Imaging Biomarkers of Lymphatic Dysfunction
Phase: NA
Conditions: Secondary Lymphedema
Sponsor: Vanderbilt University Medical Center

 
 
  
  
  
Imaging Biomarkers of Lymphatic Dysfunction  
  
NCT NUMBER: 131172  
VERSION DATE: JULY 7, 2025  
Imaging Biomarkers of Lymphatic Dysfunction  
  
Principle Investigator:  
  
Dr. Manus Donahue, Ph.D.  
Full Professor  
Vanderbilt University Institute of Imaging Science  
Email: m.donahue@vumc.org  
Phone: (615) 322-8350  
Fax: (615) 322-0735  
  
 

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Table of Contents  
  
Study Schema  
1.0 
Background 2.0   
 
2.0 
Rationale and Specific Aims  
3.0 
Inclusion / Exclusion Criteria  
4.0 
Enrollment / Randomization  
5.0 
Study Procedures  
6.0 
Adverse Event Reporting 
7.0 
Study Withdrawal / Discontinuation 
8.0 
Statistical Analysis Plan 
9.0  
Privacy / Confidentiality Issues  
10.0  Follow-up  
11.0 
Record Retention 
12.0 
References 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
1.0   
 Background  
  

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Breast cancer treatment-related lymphedema (BCRL) is a chronic, debilitating disease 
caused by lymphatic flow obstruction and lymphedema of the upper limb and torso 
secondary to necessary cancer therapies, namely lymph node removal.  BCRL has been 
reported to occur in 20-30% of breast cancer survivors following these therapies, which 
is in addition to similar incidence of secondary lymphedema in the upper limb occurring 
in persons with other therapies necessitating axillary lymph node removal and/or 
radiation. However, owing to a lack of methodology for sensitively identifying lymphatic 
system compromise, there are important gaps in our knowledge regarding which patients 
are at highest risk and how and when therapies should be applied to minimize 
impairment. Here, novel and noninvasive magnetic resonance imaging approaches for 
measuring lymphatic dysfunction are applied to improve procedures for preventing, 
predicting, and treating secondary lymphedema.  
  
The long-term goal of this work is to develop and apply novel tools to visualize lymphatic 
dysfunction, and also to incorporate these tools to provide new observables that can be 
used as much needed trial endpoints to optimize therapies and improve patient 
outcomes. This research initiative which started at VUMC over four years ago has led to 
the development of new, noninvasive imaging methods for evaluating LN function and 
morphology here at VUMC, along with approaches for visualizing lymphatic vessels and 
tissue impairment without exogenous contrast agents. These methods have been applied 
by us in healthy adults across the lifespan to evaluate normative parameter ranges, in 
surveillance imaging of BCRL patients at multiple disease stages, and in BCRL patients 
before and after manual lymphatic drainage (MLD) therapy1-4. This work has led to the 
development of multiple new biomarkers that are more sensitive than traditional 
external measures of lymphatic dysfunction.  
  
We will build on the development efforts from our previous and ongoing lymphedema 
research (IRB #131172) to apply methods to evaluate lymphatic dysfunction in multiple 
stages of secondary lymphedema management, including (i) prevention, (ii) sub-clinical 
risk identification, and (iii) therapy.   
  
Below summarizes this study's overall significance:  
1. 
We will utilize multi-parametric anatomical and molecular imaging of LNs, which 
incorporate measures of anatomy and chemical environment, to evaluate abilities to 
discriminate metastatic from benign LNs noninvasively in vivo. These approaches have 
already been applied in other applications of malignancy5-7, and we have developed 
innovative high spatial resolution variants to enable sub-millimeter LN characterization in 
vivo1. If successful, findings would have relevance for noninvasively mapping the 
topography of LN malignancy, which could initiate future investigations to ultimately 
inform surgery decisions and reduce unnecessary LN removal.  
  

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2. 
We will extend our current trial to evaluate sub-clinical internal and external risk 
factors of secondary lymphedema progression in the arm and upper torso following 
cancer therapies post-LN removal and test more refined hypotheses regarding how 
impaired lymphatic pumping, superficial axillary water accumulation, and interstitial 
adipose accumulation portend secondary lymphedema onset and progression. This work 
will be significant for understanding prognostic potential of the new internal markers 
relative to existing risk factors, and it will reinforce using these measures to triage patients 
for disease-modifying therapies or as endpoints in therapeutic trials.  
  
