Liquid biopsies offer the possibility to evaluate cancer patients using noninvasive approaches. Circulating cell-free DNA (ccfDNA) is 1 of the most used and promising sources. Detecting tumor DNA among ccfDNA (ctDNA) can be used for early cancer detection, treatment response assessment, prognosis, and predictive evaluations. Providing analyses that can increase the quality of patient treatment is very much a joint effort between laboratory scientists and clinicians. With its use approaching clinical practice, it is important for clinicians to be familiar with the basic concepts and analyses behind ctDNA results in a similar way as laboratory scientists should have knowledge of the clinical needs to provide relevant analyses. In this Perspective, we describe the whole process of ctDNA analyses, from the preanalytical standards to reporting/analyzing results, and highlight some important factors that need to be addressed in the process of implementing them to clinical practice.

Multiple clinical trials are investigating ctDNA to guide adjuvant chemotherapy (ACT) in colorectal cancer. Timely ACT initiation necessitates early ctDNA testing, but the impact of postoperative cell-free DNA (cfDNA) and ctDNA dynamics remains unclear, particularly with cost-reducing input caps employed in some assays. This study investigates ctDNA detection at day 14 versus day 30, comparing whole-sample analysis with capping the cfDNA input, and evaluates single and dual timepoint assessments for ACT allocation.

From 2019 to 2023, 611 patients with stage I to III colorectal cancer were enrolled. Blood was collected preoperatively and postoperatively on ∼day 14 and ∼day 30. The cfDNA levels were assessed using digital PCR, and ctDNA was assessed using tumor-informed digital PCR or targeted sequencing analyzing all cfDNA from 8 mL of plasma.

Despite elevated cfDNA in 85% of day 14 samples, performance was comparable between the two timepoints (sensitivity, 31% vs. 32% and specificity, both 98%). A 50-ng cfDNA input cap reduced ctDNA detection probability, affecting 78% of day 14 samples and 65% of day 30 samples. At both timepoints, ctDNA detection was prognostic of recurrence (day 14; HR, 9.0, 95% confidence interval, 5.5-14.8 and day 30: HR, 12.5, 95% confidence interval, 7.6-20.4). In 74% of ctDNA-positive recurrence patients, both samples had ctDNA detected. An increase in the ctDNA level from day 14 to day 30 was associated with a shorter time to recurrence (Pearson R = -0.63, P = 0.003). Combining the timepoints would increase sensitivity (36%) and allow earlier ACT start in 80% of patients.

Early ctDNA sampling is feasible and highly prognostic. Supplemental later testing may improve sensitivity while allowing early ACT initiation for most ctDNA-positive patients.

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.

Resectable non-small cell lung cancer (NSCLC) has a relatively poor prognosis owing to the risk of developing local or distant metastatic recurrence, even at stage I. To overcome the high recurrence rate, perioperative therapies have been rapidly developed through the combination of existing cytotoxic chemotherapies with immune checkpoint inhibitors (ICIs) and molecular targeted therapies. These new therapeutic strategies have significantly improved the prognosis of patients with stage II-III NSCLC and have been approved for clinical use. However, new challenges have emerged in the selection of the optimal perioperative treatment in clinical practice. First, it is currently difficult to determine which perioperative treatment is superior, preoperative or postoperative. Additionally, since surgery alone is curative in some patients, the addition of anticancer agents such as ICIs raises concerns regarding toxicity, as serious side effects during preoperative treatment may lead to an inability to perform the surgery itself. Moreover, because various perioperative treatments are still being developed, treatment options for perioperative care are expected to increase soon. To summarize the increasingly complex perioperative treatment of resectable NSCLC, this review provides a comprehensive summary of the clinical efficacies of current perioperative therapies and future directions based on basic background, patient selection, ongoing trials, and enhancing immunotherapy.

The Liquid Biopsy (LB) represents an ideal surrogate of tumor Tissue Biopsy (TB) when the aim is to obtain useful information on patient prognosis and personalized therapy. This technique renders it possible to isolate circulating tumor cells, circulating tumor DNA and other molecules from biological fluids. The most commonly used fluid for liquid biopsy is blood, but depending on the case it could be necessary to isolate the tumor components from other biological fluids such as urine, pleural effusion, cerebrospinal fluid, and others. The main advantages of liquid biopsy are the minimally invasive nature of the procedure and the possibility of analyzing all tumor clones. Limitations include difficulties in the isolation of tumor components and the requirement for highly sensitive analysis methods to avoid the risk of technical artifacts. In our review we will focus on describing circulating tumor biomarkers to illustrate the variety of information that can be obtained from biological fluids, particularly blood. We will then discuss the advanced biotechnological techniques suitable for the identification and analysis of Circulating Tumor DNA (ctDNA), examining both the potential and limitations of analytical methods and the clinical applicability of liquid biopsy for cancer diagnosis, monitoring, and therapeutic prediction. Additionally, we will explore strategies to enhance this valuable alternative to the more invasive tissue biopsy, with a dedicated focus on ongoing clinical studies, currently approved tests, and guideline recommendations.

