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. Author manuscript; available in PMC: 2020 Nov 4.
Published in final edited form as: Appl Physiol Nutr Metab. 2020 Jun 22;45(11):1306–1309. doi: 10.1139/apnm-2019-0836

Associations between physical activity, sedentary behavior and urinary oxidized guanine in colorectal cancer patients: Results from the ColoCare Study

Richard Viskochil 1, Biljana Gigic 2, Tengda Lin 1, Stephanie Skender 3, Jürgen Böhm 1,3, Petra Schrotz-King 3, Karen Steindorf 4, Robert Owen 3, Jane C Figueiredo 5, Christopher I Li 6, Erin M Siegel 7, Sheetal Hardikar 1,8, David Shibata 9, Adetunji T Toriola 10, Martin Schneider 2, Alexis Ulrich 2,11, Cornelia M Ulrich 1,2,8, Jennifer Ose 1,8
PMCID: PMC7609563  NIHMSID: NIHMS1614087  PMID: 32569481

Abstract

To determine associations between physical activity (PA), sedentary behavior (SB) and oxidative stress in colorectal cancer patients, ColoCare Study participants in Germany wore an accelerometer 6- and/or 12-months post-surgery. Spearman partial correlations were used to assess associations between PA and urinary concentrations of oxidized guanine, a validated marker of oxidative stress. There were no significant associations between PA or SB and oxidized guanine in n=76 measurements (ng/mgcreatinine; r=0.03, p=0.76 for PA, r=−0.05, p=0.69 for SB).

Keywords: Colorectal cancer, accelerometer, sedentary time, oxidative stress, 8-OHdG

Introduction

Colorectal cancer is the third most commonly diagnosed cancer in men and the second most commonly diagnosed cancer in women (Siegel et al. 2017). Evidence from recent epidemiological studies suggests that both physical activity (PA) and sedentary behavior (SB) influence colorectal cancer prognosis (van Zutphen et al. 2017; Brenner et al. 2018), and the pathophysiological mechanisms underlying these associations are currently being explored (Ulrich et al. 2018). One plausible biological mechanism that may link PA and SB to colorectal cancer outcomes is oxidative stress and damage to DNA (Perse 2013). The repair of the damaged DNA base guanine releases oxidized guanine species into the circulation, and concentrations of 8-hydroxyguanine, 8-hydroxyguanosine, and 8-hydroxy 2’-deoxyguanosine (8-OHdG) in serum or urine represent a validated marker of oxidative stress (Wu et al. 2004; Borrego et al. 2013). High levels of oxidized guanine species have consistently been linked to the development and prognosis of colorectal cancer (Sato et al. 2010; Dziaman et al. 2014; Guo et al. 2016).

Exercise training decreases oxidative stress in healthy, sedentary adults (Campbell et al. 2010), and acute supervised exercise can reduce urinary markers of oxidized guanine in colorectal cancer patients following primary therapy (Allgayer et al. 2008). However, the associations between habitual PA, SB and DNA/RNA oxidation have yet to be investigated in colorectal cancer survivors. The purpose of this study was to investigate the relationship between habitual, objectively-measured PA and SB derived from accelerometers and urinary markers of oxidized guanine in newly diagnosed colorectal cancer patients. We expect that daily, habitual levels of PA will be inversely associated and SB will be positively associated with urinary concentrations of oxidized guanine in colorectal cancer survivors.

Methods

The present study was conducted within the Heidelberg, Germany site of the ColoCare Study (ClinicalTrials.gov identifier: NCT02328677), an international prospective cohort of >2,500 newly diagnosed colorectal cancer patients (Ulrich et al. 2019). Inclusion criteria are: patients diagnosed with stage I-IV colon or rectal cancer age 18–89, German-speaking, and both mentally and physically able to consent to participate in the study. Baseline data collected include biospecimen (e.g. tissue, blood, urine) and self-administered questionnaires (e.g. physical activity, medication use). Participants are followed for up to five years, with additional questionnaire and/or biospecimen collection at 3, 6, 12, 24, 36, 48 and 60 months. All participants included in the present study had physical activity evaluated via accelerometry and oxidative stress evaluated using urinary oxidized guanine at the 6- and/or 12-month time-points. The study was approved by the Ethics Committee of the University of Heidelberg and all participants provided written informed consent.

