Research Article

Inflammation and Oxidative Stress Markers and EsophagealAdenocarcinoma Incidence in a Barrett's Esophagus Cohort

Sheetal Hardikar1,2, Lynn Onstad1, Xiaoling Song1, Angela M. Wilson3, Thomas J. Montine3, Mario Kratz1,2,4,Garnet L. Anderson1,5, Patricia L. Blount1,4,6, Brian J. Reid1,4,6,7, Emily White1,2, and Thomas L. Vaughan1,2

AbstractBackground: Persons with Barrett’s esophagus experience increased risk of esophageal adenocarcinoma.

Prediagnostic inflammation markers predict several cancers, but their role in predicting esophageal adeno-

carcinoma is unknown.

Methods: We investigated whether biomarkers of inflammation [C-reactive protein (CRP), interleukin-6

(IL6), soluble tumor necrosis factor (sTNF) receptors I and II], and of oxidative stress (F2-isoprostanes)

predicted progression to esophageal adenocarcinoma in a prospective cohort of 397 patients with Barrett’s

esophagus, 45 of whom developed esophageal adenocarcinoma. Biomarkers were measured in stored plasma

samples from two time points during follow-up, themean of which served as the primary predictor. Adjusted

hazard ratios (HR) and 95% confidence intervals (CI) were estimated using Cox regression.

Results: CRP level above the median was associated with an 80% increased risk of esophageal adenocar-

cinoma. The HR and 95% CI adjusted for age, gender, and further adjusted for waist–hip ratio and smoking

were 1.98 (1.05–3.73) and 1.77 (0.93–3.37), respectively, with Ptrend for continuous CRP¼ 0.04. Personswith IL6

levels above themedian also had almost 2-fold increased risk [HR and 95%CI adjusted for age and gender, and

further adjusted forwaist–hip ratio and smokingwere 1.95 (1.03–3.72) and 1.79 (0.93–3.43), respectively, but no

evidence of a trend was observed]. Concentrations of TNF receptors and F2-isoprostanes were not associated

with esophageal adenocarcinoma risk.

Conclusions: Further research is needed to evaluate the role of inflammation and associated markers in

esophageal adenocarcinoma development in persons with Barrett’s esophagus.

Impact: This prospective study suggests that inflammation markers, particularly CRP and IL6, may help

identify persons at higher risk of progression to esophageal adenocarcinoma.Cancer Epidemiol Biomarkers Prev;

23(11); 2393–403. �2014 AACR.

IntroductionChronic inflammation has beenhypothesized toplay an

important role in the pathogenesis of cancers of the lung,colon, and other organs (1–5). Similarly, oxidative stresshas been implicated in cancers of the lung (6), breast (7),and prostate (8). Inflammation may contribute to cancerdevelopment through multiple mechanisms, includingDNA damage, angiogenesis, promotion of cellular pro-

liferation, and inhibition of apoptosis (1). Inflammatoryprocesses also lead to generation of reactive oxygen spe-cies (ROS), which may cause inactivating mutations intumor-suppressor genes or posttranslational modifica-tions in DNA repair proteins, thus promoting carcinogen-esis (1, 5).

In the gastrointestinal tract, several chronic inflamma-tory conditions have been associated with cancer: inflam-matory bowel disease with colorectal cancer (9), hepatitisB and C with liver cancer (10), and chronic Helicobacterpylori gastritis with gastric cancer (11). Similarly, inflam-matory conditions of the esophagus, namely reflux esoph-agitis and Barrett’s esophagus, are implicated in thedevelopment of esophageal adenocarcinoma, withBarrett’s esophagus widely considered to be a premalig-nant lesion for esophageal adenocarcinoma (12–14). Theinflammatory link with esophageal adenocarcinoma isfurther strengthened by the observation that regular useof nonsteroidal anti-inflammatory drugs (NSAID) andaspirin is associated with decreased risk (15–17).

The incidence of esophageal adenocarcinoma hasincreased dramatically over the past four decades (18).

1Public Health Sciences Division, Fred Hutchinson Cancer Research Cen-ter, Seattle, Washington. 2Department of Epidemiology, University ofWashington, Seattle, Washington. 3Department of Pathology, Universityof Washington, Seattle, Washington. 4Department of Medicine, Universityof Washington, Seattle, Washington. 5Department of Biostatistics, Univer-sity of Washington, Seattle, Washington. 6Human Biology Division, FredHutchinsonCancer ResearchCenter, Seattle,Washington. 7Department ofGenome Sciences, University of Washington, Seattle, Washington.

Corresponding Author: Sheetal Hardikar, Fred Hutchinson CancerResearch Center, 1100 Fairview Avenue North, M4-B402, PO Box19024, Seattle, WA 98109-1024. Phone: 206-667-7140; Fax: 206-667-4787; E-mail: shardika@fhcrc.org

doi: 10.1158/1055-9965.EPI-14-0384

�2014 American Association for Cancer Research.

CancerEpidemiology,

Biomarkers& Prevention

www.aacrjournals.org 2393

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

Although the relative risk of esophageal adenocarcinomais at least 30-times higher in individuals with Barrett’sesophagus compared with those without (19), absoluterisk of progression is relatively low (0.12%–0.6% per year;refs. 20–22), and it is not yet clear which persons withBarrett’s esophagus aremost likely to develop esophagealadenocarcinoma, orwhether lifestylemodificationsmighthelp prevent esophageal adenocarcinoma within thishigher-risk population. Previous studies identified obe-sity, cigarette smoking, gastroesophageal reflux, and dietas potential modifiable risk factors for esophageal adeno-carcinoma (14, 23). However, the roles of inflammationand oxidative stress as potentially modifiable risk factorsor predictors for esophageal adenocarcinoma have notbeen studied directly. In this report, we assess the asso-ciation between markers of systemic inflammation andoxidative stress and the subsequent risk of esophagealadenocarcinoma in a well-characterized Barrett’s esoph-agus cohort followed for up to 14.5 years.

Materials and MethodsStudy population, follow-up, and cancerascertainment

The Seattle Barrett’s Esophagus Study (SBES), is aprospective cohort study aimed at understanding the riskfactors and mechanisms underlying neoplastic progres-sion to esophageal adenocarcinoma among persons withBarrett’s esophagus. The SBES cohort is one of the largestand longest-running well-characterized cohorts of per-sons with Barrett’s esophagus in the world. Details of thecohort have been described previously (23, 24). The studyinvolves periodic endoscopic surveillance for all partici-pants, withmultiple biopsies of the Barrett’s segment. Thecurrent report includes the 427 SBES participants withBarrett’s esophagus and no history of esophageal cancerenrolled between February 1995 and September 2009, ofwhom411 (96.3%) had at least one follow-up visit. At theirbaseline visit, participants underwent an extensive per-sonal interview, anthropometric assessment, endoscopywith biopsy, and blood draw. At subsequent follow-upvisits, baseline information was updated, additionalbloodwas collected, anda repeat endoscopywith biopsieswas performed (12, 13, 25). Specimens were fixed, pro-cessed, and interpreted by a single pathologist blinded tothe participants’ exposure status (12). Individuals wereclassified as having Barrett’s esophagus, low-grade dys-plasia, high-grade dysplasia, or esophageal adenocarci-noma based on their most severe histologic diagnosis.Participants with high-grade dysplasia at their initialendoscopy (80 of 411 participants; 19.46%) were endos-coped twice more within 5 months to detect any occultcancers missed at baseline. Of the 411 SBES participantseligible for analyses, 14 individuals had less than 5monthsof follow-up and 11 of these developed cancer. Because ofan a priori concern that cancers diagnosed during thisearly period of intensive search for occult malignanciesmay have been present at baseline, these 14 individuals

were excluded from the primary statistical analyses.Esophageal adenocarcinoma was defined as invasion ofneoplastic epithelium beyond the basement membrane ofthe esophageal mucosa into the surrounding lamina pro-pria, muscularis mucosa, or submucosa (12). This studywas approved by the Institutional Review Boards at theUniversity of Washington and Fred Hutchinson CancerResearch Center.