Although the 5-year relative survival rates for resectable solid tumors have improved over the past few years, the risk of postoperative recurrence necessitates effective monitoring strategies. Recent advancements in molecular residual disease (MRD) testing based on circulating tumor DNA (ctDNA) analysis have shown considerable promise in the context of predicting recurrence; however, significant barriers to widespread clinical implementation remain-mainly, low awareness among healthcare professionals, high costs, and lack of standardized assays and comprehensive evidence. This position paper, led by the Japan Society of Clinical Oncology, aims to establish a common framework for the appropriate clinical use of MRD testing in a tumor type-agnostic manner. It synthesizes currently available evidence, reviews region-specific clinical trends, addresses critical clinical questions related to MRD testing, and offers recommendations to guide healthcare professionals, biotechnology and pharmaceutical companies, and regulatory authorities. These recommendations were developed based on a voting process involving 15 expert members, ensuring a consensus-driven approach. These findings underscore the importance of collaborative efforts among various stakeholders in enhancing the clinical utility of MRD testing. This project aimed to foster consensus and provide clear guidelines to support the advancement of precision medicine in oncology and improve patient outcomes in the context of perioperative care.

In the practice of colorectal cancer (CRC), traditional tumor tissue analysis is limited by intratumoral and intertumoral heterogeneity and its invasive nature. Circulating tumor DNA (ctDNA) analysis, a promising liquid biopsy approach, has been increasingly explored in clinical studies. Biologically, ctDNA is characterized by tumor-specific diversity and rapid clearance from circulation, enabling real-time, dynamic, and repeatable assessments. Technologically, PCR- and NGS-based downstream analysis methods have been developed and validated. However, variables in pre-analytical and analytical procedures underscores the need for standardized protocols. Compared with clinicopathology-based risk stratification, ctDNA-based molecular residual disease detection has demonstrated significant potential in guiding treatment decisions. Qualitative and quantitative changes in ctDNA have also shown predictive and prognostic value during neoadjuvant or adjuvant treatment, as well as in later-line treatment for metastatic CRC. Specific molecular aberrations in ctDNA can not only assist in identifying candidates for targeted therapies but also reveal resistance mechanisms. Additionally, emerging research is exploring the potential of ctDNA in early cancer detection. Overall, as a novel biomarker, ctDNA holds substantial promise in advancing clinical practice. This review focuses on the biological characteristics, pre-analytical variables, and downstream analysis methods of ctDNA and summarizes its role across various clinical scenarios in CRC.

The leucine-rich repeat containing protein (NLR) canonical inflammasome family includes Nod-like receptor protein 3 (NLRP3). Via the mediation of apoptosis proteins and immunological reactions, it controls the pathogenesis of malignancy. Experimental studies showed a relationship among lymphogenesis, cancer metastasis, and NLRP3 expression. Natural products have also been used as lead-based substances in a number of investigations to speed up the creation of novel, specific NLRP3 inhibitors. Via the mediation of apoptotic proteins and immunological responses, it controls the pathogenesis of malignancy. Moreover, it was recently noted that among human cancers, chemotherapy activates NLRP3. Induction of NLRP3 could encourage the generation of IL-1β and IL-22 to facilitate the propagation of malignancy. Additionally, prior research has demonstrated that the usage of NLRP3 in cancer therapy may result in resistance to drugs. The depletion of NLRP3 could affect the survival of cells. Natural products have been used as lead materials in a number of studies to help generate novel, specific NLRP3 antagonists more quickly. In the present review, we examine the mechanism behind the beneficial effects of the natural substances on the inhibition of cancer growth and progression, with special focus on NLRP3 regulation.

The role of adjuvant chemotherapy (adj-CT) in stage II colon cancer remains controversial. Circulating tumor DNA (ctDNA) is a promising biomarker for detecting minimal residual disease (MRD) and predicting recurrence. This systematic review and meta-analysis evaluated the prognostic value of ctDNA in stage II colorectal cancer (CRC), focusing on postoperative detection, post adj-CT outcomes, and dynamic surveillance. A literature search identified studies correlating ctDNA positivity in stage II CRC with recurrence risk, recurrence-free survival (RFS), and disease-free survival (DFS). Seven studies met the inclusion criteria. Postoperative ctDNA positivity significantly increased the risk of recurrence (pooled risk ratio [RR:] 3.66; 95% confidence interval [CI]: 1.25-10.72; p = 0.002). CtDNA positivity after adj-CT was strongly associated with poor survival, while dynamic ctDNA monitoring detected recurrence earlier than conventional methods, including carcinoembryonic antigen (CEA) and imaging. CtDNA is a robust prognostic biomarker in stage II CRC, enabling personalized treatment. High-risk ctDNA-positive patients may benefit from intensified therapy, while ctDNA-negative patients could avoid unnecessary treatments. However, the standardization of detection methods and large-scale validation studies are needed before integrating ctDNA into routine clinical practice as a non-invasive, dynamic tool for personalized care.

Circulating tumor DNA (ctDNA) analysis has emerged as a minimally invasive tool for detecting minimal residual disease (MRD) in colorectal cancer (CRC) patients. This enables dynamic risk stratification, earlier recurrence detection and optimized post-surgical treatment. Two primary methodologies have been developed for ctDNA-based MRD detection: tumor-informed strategies, which identify tumor-specific mutations through initial tissue sequencing to guide ctDNA monitoring, and tumor-agnostic approaches, which utilize predefined panels to detect common cancer-associated genomic or epigenomic alterations directly from plasma without prior tissue analysis. The debate over which is superior in terms of sensitivity, specificity, cost-effectiveness and clinical feasibility remains unsolved.