3. 
Emerging therapies that utilize CDT with refined approaches in the individual 
components (i.e. incorporating graded negative pressure) are being utilized in specialized 
lymphedema outpatient clinics including Vanderbilt Lymphedema Clinic, but formal 
efficacy trials have not been performed. We will perform a multiple-measures cross-over 
trial to evaluate the impact of CDT with vs. without graded negative pressure on reducing 
lymph stasis and internal markers of fibrosis in chronic BCRL states, along with a subgroup 
of 1 time treatment  before and after MRI and nonMRI measures for immediate treatment 
impact investigation. Improved lymphatic flow has demonstrated the ability to induce 
lymphangiogenesis in pre-clinical studies8, and it will be crucial to demonstrate internal 
improvements in vivo, relative to basic symptomatology changes which can be subjective 
and difficult to quantify, to underscore the need for expanding healthcare coverage of 
these disease-modifying therapies.  
  
4. 
The imaging methods developed evaluate functional and anatomical lymphatic 
system properties and will be applied in hypothesis-driven studies of BCRL in this work. 
Owing to the known etiology of overt secondary lymphedema such as BCRL, this condition 
represents an ideal system to evaluate relationships between tissue physiology, such as 
adiposity and interstitial protein accumulation, and lymphatic insufficiency. However, the 
findings should have relevance for improving our understanding of the physiological 
changes that occur in lymphatic vessels and dependent tissues in other cases of 
lymphedema or disorders where lymphatic dysfunction is suspected9.    
  
  
2.0   
Rationale and Specific Aims  
  
The goal of this work is to apply novel, noninvasive magnetic resonance imaging (MRI) 
methods for visualizing lymphatic circulation dysfunction to test fundamental hypotheses 
about secondary lymphedema risk factors and therapies. Breast cancer treatment-related 
lymphedema (BCRL) arises secondary to surgical axillary lymph node (LN) dissection and 
irritation, and is a chronic and lifelong condition affecting a high 21.4% of patients 
receiving common breast cancer therapies. Persons at risk for secondary lymphedema 
following axillary lymph node removal also include undergoing treatment for other 
cancers, such as melanoma involving the upper body, necessitating the need for axially 

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lymph node removal. Reducing condition onset and improving management represent 
major unmet clinical needs, and emerging efforts focus on improving quality of life 
through more informed LN dissection and biopsy decisions, optimizing post-surgical 
complex decongestive therapy (CDT), and exploring novel pharmacological and surgical 
procedures. However, fundamental gaps in our knowledge persist regarding optimized 
implementation of these therapies and details of the physiological changes they elicit. 
The major underlying limitation is that there is a shortage of imaging methods available 
that can be used to evaluate lymphatic function directly, and there is currently no 
consensus regarding effective outcome measures for therapeutic efficacy evaluation. 
Rather, LN removal is frequently based on sentinel LN biopsy and additional subjectivity 
of the surgeon. Therapy evaluation is frequently based on coarse measurements such as 
changes in limb volume or patient-reported symptoms, which provide little information 
on underlying mechanistic changes that could be used to further refine these therapies. 
As part of our ongoing INFORM clinical trial of lymphedema progression and therapy, we 
have demonstrated potential for new, noninvasive MRI approaches to identify 
lymphedema risk in sub-clinical disease stages, as well as to visualize internal changes in 
lymphatic functioning as a result of emerging therapies. Here, we propose to extend these 
studies to improve abilities for lymphedema theranostics.   
  
Aim 1: Prevention. We will apply new anatomical and functional LN imaging approaches 
to identify LN profiles specific to biopsy-confirmed metastatic LNs; findings could be used 
to better inform LN removal and reduce the incidence of benign LN dissection. We will 
incorporate healthy female controls to serve as a comparison to the patient population.  
  
Aim 2: Progression. We will improve our understanding of secondary lymphedema risk 
progression following cancer therapy by testing the hypothesis that superficial tissue 
profiles and subcutaneous adipose accumulation are more prevalent in patients 
experiencing lymphedema progression, and thereby could be used to triage patients for 
aggressive therapies prior to overt symptoms and irreversible damage.   
  
