsor has taken specific national insurance. 

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12 REFERENCE LIST 
1. 
Klingensmith, W.C., 3rd, et al., Normal appearance and reproducibility of liver-spleen 
studies with Tc-99m sulfur colloid and Tc-99m microalbumin colloid. J Nucl Med, 
1983. 24(1): p. 8-13. 
2. 
Nalbantoglu, I.L. and E.M. Brunt, Role of liver biopsy in nonalcoholic fatty liver 
disease. World J Gastroenterol, 2014. 20(27): p. 9026-37. 
3. 
Lazaridis, N. and E. Tsochatzis, Current and future treatment options in non-alcoholic 
steatohepatitis (NASH). Expert Rev Gastroenterol Hepatol, 2017. 11(4): p. 357-369. 
4. 
Caligiuri, A., A. Gentilini, and F. Marra, Molecular Pathogenesis of NASH. Int J Mol 
Sci, 2016. 17(9). 
5. 
Oseini, A.M. and A.J. Sanyal, Therapies in non-alcoholic steatohepatitis (NASH). 
Liver Int, 2017. 37 Suppl 1: p. 97-103. 
6. 
Sanyal, A.J., et al., Endpoints and clinical trial design for nonalcoholic steatohepatitis. 
Hepatology, 2011. 54(1): p. 344-53. 
7. 
Sancilio. Non-Alcoholic Fatty Liver Disease and Non-Alcoholic Steatohepatitis – 
SC410 2017  [cited 2017 August 25]; Available from: https://www.sancilio.com/non-
alcoholic-fatty-liver-disease-nash/. 
8. 
Charytoniuk, T., et al., Alternative treatment methods attenuate the development of 
NAFLD: A review of resveratrol molecular mechanisms and clinical trials. Nutrition, 
2017. 34: p. 108-117. 
9. 
Manns, M.P., Liver cirrhosis, transplantation and organ shortage. Dtsch Arztebl Int, 
2013. 110(6): p. 83-4. 
10. 
Parikh, N.D., et al., Projected increase in obesity and non-alcoholic steatohepatitis-
related liver transplantation waitlist additions in the United States. Hepatology, 2017. 
11. 
Younossi, Z.M., et al., Global epidemiology of nonalcoholic fatty liver disease-Meta-
analytic assessment of prevalence, incidence, and outcomes. Hepatology, 2016. 64(1): 
p. 73-84. 
12. 
Nugent, C. and Z.M. Younossi, Evaluation and management of obesity-related 
nonalcoholic fatty liver disease. Nat Clin Pract Gastroenterol Hepatol, 2007. 4(8): p. 
432-41. 
13. 
Kabbany, M.N., et al., Prevalence of Nonalcoholic Steatohepatitis-Associated 
Cirrhosis in the United States: An Analysis of National Health and Nutrition 
Examination Survey Data. Am J Gastroenterol, 2017. 112(4): p. 581-587. 
14. 
Miura, K., et al., Hepatic recruitment of macrophages promotes nonalcoholic 
steatohepatitis through CCR2. Am J Physiol Gastrointest Liver Physiol, 2012. 302(11): 
p. G1310-21. 
15. 
Krenkel, O. and F. Tacke, Liver macrophages in tissue homeostasis and disease. Nat 
Rev Immunol, 2017. 17(5): p. 306-321. 
16. 
Virgilio A. Valdez, R.N.K., Thomas W. Crucitti and Nilo E. Herrera, Case Reports: 
Nonvisualization of the Liver with Tc-99m Sulfur Colloid. J. Nucl. Med. Technol., 
1978. 6(3): p. 161-163. 
17. 
Al-Saeedi, F. and I. Loutfi, (99m)Tc sulfur colloid and (99m)Tc mebrofenin 
hepatobiliary functional liver imaging in normal and diabetic rats. Med Princ Pract, 
2011. 20(2): p. 129-32. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
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18. 
