Research is ongoing to explore all potential uses for circulating tumor DNA to help reduce the global burden of colorectal cancer.
Circulating tumor (ctDNA) technology has the potential to improve diagnosis, treatment, and monitoring of patients with colorectal cancer (CRC).1 Research is ongoing to explore all potential uses for ctDNA to help reduce the global burden of CRC.1-4
Although CRC is preventable and curable when diagnosed early, most patients receive diagnoses at advanced stages.2 Early and accurate detection of CRC tumors is critical for success, and biopsy with tissue pathology remains the gold standard for diagnosis despite the associated costs, invasive and uncomfortable nature, technical limitations based on tumor location, and ineffi ciency in targeting tumor subpopulations. Fragmented DNA in bodily fluids originating from a primary or metastatic tumor, known as ctDNA, has the potential to provide a simple, cost-effective, non-invasive (or limited) approach to diagnosing and characterizing CRC and is being evaluated and incorporated in numerous clinical trials.
Current assays for the detection of ctDNA are either tumor informed or tumor uninformed.3 Newer tumor-uninformed liquid biopsies are cancer specific, require a simple blood sample, and pair an epigenomic signature with the ctDNA component to identify neoplasm-specific genetic and epigenetic abnormalities.1,3 Tumor-informed assays are patient specific, requiring a matched sample of the tumor to generate a custom genetic panel for identifying ctDNA within the patient.3 Emerging ctDNA technologies provide opportunities for minimal residual disease (MRD) detection in patients with early-stage disease, enhanced risk stratifi cation, identification of tumor heterogeneity and genomic evolution, and improved ctDNA-guided use of neoadjuvant and adjuvant therapies.1,4,5
Although much of the research for ctDNA in CRC to date has been focused on MRD and the metastatic setting, there are also potential applications in the adjuvant setting.6 The detection of ctDNA after surgery correlates with a high risk of recurrence. After curative resection, 10% to 15% of stage II patients will have detectable ctDNA compared with almost 50% of stage IV patients.4
In addition, ctDNA is proving a valuable prognostic indicator. One meta-analysis of patients with CRC determined a strong association between ctDNA and recurrence-free survival (RFS; HR, 2.78; 95% CI, 2.08-3.72) and overall survival (OS; HR, 3.03; 95% CI, 2.51-3.66), suggesting the presence of ctDNA as a negative prognostic indicator for RFS and OS.7
Two experts discussed the latest research and opinions regarding ctDNA within CRC in interviews with Targeted Therapies in Oncology™. Pashtoon M. Kasi, MD, MS, is director for colon cancer research and precision medicine and director of liquid biopsy research at the Sandra and Edward Meyer Cancer Center at Weill Cornell Medicine Englander Institute of Precision Medicine, New York, New York. Tanios S. Bekaii-Saab, MD, is medical director of the Cancer Clinical Research Offi ce and vice chair and section chief for medical oncology in the Department of Internal Medicine at Mayo Clinic in Phoenix, Arizona.
The CIRCULATE-Japan collection of trials aims to refine precision medicine for patients with CRC by improving adjuvant therapy choice through the utilization of ctDNA technology.8 The study involves the GALAXY, VEGA, and ALTAIR trials.
GALAXY is a prospective large-scale registry trial designed to monitor ctDNA for patients with stage II to IV or recurrent CRC who are candidates for complete resection.8,9 The VEGA trial is a randomized phase 3 trial open to patients from GALAXY with high-risk stage II or low-risk stage III colon cancer who are ctDNA negative 4 weeks postoperatively, and aims to determine whether surgery alone is noninferior to standard post-operative therapy with capecitabine plus oxaliplatin for 3 months.8 The ALTAIR trial (NCT04457297) is a double-blind phase 3 trial open to ctDNA positive patients from the GALAXY trial and will evaluate the use of trifl uridine/tipiracil (TAS-102, Lonsurf) compared with placebo. Collectively, the CIRCULATE-Japan trials involve both de-escalation and escalation trials for ctDNA-negative and ctDNA-positive patients to help determine the postoperative prognostic and/or predictive value of ctDNA.
A total of 1365 patients with CRC were enrolled in the GALAXY study, comprising 116 with stage I disease, 478 with stage II, 503 with stage III, and 268 with oligometastatic resectable stage IV disease.9 Six-month disease-free survival (6M-DFS) rate analysis excluded patients enrolled in VEGA or ALTAIR. The ctDNA dynamics (positive or negative) were evaluated from week 4 to week 12 to assess changes from baseline.
