Clinical trial NCT02865434
Conditions: Arthritis, Rheumatoid
Sponsor: Navidea Biopharmaceuticals
Start date: 2017-01-01; Completion date: 2018-06-01
Enrollment: 39 participants; Allocation: NON_RANDOMIZED; Intervention model: PARALLEL

Brief summary:
Prospective, open-label, multicenter, dose escalation, safety with pharmacokinetics (PK) and dosimetry study of injected Tc 99m tilmanocept in the detection of and assessment of localization to skeletal joints in subjects with and without active RA by SPECT imaging.

Detailed description:
A Manocept Platform prospective, open-label, multicenter, dose escalation, safety with PK and dosimetry study of injected Tc 99m tilmanocept in the detection of and assessment of localization to skeletal joints in subjects with and without active RA by SPECT imaging. All subjects will receive IV administration at one of 3 mass doses: 50 µg, 200 µg, or 400 µg. Within each mass dose group, subjects will receive Tc 99m tilmanocept labeled with one of 3 radiolabel doses: 1 mCi, 5 mCi, or 10 mCi.

All subjects will have a whole body planar SPECT scan. Subjects enrolled in Groups 1-9 will receive a whole body and planar hands scan followed by SPECT/CT scans on areas of interest post injection at 60 minutes ± 15 minutes and 180 minutes ± 15 minutes. Subjects enrolled in Groups 10-11 will receive a whole body planar SPECT scan performed at 4 specified time points post injection: 15 ± 5 minutes, 60 ± 15 minutes, 180 ± 15 minutes and 18-20 hours. Planar hand scans will be collected at 60 ± 15 minutes and 180 ± 15 minutes post-injection. PK blood sampling will be performed before injection (within 15 minutes), immediately following injection (within 5 minutes) and at each scanning timepoint. Dosimetry tests will be performed at each scanning timepoint. PK of urine will be assessed through counts of the bladder wall obtained from cumulative quantitative planar imaging from radiation dosimetry.

Arms: arm 1: Group 1 will receive 50 µg Tc99m-tilmanocept radiolabeled with 10 mCi Tc99m as a single IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 2: Group 2 will receive 200 µg Tc99m-tilmanocept radiolabeled with 10 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 3: Group 3 will receive 400 µg Tc99m-tilmanocept radiolabeled with 10 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 4: Group 4 will receive 50 µg Tc99m-tilmanocept radiolabeled with 5 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 5: Group 5 will receive 200 µg Tc99m-tilmanocept radiolabeled with 5 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 6: Group 6 will receive 400 µg Tc99m-tilmanocept radiolabeled with 5 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 7: Group 7 will receive 50 µg Tc99m-tilmanocept radiolabeled with 1 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 8: Group 8 will receive 200 µg Tc99m-tilmanocept radiolabeled with 1 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 9: Group 9 will receive 400 µg Tc99m-tilmanocept radiolabeled with 1 mCi Tc99m via IV injection. Subjects will receive SPECT Imaging (60 Minutes post-injection) and SPECT Imaging (180 Minutes post-injection). arm 10: Group 10 will receive Tc99m-tilmanocept at the MTD via IV injection, Whole body planar SPECT imaging (15 Minutes post-injection) , Whole body planar SPECT imaging (60 minutes post-injection) , Whole body planar SPECT imaging (180 minutes post-injection), and Whole body planar SPECT imaging (18-20 hours post-injection), Planar Image with both Hands in Field of View (60 minutes and 180 minutes post-injection), Blood Collection for PK Testing (15 Mins Before Injection), Blood Collection for PK Testing (after Injection) , Blood Collection for PK Testing (15 minutes post injection) , Blood Collection for PK Testing (60 minutes post injection) , Blood Collection for PK Testing (180 minutes post injection) , and Blood Collection for PK Testing (18-20 hours post injection). arm 11: Group 11 will receive Tc99m-tilmanocept at the MTD via IV injection, Whole body planar SPECT imaging (15 Minutes post-injection) , Whole body planar SPECT imaging (60 minutes post-injection) , Whole body planar SPECT imaging (180 minutes post-injection), and Whole body planar SPECT imaging (18-20 hours post-injection), Planar Image with both Hands in Field of View (60 minutes and 180 minutes post-injection), Blood Collection for PK Testing (15 Mins Before Injection), Blood Collection for PK Testing (after Injection) , Blood Collection for PK Testing (15 minutes post injection) , Blood Collection for PK Testing (60 minutes post injection) , Blood Collection for PK Testing (180 minutes post injection) , and Blood Collection for PK Testing (18-20 hours post injection).
Interventions: intervention 1: Tc99m-tilmanocept intervention 2: SPECT Imaging (60 Minutes post-injection) intervention 3: SPECT Imaging (180 Minutes post-injection) intervention 4: Whole body planar SPECT imaging (15 Minutes post-injection) intervention 5: Whole body planar SPECT imaging (60 Minutes post-injection) intervention 6: Whole body planar SPECT imaging (180 Minutes post-injection) intervention 7: Whole body planar SPECT imaging (18-20 Hours post-injection) intervention 8: Blood Collection for PK Testing (15 Mins Before Injection) intervention 9: Blood Collection for PK Testing (after injection) intervention 10: Blood Collection for PK Testing (15 minutes post injection) intervention 11: Blood Collection for PK Testing (60 minutes post injection) intervention 12: Blood Collection for PK Testing (180 minutes post injection) intervention 13: Blood Collection for PK Testing (18-20 hours post injection) intervention 14: Planar Image with both Hands in Field of View
Intervention descriptions: intervention 1: Tilmanocept is a radiotracer that accumulates in macrophages by binding to a mannose binding receptor that resides on the surface. intervention 2: Subjects enrolled in Groups 1-9 will receive whole body planar SPECT imaging with 3D SPECT or SPECT/CT for targeted joints of interest 60 Minutes post-injection. intervention 3: Subjects enrolled in Groups 1-9 will receive whole body planar SPECT imaging with 3D SPECT or SPECT/CT for targeted joints of interest 180 Minutes post-injection. intervention 4: Subjects enrolled in Groups 10-11 will receive a whole body planar SPECT scan performed 15 minutes post-injection for dosimetry. intervention 5: Subjects enrolled in Groups 10-11 will receive a whole body planar SPECT scan performed 60 minutes post-injection for dosimetry. intervention 6: Subjects enrolled in Groups 10-11 will receive a whole body planar SPECT scan performed 180 minutes post-injection for dosimetry. intervention 7: Subjects enrolled in Groups 10-11 will receive a whole body planar SPECT scan performed 18-20 Hours post-injection for dosimetry. intervention 8: Blood will be collected for subjects enrolled in Groups 10-11 for the purpose of PK analysis within 15 minutes prior to administration of Tc 99m tilmanocept intervention 9: Blood will be collected for subjects enrolled in Groups 10-11 for the purpose of PK analysis immediately following administration of Tc 99m tilmanocept (00:00) intervention 10: Blood will be collected for subjects enrolled in Groups 10-11 for the purpose of PK analysis at 15 ± 5 minutes post injection of Tc 99m tilmanocept intervention 11: Blood will be collected for subjects enrolled in Groups 10-11 for the purpose of PK analysis at 60 ± 15 minutes post injection of Tc 99m tilmanocept intervention 12: Blood will be collected for subjects enrolled in Groups 10-11 for the purpose of PK analysis at 180 ± 15 minutes post injection of Tc 99m tilmanocept intervention 13: Blood will be collected for subjects enrolled in Groups 10-11 for the purpose of PK analysis at 18-20 hours post injection of Tc 99m tilmanocept intervention 14: Subjects in Groups 1-9 and Groups 10 and 11 will receive planar imaging with both hands in the field of view at 60 and 180 minutes post-injection.

Protocol documents:
Prot_SAP_000.pdf: 
 
 
Clinical Study Protocol: NAV3-21 
Study Title: 
An Evaluation of the Safety of Escalating Doses of Tc 99m 
Tilmanocept by Intravenous (IV) Injection and Skeletal Joint Imaging 
with SPECT in Subjects with Active Rheumatoid Arthritis (RA) and 
Healthy Controls 
Study Number: 
NAV3-21 
Study Phase: 
1 and 2 
Product Name: 
  Technetium Tc 99m tilmanocept 
IND Number: 
132943 
Investigator(s): 
Multicenter 
Sponsor: 
Navidea Biopharmaceuticals, Inc 
4995 Bradenton Avenue, Suite 240 Dublin, OH 43017 
Sponsor Contact: 
Allison Kissling 
Associate Clinical Research Project Manager 
+1 614-973-7551 
Medical Monitor: 
Michael Blue, MD 
Senior Medical Director 
+1 614-571-4313 
 
Date 
Original Protocol: 
Amendment 1: 
Amendment 2:  
Amendment 3: 
Amendment 4: 
Amendment 5: 
Amendment 6: 
23 May 2016  
12 September 2016 
22 December 2016 
09 February 2017 
23 August 2017 
18 April 2018 
19 June 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: NAV3-21 
 
Date: 19 June 2018 
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SYNOPSIS 
Study title 
An Evaluation of the Safety of Escalating Doses of Tc 99m 
Tilmanocept by Intravenous (IV) Injection and Skeletal Joint 
Imaging with SPECT in Subjects with Active Rheumatoid 
Arthritis (RA) and Healthy Controls 
Study phase 
Phase 1 and 2 
Study objective(s) 
Primary  
• Determine the safety and tolerability of escalating 
doses of Tc 99m tilmanocept.  
Secondary  
• Determine the localization of Tc 99m tilmanocept by 
SPECT 
(single 
photon 
emission 
computed 
tomography) imaging in subjects with active RA and 
asymptomatic 
controls 
and 
concordance 
of 
localization with clinical symptomology. 
• Comparison of localization intensity between Tc 99m 
tilmanocept dose groups for optimal dose selection and 
determination of optimal imaging time. 
• To estimate the pharmacokinetics (PK) and dosimetry 
of Tc 99m tilmanocept in subjects with and without 
active RA.  
Duration of treatment 
Up to 53 days 
Study drug 
Technetium Tc 99m tilmanocept  
Dose(s) and Route of 
administration 
Route of Administration: All administrations of Tc 99m 
tilmanocept will occur through an intravenous (IV) route of 
injection. For the 50 and 200 µg doses a single syringe will be 
used and injected as a slow push into the Y-port. For the 400 
µg dose 2 syringes will be used and injected sequentially as a 
slow push into the Y-port. At the completion of the 
injection(s), a 10 cc sterile normal saline flush will be 
administered. 
The preferred site of IV placement will be the left or right 
antecubital vein unless both elbows are primary sites for RA 
imaging, and then the site for IV placement will be between 
the elbow and the wrist. 
Doses: Tc 99m tilmanocept will be received by subject 
through IV administration at one of 3 mass doses: 50 µg, 200 
µg, or 400 µg. Within each mass dose group, subjects will 
receive Tc 99m tilmanocept labeled with one of 3 radiolabel 
doses: 1 mCi, 5 mCi, or 10 mCi. 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 3 of 83 
 
 
Tilmanocept Mass Dose 
Specific 
Radioactivity  
50 µg 
200 µg 
400 µg 
 
10 mCi 
Group 1 
n = 3 RA 
Group 2 
n = 3 RA 
 Group 3 
n = 3 RA 
 
5 mCi 
Group 4 
n = 3 RA 
Group 5 
n = 3 RA 
Group 6 
n = 3 RA 
 
1 mCi 
Group 7 
n = 3 RA 
Group 8 
n = 3 RA 
Group 9 
n = 3 RA 
* Group 10 will consist of 6 Healthy Controls (HC) (3  
female subjects and 3 male subjects) and Group 11 will 
consist of 6 subjects with active RA (3  female subjects and 
3 male subjects). Subjects in Groups 10 and 11 will be 
enrolled in parallel and will be dosed at the maximum 
tolerated dose (MTD) as determined from Groups 1-9. 
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. Has a negative urine drug screening for illicit or 
unprescribed drugs suggestive of drug abuse as 
determined by the investigator. 
3.  All subjects shall be ≥18 years of age at the time of 
consent.   
 
CONTROL SUBJECTS: 
4. The subject is deemed to be clinically free of any 
inflammatory disease (s) and has not experienced joint 
pain for at least 4 weeks prior to the consent date. 
 
ACTIVE RHEUMATOID ARTHRITIS SUBJECTS: 
4. The subject has moderate to severe RA as determined 
by the 2010 ACR/EULAR (score of ≥ 6/10) that 
includes an affected hand and at least one large jointing 
(elbow, knee or shoulder). 
5. The subject has a DAS28 of ≥ 3.2 (includes the 
Erythrocyte Sedimentation Rate [ESR] test and Visual 
Analog Scale [VAS]) . 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 4 of 83 
 
6. If the subject is receiving methotrexate, they have been 
at a stable dose for > 4 weeks prior to the  Baseline Visit 
2 (Day 1). 
7. If the subject is receiving biologic therapy, they have 
been at a stable dose > 8 weeks prior to the Baseline 
Visit 2 (Day 1). 
8. If the subject is receiving NSAIDS (nonsteroidal anti-
inflammatory drug) or oral corticosteroids, the dose has 
been at a stable dose for > 4 weeks prior to the Baseline 
Visit 2 (Day 1). The corticosteroid dose should be ≤ 
10mg/day of prednisone or an equivalent steroid dose. 
Exclusion criteria 
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 had or is currently receiving radiation 
therapy or chemotherapy for a condition other than 
rheumatoid arthritis. 
4. The subject has renal insufficiency as demonstrated by 
serum creatinine clearance of < 60 mL/min. 
5. 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. 
6. The subject has a chronic or persistent infection or has any 
condition that would, in the opinion of the examining 
physician, preclude their participation. 
7. The subject has a known allergy to or has had an adverse 
reaction to dextran exposure. 
8. The subject has received an investigational product within 
30 days prior to the Tc 99m tilmanocept administration. 
9. The subject has received any radiopharmaceutical within 
7 days prior to the administration of Tc 99m tilmanocept. 
Study design 
Prospective, open-label, multicenter, dose escalation, and 
safety with PK and dosimetry study of injected Tc 99m 
tilmanocept in the detection of and assessment of localization 
to skeletal joints in subjects with and without active RA by 
SPECT imaging. All subjects will receive IV administration at 
one of 3 mass doses: 50 µg, 200 µg, or 400 µg. Within each 
mass dose group, subjects will receive Tc 99m tilmanocept 
labeled with one of 3 radiolabel doses: 1 mCi, 5 mCi, or 10 
mCi.  
All subjects will have a whole body planar SPECT scan. 
Subjects enrolled in Groups 1-9 will receive a whole body 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 5 of 83 
 
planar SPECT scan followed by a 3D SPECT or SPECT/CT 
scan on areas of interest at two specified time points post 
injection: 60 minutes ± 15 minutes and 180 minutes ± 15 
minutes. Subjects enrolled in Groups 10-11 will receive a 
whole body planar SPECT scan performed at 4 specified time 
points post injection: 15 ± 5 minutes, 60 ± 15 minutes, 180 ± 
15 minutes and 18-20 hours. Subjects enrolled in Groups 10-
11 will also receive a planar SPECT hands scan at 60 ± 15 
minutes and 180 ± 15 minutes post-injection. PK blood 
samplingwill be performed before injection (within 15 
minutes), immediately following injection (within 5 minutes), 
and at each scanning timepoint. Dosimetry tests will be 
performed at each scanning timepoint. PK of urine will be 
assessed through counts of the bladder wall obtained from 
cumulative quantitative planar imaging  from radiation 
dosimetry. 
Methodology 
The study includes: screening, enrollment, pre- and post-
injection assessments, injection, imaging and follow-up. 
All Subjects: 
Visit 1, Screening (Day -44 to Day 0): The screening visit 
will include informed consent, preliminary review of study 
eligibility, 
collection 
of 
medical 
history 
including 
medications, 
urine 
drug 
screen, 
vital 
signs, 
ECG 
(electrocardiogram), physical exam including height and 
weight, clinical labs, RA specific labs, urinalysis, urine 
pregnancy test for subject of childbearing potential, 2010 
ACR/EULAR score, DAS28 evaluation.  
 
