The Exercise And Colorectal Cancer Treatment (EXACT) Trial
Phase: NA
Conditions: Colon Cancer, Rectal Cancer
Sponsor: Pennington Biomedical Research Center

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Protocol Title: The Exercise And Colorectal Cancer Treatment (EXACT) Trial
PI Name: Justin C. Brown, PhD
Sub-Investigator’s Name: Anne Gilmore, PhD; Lauren Maniscalco, MPH
Medical Investigator: Frank Greenway, MD
Study-site contacts: Shengping Yang, PhD; Grace Bella; Alison Carville; Melissa Harris; Phillip 
Nauta; Elizabeth Zunica
Protocol Version Date: March 27, 2024 (Version 13)
Objectives
This randomized trial will examine the biological effects of 12 weeks of moderate-intensity 
aerobic exercise versus wait-list control in 60 subjects who have completed standard medical 
therapy for stage I- III colorectal cancer. The primary objective is to determine if aerobic 
exercise can reduce systemic inflammation, quantified using plasma concentrations of high-
sensitivity C-reactive protein and interleukin-6. The secondary objectives are to determine if 
exercise can reduce 1) insulin resistance quantified using an oral glucose tolerance test and 2) 
circulating tumor cells quantified using a microfluidic antibody-mediated capture platform. The 
exploratory objective is to determine if exercise can improve mitochondrial respiration rates 
and fatty acid oxidation from peripheral blood mononuclear cells, and tumor fraction.
Background
Colorectal Cancer Prognosis. Each year, 145,600 people are diagnosed with colorectal cancer in 
the United States.1 Despite efficacious surgical and chemotherapeutic interventions, 25–30% of 
patients will experience recurrent and metastatic disease within three years of diagnosis, and 
91% of those who recur within three years die by five years.2 Additional therapies are critically 
needed to maximize the likelihood of cure for these patients. One such additional therapy may 
be the prescription of physical activity.3
Physical Activity and Colorectal Cancer. Physical activity or exercise is a modifiable lifestyle 
behavior that is associated with disease outcomes among colorectal cancer patients. A meta-
analysis of six prospective cohort studies that included 7,523 colorectal cancer patients 
demonstrated that participation in physical activity after cancer diagnosis was associated with a 
39% lower risk of colorectal cancer-specific mortality [Hazard Ratio (HR): 0.61, 95% CI: 
0.40−0.92] and 42% lower risk of all-cause mortality [HR: 0.58, 95% CI: 0.48−0.70].4 The 
relationship between physical activity and mortality is independent of various demographic, 
clinical, pathologic, treatment, and behavioral factors (e.g., smoking and obesity). Colorectal 
cancer patients who derive the largest survival benefits from physical activity are those who 
were sedentary before diagnosis and adopt a physically active lifestyle after diagnosis.5 This 
observation substantiates the hypothesis that the prescription of physical activity after 
 
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colorectal cancer diagnosis may decrease disease-specific morbidity and mortality. However, 
the biological pathways through which physical activity may influence cancer recurrence and 
mortality are not yet completely understood.6
Inflammation and Colorectal Cancer. One biological pathway hypothesized to mediate the 
relationship between physical activity and cancer recurrence and mortality in colorectal cancer 
is systemic inflammation. Systemic inflammation, such as elevated concentrations of C-reactive 
protein (CRP),7 interleukin-6 (IL-6),8 and soluble tumor necrosis factor-alpha receptor 2 (sTNF-
αR2),9 is associated with a higher risk of cancer recurrence and mortality among colorectal 
cancer patients. Systemic inflammation induces signaling pathways, including PI3K-Akt-mTOR, 
JAK-STAT, and NF-κB, resulting in cell growth, migration and invasion, anti-apoptosis, and 
angiogenesis.10-12 Elevated IL-6 and TNF-α induce the hepatic expression of CRP, producing a 
state of chronic low-grade systemic inflammation.13 CRP is a robust overall measure of systemic 
inflammation given its strong correlation with IL-6 and sTNF-αR2.14,15 The observations derived 
from cohort studies are supported by preclinical data that demonstrate experimentally 
manipulating inflammatory-related pathways, including IL-616,17 and TNF-α/sTNF-αR2,18-22 
regulate colorectal cancer growth and progression. Collectively, these data suggest that 
systemic inflammation is an important mechanism associated with a higher risk of cancer 
recurrence and mortality in colorectal cancer.
Insulin Resistance and Colorectal Cancer. Another biological pathway hypothesized to mediate 
the relationship between physical activity and cancer recurrence and mortality in colorectal 
cancer is insulin resistance. The hallmarks of insulin resistance are high insulin concentrations 
and an exaggerated insulin response to glucose.23 Colorectal cancer patients have fasting insulin 
concentrations 58% higher than age- and sex-matched controls (P<0.001).24 Hyperinsulinemia is 
often a symptom among people diagnosed with type 2 diabetes. Patients with colorectal cancer 
who also have type 2 diabetes have a worse prognosis than those without type 2 diabetes.25,26 
Markers of insulin secretion, such as C-peptide, predict mortality in colorectal cancer 
patients.27,28 In vitro studies demonstrate that insulin increases cell resistance to chemotherapy 
agents used to treat colorectal cancer, including 5-fluorouracil29 and oxaliplatin,30,31 and 
preclinical models demonstrate that exposure to insulin promotes colorectal tumor 
multiplicity.32 Collectively, these data suggest that insulin resistance is an important mechanism 
associated with a higher risk of cancer recurrence and mortality in colorectal cancer.
Circulating Tumor Cells in Colorectal Cancer. The development of metastases from colorectal 
cancer are hypothesized to result from tumor cells entering the circulation, migrating to distant 
organs, extravasating, multiplying, and eventually manifesting as clinically detectable lesions. 
This process marks the transition from localized, potentially curable to disseminated, usually 
incurable disease.33 Autopsy studies suggest 66% of metastases in colorectal cancer are 
explained by vascular blood flow alone.34 The early detection and characterization of circulating 
tumor cells (CTCs) are important to monitor and prevent the development of metastases.35 
CTCs predict disease recurrence and mortality among patients with stage I-III colorectal 
cancer.36-39 For example, among 438 patients with stage I-III colorectal cancer, 31% had tumor 
cells in peripheral blood after completing treatment, and the presence of CTCs was associated 
with a 29-fold increase in the risk of cancer recurrence (P<0.001).40 However, few therapeutic 
 