Aim 3: Therapy. We will perform a repeated-measures cross-over trial to test the 
hypothesis that mobilization of protein enriched hardened tissue using graded negative 
pressure therapy in conjunction with complete decongestive therapy (CDT) is more 
effective to standard CDT alone for secondary lymphedema management. Both of these 
conservative physical therapy treatments are commonly employed for treatment of 
secondary lymphedema and the Vanderbilt Lymphedema Clinic utilizes the graded 
negative pressure therapy as a component of treatment. Additionally we will perform a 
subgroup of 1-time only hour treatment with before and after MRI and nonMRI 
measurements for additional immediate treatment impact in CDT, CDT with graded 
negative pressure, or exercise and self management treatment strategies alone as 
commonly performed in outpatient physical therapy clinics and at home for the 
management of BCRL.  

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The overarching goal is to develop objective markers of lymphatic dysfunction that can 
be used in emerging therapeutic trials of cancer and lymphedema therapies to be used to 
reduce both the incidence and severity of symptoms associated with this prevalent and 
chronic condition.  
  
Aim (1). To compare in vivo anatomical and novel molecular imaging contrasts 
between LNs with versus without biopsy-confirmed metastasis.  
  
Hypothesis (1). Biochemical LN profiles quantified from noninvasive chemical exchange 
saturation transfer (CEST) MRI provide discriminatory potential for distinguishing 
metastatic versus benign LNs pre-operatively. These biochemical profiles are more 
discriminatory than anatomical measures of LN morphology.   
  
Aim (2). To identify sub-clinical biomarkers of lymphatic insufficiency using 
quantitative evaluation of lymphatic vessel anatomy, axillary edema, and interstitial 
adiposity.   
  
Hypothesis (2). Sub-acute internal imaging markers reflective of tissue microenvironment 
and interstitial adipose accumulation provide superior discriminatory abilities for 
portending BCRL progression compared with established risk factors such as BMI, age, 
and quantity of LNs removed.   
  
Aim (3). To evaluate the impact of CDT with and without graded negative pressure 
therapy on internal measures of lymphatic dysfunction and edema in patients with 
chronic BCRL.   
  
Hypothesis (3). Mobilization of protein-enriched hardened tissue using graded negative 
pressure therapy in conjunction with CDT will reduce (i) superficial markers of interstitial 
protein accumulation and (ii) lymph stasis, and improve (iii) patient-valued functional 
outcomes compared to standard CDT and/or exercise and self management treatment 
strategies.    
  
3.0  
Inclusion/Exclusion Criteria  
  
The following subjects will be excluded from all aims of this study:  
1. Subjects who have any type of non-MRI compatible bioimplant activated by 
mechanical, electronic, or magnetic means (e.g., cochlear implants, pacemakers, 
neurostimulators, biostimulators, electronic infusion pumps, etc.), because such 
devices may be displaced or malfunction.  
2. Subjects who have any type of ferromagnetic bioimplant that could potentially be 
displaced.   
3. Subjects who have cerebral aneurysm clips.  

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4. Subjects who may have shrapnel imbedded in their bodies (such as from war 
wounds), metal workers and machinists (potential for metallic fragments in or 
near the eyes).  
5. Minors (younger than 18 years)  
6. Pregnant women  
7. Subjects who have open wounds on either wrist and / or right ankle because of 
contraindications with placement of electrodes to obtain the L-DEX U400 readings.  
  
Also excluded are subjects incapable of giving informed written consent:  
  
1. Subjects who are non-English speaking, Children/minors* (Form #1117),  Pregnant 
women/fetal tissue/placenta* (Form #1116),  Cognitively impaired* (Form #1118),  
Prisoners* (Form #1115)  
2. Subjects who cannot adhere to the experimental protocols for any reason, or have 
an inability to communicate with the researcher.  
3. Subjects who have limited mental ability to give informed consent, mentally 
retarded, altered mental status, mental disability, confusion, or psychiatric 
disorders.  
4. Prisoners  
To minimize discomfort, we will also exclude subjects who exhibit noticeable anxiety 
and/or claustrophobia or who exhibit severe vertigo when they are moved into the 
magnet.  
  
Inclusion criteria for Aim 1:  
1. Females (over the age of 18 years) planning to have unilateral or bilateral axillary 
lymph node removal surgery as a component of their cancer regime  
2. or Females (over the age of 18 years) without history of cancer or lymphedema to 
serve as study controls  
  
Exclusion criteria for Aims 1:  
1. Any contraindication to MRI.  
2. Any axillary lymph node removal procedure already performed before study visit.  
3. Persons who are deemed clinically unsuitable for an MRI by their treating 
physician  
4. Severe claustrophobia  
5. Minors (younger than 18 years)  
6. To minimize discomfort, we will also exclude subjects who exhibit noticeable 
anxiety and/or claustrophobia or who exhibit severe vertigo when they are moved 
into the magnet. All participants will be screened with the safety screening form 
currently used for research studies on all research MRI scanner  
7. Open wounds on right ankle or bilateral wrists for purposes of obtaining external 
L-Dex extracellular fluid reading.  