At a median follow-up of 11.4 months, a significant difference was observed between the “positive to negative” and “positive to positive” groups with an HR of 15.8 (95% CI, 5.7-44.2; P < .001), and compared with the “negative to positive” group with (HR, 9.2; 95% CI, 3.0-28.4; P < .001).9 Ninety-six of 183 evaluable patients who were positive at 4 weeks and had an available 12-week status were treated with standardof-care (SOC) adjuvant chemotherapy (ACT).9 Across most subgroups, the ctDNA clearance rate was higher for those who received ACT vs those without ACT. However, in patients with stage IV disease (n = 68), the clearance rate was 23% in patients who received ACT vs 53% in those who did not receive ACT (P < .001), whereas the rates were 68% and 29% (P < .001), respectively, among patients with stage III disease (n = 90). The cumulative incidence of ctDNA clearance by 24 weeks was significantly higher in the ACT group vs the non-ACT group (68% vs 10%; HR, 9.3; 95% CI, 4.6-18.9; P < .001) and the 6M-DFS rate among patients who were ctDNA-positive at 4 weeks was significantly higher in the ACT vs non-ACT group across disease stages.
GALAXY is the largest MRD study to date and demonstrates a relationship between ctDNA dynamics and improved clinical outcomes, suggesting ctDNA can help identify patients more likely to benefit from ACT therapy. VEGA and ALTAIR are ongoing and will help further establish ctDNA-guided ACT strategies.
“ctDNA is a dynamic variable,” stated Kasi, “and with GALAXY we were able to see patients who were positive convert to negative with ‘mop-up’ chemotherapy. The RFS was the same for those who remained negative and those who were positive but converted to negative.”
Kasi and Bekaii-Saab were interested in the data regarding ACT. “The one group that is very intriguing is the ctDNA-negative cohort,” Bekaii-Saab commented, “where there wasn’t much difference between patients receiving ACT and those who did not. [Although] this study wasn’t randomized, it indicates a possible predictive value for this test.”
Kasi added, “The GALAXY results are very provocative and give rationale for future de-escalation evaluation and support ongoing studies, such as CIRCULATE-US.” ctDNA
A retrospective, single-center cohort study evaluated a ctDNA assay (Signatera; Natera) compared with standard radiographic imaging and carcinoembryonic antigen (CEA) level measurements as evaluation of recurrence in patients with resected CRC.10 Of the 48 patients included in the study, 15 had disease recurrence. The sensitivity of the ctDNA assay (53.3%; 95% CI, 27.4%-77.7%) was numerically less than imaging alone (60.0%; 95% CI, 32.9%-82.5%) and the combination of imaging with CEA measurement (73.3%; 95% CI, 44.8%-91.1%; P = .55). Additionally, there was no significant differenced noted in the median time to detection of recurrence between ctDNA (14.3 months), imaging (15.0 months), or the combination of imaging with CEA (15.0 months). Although these results are retrospective, taken from a single-center, and comprise a relatively small patient population, they suggest that ctDNA assays may not improve surveillance compared with imaging and CEA levels. However, this study did find a low correlation between low-burden lung recurrence and negative ctDNA results that should be further investigated in larger prospective studies.
However, Bekaii-Saab and Kasi were equally surprised by the amount of attention this study received, given bias, vague end points, and lack of blindness. Bekaii-Saab commented, “I personally put it in the interesting observation category with no real practice application. It isn’t just how early you detect cancer; it’s [whether] a difference can be made in survival outcomes.”
A small study consisting of 14 patients with metastatic CRC who were treated using chemotherapy alone or in combination with a targeted agent indicates ctDNA analysis of plasma samples obtained within 4 to 10 weeks after treatment initiation can support molecular response assessments.11 A consistent reduction in mean variant allele fraction (mVAF) as assessed by the GuardantOMNI 500-gene liquid biopsy assay was observed in 11 of 13 evaluable patients. One outlier had an initial increase in mVAF followed by consistent decrease thereafter, whereas the second outlier had elements of KRAS alterations that were not previously detected by tissue biopsy. These results suggest patient-centric liquid biopsy has an advantage in detecting potential resistance mutations prior to therapy over genetic testing of tumor samples.
A recent study evaluating ctDNA via ultradeep sequencing found ctDNA measurements correlated with the stage of disease, with lower levels of ctDNA concentration in stage I patients vs stage IV patients.12 Additionally, ctDNA testing identified 112 alterations in 14 genes in 69% of patients studied (69/100). The most frequent mutations were in APC (29%), TP53 (25%), KRAS (19%), and EGFR (5%). RAS family mutations were identified in 37 patients; 89% of those identified by tissue sampling were identified via ctDNA testing. Overall, ctDNA was found to be a greater performance predictor than CEA measurement, and ctDNA mutation levels reflected 5 patients’ clonal evolution and clinical outcomes. Therefore, monitoring disease progression and clonal evolution via ctDNA testing has the potential to improve patient management.
A recent retrospective study evaluated data from 595 patients from 3 earlier CRC clinical trials that included information on high-risk factors and postsurgical ctDNA status.13 RFS in patients with high-risk stage II CRC who were ctDNA negative did not differ between patients who did or did not receive ACT (P = .1758) despite high-risk patients having significantly reduced RFS compared with low-risk patients (HR, 3.283; 95% CI, 1.398-14.18; P = .0116). These results suggest ctDNA alone is not enough to suggest which patients will experience recurrence and post-operative ctDNA status, and risk factor combinations may need to be assessed for more accurate therapeutic decisions.
“Currently, negative predictive value can be problematic and varies,” stated Bekaii-Saab. “We would want to rely on these tests, because of the potential to prevent unnecessary chemotherapy exposure if negative. However, if the negative predictive value isn’t as accurate then it becomes more challenging for decision-making and conversations with the patient.”
Kasi added, “For the negative patient, you can add 2 additional points for prolonged tumor assessment, increasing the sensitivity and specificity.”
The BESPOKE CRC trial (NCT04264702) is a prospective, multicenter trial evaluating the clinical impact of serial testing with Signatera in patients with resectable CRC, including as a potential guide for adjuvant therapy use.14 Eligible patients have undergone surgery for stage I to IV CRC and have formalin-fixed paraffin-embedded tissue available. Also, ctDNA will be evaluated using the Signatera test, and patients may be recommended for post-operative systemic therapy or observation with a follow-up of up to 2 years. A control arm will consist of matched stage I to IV CRC cases with a minimum of 2 years’ follow-up data available. Currently, the study has over 100 participating locations and 1000 patients enrolled of the 2000 enrollment target with an interim analysis planned for later this year.
The Prospective Dutch Colorectal Cancer cohort study (NCT02070146) will analyze patients with CRC in nearly all hospitals within the Netherlands to further research in the field; the trial includes subcohorts for varying stages of disease.15
Under this trial umbrella, the MEDOCC-CrEATE study aims to evaluate how many patients with stage II colon cancer who are ctDNA-positive postoperatively will accept ACT, and whether ACT reduces the rate of recurrence.16 The study will enroll 1320 patients without an indication for ACT who will be randomized 1:1 into the experimental and control arms. In the experimental arm, ctDNA is identifi ed using the PGDx elio platform, and patients with detectable ctDNA are offered 8 cycles of adjuvant capecitabine and oxaliplatin. The control arm and patients without detectable ctDNA will receive standard follow-up care. The proportion of patients accepting ACT when ctDNA is detected is the primary end point with secondary end points of 2-year and 5-year response rates.
The PROVENC3 study aims to determine the prognostic value of post-operative ctDNA in stage III colon cancer in patients treated with ACT.17 The observational study consists of 267 patients with stage III colon cancer who will be evaluated by PGDx elio plasma resolve assay to determine the association between postoperative ctDNA and progressionfree survival and OS. A ctDNA-guided interventional trial will be designed based on this study’s results.
The CIRCULATE-US study (NCT05174169) will evaluate approximately 2000 patients with stage II or III colon cancer post resection.18 MRD-negative patients by ctDNA will be randomized to SOC ACT or observation and ctDNA-positive patients will be randomized to SOC chemotherapy or intensifi ed chemotherapy with modified FOLFIRINOX (oxaliplatin, leucovorin, irinotecan, and 5-fluorouracil). The study’s primary aim is to compare DFS in MRD-negative patients treated with either immediate or delayed chemotherapy and in MRD-positive patients treated with either SOC or intensified ACT.
More data are needed before clinicians can rely on ctDNA for anything more than diagnosis or prognosis. “What we need are prospective studies,” said Bekaii-Saab. He also commented on additional considerations if ctDNA continues to show promise. “We need to think about how to improve the logistics, and one of my concerns is the need for a strong value but with a controlled cost for these tests. Tissue-informed tests require collaboration with surgeons to ensure the tissue is obtained as early as possible. Liquid-only tests can be cheaper and faster, but the loss of negative-predictive value is a huge problem. Hopefully as we refine our methods we can improve the sensitivity of liquid-only testing.”
For the community oncologist, Kasi and Bekaii-Saab discussed ctDNA as an added piece to the puzzle. “ctDNA is not meant to be in isolation and should be used in conjunction with clinical assessment,” said Kasi. “It boils down to informed decision-making. A positive ctDNA [result] could encourage greater surveillance or different imaging modalities than CT such as MRI or PET scans.”
Bekaii-Saab added that lack of knowledge of what to do with these test results can increase anxiety in decision-making.
“If you decide to use the test, this will require extensive conversation with the patient concerning the ‘what-ifs’ and the available data,” he said.
He added that he will use ctDNA in certain cases. For example, a patient with stage IIIA colon cancer who is reluctant to undergo chemotherapy may be able to make a more confi dent decision based on a tumor-informed assay.
“Even if the test is negative, there’s a 15% chance of false negative across all stages, and this is still within the margin of recurrence rate.”
There is little question regarding the prognostic value of ctDNA. “Most experts agree this is likely the single most important prognostic indicator,” stated Kasi.
However, more data are needed to determine its use as predictive value, assessing for therapeutic targets, or determining treatment response.
Kasi concluded, “There is need for prospective data, but it is often hard to find funding given you aren’t evaluating a novel treatment. If we don’t perform systematic randomized studies free of bias, a lack of good data to support either direction for ctDNA will result.”