Inflammation marker measurementsFasting blood samples were collected, processed

(most within 2 hours after collection), and stored at�80�C until analysis. Potential inflammation markerswere identified from the literature, and selected for usein this study based on the sensitivity and accuracy ofavailable assays, giving consideration to their cost.Plasma levels of C-reactive protein (CRP), IL6, andsoluble tumor necrosis factor (sTNF) receptors weremeasured using samples from the first two availabletime points (baseline and first follow-up for most par-ticipants, mean duration between two samples, 1.8years). F2-isoprostanes were measured at a single timepoint (earliest available, baseline for most). Intra- andinterbatch coefficients of variation (CV) were calculatedusing blinded pooled plasma samples included as qual-ity controls within each batch. Intraclass correlationcoefficients (ICC) and 95% confidence intervals (CI)were calculated using the two cytokine measurementsper participant, as described previously (26).

Briefly, CRP concentrations were measured in never-thawed plasma samples with a high-sensitivity assayusing immunonephelometry [Dade Behring Inc; Inter-batch CV; 2.9%; ICC (95% CI), 54.9% (47.4–62.5)]. Theassay detectable limit was 0.2 mg/L; a value of 0.1 mg/L was assigned to all participants with CRP concentra-tions below the detection limit (<1% samples). IL6 wasassayed in never-thawedplasma samples using theQuan-tikine HS human IL6 Elisa kit [R&D Systems Inc; HS600B;interbatch CV, 4.4%; intrabatch CV, 4.1%; ICC (95% CI),56.4% (49.4–63.4)]. Samples were run in duplicate with amedian duplicate CV of 2.7%; samples with CVs greaterthan 12% were rerun and the repeat measurements wereused for analysis. sTNFR-I and sTNFR-II were measuredon previously thawed and refrozen plasma samples usingthe MILLIPLEX MAP Human Soluble Cytokine ReceptorPanel [Millipore; HSCR-32K; sTNFR-I: interbatch CV,8.9%; intrabatch CV, 5.9%; ICC (95% CI), 89.2% (87.2–91.3); sTNFR-II: interbatch CV, 6.1%; intrabatch CV, 2.4%,ICC (95% CI), 84.9% (82.1–87.8)]. Samples were run induplicate with a median duplicate CV of 4.1% and 3.4%for sTNFRI and sTNFRII, respectively. F2-isoprostaneswere estimated in never-thawed plasma samples usinggas chromatography/negative ion chemical ionizationmass spectrometry (GC/NICI-MS) as describedprevious-ly (6890N Agilent gas chromatograph, 5973 quadruplemass spectrometer; refs. 27, 28). This assay had a precisionof �3%, accuracy of 97%, and a detection limit of 20 pg/mL.

Hardikar et al.

Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 Cancer Epidemiology, Biomarkers & Prevention2394

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

Other covariatesDetailed information on medical, family, and medica-

tion histories, as well as on lifestyle exposures such astobacco use, were collected in a personal interview con-ducted by trained staff. Anthropometric measurements,including height, weight, and waist, hip, thigh, andabdominal circumferences, were determined using anestablished protocol (23) and were used to calculatewaist–hip ratio (WHR) andbodymass index [BMI;weight(kg)/height (m)2], which was categorized as normal (<25kg/m2), overweight (�25 and <30 kg/m2), and obese (�30kg/m2). Cumulative pack-years of smoking were com-puted using the number of packs smoked per day and thenumber of years smoked.

Statistical analysisWe estimated the associations between markers of

inflammation and oxidative stress and the risk of esoph-ageal adenocarcinoma by calculating hazard ratios (HR)and 95%CIs using separate Coxmodels for each biomark-er. As we dropped individuals with less than 5 months offollow-up from our primary statistical analyses, time atrisk todevelopment of esophageal adenocarcinomabegan5monthspostbaseline for all participants. Forparticipantswith inflammationmarkermeasurements available at twotime points during the follow-up, the mean of the twomeasures was used as the primary predictor; otherwise,the lone measure was used. If the outcome occurredduring the same visit as a biomarker measurement, onlythe first measurement was used for risk prediction. Apriori, we decided to exclude CRP values more than 10mg/L from analyses due to the possibility that suchelevated concentrations may be the result of acute ratherthan chronic inflammation (29, 30). Eight participants hadboth their CRP measures more than 10 mg/L and, hence,were dropped from all CRP-related analyses. Biomarkerconcentrations were tested for associations with esoph-ageal adenocarcinoma risk using three models: unadjust-ed; age- and gender-adjusted; and adjusted for obesity,smoking, and NSAID use in addition to age and gender.These specific confounders were selected on the basis ofthe previously established risk factors of esophageal ade-nocarcinoma (increasing age, male gender, smoking,higherWHR, and use of NSAIDs; refs. 14, 23) determineda priori, and their correlations with the biomarkers, basedon Pearson correlation coefficients. Although it is moreconventional to adjust for the effects of obesity by adjust-ing for BMI, we chose to adjust for WHR (a measure ofcentral adiposity) in the current analyses as there is goodevidence that WHR is more strongly associated withBarrett’s esophagus as comparedwith BMI (31). Analyseswere conducted on quartiles of the various biomarkers, aswell as bymedian level.Analyseswere repeated restrictedtomales (number of femaleswas too small formeaningfulresults). Tests for trendwere based on the likelihood-ratiotest associated with addition of the biomarker beingevaluated in its continuous form. A two-sided P valueless than 0.05 was considered statistically significant. The

assumption of proportional hazards over timewas tested.All analyses were performed using STATA statisticalpackage, release 12 (StataCorp).

ResultsParticipant characteristics for the 411 individuals eligi-

ble for current analyses are presented in Table 1. Themajority of the cohort was Caucasian (96.6%) and male(81.3%), with a mean age of 61.2 years. More than 39% ofthe participants were obese, 64% were current or formersmokers, 81.5% reported regular alcohol use in theirlifetime, and 60.6% had regularly taken NSAIDs at somepoint in their life. ThemeanWHR for the entire cohortwas0.95 (males, 0.96; females, 0.87).

The medians and interquartile ranges for the variousbiomarkers overall and by gender are presented in Table 2(distributions by gender were based on sex-specific quar-tiles). Overall, the distributions of the various biomarkersevaluated in this study are comparable with other studiesof older populations and obese individuals (32–34).

The Pearson correlation coefficients between the bio-markers and risk factors for esophageal adenocarcinomaare shown in Table 3. The correlations between the bio-markers themselves were small to moderate (0.12–0.63),and were statistically significant (except IL6 and F2-iso-prostanes). Therefore, these biomarkerswere evaluated inseparate models with respect to their esophageal adeno-carcinoma risk. Most of the biomarkers were significantlycorrelated with age, WHR, and cigarette pack-years, sug-gesting that these factors might confound the biomarker–esophageal adenocarcinoma associations.