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8. Subjects incapable of giving informed written consent:  
a. Subjects who are non-English speaking  
b. Subjects who cannot adhere to the experimental protocols for any reason, 
or have an inability to communicate with the researcher.  
c. Subjects who have limited mental ability to give informed consent, 
mentally retarded, altered mental status, mental disability, confusion, or 
psychiatric disorders.  
9. Prisoners  
  
Inclusion criteria for Aims 2:  
1. Females (over the age of 18 years) who have already had unilateral axillary lymph 
node removal surgery as a component of their cancer regime in the “at risk” side 
being monitored for lymphedema development.  
  
Exclusion criteria for Aims 2:  
1. Any contraindication to MRI.   
2. Persons with signs or symptoms of lymphedema before their first MRI study visit 
for their “at risk side” being monitored for lymphedema development.  
3. Persons who are deemed clinically unsuitable for an MRI by their treating 
physician  
4. Severe claustrophobia  
5. Minors (younger than 18 years)  
6. To minimize discomfort, we will also exclude subjects who exhibit noticeable 
anxiety and/or claustrophobia or who exhibit severe vertigo when they are moved 
into the magnet. All participants will be screened with the safety screening form 
currently used for research studies on all research MRI scanners.  
7. Open wounds on right ankle or bilateral wrists for purposes of obtaining external 
L-Dex extracellular fluid reading.  
8. Also excluded are subjects incapable of giving informed written consent:  
a. Subjects who are non-English speaking  
b. Subjects who cannot adhere to the experimental protocols for any reason, 
or have an inability to communicate with the researcher.  
c. Subjects who have limited mental ability to give informed consent, 
mentally retarded, altered mental status, mental disability, confusion, or 
psychiatric disorders.  
9. Prisoners  
  
Inclusion criteria for Aim 3:  
1. Females (over the age of 18 years) diagnosed with unilateral secondary 
lymphedema in the upper quadrant and/or extremity following cancer related 
surgery involving removal of any axillary lymph nodes and/or radiation therapy.  
  

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Exclusion criteria for Aims 3:  
1. Any contraindication to MRI.  
2. Persons who have undergone bilateral axillary lymph node removal and thus are 
at risk for bilateral secondary lymphedema  
3. Persons who are deemed clinically unsuitable for an MRI or lymphedema therapy 
by their treating physician  
4. Severe claustrophobia  
5. Minors (younger than 18 years)  
6. To minimize discomfort, we will also exclude subjects who exhibit noticeable 
anxiety and/or claustrophobia or who exhibit severe vertigo when they are moved 
into the magnet. All participants will be screened with the safety screening form 
currently used for research studies on all research MRI scanners and the manual 
lymphatic drainage screening form administered by a licensed lymphedema 
physical therapist.  
7. Open wounds on right ankle or bilateral wrists for purposes of obtaining external 
L-Dex extracellular fluid reading.  
8. Also excluded are subjects incapable of giving informed written consent:  
9. Subjects who are non-English speaking  
a. Subjects who cannot adhere to the experimental protocols for any reason, 
or have an inability to communicate with the researcher.  
b. Subjects who have limited mental ability to give informed consent, 
mentally retarded, altered mental status, mental disability, confusion, or 
psychiatric disorders.  
10. Prisoners  
  
  
4.0  
  
Enrollment / Randomization  
  
1. Subjects with a clinical diagnosis of lymphedema following axillary lymph node 
removal including breast cancer related surgery will be made aware of the study 
by recruitment flyers and / or their health care clinician, who will ask the patient 
if they would like for a member of the research team to contact them regarding 
potential participation in the research study.  If the patient would like to receive 
more information about participating in the study and like to be contacted by a 
KSP in the study, then the clinician will provide the patient’s preferred contact 
information to a KSP member.  The patient will then be contacted by the KSP 
member and the risks and benefits of the study will be described.  Importantly, 
the treating clinician will only ask if the patient would like to receive more 
information about the study, but will not be involved in obtaining consent or 
coercing the patient to participate and the patient will understand there is no 
change in their clinician care regardless of participation or not in the study.  
Furthermore, to recruit patients receiving these cancer therapies in local 