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interventions exist to reduce CTCs and improve clinical outcomes.41 We hypothesize that cancer 
cells disseminate via the circulation during the earliest stages of recurrent metastatic growth, 
mirroring what occurs during the primary tumor setting. If this occurs and exercise interrupts 
this process, we are uniquely situated to detect this phenomenon by quantifying CTCs. 
Collectively, these data suggest that CTCs are a key constituent of cancer recurrence and 
mortality in colorectal cancer.
Circulating Cell-Free DNA (Tumor Fraction). Circulating cell-free DNA allows for detailed 
genetic characterization of residual tumor burden.42 Cell-free DNA is released into the 
circulation through apoptosis and necrosis. Cell-free DNA are generally double-stranded 
fragments of 150 to 200 base pairs in length.42 Cell-free DNA that is released from tumor cells is 
classified as circulating tumor DNA. Estimates of the tumor’s contribution to cell-free DNA, 
defined as the tumor fraction, offer greater sensitivity compared to that of CTCs alone, and the 
combination of both CTCs and cell-free DNA provides complementary biological insight.43 It is 
estimated that 70% of patients with stage I to III colorectal cancer have measurable cell-free 
DNA concentrations with cancer cell origin (i.e., a tumor fraction 0.1).43,44 Tumor fraction is a 
pharmacodynamic biomarker that correlates with response to chemotherapy,45 and is 
prognostic of disease recurrence and mortality in stage I to III colorectal cancer.46 For example, 
in patients with stage II colorectal cancer, a higher tumor fraction after surgical resection is 
associated with an 11-fold to 18-fold higher risk of systemic recurrence.47 The advantages of 
cell-free DNA include a high sensitivity to accurately quantify tumor burden in patients with 
curatively resected cancer (e.g., high signal-to-noise ratio), the capacity to characterize intra-
tumor heterogeneity, and the ability to predict future treatment resistance.44 However, the 
effect of exercise on circulating cell-free DNA in cancer patients has not been evaluated.
Preliminary Data. The pilot data to justify the current trial was a dose-response study that 
randomized 39 subjects with stage I-III colon cancer to 150 min/wk or 300 min/wk of aerobic 
exercise or a usual-care control group for 24 weeks.48 We identified an interaction between 
cancer stage (a randomization stratification factor in the pilot study) and randomized group for 
changes in high-sensitivity [hs]-CRP (Pinteraction=0.051) and IL-6 (Pinteraction=0.002). Compared to 
baseline, 150 min/wk exercise reduced hs-CRP (−0.79±0.38 mg/L; P=0.038) and IL-6 (−0.57±0.29 
pg/mL; P=0.047) in subjects with stage III but not in subjects with stage I/II disease. Compared 
to baseline, 300 min/wk of aerobic exercise did not reduce hs-CRP or IL-6. Additional analyses 
demonstrated that compared to baseline, 150 min/wk of aerobic exercise reduced fasting 
plasma insulin (−28.0±8.4 pmol/L; P=0.001) and glucose (−0.39±0.1 mmol/L; P=0.011),49 and 
CTCs (−1.3±0.3 cells/mL; P<0.001).50 However, due to the multiple randomized groups of this 
pilot trial, the sample size within each group was small (n=12−14), making between-group 
comparisons statistically underpowered. The results obtained from this pilot study inform the 
primary and secondary aims for the proposed trial, as detailed below.
Experimental Design & Methods
A. Number of Subjects
 
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Accrual is 60 subjects (see power and sample size section below), recruited over 24 
months, with a monthly accrual goal of approximately 23 subjects per month. In our 
prior trial, we successfully accrued 5.5 subjects per month.1 Using estimates from our 
prior trial, we predict we will need to screen approximately 160 potential subjects to 
successfully randomize 60. We aim to recruit an equal ratio of males and females and 
≥25% non-white minority subjects. 
B. Inclusion and Exclusion Criteria
Many oncology clinical trials do not reach their minimum projected accrual goal.51 To 
ensure trial completion, we implement prospectively specified stepped inclusion and 
exclusion criteria that enable the efficient recruitment of the most homogeneous study 
population that is feasible within the allowable time horizon without compromising 
subject safety or scientific validity. 
We will begin with three months of strict eligibility criteria (Phase 1). At the end of three 
months, if six subjects are not accrued, eligibility criteria will be expanded for another 
three-month period (Phase 2). If, at the end of six months, we have still not accrued an 
average of two subjects per month, we will further expand eligibility criteria (Phase 3). 
We successfully implemented these prospectively specified stepped inclusion and 
exclusion criteria in the pilot study.48
Phase 1 (most strict) inclusion criteria: 
1.1
Age ≥18 years
1.2
Histologically-confirmed stage III colorectal cancer 
a. Self-report during screening is acceptable if confirmed with pathology 
report, state tumor registry file, or physician documentation prior to 
randomization
1.3
Completed surgical resection within 124 months or completed chemotherapy 
(if applicable) within 1-24 months
1.4
Provide written approval by a physician or other qualified healthcare provider 
1.5
No planned major surgery during the study period (including colostomy 
reversal; however, chemotherapy infusion port removal is permitted) 
1.6
Readiness to exercise [as determined by a modified version of the Physical 
Activity Readiness Questionnaire (PAR-Q)]
a. If there are any indications that home-based exercise would be unsafe 
based on the PAR-Q, the patient will not be enrolled until approval from 
the patient’s provider (e.g., oncologist, cardiologist) confirms that it is 
safe to exercise
1.7
Allow the collection and storage of specimens and data for future use
1.8
Willing to be randomized
Phase 2 (less restrictive) inclusion criteria will expand:
2.1
Histologically-confirmed stage to include stage I, II, or III colorectal cancer
 