Physical activity measurement

A complete description of the objectively measured physical activity protocol and description of the ColoCare Heidelberg cohort has been published elsewhere (Skender et al. 2017). Briefly, PA and SB were evaluated using the Actigraph GT3x accelerometer (Actigraph, Pensacola FL), worn around the chest during waking hours for at least five consecutive days. Data were collected at 30 Hertz (Hz) intervals, and raw data were downloaded using the ActiLife software package (v. 6.6.3) and summed into 10 second epochs. The chest was chosen as the wear location instead of the waist in order to avoid interference with ostomy bags, stoma or surgical scars within this population. Additionally, self-reported physical activity was collected using a modified Vitamins and Lifestyle (VITAL) questionnaire incorporated into the baseline, 6- and 12-month ColoCare Study questionnaire (Littman et al. 2004).

Biomarker measurement

Urine was collected in sterile cups at the 6- and 12-month time-points and stored at −80˚C for future analysis. Urine collection occurred within a one week window surrounding the period of accelerometer wear. Concentrations of oxidized guanine species were quantified using a validated enzyme-linked immunosorbent assay (ELISA; Cayman Chemical Co., high sensitivity DNA/RNA oxidative damage kit product #589230, Ann Arbor, MI). This assay is based on competition between the oxidized guanine species within the sample and an 8-OHdG conjugate binding to anti-mouse clonal IgG. Cross-reactivity with each species of oxidized guanine is 100%, 38% and 23% for 8-OHdG, 8-hydroxyguanosine and 8-hydroxyguanine, respectively. Samples were run in duplicate, and control samples were included across the multiple assay plates. Coefficients of variation (CV’s) were determined using the standard deviation and mean of these duplicate (intra-assay) and control (inter-assay) samples. All intra- and inter-assay CVs were required to be <10% in order for the assay to be considered valid. Urinary oxidized guanine concentrations were normalized to concentrations of urinary creatinine, determined via a colorimetric creatinine assay (Cayman Chemical Co, Ann Arbor MI), to eliminate differences in original sample dilution of urine and log transformed for analysis.

Physical activity and sedentary behavior analysis

Accelerometer data were considered valid if participants wore the activity monitor on >4 consecutive days for at least 8 hours per day. Non-wear time was defined as >60 minutes of consecutive zero counts with a 2 minute interruption tolerance (Skender et al. 2017). Daily minutes and percent of daily accelerometer wear time spent in sedentary, light intensity physical activity (LIPA), and moderate to vigorous intensity physical activity (MVPA) were calculated using Freedson cut-points (Freedson et al. 1998). Additional PA-specific variables included steps, energy expenditure (kcal), and exercise minutes (MVPA in bouts of >10 consecutive minutes). SB-specific variables included the number of daily sedentary periods >30 consecutive minutes (bouts), as well as the number of interruptions in sedentary time (breaks). Self-reported physical activity metabolic equivalent (MET) hours/week at baseline and each accelerometer time-point were derived from the VITAL questionnaire.

Statistical analysis

Spearman partial correlations were calculated to assess the association between minutes per week of physical activity at each intensity level, weekly exercise minutes, SB-specific variables and urinary oxidized guanine, adjusting for age, sex, body mass index (BMI; kg/m2), tumor stage and baseline physical activity levels. Correlations were evaluated independently at 6-month, 12-month time-points as well as a combination of 6- and 12-month time-points. All statistical tests were two-sided with a p<0.05 considered to be significant. All statistical analyses were performed using SAS v. 9.4 (Raleigh, NC).

Results

Seventy-six patients had available physical activity and oxidized guanine measurements at 6- (n=44) and 12- (n=32) month time-points. Fourteen participants had physical activity and oxidized guanine measurements at both the 6- and 12-month time-points. At both the 6- and 12-month follow-up time-points, patients were older (mean age >60 years), overweight (BMI averaging between 25–29.9 kg/m2), and inactive, with less than 10% meeting the American College of Sports Medicine Physical Activity Recommendations (150+ min/wk of MVPA in bouts of >10 consecutive minutes, Table 1). Participants spent a similar percentage of accelerometer wear time at each time-point in sedentary (74 ± 6% and 73 ± 7%), light-intensity (21 ± 5% and 22 ± 7%) and moderate-to-vigorous (6 ± 1% and 5 ± 2%) physical activity. Oxidized guanine was comparable between 6-month and 12-month time-points (6.92 ± 0.5 and 6.85 ± 0.4 ng/mg creatinine). There were no statistically significant differences observed in participant characteristics, PA/SB measures, or oxidized guanine across the 6-month and 12-month time-points (Table 1).