Table 4 presents the HRs and 95% CIs for developingesophageal adenocarcinoma according to biomarkerlevels among the 397 participants in the primary analysis.They were followed for a median of 6.14 years (31,677person-months), and 45 developed cancer. Plasma sam-ples were not available for 3 individuals. For analysesinvolving CRP, we omitted 8 participants for whom allavailable CRP values were more than 10 mg/L (2 out ofthese 8 developed esophageal adenocarcinoma; total 43cancers in analyses based on CRP). Ultimately, analyseswere conducted on 394 participants for IL6 and sTNFreceptors, 386 participants for CRP, and 377 participantsfor F2-isoprostanes.

Mean CRP levels above the median of 1.9 mg/L wereassociated with a 2-fold increased esophageal adenocarci-noma risk compared with those with values below themedian, after adjustment for age andgender (HR, 1.98; 95%CI, 1.05–3.73, Ptrend for continuous CRP ¼ 0.01). Furtheradjustment forWHR, smoking, andNSAIDsattenuated theassociation somewhat (HR, 1.77; 95% CI, 0.93–3.37; Ptrend

for continuous CRP ¼ 0.04). Analyses limited to menrevealed slightly stronger associations with CRP.

Participants with average IL6 levels above the medianhad a 2-fold increased risk for esophageal adenocarcino-ma (HR, 1.95; 95%CI, 1.03–3.72) but no evidence of a trendwas observed (Ptrend¼ 0.94). The increase in riskwasmore

Chronic Inflammation and Risk of Esophageal Adenocarcinoma

www.aacrjournals.org Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 2395

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

pronounced among males, with an almost 3-foldincreased risk (HR, 2.85; 95% CI, 1.38–5.92) after adjust-ment for age and gender. Overall as well as in thesubgroup analysis for males, additional adjustmentfor obesity, smoking, and NSAIDs had little effect.No evidence of an association between sTNF-RIand esophageal adenocarcinoma risk was observed. ForsTNF-RII, although univariate analyses overall andamong males revealed statistically significant associa-tions, adjustment for confounders attenuated the asso-ciation substantially such that it was no longer statis-tically significant. Circulating levels of F2-isoprostaneswere not associated with increased risk of esophagealadenocarcinoma in this cohort.

To evaluate if the significant associations observedwithCRP and IL6 were more pronounced within the first fewyears of follow-up, we conducted subanalyses restrictingthe follow-up time to 3 and 5 years from baseline (Fig. 1).The associations with CRP were stronger and the statis-tical trends more pronounced (Ptrend for continuousCRP ¼ 0.02 in a fully adjusted model) with the restricted

5-year follow-up. The associations with IL6 did not altermuch after restricting the follow-up to 3 or 5 years. Werepeated our main analyses after adding the 14 indivi-duals with less than 5 months of follow-up that we hadinitially excluded, with no important differences found(data not shown). To examine whether prolonged storageof biologic samples affectedour results,wealso conducteda sensitivity analysis restricted to only those individualswhose biologic samples had been stored for 10 years orless (mean storage time for the earlier of the two samplesfrom an individual was 12 years).We found that althoughthe results forCRPwere slightly stronger and those for IL6were weaker as compared with the main statistical anal-ysis, the overall conclusions remained the same (data notshown). As IL6 may be produced by the metaplasticepithelium (35), we also investigated the effect of adjust-ment for Barrett’s esophagus segment length in modelsinvolving IL6 and found that the point estimate wasreduced by a small amount—from 1.79 to 1.60.

We also computed an "inflammation score" based onthe quartile categories for various biomarkers such that

Table 1. Baseline characteristics of all participants, males and females, in the SBES cohort (n ¼ 411)

Variable All participants (n ¼ 411) Males (n ¼ 334) Females (n ¼ 77)

N [% or median (IQR)] N [% or median (IQR)] N [% or median (IQR)]

Agea, y30–44.9 30 (7.3) 25 (7.5) 5 (6.5)45–54.9 97 (23.6) 83 (24.9) 14 (18.2)55–64.9 110 (26.8) 84 (25.2) 26 (33.8)65–74.9 114 (27.7) 93 (27.8) 21 (27.3)�75 60 (14.6) 49 (14.7) 11 (14.3)

Racea

White 397 (96.6) 324 (97.0) 73 (94.8)Non-White 14 (3.4) 10 (3.0) 4 (5.2)

BMIa (kg/m2)�25 56 (13.6) 41 (12.3) 15 (19.5)25.1–�30 194 (47.2) 167 (50.0) 27 (35.1)30.1–�35 125 (30.4) 102 (30.5) 23 (29.9)>35 36 (8.8) 24 (7.2) 12 (15.6)

Cigarette smokinga,c

Current 40 (9.7) 28 (8.4) 12 (15.6)Former 223 (54.3) 192 (57.5) 31 (40.3)Never 148 (36.0) 114 (34.1) 34 (44.1)

NSAID usea,d

Current 169 (41.1) 145 (43.4) 24 (31.2)Former 79 (19.2) 58 (17.4) 21 (27.3)Never 162 (39.4) 130 (38.9) 32 (41.6)

WHRb,e 409 [0.95 (0.91–0.99)] 332 [0.96 (0.93–1.00)] 77 [0.87 (0.81–0.91)]

aFrequency (%) of categorical variables.bMedian (interquartile range) for continuous variables.cCigarette smokingwasbasedonwhetherparticipants smokedonecigarette/day for 6monthsor longer, currently, in thepast, or never.dNSAID use was defined as having taken NSAID at least once a week for 6 months or longer; NSAID use history for 1 male participantwas missing.eTwo male participants had missing information on WHR.

Hardikar et al.

Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 Cancer Epidemiology, Biomarkers & Prevention2396

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

individuals in the lowest quartile for a biomarkerreceived a score of 0 and those in the highest quartilereceived a score of 3. Using the summed combined scoreas the primary predictor, we found that esophagealadenocarcinoma risk nonsignificantly increased by 2%per unit increase in the score in adjusted models (datanot shown).

DiscussionIn this prospective study, we observed that elevated

prediagnostic blood concentrations of CRP and possiblyIL6 are associated with subsequent increased incidenceof esophageal adenocarcinoma among persons withBarrett’s esophagus. Plasma levels of sTNF receptors orisoprostaneswere not statistically significantly associatedwith esophageal adenocarcinoma risk.

The role of inflammation and resulting oxidative stressin the development of cancer has been the focus of exten-sive research (3, 36, 37). More recently, inflammationmarkers, particularly CRP and IL6, have been reportedto be associatedwith all cancers in prospective and nestedcase–control studies. In a prospective Danish cohort of10,000 individuals, the risk for any cancer associated withCRP levels more than 3 mg/L was 1.3 (95% CI, 1.0–1.6;ref. 38); a similar 1.2-fold (95%CI, 1.10–1.32) increased riskfor any cancer with increasing CRP was observed in aGreek cohort (39). In another prospective study involvingthe Health Aging and Body Composition cohort, risk forall cancers increased with higher levels of CRP, IL6, andTNFa (40). In studies looking at individual cancer sites,the evidence for a possible role of chronic inflammation isthe strongest for colon cancer (38, 40–43). Elevated levels

Table 3. Relationship between the biomarkers and correlates of esophageal adenocarcinoma (n ¼ 411)

CRP IL6 sTNF-RI sTNF-RII Isoprostanes

Agea 0.08 0.18d 0.37d 0.43d �0.03WHRa 0.10d 0.08 0.13d 0.08 �0.12d

Cigarette pack-yearsb 0.24d 0.22d 0.22d 0.12d 0.07NSAID usec �0.15 0.21 0.01 0.04 �0.06CRPa

— 0.44d 0.18d 0.20d 0.22d

IL6a — — 0.21d 0.26d 0.08sTNF-RIa — — — 0.63d 0.12d

sTNF-RIIa — — — — 0.14d

Isoprostanesa — — — — —

NOTE: NSAIDs are coded as noncurrent vs. current. Non-smokers are coded as have smoked zero pack-years.aPearson correlation coefficient.bSpearman correlation coefficient.cDifference in biomarker (b coefficient) between current users and noncurrent users of NSAIDs.dP < 0.05.