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nonVUMC clinics, we will contact the local clinics using the IRB approved letter and 
distribute IRB approved flyers to the clinics. Additional patients will be identified 
using the lymphedema research database where eligible persons request to be on 
the database to be informed when potential studies become available for the KSP 
to contact them regarding participation.   
2. For Aim 1 and 2 of the study tracking patients near the time of their breast cancer 
related surgery, we will recruit patients from the One Hundred Oaks Breast Center 
outpatient clinics seen by the VUMC surgical oncologists and oncologists where 
the patients are seen for their cancer care. Furthermore, to recruit patients 
receiving these cancer surgeries in local non-VUMC cancer centers, we will contact 
the local clinics using the IRB approved letter and distribute IRB approved flyers to 
the clinics.   
3. We will also utilize research databases additional recruitment tools for this 
protocol. ResearchMatch.org will be utilized as a recruitment tool for this protocol.  
ResearchMatch.org is a national electronic, web-based recruitment tool that was 
created through the Clinical & Translational Science Awards Consortium in 2009 
and is maintained at Vanderbilt University as an IRB-approved data repository (see 
IRB #090207). SUBJECT LOCATOR will be used as a recruitment tool to identify 
potential research subjects based on data available in VU clinical systems (e.g. 
STAR Panel, WizOrder, Clinic Scheduling).  The SUBJECT LOCATOR program is part 
of a toolset available through VICTR that enables teams to specify 
inclusion/exclusion criteria for a specific study.  The inclusion/exclusion criteria are 
codified for computable use and combined with data coming through VU Clinical 
Systems to proactively identify individuals who might qualify for a study.  Once a 
'match' is made, research study personnel are alerted using confidential 
messaging or a secure web portal and they may then use the information to 
further review the patient's information using the Vanderbilt Electronic Medical 
Record.  If the patient is further deemed a candidate for the study, the study 
personnel will notify the patient's care provider who will then ask the patient if 
they would be interested in communicating with study personnel about research 
population.  
4. IRB approved flyers/advertisements of the study may also be distributed to 
outpatient clinics and lymphedema support groups/networks as additional means 
of subject recruitment for the aims of the study.  
5. Recruitment of patients will be done through flyers, posted adverts, research 
match, email, departmental research boards, and those who contact KSP directly 
using 
our 
VUMC 
website 
link 
for 
interested 
patient 
volunteers 
(https://ww2.mc.vanderbilt.edu/donahuelab/50860).    
6. Reporting Workbench Reports (RWB) are available/viewable in eStar. 
These reports are developed using real-time data and can be customized 
to meet study-specific inclusion/exclusion criteria. Research team members 
with eStar access and access to the Report Groups can view/run the reports 

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as frequently as needed. While the report is generally limited to certain 
variables, it provides easy access to a patient's record for additional 
screening and confirmation of eligibility.   
  
A custom RWB report will be utilized to identify potential participants. The 
report will filter on study inclusion/exclusion criteria computable in the EHR 
and EMR . Additionally, the report will include OK to Contact status, MHAV 
account status, and ability to receive MHAV messages. The report may 
also display additional information that will facilitate screening.  We will 
utilize these systems for MHAV mass messaging and Individual Direct to 
Patient Messaging for out-reach communication.  
  
  
5.0  
  
 Study procedures  
  
1. Informed consent will be obtained via e-consent initially and then during initial 
visit reviewed in a private setting (e.g., an otherwise empty conference room or 
waiting area) by one of the study Investigators. This initial e-consent will occur as 
far in advance of the exam as possible, preferably before the day of the study and 
a KSP will review any questions with patient remotely as needed prior to onset of 
study. The study participant will be given a copy of the signed consent form at first 
visit.  The subject will be screened for the study inclusion and exclusion criteria 
including MRI contraindications or MLD contraindications or precautions to 
determine eligibility of the study.  
  
2. All subjects will complete an MRI screening form in advance of their scheduled 
visits for any MRI.  The MRI screening form will be cleared by the KSPs and MRI 
technicians prior to scheduling their 1st MRI visit. This MRI screening form will be 
completed for each MRI scan a subject may have during the course of the study.   
  
3. The informed consent andthe MRI screening form will be sent electronically via 
email to the participant, to be completed before their study visit.   
  