Groups 1-9: 
Visit 2, Baseline (Day 1):Pre-Tc 99m Tilmanocept 
Injection:  
• Before injection (Day 1) 
o Urine pregnancy test will be performed for 
subjects of childbearing potential 
o Concomitant medication review 
o ECG - within 15 minutes prior to injection 
o Vital Signs - within 15 minutes prior to 
injection 
 
 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 6 of 83 
 
Tc 99m Tilmanocept Injection : Subjects will receive their 
open label serial dose assignment in accordance with Table 1. 
Adverse event monitoring will continue through completion 
of the trial.  
Post-Tc 99m Tilmanocept Injection:  
• 0 - 15 Minutes Post Injection: 
o Assessment of adverse events 
o ECG (completed before vital signs) 
o Vital Signs 
• 60 ± 15 Minutes Post Injection: 
o Assessment of adverse events 
o Whole body planar imaging 
o Planar image with both hands in the field of 
view (FOV)  
 Additional 3D SPECT or SPECT/CT 
will be obtained on identified joints of 
interest. 
• 180 ± 15 Minutes Post Injection 
o Assessment of adverse events 
o Whole body planar imaging 
o Planar image with both hands in the FOV 
 Additional 3D SPECT or SPECT/CT 
will be obtained on identified joints of 
interest. 
5± 2 Days Post-injection Follow-Up Visit (Visit 3):  
• Vital Signs  
• ECG 
• Physical Exam, including body weight 
• Clinical labs and urinalysis 
• Concomitant medication review 
• Assessment of adverse event 
 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 7 of 83 
 
Groups 10-11: 
Visit 2, Baseline (Day 1): 
Pre-Tc 99m Tilmanocept Injection:  
• Before injection (Day 1) 
o Urine pregnancy test will be performed for 
subjects of childbearing potential 
o Concomitant medication review 
o ECG - within 15 minutes prior to injection 
o Vital Signs - within 15 minutes prior to 
injection 
o PK blood sample – within 15 minutes prior to 
injection 
 
Tc 99m Tilmanocept Injection: Subjects will receive the 
maximum tolerated dose, as determined from Table 1. 
Immediately following Tc 99m tilmanocept administration, a 
PK blood sample will be drawn. Adverse event monitoring 
will continue through completion of the trial.  
Post-Tc 99m Tilmanocept Injection Procedures: 
• 0 - 15 Minutes Post Injection: 
o Assessment of adverse events 
o ECG (completed before vital signs) 
o Vital Signs 
• 15 ± 5 Minutes Post Injection: 
o Whole body planar imaging 
o PK blood sampling 
o Assessment of adverse events 
• 60 ± 15 Minutes Post Injection: 
o Whole body planar imaging 
o Planar image with both hands in the FOV 
o PK blood sampling 
o Assessment of adverse events 
• 180 ± 15 Minutes Post Injection: 
o Whole body planar imaging 
o Planar image with both hands in the FOV 
o PK blood sampling 
o Assessment of adverse events 
 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 8 of 83 
 
Post-Tc 99m Tilmanocept Injection Procedures and 
Follow-Up Visit (Visit 3): 
• 18-20 Hours Post Injection:  
o Whole body planar imaging 
o PK blood sampling 
o Vital Signs 
o ECG 
o Physical Exam, including body weight 
o Clinical labs and urinalysis 
o Concomitant medication review 
o Assessment of Adverse Events  
Planned study dates 
Start of study 
recruitment / August 
2016 
End of Recruitment/July 2018 
End of Study/September 2018 
Planned number of 
study centers 
Approximately 1-3 centers in the United States  
Number of subjects 
Up to 48 evaluable subjects may be enrolled. The dose 
escalation portion of this study is a modified 3+3 design with 
no dose de-escalation.  The sequential cohort enrollment 
characteristics of this design do not allow a fixed computation 
of sample size.  The parameters of the design that can be 
calculated are shown in Table 4.  Based on an assumed vector 
of probabilities of no subjects experiencing pharmacologic 
activity or an Adverse Drug Reaction (ADR) in each dose 
group, the average sample size of the dose escalation phase of 
the study is n=26.3 patients and the probability that dose group 
7 will be selected as the MTD (maximum tolerated dose) is 
0.152. 
Primary endpoint 
Proportion of subjects not experiencing pharmacologic 
activity or ADR in each dose group.  
Secondary endpoints 
The secondary endpoints for this study are: 
• Per subject localization rate of Tc 99m tilmanocept by 
SPECT imaging  
• Per joint location clinically RA-identified joint 
localization rate of Tc 99m tilmanocept by SPECT 
imaging  
• Concordance of Tc 99m localization with anatomical 
areas of active RA defined by clinical symptomology. 
• Localization intensity for each clinically RA-defined 
joint (as determined by quantitative SPECT gamma 
counts)  


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 9 of 83 
 
• Per subject localization rate of Tc 99m tilmanocept in 
areas other than RA-positive joints by SPECT 
imaging  
• The following PK parameters will be calculated, 
whenever possible, using Tc 99m tilmanocept total 
radioactivity in whole blood and urine: maximum 
observed concentration (Cmax), time to Cmax (tmax), 
area under the concentration-time curve (AUC) from 
Hour 0 to the last measureable concentration (AUC0-
t), AUC extrapolated to infinity (AUC0-∞), apparent 
terminal elimination rate constant (λZ), and apparent 
terminal elimination half-life (t1/2).   
• Radiation dosimetry of Tc 99m tilmanocept will 
include radiation doses per organ (μGy/MBq), organs 
receiving the highest organ dose (HT), effective doses 
per organ (μSv/MBq), mean effective dose 
(μSv/MBq), effective dose resulting from a diagnostic 
dose  ± 20%.  Dosimetry will be based on target 
organ image counts and modeling per 
OLINDA/EXM® 2.0 (Organ Level INternal Dose 
Assessment/EXponential Modeling) software. 
Data Analyses  
The following analysis population will be defined for the 
study: 
• Intent-to-Diagnose (ITD) Population - Subjects who 
are enrolled in the study in Groups 1-9, injected with 
Tc 99m tilmanocept, and received whole body planar 
imaging will be included in the ITD analysis 
population. 
• Radiopharmacokinetic 
(RPK) 
Population 
– 
Subjects that have been enrolled in the study in Groups 
10-11, injected with Tc 99m tilmanocept, and received 
whole body planar imaging will be included in the 
RPK analysis population. 
• Safety Population – All patients who are enrolled and 
injected with Tc 99m tilmanocept in the study will be 
included in the safety population. 
• Per Protocol (PP) Population – The PP population 
will include all safety subjects without major protocol 
violations. 
All safety data analyses will be conducted on the safety 
population.  All efficacy data analyses will be conducted on 
the ITD and PP population with the ITD population being the 
primary analysis set. All analyses of RPK data will be carried 
out on the RPK population. 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 10 of 83 
 
The number and percentage of subjects experiencing 
pharmacologic activity or ADRs will be tabulated by dose 
group and overall. 
Per subject localization rates and percentage will be calculated 
by time point by dose group and overall.   
Per joint location localization rates and percentages will be 
calculated by time point by dose group and overall.   
Per location concordance of clinical diagnosis to nuclear 
medicine specialist illumination indication will be computed 
across all RA positive sites of all subjects at all time points.   
Localization intensities will be summarized by descriptive 
statistics (mean, median, standard deviation, minimum, 
maximum and range) by joint location at each time point.   
Radiopharmacokinetic (RPK) parameters will be estimated for 
each subject using noncompartmental models.  Summary 
statistics (n, mean, std, CV, geometric mean) will be computed 
for each RPK parameter across all subjects  by Group (10, 11). 
For dosimetry data, for each dosimetry parameter, summary 
statistics (n, mean, std, and a one standard deviation width 
confidence interval) will be computed by organ type and 
across all subjects by Group (10,11). 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 11 of 83 
 
TABLE OF CONTENTS 
SYNOPSIS .................................................................................................................................2 
LIST OF IN-TEXT TABLES ..................................................................................................13 
LIST OF APPENDICES ..........................................................................................................14 
LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS ..........................................15 
STUDY ADMINISTRATIVE STRUCTURE .........................................................................19 
1 
INTRODUCTION ...........................................................................................................20 
1.1 
Background ............................................................................................................20 
1.2 
Preliminary Data ....................................................................................................23 
1.3 
Previous Nonclinical Research and Clinical Trial Experience Tilmanocept and 
Lymphoseek ...........................................................................................................25 
1.3.1 
Nonclinical Evaluations – Injection Site Subcutaneous ..............................25 
1.3.2 
Nonclinical Evaluations – IV AND IP Administration ...............................26 
1.3.3 
Clinical Pharmacokinetics and Pharmacodynamics ....................................29 
1.3.4 
Clinical Efficacy ..........................................................................................29 
1.3.5 
Clinical Safety ..............................................................................................30 
2 
STUDY OBJECTIVES ....................................................................................................31 
2.1 
Primary Objective(s) ..............................................................................................31 
2.2 
Secondary Objective(s) ..........................................................................................31 
3 
OVERVIEW OF METHODOLOGY AND DESIGN .....................................................32 
3.1 
Overall Study Design .............................................................................................32 
3.2 
Justification for Study Design and Population.......................................................32 
3.3 
Protocol Adherence ................................................................................................33 
3.4 
Study Duration .......................................................................................................33 
4 
STUDY POPULATION ..................................................................................................34 
4.1 
Eligibility ...............................................................................................................34 
4.1.1 
Inclusion Criteria .........................................................................................34 
4.1.2 
Exclusion Criteria ........................................................................................35 
4.2 
Recruitment ............................................................................................................35 
4.3 
Withdrawal .............................................................................................................35 
4.4 
Replacement ...........................................................................................................36 
4.5 
Subject Identification .............................................................................................36 
5 
INVESTIGATIONAL PRODUCT ..................................................................................37 
5.1 
Identification of Investigational Product ...............................................................37 
5.2 
Investigational Product Dosage and Administration .............................................37 
5.3 
Treatment Assignment ...........................................................................................38 
5.4 
Packaging and Labeling .........................................................................................38 
5.5 
Drug Logistics and Investigational Product Accountability ..................................39 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 12 of 83 
 
6 
THERAPIES OTHER THAN INVESTIGATIONAL PRODUCT .................................40 
6.1 
Prior and Concomitant Therapy .............................................................................40 
6.2 
Post-Study Therapy ................................................................................................40 
7 
STUDY PROCEDURES .................................................................................................41 
7.1 
Schedule of Evaluations .........................................................................................41 
7.2 
Visit Description ....................................................................................................41 
7.2.1 
Visit 1, Screening (Day -44 to Day 0) .........................................................41 
7.2.2 
Visit 2, Baseline (Day 1) ..............................................................................42 
7.2.2.1 
Before Administration of Tc 99m Tilmanocept.................................42 
7.2.2.2 
Administration of Tc 99m Tilmanocept ............................................42 
7.2.2.3 
After Administration of Tc 99m Tilmanocept ...................................43 
7.2.3 
Visit 3 Follow Up and End of Study ............................................................44 
8 
PROCEDURES AND VARIABLES ..............................................................................45 
8.1 
Population Characteristics .....................................................................................45 
8.1.1 
Demographic and Other Baseline Characteristics .......................................45 
8.1.2 
Medical and Surgical History ......................................................................45 
8.1.3 
DAS28 Evaluation .......................................................................................45 
8.1.4 
2010 ACR/EULAR Classification Criteria ..................................................45 
8.1.5 
Prior and Concomitant Medication ..............................................................46 
8.2 
Tc 99m Tilmanocept Preparation and Administration...........................................46 
8.3 
SPECT/SPECT-CT Image Acquisition .................................................................46 
8.4 
Pharmacokinetics ...................................................................................................48 
8.4.1 
Sample Processing .......................................................................................49 
8.5 
Dosimetry ...............................................................................................................49 
8.6 
Safety .....................................................................................................................49 
8.6.1 
Adverse Events ............................................................................................49 
8.6.1.1 
Definition of Adverse Event ..............................................................49 
8.6.1.2 
Categories for Adverse Event Assessment ........................................50 
8.6.1.3 
Assessments and Documentation of Adverse Events ........................51 
8.6.1.4 
Expected Adverse Events ..................................................................52 
8.6.1.5 
Serious Adverse Events .....................................................................53 
8.6.2 
Further Safety Assessments .........................................................................54 
8.6.2.1 
Physical Exam ....................................................................................54 
8.6.2.2 
Vital Signs ..........................................................................................54 
8.6.2.3 
Electrocardiogram ..............................................................................55 
8.6.2.4 
Clinical Laboratory Parameters for Screening and Safety .................55 
9 
STATISTICAL METHODS ............................................................................................57 
9.1 
Randomization Methods ........................................................................................57 


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9.2 
Safety Variables .....................................................................................................57 
9.3 
Efficacy Variables ..................................................................................................57 
9.4 
Sample Size Justification .......................................................................................58 
9.5 
Statistical Analyses ................................................................................................59 
9.5.1 
Analysis Populations ....................................................................................59 
9.5.2 
Analysis of Baseline and Demographic Characteristics ..............................59 
9.5.3 
Analysis of Efficacy Variables ....................................................................59 
9.5.4 
Analysis of RPK and Dosimetry Variables .................................................60 
9.5.5 
Handling Missing Values .............................................................................61 
9.5.6 
Interim Analysis ...........................................................................................61 
10 DATA HANDLING AND QUALITY ASSURANCE ...................................................62 
10.1 
Data Recording ......................................................................................................62 
10.1.1 
CRF Design ..................................................................................................62 
10.2 
Monitoring .............................................................................................................62 
10.3 
Data Processing ......................................................................................................62 
10.4 
Auditing .................................................................................................................62 
10.5 
Archiving ...............................................................................................................63 
10.6 
Premature Termination of the Study ......................................................................63 
10.6.1 
Study as a Whole .........................................................................................64 
10.6.2 
Center ...........................................................................................................64 
10.6.3 
Study Participant ..........................................................................................64 
11 ETHICAL AND LEGAL ASPECTS...............................................................................65 
11.1 
Ethical and Legal Conduct of the Study ................................................................65 
11.2 
Subject Information and Consent...........................................................................65 
11.3 
Financing/Financial Disclosure .............................................................................66 
11.4 
Publication Policy ..................................................................................................66 
11.5 
Subject Injury .........................................................................................................66 
12 REFERENCE LIST .........................................................................................................67 
 
LIST OF IN-TEXT TABLES 
Table 1. 
Dose Escalation Matrix .......................................................................................37 
Table 2. 
Clinical Laboratory Parameters ..........................................................................55 
Table 3. 
Approximate Amount of Blood Withdrawn .......................................................56 
Table 4. 
Design Characteristics for the Study Design ......................................................59 
 


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LIST OF IN-TEXT FIGURES 
Figure 1. 
Significantly increased Cy3-tilmanocept mediated joint fluorescence in 
arthritic joints (right) compared with control (left).............................................23 
Figure 2. 
Significantly increased Cy3-tilmanocept mediated joint fluorescence in 
arthritic joints compared with controls ...............................................................24 
Figure 3. 
A, B – (A) Significant staining of a CD16 and CD206 concordance in cells 
in synovial fluid (RA (right) but not OA (left) patients) and (B) - Staining of 
synovial tissue frozen sections from a patient with rheumatoid arthritis............24 
 
LIST OF APPENDICES 
Appendix 1 Schedule of Events ..............................................................................................76 
Appendix 2 Groups 1-9 Visit 2, Day 1 Diagram (example) ...................................................77 
Appendix 3 Groups 10-11 Visits 2 and 3 Diagram (example) ...............................................78 
Appendix 4 2010 ACR/ EULAR Classification Criteria ........................................................79 
Appendix 5 DAS-28 Scale......................................................................................................81 
Appendix 6 Sponsor Signatures..............................................................................................82 
Appendix 7 Investigator’s Signature ......................................................................................83 
 


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LIST OF ABBREVIATIONS AND DEFINITIONS OF TERMS 
ACR/EULAR American College of Rheumatology/ European League Against 
Rheumatism 
ACPA 
Anti-citrullinated peptide antibody 
ADR 
adverse drug reaction 
AE 
adverse event 
ALT 
alanine aminotransferase (SGPT) 
AST 
aspartate aminotransferase (SGOT) 
AUC 
area under the concentration-time curve 
AUC0-t 
AUC from hour 0 to last measurable concentration 
AUC0-∞ 
AUC extrapolated to infinity 
BMI 
body mass index 
BUN 
blood urea nitrogen 
Cmax 
maximum observed concentration 
CD206 
Mannose-binding receptor (Ca2+-binding lectin) 
 