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a. Self-report during screening is acceptable if confirmed with tumor 
registry, pathology report, or physician documentation prior to 
randomization
2.2
Time since completing surgical resection to 136 months or time since 
completing chemotherapy (if applicable) to 1-36 months
Phase 3 (least restrictive) inclusion criteria will expand:
3.1
Time since completing surgical resection 1 month or time since completing 
chemotherapy (if applicable) 1 month
3.2
Histologically confirmed stage I, II, or III colorectal cancer 1 month since    
completing surgical resection or 1 month since completing chemotherapy 
and/or radiotherapy 
Exclusion criteria for all three accrual phases of recruitment include: 
1.1
Evidence of metastatic colorectal cancer (biopsy-proven or radiologic evidence 
that results in the initiation of local or systemic therapy) 
a. Elevated tumor markers (e.g., CEA 5 ng/mL), in the absence of an 
identified lesion(s) on imaging, that is monitored with surveillance only is 
not considered evidence of metastatic disease
b. Lesion(s) classified on imaging as “suspicious” that are monitored with 
surveillance only are not considered evidence of metastatic disease
1.2
Concurrently actively treated other cancers (except non-melanoma skin cancer 
or in situ cancers)
1.3
Currently enrolled in another clinical trial of weight loss, physical activity, or 
dietary intervention
1.4
Current body mass 181 kg
1.5
Unable to provide a baseline fasting blood sample 
1.6
Unable or unwilling to give informed consent 
1.7
Unable or unwilling to be randomized
1.8
Or any other condition that may impede the testing of the study hypothesis or 
make it unsafe to engage in the exercise program (as determined by the 
investigative team)
C. Recruitment Methods
Potentially eligible study subjects may be identified and contacted using the Louisiana 
Tumor Registry and the Gulf South Community Oncology Research Program. 
The Louisiana Tumor Registry is part of the Surveillance, Epidemiology and End Results 
(SEER) Program, funded by the National Cancer Institute. The registry collects and 
reports high-quality and timely population-based cancer data across the state to 
support research activities in cancer prevention and control. The Louisiana Tumor 
Registry Director is Xiaochen Wu, MD, MPH, who is a Professor of Epidemiology at the 
School of Public Health in the Louisiana Health Sciences Center. Potentially eligible 
subjects will be identified in the Louisiana Tumor Registry (e.g., using tumor codes 
 
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affiliated with the diagnosis of colorectal cancer) and contacted using mail and 
telephone.
The Gulf South Minority Underserved Community Oncology Research Program is part of 
the National Cancer Institute-funded Community Oncology Research Program (NCORP). 
The Gulf South NCORP is a state-wide initiative that brings together an integrated 
clinical trials program in the state of Louisiana to promote and facilitate participation 
and enrollment of cancer patients into clinical trials. The Gulf South Minority 
Underserved NCORP Director is Augusto Ochoa, MD, who is the Director of the Stanley 
S. Scott Cancer Center at the Louisiana State University Health Sciences Center. The Gulf 
South Minority Underserved NCORP currently includes 25 oncology clinics throughout 
the state. Subjects will be recruited from these clinics using flyers placed in patient 
waiting areas, mailed postal letters, direct physician referrals, or other methods deemed 
appropriate.
Potential subjects will be directed to complete a web screening form on the PBRC 
landing page, call the PBRC Recruitment Core directly, or e-mail the Recruitment Core. 
Potential subjects will then undergo a phone screen to answer a series of yes or no 
questions to determine eligibility. Individuals meeting the criteria from the phone 
screen will be scheduled for a screening visit at PBRC.
Other approved recruitment materials may include a landing page, listserv, flyers, and 
social media developed specifically for the study. Our community outreach efforts will 
include but are not limited to, presentations and community events. We will also utilize 
media outlets, including newspapers, radio, and social media. 
The Principal Investigator and study staff will also notify members of local cancer 
support groups, and additional cancer services organizations may be utilized.
D. Study Timelines
Subjects are expected to undergo screening, baseline, and randomization visits at 
Pennington Biomedical Research Center and participate in a 12-week, home-based 
exercise intervention or wait-list control. A final clinic visit after the completion of the 
12-week intervention period will conclude study participation. The duration of an 
individual subject’s participation will be approximately five months. The study will be 
conducted over a three-year period.
E. Study Procedures 
Table 1. Schedule of Procedures
Procedure
Screening
(SV1)
Baseline
(BL)a,b
12 Week 
Intervention
(W1-W12)
Month 3
(M3)a,c
Review Phone Screen 
X
 
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Consent
X
History & Physical
X
ECG
X
Vital Signs
X
X
X
Height
X
Weight
X
X
X
Waist/Hip Circumference
X
X
Questionnaires
Xh
Xh
Concomitant Meds
X
X
X
Accelerometer
Xd
Xe
Lifestyle Consultation
X
Adverse Events
X
X
X
Urine Pregnancy Test
Xf
Xf
DXA
X
X
IV OGTT
X
X
Lab Work
X
X
Fitness Assessment
X
X
Randomization
Xg
Exercise Instruction
X
Weekly Telephone Calls
X
Exercise Program/
Usual Care
X
aBaseline and Month 3 visits can each be split into two days of measures if requested by the 
participant.
bBL is to be completed within 30 days of SV1. 
cM3 is to be completed within 30 days W12.
dAccelerometer is provided to the participant at SV1 and is to be returned at the BL visit.
eAccelerometer is mailed to the participant at W10 and is to be returned at the M3 visit.
fFemales of child-bearing potential only.
gRandomization can occur up to 7 days after BL. 
hQuestionnaires will be given to participant at BL and M3 to complete.
1. Visit Description
Screening Visit (SV1): As part of the screening visit, each subject will review their 
answers provided during the phone screen and the informed consent with a 
member of the clinical trials unit staff. Once the consent is signed, the following will 
be completed: 
•
Medical History and Physical;
•
Resting 12 lead ECG;
•
Height, weight, and vital signs;
•
Concomitant Meds;
•
Lifestyle consultation to assess subject readiness to initiate a physical activity 
program and assess barriers to participation;
 