4. Persons will undergo external measurements obtaining weight, height, arm 
volumetric measurements (Perometer), skin thickness (SkinFibroMeter), and 
extracellular 
fluid 
levels 
in 
the 
arms 
(InBODY 
770, 
L-Dex 
and 
MoistureMeterD/LymphScanner) 
and 
upper 
body 
(InBODY 
770, 
MoistureMeterD/LymphScanner) and then undergo a non-invasive MRI to 
evaluate lymph fluid properties in the upper extremities. No MR methods used 
require the administration of any exogenous contrast agents, and therefore will 
be completely non-invasive. The visit for non-MRI and MRI measures lasts 

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approximately 90minutes. Patients undergoing 2-year monitoring will also 
undergo these non-MRI measurements at baseline, and every 5-7 months 
(targeting at 6months) up to 5 total visits within 2 years; and for this same group 
an MRI will be done at Visit 1 and then as needed based on symptoms, clinical 
therapies provided increasing risk of lymphedema such as radiation, and non-MRI 
measures at subsequent follow-up visits within the 2 year time period.  For these 
follow-up visits non-involving MRI, the visits last approximately 30 minutes.  
  
  
  
  
  
  
  
 
Perometer  
  
  
  
  
  
  
L-Dex  
                                                                  
  
  
  
  
 
LymphaTouch   
  
  
  
MoistureMeter D/LymphaScanner  
  
  
 
InBODY 770 (Body Composition)  
  
SkinFibroMeter  
   

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5. For subjects enrolled in Aim 3 of this study, they will receive the above mentioned 
non-MRI and MRI measures (#2) at Visits 1, 11, 12 and 22.  In addition the MLD 
screening form will be completed on Visit 1 in preparation for this component of 
their CDT during subsequent treatment visits, as well as completion of the 
Functional Outcome Measures (PSFS and UEFI) patient self-reports will be done 
on Visits 1, 11, 12 and 22 to establish baseline functional level and monitor during 
treatments. Then they will receive their assigned therapy intervention (CDT alone 
or CDT with graded negative pressure) at Visits 2-10 and 13-21.  For Group 1, the 
first course of treatment will be CDT alone, then the second course of treatment 
will be CDT with graded negative pressure.  For Group 2, the first course of 
treatment will be CDT alone, then the second course of treatment will be CDT with 
graded negative pressure.  During all courses of treatment, it will consist of 9 visits 
within 6-8 weeks at 75minutes duration each visit, twice per week for three weeks 
and then once per week for 3-5 weeks.  There will be a 4-8 week washout period 
between Visit 11 and Visit 12.   
6. For the subgroup of subjects enrolled in the 1 time hourly treatment visit of Aim 
3, they will receive the above mentioned MRI and nonMRI measures before and 
immediately following the 1x hourly treatment. The PSFS and UEFI questionnaires 
will not be administered for this 1x treatment visit only.  The participants will 
undergo (i.) CDT alone, (ii.) CDT with graded negative pressure, or (iii.) exercise 
and self management treatment. This subgroup participation may involve 
undergoing all three of the single treatment options on different study dates if the 
participant is interested oterhwise the study only only involves this one-time study 
visit.  
7. Post processing of this MRI data will be overseen by Dr. Manus Donahue and 
performed by Dr. Manus Donahue or a member of the KSP team. The KSPs 
analyzing the data in Aim 3 will be blinded of the subjects’ group assignments until 
after data processing and statistically analysis.   
8. Images from cases showing examples of findings that correlate with changes in 
lymphatic structure and function will be de-identified before inclusion in any 
potential publication or presentation of the results of this pilot study and saved by 
Dr. Manus Donahue.  

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9. Compensation will be as follows:   
a. Aim 1 = $60  
b. Aim 2 = $20 (non-MR measures) / $40 (MRI)  
c. Aim 3  
i. Scan 1 for 1st Treatment = Choice of check for $60 OR 1 Velcro day 
and night-time compression wrap for arm and hand  
ii. If all 9 treatments + exit visit MRI/non-MRI measures complete =  
$60 iii. After 4-8 month 
washout period iv. Scan 1 for 2nd 
Treatment = $60  
v. 
If all 9 treatments + secondary MRI complete = $60  
vi. If completes all parts of the study, will earn additional $50  
vii. Total = $120+$120+$50 = $290 OR Total = 1 Velcro compression 
garment (free) + $60 + 120 + $50 = $230 + 1 Velcro compression 
garment  
d. Subgroup Aim 3 each one-time visit treatment (pre and post MRI and non 
MRI measures = $80  
  
6.0   
 Adverse event reporting  
  
Adverse events (AEs) will be reported according to IRB policies and procedures. Reporting 
will depend on adverse event severity:  
  
Grading of Severity  
0. No AE or within normal limits.  
1. Mild AE.  
2. Moderate AE.  
3. Severe AE resulting in inpatient hospitalization, or a persistent or significant 
disability/incapacity.  
4. Life-threatening or disabling AE.  
5. Fatal AE.  
  