CO2 
carbon dioxide 
CRF 
case report form 
CRA 
clinical research associate 
CRO 
contract research organization 
CRP 
C-Reactive protein 
CT 
X-ray computed tomography 
Cy3 
Cy3 cyanine dyes fluoresce orange (~550 nm excitation, ~570 nm 
emission) 


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DAS28 
Disease activity score used with the ACR/EULAR 2010 RA guidelines 
DICOM 
Digital Imaging Communications in Medicine 
DMARD(s) 
disease-modifying antirheumatic drug(s)  
DTPA 
diethylene triamine pentaacetic acid  
ECG 
electrocardiogram 
eCRFs 
Electronic case report forms 
ESR 
Erythrocyte sedimentation rate 
EU 
European Union 
FDA 
Food and Drug Administration 
FNR 
false negative rate 
FOV 
field of view 
GCP 
Good Clinical Practice 
HC 
healthy controls  
HCT 
hematocrit 
Hgb 
hemoglobin 
HIPAA 
Health Information Portability and Accountability Act 
HT 
highest organ dose 
ICF 
informed consent form 
ICH 
International Conference on Harmonization  
%IDSN 
percentage of Injected Dose in the Sentinel Node 
ILM 
intraoperative lymphatic mapping 
IRB 
Institutional Review Board 
ISF 
investigator site file 
ITD 
intent-to-diagnose population 
IV 
intravenous 
kDa 
kiloDalton (molecular weight designation) 
mCi 
milliCurie (37x106 becquerels; 37megabecquerels) 


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MedDRA 
Medical Dictionary for Regulatory Activities 
MTD 
maximum tolerated dose 
NOAEL 
no-observed adverse-effect level 
NSAIDS 
Nonsteroidal anti-inflammatory drug 
OA 
osteoarthritis 
OLINDA/EX
M® 
Organ Level INternal Dose Assessment/EXponential Modeling 
PI 
principal investigator 
PK 
Pharmacokinetics  
PP 
per protocol 
RA 
Rheumatoid arthritis 
RBC 
red blood cells 
RF 
Rheumatoid Factor 
RPK 
Radiopharmacokinetic  
SAE 
serious adverse event 
SAP 
statistical analysis plan 
SCC 
squamous cell carcinoma 
SD 
study day 
SJC 
swollen joint count 
SLN 
sentinel lymph node 
SLNB 
sentinel lymph node biopsy 
SPECT 
single photon emission computed tomography 
SUSARs 
Suspected, Unexpected, Serious Adverse Reactions 
t1/2 
apparent terminal elimination half life 
tmax 
time to Cmax 
Tc 99m 
technetium-99m metastable isotope; γ-emitting (t½ = 6.02 h) 
TcSC 
Tc 99m sulfur colloid 
Tilmanocept 
DTPA Mannosyl Dextran (the US Adopted Name for the drug substance 
of Lymphoseek) 
TJC 
tender joint count 
TMF 
trial master file 


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TNFα 
Tumor necrosis factor alpha 
ULN 
upper limit of normal 
US 
United States 
VAS 
visual analog scale 
VBD 
vital blue dye 
WBC  
white blood cell 
λz 
apparent terminal elimination rate constant 
 


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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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1 
INTRODUCTION 
1.1 
Background 
Rheumatoid arthritis (RA) is a common, chronic, systemic, progressive, autoimmune disease 
causing inflammation and pathology throughout the body, but perhaps most noticeably in the 
peripheral joints of the skeleton (i.e. hands, feet, knees, hips, etc.). It has been estimated that 
52.2 million U.S. adults suffer from arthritis of which 1.3 million adults suffer from RA (CDC, 
Helmick et al.). In the affected joints, RA is characterized by macrophage and lymphocyte 
infiltration, proliferation of synovial fibroblastic tissue (pannus), and joint destruction (Ma et 
al.). RA causes joint pain, stiffness and reduced mobility. If not successfully treated, joint 
inflammation and destruction in RA patients can lead to crippling loss of function, severe 
chronic pain, and disfigurement of the joints. Also, RA patients have significantly higher risk 
of coronary heart disease including acute myocardial infarction (odds ratio = 3.17) (Maradit-
Kremers et al.). Patients with uncontrolled RA may also suffer a reduction of 3-10 years in 
their life expectancies (Carmona et al.). RA can strike anyone at any age, but is diagnosed most 
frequently in women in their 40s and 50s (Oliver et al.). Worldwide, about one adult in every 
200 has RA. In the United States, approximately 1.3 million adults have RA (Helmick et al., 
Scott et al.). Because RA is a chronic disease, the prevalence of RA increases with age. Because 
of aging demographics in the United States and elsewhere, the number of patients with RA and 
the burden of RA on society are expected to increase in the coming decades (Hootman et al.). 
Therefore, there is a significant current and growing need to manage RA patients more 
effectively to limit the morbidity and mortality caused by RA. 
 
Over the last decade or so, disease-modifying antirheumatic drugs (DMARDs) have 
transformed RA therapy. First line DMARD therapy is usually methotrexate (Scott et al.). In 
addition, a number of biologic DMARDs have been developed. Basic research on the 
pathobiology of RA revealed that the inflammation observed in RA is the consequence of a 
self-perpetuating disturbance in the expression and downstream signaling of a network of 
cytokines (Raza et al., Olszewski et al., Meyer et al.). At the center of this cytokine network is 
the overexpression of tumor necrosis factor alpha (TNFα) by synovial macrophages (Leizer et 
al., Westra et al., Hamilton et al., Keffer et al., Noack et al., Bugatti et al., Boissier et al., Tran 
et al., Kinne et al.). TNFα then induces IL-1 and other cytokines that together with TNFα 
mediate the inflammation and joint damage characteristic of RA (Leizer et al., Zwerina et al., 
Feldmann et al., Schett et al.). These findings led to the development of biologic therapies for 
RA that block TNFα (i.e., etanercept, infliximab, adalimumab, certolizumab pegol, and 
golimumab) (Scott et al., Kalden et al., Wiedmann et al., Chen et al.). In addition, other biologic 
therapies have been introduced that target other aspects of the immune system that are 
pathologically dysfunctional in RA. These include abatacept (targeting T-cell stimulation), 
rituximab (targeting B-cells), and tocilizumab (targeting the IL-6 receptor) (Vivar et al.). 
Generally, the biologic DMARDs are used as second line therapies for RA, either alone or with 
methotrexate, in patients who have failed methotrexate monotherapy (Wasserman et al., Saag 
et al.). All of the DMARDs can cause serious adverse side effects (Choy et al., Donahue et al.). 
Therefore, patients on DMARDs require close monitoring to manage and minimize their 
adverse side effect risks. Also, there is a need, not yet fully satisfied, to monitor the 
effectiveness of DMARD therapy to further minimize these risks and to optimize the benefits 


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of therapy. Conversely, the benefits of DMARD therapy can far outweigh its risks. Most 
patients on DMARDs receive a significant reduction in symptoms, and some patients 
experience a disease remission. In the past, the goals of RA therapy were to reduce the severity 
of symptoms and limit the rate of progression of the illness. Today, the goal is remission 
(Gramling et al.). Also, RA patients taking TNFα antagonists have a reduced risk for 
cardiovascular disease events (Barbhaiya et al.). 
 
There are two barriers preventing more RA patients from achieving disease remission (Bykerk 
et al.). Both of these barriers may be remedied by tilmanocept. Firstly, DMARD therapy is 
much more effective when provided soon after the onset of RA symptoms and declines in 
efficacy if initiation of DMARD therapy is delayed (Demoruelle et al.). Currently, and as 
described in the next section, early diagnosis of RA is problematic. Secondly, even when 
DMARD therapy is provided soon after the onset of symptoms, a significant  minority of 
patients do not respond or respond poorly, demonstrating the continuing need for new and 
better DMARD therapies (Sacre et al.). For example, in a study reported by Anderson et al. 
(Anderson et al.), the authors found that the time between the onset of RA symptoms and the 
initiation of methotrexate therapy was inversely associated with the response rate for therapy 
(p = 0.001). In another study of 598 RA patients who were followed for a number of years after 
their RA diagnosis (van der Linden et al.), patients who initiated therapy more than 12 weeks 
from the onset of symptoms experienced higher probability of failure to achieve DMARD-free 
remission (HR= 1.87) and experienced greater long term joint damage when compared with 
patients who initiated therapy within 12 weeks of symptom onset. These studies and the 
observations of others (Nell et al., Cush et al.) indicate that early diagnosis of RA affords a 
“window of opportunity” for the greatest probability of effective RA therapy and the possibility 
of disease remission.  Delayed diagnosis increases the chance of DMARD treatment failure. 
Furthermore, even in the best of circumstances, the response rate for RA therapies is in the 
50%-70% range (Choy et al., Bykerk et al., Sacre et al., Zwerina et al.),indicating the continued 
need for better DMARD therapies. 
 
The growing realization that early diagnosis of RA can positively impact treatment efficacy 
and long term patient outcomes motivated the American College of Rheumatology and the 
European League Against Rheumatism (ACR/EULAR) to collaborate on the development of 
new diagnostic criteria for RA. The results of this collaboration were published in 2010 as the 
ACR/EULAR 2010 criteria (Aletaha et al.). These new criteria replaced the older 1987 ACR 
criteria and were largely hailed as a “game changer” for the diagnosis and management of RA 
(Cohen et al.). Since its publication, the ACR/EULAR 2010 criteria have been evaluated in a 
number of studies to determine which patients presenting with polyarthralgia progress to RA 
(Humphreys et al.). For example, in one study 313 undifferentiated arthritis patients were 
evaluated at presentation and again after 1 year of follow up for a clinical diagnosis of RA 
and/or the initiation of methotrexate therapy (Berglin et al.). At baseline, 56% of patients 
fulfilled the 1987 ACR criteria, while 74% fulfilled the ACR/EULAR 2010 criteria for having 
RA. Compared with the diagnosis at follow up, the ACR/EULAR 2010 had a sensitivity and 
specificity for diagnosis of RA of 0.79 and 0.54, respectively. This compared with a sensitivity 
and specificity for the 1987 ACR criteria of 0.68 and 0.84, respectively. Other studies have 
made similar observations (Varache et al., Liao et al., Biliavska et al., de Hair et al., Alves et 
al., Britsemmer et al., Cader et al., Kaneko et al., van der Linden et al.). Sakellariou et al. 


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recently published a meta-analysis of this literature. They observed that the sensitivity and 
specificity of the ACR/EULAR 2010 criteria were 0.73 (CI: 0.64,0.80) and 0.74 (CI: 0.68,0.79) 
respectively. Clearly, there is significant risk of misclassification. As stated by Zeidler, 
“overdiagnosis and under-diagnosis may become important issues if the (ACR/EULAR 2010) 
criteria recommend” DMARD therapies. While the ACR/EULAR 2010 criteria may be a little 
better than the 1987 ACR criteria, the current situation is that diagnostic misclassification 
currently results in about a quarter of early RA patients not being given the DMARD therapies 
from which they could receive great benefit while a significant portion of arthritis patients 
without RA are being given DMARD therapy from which they will not benefit and which 
exposes them to risks from adverse side effects (Vonkeman et al.). Therefore, there is a critical 
unmet need for a more sensitive and specific diagnostic for RA that can differentiate patients 
first presenting with polyarthralgia and identify those with RA to facilitate the more efficacious 
delivery of DMARD therapies. 
 
While many types of cells, including T-cells, B-cells, dendritic cells and activated synovial 
fibroblasts contribute significantly to the establishment and maintenance of the pathology of 
RA, (Ma et al., Noack et al., Bugatti et al., Boissier et al., Tran et al.) macrophages play a 
critical role in RA (Kinne et al.). They produce most of the TNFα that drives and perpetuates 
the inflammatory cycle in RA (Leizer et al., Westra et al., Hamilton et al., Keffer et al., Noack 
et al., Bugatti et al., Boissier et al., Tran et al., Kinne et al., Zwerina et al., Feldman et al., 
Schett et al.). In the synovial sublining of a joint affected by RA, macrophages are the dominant 
cell type (Kraan et al., Cutolo et al.). In the inflamed joint as a whole, macrophages in RA 
patients make up at least 30%-40% of all cells (Kennedy et al.). Furthermore, macrophages 
participate directly in the destruction of bone and cartilage (Ma et al.). Activated macrophage 
populations and synoviocytes are the predominant cell types at the interface between pannus 
and cartilage and secrete destructive proteases in abundance (Bresnihan et al.). As a result, it 
may not be surprising that synovial macrophage numbers—but not the numbers of other 
immune cell types—correlate with radiographically determined joint destruction in RA 
(Mulherin et al., Yanni et al.). While  macrophages may play a role in other pathologies that 
cause joint pain and inflammation, the degree to which macrophages are involved in the 
pathological process of RA and the sheer mass or volume of macrophages that infiltrate the 
joints inflamed due to RA differentiates RA from other rheumatic diseases. Therefore, 
detection of the density or numbers of macrophages in inflamed joints may permit 
differentiation of patients with RA from those with other causes of arthritis. In addition, it is 
known that the RA pathology begins significantly before, perhaps years before, the onset of 
symptoms (i.e., joint pain and inflammation) and well before the beginning of bone destruction 
(Deane et al., El-Gabalawy et al.). Macrophage infiltration of synovial tissues precedes 
development of clinical signs of RA in animal studies (Kraan et al.). In humans, macrophage 
infiltrations of synovial tissues are present when RA patients first develop clinical symptoms 
(Demoruelle et al., van de Sande et al.). Therefore, detection of the density or numbers of 
macrophages in inflamed joints may facilitate more sensitive and specific identification of RA 
patients as soon as they present with symptoms and early in the course of their illnesses when 
DMARDs are likely to be most effective. 
 
An interesting and important observation that has been made in many studies is that the number 
of macrophages in synovial tissue, and particularly in the synovial sublining, declines in RA 


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patients when they are given DMARD therapy (Hamilton et al.). Furthermore, the degree to 
which synovial macrophage numbers decline is correlated with the magnitude of the DMARD 
treatment response. One study compared changes in the 28 joint count Disease Activity Score 
(DAS28) with changes in sublining macrophage numbers as determined by biopsies and found 
a significant correlation between the change in the number of macrophages and the change in 
DAS28 (Pearson correlation 0.874, p < 0.01) (Haringman et al.). The authors of this study have 
confirmed these findings in two additional studies, which used slightly different methodologies 
(Bresnihan et al., Bresnihan et al.). This correlation between declining macrophage numbers 
and the efficacy of DMARD therapy appears to be largely independent of the kind of DMARD 
therapy being investigated (Hamilton et al., Kinne et al., Franz et al., Kraan et al., Catrina et 
al., Cunnane et al., Vieira-Sousa et al.). These findings indicate that assaying the number of 
macrophages in inflamed joints of patients with RA could be used as an objective measure of 
the efficacy of DMARD therapy. These findings further suggest that assaying the number of 
macrophages in inflamed joints of patients with RA could be used in clinical studies as a 
biomarker of clinical response for potential new RA therapeutics. The problem with current 
methodologies is that macrophage numbers and densities need to be determined with synovial 
biopsies. This is obviously an invasive procedure that samples only a small portion of the 
inflamed synovial tissue and is painful and unpleasant for the patient. What would be 
preferable and likely more accurate is an imaging protocol, such as the one proposed in this 
application, that can assay synovial macrophages more completely and less invasively. 
 