Table 2. Distribution of mean biomarker levels overall and by gender in the SBES cohort (n ¼ 411)

All participants (n ¼ 411) Males (n ¼ 334) Females (n ¼ 77)

N Median IQR N Median IQR N Median IQR

CRPa (mg/L) 398 1.90 0.95–3.60 329 1.80 0.90–3.35 69 2.95 1.7–4.35IL6b (pg/mL) 407 1.88 1.34–3.06 332 1.77 1.31–2.87 75 2.62 1.48–3.79sTNF-receptor Ib (ng/mL) 407 1.39 1.14–1.65 332 1.42 1.15–1.66 75 1.31 1.08–1.61sTNF-receptor IIb (ng/mL) 407 5.35 4.71–6.44 332 5.34 4.68–6.40 75 5.49 4.74–6.62Isoprostanesc (pg/mL) 390 53 41–74 320 51 40–66 70 74 47–107

Abbreviation: N, number/frequency.aCRPwas notmeasured in 4 people due to exhausted baseline plasma. CRP levelsmore than 10mg/Lwere excluded as per an a priorihypothesis. Four people had plasma measured only at one time point and it was greater than 10 mg/L; 5 people had both CRPmeasurements greater than 10 mg/L.bIL6 and TNF receptors were not measured in 4 people due to exhausted baseline plasma samples.cIsoprostaneswere notmeasured in 16 people due to exhaustedbaseline plasma samples; 5 sampleswere not successfullymeasuredby the laboratory.

Chronic Inflammation and Risk of Esophageal Adenocarcinoma

www.aacrjournals.org Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 2397

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

Table 4. HRs and 95% CI for esophageal adenocarcinoma associated with markers of systemicinflammation (CRP, IL6, sTNF-RI, and sTNF-RII) and markers of oxidative stress (F2-isoprostanes) in theSBES cohort (n ¼ 397)

Biomarker

EAcases/total

UnadjustedHR (95% CI)

Adjusted for agea

and genderHR (95% CI)

Adjusted for agea,gender, WHRb,smokingc, and NSAIDused

HR (95% CI)

CRP (mg/L)e

All participantsMedianBelow 15/192 REF REF REFAbove 28/194 1.83 (0.98–3.43) 1.98 (1.05–3.73) 1.77 (0.93–3.37)

QuartilesQ1 (0.1–) 6/95 REF REF REFQ2 (0.9–) 9/97 1.29 (0.46–3.63) 1.18 (0.42–3.32) 1.05 (0.37–2.99)Q3 (1.9–) 15/95 2.35 (0.91–6.06) 2.29 (0.89–5.92) 2.12 (0.81–5.56)Q4 (3.6–) 13/99 1.90 (0.72–5.01) 2.06 (0.78–5.44) 1.55 (0.56–4.24)Continuous Ptrend

f 0.03 0.01 0.04MalesMedianBelow 12/155 REF REF REFAbove 28/165 2.31 (1.18–4.55) 2.21 (1.12–4.36) 1.93 (0.96–3.89)

QuartilesQ1 (0.1–) 5/79 REF REF REFQ2 (0.9–) 7/76 1.20 (0.38–3.77) 1.20 (0.38–3.79) 1.09 (0.34–3.49)Q3 (1.8–) 15/84 2.74 (0.99–7.55) 2.55 (0.92–7.02) 2.33 (0.83–6.51)Q4 (3.4–) 13/81 2.37 (0.85–6.66) 2.36 (0.84–6.62) 1.76 (0.60–5.15)Continuous Ptrend

f 0.01 0.01 0.05IL6 (pg/mL)All participantsMedianBelow 15/197 REF REF REFAbove 30/197 2.06 (1.11–3.82) 1.95 (1.03–3.72) 1.79 (0.93–3.43)

QuartilesQ1 (0.4–) 7/98 REF REF REFQ2 (1.3–) 8/99 1.14 (0.41–3.15) 0.95 (0.34–2.66) 0.82 (0.29–2.34)Q3 (1.9–) 19/99 2.74 (1.15–6.52) 2.35 (0.96–5.77) 1.93 (0.78–4.79)Q4 (3.1–) 11/98 1.65 (0.64–4.24) 1.40 (0.52–3.78) 1.17 (0.42–3.26)Continuous Ptrend

f 0.64 0.94 0.87MalesMedianBelow 10/161 REF REF REFAbove 31/161 3.19 (1.57–6.52) 2.85 (1.38–5.92) 2.52 (1.19–5.33)

QuartilesQ1 (0.4–) 6/81 REF REF REFQ2 (1.3–) 4/80 0.67 (0.19–2.36) 0.56 (0.16–2.01) 0.53 (0.15–1.89)Q3 (1.8–) 18/80 3.06 (1.21–7.71) 2.51 (0.96–6.51) 2.04 (0.77–5.41)Q4 (2.9–) 13/81 2.25 (0.86–5.93) 1.77 (0.64–4.86) 1.56 (0.55–4.43)Continuous Ptrend

f 0.66 0.99 0.81sTNF-I (ng/mL)All participantsMedianBelow 18/197 REF REF REFAbove 27/197 1.62 (0.89–2.93) 1.29 (0.69–2.42) 0.99 (0.51–1.92)

(Continued on the following page)

Hardikar et al.

Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 Cancer Epidemiology, Biomarkers & Prevention2398

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

Table 4. HRs and 95% CI for esophageal adenocarcinoma associated with markers of systemic inflam-mation (CRP, IL6, sTNF-RI, and sTNF-RII) and markers of oxidative stress (F2-isoprostanes) in the SBEScohort (n ¼ 397) (Cont'd )

Biomarker

EAcases/total

UnadjustedHR (95% CI)

Adjusted for agea

and genderHR (95% CI)

Adjusted for agea,gender, WHRb,smokingc, and NSAIDused

HR (95% CI)

QuartilesQ1 (0.3–) 11/98 REF REF REFQ2 (1.1–) 7/99 0.64 (0.25–1.64) 0.56 (0.22–1.45) 0.60 (0.23–1.56)Q3 (1.4–) 13/98 1.24 (0.55–2.76) 0.94 (0.41–2.15) 0.87 (0.38–1.98)Q4 (1.7–) 14/99 1.41 (0.64–3.10) 1.02 (0.44–2.37) 0.68 (0.27–1.68)Continuous Ptrend

f 0.18 0.68 0.69MalesMedianBelow 17/161 REF REF REFAbove 24/161 1.56 (0.84–2.90) 1.23 (0.63–2.41) 0.95 (0.47–1.94)