AE’s of grade three or higher will be reported immediately to the IRB using the Report of 
Adverse Events per IRB policies and procedures.  Every six months the PI will summarize 
any AE’s to the IRB.  In conjunction with the IRB, the PI will determine if modifications to 
the protocol are warranted.  
  
 Serious and unexpected adverse events will be reported to our IRB within seven calendar 
days. All our adverse events will be reported as required at the time of continuing review. 
Protocol deviations and violations will be reported to our IRB within ten working days.  
  
  

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7.0   
Study withdrawal/discontinuation  
  
Subjects will be withdrawn from the study if they will not lie still in the scanner during 
data collection. In addition, subjects can voluntarily withdraw from the study at any time.   
  
8.0   
Statistical analysis plan 
  
Statistical analysis will be overseen by statistician co-investigators. Means, standard 
deviations, and ranges for continuous variables will be recorded, along with tests for 
normality and homoscedasticity. The primary statistical objective is to assess whether the 
changes, pre- vs. post-intervention, in internal imaging metrics of lymphatic clearance or 
PSFS score are different for the standard CDT therapy or CDT therapy with negative 
pressure. First, a Student’s t-test or Wilcoxon signed-rank test (depending on normality 
results) will be applied to test the difference between each measurement for the same 
subject for each of the two therapy sessions. We will also test the difference of any potential 
carryover effect and we will apply a general linear model approach, in which we include 
the treatment and treatment-period interaction in the model. Power calculations are based 
on our preliminary data that observed an improvement in PSFS in five patients on CDT 
with negative pressure vs. CDT alone, as well as an ability to detect a mean change in T2 
and lymphangiography score pre- vs. post-MLD of N. With the premise that CDT with 
negative pressure will be at least as good as CDT without negative pressure, with the 
proposed sample size, we will have 80% power to detect a 40% reduction rate with one-
sided type I error = 5%.  
  
9.0   
Privacy/confidentiality issues  
  
The data files generated by the MRI scanner, Perometer and L-Dex software are coded 
using a project name and unique number generated sequentially.  Only the investigators 
will have the key to the subject codes. Image analysis will use the coded files. The safety 
surveys described above include questions about sensitive health issues. This information 
will remain with the signed consent form in a secured area near the scanner. These 
records will remain in the investigators’ possession for ten years following the 
termination of the study and will then be destroyed.  
  
Risks to participants are minimized by limiting this prospective study to subjects meeting 
the inclusion/exclusion criteria. Access to PHI will be limited to the PI and co-investigators. 
All data will be de-identified before publication and/or presentation of the results of this 
study. All PHI used in this study will be kept under lock and key in Dr. Paula Donahue’s or 
Dr. Manus Donahue’s office. The primary risk in this study involves the disclosure of 
protected health information. This risk is reasonable, given the above precautions, in 
relation to the potential benefit to both future patients and society in general by the 
possibility of better understanding and treating lymphedema.  
  

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10.0   Follow-up  
  
No specific follow-up scans are required for Aim 1 unless patients are eligible and would 
like to enroll in the 2-year monitoring aim of the study (Aim2).  For Aim 2, subjects will be 
able to contact the PI and co-investigators with any questions and a KSP will contact them 
to schedule their 6-8 month follow up visits for a total of 4 within the 2 year monitoring 
period.  For Aim 3, subjects will be scheduled for all treatment visits prior to start of each 
course of treatment (Visits 2-10 and 13-21).  The subjects will be able to contact the KSP 
with any questions and needs to change their assigned treatment times and dates should 
any adjustments be necessary. The subjects will be contacted during their washout period 
to schedule their second course of treatment visits and intake and exit measurement 
visits. An unwillingness to participate in any future scans or treatments will not influence 
or disqualify a volunteer from participating in the first instance. For the one-time 
treatment subgroup of Aim 3, this is a one-time visit for the pre and post MRI and nonMRI 
measures with no further study follow-up unless the participant wishes to undergo all 
three separate treatment interventions at different dates for interperson treatment 
comparison. For participants in Aim 2 (monitoring period), if the lymphedema specialist 
recommends lymphedema management, then the subject will be encouraged to seek out 
clinical care which does not prevent them from being able to participate in this study, 
including patients being monitored over 2-years.  The lymphedema specialist will help the 
client obtain orders and location of treatment if subject would like assistance.  
  