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•
An accelerometer with instructions will be given to the participant and 
returned at the baseline visit.
•
Clinic staff will send a fax to the subject’s medical providers that describes 
the study, reviews the subject’s answers to the PAR-Q, and requests that the 
medical provider approve participation in the study.
Baseline Visit (BL): At the baseline visit, the participant will report to the inpatient 
unit after fasting for approximately 10 hours (minimum of 8 hours). The following 
will be completed:
•
Weight and circumferences of the hip and waist 
•
Vital signs
•
Concomitant medications
•
Collection of accelerometer
•
Females of child-bearing potential will complete a urine pregnancy test
•
DXA*
•
Oral glucose tolerance test (OGTT)**
•
Submaximal fitness assessment
•
Administer and review questionnaires  
*After completing the DXA, the participant will be provided a snack prior to the 
fitness assessment.
**OGTT will take blood samples at 0, 60, and 120 minutes. 
Randomization: After the baseline visit, all collected data will be reviewed for 
completeness and accuracy. Once all baseline data is confirmed to be complete, 
accurate, and the participant is deemed eligible, he/she will be randomized into one 
of the two study groups. The interventional resources staff will call the subject and 
explain their randomly assigned group assignment and expectations. At that time, 
participants randomized into the intervention group will be shipped a heart rate 
monitor with instructions for use. A treadmill will also be shipped to the subject’s 
home. Instructions on the exercise program will be provided in brief along with a 
plan for future contact. Upon confirmation of delivery and setup of the treadmill and 
receipt of the heart rate monitor, the interventional resources staff will re-contact 
the participant and review the intervention in greater detail. 
Intervention. Participants will be randomized into 1 of 2 intervention groups: 
Exercise Intervention Group or Usual Care Control Group.
a. Exercise Intervention. The exercise intervention will consist of moderate-
intensity (50−70% age-predicted maximum heart rate) treadmill walking. This 
exercise protocol balances participant safety with behavioral feasibility 
towards the goal of accumulating 150 min/wk of moderate-intensity aerobic 
exercise. Participants with residual lower-extremity neuropathy from 
chemotherapy may elect to receive an elliptical or cycle rather than a 
treadmill. After treadmill delivery (or other equipment) and set up, 
participants will speak on the phone with an exercise specialist to introduce 
 
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the exercise prescription and familiarize the participant with the use of the 
treadmill, the completion of exercise logs, the use of a heart rate monitor 
(described below), appropriate warm-up and cool-down, stretches, and 
proper footwear for aerobic exercise. In subsequent weeks, participants will 
be permitted to complete supervised exercise sessions at Pennington 
Biomedical’s Fitness Center if they choose to do so. All other exercises will be 
completed at home with the study-provided treadmill (or other equipment of 
preferred exercise modality). All exercise sessions will begin with a five-
minute warm-up of slow walking, 30-60 minutes of moderate-intensity 
walking, and a five-minute cool-down of slow walking. In our pilot study, the 
study-provided treadmill was used for 77% of exercise sessions; most of the 
remaining exercise included outdoor walking (19%). No serious adverse 
events occurred.
Monitoring Exercise Adherence. Each participant will be provided a Polar 
heart rate monitor (M430) for the duration of the study. The M430 heart 
rate monitor can record up to 12 weeks of exercise using a one-minute 
epoch with a failure rate of <3%. The exercise specialist will download 
and review the heart rate data to objectively monitor exercise adherence 
(frequency, duration, and intensity) approximately bi-weekly. The 
exercise specialist will also inquire about any adverse events or barriers 
to completing exercise via weekly phone calls. The compliance 
information gathered from the heart rate monitors will be reviewed with 
the participant for positive reinforcement and to develop and tailor the 
desired dose of exercise for the following week. Participants who 
demonstrate poor adherence to the study protocol will be reminded of 
the study objectives, discuss any barriers or obstacles to completing the 
exercise, and be provided with encouragement to promote exercise self-
efficacy, participation, and compliance. 
b. Usual Care Control Group. Participants randomized into the wait-list control 
group are asked to maintain their pre-study levels of physical activity and 
follow the recommendations provided by their physician. The study 
interventionist will contact control group participants each week to maintain 
ongoing communication about the study and inquire about any adverse 
events. After completing three-month measures, control group participants 
are provided with an in-home treadmill and individualized exercise program 
similar to that prescribed to the exercise group. 
Month 3 Visit: The Month 3 Visit is a repeat of the Baseline Visit. At week 10 of the 
study, an accelerometer will be mailed to the participant with instructions for use. 
The participant will report to the inpatient unit after fasting for approximately 10 
hours (minimum of 8 hours). The following will be completed: 
•
Weight and circumferences of the hip and waist 
•
Vital signs
 
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•
Concomitant medications
•
Collection of accelerometer
•
Females of child-bearing potential will complete a urine pregnancy test   
•
DXA*
•
Oral glucose tolerance test (OGTT)**
•
Submaximal fitness assessment
•
Administer and review questionnaires 
*After completing the DXA, the participant will be provided a snack prior to the 
fitness assessment. 
**OGTT will take blood samples at 0, 60, and 120 minutes.
Upon completion of delivery and acceptance by the participant, the treadmill will be the 
participant’s responsibility and will no longer be the responsibility of Pennington Biomedical 
Research Center or any of the researchers. 
2. Procedure Descriptions
•
Laboratory Collection. At baseline and three months, all participants will 
undergo a blood draw. Blood will be performed after fasting for 
approximately 10 hours (minimum of 8 hours), abstinence from alcohol 
consumption, and any structured exercise for 24 hours. 70 mL of blood will 
be collected, centrifuged, and aliquoted into 1.8 mL cryovials at 0.5 mL/vial. 
Whole blood, plasma, serum, and buffy coat will be collected. Blood samples 
will be stored at −80°C. Females of child-bearing potential will complete a 
urine pregnancy test.
Plasma Inflammatory Marker Concentrations. 
•
The concentration of high-sensitivity CRP will be determined using an 
immunoturbidimetric assay (Roche Diagnostics, Indianapolis, IN). This high-
sensitivity assay has a limit of detection of 0.03 mg/L. The day-to-day 
variability of the assay at concentrations of 0.91, 1.60, and 18.40 mg/L are 
3.8, 3.3 and 1.9%, respectively.
•
IL-6 is measured by an ultra-sensitive quantitative sandwich enzyme 
immunoassay (R&D Systems, Minneapolis, MN). The assay has a sensitivity of 
0.094 pg/mL, and the day-to-day variability of the assay at concentrations of 
0.49, 2.78, and 5.65 pg/mL are 9.6, 7.2, and 6.5%, respectively. 
•
sTNFα-R2 is measured by a quantitative sandwich enzyme immunoassay 
assay (R&D Systems). The assay has a sensitivity of 0.6 pg/mL. The day-to-day 
variability of the assay at concentrations of 89.9, 197, and 444 pg/mL are 5.1, 
3.5, and 3.6%, respectively. Masked quality-control samples are interspersed 
among cases, and all lab personnel are blinded to patient outcomes.
 