This review summarizes studies published up to November 2024, exploring the utility and performance of tumor-informed and tumor-agnostic approaches for ctDNA analysis in CRC. We evaluate the strengths and limitations of each methodology, focusing on sensitivity, specificity and clinical outcomes.

Both strategies demonstrate clinical utility in post-operative risk stratification and guiding adjuvant chemotherapy decisions in CRC patients. Tumor-informed approaches generally exhibit superior sensitivity and specificity for recurrence prediction, attributed to their personalized tumor profile designs. However, these methods are limited by the need for prior tissue sequencing and higher associated costs. In contrast, tumor-agnostic approaches offer broader applicability due to their reliance on plasma-only analysis, although with relatively lower sensitivity. Technological advancements, including fragmentomics and multi-omic integrations, are expanding the capabilities of ctDNA-based MRD detection, enhancing the performance of both approaches.

While tumor-informed strategies currently offer higher precision in MRD detection, tumor-agnostic approaches are gaining traction due to their convenience and improving performance metrics. The integration of novel technologies in ongoing clinical trials may redefine the optimal approach for MRD detection in CRC, paving the way for more personalized and adaptive patient management strategies.

In this editorial, we reviewed the article by Fadlallah et al that was recently published in the World Journal of Clinical Oncology. The article provided a comprehensive and in-depth view of the management and treatment of colorectal cancer (CRC), one of the leading causes of cancer-related morbidity and mortality worldwide. The article analyzed the therapeutic modalities and their sequencing, focusing on total neoadjuvant therapy for locally advanced rectal cancer. It highlighted the role of immunotherapy in tumors with high microsatellite instability or deficient mismatch repair, addressing recent advances that have improved prognosis and therapeutic response in localized and metastatic CRC. Innovations in surgical techniques, advanced radiotherapy, and systemic agents targeting specific mutational profiles are also discussed, reflecting on how they revolutionized clinical management. Circulating tumor DNA has emerged as a promising tool for detecting minimal residual disease, prognosis, and therapeutic monitoring, solidifying its role in precision oncology. This review emphasized the importance of technological and therapeutic advancements in improving clinical outcomes and personalizing CRC treatment.

Gastrointestinal (GI) malignancies remain the culprit behind a substantial portion of cancer-related mortality worldwide, and outcomes for patients with advanced or metastatic GI cancers remain poor despite continued efforts to improve care. In 2024, the ongoing clinical trials to optimize and improve GI cancer care showcased progress in molecular diagnostics, systemic therapies, and localized treatment approaches, providing hope for continued progress toward improved patient outcomes.

This review summarizes selected updates in GI cancer care from the 2024 ASCO and ESMO Annual Meetings, including both positive and negative trials that, while not universally practice-changing, contribute to shaping GI cancer care, clinical management, or address key questions in the field. The selected trials cover early detection and diagnostic advances, perioperative management, metastatic disease management, and immune checkpoint inhibitor (ICI)'s emerging role in GI cancer.

Various clinical trials in perioperative management and their results continue to reshape or strengthen the current treatment paradigms. The use of ICIs for microsatellite instability-high colorectal cancer (CRC) presented a notable advancement with the potential to improve patient outcomes. Localized treatments such as thermal ablation appear to benefit some patients with CRC and liver metastases.

The collection of trial results presented at the 2024 ASCO and ESMO Annual Meetings denote the ongoing efforts of the medical and scientific community for optimizing GI cancer care. The ongoing efforts of the GI cancer research and patient community provide hope for continued progress toward improved patient outcomes and new standards of care.

Identification of colorectal cancer (CRC) patients at high risk of recurrence could be of substantial clinical use. We evaluated the association of ctDNA status, using a tumor-informed assay, with recurrence-free survival (RFS).

Stage III CRC patients were enrolled between 2016 and 2020. Tumor tissue and serial (every 3 months for years 1-3, biannually for years 4-5) blood samples were collected. Utilizing whole-exome sequencing and selection of 50-200 variants for tumor informed assays, ctDNA status was determined using plasma cell-free DNA.

Of 137 patients enrolled, 124 with 1029 ctDNA results were included in the analyses. Median follow-up was 4.8 years. Plasma ctDNA status was strongly associated with risk of recurrence during the surveillance period (hazard ratio (HR) 49.6, 95% CI: 16.6-148.3; p < 0.0001), and at the postsurgical (HR 9.6, 95% CI: 3.2-29.5) and postdefinitive therapy timepoints (HR: 16.7, 95% CI: 6.9-40.3). The estimated 3-year RFS for ctDNA positive and ctDNA negative patients were, respectively, 54.5% and 96.1% after surgery, and 18.2% and 90.0% after definitive therapy. Multivariable analysis indicated ctDNA but not CEA was strongly prognostic for recurrence.

Our tumor-informed ctDNA assay was strongly prognostic for recurrence in patients with stage III colorectal cancer at all timepoints.