11.0   Record retention  
  
Image data will be acquired by the 3 Tesla scanner and will be stored both on digital media 
(which will not leave the secured scanner area) and on the Institute of Imaging Science 
server, which is protected by institutional firewalls and is password protected. 
Investigators may archive these data for their own use, in which case they will be kept in 
the relevant investigator’s locked office. Ten years after the study is concluded, the image 
data will be destroyed.   
  
  
In compliance with the National Institute of Health data sharing initiative, imaging data 
without any personal information attached may be shared with other investigators or 
public data repositories, which provides the research community with open access to 
datasets contributed by labs around the world. Information will be completely 
anonymized with demographics limited to age (accurate to the year up to 90 years old, or 
“90+” for older individuals), gender (male, female), group membership (e.g., 
disease/treatment state) and handedness. Data will be transferred using secure file 
transfer protocols.  
  
 
 

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12.0      References:  
1Crescenzi R, Donahue PMC, Hartley KG, Desai AA, Scott AO, Braxton V, Mahany H, Lants 
SK, Donahue MJ. Lymphedema evaluation using noninvasive 3T MR lymphangiography. 
Journal of magnetic resonance imaging: JMRI. 2017;46(5):1349-60. doi:  
10.1002/jmri.25670. 5573666.  
  
2Donahue PM, Crescenzi R, Scott AO, Braxton V, Desai A, Smith SA, Jordi J, Meszoely IM, 
Grau AM, Kauffmann RM, Sweeting RS, Spotanski K, Ridner SH, Donahue MJ. Bilateral 
Changes in Deep Tissue Environment After Manual Lymphatic Drainage in Patients with 
Breast Cancer Treatment-Related Lymphedema. Lymphatic research and biology. 
2017;15(1):45-56. doi: 10.1089/lrb.2016.0020. 5369395.  
  
3Rane S, Donahue PM, Towse T, Ridner S, Chappell M, Jordi J, Gore J, Donahue MJ. Clinical 
feasibility of noninvasive visualization of lymphatic flow with principles of spin labeling 
MR imaging: implications for lymphedema assessment. Radiology. 2013;269(3):893-902. 
doi: 10.1148/radiol.13120145. 4485559.  
  
4Donahue MJ, Donahue PC, Rane S, Thompson CR, Strother MK, Scott AO, Smith SA. 
Assessment of lymphatic impairment and interstitial protein accumulation in patients 
with breast cancer treatment-related lymphedema using CEST MRI. Magnetic resonance 
in medicine. 2016;75(1):345-55. doi: 10.1002/mrm.25649. 4561605.  
  
5Zhou J, Tryggestad E, Wen Z, Lal B, Zhou T, Grossman R, Wang S, Yan K, Fu DX, Ford E, 
Tyler B, Blakeley J, Laterra J, van Zijl PC. Differentiation between glioma and radiation 
necrosis using molecular magnetic resonance imaging of endogenous proteins and 
peptides. Nature medicine. 2011;17(1):130-4. doi: 10.1038/nm.2268. 3058561.  
  
6Klomp DW, Dula AN, Arlinghaus LR, Italiaander M, Dortch RD, Zu Z, Williams JM, 
Gochberg DF, Luijten PR, Gore JC, Yankeelov TE, Smith SA. Amide proton transfer imaging 
of the human breast at 7T: development and reproducibility. NMR in biomedicine. 
2013;26(10):1271-7. doi: 10.1002/nbm.2947. 3726578.  
  
7van Zijl PCM, Lam WW, Xu J, Knutsson L, Stanisz GJ. Magnetization Transfer Contrast and 
Chemical Exchange Saturation Transfer MRI. Features and analysis of the field-dependent 
saturation spectrum. NeuroImage. 2017. doi: 10.1016/j.neuroimage.2017.04.045. 
5650949.  
  
8Boardman KC, Swartz MA. Interstitial flow as a guide for lymphangiogenesis. Circ Res.  
2003;92(7):801-8. Epub 2003/03/08. doi: 10.1161/01.RES.0000065621.69843.49.  
   

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9Mortimer PS, Rockson SG. New developments in clinical aspects of lymphatic disease. J 
Clin Invest. 2014;124(3):915-21. doi: 10.1172/JCI71608. PMC3938261.