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Insulin Resistance. An assessment of insulin resistance will be performed using a 75-
gram oral glucose tolerance test. Plasma glucose and insulin concentrations will be 
determined at pre-specified intervals to calculate the incremental area under the 
curve during the oral glucose tolerance test. 
Circulating Tumor Cells (CTCs). CTCs from participant venous blood will be isolated 
using a geometrically enhanced differential immunocapture (GEDI) platform. GEDI is 
a microfluidic platform that utilizes antibody-coated obstacles to capture rare cells 
within the blood. This allows for anticoagulated whole blood to be applied to the 
“chip” the size of a microscope slide in the laboratory using standard syringe pumps. 
To capture CTCs, obstacles will be coated with an antibody specific to epithelial cell-
adhesion molecule (EpCAM), an epithelial cell-specific marker. After washes, 
captured cells will be stained with the nuclear marker DAPI and fluorescently labeled 
antibodies to the leukocyte marker CD45 and the epithelial cell marker Pdx-1. Using 
fluorescence microscopy, Pdx1+/DAPI+/CD45- EpCAM captured cells with intact 
cellular morphology will be counted as CTCs by a blinded technician. Cells will be 
stored at -80°C for additional analyses in subsequent grant submissions.
Circulating Cell-Free DNA (Tumor Fraction). The Broad Institute of MIT and Harvard 
has developed a comprehensive, versatile, and scalable platform to measure 
circulating cell-free DNA.52 Venous blood samples will be drawn into 10 mL cell-free 
DNA blood collection tubes (Streck, Inc., La Vista, NE). Blood will be processed into 
component plasma, buffy coat, and erythrocytes within four hours of collection 
through standard density gradient centrifugation. Plasma samples will be subjected 
to additional high-speed centrifugation and frozen at –80°C until processing.
Mitochondrial Function and Fatty Acid Oxidation. The Buffy coat will be isolated 
within 24 hours of sample collection and stored at room temperature. The Buffy 
coat will be diluted with an equal amount of PBS, layered over a density gradient 
medium, and centrifuged according to the manufacturer’s guide. Mononuclear cells 
will be washed with PBS and centrifuged, and any remaining red blood cells will be 
lysed and separated via centrifugation, followed by a final PBS wash and 
centrifugation. The mononuclear cell pellet will be prepared for oxidative 
phosphorylation by suspending it in a mitochondria respiration medium (MiR05), 
and the number of live mononuclear cells will be quantified. The integrated 
mitochondrial function will be assessed using a high-resolution respirometer, 
Oxygraph-2 K using DatLab 4 software (Oroboros Instruments, Innsbruck, Austria) to 
record the oxygen consumption rate for the transport of substrates into the 
mitochondria, the generation of reducing equivalents by specific dehydrogenases, 
entry into ETC (electron transport chain), and coupling to ATP synthesis.53 Using two 
established protocols, the respiration rates of complex I, II, III, and IV as well as fatty 
acid oxidation will be quantified.54
Fitness Assessment. An assessment of sub-maximal fitness will be performed at 
baseline and three months with a treadmill using the modified Bruce Protocol.9 The 
 
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speed and elevation will slowly increase until 80% of the age-predicted maximal 
heart rate (or RPE of 18 for patients on beta-blocker therapy), adhering to guidelines 
by the American College of Sports Medicine’s Guidelines for Exercise Testing and 
Prescription.    
Body Composition. Body mass, height, weight, waist, and hip circumferences will be 
assessed using a digital scale, scale-mounted stadiometer, and Gulick tape measure, 
respectively. DXA (Hologic) will be used to quantify changes in bone mineral density 
and body composition. 
Accelerometry. Participants will be asked to wear an Actigraph GT3X+ triaxial 
accelerometer for one week before and after the study to objectively assess activity 
levels.
Clinical Characteristics. Clinical characteristics, including tumor pathology, surgical 
history, and chemotherapy treatment plan, will be obtained from the clinical and 
cancer registry data files prior to randomization.
Patient Outcomes. Participants will complete questionnaires relating to 
demographics, alcohol and smoking habits, current medicine, and supplement 
usage. Participants will complete a patient-reported version of the Common 
Terminology Criteria for Adverse Events.
Data and Specimen Banking
The participant will be asked to allow blood specimens and data to be stored and used for 
research at a later time. Participants who refuse to have this information kept for future 
research will be excluded from the study. The specimens and data will be stored indefinitely. 
Future research may take place at Pennington Biomedical and may involve Pennington 
Biomedical researchers in this study. The future research may not take place at Pennington 
Biomedical Research Center and may not be reviewed by Pennington Biomedical Research 
Center’s Institutional Review Board. 
For privacy and confidentiality, all specimens and scans will be labeled with a unique series of 
letters and numbers. Pennington Biomedical will store the information with this unique 
identifier. The research done with data from these specimens and scans may help to develop 
new products in the future or may be used to establish a diagnostic test that could be patented 
or licensed. The participant will not receive any financial compensation for any patents, 
inventions, or licenses developed from this research.
The de-identified data will be stored indefinitely on the network shares at PBRC. Hard copy 
records for each subject will be kept in the participant’s chart located in the Medical Records
Department. These data will be used in publications but not shared outside of the PI’s.
 