1.2 
Preliminary Data 
Dr. Thomas Rosol, DVM PhD FACVP, Professor, Veterinary Biosciences investigated CD206 
expression and tilmanocept binding to synovial macrophages in an anti-type II collagen 
monoclonal antibody induced mouse model of RA (Hutamekalin et al.).  Evidence of arthritis 
in this model (joint swelling and redness) developed in 5-6 days, and on days 9 or 11, mice 
were imaged 1-2 hours after they had received an intravenous (IV) injection of Cy3-
tilmanocept. The mice were then euthanized followed by limb dissection and reimaging. The 
primary result of this experiment was that Cy3-tilmanocept administered intravenously 
localized to synovial macrophages in the affected joints of arthritic mice but not control mice 
(Figure 1 and Figure 2). 
Figure 1. 
Significantly increased Cy3-tilmanocept mediated joint 
fluorescence in arthritic joints (right) compared with control (left) 
 
 


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Figure 2. 
Significantly increased Cy3-tilmanocept mediated joint fluorescence 
in arthritic joints compared with controls 
 
The success of these studies in mice strongly supports the feasibility and likely success of the 
proposed studies in humans.  Further support for this notion could be derived from ex vivo 
experiments carried out that showed specific binding of Cy3-tilmanocept to human synovial 
macrophages obtained from patients with active RA but not osteoarthritis (OA) patients who 
were undergoing therapeutic/diagnostic arthrocentesis (Figure 3).   In these experiments the 
synovial cells were isolated by a ficoll gradient and the cells stained with a variety of surface 
markers to identify the cell type that can stain with Cy3-tilmanocept.  Flow cytometry contour 
plots from two representative RA and OA patients demonstrate a significant population of 
macrophages that stain with Cy3-tilmanocept in RA but is largely absent in OA (A) and 
staining of synovial tissue frozen sections from a patient with rheumatoid arthritis with Cy3-
tilmanocept, a fluorescent version of the radio-tracer version, Tc99m tilmanocept. 
Figure 3. 
A, B – (A) Significant staining of a CD16 and CD206 concordance in 
cells in synovial fluid (RA (right) but not OA (left) patients) and (B) - 
Staining of synovial tissue frozen sections from a patient with 
rheumatoid arthritis.    
 
B  - Sections were stained with 4',6-diamidino-2-
phenylindole (DAPI = DNA) and Cy3-tilmanocept 
(CD206) and examined under fluorescence 
microscopy. Image Key: Red, Cy3-tilmanocept; 
blue (DAPI). 


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Lymphoseek (technetium Tc 99m tilmanocept) is a radiotracer that accumulates in lymphatic 
tissue by binding to a mannose binding receptor that resides on the surface of dendritic cells 
and macrophages within the nodes. Tilmanocept is a macromolecule consisting of multiple 
units of DTPA (diethylenetriaminepentaacetic acid) and mannose, each synthetically attached 
to a 10 kDa dextran backbone. The mannose acts as a substrate for the receptor and the DTPA 
serves as a chelating agent for labeling with Tc 99m. Standard lymphoscintigraphy has been 
performed using Tc 99m sulfur colloid (TcSC) particles (50 to 800 nm in size) that are 
nonselectively removed by lymph node phagocytosis. Tilmanocept has a diameter of about 7 
nm, which is substantially smaller than current radiolabeled agents used for targeting lymphoid 
tissue. Tilmanocept’ s small diameter permits enhanced diffusion into lymph nodes and blood 
capillaries, resulting in a rapid injection site clearance. Lymphoseek has demonstrated less 
injection pain perception by patients, faster injection site transit, more successful localization 
of lymph nodes, and detection of more lymph nodes in the lymphatic drainage pathways in 
subsets of patients receiving Lymphoseek compared with other subsets having sulfur colloid. 
Lymphoseek is approved by the US (United States) Food and Drug Administration (FDA) for 
use in 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) in patients with clinically 
node negative squamous cell carcinoma (SCC) of the oral cavity, breast cancer, or melanoma. 
Lymphoseek is approved in the European Union (EU) for use in imaging and intraoperative 
detection of sentinel lymph nodes draining a primary tumor in adult patients with breast cancer, 
melanoma, or localized squamous cell carcinoma of the oral cavity. 
1.3 
Previous Nonclinical Research and Clinical Trial Experience 
Tilmanocept and Lymphoseek 
A detailed evaluation of safety and efficacy data from nonclinical and clinical studies can be 
found in the accompanying Investigator’s Brochure supplied by Navidea Biopharmaceuticals, 
Inc.  In addition, the data described in this protocol are reported in peer-reviewed publications, 
including nonclinical studies (Hoh et al., 2003), the Phase 1 clinical trials (Ellner et al.; Wallace 
et al.; Wallace et al.; Wallace et al.; Wallace et al.), the Phase 2 clinical trial (Leong et al.) and 
the Phase 3 studies (Tokin et al.; Sondak et al.; Wallace et al.; Marcinow et al., Agrawal et al., 
2015).   
1.3.1 
Nonclinical Evaluations – Injection Site Subcutaneous 
Nonclinical studies with Lymphoseek or tilmanocept demonstrated that the drug selectively 
binds to its intended receptor (the CD206 mannose binding receptor), is appropriately 
distributed for radio-detection of lymphatic tissue, and is well tolerated by rats, rabbits, guinea 
pigs, and dogs. 
Pharmacokinetics data obtained from nonclinical studies demonstrated rapid absorption into 
the plasma. Urinary excretion was a major pathway of elimination. Lymphoseek exhibited 
rapid clearance from the injection site, rapid uptake by the local lymph node, and low uptake 
by the remaining lymph nodes. Tilmanocept was well tolerated at all doses tested in nonclinical 


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safety pharmacology studies and in single and repeated dose toxicology studies in rats, rabbits, 
and dogs. In some studies in rabbits and dogs, tilmanocept acted as a local irritant of the 
subcutis or skeletal muscle, and induced mild inflammation and tissue degeneration. The no-
observed adverse-effect level (NOAEL) was 42μg/kg/day. Tilmanocept was not mutagenic or 
genotoxic in vitro or in vivo. No signs or symptoms of hypersensitivity were observed in a 
study in guinea pigs.  
1.3.2 
Nonclinical Evaluations – IV AND IP Administration 
Dosing in Dogs, Guinea Pigs and Mice: IV dosing in IND submitted non-clinical studies 
represents, cumulative dosing in male and female beagle dogs [eight dogs (4/sex; wt ♀= 7.6 
kg;  wt ♂ = 10.5 kg ) given an intravenous injection of saline on Study Day (SD) 1 and 2 and 
DTPA-mannosyl-dextran at 0.084 mg/kg on SD 4, 0.42 mg/kg on SD 6, and 0.84 mg/kg on 
SD 8 and 10] 1,560X the 50 µg and 195X the 400 µg dose (female dogs) and 1706X the 50 µg 
dose and 213X the 400µg dose (male dogs).   Based on the highest single dose administration 
(0.84 mg/kg) the dogs received 600X the 50 µg and 75X the 400 µg dose (female dogs) and 
657X the 50 µg and 82X the 400 µg dose (male dogs). [NOTE: In a separate dog study the 
dose was 0.56 mg/kg in 2 each ♂ and ♀ dogs of similar weight/sex), no effect was noted in 
that study, consistent with the higher dose dog study]. 
With regard to the sensitization test (IV administration in guinea pigs, wt = 0.415 kg) the 
maximum dose was the equivalent of 96X of the anticipated high dose tilmanocept. 
Lastly, although IP administered in mice, based on the highest single dose administration (2000 
mg/kg) the mice received 300,0000X the 50 µg and 37,500X the 400 µg dose (♀ mice, wt = 
22 gm) and 340,000X the 50 µg and 42,500X the 400 µg dose (♂ mice, wt = 29 gm). 
Dosing In Rats: 
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 
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 


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Type of Study / 
Description 
Test System 
Method of 
Administration 
Dosing 
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 
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 


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Type of Study / 
Description 
Test System 
Method of 
Administration 
Dosing 
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 intravenous 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. 
Single Bolus Toxicity: Tilmanocept-related clinical pathology changes were limited to 
minimally greater, dose-related, aspartate aminotransferase 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 


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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). 
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). 
1.3.3 
Clinical Pharmacokinetics and Pharmacodynamics 
In Phase 1 clinical trials, Lymphoseek showed rapid injection site clearance (approximately 2 
to 3 hours). Absolute uptake in the primary sentinel lymph node was dose-related for 
Lymphoseek, although relative nodal uptake (%IDSN) overall was independent of dose and 
ranged from 0.05%IDSN to 1.81%IDSN. 
In a Phase 2 trial, Lymphoseek was highly effective in identifying tumor-draining lymph nodes 
(i.e., overall, a “hot spot” from Lymphoseek was identified in 93.0% of patients for whom 
preoperative lymphoscintigraphy was performed, and the per-patient intraoperative 
localization rate was 96.2%). Diagnostic performance of in vivo Lymphoseek findings relative 
to pathology assessment of tumor tissue indicated a high per tissue sensitivity estimate (overall, 
92.0%). The overall false negative rate (FNR) for pathology was 8.0%, supporting the accuracy 
of Lymphoseek in identifying lymph nodes with a high potential for containing tumor 
metastases in the lymphatics draining the tumor bed. 
1.3.4 
 Clinical Efficacy 
Clinical efficacy was evaluated in two pivotal Phase 3 clinical trials in subjects with breast 
cancer and melanoma, comparing lymph node detection of Lymphoseek and vital blue dye 
(VBD). Lymphoseek demonstrated a statistically significant concordance rate with VBD 
(meta-analysis concordance rate = 99.99%). 
The detection concordance between Lymphoseek and VBD was similar among subjects with 
melanoma and subjects with breast cancer (meta-analysis concordance rate = 99.99% for both 
populations). Lymphoseek also demonstrated a higher sensitivity for detecting tumor-positive 
(as confirmed by pathology) lymph nodes, corresponding to a decreased FNR when compared 
with VBD on a per node basis. The corresponding sensitivity rate in detection of lymph nodes 
most likely to be positive for tumor cells for Lymphoseek was 99.99%, compared with 78.02% 
for VBD. The FNR for VBD (21.98%) was higher than the FNR for Lymphoseek (0.01%). 
These findings suggest that, when VBD is used as the imaging agent, there is an increased risk 
of missing the detection of tumor-involved lymph nodes and incorrectly staging cancer 
patients. The Lymphoseek-only findings (pathology-positive nodes that were hot/not blue) 
suggest that Lymphoseek was markedly more effective in identifying lymph nodes that 
harbored disease than was VBD. 


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A third pivotal Phase 3 clinical trial was conducted in subjects with oral cavity or cutaneous 
SCC.  Subjects received an injection of Lymphoseek prior to surgery for excision of the 
primary tumor and Lymphoseek-guided sentinel lymph node (SLN) dissection followed by 
planned elective neck dissection. All excised lymph nodes (Lymphoseek-identified SLNs as 
well as non-SLNs) underwent histopathologic evaluation for presence of metastatic disease. 
The primary endpoint in this trial was the Lymphoseek FNR (subjects with pathology-positive 
lymph node(s) not identified by Lymphoseek). Thirty-nine subjects had at least one 
pathology-positive lymph node, and Lymphoseek detected nodes with positive pathology in 
all but one patient. The FNR in this trial was 2.56%.  
1.3.5 
 Clinical Safety 
Well over 180,000 patients in clinical trials and US commercial use for ILM (intraoperative 
lymphatic mapping) with SLNB have been exposed to Lymphoseek, and there have been no 
safety signals, no deaths due to drug, and no SAEs (Serious Adverse Events) 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 
adverse events, 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 escalating doses of Tc 99m tilmanocept 
 
2.2 
Secondary Objective(s)     
• Determine the localization of Tc 99m tilmanocept by SPECT (single photon emission 
computed tomography) imaging in subjects with active RA and asymptomatic controls 
and concordance of localization with clinical symptomology. 
• Comparison of localization intensity between the dose groups of Tc 99m tilmanocept 
by SPECT imaging for optimal dose selection and determination of optimal imaging 
time. 
• To estimate the pharmacokinetics (PK) and dosimetry of Tc 99m tilmanocept in 
subjects with and without active RA. 
 


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3 
OVERVIEW OF METHODOLOGY AND DESIGN 
3.1 
Overall Study Design 
Prospective, open-label, multicenter, dose escalation, and safety with PK and dosimetry study 
of injected Tc 99m tilmanocept in the detection of and assessment of localization to skeletal 
joints in subjects with and without active RA by SPECT imaging. Subjects will receive IV 
administration at one of 3 mass doses: 50 µg, 200 µg, or 400 µg. Within each mass dose group, 
subjects will receive Tc 99m tilmanocept labeled with one of 3 radiolabel doses: 1 mCi, 5 mCi, 
or10 mCi. All subjects will have a whole body planar SPECT scan. Subjects enrolled in Groups 
1-9 will receive a whole body planar SPECT scan and planar of hands at two specified time 
points: 60 minutes ± 15 minutes, and 180 minutes ± 15 minutes followed by a 3D SPECT or 
SPECT/CT scan on areas of interest. Subjects enrolled in Groups 10-11 will have a whole body 
planar SPECT scan with pharmacokinetic and dosimetry tests performed at 4 specified time 
points post injection: 15± 5 minutes, 60 ± 15 minutes, 180 ± 15 minutes, and 18-20 hours. 
Subjects enrolled in Groups 10-11 will also receive a planar SPECT hands scan at 60 ± 15 
minutes and 180 ± 15 minutes post-injection. PK blood sampling will be performed before 
injection (within 15 minutes), immediately following injection (within 5 minutes),and at each 
scanning timepoint. Dosimetry tests will be performed at each scanning timepoint. PK of urine 
will be assessed through counts of the bladder wall obtained from cumulative quantitative 
planar imaging  from radiation dosimetry.The proposed study includes 3 visits: A screening 
visit for initial determination of eligibility and evaluation of clinical status (Visit 1), a baseline: 
Day 1 (Visit 2) on the day of tilmanocept administration and imaging, and a safety follow-up 
(Visit 3).  
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.  
3.2 
Justification for Study Design and Population 
This study is designed to evaluate the safety and tolerability of escalating doses of Tc 99m 
tilmanocept. Whole body planar as well as a planar scan of the hands using a SPECT camera 
will also be combined with 3D SPECT or SPECT/CT imaging to provide greater resolution of 
areas of Tc 99m tilmanocept localization.  Additionally, the PK and dosimetry of the MTD of 
Tc 99m tilmanocept will be evaluated in twelve subjects from Groups 10 and 11. Planar gamma 
camera imaging will be used to assess dosimetry. 
This study is designed to evaluate the use of Tc 99m tilmanocept as an imaging agent in 
subjects with known active rheumatoid arthritis by evaluating the localization in inflamed 
joints.   
No clinical trials have been conducted to evaluate IV administration of Tc 99m tilmanocept 
performance in subjects with rheumatoid arthritis. The rationale for evaluating Tc 99m 
tilmanocept in this subject population is discussed in Section 1.1 and Section 1.2.  


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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 
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 53 days depending on the duration of the screening 
window (up to 45 days) and Group assignment. 


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4 
STUDY POPULATION 
4.1 
Eligibility 
Subjects who fulfill all respective inclusion and none of the 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 Tc 99m tilmanocept and imaging (Day 1 
procedures).  A subject will be considered enrolled in the study on the morning of study day 1 
when they arrive at the study site.  Written, 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 1 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. Has a negative urine drug screening for illicit or unprescribed drugs suggestive of drug 
abuse as determined by the investigator. 
3. All subjects shall be ≥18 years of age at the time of consent.   
 
CONTROL SUBJECTS: 
4. The subject is deemed to be clinically free of any inflammatory disease (s) and has not 
experienced joint pain for at least 4 weeks prior to the consent date. 
 
ACTIVE RHEUMATOID ARTHRITIS SUBJECTS: 
4. The subject has moderate to severe RA as determined by the 2010 ACR/EULAR (score 
of ≥ 6/10) that includes an affected hand and at least one large jointing (elbow, knee or 
shoulder). 
5. The subject has a DAS28 of ≥ 3.2 (includes the Erythrocyte Sedimentation Rate [ESR] 
test and Visual Analog Scale [VAS]) . 
6. If the subject is receiving methotrexate, they have been at a stable dose for > 4 weeks 
prior to the Baseline Visit 2 (Day 1). 
7. If the subject is receiving biologic therapy, they have been at a stable dose > 8 weeks 
prior to the Baseline Visit 2 (Day 1). 
8. If the subject is receiving NSAIDS (nonsteroidal anti-inflammatory drug) or oral 
corticosteroids, the dose has been at a stable dose for > 4 weeks prior to the Baseline 
Visit 2 (Day 1). The corticosteroid dose should be ≤ 10mg/day of prednisone or an 
equivalent steroid dose. 


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4.1.2 
Exclusion Criteria 
Subjects who meet any of the following criteria will be excluded from the study. 
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 had or is currently receiving radiation therapy or chemotherapy for a 
condition other than rheumatoid arthritis. 
4. The subject has renal insufficiency as demonstrated by serum creatinine clearance of < 60 
mL/min. 
5. 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. 
6. The subject has a chronic or persistent infection or has any condition that would, in the 
opinion of the examining physician, preclude their participation. 
7. The subject has a known allergy to or has had an adverse reaction to dextran exposure. 
8. The subject has received an investigational product within 30 days prior to the Tc 99m 
tilmanocept administration. 
9. The subject has received any radiopharmaceutical within 7 days prior to the administration 
of Tc 99m tilmanocept. 
 