Table 1:

Participant characteristics

Patient Characteristics 6 months (n=44) 12 months (n=32) Combined (n=76)
Age (y) 60.1 ± 13.9 62.1 ± 10.0 61.0 ± 12.3
AJCC Stage, n (%)
I 7 (16) 8 (25) 15 (20)
II 16 (36) 11 (34) 27 (36)
III 15 (34) 10 (31) 25 (33)
IV 6 (14) 3 (7) 9 (12)
Sex, n (%)
Male 28 (64) 22 (69) 50 (66)
Female 16 (36) 10 (31) 26 (34)
BMI (kg/m2) 25.6 ± 4.8 26.3 ± 3.4 25.9 ± 3.9
Oxidized guanine (ng/mL) 6.92 ± 0.5 6.85 ± 0.4 6.90 ± 0.4
Total MVPA* (min/week) 264 ± 184 263 ± 182 264 ± 180
Steps (steps/day) 6184 ± 2898 6646 ± 3358 6332 ± 3151
Exercise† minutes (min/week) 86 ± 130 74 ± 123 81 ± 126
Meeting PA^ guidelines, n (%) 7 (16) 5 (16) 12 (16)
Daily light intensity (%) 20.8 ± 4.6 21.7 ± 7.0 21.2 ± 5.7
Daily sedentary (%) 73.7 ± 6.4 73.2 ± 7.0 73.5 ± 6.5
Sedentary bouts (bouts/day)‡ 10.3 ± 2.6 10.1 ± 3.1 10.2 ± 2.8
Sedentary breaks (breaks/day) 10.9 ± 2.7 10.9 ± 3.2 10.9 ± 2.8
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Data presented as mean ± SD.

*

Moderate to vigorous physical activity,

†

Exercise minutes defined as MVPA in bouts of ≥10 minutes.

^

≥150 minutes of weekly exercise minutes.

‡

Bout defined as ≥20 consecutive minutes of sitting time.

We did not observe statistically significant correlations between oxidized guanine and metrics of PA, including: 1) total MVPA, 2) weekly exercise minutes, 3) (%) daily MVPA, 4) steps/day, and 5) questionnaire-derived PA (METs, Table 2). There were also no significant associations between oxidized guanine and (%) sedentary time, bouts of sedentary time or any other measure of SB (Table 2). Additionally, when patients were grouped into quartiles of weekly exercise minutes, we did not observe any significant differences in oxidized guanine between those with the highest and those with the lowest volume of weekly exercise (data not shown). Notably, we did observe several significant inverse associations between objectively measured PA and objectively measured SB (Table 2). Correlations between percent of daily sedentary time and metrics of PA that are independent of the proportion of wear time, such as exercise minutes (r=−0.26, p=0.04), steps per day (r=−0.62, p<0.01) and energy expenditure (r=−0.54, p<0.01) were inversely related.

Table 2:

Associations between physical activity, sedentary behavior and oxidative stress, adjusted for sex, stage, BMI and baseline physical activity.

Variable 1 Variable 2 Corr. Coefficient p-value
Oxidized guanine MVPA (min/week) 0.04 0.76
Exercise† minutes (min/week) 0.02 0.86
Steps (steps/day) 0.08 0.52
Energy expenditure (kcal/day) −0.01 0.95
Daily sedentary time (%) −0.05 0.69
MET h/wk (Self-report) Exercise† minutes (min/week)* 0.35 <0.01
Steps (steps/day)* 0.37 <0.01
Energy expenditure (kcal/day)* 0.29 0.02
Daily sedentary time (min/day) 0.15 0.25
Daily sedentary time (%) −0.03 0.81
Daily sedentary time (%) Exercise† minutes (min/week)* −0.26 0.04
Steps (steps/day)* −0.62 <0.01
Energy expenditure (kcal/day)* −0.54 <0.01
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*

p<0.05.