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Long-term storage of specimens will be at -80 C. The samples will be stored in a designated 
freezer housed at Pennington Biomedical Research Center. The Principal Investigator, unless 
otherwise noted, will be the person who has access to the samples. Each sample will be labeled 
with the subject’s ID and date.
Power analysis
After completion of baseline measurements, participants will be stratified by sex (male vs 
female) and then randomized in a 1:1 ratio to either the exercise or control group using a 
computer-generated algorithm. The co-primary outcomes for this study are hs-CRP and IL-6. For 
the CRP outcome, assuming a baseline concentration of 1.87 mg/L with a standard deviation of 
1.32 mg/L, correlation of 0.8 between time points, a reduction of −0.71 mg/L in the exercise 
group over 12 weeks, and a type-I error rate of 5%, 46 participants total will provide 80% 
statistical power to detect between-group differences. For the IL-6 outcome, assuming a 
baseline concentration of 1.7 pg/mL with a standard deviation of 0.7 pg/mL, a correlation of 
0.58 between time points, a reduction of −0.57 pg/mL in the exercise group over 12 weeks, and 
a type-I error rate of 5%, 48 participants will provide 80% statistical power to detect between-
group differences. sTNF-αR2 is a secondary inflammatory measure. In the pilot study, we 
observed a 3% dropout rate (defined as participants not providing endpoint data), an 8% 
crossover rate (defined as control group participants who disclose initiating a structured, 
progressive exercise program during the trial), and 8% recurrence rate (defined as the clinical 
discovery of locoregional or distant metastatic disease).70 We will inflate these estimates to 
accommodate a higher-than-anticipated dropout, experience a recurrence, and/or crossover. 
Enrolling 60 study participants provides a robust estimate of the largest necessary sample size 
needed for this trial. This sample size provides adequate statistical power to detect moderate 
between-group effect size differences (e.g., Cohen’s d of ≥0.3) for measures of insulin 
resistance (Aim 2) and CTCs (Aim 3).
Statistical Analysis
The primary hypothesis in this trial is that exercise will significantly lower plasma inflammation 
concentrations over 12 weeks compared to control. All inferential analyses will be conducted 
on an intention-to-treat basis. Plasma inflammatory concentrations will be log-transformed in 
the inferential analysis to improve normality. Changes in outcomes will be evaluated from 
baseline to follow-up in the two groups using a repeated-measures mixed-effects regression 
model that accounts for the correlation between repeated measures and is robust to missing 
data. The baseline value of the dependent variable and sex (randomization stratification factor) 
will be included as a covariate in the regression models.55 Group-by-time interaction terms will 
be included as fixed effects in the regression model. Model fit will be assessed using standard 
methods. A closed testing procedure for the two study outcomes (hs-CRP and IL-6) will be used, 
which is hierarchically ordered to test CRP and then IL-6 to maintain the overall experiment-
wise error rate at 5%.56 Other inflammatory outcomes and measures of insulin resistance (Aim 
2) and CTCs (Aim 3) will be analyzed using the same statistical techniques and reported at the 
nominal type I error rate of 5%.
 
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Data and Specimen Management
Study participants will be assigned unique subject identification (ID) numbers. Study subject ID 
numbers will be used on all data collection instruments, including questionnaires, data 
collection forms, biological specimen tubes, and computer records. A master list linking the 
participants’ names and ID numbers will be kept in a password-protected computer file with 
access restricted to the PI and co-PIs. Biological samples that are moved off-site for analysis will 
not contain any personally identifiable information and will be labeled with only the unique 
subject ID numbers. Staff at these sites will not have access to the master list at any time.
Data collection forms will be kept under lock and key or password-protected if computerized 
and under the control of the PI, co-PIs, and medical investigator. Only personnel assigned to the 
research study by the PI will have access to the data. Hard-copy data records will be stored for a 
minimum of three years after study completion.
The PBRC has a fully integrated, campus-wide, automated data management system, REDCap. 
REDCap, a web-based application used to build and manage surveys and databases, will be used 
for questionnaire collection in this study. REDCap (Research Electronic Data Capture) is a 
secure, web-based application that is flexible enough to be used for a variety of types of 
research. REDCap provides an intuitive user interface that streamlines project development and 
improves data entry through real-time validation rules (with automated data type and range 
checks). REDCap also provides easy data manipulation (with audit trails for reporting, 
monitoring, and querying patient records) and an automated export mechanism to common 
statistical packages (SPSS, SAS, Stata, R/S-Plus). 
All data are entered into a central database using existing methodology that has been fully 
validated and undergoes continuous quality assurance by the PBRC Research Computing Core. 
All data are backed up daily, and the Research Computing Core at the PBRC oversees all data 
management. The research team has extensive experience using the procedures and methods 
required to conduct this study. Standard operating procedures in place throughout the units at 
Pennington Biomedical will be utilized for repeatable, valid data collection and quality.  
In accordance with the standards of practice followed by the Clinical Chemistry core, blood 
samples will be stored frozen at PBRC until analysis can be completed. Specific blood samples 
will be shipped to Boston Children’s Hospital and Cornell University for further analysis of 
plasma inflammatory markers and circulating tumor cells, respectively. Packaging and shipping 
of biological samples will be overseen by the PI, and they will be shipped by the laboratory and 
completed in accordance with International Air Transport Association regulations to ensure that 
viable biological samples reach their intended destination.  
Provisions to Monitor the Data to Ensure the Safety of Subjects
The study has a minimal level of risk to study participants and does not warrant the 
establishment of an independent Data and Safety Monitoring Board. This plan describes the 
safety monitoring procedures for the proposed study, including a description of how often and 
 