4.2 
Recruitment 
Subjects will be recruited from rheumatology 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, “RA-free” subjects 
(controls) will be recruited via IRB approved advertisements and clinically assessed for the 
“absence” of painful and/or swollen joints.     
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 arrival at the study site on Day 1 will be considered 
a screen failure. 
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 
possible. An explanation should be given of why the subject is withdrawing or being 
withdrawn from the study.  
 


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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 study drug administration or did not proceed to imaging 
 
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 “21”  
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 “21-01-001.”  
 
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 
Technetium 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 
Subjects will receive the administration of Tc 99m tilmanocept through an intravenous (IV) 
route of administration. For the 50 and 200 µg doses a single syringe will be used and injected 
as a slow push into the Y-port. For the 400 µg dose 2 syringes will be used and injected 
sequentially as a slow push into the Y-port. At the completion of the injection(s), a 10 cc sterile 
normal saline flush will be administered. Injection volume will be between 0.5 - 4.0 ml in 
sterile normal saline based on the mass dose group assignment. The preferred site of IV 
placement will be the left or right antecubital vein unless both elbows are primary sites for RA 
imaging, and then the site for IV placement will be between the elbow and the wrist. 
Tc 99m tilmanocept will be received by subjects through IV administration at one of 3 mass 
doses: 50 µg, 200 µg, or 400 µg. Within each mass dose group, subjects will receive Tc 99m 
tilmanocept labeled to one of 3 radiolabel doses: 1 mCi, 5 mCi, or 10 mCi (see Table 1). 
Table 1. 
Dose Escalation Matrix 
 
Tilmanocept Mass Dose 
Specific 
Radioactivity 
Tc 99m  
50 µg 
200 µg 
400 µg 
 
10 mCi 
Group 1 
n = 3 RA 
Group 2 
n = 3 RA 
 Group 3 
n = 3 RA 
 
5 mCi 
Group 4 
n = 3 RA 
Group 5 
n = 3 RA 
Group 6 
n = 3 RA 
 
1 mCi 
Group 7 
n = 3 RA 
Group 8 
n = 3 RA 
Group 9 
n = 3 RA 
*Group 10 will consist of 6 HC (3 female subjects and 3 male subjects) and Group 11 will 
consist of 6 subjects (3 female subjects and 3 male subjects) with active RA. Subjects in 
Groups 10 and 11 will be enrolled in parallel and will be dosed at the MTDas determined 
from Groups 1-9. 
The final administered dose for groups will be ± 20% of the tilmanocept mass dose and 
radiolabel mCi dose assignment.  


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5.3 
Treatment Assignment 
In this open-label, non-randomized, multicenter, dose-escalation safety study all subjects will 
receive IV administration at one of 3 tilmanocept mass doses: 50 µg, 200 µg, or 400 µg. Within 
each mass dose group, subjects will receive tilmanocept labeled to one of 3 radiolabel Tc 99m 
doses: 1 mCi, 5 mCi, or10 mCi. 
Group 1 will be the starting dose group. Three subjects will be enrolled into Group 1.  If none 
of the 3 subjects experiences pharmacologic activity or an ADR, 3 subjects will be enrolled in 
Group 2. However, if one of the first three subjects experiences a pharmacologic activity or an 
ADR, three more subjects will be treated at the same dose level. If 2 out of 3 or 3 out of 6 
subjects experiences a pharmacologic activity or an ADR in Group 1 the MTD cannot be 
determined. If no additional pharmacologic activity or ADRs are observed in the expanded 
group, the dose escalation will continue.  If one additional pharmacologic activity or ADR is 
observed, the current dose will be deemed the maximum tolerated dose (MTD). If more than 
one additional pharmacologic activity or ADR is observed in the expanded group, the previous 
dose will be deemed the MTD. Dose escalation will continue until the MTD is found or until 
Group 9 is reached with only one ADR or without any ADRs. If no ADRs are experienced 
from Groups 1-9, then the MTD will be highest tilmanocept mass dose radiolabeled at the 
hightest level (400 µg/10 mCi ).  
Group 10 will consist of 6 HC subjects and Group 11 will consist of 6 subjects with active RA. 
Subjects enrolled in Groups 10 and 11 will be dosed at the dose level that is established as the 
MTD from Groups 1 through 9.  The maximum tolerated dose is defined as the dose below 
where >33% of subjects experienced an ADR.  
NOTE: Adverse Drug Reaction 
An Adverse drug reaction (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 adverse event is at least 
a reasonable possibility, i.e. the relationship cannot be ruled out. 
In this multi-center trial, doses will be serially assigned in an open label fashion in accordance 
with Table 1. Investigators who have identified a subject for participation will receive the dose 
assignment from the sponsor and request that dose from the supporting  Cardinal Health 
pharmacy. 
5.4 
Packaging and Labeling 
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 is one tilmanocept vial, should be 
used for no more than one subject. The 400 µg dose groups will require two vials for each 
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. 


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Cardinal Health will radiolabel tilmanocept with the protocol group specified tilmanocept dose 
and the group assigned millicurie dose Tc 99m in 0.5 mL – 2.0 mL and deliver one syringe for 
the 50 and 200 µg doses and  up to 2 syringes for the 400 µg dose 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 Tc 99m tilmanocept injection through the post-injection 
safety follow-up will be documented. Subjects receiving radiation therapy or chemotherapy 
for a condition other than rheumatoid arthritis are not eligible for participation in the trial. The 
subject’s  history of treatments for rheumatoid arthritis 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 -44 to Day 0) 
All Subjects:  
• 
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. 
• 
Concomitant Medications (within 30 days before injection). 
• 
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. 
• 
Clinical laboratory tests – study subjects will have blood obtained for hematology, 
chemistry, and RA panel (see Section 8.6.2.4). 
• 
Urine collection for routine urinalysis  
• 
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. 
• 
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, lymph nodes, 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. 
• 
RA Evaluation: The swollen and tender joints will be identified and documented 
during by physical examination as established by the 2010 ACR/EULAR, a DAS28 
evaluation and a characterization of the subject’s RA history including previous 
treatments, date of symptom onset and date of diagnosis. 
• 
Final confirmatory review of inclusion and exclusion criteria 


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Changes in health occurring after consent and prior to the day of injection will be added to the 
subject’s medical history unless related to a study procedure. 
7.2.2 
Visit 2, Baseline (Day 1) 
All subjects will be assessed for adverse events in an ongoing manner from the day of injection 
through the end of participation.  
7.2.2.1 
Before Administration of Tc 99m Tilmanocept 
The following procedures will be completed for all subjects 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 adverse events 
• 
Concomitant medication review 
• 
ECG within 15 minutes prior to administration of Tc 99m tilmanocept 
• 
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 
 
A PK blood sample will be collected for Subjects in Groups 10 and 11 within 15 minutes 
prior to the administration of Tc 99m tilmanocept.  
 
7.2.2.2 
Administration of Tc 99m Tilmanocept 
IV administration of Tc 99m tilmanocept will be at study time 00:00. The preferred site of IV 
placement will be the left or right antecubital vein unless both elbows are primary sites for RA 
imaging, then the site for IV placement will be between the elbow and the wrist. 
The filled syringe(s) will be connected to the Y-port for a slow push injection. At the 
completion of the injection(s), 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 adverse events during the Day 1 Baseline Visit. 


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7.2.2.3 
After Administration of Tc 99m Tilmanocept 
Groups 1-9 
For subjects enrolled in Groups 1-9, the following procedures will be completed at the 
specified times:  
• 
Within 15 Minutes (00:01 to 00:15) Post Injection: 
o Assessment of adverse events  
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) 
 
• 
60 ± 15 Minutes Post Injection: 
o Assessment of adverse events  
o Whole body planar imaging (anterior/posterior views) 
o Planar image with both hands in the FOV (5 min anterior/posterior views) 
 Additional 3D SPECT or SPECT/CT will be obtained on identified 
joints of interest. 
 
• 
180 ±15 Minutes Post Injection: 
o Assessment of adverse events  
o Whole body planar imaging (anterior/posterior views) 
o Planar image with both hands in the FOV (5 min anterior/posterior views) 
 Additional 3D SPECT or SPECT/CT will be obtained on identified 
joints of interest. 
Groups 10-11 
For subjects enrolled in Groups 10-11, the following procedures will be completed at the 
specified times: 
• 
Immediately following administration of Tc 99m Tilmanocept (00:00), a PK blood 
sample will be drawn. 
• 
Within 15 Minutes (00:01 to 00:15) Post Injection: 
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 Assessment of adverse events 
• 
15 ± 5 Minutes Post Injection: 
o Assessment of adverse events 
o PK blood sampling 
o Whole body planar imaging (anterior/posterior views) 


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• 
60 ± 15 Minutes Post Injection: 
o Assessment of adverse events 
o PK blood sampling 
o Whole body planar imaging (anterior/posterior views) 
o Planar image with both hands in the FOV (5 min anterior/posterior views) 
 
• 
180 ±15 Minutes Post Injection: 
o Assessment of adverse events 
o PK blood sampling  
o Whole body planar imaging (anterior/posterior views) 
o Planar image with both hands in the FOV (5 min anterior/posterior views) 
 
7.2.3 
Visit 3 Follow Up and End of Study 
18-20 Hours Post-Injection (Groups 10-11): 
The following procedures will be conducted 18-20 hours after administration of Tc 99m 
tilmanocept at the final end of study visit for subjects enrolled in Groups 10-11: 
o PK blood sampling  
o Whole body planar imaging (anterior/posterior views) 
o Vital signs 
o ECG 
o Physical Exam, including body weight 
o Clinical Labs 
o Urinalysis 
o Concomitant medication review 
o Assessment and review of adverse events 
 
5± 2 Days Post-injection (Groups 1-9): 
The following procedures will be conducted 3 to 7 days after administration of Tc 99m 
tilmanocept and the final end of study visit for Subjects in Groups 1-9: 
o Vital signs 
o ECG 
o Physical Exam, including body weight 
o Clinical Labs 
o Urinalysis 
o Concomitant medication review 
o Assessment and review of adverse events 
 


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8 
PROCEDURES AND VARIABLES 
8.1 
Population Characteristics 
8.1.1 
Demographic and Other Baseline Characteristics 
Up to 48 evaluable subjects may be enrolled. Up to thirty-six (36) female or male subjects 
≥18 years of age with evidence of active RA will be enrolled in Groups 1-9.  Three (3) 
female and three (3) male subjects aged ≥18 without evidence of active RA (HC) will be 
enrolled in Group 10. Three (3) female and three (3) male subjects aged ≥18 with evidence of 
active RA will be enrolled in Group 11. 
 
8.1.2 
Medical and Surgical History 
Relevant medical and surgical histories will be obtained on all study subjects.   
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 
DAS28 Evaluation 
All subjects will be evaluated a single time at screening (Visit 1) for the Disease Activity Score 
(DAS) 28 (Prevoo et al, 1995) assessment. 
The DAS28 will be calculated from four components: tender joint count (TJC), swollen joint 
count (SJC), Visual analogue scale (VAS) of the subject’s global health, and the laboratory 
parameter erythrocyte sedimentation rate (ESR).  The formula used to calculate the DAS28 
score is: DAS28 = 0.56*√(TJC) + 0.28*√(SJC) + 0.70*LN(ESR) + 0.014*VAS).  See 
Appendix 5 for details. 
A DAS28 score of higher than 5.1 is indicative of high disease activity, whereas a DAS28 
below 3.2 indicates low disease activity. A subject is considered to be in remission if they have 
a DAS28 lower than 2.6 
8.1.4 
2010 ACR/EULAR Classification Criteria  
All subjects will be evaluated at screening using the 2010 ACR/EULAR Classification Criteria 
as part of eligibility and inclusion.  
The 2010 American College of Rheumatology (ACR)/European League Against Rheumatism 
(EULAR) classification criteria includes four components: number and site of involved joints, 
serologic abnormality, elevated acute-phase response and symptom duration. See Appendix 4 
for details. 
A total score of 6 or higher (out of a possible ten) combined with clinical synovitis not better 
explained by another disease confirms a diagnosis of “definite RA” (Aletaha et al). 


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8.1.5 
Prior and Concomitant Medication 
All prior medications used within the last 30 days before the first screening examination 
through the follow-up safety visit will be documented. An assessment of RA treatments will 
also be collected. 
8.2 
Tc 99m Tilmanocept Preparation and Administration 
Based on the dose group the subject is enrolled in, the dose of Tc 99m tilmanocept should be 
ordered from Cardinal Health once the subject has been scheduled for IV administration and 
imaging. 
In Nuclear Medicine, prior to initial imaging, an indwelling catheter will be placed for venous 
access. The preferred site of IV placement will be the left or right antecubital vein unless both 
elbows are primary sites for RA imaging, and then the site for IV placement will be between 
the elbow and the wrist. 
The filled syringe(s) will be connected to the Y-port for a slow push injection. Immediately 
after the completion of the injection, a 10 cc sterile normal saline flush will be administered. 
Injection of Tc 99m tilmanocept will be at study time 0:00. 
8.3 
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 whole body (head to toe) planar scan using auto-contour should be obtained at the following 
time points: 
Groups 1-9 (see Appendix 2): 
• 
60 ±  15 minutes post administration 
• 
180 ± 15 minutes post administration 
 


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Groups 10-11 (see Appendix 3): 
• 
15 ± 5 minutes post administration 
• 
60 ± 15 minutes post administration 
• 
180 ± 15 minutes post administration 
• 
18-20 hours post administration 
 
It is anticipated that the whole body 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 the 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 whole body scan. 
Following the whole body acquisition, a planar image with both hands in the FOV should be 
acquired for ~ 5-7 minutes in both the anterior and posterior view for subjects in Groups 1-9 
and subjects in Groups 10 and 11 at 60 ± 15 and 180 ± 15 minutes post-administration .  
For subjects enrolled in Groups 1-9: Following completion of the the planar imaging session 
at 60 minutes, a 3D SPECT or SPECT/CT should be obtained on Joints A and B, followed by 
a patient break. Following the 180 minute whole body scan, and planar image of the hands, 3D 
SPECT or SPECT/CT will be repeated for joint A and completed for joint B and/or C in 
accordance with the prioritization below. No greater than 3 SPECT or SPECT/CT imaging 
sessions will be obtained after the whole body planar imaging session. 
In the event that multiple joints of interest are identified during whole body planar imaging, 
and the planar image of the hands, the following prioritization will be applied for the selection 
of joints for SPECT or SPECT/CT imaging. Prioritization for joint selection may be revised 
after review of the first group of images. Any changes in the prioritization for joint selection 
will be communicated to investigators prior to the fulfillment of additional doses. Images will 
be obtained with bilateral joints in the field of view, including both hands, shoulders, elbows, 
knees.   
“Greatest interest” will be defined as the joint of highest Tc99m localization on imaging and 
as guided by concordance with DAS 28 finding.  
1. Joint A will be a the joint of greatest interest as defined above from the planar 
imaging session. In the event a determination could not be made due to equivalent 
localization in multiple joints, the hands  will be selected as joint A.  3D SPECT or 
SPECT/CT  of Joint A will occur after the 60 and 180 minute whole body scan. 
2. Joint B will be the joint of second greatest interest (knee or elbow or shoulder) or the 
hands if not already imaged as Joint A. 3D SPECT or SPECT/CT of Joint B will 
follow the 180 minute whole body scan. 


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3. Joint C would be imaged in the event that Joints A or B were no longer identified as a 
joint of interest from the 180 minutes whole body imaging. Joint C would be 
determined by the interest and the discretion of the investigator. 
 