Analysis of ctDNA may enable early identification of patients likely to relapse, presenting an opportunity for early interventions and improved outcomes. Tumor-naïve plasma-only approaches for minimal residual disease (MRD) assessment accelerate turnaround time, enabling rapid treatment decisions and ongoing surveillance.

Plasma samples were obtained from 80 study participants with stage II or III colorectal cancer selected from CIRCULATE-Japan GALAXY. MRD status was assessed using a tumor-naïve ctDNA assay (xM) that integrates methylation and genomic variant data, delivering a binary call. MRD was assessed at 4 weeks postsurgery [landmark time point (LMT)] using methylation and genomic variant data and longitudinally (median, 22.1 months) using only methylation data.

At LMT, 69/80 study participants were evaluable (36 recurrent; 33 nonrecurrent). Of recurrent study participants, 22/36 had detectable ctDNA (MRD-positive) at LMT and 29/33 nonrecurrent study participants had undetectable ctDNA (MRD-negative), yielding a clinical sensitivity of 61.1% and specificity of 87.9%. Additionally, 74 study participants were evaluable for longitudinal performance with a clinical sensitivity of 83.3% and specificity of 89.5%. The median lead time from the first MRD-positive result to recurrence was 4.77 months overall, and 5.30 months for study participants with no adjuvant treatment. At 12 weeks postsurgery, MRD status strongly correlated with disease-free survival (adjusted HR, 9.69), outperforming carcinoembryonic antigen correlation (HR, 2.13).

This tumor-naïve MRD assay demonstrated clinically meaningful performance at LMT and longitudinally, accurately predicting clinical recurrence. MRD status was a stronger prognostic biomarker for disease-free survival compared with standard-of-care carcinoembryonic antigen.

Colorectal cancer (CRC) is a significant global health concern. We need ways to detect it early and determine the best treatments. One promising method is liquid biopsy, which uses cancer cells and other components in the blood to help diagnose and treat the disease. Liquid biopsies focus on three key elements: circulating tumor DNA (ctDNA), circulating microRNA (miRNA), and circulating tumor cells (CTC). By analyzing these elements, we can identify CRC in its early stages, predict how well a treatment will work, and even spot signs of cancer returning. This study investigates the world of liquid biopsy, a rapidly growing field. We want to understand how it can help us better recognize the molecular aspects of cancer, improve and diagnostics, tailor treatments to individual patients, and keep track of the disease over the long-term. We explored specific components of liquid biopsy, like extracellular vesicles and cell-free DNA, and how they are used to detect CRC. This review sheds light on the current state of knowledge and the many ways a liquid biopsy can be used in treating colorectal cancer. It can transform patient care, disease management, and clinical outcomes by offering non-invasive cancer-targeting solutions.

Biomarkers have revolutionized the management of colorectal cancer (CRC), facilitating early detection, prevention, personalized treatment, and minimal residual disease (MRD) monitoring. This review explores current CRC screening strategies and emerging biomarker applications.

We summarize the landscape of non-invasive CRC screening and MRD detection strategies, discuss the limitations of the current approaches, and highlight the promising potential of novel biomarker solutions. The fecal immunochemical test remained the cornerstone of CRC screening, but its sensitivity has been improved by assays that combined its performance with other stool analytes. However, their sensitivity for advanced adenomas and the patient compliance both remain suboptimal. Blood-based tests promise to increase compliance but require further refinement to compete with stool-based biomarker tests. The ideal scenario involves leveraging blood tests to increase screening participation, and simultaneously promote stool- and endoscopy-based screening among those who are compliant. Once solely reliant on upfront surgery followed by stage and pathology-driven adjuvant chemotherapy, the treatment of stage II and III colon cancer has undergone a revolutionary transformation with the advent of MRD testing after surgery. A decade ago, the concept of using a post-surgical test instead of stage and pathology to determine the need for adjuvant chemotherapy was disruptive. Today, a blood test may be more informative of the need for chemotherapy than the stage at diagnosis.

Biomarker research is not just improving, but bringing a transformative change to CRC clinical management. Early detection is not just getting better, but improving thanks to a multi-modality approach, and personalized treatment plans are not just becoming a reality, but a promising future with MRD testing.

The prognostic role of circulating tumor DNA (ctDNA)-based molecular residual disease (MRD) detection and its utility for postsurgical risk stratification has been reported in colorectal cancer. In this study, we explored the use of ctDNA-based MRD detection in patients with colorectal liver metastases (CLM), for whom the survival benefit of adjuvant chemotherapy (ACT) after surgical resection remains unclear.

Patients with CLM without extrahepatic disease from the GALAXY study (UMIN000039205) were included. The disease-free survival (DFS) benefit of ACT was evaluated in MRD-positive and -negative groups after adjusting for age, gender, number, and size of liver metastases, RAS status, and previous history of oxaliplatin for primary cancer. ctDNA was detected using a personalized, tumor-informed 16-plex polymerase chain reaction-next-generation sequencing (mPCR-NGS) assay. ctDNA-based MRD status was evaluated 2-10 weeks after curative surgery, before the start of ACT.

Among 6061 patients registered in GALAXY, 190 surgically resected CLM patients without any preoperative chemotherapy were included with a median follow-up of 24 months (1-48 months). ctDNA positivity in the MRD window was 32.1% (61/190). ACT was administered to 25.1% (48/190) of patients. In the MRD-positive group, 24-month DFS was higher for patients treated with ACT [33.3% versus not reached, adjusted hazard ratio (HR): 0.07, P < 0.0001]; whereas no benefit of ACT was seen in the MRD-negative group (24-month DFS: 72.3% versus 62.2%, adjusted HR: 0.68, P = 0.371). Multivariate analysis showed that the size of liver metastases (HR: 3.94, P = 0.031) was prognostic of DFS in the MRD-positive group. In the MRD-negative group, however, none of the clinicopathological factors were prognostic of DFS.

Our data suggest that ACT may offer notable clinical benefits in MRD-positive patients with CLM. MRD status-based risk stratification could be potentially incorporated in future clinical trials for CLM.

The interim analysis of the CIRCULATE-Japan GALAXY observational study demonstrated the association of circulating tumor DNA (ctDNA)-based molecular residual disease (MRD) detection with recurrence risk and benefit from adjuvant chemotherapy (ACT) in resectable colorectal cancer (CRC). This updated analysis with a 23-month median follow-up, including 2,240 patients with stage II-III colon cancer or stage IV CRC, reinforces the prognostic value of ctDNA positivity during the MRD window with significantly inferior disease-free survival (DFS; hazard ratio (HR): 11.99, P < 0.0001) and overall survival (OS; HR: 9.68, P < 0.0001). In patients who experienced recurrence, ctDNA positivity correlated with shorter OS (HR: 2.71, P < 0.0001). The significantly shorter DFS in MRD-positive patients was consistent across actionable biomarker subsets. Sustained ctDNA clearance in response to ACT was an indicator of favorable DFS and OS compared to transient clearance (24-month DFS: 89.0% versus 3.3%; 24-month OS: 100.0% versus 82.3%). True spontaneous clearance rate with no clinical recurrence was 1.9% (2/105). Overall, our findings provide evidence for the utility of ctDNA monitoring for post-resection recurrence and mortality risk stratification that could be used for guiding adjuvant therapy.

Over the next few years, the analysis of circulating tumor DNA (ctDNA) through liquid biopsy is expected to enter clinical practice and revolutionize the approach to biomarker testing and treatment selection in GI cancers. In fact, growing evidence support the use of ctDNA testing as a noninvasive, effective, and highly specific tool for molecular profiling in GI cancers. Analysis of blood ctDNA has been investigated in multiple settings including early tumor detection, minimal residual disease evaluation, tumor diagnosis and evaluation of prognostic/predictive biomarkers for targeted treatment selection, longitudinal monitoring of treatment response, and identification of resistance mechanisms. Here, we review the clinical applications, advantages, and limitations of ctDNA profiling for precision oncology in GI cancers.

Circulating tumor (ctDNA) can be used to detect residual disease after cancer treatment. Detecting low-level ctDNA is challenging, due to the limited number of recoverable ctDNA fragments at any target loci. In response, we applied tumor-informed whole-genome sequencing (WGS), leveraging thousands of mutations for ctDNA detection.

Performance was evaluated in serial plasma samples (n = 1283) from 144 stage III colorectal cancer patients. Tumor/normal WGS was used to establish a patient-specific mutational fingerprint, which was searched for in 20x WGS plasma profiles. For reproducibility, paired aliquots of 172 plasma samples were analyzed in two independent laboratories. De novo variant calling was performed for serial plasma samples with a ctDNA level > 10 % (n = 17) to explore genomic evolution.

WGS-based ctDNA detection was prognostic of recurrence: post-operation (Hazard ratio [HR] 6.75, 95 %CI 3.18-14.3, p < 0.001), post-adjuvant chemotherapy (HR 28.9, 95 %CI 10.1-82.8; p < 0.001), and during surveillance (HR 22.8, 95 %CI 13.7-37.9, p < 0.0001). The 3-year cumulative incidence of ctDNA detection in recurrence patients was 95 %. ctDNA was detected a median of 8.7 months before radiological recurrence. The independently analyzed plasma aliquots showed excellent agreement (Cohens Kappa=0.9, r = 0.99). Genomic characterization of serial plasma revealed significant evolution in mutations and copy number alterations, and the timing of mutational processes, such as 5-fluorouracil-induced mutations.

Our study supports the use of WGS for sensitive ctDNA detection and demonstrates that post-treatment ctDNA detection is highly prognostic of recurrence. Furthermore, plasma WGS can identify genomic differences distinguishing the primary tumor and relapsing metastasis, and monitor treatment-induced genomic changes.

The SCRUM-Japan MONSTAR-SCREEN consortium is a nationwide molecular profiling project employing artificial intelligence-driven multiomics analyses for patients with advanced malignancies, aiming to develop novel therapeutics and diagnostics and deliver effective drugs to patients. Concurrently, studies assessing molecular residual disease-based precision medicine for resectable solid tumors, including CIRCULATE-Japan, are ongoing. The substantial data generated by these platforms are stored within a state-of-the-art supercomputing infrastructure, VAPOR CONE. Since 2015, our project has registered over 24,000 patients as of December 2023. Among 16,144 patients with advanced solid tumors enrolled in MONSTAR-SCREEN projects, 5.0% have participated in matched clinical trials, demonstrating a 29.2% objective response rate and 14.8-month median survival (95% CI, 13.4-16.3) for patients treated in the matched clinical trials. Notably, patients who received matched therapy demonstrated significantly prolonged overall survival compared with those who did not (hazard ratio 0.77; 95% confidence interval, 0.71-0.83). Significance: Our nationwide molecular profiling initiative played pivotal roles in facilitating the enrollment of patients with advanced solid tumors into matched clinical trials and highlighted the substantial survival benefits of patients treated with matched therapy. We aim to facilitate an industry-academia data-sharing infrastructure ecosystem, fostering new drug discovery paradigms and precision medicine.

Approximately 50% of patients diagnosed with colorectal cancer develop colorectal cancer liver metastases (CRLM). Although curative intent liver resection provides 5-year survival of 40-50%, up to 70% of patients develop recurrence of CRLM. Detection of minimal residual disease (MRD) is essential for timely, optimized treatment. This study evaluated the feasibility and utility of using circulating tumor DNA (ctDNA) to identify MRD and predict disease recurrence.

Patients with CRLM that underwent liver resection and had known KRAS or PIK3CA mutations were retrospectively identified. Serial blood samples were collected every 3 months following surgery for disease surveillance. ctDNA was isolated from the samples and analyzed with digital PCR (dPCR).

KRAS and PIK3CA mutations were identified by dPCR in 29 patients over 115 timepoints. In patients with detectable ctDNA at time of liver resection, 81% (13/16) developed disease recurrence, while 46% (6/13) of the patients with undetectable ctDNA recurred (p=0.064). Presence of ctDNA was detected in 27.6% (8/29) of the initial postoperative samples. Radiologic recurrence was later diagnosed in 100% (8/8) of these patients, while 52% (11/21) who had undetectable ctDNA postoperatively recurred (p=0.026). Detectable ctDNA postoperatively was associated with a shorter disease-free survival (DFS) of 9 months vs 13 months in patients who had undetectable ctDNA (HR 2.95, 95% CI 1.16-7.49; p=0.02).

Liquid biopsy using dPCR can identify low levels of ctDNA, enabling early detection of disease recurrence. Additionally, the presence of ctDNA postoperatively was predictive of recurrence. This study corroborates current literature and provides rational for moving toward a clinical trial using ctDNA and dPCR to detect MRD after CRLM resection.

We studied the poorly-known dynamics of circulating DNA (cir-nDNA), as monitored prospectively over an extended post-surgery period, in patients with cancer.

On patients with stage III colon cancer (N = 120), using personalised molecular tags we carried out the prospective, multicenter, blinded cohort study of the post-surgery serial analysis of cir-nDNA concentration. 74 patients were included and 357 plasma samples tested.

During post-operative follow-up, the patients' median cir-nDNA concentration was greater (P < 0.0001 in the [43-364 days range]) than both the median value in healthy individuals and the pre-surgery value. These cir-nDNA levels were highly associated with NETs markers (P-value associating MPO and cir-nDNA, and NE and cir-nDNA are 6.6 x 10-17, and 1.9 x 10-7), in accordance with previous reports which indicate that cir-nDNA are NETs by-products. Unexpectedly, in 34 out of 50 patients we found that NETs continued to be formed for an extended duration post-surgery, even in patients without disease progression. Given that this phenomenon was observed in patients without adjuvant CT, and in patients >18 months post-surgery, the data suggest that the persistence of NETs formation is not due to the adjuvant CT.

(1), Given the inter-patient heterogeneity, the post-surgery cir-nDNA level cannot be considered a reliable value, and caution must be exercised when determining mutation allele frequency or the mutation status; and (2), specific studies must be undertaken to investigate the possible clinical impact of the persistent, low-grade inflammation resulting from elevated NETs levels, such as observed in these post-surgery patients, given that such levels are known to potentially induce adverse cardiovascular or thrombotic events.

This work was supported by the H2020 European ERA-NET grant on Translational Cancer Research (TRANSCAN-2).

Approximately 15% of colorectal cancers (CRCs) are associated with germline mutations. There is increasing adoption of DNA-based assays for molecular residual disease (MRD) and growing evidence supporting its clinical utility, particularly for CRC by oncologists in the U.S. We assessed the uptake of germline multi-gene panel testing (MGPT) for hereditary cancer in CRC patients receiving MRD analyses in community oncology settings.

This retrospective study included 80 patients receiving care for CRC through community oncology practices who were referred for MRD testing at a commercial laboratory (January-March 2022). Clinical data, including test requisition forms, pathology reports, and clinical notes were reviewed. Documentation of tumor microsatellite instability and/or immunohistochemical (IHC) testing for mismatch repair (MMR) deficiency, age of CRC diagnosis, family history of cancer, and any order or recommendation for MGPT were assessed.

Overall, 5/80 (6.3%) patients in the study have documented germline MGPT; 65/80 (81.3%) patients have documented MMR testing of their colorectal tumor. Among the 5 cases with abnormal MMR IHC, 2 have MGPT. Of the 33 patients meeting the 2021 National Comprehensive Cancer Network (NCCN) criteria for genetic/familial high-risk assessment, only 2 have MGPT.

Our real-world data suggest that many CRC patients receiving MRD testing and meeting NCCN (v. 2021) criteria for germline MGPT may not be receiving evaluation beyond routine MMR status. Process and educational improvements are needed in community health settings to increase access and uptake of germline testing among CRC patients regardless of age at diagnosis or MMR status.

The sampling of circulating biomarkers provides an opportunity for non-invasive evaluation and monitoring of cancer activity. In modern day practice, this has typically been in the form of circulating tumor DNA (ctDNA) detected in plasma. The field of ctDNA has been a burgeoning technology, with prominent applications for blood-based cancer screening and in disease status assessment, especially after curative-intent surgery to evaluate for minimal residual disease (MRD). Clinical applications for the latter show an incredibly high sensitivity in certain cancer types with a need for additional studies to determine how much clinical decision-making should be adapted based on ctDNA results and which cancer types, stages, and treatments are best informed by ctDNA results. This chapter provides an overview of ctDNA detection as tool for cancer screening, detecting MRD, and/or molecularly characterizing a cancer, highlighting the rapidly amassing research as a prognostic biomarker and emerging data on ctDNA as a predictive biomarker.

Although there has been significant advancement in the identification and management of colorectal cancer (CRC) in recent years, there is still room for improvement in the current standard treatment regimen. One area of concern is the lack of reliable tumor markers to predict treatment efficacy and guide tailored care. Due to its dynamic, effective, and non-invasive benefits over tissue biopsy, the detection of minimal or molecular residual lesions (MRD) based on circulating tumor DNA (ctDNA) is beneficial to the clinical development of drugs for patients with CRC after radical treatment, as well as for continuous monitoring of tumor recurrence and malignancy molecular gene evolution. The detection of ctDNA can currently be used to guide individual postoperative auxiliary treatment decisions (upgrade or downgrade treatment) in CRC, stratify the risk of clinical recurrence more precisely, and predict the risk of recurrence in advance of imaging examination, according to a large number of observational or prospective clinical studies. With increasing clarity comes the possibility of selecting a regimen of treatment based on postoperative ctDNA, which also improves the accuracy of clinical recurrence risk assessment for CRC. Therefore, it is anticipated that the identification of ctDNA would alter the current framework for dealing with CRC and lead to individualized, stratified precision therapy; however, additional confirmation will require subsequent high-quality, prospective, large-scale randomized controlled studies. This article will provide an overview of the definition and clinical significance of MRD, the primary indications and technological challenges for MRD detection, along with the advancement in clinical research about ctDNA detection following radical resection of the CRC.

Liquid biopsy has emerged as a promising noninvasive approach for colorectal cancer (CRC) management. This review focuses on technologies detecting circulating nucleic acids, specifically circulating tumor DNA (ctDNA) and circulating RNA (cfRNA), as CRC biomarkers. Recent advancements in molecular technologies have enabled sensitive and specific detection of tumor-derived genetic material in bodily fluids. These include quantitative real-time PCR, digital PCR, next-generation sequencing (NGS), and emerging nanotechnology-based methods. For ctDNA analysis, techniques such as BEAMing and droplet digital PCR offer high sensitivity in detecting rare mutant alleles, while NGS approaches provide comprehensive genomic profiling. cfRNA detection primarily utilizes qRT-PCR arrays, microarray platforms, and RNA sequencing for profiling circulating microRNAs and discovering novel RNA biomarkers. These technologies show potential in early CRC detection, treatment response monitoring, minimal residual disease assessment, and tumor evolution tracking. However, challenges remain in standardizing procedures, optimizing detection limits, and establishing clinical utility across disease stages. This review summarizes current circulating nucleic acid detection technologies, their CRC applications, and discusses future directions for clinical implementation.

Now, genomics forms the core of the precision medicine concept. Comprehensive investigations of tumor genomes have made it possible to characterize tumors at the molecular level and, specifically, to identify the fundamental processes that cause condition. A variety of kinds of tumors have seen better outcomes for patients as a result of the development of novel medicines to tackle these genetic-driving processes. Since therapy may exert selective pressure on cancers, non-invasive methods such as liquid biopsies can provide the opportunity for rich reservoirs of crucial and real-time genetic data. Liquid biopsies depend on the identification of circulating cells from tumors, circulating tumor DNA (ctDNA), RNA, proteins, lipids, and metabolites found in patient biofluids, as well as cell-free DNA (cfDNA), which exists in those with cancer. Although it is theoretically possible to examine biological fluids other than plasma, such as pleural fluid, urine, saliva, stool, cerebrospinal fluid, and ascites, we will limit our discussion to blood and solely cfDNA here for the sake of conciseness. Yet, the pace of wider clinical acceptance has been gradual, partly due to the increased difficulty of choosing the best analysis for the given clinical issue, interpreting the findings, and delaying proof of value from clinical trials. Our goal in this review is to discuss the current clinical value of ctDNA in cancers and how clinical oncology systems might incorporate procedures for ctDNA testing.

Despite decreased incidence rates in average-age onset patients in high-income economies, colorectal cancer is the third most diagnosed cancer in the world, with increasing rates in emerging economies. Furthermore, early onset colorectal cancer (age ≤50 years) is of increasing concern globally. Over the past decade, research advances have increased biological knowledge, treatment options, and overall survival rates. The increase in life expectancy is attributed to an increase in effective systemic therapy, improved treatment selection, and expanded locoregional surgical options. Ongoing developments are focused on the role of sphincter preservation, precision oncology for molecular alterations, use of circulating tumour DNA, analysis of the gut microbiome, as well as the role of locoregional strategies for colorectal cancer liver metastases. This overview is to provide a general multidisciplinary perspective of clinical advances in colorectal cancer.

To assess the efficacy of ctDNA measurement at different time intervals in predicting response and prognosis in patients diagnosed with locally advanced rectal cancer (LARC) who underwent neoadjuvant treatment prior to curative resection.

English language randomized controlled trials and observational studies, published from 1946 to January 2024, comparing outcomes between ctDNA-positive and ctDNA-negative patients with LARC undergoing neoadjuvant treatment prior to curative surgical resection were included in the search. The search included Ovid MEDLINE, Embase, Cochrane Central Register of Controlled Trials (CENTRAL), and the Cochrane Database of Systematic Reviews (CDSR).

Data for 1022 patients were analysed. Patients with positive ctDNA in the preoperative period had more than five times the risk of developing distant metastasis (RR [95% CI] 5.03 [3.31-7.65], p < 0.001), while those with positive ctDNA in the postoperative period had more than six times the risk (RR [95% CI] 6.17 [2.38-15.95], p < 0.001). There was no significant relationship between ctDNA status at baseline, pre-, or postoperative periods and achievement of pCR (RR [95% CI] 1.21 [0.86-1.7], 1.82 [0.94-3.55], 1.48 [0.78-2.82], p = 0.27, 0.08, and 0.23, respectively). However, patients with positive ctDNA in the pre- and postoperative periods had more than 13 and 12 times the risk of overall disease relapse after curative-intent treatment (RR [95% CI] 13.55 [7.12-25.81], 12.14 [3.19-46.14], p < 0.001), respectively.

ctDNA could potentially guide treatment and follow-up in LARC, predicting high-risk patients for disease relapse, allowing individualized surveillance and treatment strategies. Prospective studies are needed for standardization.

Circulating tumor DNA (ctDNA) is the fraction of cell-free DNA in patient blood that originates from a tumor. Advances in DNA sequencing technologies and our understanding of the molecular biology of tumors have increased interest in exploiting ctDNA to facilitate detection of molecular residual disease (MRD). Analysis of ctDNA as a promising MRD biomarker of solid malignancies has a central role in precision medicine initiatives exemplified by our CIRCULATE-Japan project involving patients with resectable colorectal cancer. Notably, the project underscores the prognostic significance of the ctDNA status at 4 weeks post-surgery and its correlation to adjuvant therapy efficacy at interim analysis. This substantiates the hypothesis that MRD is a critical prognostic indicator of relapse in patients with colorectal cancer. Despite remarkable advancements, challenges endure, primarily attributable to the exceedingly low ctDNA concentration in peripheral blood, particularly in scenarios involving low tumor shedding and the intrinsic error rates of current sequencing technologies. These complications necessitate more sensitive and sophisticated assays to verify the clinical utility of MRD across all solid tumors. Whole genome sequencing (WGS)-based tumor-informed MRD assays have recently demonstrated the ability to detect ctDNA in the parts-per-million range. This review delineates the current landscape of MRD assays, highlighting WGS-based approaches as the forefront technique in ctDNA analysis. Additionally, it introduces our upcoming endeavor, WGS-based pan-cancer MRD detection via ctDNA, in our forthcoming project, SCRUM-Japan MONSTAR-SCREEN-3.

No reliable marker has been identified to predict postoperative recurrence of gastric cancer. We designed a clinical trial to investigate the utility of serum NY-ESO-1 antibody responses as a predictive marker for postoperative recurrence in gastric cancer.

A multicenter prospective study was conducted between 2012 and 2021. Patients with resectable cT3-4 gastric cancer were included. Postoperative NY-ESO-1 and p53 antibody responses were serially evaluated every 3 months for 1 year in patients with positive preoperative antibody responses. The recurrence rate was assessed by the positivity of antibody responses at 3 and 12 months postoperatively.

Among 1001 patients, preoperative NY-ESO-1 and p53 antibody responses were positive in 12.6% and 18.1% of patients, respectively. NY-ESO-1 antibody responses became negative postoperatively in non-recurrent patients (negativity rates; 45% and 78% at 3 and 12 months, respectively), but remained positive in recurrent patients (negativity rates; 9% and 8%, respectively). p53 antibody responses remained positive in non-recurrent patients. In multivariate analysis, NY-ESO-1 antibody positivity at 3 months (P  < 0.03) and 12 months (P  < 0.001) were independent prognostic factors for a shorter recurrence-free interval.

Serum NY-ESO-1 antibodies may be a useful predictive marker for postoperative recurrence in gastric cancer.