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to whom serious and unexpected adverse events will be reported. The plan will help ensure the 
safety of all participants. The PI will communicate via electronic submission to the IRB all 
unanticipated problems as defined by the IRB and all serious adverse events (SAEs) to the 
medical investigator within 72 hours.
The investigators will monitor the study's conduct. Study staff will report adverse events or 
other problems directly to the PI as they occur. The PI will schedule monthly meetings with 
study staff to review data on adverse events and recruitment or adherence to regimen 
problems. Any significant health problems coming to our attention during the study will be 
referred to the participant’s usual source of medical care with his/her permission. We will 
cooperate fully with his/her physician by providing relevant medical records. The following 
criteria, if detected during any part of the study regimen, will lead to referral to the 
participant’s usual source of medical care:
•
Clinical symptoms or signs of CVD include chest pain suggestive of angina pectoris, 
unusual dyspnea on exertion, severe ankle edema, and symptoms suggestive of 
transient ischemic attacks or intermittent claudication.
•
Musculoskeletal injuries or problems causing severe pain during exercise or interference 
with daily activities.
A. Adverse Events
Adverse events that occur will be classified as a Serious Adverse Event (SAE) or an 
Adverse Event (AE). Serious adverse events are defined to include:
•
Death
•
A life-threatening event
•
Severe illness, including worsening of a pre-existing condition, injury, or 
accidents
•
An inpatient hospitalization, surgical procedure, or treatment to prevent an SAE
•
A permanent disability or incapacity
•
A clinically significant abnormal laboratory or diagnostic test result
•
Any other event that, in the opinion of the principal investigator or study 
physician, might have resulted in a serious adverse event if medical intervention 
had not been initiated
Serious adverse events will be reported to the study PI, Project Manager, and MI 
throughout the trial. Serious adverse events will be collected from randomization until 
the final visit. SAE data will be analyzed quarterly, but serious or life-threatening 
adverse events require immediate reporting and follow-up in the event that an adverse 
event occurs on campus and results in a serious or life-threatening situation, the 
investigator or other project staff present will begin emergency measures, as 
appropriate, and call 911.
 
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•
For minor physical injury, the individual will be encouraged to see a health care 
practitioner of his or her choice.
•
If the study participant experiences psychological or emotional distress, the 
project staff will cease research activities and attempt to calm and reassure the 
participant. The participant will be directed to an appropriate healthcare 
practitioner for further assessment and treatment as needed.
•
The investigator and/or project staff will record detailed narrative notes 
describing the adverse event they witnessed or that was reported by the 
participant. The medical investigator will complete the Notification form for a 
Serious Adverse Event.
Serious adverse event reporting will follow the requirements of the IRB of the 
Pennington Biomedical Research Center. Serious adverse events that are unanticipated 
problems will be reported within 48 hours. 
For this trial’s purposes, an adverse event is defined as any health-related unfavorable 
or unintended medical occurrence that happens after randomization. Examples of 
Adverse Events include but are not limited to the following:
•
An event that requires a visit to a physician because it alters the participant’s 
ability to do physical activity
•
An event that occurs as a result of a study procedure which is not listed in the 
risks section of the consent
Adverse events will be reported to the study PI and the MI throughout the trial as 
necessary. Adverse event data will be collected from randomization until the final visit. 
Adverse events classified as serious will be reported from the date of consent through 
the final closeout visit. Adverse event data will be analyzed quarterly, but serious or life-
threatening adverse events require immediate reporting and follow-up. It is anticipated 
that most adverse events will be mild, and the participant will be able to resume 
intervention activities within a week of reporting the event.
B. Stopping Rules
There is minimal risk for participating in this trial. The most likely scenario that would 
indicate a cessation of the study would be a failure to recruit participants or implement 
the intervention as planned. Nevertheless, in addition to monitoring recruitment and 
compliance with the intervention, the staff will monitor the participant rates of injury. 
The medical investigator, in conjunction with the study investigators, will alert the IRB 
and funder if a larger than reasonably expected injury rate occurs in the treatment 
groups. 
Other issues that are related to the stopping rules include:
 
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•
New information – It is unlikely that new information will become available 
during this study, which would result in the discontinuation of the trial.
•
Limits of assumption – It is possible that the value of data analysis will be limited 
by differences between the intervention groups because of study dropouts or 
missing data.
•
Limit of rules – We acknowledge that circumstances other than what is listed 
may justify stopping the study.
Withdrawal of Subjects
There is no risk associated with withdrawal from the study. Participants may voluntarily 
withdraw or be withdrawn from the study if either the PI or MI feels that their continued 
participation would compromise the participant’s safety or the results of the study.
Risks to Subjects
The following is a list of reasonably foreseeable risks, discomforts, hazards, or inconveniences 
to the subjects related to the procedures in this study:
OGTT:  There is a possibility of pain, bruising, or infection at the site of the needle insertion for 
the IV line. Trained personnel minimize this risk. The drink may cause nausea, vomiting, 
abdominal bloating, or a headache.
DXA: The amount of radiation used for this procedure is very small. The radiation dose for this 
scan is equivalent to the radiation you are naturally exposed to in the environment in less than 
one day. Scans will not be performed on any subject who is pregnant. A pregnancy test will be 
performed within 72 hours before the scan on females of child-bearing potential.
Electrocardiogram (EKG or ECG):  There are minimal risks associated with this test. There is a 
small possibility there may be some redness or irritation while cleaning the skin prior to 
applying the electrodes or if you happen to be allergic to the adhesive on the electrodes.  
Blood Draws: There is the possibility of infection and/or pain and bruising at the vein on your 
arm where the needle is inserted.  Aseptic (sterile) technique and trained personnel minimize 
these risks.
Submaximal Fitness Assessment: All exercise testing core operating procedures are in 
accordance with the American College of Sports Medicine’s Guidelines for Exercise Testing and 
Prescription, as well as the American Heart Association. There is minimal risk of injury or a 
cardiovascular event during exercise testing. We believe the risk of an event during exercise 
testing is minimized with a pretest review of the medical history, physical examination, use of 
highly trained staff, and well-defined emergency procedures. Participants may experience 
temporary discomfort during blood pressure recordings due to the pressure of the blood 
pressure cuff on the arm. All tests are conducted in the presence of Exercise Testing Core 
personnel with extensive experience in conducting maximal exercise testing. All laboratory staff 
 
