Background:

Post curative-intent surgery and therapy, the presence of circulating tumor DNA (ctDNA) load represents minimal residual disease (MRD). Conversely, the presence of circulating tumor cells (CTCs) in stage I–II cancer or even in disease-free survival (DFS) patients indicates occult cellular residual disease (CRD) with undetectable micrometastasis. These complementary biomarkers in patients undergoing treatment act as indicators of non-responsiveness, suggesting the need for treatment modifications.

Methods:

Retrospectively, we monitored a cohort of 46 cancer patients for MRD using ctDNA and CTCs who were treated or undergoing treatment (e.g., lung, breast, colon, and head and neck cancer; n = 14, 7, 6, and 4, respectively). The OncoMonitor test detected single nucleotide variations (SNVs), small insertions and deletions (INDELs), copy number variations (CNVs), and translocations (fusions). Libraries were prepared using a hybridization-capture method covering 1000 targets with a mean sequencing depth of 5000× on the Illumina NextSeq 2000 in paired-end mode (150 × 2). Variant calling was performed using a proprietary bioinformatics pipeline, iCare. CTCs were isolated using the OncoDiscover platform, which possesses an anti-EpCAM antibody-based immunomagnetic system per 1.5 mL of blood. CTCs were confirmed using CK18+, PD-L1, and CD45 markers with a motorized fluorescence microscope.

Results:

From ctDNA analysis, 47.82% (n = 22) of patients were identified with at least one actionable genomic finding. Among these, 13.04% (n = 6) of patients showed EGFR driver mutations. Additionally, 19.56% (n = 9) of patients were identified with either EGFR driver, KRAS, or PI3K passenger mutations, while 4.34% (n = 2) were identified with ALK–EML4 fusion. The average ctDNA load obtained in patients with progressive disease (n = 26) was 8.2 molecules per 1 mL of plasma. At least one CTC was detected in 61.53% (n = 16) of progressive disease patients, with the highest count of four CTCs identified in 7.69% (n = 2) of patients.

Only 30% (n = 6) of patients with stable disease were identified with at least one genomic finding from a total of 20 patients upon ctDNA analysis, with an average ctDNA load of 2.2 molecules per 1 mL of plasma. Patients with clinically progressive disease showed ctDNA load approximately fourfold higher than those with stable disease during treatment. No patients with stable disease were identified with four CTCs, as opposed to 7.69% in the progressive disease cohort during treatment.

Conclusions:

We observed that ctDNA and CTCs complement MRD status even after curative-intent surgery and therapy, with the potential to identify patients likely to experience disease progression. Our findings strongly indicate a positive correlation between ctDNA load, the number of detected CTCs, and disease progression based on radiological findings. These biomarkers can support practical clinical decision-making. Further studies are necessary to validate these findings and improve follow-up strategies for better clinical outcomes.