In the event all DAS 28 joints are negative for joint localization on whole body imaging, 
SPECT/CT may be performed on other areas of localization. A visual representation of the 
Visit 2, Day 1 imaging procedures for subjects enrolled in Groups 1-9 can be found in 
Appendix 2.  
It is anticipated that each SPECT or SPECT/CT scan would be acquired (at one bed position) 
for 25-30 minutes (30 minutes preferred). A break should address the comfort of the patient, 
encouraging the subject to void, consume fluids or food and to move or rest freely. For areas 
of low activity uptake, the SPECT scan may need to be increased. It is very important that the 
subject remain motionless for the duration of the 3D SPECT 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 whole body scan, planar imaging,  as well as the SPECT and 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 eCRFs (electronic case report forms). 
8.4 
Pharmacokinetics 
Blood will be collected in the contralateral arm that study drug was administered for the 
purpose of PK analysis for subjects enrolled in Groups 10 and 11 at the timepoints described 
below:   
• 
Within 15 minutes prior to administration of Tc 99m tilmanocept 
• 
Immediately following administration of Tc 99m Tilmanocept (00:00-00:05) 
• 
15 ± 5 minutes post injection 
• 
60 ±  15 minutes post injection  
• 
180 ±15 minutes post injection 
• 
18-20 hours post injection 
 
Use of an intravenous catheter is not required for PK blood sampling.  If an IV catheter is used, 
at least 3 mL of blood must be drawn and discareded prior to collecting the PK blood sample. 
Blood samples will be be collected in 5.5 mL heparinized vacutainer tubes.  


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Radioactivity in approximately 2 mL of whole blood will be quantitated in a calibrated well 
counter and decay corrected to the time of study drug administration. The radioactivity in urine 
will be obtained from cumulative quantitative planar imaging counts of the bladder wall 
obtained from radiation dosimetry.  
8.4.1 
Sample Processing  
Whole blood samples will be processed in a well counter immediately following collection. 
8.5 
Dosimetry 
Gamma camera planar images will undergo dosimetry analysis of all target organs. Dosimetry 
will be based on target organ image counts and modeling per OLINDA/EXM®2.0 (Organ 
Level INternal Dose Assessment/EXponential Modeling) software supplied by Hermes 
Medical Solutions (https://www.hermesmedical.com/products/oncology-dosimetry-olinda-
theranostics/). 
8.6 
Safety  
8.6.1 
Adverse Events 
8.6.1.1 
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 2 
Baseline (Day 1) through the assessment of Visit 3 Follow-up () are to be reported as AEs.  
AEs continuing after study completion will be followed to normalization or stabilization. 
Additionally, untoward medical events occurring prior to the day of Tc 99m tilmanocept 
administration will be collect and added to the subject’s medical history unless they are related 
to a study procedure in which case the event will be recorded as an AE.    SAEs will be reported 
from the time of consent through the end of participation. 


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8.6.1.2 
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 8.6.1.5. 
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 adverse event is transient and easily tolerated by the subject. 
• 
Moderate -  The adverse event causes the subject discomfort and interrupts the 
subject’s usual activities 
• 
Severe -  
The adverse event 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 adverse 
event 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. 


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Causal relationship to study procedure 
The investigator will use the following definitions to assess the relationship of the adverse 
event tostudy procedure:  
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. 
8.6.1.3 
Assessments and Documentation of Adverse Events 
Attention shall be paid to the occurrence of AEs for the duration of subject participation. Events 
occurring prior to Visit 2 (day of Tc 99m tilmanocept administration) will be recorded in the 
subject’s medical history. Untoward medical events beginning on Visit 2 (day of Tc 99m 
tilmanocept administration) through the completion of the Visit 3 Follow-up will be reported 
as adverse events.  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 serious adverse events is provided in Section 8.6.1.5.   
• 
The maximum intensity (mild, moderate, or severe). 
• 
Whether drug treatment was administered for the event, any specific drug treatment 
must be documented. 


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• 
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]). 
8.6.1.4 
Expected Adverse Events 
Investigational Product-Related Risks 
In all completed studies of Lymphoseek, involving 553 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 Lymphoseek by the investigator. The most common adverse reactions 
(incident <1%) have been lack of effect (<0.067%), injection site pain (<0.02%) and rash 
(<0.02%). Adverse events from the radioactive dose are not expected, since the applied 
radiation doses are far below doses that can cause acute effects in human tissues.  
In addition to the Lymphoseek pre-approval clinical studies, post-marketing surveillance 
shows that Lymphoseek has been administered to more than 180,000  patients with not a single 
drug-related SAE.  Routes of administration included: subcutaneous, intradermal, and 
peritumoral. The intended route of administration in this study is intravenous.   
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 adverse event 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. 


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8.6.1.5 
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 Food 
and Drug Administration (FDA) and effective 06 Apr 1998.   
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 
adverse events 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 Adverse Event eCRFs. 


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Notification of the IRB(s) 
The sponsor and/or the investigator will notify the IRB(s) about all relevant events (e.g., 
serious adverse events [SAEs] and Suspected, Unexpected, Serious Adverse Reactions 
[SUSARs]) according to all applicable regulations. 
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. 
8.6.2 
Further Safety Assessments 
8.6.2.1 
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). Only body 
weight will be repeating at the Visit 3 (Follow-Up). 
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) 
• 
Lymph nodes 
• 
Musculoskeletal 
• 
Nervous system 
 
8.6.2.2 
Vital Signs 
Vital signs comprise the measurement of body temperature, heart rate, 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 investigational product 
injection, within 15 minutes post injection, and at the follow-up safety visit. 
  


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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. 
8.6.2.3 
Electrocardiogram 
A standard 12-lead electrocardiogram (ECG) will be obtained at screening, as well as 15 
minutes before investigational product injection, within 15 minutes after investigational 
product injection, and at the follow-up safety visit. 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 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.   
8.6.2.4 
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, 
potassium, sodium, total protein, albumin, carbon dioxide 
(CO2)/bicarbonate, blood urea nitrogen (BUN), glucose  
Urinalysis 
pH, specific gravity 
Rheumatoid Panel* 
Erythrocyte Sedimentation Rate (ESR), C-Reactive Protein 
(CRP), Rheumatoid Factor (RF); Anti-Citrullinated Peptide 
Antibody (ACPA) 
* Only at screening; The site laboratory will provide the necessary supplies to collect the blood 
and urine samples. 


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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. 
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 
RA Panel     4 mLs 
 
13 mL (~2.6 tsp) 
Pre-Injection 
PK Sample 
5.5 mL (~1.1 tsp) 
Post-Injection  
PK Samples  
26 mL (~5.3 tsp) 
Follow-up 
Chemistry    5 mLs 
Hematology 4 mLs 
9 mL (~2 tsp) 
TOTAL: 
 
54 mL (~11 tsp) 
 


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9 
STATISTICAL METHODS 
The study is a prospective, open-label, multicenter, dose escalation, and safety with PK and 
dosimetry study of injected Tc 99m tilmanocept in the detection of and assessment of 
localization to skeletal joints in subjects with and without active RA by SPECT and SPECT/CT 
imaging.  The dose escalation portion of the study is a modified 3+3 design with no dose de-
escalation as described in Chevret (2006).  The statistical objective of the study is to determine 
the maximum tolerated dose (MTD) of Tc 99m tilmanocept and to compare the capabilities of 
Tc99m tilmanocept between subjects with and without RA. 
A study center is defined as a treatment administration site or group of treatment administration 
sites under the control and supervision the same Primary Investigator.   
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 pharmacologic activity 
or an 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. 
9.3 
Efficacy Variables 
The efficacy variables for this study are as follows: 
• 
Nuclear medicine specialist qualitative determination of presence/absence of radiotracer 
uptake (i.e., Tc 99m tilmanocept localization positive/negative) relative to background 
from SPECT imaging results for each DAS28 joint location for each subject (deemed the 
“gold standard” result, 28 joint locations per subject) 
• 
Clinically identified presence/absence of RA using the DAS28 instrument (28 joint 
locations per subject) 
• 
Quantitative planar imaging intensity of radiotracer uptake for each joint location 
 
A location/joint would be considered clinically identified as RA positive if it is either swollen 
or tender as measured by the DAS28 instrument.  The presence of radiotracer uptake relative 
to background as determined by the nuclear medicine specialist at a location would indicate 
the location is RA positive. 
The efficacy endpoints for this study are as follows: 
• 
Per subject localization rate of Tc 99m tilmanocept by SPECT imaging  
• 
Per joint location clinically RA-identified joint localization rate of Tc 99m 
tilmanocept by SPECT imaging  


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• 
Concordance of Tc 99m localization with anatomical areas of active RA defined by 
clinical symptomology. 
 
• 
Localization intensity for each clinically RA-defined joint (as determined by 
quantitative SPECT gamma counts)  
• 
Per subject localization rate of Tc 99m tilmanocept in areas other than RA-positive 
joints by SPECT imaging  
• 
The following PK parameters will be calculated for each subject, whenever possible, 
using Tc 99m tilmanocept total radioactivity in whole blood and urine: maximum 
observed concentration (Cmax), time to Cmax (tmax), area under the concentration-time 
curve (AUC) from Hour 0 to the last measureable concentration (AUC0-t), AUC 
extrapolated to infinity (AUC0-∞), apparent terminal elimination rate constant (λZ), 
and apparent terminal elimination half-life (t1/2).   
• 
Radiation dosimetry of Tc 99m tilmanocept will include radiation doses per organ 
(μGy/MBq), organs receiving the highest organ dose (HT), effective doses per organ 
(μSv/MBq), mean effective dose (μSv/MBq), effective dose resulting from a 
diagnostic dose  ± 20%. Dosimetry will be based on target organ image counts and 
modeling per OLINDA/EXM® 2.0 software. 
 
Per subject localization of Tc99m by SPECT imaging is defined as the illumination of at least 
one site relative to background in a subject as determined by the nuclear medicine specialist.   
Per joint location RA-identified localization rate is the number of joint locations that are 
clinically RA positive and SPECT illuminated divided by the total number of joints. 
Per joint location concordance of Tc99m localization to clinically positive RA joints is the 
number of joint locations that are clinically RA positive and SPECT illuminated divided by 
the number of clinically RA positive joints. 
Per subject localization rate of Tc99m by SPECT imaging in areas other than RA-positive 
joints is defined as the illumination of at least one joint location relative to background that is 
clinically RA negative in a subject divided by the number of subjects. 
9.4 
Sample Size Justification 
The dose escalation portion of this study is a modified 3+3 design with no dose de-escalation.  
The sequential cohort enrollment characteristics of this design do not allow a fixed 
computation of sample size.  The parameters of the design that can be calculated are shown in 
Table 4.  Based on an assumed vector of probabilities of any pharmacologic activity or an ADR 
for a subject, the probability of each dose being the MTD is shown in the table.  The average 
sample size of the dose escalation phase of the study is n=26.3 patients for the assumed vector 
of probabilities of any pharmacologic activity or an ADR. 


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Table 4. 
Design Characteristics for the Study Design 
Dose Group 
1 
2 
3 
4 
5 
6 
7 
8 
9 
Total 
Assumed Probability 
of any pharmacologic 
activity or a ADR for 
a subject 
.01 
.02 
.05 
.08 
.10 
.13 
.16 
.19 
.22 
--- 
Probability of the 
Dose Being the MTD 
.002 
.012 
.039 
.071 
.103 
.136 
.152 
.148 
.065 
--- 
Average Sample Size 
for Each Dose and 
Total 
3.1 
3.2 
3.4 
3.5 
3.3 
3.2 
2.8 
2.2 
1.6 
26.3 
 
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 in 
Groups 1-9, injected with Tc 99m tilmanocept, and received whole body planar 
imagingwill be included in the ITD analysis population. 
• 
Radiopharmacokinetic (RPK) Population – Subjects that have been enrolled in the 
study in Groups 10-11, injected with Tc 99m tilmanocept, and received whole body 
planar imaging will be included in the RPK analysis population. 
• 
Safety Population – All patients who are enrolled and injected with Tc 99m 
tilmanocept in the study will be included in the safety population. 
• 
Per Protocol (PP) Population – The PP population will include all safety subjects 
without major protocol violations. 
 
All safety data analyses will be conducted on the safety population.  All efficacy data analyses 
will be conducted on the ITD and PP population with the ITD population being the primary 
analysis set. All analyses of RPK data will be carried out on the RPK 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 
Per subject localization rates and percentage will be calculated by imaging time point by dose 
group and overall.   


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Per joint location localization rates and percentages will be calculated by time point by dose 
group and overall.   
Per location concordance of clinical diagnosis to nuclear medicine specialist illumination 
indication will be computed across all RA positive sites of all subjects at all time points.   
Localization intensities will be summarized by descriptive statistics (mean, median, standard 
deviation, minimum, maximum and range) by joint location at each time point.   
Other efficacy analyses will be described in the Statistical Analysis Plan (SAP) for the study. 
9.5.4 
Analysis of RPK and Dosimetry Variables 
The RPK variables for Groups 10 and 11 are the whole blood and urine radioactivity levels per 
time point.  The radioactivity in whole blood will be measured directly using the blood 
samples.  The radioactivity of urine will be obtained from counts of the bladder wall obtained 
from radiation dosimetry.  RPK parameters will be estimated for each subject using 
noncompartmental models.  Summary statistics (n, mean, std, CV, geometric mean) will be 
computed for each RPK parameter across all subjects by Group (10, 11). 
For dosimetry data, for each dosimetry parameter, summary statistics (n, mean, std, and a one 
standard deviation width confidence interval) will be computed by organ type and overall 
across all subjects by Group (10, 11). Analysis of Safety Variables 
The number and percentage of subjects with any pharmacologic activity or ADRs will be 
tabulated by dose group and overall. 
All adverse events (AEs) will be observed for each subject from the time of signing of informed 
consent until exit from the study.  A treatment emergent adverse event is defined as an adverse 
event 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 adverse events (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 Statistical Analysis Plan (SAP) for the study. 


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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.  For a specific location/joint, the patient must have data for the both 
the SPECT response value and the clinical evaluation of RA in order to be used in the statistical 
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 entered onto the electronic CRFs (provided by the sponsor) as soon as possible. 
10.1.1 
CRF Design 
Electronic CRFs (eCRFs) 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 investigator site file 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 trial master file and the investigator site file, 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. Centers for Disease Control and Protection. Prevalence of doctor-diagnosed arthritis-
attributable activity information- United States, 2010-2012. MMWR. 2012;62(44); 
869-873. Information available at http://www.cdc.gov/mmwr/PDF/wk/mm6244.pdf. 
2. Ma Y, Pope RM. The role of macrophages in rheumatoid arthritis. Curr Pharm Des. 
2005;11(5):569-80. PMID: 15720276 
3. Maradit-Kremers H, Crowson CS, Nicola PJ, Ballman KV, Roger VL, Jacobsen SJ, 
Gabriel SE. Increased unrecognized coronary heart disease and sudden deaths in 
rheumatoid arthritis: a population-based cohort study. Arthritis Rheum. 2005 
Feb;52(2):402-11. PMID: 15693010 
4. Carmona L, Cross M, Williams B, Lassere M, March L. Rheumatoid arthritis. Best 
Pract Res ClinRheumatol. 2010 Dec;24(6):733-45. PMID: 21665122 
5. Oliver JE, Silman AJ. Why are women predisposed to autoimmune rheumatic 
diseases? Arthritis Res Ther. 2009;11(5):252. PMID: 19863777 
6. Helmick CG, Felson DT, Lawrence RC, Gabriel S, Hirsch R, Kwoh CK, Liang MH, 
Kremers HM, Mayes MD, Merkel PA, Pillemer SR, Reveille JD, Stone JH; National 
Arthritis Data Workgroup. Estimates of the prevalence of arthritis and other 
rheumatic conditions in the United States. Part I. Arthritis Rheum. 2008 Jan;58(1):15-
25. 18163481 
7. Scott DL, Wolfe F, Huizinga TW. Rheumatoid arthritis. Lancet. 2010 Sep 
25;376(9746):1094-108. PMID: 20870100 
8. Hootman JM, Helmick CG. Projections of US prevalence of arthritis and associated 
activity limitations. Arthritis Rheum 2006; 54:226–9. PMID: 16385518 
9. Raza K, Falciani F, Curnow SJ, Ross EJ, Lee CY, Akbar AN, Lord JM, Gordon C, 
Buckley CD, Salmon M. Early rheumatoid arthritis is characterized by a distinct and 
transient synovial fluid cytokine profile of T cell and stromal cell origin. Arthritis Res 
Ther. 2005;7(4):R784-95. PMID: 15987480 
10. Olszewski WL, Pazdur J, Kubasiewicz E, Zaleska M, Cooke CJ, Miller NE. Lymph 
draining from foot joints in rheumatoid arthritis provides insight into local cytokine 
and chemokine production and transport to lymph nodes. Arthritis Rheum. 2001 
Mar;44(3):541-9. PMID: 11263768 
11. Meyer PW, Hodkinson B, Ally M, Musenge E, Wadee AA, Fickl H, Tikly M, 
Anderson R. Circulating cytokine profiles and their relationships with autoantibodies, 
acute phase reactants, and disease activity in patients with rheumatoid arthritis. 
Mediators Inflamm. 2010;2010:158514. PMID: 21437211 
12. Leizer T, Cebon J, Layton JE, Hamilton JA. Cytokine regulation of colony-
stimulating factor production in cultured human synovial fibroblasts: I. Induction of 
GM-CSF and G-CSF production by interleukin-1 and tumor necrosis factor. Blood. 
1990 Nov 15;76(10):1989-96. PMID: 1700731 