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are trained in BLS (basic life support – CPR) and/or ACLS (advanced cardiac life support). In the 
event of a life-threatening emergency, the subject would be treated with BLS/ACLS by trained 
individuals and subsequently be transported to the nearest acute care medical-surgical facility 
via Emergency Medical Services, which is a parish-wide paramedic response unit. 
Accelerometry & other activity monitoring devices: There is no known risk associated with 
measuring activity with accelerometers and other activity monitoring devices. Accelerometers 
fit comfortably on your waist or arm and can easily be adjusted should they become 
uncomfortable. In rare cases, the device may irritate the skin. If this should happen, the device 
can easily be repositioned to be more comfortable.
Blood Pressure Testing: Participants may experience temporary discomfort during blood 
pressure recordings due to the pressure of the cuff on their arm.
Height measurement: There is no risk to participants who record their height. 
Body weight: There is no risk to participants who record their body weight. Measurements will 
be taken in a secluded area to protect the participant’s privacy.
Self-report Questionnaires:  There are no anticipated risks from completing self-report 
questionnaires. If signs of minor stress or fatigue are apparent, participants will be given time 
to take a break from completing the questionnaires. It is estimated that the questionnaires will 
take approximately 30 minutes to complete. The questions contained in some of the 
questionnaires may make people feel uncomfortable. Responses to the questions will be coded 
to protect confidentiality, and participants may choose not to answer questions.
Exercise interventions:  The proposed exercise interventions are unlikely to cause major harm. 
We have conducted numerous exercise training studies and have never had a serious adverse 
event. There is the possibility of adverse events ranging from minor musculoskeletal problems 
to, in very rare cases, cardiovascular events. Exercise bouts will include aerobic exercise in the 
form of treadmill walking at moderate intensity. The exercise prescription will be at the 
discretion of the trainer to ensure compliance with the intervention and minimize injury.  
Occasionally, study participants experience minor orthopedic problems, but most self-correct 
with rest and standard first aid. These orthopedic injuries will be minimized by gradually 
progressing participants to their abilities as necessary. Exercise staff are trained in first aid and 
basic CPR. Each staff member is trained in either advanced or basic life support, and an 
automated external defibrillator and a fully stocked crash cart are kept on site. Although some 
study participants will be at moderately elevated risk for CVD, they will receive a thorough 
health screen, including a physical examination and an EKG. According to the available data on 
adverse events resulting from exercise, risk should be low. Fatal events during exercise are 
extremely rare. 
Potential Benefits to Subjects
 
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There are no direct benefits to the participants; however, the knowledge to be gained is critical 
for the advancement of science.
Vulnerable Populations
No vulnerable populations will be included in the study.
Multi-Site Research
This is a single-site research study.
Sharing of Results with Subjects
Subjects will be provided reports of medical information obtained from the body composition 
and exercise stress test following the completion of the study if requested.
Resources Available
The outpatient and inpatient research units are well-equipped and staffed to carry out the 
requirements of this study, and appropriate standards of practice are in place to ensure 
appropriate research procedures are followed.
Setting
This study will be conducted at Pennington Biomedical Research Center.
Compensation
Enrolled participants will be compensated up to $300 for the completion of the study. 
Participants will receive $100 after the completion of baseline and randomization if eligible. 
Participants will receive up to $100 for completion of all intervention-related activities (e.g., 
returning telephone calls, tracking, and adhering to the exercise protocol). The remaining $100 
will be reimbursed upon the completion of the study (defined as providing complete endpoint 
data measures). Reimbursements may be prorated according to the degree of adherence and 
willingness of participants to accommodate staff requests. Reimbursement will be requested 
from the LSU payroll department, and it usually takes about 3-4 weeks for it to arrive at 
Pennington Biomedical Research Center.
As part of the study, all exercise intervention participants will receive a treadmill for home use.  
After completing the three-month measures, control group participants will be provided with 
an in-home treadmill and individualized exercise program similar to that prescribed to the 
exercise group. As additional compensation for participation in the study, participants will keep 
the treadmill provided.
Confidentiality
 
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All participants are assured of their confidentiality, both verbally and in the informed consent 
form. The clinical facilities are strictly limited to the staff of the research institution and to 
research participants. This is accomplished by a variety of stringent security measures. All 
medical records are stored in locked areas. Access to these areas is limited to the clinical 
support staff, the director of the clinical facilities, and the PIs. Participants’ medical records are 
filed according to ID numbers. All forms on the chart display the ID number. Electronic data 
storage is similarly restricted, with only the PIs and authorized persons having access to 
databases containing confidential clinical records, i.e., those containing names OR other 
identifying information.
Data, including body weight, body composition, exercise testing, etc., will be collected from 
participants. Data are confidentially collected from study participants and are only used for 
research purposes. All records are kept in locked file cabinets, and participant data can be 
identified only by number. Data are used only in aggregate, and no identifying characteristics of 
individuals are published or presented.
Provisions to Protect the Privacy Interests of Subjects
All attempts will be made to maintain a subject’s privacy. Safeguards such as password-
protected computers and networks have been put in place to limit access to subject 
data.  Subjects will be given ample time to read over the consent, ask questions, and 
agree to participate in the research study. Subjects may decline to answer questions 
they are not comfortable with. Each procedure will be explained to the subject before it 
is performed. All discussions will be held with study subjects in examination or private 
rooms to protect their privacy.  
Compensation for Research-Related Injury
No form of compensation for medical treatment or for other damages (i.e., lost wages, time 
lost from work, etc.) is available from the Pennington Biomedical Research Center. In the event 
of injury or medical illness resulting from the research procedures in which a subject 
participates, they will be referred to a treatment facility. Medical treatment may be provided at 
the subject’s expense or at the expense of their health care insurer (e.g., Medicare, Medicaid, 
Blue Cross-Blue Shield, Dental Insurer, etc.), which may or may not provide coverage. The 
Pennington Biomedical Research Center is a research facility and provides medical treatment 
only as part of research protocols. Should the study subject require ongoing medical 
treatments, they must be provided by community physicians and hospitals.
Economic Burden to Subjects
All study-related tests and procedures will be at no cost to the subject. The subject will incur 
transportation costs to and from PBRC.
Consent Process
 
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The clinic staff will obtain informed consent during the first screening visit as per established 
policies and procedures. All clinic staff have completed the required GCP training and have 
trained on the consenting policies and procedures. An ample opportunity will be given for the 
subject to review the consent form and ask any questions prior to signing it. If subjects wish, 
they can take the form home and return it at a different time of the visit. A copy of the consent 
will be provided to the subject.
 
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