QuartilesQ1 (0.3–) 11/80 REF REF REFQ2 (1.1–) 6/81 0.51 (0.19–1.37) 0.47 (0.17–1.26) 0.51 (0.19–1.38)Q3 (1.4–) 12/80 1.10 (0.48–2.49) 0.85 (0.36–2.00) 0.73 (0.31–1.74)Q4 (1.7–) 12/81 1.23 (0.54–2.80) 0.88 (0.36–2.14) 0.66 (0.26–1.71)Continuous Ptrend

f 0.30 0.83 0.57sTNF-II (ng/mL)All participantsMedianBelow 15/197 REF REF REFAbove 30/197 2.23 (1.20–4.15) 1.90 (0.98–3.67) 1.78 (0.90–3.52)

QuartilesQ1 (1.6–) 8/98 REF REF REFQ2 (4.7–) 7/99 0.83 (0.30–2.29) 0.78 (0.28–2.17) 0.81 (0.29–2.25)Q3 (5.4–) 13/98 1.68 (0.70–4.05) 1.46 (0.59–3.61) 1.47 (0.59–3.70)Q4 (6.4–) 17/99 2.44 (1.05–5.66) 1.95 (0.76–4.95) 1.75 (0.68–4.49)Continuous Ptrend

f 0.22 0.75 0.86MalesMedianBelow 14/161 REF REF REFAbove 27/161 2.18 (1.14–4.16) 1.81 (0.91–3.62) 1.63 (0.80–3.34)

QuartilesQ1 (1.6–) 8/80 REF REF REFQ2 (4.7–) 6/81 0.69 (0.24–1.98) 0.62 (0.21–1.80) 0.69 (0.24–2.00)Q3 (5.3–) 12/80 1.52 (0.62–3.72) 1.27 (0.50–3.23) 1.28 (0.50–3.30)Q4 (6.4–) 15/81 2.17 (0.92–5.14) 1.59 (0.60–4.19) 1.44 (0.54–3.83)Continuous Ptrend

f 0.20 0.78 0.78Isoprostanes (pg/mL)All participantsMedianBelow 27/186 REF REF REFAbove 18/191 0.60 (0.33–1.09) 0.69 (0.38–1.26) 0.56 (0.30–1.03)

QuartilesQ1 (14–) 13/90 REF REF REFQ2 (41–) 14/96 0.98 (0.46–2.09) 1.18 (0.55–2.53) 0.93 (0.43–2.02)

(Continued on the following page)

Chronic Inflammation and Risk of Esophageal Adenocarcinoma

www.aacrjournals.org Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 2399

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

of inflammatory markers have also been shown to beassociated with cancers of the lung (38–40, 44, 45), breast(39, 40, 45) and ovary (46). Oxidative stress markers suchas F2-isoprostanes and oxodeoxyguanosine have beenshown to be associated with cancers of colon, lung, pros-tate, and breast (6–8, 47, 48).

Our study adds to the accumulating evidence for a keyrole of inflammation in esophageal adenocarcinomadevelopment, even among persons already diagnosedwith Barrett’s esophagus. Exposure of the esophagealepithelium to bile salts and acid resulting from gastro-esophageal reflux can cause chronic inflammation of thelower esophagus and result in increased release of proin-flammatory mediators (49–51), which may, in turn, causeDNAdamage and promote progression (1, 49). In supportof this hypothesis, an experimental study showed thatBarrett’s esophagus tissue secretes significant amounts ofIL6 resulting in an increased expression of STAT3 tran-scription factor that may lead to neoplastic conversion(35). In another study, the levels of ROS were higher inbiopsy specimens from patients with Barrett’s esophagusthan those from controls, suggesting that they play a rolein the tissue injury associated with Barrett’s esophagus(52). In our own cohort, we have previously shown thatleukocyte telomere length, a measure of person’s long-term inflammation level and oxidative damage (53), was

associated with more than a 3-fold increased risk ofesophageal adenocarcinoma (Ptrend ¼ 0.009; ref. 54), andthat anti-inflammatory drugs such as NSAIDs reduce therisk of esophageal adenocarcinoma even among thosewith dysplastic changes in their Barrett’s segment (15).Here, we show a link between prediagnostic concentra-tions of inflammation markers and esophageal adenocar-cinoma among patients with Barrett’s esophagus, sug-gesting that pathways involving inflammatory biomar-kers present prevention targets.

Blood-basedmarkers of chronic systemic inflammationmay also reflect systemic response to other exposures thatpredispose to esophageal cancer, such as obesity. Inflam-matory cytokines, including IL6 and TNF-a, have beenobserved to be systematically elevated in obesity (55, 56),and dietary intervention studies have shown that weightloss reduces CRP levels among obese individuals (57).Wehave previously reported amodest increase in esophagealadenocarcinoma risk associated with measures of centraladiposity (23). Results from thepresent study indicate thatCRP and IL6 may be predictive in esophageal adenocar-cinoma development even after adjustment for confound-ing effects of obesity in patients with Barrett’s esophagus.Taken together, these results suggest that inflammationmarkers may mediate the association of obesity withesophageal adenocarcinoma, but they also have some

Table 4. HRs and 95% CI for esophageal adenocarcinoma associated with markers of systemic inflam-mation (CRP, IL6, sTNF-RI, and sTNF-RII) and markers of oxidative stress (F2-isoprostanes) in the SBEScohort (n ¼ 397) (Cont'd )

Biomarker

EAcases/total

UnadjustedHR (95% CI)

Adjusted for agea

and genderHR (95% CI)

Adjusted for agea,gender, WHRb,smokingc, and NSAIDused

HR (95% CI)

Q3 (53–) 10/98 0.68 (0.30–1.54) 0.76 (0.33–1.74) 0.61 (0.26–1.41)Q4 (74–) 8/93 0.52 (0.22–1.26) 0.74 (0.30–1.85) 0.46 (0.18–1.20)Continuous Ptrend

f 0.34 0.92 0.41MalesMedianBelow 23/143 REF REF REFAbove 18/157 0.74 (0.40–1.36) 0.79 (0.42–1.46) 0.64 (0.34–1.20)

QuartilesQ1 (14–) 11/74 REF REF REFQ2 (40–) 12/79 1.03 (0.45–2.33) 1.18 (0.52–2.69) 0.98 (0.43–2.27)Q3 (51–) 11/80 0.96 (0.42–2.21) 1.04 (0.45–2.41) 0.80 (0.34–1.88)Q4 (66–) 7/77 0.55 (0.21–1.43) 0.66 (0.25–1.72) 0.47 (0.18–1.25)Continuous Ptrend

f 0.88 0.79 0.54

Abbreviation: EA, esophageal adenocarcinoma.aAge was modeled as a continuous variable.bWHR was modeled as a continuous variable.cCigarette smoking was modeled as pack-years smoked, never smokers were assigned a pack-year value of 0.dNSAID use was modeled as current vs. noncurrent users of NSAIDs at the baseline visit.eCRP measurements more than 10 mg/L were excluded from the analysis.fTest for trendwasbasedon the likelihood-ratio test associatedwith addition of the variable under consideration in its continuous form.

Hardikar et al.

Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 Cancer Epidemiology, Biomarkers & Prevention2400

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

independent effect on esophageal adenocarcinoma devel-opment beyond their effect through obesity.This study has several strengths. Its prospective design

allowed for measurement of multiple markers of inflam-mation and oxidative stress before the development ofcancer,minimizing thepossibility of reverse causality. Forthemajority of the biomarkers, we alsowere able to assessplasma levels at two time points during follow-up, thusreducing randommeasurement error and thepotential forregression dilution bias (58, 59). In addition, measuringthe biomarkers at two time points enabled us to improveon the ICCs reported earlier (Materials and Methods) bycapturing some of the intraperson variation. For example,the ICC of 0.55 for CRP that we reported earlier is actuallyimproved to 0.71 just by averaging over two CRP mea-surements (60). We also blinded the laboratory personnelto participants’ disease status. Comprehensive measure-ment of covariates such asWHR and pack-years of smok-ing enabled us to limit confounding. We also carried outanalyses restricting to the first 3 and 5 years of follow-up,so as to minimize the misclassification of inflammationmarker status due to prolonged duration between mea-surement of inflammation markers and occurrence ofesophageal adenocarcinoma events, and observed stron-ger associations and more pronounced trends.Our study is limited by the relatively small number of

incident esophageal adenocarcinoma cases, despite beingone of the largest, well-characterized cohorts of patientswith Barrett’s esophagus reported in the literature.Although we controlled for potential confounding effectsof smoking, obesity, and NSAID use in multivariableanalyses, we cannot exclude the possibility of residual

confounding by measured and unmeasured risk factors.In particular, we did not collect data onHelicobacter pyloristatus, which has been shown to be inversely related toesophageal adenocarcinoma risk (61, 62), while at thesame time being associated with systemic inflammation(63) such that H. pylori eradication therapies reduce theblood levels of proinflammatory cytokines, includingCRP (64).We did not adjust for confounding by the lengthof Barrett’s segment in our analysis. Although a recentstudy showed that presence of long-segment Barrett’sesophagus carried a 7-fold increased risk of progressionto esophageal adenocarcinoma (65), Barrett’s esophagussegment length is only modestly associated with the riskof esophageal adenocarcinoma in our cohort (66). Weattempted to limit measurement error by using highsensitivity and reliable assays for biomarker measure-ment. However, we cannot exclude the possibility oferrors in biomarker measurement resulting from degra-dation of biologic samples during storage. Finally, as thestudy cohort represents a specialty clinic, results pre-sented in the current report should be cautiously inter-preted in terms of their generalizability to the generalpopulation.

In conclusion, our results indicate that systemiclevels of CRP and, to some extent, IL6 are associatedwith progression to esophageal adenocarcinoma in per-sons with Barrett’s esophagus. Soluble TNF receptors andF2-isoprostanes were not found to be associated withincreased esophageal adenocarcinoma risk. Our findingswith CRP and IL6 are consistent with the literature thatsupports the role of chronic inflammation in the devel-opment of cancer, and esophageal adenocarcinoma in

A

B

CRP

IL6

10.005.00

2.001.000.50

0.200.10

Median Q2 Q3 Q4 Median Q2 Q3H

R, l

og s

cale

10.005.00

2.001.000.50

0.200.10

HR

, log

sca

le≤ 5 years≤ 3 years Entire FU

Q4 Median Q2 Q3 Q4

Median Q2 Q3 Q4 Median Q2 Q3≤ 5 years≤ 3 years Entire FU

Q4 Median Q2 Q3 Q4

Time trends in the relationships between circulating levels of biomarkers and risk of EACRP (A) and IL6 (B). FU, follow-up

Figure 1. Relationship betweenassociation of esophagealadenocarcinoma (EA) with plasmaCRP (A) and IL6 (B) with follow-uprestricted to 3 and 5 years frombaseline. All models are adjustedfor confounding effects of age,gender, smoking (pack-years), andobesity (WHR).

Chronic Inflammation and Risk of Esophageal Adenocarcinoma

www.aacrjournals.org Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 2401

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

particular. Additional analyses involving further follow-up of this and other cohorts are needed to confirm thesefindings, as well as to evaluate the utility of biomarkerassessment in clinical prediction and risk stratification ofesophageal adenocarcinoma.

Disclosure of Potential Conflicts of InterestNo potential conflicts of interest were disclosed.

DisclaimerThe funders had no role in the study design, data collection and

analysis, decision to publish, or preparation of the article.

Authors' ContributionsConception and design: S. Hardikar, M. Kratz, B.J. Reid, T.L. VaughanDevelopment of methodology: S. Hardikar, T.J. Montine, T.L. VaughanAcquisition of data (provided animals, acquired and managed patients,provided facilities, etc.): S. Hardikar, X. Song, A.M. Wilson, T.J. Montine,P.L. Blount, B.J. Reid, T.L. VaughanAnalysis and interpretation of data (e.g., statistical analysis, biostatis-tics, computational analysis): S. Hardikar, M. Kratz, G.L. Anderson,B.J. Reid, E. White, T.L. Vaughan

Writing, review, and/or revision of the manuscript: S. Hardikar,L. Onstad, X. Song, T.J. Montine, M. Kratz, G.L. Anderson, P.L. Blount,B.J. Reid, E. White, T.L. VaughanAdministrative, technical, or material support (i.e., reporting or orga-nizing data, constructing databases): S. Hardikar, L. Onstad, X. SongStudy supervision: X. Song, T.L. Vaughan

AcknowledgmentsThe authors thank Tricia Christopherson for projectmanagement; Terri

Watson for databasemanagement; and Christine Karlsen for coordinationof patient care.

Grant SupportThis work was supported by NIH grants P01CA091955 (to B.J. Reid),

K05CA124911 (to T.L. Vaughan), and R25CA094880 (to E. White).The costs of publication of this article were defrayed in part by the

payment of page charges. This article must therefore be hereby markedadvertisement in accordance with 18 U.S.C. Section 1734 solely to indicatethis fact.

Received April 15, 2014; revised June 25, 2014; accepted July 17, 2014;published OnlineFirst August 8, 2014.

References1. Coussens LM, Werb Z. Inflammation and cancer. Nature 2002;420:

860–7.2. Balkwill F, Mantovani A. Inflammation and cancer: back to Virchow?

Lancet 2001;357:539–45.3. Federico A, Morgillo F, Tuccillo C, Ciardiello F, Loguercio C. Chronic

inflammation and oxidative stress in human carcinogenesis. IntJ Cancer 2007;121:2381–6.

4. Klaunig JE, Wang Z, Pu X, Zhou S. Oxidative stress and oxidativedamage in chemical carcinogenesis. Toxicol Appl Pharmacol2011;254:86–99.

5. Hussain SP, Harris CC. Inflammation and cancer: an ancient link withnovel potentials. Int J Cancer 2007;121:2373–80.

6. Epplein M, Franke AA, Cooney RV, Morris JS, Wilkens LR, Good-man MT, et al. Association of plasma micronutrient levelsand urinary isoprostane with risk of lung cancer: the multiethniccohort study. Cancer Epidemiol Biomarkers Prev 2009;18:1962–70.

7. Rossner P Jr, Gammon MD, Terry MB, Agrawal M, Zhang FF, Teitel-baum SL, et al. Relationship between urinary 15-F2t-isoprostane and8-oxodeoxyguanosine levels andbreast cancer risk.Cancer EpidemiolBiomarkers Prev 2006;15:639–44.

8. Gill JK, Franke AA, Steven Morris J, Cooney RV, Wilkens LR, LeMarchand L, et al. Association of selenium, tocopherols, carote-noids, retinol, and 15-isoprostane F(2t) in serum or urine withprostate cancer risk: the multiethnic cohort. Cancer Causes Control2009;20:1161–71.

9. Triantafillidis JK, Nasioulas G, Kosmidis PA. Colorectal cancer andinflammatory bowel disease: epidemiology, risk factors, mechanismsof carcinogenesis and prevention strategies. Anticancer Res 2009;29:2727–37.