Sofroniou, T. Digestive System. 2012  [cited 2012 24 Decemeber]; Available from: 
http://tsbiomed.blogspot.com/2012/12/digestive-system.html. 
19. 
Prieto Valtũena, J.s.M., J. Rodë s, and D.A. Shafritz, Hepatobiliary diseases. 1992, 
Berlin ; New York: Springer-Verlag. xxv, 1128 p. 
20. 
Takahashi, Y. and T. Fukusato, Histopathology of nonalcoholic fatty liver 
disease/nonalcoholic steatohepatitis. World J Gastroenterol, 2014. 20(42): p. 15539-
48. 
21. 
Sleyster, E.C. and D.L. Knook, Relation between localization and function of rat liver 
Kupffer cells. Lab Invest, 1982. 47(5): p. 484-90. 
22. 
Dooley, S. and P. ten Dijke, TGF-beta in progression of liver disease. Cell Tissue Res, 
2012. 347(1): p. 245-56. 
23. 
Nguyen-Lefebvre, A.T. and A. Horuzsko, Kupffer Cell Metabolism and Function. J 
Enzymol Metab, 2015. 1(1). 
24. 
Hamer, O.W., et al., Fatty liver: imaging patterns and pitfalls. Radiographics, 2006. 
26(6): p. 1637-53. 
25. 
Lapadat, A.M., et al., Non-invasive imaging techniques in assessing non-alcoholic fatty 
liver disease: a current status of available methods. J Med Life, 2017. 10(1): p. 19-26. 
26. 
Hennedige, T. and S.K. Venkatesh, Advances in computed tomography and magnetic 
resonance imaging of hepatocellular carcinoma. World J Gastroenterol, 2016. 22(1): 
p. 205-20. 
27. 
Chakraborty, D., et al., Role of Tc99m sulfur colloid scintigraphy in differentiating non-
cirrhotic portal fibrosis from cirrhosis liver. Indian J Nucl Med, 2010. 25(4): p. 139-
42. 
28. 
Taylor, M.E. and K. Drickamer, Structural requirements for high affinity binding of 
complex ligands by the macrophage mannose receptor. J Biol Chem, 1993. 268(1): p. 
399-404. 
29. 
Wallace, A.M., et al., Sentinel lymph node accumulation of Lymphoseek and Tc-99m-
sulfur colloid using a "2-day" protocol. Nucl Med Biol, 2009. 36(6): p. 687-92. 
30. 
Leong, S.P., et al., A phase 2 study of (99m)Tc-tilmanocept in the detection of sentinel 
lymph nodes in melanoma and breast cancer. Ann Surg Oncol, 2011. 18(4): p. 961-9. 
31. 
Tokin, C.A., et al., The efficacy of Tilmanocept in sentinel lymph mode mapping and 
identification in breast cancer patients: a comparative review and meta-analysis of the 
(9)(9)mTc-labeled nanocolloid human serum albumin standard of care. Clin Exp 
Metastasis, 2012. 29(7): p. 681-6. 
32. 
Sondak, V.K., et al., Combined analysis of phase III trials evaluating 
[(9)(9)mTc]tilmanocept and vital blue dye for identification of sentinel lymph nodes in 
clinically node-negative cutaneous melanoma. Ann Surg Oncol, 2013. 20(2): p. 680-8. 
33. 
Wallace, A.M., et al., Comparative evaluation of [(99m)tc]tilmanocept for sentinel 
lymph node mapping in breast cancer patients: results of two phase 3 trials. Ann Surg 
Oncol, 2013. 20(8): p. 2590-9. 
34. 
Marcinow, A.M., et al., Use of a novel receptor-targeted (CD206) radiotracer, 99mTc-
tilmanocept, and SPECT/CT for sentinel lymph node detection in oral cavity squamous 
cell carcinoma: initial institutional report in an ongoing phase 3 study. JAMA 
Otolaryngol Head Neck Surg, 2013. 139(9): p. 895-902. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date: 23 April 2018 
 