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13. Westra J, Doornbos-van der Meer B, de Boer P, van Leeuwen MA, van Rijswijk MH, 
Limburg PC. Strong inhibition of TNF-alpha production and inhibition of IL-8 and 
COX-2 mRNA expression in monocyte-derived macrophages by RWJ 67657, a p38 
mitogen-activated protein kinase (MAPK) inhibitor. Arthritis Res Ther. 2004;6(4): 
R384-92. PMID: 15225374 
14. Hamilton JA, Tak PP. The dynamics of macrophage lineage populations in 
inflammatory and autoimmune diseases. Arthritis Rheum. 2009 May;60(5):1210-21. 
PMID: 19404968 
15. Keffer J, Probert L, Cazlaris H, Georgopoulos S, Kaslaris E, Kioussis D, Kollias G. 
Transgenic mice expressing human tumour necrosis factor: a predictive genetic model 
of arthritis. EMBO J. 1991 Dec;10(13):4025-31. PMID: 1721867 
16. Noack M, Miossec P. Th17 and regulatory T cell balance in autoimmune and 
inflammatory diseases. Autoimmun Rev. 2014 Jun;13(6):668-77. PMID: 24418308 
17. Bugatti S, Vitolo B, Caporali R, Montecucco C, Manzo A. B cells in rheumatoid 
arthritis: from pathogenic players to disease biomarkers. Biomed Res Int. 
2014;2014:681678. PMID: 24877127 
18. Boissier MC, Semerano L, Challal S, Saidenberg-Kermanac'h N, Falgarone G. 
Rheumatoid arthritis: from autoimmunity to synovitis and joint destruction. J 
Autoimmun. 2012 Sep;39(3):222-8. PMID: 22704962 
19. Tran CN, Lundy SK, Fox DA. Synovial biology and T cells in rheumatoid arthritis. 
Pathophysiology. 2005 Oct;12(3):183-9. PMID: 16112560 
20. Kinne RW, Stuhlmüller B, Burmester GR. Cells of the synovium in rheumatoid 
arthritis. Macrophages. Arthritis Res Ther. 2007;9(6):224. PMID: 18177511 
21. Zwerina J, Redlich K, Polzer K, Joosten L, Krönke G, Distler J, Hess A, Pundt N, Pap 
T, Hoffmann O, Gasser J, Scheinecker C, Smolen JS, van den Berg W, Schett G. 
TNF-induced structural joint damage is mediated by IL-1. Proc Natl Acad Sci U S A. 
2007 Jul 10;104(28):11742-7. PMID:17609389 
22. Feldmann M, Brennan FM, Maini RN. Role of cytokines in rheumatoid arthritis. 
Annu Rev Immunol. 1996;14:397-440. 
23. Schett G, Zwerina J, Firestein G. The p38 mitogen-activated protein kinase (MAPK) 
pathway in rheumatoid arthritis. Ann Rheum Dis. 2008 Jul;67(7):909-16. PMID: 
17827184 
24. Kalden JR. Emerging role of anti-tumor necrosis factor therapy in rheumatic diseases. 
Arthritis Res. 2002;4 Suppl 2:S34-40. PMID: 12110156. 
25. Wiedmann MW1, Mössner J, Baerwald C, Pierer M. TNF alpha inhibition as 
treatment modality for certain rheumatologic and gastrointestinal diseases. Endocr 
Metab Immune Disord Drug Targets. 2009 Sep;9(3):295-314. PMID: 19594416 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
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Page 69 of 83 
 
26. Chen YF, Jobanputra P, Barton P, Jowett S, Bryan S, Clark W, Fry-Smith A, Burls A. 
A systematic review of the effectiveness of adalimumab, etanercept and infliximab 
for the treatment of rheumatoid arthritis in adults and an economic evaluation of their 
cost-effectiveness. Health Technol Assess. 2006 Nov;10(42):iii-iv, xi-xiii, 1-229. 
PMID: 17049139 
27. Vivar N, Van Vollenhoven RF. Advances in the treatment of rheumatoid arthritis. 
F1000Prime Rep. 2014 May 6; 6:31. doi: 10.12703/P6-31. eCollection 2014. Review. 
PMID: 24860653 
28. Wasserman AM. Diagnosis and management of rheumatoid arthritis. Am Fam 
Physician. 2011 Dec 1;84(11):1245-52. PMID: 22150658 
29. Saag KG, Teng GG, Patkar NM, Anuntiyo J, Finney C, Curtis JR, Paulus HE, 
Mudano A, Pisu M, Elkins-Melton M, Outman R, Allison JJ, Suarez Almazor M, 
Bridges SL Jr, Chatham WW, Hochberg M, MacLean C, Mikuls T, Moreland LW, 
O'Dell J, Turkiewicz AM, Furst DE; American College of Rheumatology. American 
College of Rheumatology 2008 recommendations for the use of nonbiologic and 
biologic disease-modifying antirheumatic drugs in rheumatoid arthritis. Arthritis 
Rheum. 2008 Jun 15;59(6):762-84. PMID: 18512708 
30. Choy EH, Smith C, Doré CJ, Scott DL. A meta-analysis of the efficacy and toxicity 
of combining disease-modifying anti-rheumatic drugs in rheumatoid arthritis based 
on patient withdrawal. Rheumatology (Oxford). 2005 Nov;44(11):1414-21. PMID: 
16030080 
31. Donahue KE, Gartlehner G, Jonas DE, Lux LJ, Thieda P, Jonas BL, Hansen RA, 
Morgan LC, Lohr KN. Systematic review: comparative effectiveness and harms of 
disease-modifying medications for rheumatoid arthritis. Ann Intern Med. 2008 Jan 
15;148(2):124-34. PMID: 18025440 
32. Gramling A, O'Dell JR. Initial management of rheumatoid arthritis. Rheum Dis Clin 
North Am. 2012 May;38(2):311-25. PMID: 22819086 
33. Barbhaiya M, Solomon DH. Rheumatoid arthritis and cardiovascular disease: an 
update on treatment issues. Curr Opin Rheumatol. 2013 May;25(3):317-24. PMID: 
23466960 
34. Bykerk V. Unmet needs in rheumatoid arthritis. J Rheumatol Suppl. 2009 Jun;82:42-
6. PMID:19509330 
35. Demoruelle MK, Deane KD. Treatment strategies in early rheumatoid arthritis and 
prevention of rheumatoid arthritis. Curr Rheumatol Rep. 2012 Oct;14(5):472-80. 
PMID: 22773387 
36. Sacre SM, Drexler SK, Andreakos E, Feldmann M, Brennan FM, Foxwell BM. Could 
toll-like receptors provide a missing link in chronic inflammation in rheumatoid 
arthritis? Lessons from a study on human rheumatoid tissue. Ann Rheum Dis. 2007 
Nov;66 Suppl 3:iii81-6. PMID: 17934103 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
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Page 70 of 83 
 
37. Anderson JJ, Wells G, Verhoeven AC, Felson DT. Factors predicting response to 
treatment in rheumatoid arthritis: the importance of disease duration. Arthritis 
Rheum. 2000 Jan;43(1):22-9. PMID:10643696 
38. van der Linden MP, le Cessie S, Raza K, van der Woude D, Knevel R, Huizinga TW, 
van der Helmvan Mil AH. Long-term impact of delay in assessment of patients with 
early arthritis. Arthritis Rheum. 2010 Dec;62(12):3537-46. PMID: 20722031 
39. Nell VP, Machold KP, Eberl G, Stamm TA, Uffmann M, Smolen JS. Benefit of very 
early referral and very early therapy with disease-modifying anti-rheumatic drugs in 
patients with early rheumatoid arthritis. Rheumatology (Oxford). 2004 Jul;43(7):906-
14. PMID: 15113999 
40. Cush JJ. Early rheumatoid arthritis -- is there a window of opportunity? J Rheumatol 
Suppl. 2007 Nov;80:1-7. PMID: 17985417 
41. Zwerina J, Redlich K, Schett G, Smolen JS. Pathogenesis of rheumatoid arthritis: 
targeting cytokines. Ann N Y Acad Sci. 2005 Jun;1051:716-29. PMID: 16127012 
42. Aletaha D, Neogi T, Silman AJ, Funovits J, Felson DT, Bingham CO 3rd, Birnbaum 
NS, Burmester GR, Bykerk VP, Cohen MD, Combe B, Costenbader KH, Dougados 
M, Emery P, Ferraccioli G, Hazes JM, Hobbs K, Huizinga TW, Kavanaugh A, Kay J, 
Kvien TK, Laing T, Mease P, Ménard HA, Moreland LW, Naden RL, Pincus T, 
Smolen JS, Stanislawska-Biernat E, Symmons D, Tak PP, Upchurch KS, Vencovský 
J, Wolfe F, Hawker G. 2010 Rheumatoid arthritis classification criteria: an American 
College of Rheumatology/European League Against Rheumatism collaborative 
initiative. Arthritis Rheum. 2010 Sep;62(9):2569-81. PMID: 20872595 
43. Cohen S, Emery P. The American College of Rheumatology/European League 
Against Rheumatism criteria for the classification of rheumatoid arthritis: a game 
changer. Arthritis Rheum. 2010 Sep;62(9):2592-4. PMID: 20872597 
44. Humphreys JH, Symmons DP. Postpublication validation of the 2010 American 
College of Rheumatology/European League Against Rheumatism classification 
criteria for rheumatoid arthritis: where do we stand? Curr Opin Rheumatol. 2013 
Mar;25(2):157-63. PMID: 23274519 
45. Berglin E, Dahlqvist SR. Comparison of the 1987 ACR and 2010 ACR/EULAR 
classification criteria for rheumatoid arthritis in clinical practice: a prospective cohort 
study. Scand J Rheumatol. 2013;42(5):362-8. PMID: 23607599 
46. Varache S, Cornec D, Morvan J, Devauchelle-Pensec V, Berthelot JM, Le Henaff-
Bourhis C, Hoang S, Thorel JB, Martin A, Chalès G, Nowak E, Jousse-Joulin S, 
Youinou P, Saraux A. Diagnostic accuracy of ACR/EULAR 2010 criteria for 
rheumatoid arthritis in a 2-year cohort. J Rheumatol. 2011 Jul;38(7):1250-7. PMID: 
21572146 
47. Liao KP, Bykerk V. All that glitters is not gold--standardizing diagnosis in 
rheumatoid arthritis studies. J Rheumatol. 2011 Jul;38(7):1223-4. PMID: 21724709 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
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Page 71 of 83 
 
48. Biliavska I, Stamm TA, Martinez-Avila J, Huizinga TW, Landewé RB, Steiner G, 
Aletaha D, Smolen JS, Machold KP. Application of the 2010 ACR/EULAR 
classification criteria in patients with very early inflammatory arthritis: analysis of 
sensitivity, specificity and predictive values in the SAVE study cohort. Ann Rheum 
Dis. 2013 Aug;72(8):1335-41. PMID: 22984174 
49. de Hair MJ, Lehmann KA, van de Sande MG, Maijer KI, Gerlag DM, Tak PP. The 
clinical picture of rheumatoid arthritis according to the 2010 American College of 
Rheumatology/European League Against Rheumatism criteria: is this still the same 
disease? Arthritis Rheum. 2012 Feb;64(2):389-93. PMID: 21918954 
50. Alves C, Luime JJ, van Zeben D, Huisman AM, Weel AE, Barendregt PJ, Hazes JM. 
Diagnostic performance of the ACR/EULAR 2010 criteria for rheumatoid arthritis 
and two diagnostic algorithms in an early arthritis clinic (REACH). Ann Rheum Dis. 
2011 Sep;70(9):1645-7. PMID: 21622769 
51. Britsemmer K, Ursum J, Gerritsen M, van Tuyl LH, van Schaardenburg D. Validation 
of the 2010 ACR/EULAR classification criteria for rheumatoid arthritis: slight 
improvement over the 1987 ACR criteria. Ann Rheum Dis. 2011 Aug;70(8):1468-70. 
PMID: 21586440 
52. Cader MZ, Filer A, Hazlehurst J, de Pablo P, Buckley CD, Raza K. Performance of 
the 2010 ACR/EULAR criteria for rheumatoid arthritis: comparison with 1987 ACR 
criteria in a very early synovitis cohort. Ann Rheum Dis. 2011 Jun;70(6):949-55. 
PMID: 21285117 
53. Kaneko Y, Kuwana M, Kameda H, Takeuchi T. Sensitivity and specificity of 2010 
rheumatoid arthritis classification criteria. Rheumatology (Oxford). 2011 
Jul;50(7):1268-74. PMID: 21292733 
54. van der Linden MP, Knevel R, Huizinga TW, van der Helm-van Mil AH. 
Classification of rheumatoid arthritis: comparison of the 1987 American College of 
Rheumatology criteria and the 2010 American College of Rheumatology/European 
League Against Rheumatism criteria. Arthritis Rheum. 2011 Jan;63(1):37-42. PMID: 
20967854 
55. Sakellariou G, Scirè CA, Zambon A, Caporali R, Montecucco C. Performance of the 
2010 classification criteria for rheumatoid arthritis: a systematic literature review and 
a meta-analysis. PLoS One. 2013;8(2):e56528. PMID: 23437156 
56. Zeidler H. The need to better classify and diagnose early and very early rheumatoid 
arthritis. J Rheumatol. 2012 Feb;39(2):212-7. PMID: 22174195 
57. Vonkeman HE, van de Laar MA. The new European League Against 
Rheumatism/American College of Rheumatology diagnostic criteria for rheumatoid 
arthritis: how are they performing? Curr Opin Rheumatol. 2013 May;25(3):354-9. 
PMID: 23492742 


Tc 99m tilmanocept 
 
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58. Kraan MC, Reece RJ, Smeets TJ, Veale DJ, Emery P, Tak PP. Comparison of 
synovial tissues from the knee joints and the small joints of rheumatoid arthritis 
patients: Implications for pathogenesis and evaluation of treatment. Arthritis Rheum. 
2002 Aug; 46(8):2034-8. PMID: 12209505 
59. Cutolo M, Sulli A, Barone A, Seriolo B, Accardo S. Macrophages, synovial tissue 
and rheumatoid arthritis. Clin Exp Rheumatol. 1993 May-Jun;11(3):331-9. PMID: 
8394794 
60. Kennedy A, Fearon U, Veale DJ, Godson C. Macrophages in synovial inflammation. 
Front Immunol. 2011 Oct 10;2:52. PMID: 22566842 
61. Bresnihan B. Pathogenesis of joint damage in rheumatoid arthritis. J Rheumatol. 1999 
Mar;26(3):717-9. PMID: 10090189 
62. Mulherin D1, Fitzgerald O, Bresnihan B. Synovial tissue macrophage populations 
and articular damage in rheumatoid arthritis. Arthritis Rheum. 1996 Jan;39(1):115-24. 
PMID: 8546720 
63. Yanni G, Whelan A, Feighery C, Bresnihan B. Synovial tissue macrophages and joint 
erosion in rheumatoid arthritis. Ann Rheum Dis. 1994 Jan;53(1):39-44. PMID: 
8311554 
64. Deane KD, Norris JM, Holers VM. Preclinical rheumatoid arthritis: identification, 
evaluation, and future directions for investigation. Rheum Dis Clin North Am. 2010 
May;36(2):213-41. PMID: 20510231 
65. El-Gabalawy H. The preclinical stages of RA: lessons from human studies and animal 
models. Best Pract Res Clin Rheumatol. 2009 Feb;23(1):49-58. PMID: 19233045 
66. Kraan MC, Versendaal H, Jonker M, Bresnihan B, Post WJ, t Hart BA, Breedveld 
FC, Tak PP. Asymptomatic synovitis precedes clinically manifest arthritis. Arthritis 
Rheum. 1998 Aug; 41(8):1481-8. PMID: 9704648 
67. Demoruelle MK, Deane KD, Holers VM. When and where does inflammation begin 
in rheumatoid arthritis? Curr Opin Rheumatol. 2014 Jan;26(1):64-71. PMID: 
24247116 
68. van de Sande MG1, de Hair MJ, Schuller Y, van de Sande GP, Wijbrandts CA, 
Dinant HJ, Gerlag DM, Tak PP. The features of the synovium in early rheumatoid 
arthritis according to the 2010 ACR/EULAR classification criteria. PLoS One. 
2012;7(5):e36668. PMID: 22574210  
69. Haringman JJ, Gerlag DM, Zwinderman AH, Smeets TJ, Kraan MC, Baeten D, 
McInnes IB, Bresnihan B, Tak PP. Synovial tissue macrophages: a sensitive 
biomarker for response to treatment in patients with rheumatoid arthritis. Ann Rheum 
Dis. 2005 Jun;64(6):834-8. PMID: 15576415 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
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Page 73 of 83 
 