10. Szabo G, Lippai D. Molecular hepatic carcinogenesis: impact ofinflammation. Dig Dis 2012;30:243–8.

11. Bornschein J, Selgrad M, Warnecke M, Kuester D, Wex T, Malferthei-ner P. H. pylori infection is a key risk factor for proximal gastric cancer.Dig Dis Sci 2010;55:3124–31.

12. Reid BJ, Levine DS, Longton G, Blount PL, Rabinovitch PS. Predictorsof progression to cancer in Barrett's esophagus: baseline histologyand flow cytometry identify low- and high-risk patient subsets. Am JGastroenterol 2000;95:1669–76.

13. Reid BJ, Blount PL, Rubin CE, Levine DS, Haggitt RC, Rabinovitch PS.Flow-cytometric and histological progression to malignancy in Bar-rett's esophagus: prospective endoscopic surveillance of a cohort.Gastroenterology 1992;102:1212–9.

14. Reid BJ, Li X, Galipeau PC, Vaughan TL. Barrett's oesophagus andoesophageal adenocarcinoma: time for a new synthesis. Nat RevCancer 2010;10:87–101.

15. VaughanTL, Dong LM,Blount PL, AyubK,OdzeRD,SanchezCA, et al.Non-steroidal anti-inflammatory drugs and risk of neoplastic progres-sion in Barrett's oesophagus: a prospective study. Lancet Oncol2005;6:945–52.

16. Liao LM, Vaughan TL, Corley DA, Cook MB, Casson AG, Kamangar F,et al. Nonsteroidal anti-inflammatory drug use reduces risk of adeno-carcinomas of the esophagus and esophagogastric junction in apooled analysis. Gastroenterology 2012;142:442–52.

17. Bosetti C, Rosato V, Gallus S, Cuzick J, La Vecchia C. Aspirin andcancer risk: a quantitative review to 2011. Ann Oncol 2012;23:1403–15.

18. Cook MB, Chow WH, Devesa SS. Oesophageal cancer incidence inthe United States by race, sex, and histologic type, 1977–2005. BrJ Cancer 2009;101:855–9.

19. Oh DS, Demeester SR. Pathophysiology and treatment of Barrett'sesophagus. World J Gastroenterol 2010;16:3762–72.

20. Yousef F, Cardwell C, Cantwell MM,GalwayK, JohnstonBT,Murray L.The incidence of esophageal cancer and high-grade dysplasia inBarrett's esophagus: a systematic review and meta-analysis. AmJ Epidemiol 2008;168:237–49.

21. Bhat S, ColemanHG, Yousef F, Johnston BT,McManus DT, Gavin AT,et al. Risk of malignant progression in Barrett's esophagus patients:results from a large population-based study. J Natl Cancer Inst2011;103:1049–57.

22. Hvid-Jensen F, Pedersen L, Drewes AM, Sorensen HT, Funch-JensenP. Incidence of adenocarcinoma among patients with Barrett's esoph-agus. N Engl J Med 2011;365:1375–83.

23. Hardikar S, Onstad L, Blount PL, Odze RD, Reid BJ, Vaughan TL. Therole of tobacco, alcohol, and obesity in neoplastic progression toesophageal adenocarcinoma: a prospective study of Barrett's esoph-agus. PLoS ONE 2013;8:e52192.

24. Galipeau PC, Li X, Blount PL, Maley CC, Sanchez CA, Odze RD,et al. NSAIDs modulate CDKN2A, TP53, and DNA content risk forprogression to esophageal adenocarcinoma. PLoS Med 2007;4:e67.

25. Rabinovitch PS, Longton G, Blount PL, Levine DS, Reid BJ. Predictorsof progression in Barrett's esophagus III: baseline flow cytometricvariables. Am J Gastroenterol 2001;96:3071–83.

26. Hardikar S, Song X, Kratz M, Anderson GL, Blount PL, Reid BJ, et al.Intraindividual variability over time in plasma biomarkers of

Hardikar et al.

Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 Cancer Epidemiology, Biomarkers & Prevention2402

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

inflammation and effects of long-term storage. Cancer CausesControl2014;25:969–76.

27. Milatovic D, Montine TJ, Aschner M. Measurement of isoprostanes asmarkers of oxidative stress. Methods Mol Biol 2011;758:195–204.

28. Milne GL, Yin H, Brooks JD, Sanchez S, Jackson Roberts L II, MorrowJD.Quantification of F2-isoprostanes in biological fluids and tissues asa measure of oxidant stress. Methods Enzymol 2007;433:113–26.

29. MyersGL, Rifai N, Tracy RP, RobertsWL, Alexander RW, Biasucci LM,et al. CDC/AHA workshop on markers of inflammation and cardiovas-cular disease: application to clinical and public health practice: reportfrom the laboratory science discussion group. Circulation 2004;110:e545–9.

30. Pearson TA, Mensah GA, Alexander RW, Anderson JL, Cannon RO III,Criqui M, et al. Markers of inflammation and cardiovascular disease:application to clinical and public health practice: a statement forhealthcare professionals from the Centers for Disease Control andPrevention and the American Heart Association. Circulation 2003;107:499–511.

31. Kubo A, CookMB, ShaheenNJ, Vaughan TL,Whiteman DC,Murray L,et al. Sex-specific associations between body mass index, waistcircumference and the risk of Barrett's oesophagus: a pooled analysisfrom the international BEACON consortium. Gut 2013;62:1684–91.

32. Colbert LH, Visser M, Simonsick EM, Tracy RP, Newman AB, Kritch-evskySB, et al. Physical activity, exercise, and inflammatorymarkers inolder adults: findings from the Health, Aging and Body CompositionStudy. J Am Geriatr Soc 2004;52:1098–104.

33. Hauner H, Bender M, Haastert B, Hube F. Plasma concentrations ofsoluble TNF-alpha receptors in obese subjects. Int JObesRelatMetabDisord 1998;22:1239–43.

34. Olszanecka-Glinianowicz M, Zahorska-Markiewicz B, Janowska J,Zurakowski A. Serum concentrations of nitric oxide, tumor necrosisfactor (TNF)-alpha and TNF soluble receptors in women with over-weight and obesity. Metabolism 2004;53:1268–73.

35. Dvorakova K, Payne CM, Ramsey L, Holubec H, Sampliner R, Dom-inguez J, et al. Increased expression and secretion of interleukin-6 inpatients with Barrett's esophagus. Clin Cancer Res 2004;10:2020–8.

36. Kundu JK, Surh YJ. Inflammation: gearing the journey to cancer. MutatRes 2008;659:15–30.

37. Allavena P, Garlanda C, Borrello MG, Sica A, Mantovani A. Pathwaysconnecting inflammation and cancer. Curr Opin Genet Dev 2008;18:3–10.

38. Allin KH, Bojesen SE, Nordestgaard BG. Baseline C-reactive protein isassociated with incident cancer and survival in patients with cancer. JClin Oncol 2009;27:2217–24.

39. Trichopoulos D, Psaltopoulou T, Orfanos P, Trichopoulou A, BoffettaP. Plasma C-reactive protein and risk of cancer: a prospective studyfrom Greece. Cancer Epidemiol Biomarkers Prev 2006;15:381–4.

40. Il'yasova D, Colbert LH, Harris TB, Newman AB, Bauer DC, SatterfieldS, et al. Circulating levels of inflammatory markers and cancer risk inthe health aging and body composition cohort. Cancer EpidemiolBiomarkers Prev 2005;14:2413–8.