Confidential 
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35. 
Surasi, D.S., J. O'Malley, and P. Bhambhvani, 99mTc-Tilmanocept: A Novel Molecular 
Agent for Lymphatic Mapping and Sentinel Lymph Node Localization. J Nucl Med 
Technol, 2015. 43(2): p. 87-91. 
36. 
Kit for the Preparation of Technetium Tc 99m Sulfur Colloid Injection for 
Subcutaneous, Intraperitoneal, Intravenous, and Oral Use. Full Prescribing 
Information. http://www.sulfurcolloid.com/pdf/PI.pdf. 2012.  
 
 

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Clinical Study Protocol Number: NAV3-30 
 
Date:  23 April 2018 
 
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Appendix 1 
Schedule of Events 
Evaluation 
Visit 1 
Screening  
Visit 2  
Day 0 Tc 99m sulfur colloid Administration 
Days 
 -45 to -1 
Day 0 
Prior to 
Administration 
- 00:15 to -00:01 Mins 
00:00 
20 ± 5 Mins 
Informed Consent 
x 
 
 
 
 
Entry Criteria  
x 
 
 
 
 
Medical History, Demography 
x 
 
 
 
 
Vital Sign Assessment 
x 
 
x 
 
x 
ECG 
x 
 
 
 
 
Physical Examination  
x 
 
 
 
 
Clinical Laboratory Evaluation: 
Chemistry, Hematology and UA   
x 
 
 
 
 
Hemoglobin A1c Evaluation 
x 
 
 
 
 
Pregnancy Test 
xa 
xa 
 
 
 
Transient Elastography 
x 
 
 
 
 
Sulfur Colloid Administration 
 
 
 
x 
 
Tilmanocept Administration 
 
 
 
 
 
Planar and SPECT/CT Liver Imaging 
 
 
 
 
x 
Concomitant Medications 
x 
x 
 
 
 
Adverse Event Monitoring 
x 
x 
x 
x 
x 
a. 
Pregnancy test at screening will be serum; Day 0 and Day 3 pregnancy tests will be urine.  
b. Collection of samples for laboratory evaluation will occur after completion of all imaging procedures. 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date:  23 April 2018 
 
Confidential 
Page 69 of 71 
 
Evaluation 
Visit 3  
Day 3 Tc 99m tilmanocept Administration 
Follow-up 
Telephone 
Call 
Day 3 
Prior to 
Administration 
- 00:15 to -00:01 Mins 
00:00 
20 ± 5 Mins 
75 ± 15 
Mins 
180 ± 15 Mins 
Day 8 ± 3 
Informed Consent 
 
 
 
 
 
 
 
Entry Criteria  
 
 
 
 
 
 
 
Medical History, Demography 
 
 
 
 
 
 
 
Vital Sign Assessment 
 
x 
 
x 
 
 
 
ECG 
 
 
 
x 
 
 
 
Physical Examination  
 
 
 
 
 
 
 
Clinical Laboratory 
Evaluation: Chemistry, 
Hematology and UA   
 
 
 
 
 
xb 
 
Pregnancy Test 
xa 
 
 
 
 
 
 
Transient Elastography 
 
 
 
 
 
 
 
Sulfur Colloid Administration 
 
 
 
 
 
 
 
Tilmanocept Administration 
 
 
x 
 
 
 
 
Planar and SPECT/CT Liver 
Imaging 
 
 
 
x 
x 
x 
 
Concomitant Medications 
x 
 
 
 
 
 
x 
Adverse Event Monitoring 
x 
x 
x 
x 
x 
x 
x 
a. 
Pregnancy test at screening will be serum; Day 0 and Day 3 pregnancy tests will be urine. 
b. Collection of samples for laboratory evaluation will occur after completion of all imaging procedures.  

Tc 99m tilmanocept 
Clinical Study Protocol Number: NA V3-30 
Appendix 2 
Sponsor Signatures 
Navidea Biopharmaceuticals, Inc 
Date: 23 April 2018 
Study Title: 
An Evaluation of the Safety of Intravenous (IV) Tc 99m 
Tilmanocept and a Comparison of Localization to Tc 99m Sulfur 
Colloid by Planar and SPECT/CT Imaging in Subjects with 
Nonalcoholic Steatohepatitis (NASH) and Healthy Controls (HC) 
Study Number: 
Original Protocol 
Date: 
Amendment 1 Date: 
NAV3-30 
06 October 2017 
23 April 2018 
This clinical study protocol was subject to critical review and has been approved by the 
sponsor. The following per 
nel contributed to writing and/or approving this protocol: 
Signed: 
c[Y' 
Michael S. Blue, MD 
Senior Medical Director 
Navidea Biopharmaceuticals 
William J. Regan 
Senior VP, Global Regulator)". 
Navidea Biopharmaceuticals 
Confidential 
Page 70 of71 
Date: ~7·-M?R-&Olo 

Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-30 
 
Date:  23 April 2018 
 
Confidential 
Page 71 of 71 
 
Appendix 3 
Investigator’s Signature 
Study Title: 
An Evaluation of the Safety of Intravenous (IV) Tc 99m 
Tilmanocept and a Comparison of Localization to Tc 99m Sulfur 
Colloid by Planar and SPECT/CT Imaging in Subjects with 
Nonalcoholic Steatohepatitis (NASH) and Healthy Controls (HC) 
Study Number: 
NAV3-30 
Original Protocol 
Date: 
06 October 2017 
Amendment 1 Date: 
23 April 2018 
I have read the protocol described above. I agree to comply with all applicable regulations and 
to conduct the study as described in the protocol.  
 
 
Signed: 
 
 
Date: