Utility of circulating tumor DNA to detect minimal residual disease in colorectal cancer: A systematic review and network meta-analysis

Tung Hoang1.2 | Moon Ki Choi? | Jae Hwan \mathsf{o h}^{3} 1 Jeongseon Kim {\mathfrak{a}}_{\oplus}

1Department of Cancer Al&Digital Health, National Cancer CenterGraduateSchool of Cancer Science and Policy,Goyang-si, Gyeonggi-do,Republic of Korea

2Faculty of Pharmacy,University of Health Sciences,Vietnam National University,Ho Chi Minh City, Vietnam

3Center for Colorectal Cancer,National Cancer Center, Goyang-si, Gyeonggi-do,Republic of Korea

Correspondence

JeongseonKim,Department of Cancer AI& DigitalHealth,National CancerCenter Graduate School of Cancer Science and Policy, Goyang-si,Gyeonggi-do,RepublicofKorea. Email: jskim@ncc.re.kr

Funding information
National Cancer Center,Korea,Grant/Award
Number: 2310470

Abstract

Circulating tumor DNA (ctDNA) is a promising biomarker for predicting minimal residual disease (MRD)and guiding treatment decisions in patients with colorectal cancer (CRC). This study aimed to examine the study designs and settings of ongoing clinical trials that use ctDNA to guide treatment decisions and to determine the best timing for detecting MRD in non-metastatic CRC. We searched PubMed, Embase, Web of Science, Cochrane Library, and clinicaltrials.gov for English language records. The ctDNA settings from the clinical trials were categorized by randomization to ctDNA testing, treatment options based on ctDNA results,and the timing of ctDNA testing relative to adjuvant therapy.For prospective studies, a network meta-analysis using a frequentist approach was conducted to examine the pairwise associations between different ctDNA timing strategies and MRD,defined as recurrence,relapse, and progression. The main approaches in ctDNA-based interventional trial designs were categorized as ctDNA-guided treatment,ctDNA-by-treatment,ctDNA-guided surveillance, and ctDNA-enriched adjuvant therapy for guiding treatment decisions, including both escalation and de-escalation strategies, and surveillance. Overall, both preoperative and postoperative ctDNA detection were linked to higher risks of progression, with pooled hazard ratios ( 95% confidence intervals) of 5.23 (2.10-13.00) and 7.95 (5.30-11.91), respectively. Among the timing strategies, ctDNA testing after adjuvant therapy was the most effective for identifying high-risk patients, strongly suggesting the presence of residual disease. This study comprehensively reviewed the clinical settings of ctDNA testing in ongoing trials and provided evidence supporting the selection of post-adjuvant therapy as the optimal timing for ctDNA testing.

KEYWORDS

clinicaltrial,colorectalcancer,ctDNA,minimal residual disease,optimal timing

What's New?

Circulating tumor DNA (ctDNA) is a promising biomarker for predicting colorectal cancer (CRC) recurrence post-surgery.Thisreview examined therole of ctDNAin guiding treatment decisions and the timing of its use in detecting minimal residual disease in CRC across 26 ongoing clinical trials.Analyses of prospectivestudiesrevealed anassociationbetweenctDNA detectionfollowing CRC surgery and poor prognosis. The most effective timing for ctDNA testing in identifying patients at highrisk of relapse was afteradjuvant chemotherapy.Thefindings cast light on strategies to optimize ctDNA testing for informing personalized treatment approaches and assisting in riskstratification.

1 INTRODUCTION

AccordingtoGLOBOCAN2022,colorectalcancer(CRC)remainsthe secondleadingcauseofcancer-relateddeathsworldwide,withan estimated 903,859 deaths.1Globally,CRC survival rates improved in many countries during 2000-2014.23 This improvement might be attributed to both early detection and advances in treatment.4.5

Currently,surgicalresectionisthestandard ofcareforearly-stage CRC,with subsequent adjuvant chemotherapy (ACT） for up to 6 months.6The American Society of Clinical Oncology (ASCO) guidelinesrecommend adjuvantoxaliplatin-containingchemotherapywithin 3-6monthsforstageIllcolon cancerinboth thehigh-and low-risk subgroups.7In contrast,it is recommended for patients with high-risk stage Il colon cancer.8 In contrast, a meta-analysis in stage Ill CRC found thatdelayingACTbeyond8weeksaftercurativesurgerywas associated withreduced overallsurvival(relative risk[RR] =1.20 95% confidence interval [Cl] 1.15-1.26).9Nevertheless, an estimated 10%-30% of patients with stages Il and Ill CRC experience recurrence within 5 years after surgery,10.11 suggesting an uncertainty of who will benefitfromACTand theoptimal durationofACT.

Previous effortshavebeenmade toimproveCRCpatient outcomes.In a pooled analysis of 22 studies,a shortened duration of chemotherapymayreducesurvivalamongpatientswithstageIllcolon cancerwhowereprescribed monotherapybut nota combinationregimen.12 In another network meta-analysis of 30 trials, compared to capecitabineplus oxaliplatin therapy,none of the systemicregimens showed significant superiorityinterms of disease-free survival(DFS) over the past fewyears.13Therefore,it is important to identifybiomarkersthat canenhance theprognosticidentificationofhigh-risk patients andactas earlyindicators of adjuvanttreatment effectiveness.

To date, emerging data have supported the use of circulating tumorDNA(ctDNA)as apromisingbiomarker forpredicting recurrence in patients with CRC post-surgery,guiding ACT decisions,and should be tailored to support personalized treatment strategies.14The ctDNA,which primarily originates from programmed apoptosis or necrosis of tumor cells,was associated with tumor development.15 In CRC,tumor size and proliferative capacity are two key factors associated with ctDNA shedding.16 In a recent randomized controlled trial (RCT),ctDNA-guided management of stage Il colon cancer was associatedwithdecreasedACTusewithoutcompromisingrecurrence-free survival.17Pooled results from observational studies further indicate thatpost-operative ctDNAdetectionis stronglylinked topoorer progression-free survival.18 However, the optimal timing of ctDNA testing to detect minimalresidual disease(MRD)remains unclear.To furtherinvestigatetheefficacyofctDNA-guidedadjuvanttreatments acrossvarioussettings,wesystematicallysummarizedongoing and published RCTs examining this approach.In addition,we conducted a networkmeta-analysisto assess theefficacyofctDNAtestingatdifferenttimepointstoevaluateCRCoutcomes.

2 MATERIALSANDMETHODS

2.1|Study design and literature search

ThissystematicreviewwasconductedinaccordancewiththePreferredReportingItemsforSystematicReviewsandMeta-analyses (PRISMA) 2020 guidelines.19 The study protocol was registered in PROSPERO (CRD42024596279).

We searched PubMed，Embase,Web of Science， Cochrane Library,and clinicaltrials.gov databases for potential studies published on October 1,2024.Weperformeda combined searchfor ctDNA, chemoradiotherapy,resection, and CRC related terms.Accordingly, the following command was used for searching:“(ctDNA OR circulating tumor DNA OR circulating tumor DNA) AND (adjuvant OR neoadjuvantORchemotherapyORradiotherapyORchemoradiotherapyOR therapy OR treatment）AND(surgicalORsurgeryOR operative OR operation OR pre-operative OR preoperative OR post-operative ORpostoperative ORresectable ORresection)AND([colorectal OR colon ORrectal]AND[cancer OR carcinoma OR adenocarcinoma])” Wefurtherreviewed thebibliographies of relevant studies toidentify additional articlesrelated to thetopic.

2.2 | Eligibility criteria

Studies wereincluded inthissystematicreviewif (i)participantswere diagnosedwithCRC andunderwentacurative surgicalresection; (i) ctDNA was performed at any time points (for cohort studies) and after surgery only (for RCTs); (i) studies reported multivariableadjusted hazard ratios(HRs) and 95% Clsfortheassociationbetween detectable ctDNA levels andCRC post-operative outcomes(for cohort studies); and (iv)adjuvant treatments were included in both the intervention and comparison arms(for RCTs).

Studies were excluded if the manuscript was not published in English andfindingswereinvestigated inpatientswithunresectable CRC.We also removed reviews,case reports,case series, and casecontrolstudies inwhich the temporal relationshipbetween ctDNA testing and neoadjuvant therapy(NAT) andACT could notbeidentified.The publication year and sample size were not restricted.

2.3 |Studyscreening and data extraction

Thestudieswere screened bytwoindependentreviewers(TH andJK) according to the predetermined criteria.First,duplicate records from electronic databases were excluded using the EndnoteX9 program. Then,title and abstract screening was performed to select relevant studies,and the final selection was assessed by screening the full texts of title- and/or abstract-relevant articles.Any differences were discussed to reach an agreement.

Tworesearchers(TH and JK)independently extracted the data fromtheincluded studies.FortheRCTs,weobtainedgeneralinformation (trial name,country,and sample size) and the PICOS question (study population, ctDNA assay,trial intervention,trial comparison, endpoint,and study setting).For cohort studies,we obtained data on the study characteristics (author, publication year, country, sample size,and follow-up duration),patient characteristics (age, sex, and stage), ctDNA sequencing arrays, ctDNA approach (tumor-informed or tumor agnostic),NAT/ACT，ctDNA sampling time,and adjusted covariates.Anydisagreementsbetweenthereviewerswereresolved by discussion to reach a consensus.

2.4 | Quality assessment

Thequalityofcohortstudiesincluded inthefinalanalysiswasevaluated by using the Newcastle-Ottawa Scale (NOs).20 The assessment toolincludesatotalofeightitemsfortheselectionofstudygroups (fouritems),comparability of groups(oneitem),and ascertainment of the outcome of interest(three items),witha maximumof nine scores.20

2.5 Statistical analysis

We extracted HRs and 95% Cls from the cohort studies.Heterogeneity across studies was measured using the Higgins I^{2} metric,for which an I^{2} value greater than 50% implies substantial heterogeneity.21 By assumingthatallstudieswereconductedonthestudypopulation withdifferentcharacteristics,logarithm-transformed HRswerepooled using a random-effects model22 for direct evidence of the overall associationbetweendetectable ctDNA andCRCprognosis.Inaddition,we tested publication bias using Begg and Egger tests and visualized with funnel plots.2324 Furthermore, to compare the efficacy of ctDNA tests at differenttime points,weperformed a network metaanalysisusingthefrequentistapproachtocombine direct and indirect evidence.25

All statistical analyses were performed using STATA (version 18.0 StataStatistical Software,CollegeStation,TX)and R(version4.0.2,R

Foundation for Statistical Computing, Vienna，Austria). Package “"metan”(in STATA)and package“netmeta”(in R)were used for the meta-analysis andnetworkmeta-analysis,respectively.

3 RESULTS

3.1|Study characteristics

Figure 1 shows thePRISMA flow diagram for study selection.A total of 3763 records fromPubMed，Embase，Web of Science，and clinicaltrials.govwererelatedtothetopic.Weremoved1820records owing to duplicates.After title and abstract screening, seeking retrieval, and assessing the eligibility criteria of thefull texts,we ultimately included 26 clinical trials and 34 cohorts and post-analyses on the utility ofctDNAin detectingMRD.

3.2|Clinical trials on the utility of ctDNA in detectingminimalresidualdisease

The main approaches of ctDNA-based interventional trial designs for identifying MRDaredisplayed inFigure2A-D.The trialsettings for each approach are summarized inTable 1. In the ctDNA-guided treatmentapproach，thetreatmentoutcomes ofpatientsinthe ctDNA-guidedarmwerecomparedwiththoseinthecontrolarm of thestandard treatment.Withthis approach,patients in the ctDNAguided arm are assigned to thecurrent standard of care,an escalation orde-escalationtreatmentdependingonthepositiveornegativefinding of ctDNA testing (Figure 2A). For example, in the TRACC Part C trial,patientswererandomizedintothectDNA testinggroup and the standardofcaregroupwitha3-monthCAPOXstrategy.Basedon ctDNA testing results,ctDNA-positive patients received ACT, whereasctDNA-negativepatientsreceivedde-escalatedtreatmentor escalatedtreatmentifctDNAbecamepositiveafter3monthsof ACT.26,27

In the ctDNA-by-treatment approach,ctDNA testing was performed in allpatients,and different therapeutic options were compared in bothctDNA-positive andctDNA-negative patients (Figure 2B).For example,in the CIRCULATE-US trial,after a ctDNA testwas performed in allpatients,those with positive ctDNA were randomized toeitherFOLFOX6/CAPEOXorFOLFIRINOX,whereas thosewithnegativectDNAwererandomized toeitherFOLFOX6/ CAPEOX or serial ctDNA.If subsequent ctDNA tests were positive, patientswererandomizedtoeitherFOLFOX6/CAPEOXor FOLFIRINOX.41.42

In the ctDNA-guided surveillance approach, patients were randomlyassignedtoundergoeitherctDNAtestingorstandardsurveillance,with the outcome of the ctDNA-positive group compared to those of the standard surveillance (Figure 2C).For example, in the IMPROVE-IT2 trial, patients were randomized to receive a dynamic ctDNA test every 4 monthsfor 2 years or a standard surveillance of computed tomographyscans at12and36monthspostoperativeand colonoscopyevery5yearsuntiltheageof75years.Patientswith positive ctDNA were further subjected to PET-CT scans every 3 months, whereas patients with negative ctDNA still followed the standard surveillance.5556

In the ctDNA-enriched adjuvant therapy approach, following ctDNA testing results afterfirst-line ACT,ctDNA-positive patients are randomly assigned toeitherreceiveadditionalsecond-lineACTor undergo watchful waiting (Figure 2D).For example,inboth the ALTAIR and ERASE-CRC Part Il trials, post-ACT ctDNA-positive patientswererandomizedtoeitheroraltrifluridine/tipiracilfor 6 months or placebo/observation.48,53.54

Table 1furthersummarizes ongoing trialsby the use of ctDNA as a surrogate of MRD in the adjuvant and surveillance setting.The role of ctDNA in guiding surveillance during thefollow-upwas examined in some studies.Forexample,the INFORM trial randomized patients with stageIl and high-riskstageIlCRC toreceiveroutinefollow-upwithorwithout ctDNAmonitoring afterradical resectionto assessthe sensitivity of postoperative ctDNA methylation indetectingrecurrence.In theIMPROVEIT2 trial,postoperative outcomesforthe ctDNA-guided surveillance strategywere compared with standard surveillance usingchest,abdomen,and pelviscomputed tomographyscanstoassesswhetherctDNA-guided surveillanceincreasestheproportionofpatientsundergoingcurativeresection or treatments targeting local metastases556

In addition,severalstudieshaveexplored thevalueof ctDNAin guiding treatment decisions as well as both escalation and de-escalation strategies. In the CIRCULATE-US trial,patients with stage lIl colon cancer with negative postoperativectDNAwererandomizedtoeitherimmediateFOLFOX6/ CAPOXtreatment or serial ctDNA monitoringwith delayed ACT,andDFS was compared.42In contrast,patients with eitherinitially or subsequently positive ctDNA wererandomized toreceive either FOLFOX6/CAPOX or FOLFIRINOX, and DFS was similarly compared.42 In the DYNAMIC trial, patientswithstagellcoloncancer wererandomizedtoreceive eithera ctDNA-guided therapeutic strategy or standard care.17 Based on ctDNA results,the ctDNA-guidedgroupwasfurtherevaluated fortreatment escalation or de-escalation, ranging from no chemotherapy to 6 months of 5-fluorouracil/capecitabine,or3-6months of FOLFOX/CAPOX.17

Several trialshavefocused on thenon-inferiority of ctDNA-guided de-escalationandthesuperiorityofctDNA-guidedescalationstrategies compared to standard care.Forinstance,intheVEGA trial,3-year DFS was compared betweenpatientswithnegativepostoperative ctDNA whoreceivedde-intensified,no-ACTtreatment andthosewhoreceived standard ACT with CAPOXfor 3 months.53 Conversely,in the PRODIGE 70 trial, 3-year DFS was compared between patients with positive postoperative ctDNA who received intensified mFOLFOX6 for 12 cycles and those receiving standard ACT.39

FIGUREClinicaltrialdesignsfortheutityofcirculatingtuorDNA(ctDNAtestinin(A)ctNA-guidedtreatentoptionapproac (B)ctDNA-by-treatment aproach,C)cDNA-gudedsurvellanceapproach,D)ctDNA-enrichedadjuvant therayapproac

Theutility of ctDNAhas alsobeen evaluated toguidepostadjuvant treatments.In the ALTAIR trial,DFSwas assessed in patients withpositivectDNAwhoreceived6monthsoftrifluridine/tipiracil following standard ACT with CAPOX compared to those receiving a placebo.53Similarly,in the ERASE-CRC trial Part Il,the efficacy of the trifluridine/tipiracilregimenwasinvestigatedinctDNA-positive patients after receiving fluoropyrimidine and oxaliplatin-based ACT, compared to observation alone.48

3.3 |Prospective analyses on the utility of ctDNA indetectingminimalresidualdisease

Table 2 presents prospective studies examining the clinical utility of ctDNA in detecting MRD.The34 studies encompassed atotal of6941

CRC patients, with ages ranging from 19 to 94 years.17.54,59-67,69-71,73-

84.86-90The ctDNAlvels ere assessed at multipe tme points, inclding baseline,before,during,and afterAT,aswellaspre-and postoperative, pre-and post-ACT, and through longitudinal or serial surveillance testing. The timing of ctDNA assessment varied across studies;forexample,postoperativectDNAtestingrangedfromasearlyas5daystoaslateas 12 weeks after surgery,while some studies did not specify the exact timingof assessment.MRDoutcomes include recurrence-free survival, relapse-free survival,DFS,and progression-free survival,as wellas overall survival,providingkey metrics for assessingpatient prognosis and monitoring cancer progression.Of the 34 studies, 32 were full-text studies, while 2 were conference abstracts.Among them, 12 studies used a tumor-informed ctDNA approach,and 20 studies used a tumor-agnostic approach.

Study quality was evaluated for 32 full-text articles (Table S1). All studiesmetthecriteriaforselectionandoutcome assessmentonthe NOS;however,they varied in cohort comparability based on study design or analysis.Fourteen studies conducted multivariable analyses adjusting for key confounders, earning a score of 1.1760-4.7073-75,77982.83 Six wentfurtherbycontrollingforadditionalfactorsinsubgrouporinteraction analyses, receiving 2 points5,697684,86.89 whereas others dd nt adjustforanyvariables orrelied solely onKaplan-Meier analyses,resulting inncre96568889calculate the ld estma the associationbetween ctDNA positivity and MRD,we stratified the analysis intobothpreoperative and postoperative ctDNA.For studies withmultiple time pointsbefore or aftersurgery,we used theeffectsize closest tothe surgery date inpooled analyses.Overall,bothpreoperative andpostoperativectDNAdetectionwereassociatedwithincreasedrisks of MRD progression, with pooled HRs 95% CIs) of 5.23 (2.10-13.00) and 7.95 (5.30-11.91),respectively (Figure S1). There was no evidence of publicationbias usingEgger's test (p=.083) and Begg's test \left(p=514\right) (Figure S2).

Three-yearDFSsurvivalratesdidnotsignificantlydifferbetween the ctDNA-positive and ctDNA-negative groups forbothpreoperative (pooled \mathsf{R R}=0.82 ， 95%\mathsf{C l}=0.64\ –1.04 three studies) and postoperative (pooled \mathsf{H R}=0.73 95%{\mathsf{C l}}=0.50{-}1.06 ，two studies)ctDNA detection(FigureS3A).There was also noevidence ofpublication bias, as evidenced by Egger's test (p=.097) and Begg's test

\left({p=.086}\right) (Figure S4).Additionally, ctDNA-positive status was associatedwithahigherriskofmortalitycomparedtoctDNA-negativestatus, with HRs ( 95% Cls) for overall survival of 7.25 (1.18-44.39) for preoperative and 2.71 (1.20-6.12) for postoperative ctDNA detection (FigureS3B).However,thisestimate exhibited substantial publication bias, as indicated by Egger's test (p<.001) and Begg's test (p=.009) (Figure S5).

Regarding ctDNA timing strategies,Figure S6 illustrates the network geometry of current studies examining various ctDNA testing strategies comparedwithwatchfulwaitingwithstandardcare.Most comparisons evaluated MRD progression through postoperative ctDNAdetection(25 comparisons),followed by serial surveillance (10 comparisons), post-ACT (8 comparisons), and post-NAT or preoperativeassessments (7comparisons).Fewer comparisons have assessed MRDdevelopment risk relative to standard care,including ctDNA detection pre-NAT (two comparisons) and during-NAT (one comparison),in comparison to standard care.

Detection of ctDNA after NAT, postoperatively,after ACT,and during longitudinal surveillance was associated with a poorerprognosis compared to cases inwhichctDNA testing wasnotperformed, with HRs( 95% Cls) of 4.02 (1.81-8.91), 8.57 (5.63-13.1), 18.7 (8.51- 41.0), and 17.6 (8.88-34.8),respectively.Additionally,positive ctDNA detectedafterACTandduringlongitudinalsurveillancewasfurther associatedwithahigherincidenceofMRDthanctDNAdetected beforeorafterNAT.PostoperativectDNAdetectionwasfurther associatedwithahigherincidenceofMRDthanctDNAdetected before NAT(Table3).However,significantlyhighheterogeneitywas observed, with an I^{2} value of 82.3% \therefore3%(95%001=77.1%-86.2%) .Among thevarious ctDNAtestingtiming strategies,post-ACTtestinghad the highest surface under the cumulativeranking curve(SUCRA)value (90.87%) ,followed by longitudinal surveillance (89.39%) and postoperativectDNAtesting (64.55%) .Thissuggests that ctDNAdetection after ACT may be a strong indicator of MRD.

4 DISCUSSION

In this systematicreview,we summarized ongoing clinical trialsthat use ctDNA to stratifypatientsintolow-and high-risk groups to informtreatmentdecisions，includingACTescalationanddeescalation.Ourpooled analyses ofprospective studiesshowedthat ctDNA detection following CRC surgery was associated with a poor prognosis, indicating an increased risk of disease relapse and recurrence.Among thetiming strategies,ctDNA testingafterACTwas the most effectiveforidentifyinghigh-riskpatients,strongly suggesting thepresence of residual disease or a higher likelihood of relapse.

One of the main concerns regarding MRD detection is the sensitivity of ctDNA assays in the postoperative setting,which may contribute to significant heterogeneity in our estimates.In a pooled analysis of 485 stages Ill and Il CRC patients across the three cohorts, false-positive results immediately following surgery were more frequentinpatientswholaterhadlocoregionalrelapse than inthose who developed distant relapse.91 Despite improvements in ctDNA assays, which can detect levels as low as 0.01% false negatives may still arise because of biological factors such as low DNA-shedding tumors,mucinous histology， and the anatomical location of hidden micrometastatic disease.92

Notably，other emerging circulating biomarkers, such as DNA methylationand microRNA,areunderdevelopment and may help address or enhance the analytical sensitivitylimitations of mutationbased ctDNA testing.9394 Compared with conventional targeted amplicon sequencing, next-generation sequencing (NGS) offers greater accuracy andprecisioninidentifying andquantifyingmutations.14 However, these advanced tests are relatively expensive, involve complex analysis procedures,and have longer turnaround times.14

Furthermore,heterogeneityexistsbetweenctDNAdetermination approaches.The tumor-informed approach,which detects mutations in tumor tissue andtracks theminplasma,demonstratesgreatersensitivity than the tumor-agnostic method, which detects mutations in the plasma.14 Additionally, the tumor-informed method requires prior tissuesequencingandisthusmorecostlythanthetumor-agnostic method.95 In contrast, tumor-agnostic approaches rely solely on plasma analysis,making themmorewidely applicablebutgenerally less sensitive.95 Nevertheless, according to the European Society for Medical Oncology, no single ctDNA assay can be used for all purposes. 14.96

Thefindings fromourpooled analysis on theuse ofctDNAfor tracking MRD align with those of previous meta-analyses.18.97 Our currentstudyfurthercontributestotheevidenceontheoptimaltimingforbloodsamplecollectiontodetect MRD.Surgicaltrauma has beenintroducedtoincreasethereleaseof totalcell-freeDNAand wild-type DNA into the plasma, which can obscure ctDNA detection.98 Therefore,blood samples 6-8 weeks after surgery to allow for the initiation of ACT,9 which is consistent with our finding that ctDNA post-ACTis thebest strategy.However,pairwise comparisons between post-ACTand postoperative strategies werenot significantly different. In the present study,serial ctDNA testing during ongoing surveillanceshowedpromisingpotentialfordetectingMRDbeyond post-ACTstrategies.Thisapproachcanenhanceearlydetectioncapabilities and improve patient outcomes.However,therepeated nature of ctDNA testing demands substantial financial resources,and the cost-effectiveness of serialtestinghasnotbeenthoroughlyestablished.Moreevidence isneededtodeterminewhether thebenefits justifythe associatedcosts,whichcould affectthefeasibility ofincorporating this method into routine clinical practice.

Economicmodellingstudieshavefurther explored thefinancial impactofctDNA-guidedACT.Toet al.evaluatedpostoperative ctDNA use in patients with stage Il CRC and found that reserving ACTfor ctDNA-detectablepatients resulted ina 13% reduction in ACT prescriptions compared to standard care.100101 Assuming a 50% adoptionrate of tumor-informed ctDNA testingin thefirstpostoperative year,ctDNA-guided ACT yielded an estimated total cost savings of $$221,684$ for commercial payers and $$123,4520for Medicare Advantage payers.102 Similarly, the GALAXY study, from a US healthcareperspective,indicated that combining ctDNA testingwithhighrisk clinical featuresimproved quality-adjusted lifeyears(QALYs)by 0.7 and saved9771 per patient.103 However, another model for stageIlcoloncancerfound thatctDNAcombinedwithcurrentguidelines (using pT4 and pMMR markers)for ACT selection was not costeffective.104 The incremental cost-effectiveness ratio was E67413 per QALY, surpassing the E50,000 per QALY willingness-to-pay threshold.104 In the UK, ctDNA-guided ACT showed greater costeffectiveness in high-risk stage Il patients than in stageIll patients in the short term,whereas long-term cost-effectiveness for stage Ill patients remained uncertain due to similar health outcomes but notable cost savings.105

In this study,we conducteda comprehensivereview ofcurrent studydesigns and applicationsofctDNAtoguidetreatmentdecisions. Asmosttrials are still ongoing,futureresearchis expected toprovide comparativeevidence ontheeffectivenessofstandard-of-caretreatments versus ctDNA-guided approaches.These studies will be crucial inevaluatingtheclinicalutilityofctDNAinguidingtreatmentdecisions,optimizing therapeuticstrategies,and identifyingpatientpopulations that maybenefitmostfromctDNA-informedinterventions. Additionally,theywill shed lightontheimpactof treatmentescalation orde-escalationstrategies,helpingtorefinepersonalizedtreatment decisions，minimize unnecessary toxicity，and ultimately improve patientoutcomes.Robustprospectiverandomized trialswillbe essentialtoestablishstandardizedguidelines and integrate ctDNA-based managementintoroutine clinicalpractice.

Drawing on prospective studies, our findings build on prior researchbyexploringthecomparativeeffectsofvarioustiming strategiesforctDNA testing.Given that ctDNAwas tested ata singletime point inmost studies,patients who experiencedctDNA clearance-converting from ctDNA-positive to negative during chemotherapy-mayindicateabetter treatmentresponse.Asa result,ctDNA-positivepatientsbeforeACTmighthavemorefavorableoutcomescomparedtothosewhoremainedpositiveafterACT, andpatientswithpersistentctDNApost-treatment mayhavea higherriskofrecurrence,contributingtotheobserved differencesin survival.Amongindividual studies,onlyKhakooet al.examined the associationbetweenpersistentlydetectablectDNAandmetastasisfree survival,reporting HRs 95% Cls) of 3.8 (1.2-11.7)for persistent ctDNA before and during NAT and 11.5(3.3-40.4)for persistence before,during,and after NAT,compared withpatients with undetectable or nonpersistent ctDNA.68 However, this review may not havefullyaccountedforleadtimeeffectsandthedynamicsof ctDNA conversion,which could introduce variability in the findings. FuturestudieswithstandardizedmonitoringprotocolsandlongitudinalctDNA assessments willbe necessarytobetter understand thesefactors.Due to theheterogeneity of the existing studies, future research should aimtovalidate thesefindingsby standardizing ctDNA analysis methods and cutoff thresholds,optimizing ctDNA collection timing,incorporating long-term follow-up data, andexaminingassociationswithinthecontextofepidemiological relationships.Theeffect of eachctDNA test and platform needsfurther validation in clinical trials,as sensitivity and specificity can vary despite technological similarities.

In summary,our combined analysis of prospective studies indicated that thedetectionofctDNA after CRCsurgeryisassociated with a poor prognosis,suggesting ahigher risk of MRD.Among the varioustimingstrategies,ctDNA testing conducted afterACTproved most effectiveinidentifyinghigh-riskpatients,stronglyimplying the presenceofrecurrenceoranelevatedchanceofrelapse.Future research should focus onstandardizing ctDNAtesting protocols, includingtheoptimaltimingandfrequencyforpersonalizedtreatment strategies,andrefiningriskstratificationtoimprovepatientoutcomes.

AUTHORCONTRIBUTIONS

TungHoang:Conceptualization;formal analysis;methodology;visualization;writing-original draft.MoonKiChoi:Conceptualization;data curation; methodology; writing - review and editing. Jae Hwan Oh: Conceptualization; data curation;methodology; visualization.JeongseonKim:Conceptualization;data curation;methodology;validation; supervision.

CONFLICTOFINTERESTSTATEMENT

The authors declare no competing interests.

DATAAVAILABILITYSTATEMENT

Thedatathatsupportthefindingsof thisstudyareavailablefromthe correspondingauthoruponreasonablerequest.

ETHICSSTATEMENT

This study utilized findings from published articles,thereby exempting it from requiring ethical approval.

ORCID

JeongseonKim \oplus https://orcid.org/0000-0002-0889-2686

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How to cite this article:HoangT,Choi MK,Oh JH,Kim J. UtilityofcirculatingtumorDNAtodetectminimalresidual diseaseincolorectalcancer:Asystematicreviewandnetwork meta-analysis.Int J Cancer.2025;1-19.doi:10.1002/ijc.35442