41. Erlinger TP, Platz EA, Rifai N, Helzlsouer KJ. C-reactive protein and therisk of incident colorectal cancer. JAMA 2004;291:585–90.

42. Gunter MJ, Stolzenberg-Solomon R, Cross AJ, Leitzmann MF, Wein-stein S, Wood RJ, et al. A prospective study of serum C-reactiveprotein andcolorectal cancer risk inmen.CancerRes 2006;66:2483–7.

43. Otani T, Iwasaki M, Sasazuki S, Inoue M, Tsugane S. Plasma C-reactive protein and risk of colorectal cancer in a nested case-controlstudy: Japan Public Health Center-based prospective study. CancerEpidemiol Biomarkers Prev 2006;15:690–5.

44. Pine SR,Mechanic LE, Enewold L, Chaturvedi AK, Katki HA, ZhengYL,et al. Increased levels of circulating interleukin 6, interleukin 8, C-reactive protein, and risk of lung cancer. J Natl Cancer Inst2011;103:1112–22.

45. Siemes C, Visser LE, Coebergh JW, Splinter TA, Witteman JC, Uitter-linden AG, et al. C-reactive protein levels, variation in the C-reactiveprotein gene, and cancer risk: the Rotterdam Study. J Clin Oncol2006;24:5216–22.

46. Poole EM, Lee IM, Ridker PM, Buring JE, Hankinson SE, Tworoger SS.Aprospective studyof circulatingC-reactiveprotein, interleukin-6, andtumor necrosis factor alpha receptor 2 levels and risk of ovarian cancer.Am J Epidemiol 2013;178:1256–64.

47. Babbs CF. Free radicals and the etiology of colon cancer. Free RadicBiol Med 1990;8:191–200.

48. Kong SY, Bostick RM, Flanders WD, McClellan WM, Thyagarajan B,Gross MD, et al. Oxidative balance score, colorectal adenoma, andmarkers of oxidative stress and inflammation. Cancer Epidemiol Bio-markers Prev 2014;23:545–54.

49. Abdel-Latif MM, Duggan S, Reynolds JV, Kelleher D. Inflammation andesophageal carcinogenesis. Curr Opin Pharmacol 2009;9:396–404.

50. Poehlmann A, Kuester D, Malfertheiner P, Guenther T, Roessner A.Inflammation and Barrett's carcinogenesis. Pathol Res Pract 2012;208:269–80.

51. Farhadi A, Fields J, Banan A, Keshavarzian A. Reactive oxygenspecies: are they involved in the pathogenesis of GERD, Barrett'sesophagus, and the latter's progression toward esophageal cancer?Am J Gastroenterol 2002;97:22–6.

52. Olyaee M, Sontag S, Salman W, Schnell T, Mobarhan S, Eiznhamer D,et al. Mucosal reactive oxygen species production in oesophagitis andBarrett's oesophagus. Gut 1995;37:168–73.

53. Houben JM, Moonen HJ, van Schooten FJ, Hageman GJ. Telomerelength assessment: biomarker of chronic oxidative stress? Free RadicBiol Med 2008;44:235–46.

54. Risques RA, Vaughan TL, Li X, Odze RD, Blount PL, Ayub K, et al.Leukocyte telomere length predicts cancer risk inBarrett's esophagus.Cancer Epidemiol Biomarkers Prev 2007;16:2649–55.

55. Yudkin JS. Adipose tissue, insulin action and vascular disease:inflammatory signals. Int J Obes Relat Metab Disord 2003;27 Suppl3:S25–8.

56. Yudkin JS, Stehouwer CD, Emeis JJ, Coppack SW. C-reactive proteinin healthy subjects: associations with obesity, insulin resistance, andendothelial dysfunction: a potential role for cytokines originating fromadipose tissue? Arterioscler Thromb Vasc Biol 1999;19:972–8.

57. Dietrich M, Jialal I. The effect of weight loss on a stable biomarker ofinflammation, C-reactive protein. Nutr Rev 2005;63:22–8.

58. Berglund L. Regression dilution bias: tools for correction methods andsample size calculation. Ups J Med Sci 2012;117:279–83.

59. Clarke R, Shipley M, Lewington S, Youngman L, Collins R, Marmot M,et al. Underestimation of risk associations due to regression dilution inlong-term follow-up of prospective studies. Am J Epidemiol1999;150:341–53.

60. White E, Armstrong BK, Saracci R. Priciples of exposuremeasurementin epidemiology:collecting, evaluating, and improving measures ofdisease risk factors. Great Britain: Oxford University Press; 2008.

61. Chow WH, Blaser MJ, Blot WJ, Gammon MD, Vaughan TL, Risch HA,et al. An inverse relation between cagAþ strains of Helicobacter pyloriinfection and risk of esophageal and gastric cardia adenocarcinoma.Cancer Res 1998;58:588–90.

62. Islami F, Kamangar F. Helicobacter pylori and esophageal cancer risk:a meta-analysis. Cancer Prev Res 2008;1:329–38.

63. Jackson L, Britton J, Lewis SA, McKeever TM, Atherton J, Fullerton D,et al. A population-based epidemiologic study of Helicobacter pyloriinfection and its association with systemic inflammation. Helicobacter2009;14:108–13.

64. Kebapcilar L, Bilgir O, Cetinkaya E, Akyol M, Bilgir F, Bozkaya G. Theeffect of Helicobacter pylori eradication on macrophage migrationinhibitory factor, C-reactive protein and fetuin-a levels. Clinics2010;65:799–802.

65. ColemanHG,Bhat SK,Murray LJ,McManusDT,O'Neill OM,GavinAT,et al. Symptoms and endoscopic features at barrett's esophagusdiagnosis: implications for neoplastic progression risk. Am J Gastro-enterol 2014;109:527–34.

66. Rudolph RE, Vaughan TL, Storer BE, Haggitt RC, Rabinovitch PS,Levine DS, et al. Effect of segment length on risk for neoplasticprogression in patients with Barrett esophagus. Ann Intern Med2000;132:612–20.

www.aacrjournals.org Cancer Epidemiol Biomarkers Prev; 23(11) November 2014 2403

Chronic Inflammation and Risk of Esophageal Adenocarcinoma

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384

2014;23:2393-2403. Published OnlineFirst August 8, 2014.Cancer Epidemiol Biomarkers Prev   Sheetal Hardikar, Lynn Onstad, Xiaoling Song, et al.   Adenocarcinoma Incidence in a Barrett's Esophagus CohortInflammation and Oxidative Stress Markers and Esophageal

  Updated version

  10.1158/1055-9965.EPI-14-0384doi:

Access the most recent version of this article at:

   

   

  Cited articles

  http://cebp.aacrjournals.org/content/23/11/2393.full.html#ref-list-1

This article cites 65 articles, 25 of which you can access for free at:

  Citing articles

  http://cebp.aacrjournals.org/content/23/11/2393.full.html#related-urls

This article has been cited by 1 HighWire-hosted articles. Access the articles at:

   

  E-mail alerts related to this article or journal.Sign up to receive free email-alerts

  Subscriptions

Reprints and

  .pubs@aacr.orgat

To order reprints of this article or to subscribe to the journal, contact the AACR Publications Department

  Permissions

  .permissions@aacr.org

To request permission to re-use all or part of this article, contact the AACR Publications Department at

on May 6, 2016. © 2014 American Association for Cancer Research. cebp.aacrjournals.org Downloaded from

Published OnlineFirst August 8, 2014; DOI: 10.1158/1055-9965.EPI-14-0384