†

Exercise minutes defined as moderate-to-vigorous physical activity (MVPA) in bouts of ≥10 minutes.

Discussion

Prior studies have incorporated accelerometers to quantify objectively measured physical activity in colorectal cancer survivors (Vallance et al. 2014; Lynch et al. 2016; van Roekel et al. 2016), however this study is the first to evaluate the associations between objectively measured habitual physical activity and a validated biomarker of oxidative stress that influences colorectal cancer prognosis. We observed no statistically significant associations between urinary oxidized guanine and levels of physical activity or sedentary behavior. These results are surprising given the reductions in oxidative stress previously observed in exercise interventions, both in healthy adults and in cancer patients.

There are several potential explanations for the lack of association between physical activity and DNA oxidation. First, this cohort is relatively active, lean, and has lower urinary oxidized guanine compared to other studies (e.g. Allgayer et al. 2008; Guo et al. 2016). Additionally, it is possible that oxidized guanine as well as creatinine concentrations in cancer survivors are tightly linked to dietary patterns and supplement use (Yeon et al. 2011). In order to accurately evaluate urinary metabolites, it is common to standardize those values to levels of creatinine. It is possible that physical activity may impact concentrations of creatinine and thus also confound the assessment of oxidized guanine. Finally, while the quantification of oxidized guanine species via ELISA represents a validated measure of oxidative stress that has been linked to colorectal cancer, many of those associations were quantified prior to primary colorectal cancer treatment and/or utilized the gold-standard technique of mass spectroscopy to identify specific forms of oxidized guanine. We were unable to address these potential confounding factors or adjust concentrations of oxidized guanine to account for differences in urinary osmolality. However the ColoCare Study is currently incorporating paired accelerometry and mass spectrometric analysis for the evaluation of metabolomics at study sites throughout the United States. It is possible that the inclusion of a population with 1) greater variation in BMI and sedentary time, 2) additional evaluation of dietary intake, and 3) utilization of gold-standard measurement techniques for the evaluation of oxidative stress and DNA damage will shed further light on the importance of physical activity and sedentary behavior in colorectal cancer patients.

We observed several novel findings regarding the relationship between physical activity and sedentary behavior in colorectal cancer survivors. Emerging evidence suggests that sedentary time and exercise are independent variables, such that high levels of daily sedentary time can negatively influence health outcomes independent of daily exercise volume (Koster et al. 2012; Arem et al. 2015). In previous accelerometer-based studies of physical activity in colorectal cancer survivors, the relationship between physical activity and sedentary time was unclear (Vallance et al. 2014; Lynch et al. 2016; Shi et al. 2018). We observed significant inverse associations between sedentary behavior and physical activity variables, including exercise time. Our findings suggest that colorectal cancer survivors who engage in exercise may also tend to spend less time sitting during the day, as opposed to increasing daily sedentary behavior (e.g. from physical activity-induced fatigue). As such, lower sedentary time throughout the day may be an additional benefit of exercise in colorectal cancer patients. Taken together, we did not observe any statistically significant associations between urinary markers of DNA damage and physical activity, but the intricate link between energy balance and oxidative stress suggest that markers of DNA damage may represent potent tools to evaluate the impact of energy imbalance longitudinally in colorectal cancer survivors.

Novelty:

Objectively-measured physical activity was not associated with a marker of oxidative stress in colorectal cancer patients

Acknowledgements and Funding

RV receives support from R01 CA207371, U01 CA206110, and the American College of Sports Medicine Paffenbarger Blair Fund for Physical Activity Epidemiology.

BG was funded by the ERA-NET on Translational Cancer Research (TRANSCAN) project 01KT1503, the National Institutes of Health/National Cancer Institute (NHI/NCI) project R01 CA189184, the Stiftung LebensBlicke, and the Matthias-Lackas Foundations.

TL, JCF, CIL, EMS, SH, DS, ATT, MS, AU, CMU and JO’s work on this project was funded by the NIH/NCI projects, R01 CA189184, R01 CA207371, U01 CA206110 and the Huntsman Cancer Foundation. CMU received additional funding from NIH P30 CA042014.

Footnotes

Conflicts of interest: CMU has as cancer center director oversight over research funded by several pharmaceutical companies, but has not received funding directly herself. The other authors declare no conflicts of interest.

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