70. Bresnihan B, Pontifex E, Thurlings RM, Vinkenoog M, El-Gabalawy H, Fearon U, 
Fitzgerald O, Gerlag DM, Rooney T, van de Sande MG, Veale D, Vos K, Tak PP. 
Synovial tissue sublining CD68 expression is a biomarker of therapeutic response in 
rheumatoid arthritis clinical trials: consistency across centers. J Rheumatol. 2009 
Aug;36(8):1800-2. PMID: 19671815 
71. Bresnihan B, Gerlag DM, Rooney T, Smeets TJ, Wijbrandts CA, Boyle D, Fitzgerald 
O, Kirkham BW,  McInnes IB, Smith M, Ulfgren AK, Veale DJ, Tak PP. Synovial 
macrophages as a biomarker of response to therapeutic intervention in rheumatoid 
arthritis: standardization and consistency across centers. J Rheumatol. 2007 
Mar;34(3):620-2. PMID: 17343309 
72. Franz JK, Burmester GR: The needle and the damage done. Ann Rheum Dis 2005, 
64:798-800. PMID: 15897300 
73. Kraan MC, Reece RJ, Barg EC, Smeets TJ, Farnell J, Rosenburg R, Veale DJ, 
Breedveld FC, Emery P, Tak PP. Modulation of inflammation and metalloproteinase 
expression in synovial tissue by leflunomide and methotrexate in patients with active 
rheumatoid arthritis. Findings in a prospective, randomized, double-blind, parallel-
design clinical trial in thirty-nine patients at two centers. Arthritis Rheum. 2000 
Aug;43(8):1820-30. PMID: 10943872 
74. Catrina AI, Trollmo C, af Klint E, Engstrom M, Lampa J, Hermansson Y, Klareskog 
L, Ulfgren AK. Evidence that anti-tumor necrosis factor therapy with both etanercept 
and infliximab induces apoptosis in macrophages, but not lymphocytes, in rheumatoid 
arthritis joints: extended report. Arthritis Rheum. 2005 Jan;52(1):61-72. PMID: 
15641091 
75. Cunnane G, Madigan A, Murphy E, FitzGerald O, Bresnihan B. The effects of 
treatment with interleukin-1 receptor antagonist on the inflamed synovial membrane 
in rheumatoid arthritis. Rheumatology (Oxford). 2001 Jan;40(1):62-9. PMID: 
11157143 
76. Vieira-Sousa E1, Gerlag DM, Tak PP. Synovial tissue response to treatment in 
rheumatoid arthritis. Open Rheumatol J. 2011;5:115-22. PMID: 22279510 
77. Hutamekalin P, Saito T, Yamaki K, Mizutani N, Brand DD, Waritani T, Terato K, 
Yoshino S. Collagen antibody-induced arthritis in mice: development of a new 
arthritogenic 5-clone cocktail of monoclonal anti-type II collagen antibodies. J 
Immunol Methods. 2009 Mar 31;343(1):49-55. PMID: 19330909 
78. Hoh C.K., A.M. Wallace, et al. Preclinical studies of [(99m)Tc]DTPA-mannosyl-
dextran. Nucl Med Biol. Jul 2003;30(5):457-464. 
79. Ellner S.J., C.K. Hoh, et al. Dose-dependent biodistribution of [(99m)Tc]DTPA-
mannosyl-dextran for breast cancer sentinel lymph node mapping. Nucl Med Biol. 
Nov 2003;30(8):805-810. 
80. Wallace A.M., C.K. Hoh, et al. Lymphoseek: a molecular imaging agent for 
melanoma sentinel lymph node mapping. Ann Surg Oncol. Feb 2007;14(2):913-921.  


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
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Page 74 of 83 
 
81. Wallace A.M., C.K. Hoh, et al. Sentinel lymph node mapping of breast cancer via 
intradermal administration of Lymphoseek. Nucl Med Biol. Oct 2007;34(7) 5:849-83. 
82. Wallace A.M., C.K. Hoh, et al. Lymphoseek: a molecular radiopharmaceutical for 
sentinel node detection. Ann Surg Oncol. Jun 2003;10(5):531-538. 
83. Wallace A.M., C.K. Hoh, et al. Sentinel lymph node accumulation of Lymphoseek 
and Tc-99m-sulfur colloid using a "2-day" protocol. Nucl Med Biol. Aug 
2009;36(6):687-692. 
84. Leong S.P., J. Kim, 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. Apr 
2011;18(4):961-969. 
85. Tokin C.A., F.O. Cope, 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 ⁹⁹mTc-labeled nanocolloid human serum albumin standard of care. 
Clin Exp Metastasis. 2012 Oct;29(7):681-6. doi:10.1007/s10585-012-9497-x. Epub 
2012 Jun 23. Review. PubMed PMID: 22729510. 
86. Sondak V.K., D.W. King, et al. Combined analysis of phase III trials evaluating 
[99mTc]tilmanocept and vital blue dye for identification of sentinel lymph nodes in 
clinically node-negative cutaneous melanoma. Ann Surg Oncol. 2013 Feb;20(2):680-
8. doi: 10.1245/s10434-012-2612-z. Epub 2012 Oct 3. PubMed PMID: 23054107; 
PubMed Central PMCID: PMC3560941. 
87. Wallace A.M., L.K. Han, 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 Aug;20(8):2590-9. doi: 10.1245/s10434-013-2887-8. Epub 
2013 Mar 17. PubMed PMID: 23504141; PubMed Central PMCID: PMC3705144. 
88. Marcinow A.M., N. Hal, 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 Sep;139(9):895-902. 
doi:10.1001/jamaoto.2013.4239. PubMed PMID: 24051744. 
89. Agrawal A, Civantos FJ, Brumund KT, Chepeha DB, Hall NC, Carroll WR, Smith 
RB, Zitsch RP, Lee WT, Shnayder Y, Cognetti DM, Pitman KT, King DW, 
Christman LA, Lai SY. [(99m)Tc]Tilmanocept Accurately Detects Sentinel Lymph 
Nodes and Predicts Node Pathology Status in Patients with Oral Squamous Cell 
Carcinoma of the Head and Neck: Results of a Phase III Multi-institutional Trial. Ann 
Surg Oncol. 2015  Oct;22(11):3708-15. doi: 10.1245/s10434-015-4382-x. Epub 2015 
Feb 11. PubMed PMID: 25670018; PubMed Central PMCID: PMC4565859. 


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90. Prevoo ML, van ‘t Hof MA, Kuper HH, can Leeuwen MA, van de Putte LB, can Riel 
PL. Modified disease activity scores that include twenty-eight-joint counts.  
Development and validation in a prospective longitudinal study of patients with 
rheumatoid arthritis.  Arthritis Rheum. 1995 Jan; 38(1):44-8. PMID: 7818570. 
91. Chevret, S (2006).  Statistical Methods in Dose Finding Experiments.  New York, 
NY: John Wiley & Sons. 
 
 


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Appendix 1 
Schedule of Events 
Evaluation 
Visit 1 
Screening  
 
Visit 2  
Baseline: Day 1 
Visit 3  
Follow-up 
Days 
 -44 to 0 
Day 1 Prior 
to Injection 
- 00:15 to 
-00:01 
00:00-
00:05 
00:01 to 
00:15 
15± 5 
Mins 
60 ± 15 
Mins 
180 ± 15 Mins  
18-20 Hours 
Post Injectionb 
Day 5 ± 2a 
Informed Consent 
x 
 
 
 
 
 
 
 
 
 
Entry Criteria  
x 
 
 
 
 
 
 
 
 
 
Medical History, RA History and 
Demography 
x 
 
 
 
 
 
 
 
 
 
Urine Drug Screen 
x 
 
 
 
 
 
 
 
 
 
ACR/EULAR 2010 Classification 
x 
 
 
 
 
 
 
 
 
 
Vital Sign Assessment 
x 
 
x 
 
x 
 
 
 
x 
x 
ECG 
x 
 
x 
 
x 
 
 
 
x 
x 
Physical Examination  
x 
 
 
 
 
 
 
 
x 
x 
DAS-28 
x 
 
 
 
 
 
 
 
 
 
Clinical Laboratory Evaluation: 
Chemistry, Hematology, UA   
x 
 
 
 
 
 
 
 
x 
x 
Rheumatoid Arthritis Panel 
x 
 
 
 
 
 
 
 
 
 
Urine Pregnancy Test 
x 
x 
 
 
 
 
 
 
 
 
Tilmanocept Administration 
 
 
 
x 
 
 
 
 
 
 
Whole Body Planar and Optional 
3D SPECT or SPECT/CT Imagea 
 
 
 
 
 
 
xa 
xa 
 
 
Whole Body Planar Image for 
Dosimetry Evaluationb 
 
 
 
 
 
xb 
xb 
xb 
x 
 
Planar SPECT Imaging of Hands 
 
 
 
 
 
 
x 
x 
 
 
Blood Collection for PK 
Analysisb 
 
 
xb 
xb  
 
xb 
xb 
xb 
x 
 
Concomitant Medications 
x 
x 
 
 
 
 
 
 
x 
x 
Adverse Event Monitoring 
x 
x 
x 
x 
x 
x 
x 
x 
x 
x 
a Groups 1-9 only 
b Groups 10 and 11 only


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Appendix 2 
Groups 1-9 Visit 2, Day 1 Diagram (example) 
 


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Appendix 3 
Groups 10-11 Visits 2 and 3 Diagram (example) 
  


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Appendix 4 
2010 ACR/ EULAR Classification Criteria 
 Score 
Target population (Who should be tested?): Patients who 
 
1) Have at least 1 joint with definite clinical synovitis (swelling)* 
 
2) With the synovitis not better explained by another disease† 
 
Classification criteria for RA (score-based algorithm: add score of categories A-D; 
 
             a score of ≥ 6/10 is needed for classification of a patient as having definite 
RA)‡ 
 
A. Joint involvement§ 
 
1 large joint 
0 
2 – 10 large joints 
1 
1 – 3 small joints (with or without involvement of large joints)# 
2 
4- 10 small joints (with or without involvement of large joints) 
3 
> 10 joints (at least 1 small joint)** 
5 
B. Serology (at least 1 test result is needed for classification)‡‡ 
 
Negative RF and negative ACPA 
0 
Low-positive RF or low-positive ACPA 
2 
High-positive RF or high-positive ACPA 
3 
C. Acute-phase reactants (at least 1 test result is needed for classification) ‡‡ 
 
Normal CRP and normal ESR 
0 
Abnormal CRP or abnormal ESR 
1 
D. Duration of symptoms§§ 
 
< 6 weeks 
0 
≥ 6 weeks 
1 
* The criteria are aimed at classification of newly presenting patients. In addition, patients 
with erosive disease typical of rheumatoid arthritis (RA) with a history compatible with prior 
fulfillment of the 2010 criteria should be classified as having RA. Patients with longstanding 
disease, including those whose disease is inactive (with or without treatment) who, based on 
retrospectively available data, have previously fulfilled the 2010 criteria should be classified 
as having RA.  
† Differential diagnoses vary among patients with different presentations, but may include 
conditions such as systemic lupus erythematosus, psoriatic arthritis, and gout. If it is unclear 
about the relevant differential diagnoses to consider, an expert rheumatologist should be 
consulted.  
‡ Although patients with a score of < 6/10 are not classifiable as having RA, their status can 
be reassessed and the criteria might be fulfilled cumulatively over time.  
§ Joint involvement refers to any swollen or tender joint on examination, which may be 
confirmed by imaging evidence of synovitis. Distal interphalangeal joints, first 
carpometacarpal joints, and first metatarsophalangeal joints are excluded from assessment. 


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Categories of joint distribution are classified according to the location and number of 
involved joints, with placement into the highest category possible based on the pattern of 
joint involvement.  
“Large joints” refers to shoulders, elbows, hips, knees, and ankles.  
# “Small joints” refers to the metacarpophalangeal joints, proximal interphalangeal joints, 
second through fifth metatarsophalangeal joints, thumb interphalangeal joints, and wrists.  
** In this category, at least 1 of the involved joints must be a small joint; the other joints can 
include any combination of large and additional small joints, as well as other joints not 
specifically listed elsewhere (e.g., temporomandibular, acromioclavicular, sternoclavicular, 
etc.).  
†† Negative refers to IU values that are less than or equal to the upper limit of normal (ULN) 
for the laboratory and assay; low-positive refers to IU values that are higher than the ULN 
but ≤ 3times the ULN for the laboratory and assay; high-positive refers to IU values that are 
> 3 times the ULN for the laboratory and assay. Where rheumatoid factor (RF) information is 
only available as positive or negative, a positive result should be scored as low-positive for 
RF. ACPA = anti-citrullinated protein antibody.  
‡‡ Normal/abnormal is determined by local laboratory standards. CRP = C-reactive protein; 
ESR = erythrocyte sedimentation rate.  
§§ Duration of symptoms refers to patient self-report of the duration of signs or symptoms of 
synovitis (e.g., pain, swelling, tenderness) of joints that are clinically involved at the time of 
assessment, regardless of treatment status. 
 


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Appendix 5 
DAS-28 Scale 
 
Courtesy of http://www.iche.edu/newsletter/DAS28.pdf 


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Appendix 6 
Sponsor Signatures 
Navidea Biopharmaceuticals, Inc 
Date: 19 June 20 1 8 
Study Title: 
An Evaluation of the Safety of Escalating Doses of Tc 99m 
Tilmanocept by Intravenous (IV) [njection and Skeletal Joint 
Imaging with SPECT in Subjects with Active Rheumatoid 
Arthritis (RA) and Healthy Controls 
Study Number: 
Original Protocol 
Date: 
Amendment I Date: 
Amendment 2 Date: 
Amendment 3 Date: 
Amendment 4 Date: 
Amendment 5 Date: 
NAV3-2l 
23 May 20 16 
12 Sep tern ber 20 I 6 
22 December 2016 
09 February 2017 
23 August 20 17 
18April2018 
rotoco was subject to critical review and has been approved by the 
net contributed to writing and/or approving th is protocol: 
Signed: __ L±z_~::...J<...~------
Michael S. Blue, MD 
Senior Medical Director 
Navidea Biopharmaceuticals 
William J. Regan 
Senior VP, Global Regulatory Af · rs and Quality 
Navidea Biopharmaceuticals 
Confidential 
Page 82 of 83 
,,,,-
Dat~ (/;d~ 
Date:_J_?-r_~_1.,,,.,,.,..,_<_?/,)l y 


Tc 99m tilmanocept 
 
Navidea Biopharmaceuticals, Inc 
Clinical Study Protocol Number: NAV3-21 
 
Date: 19 June 2018 
Confidential 
Page 83 of 83 
 
Appendix 7 
Investigator’s Signature 
Study Title: 
An Evaluation of the Safety of Escalating Doses of Tc 99m 
Tilmanocept by Intravenous (IV) Injection and Skeletal Joint 
Imaging with SPECT in Subjects with Active Rheumatoid 
Arthritis (RA) and Healthy Controls 
Study Number: 
NAV3-21 
Original Protocol 
Date: 
23 May 2016 
Amendment 1 Date: 
Amendment 2 Date: 
Amendment 3 Date: 
Amendment 4 Date: 
Amendment 5 Date: 
Amendment 6 Date: 
12 September 2016 
22 December 2016 
09 February 2017 
23 August 2017 
18 April 2018 
19 June 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: