When ctDNA Comes Back Negative, the Story Isn’t Over
Plasma ctDNA has become a go-to biomarker for detecting genomic alterations and monitoring colorectal cancer (CRC) progression.
But there’s a critical blind spot: in nearly 30% of patients, ctDNA shows no mutations detected at all, potentially leaving actionable, companion-therapy-relevant targets completely invisible.
At 1Cell.Ai, we asked a simple question:
What is ctDNA missing, and can single-cell CTC genomics see it?
Our Study: Paired sCTC and ctDNA Profiling in Advanced CRC
We retrospectively isolated live single circulating tumour cells (sCTCs) and CTC clusters from 6 patients with stage 4 colorectal cancer using our OncoRADAR technology.
Whole genomes from these sCTCs were amplified and profiled using our comprehensive 1080-gene OncoIndx panel, sequenced on the Illumina NextSeq 2000 platform. Paired ctDNA samples from the same patients were sequenced in parallel at much higher depth, allowing a direct, patient-matched comparison between single-cell and bulk liquid biopsy data.
What We Found
The genomic landscape revealed by sCTCs told a story that ctDNA alone could not:
22 sCTCs, including 4 CTC clusters, were isolated and profiled, revealing 142 clinically relevant mutations spanning missense, nonsense, frameshift, indel, splice, and structural variants.
NRAS emerged as the most frequently mutated gene across sCTCs (52%), followed by SMO, TAP1, and TP53, a markedly different mutational hierarchy than what ctDNA alone reported.
Paired ctDNA instead showed TP53, KRAS, and TAP1 as the dominant mutated genes, with only 40% concordance between sCTC and ctDNA at the individual gene level.
sCTCs were distinctly enriched for mutations in proliferative and stemness-maintenance signalling pathways, including NRAS and SMO hotspot variants, pointing to active therapy-evasion biology.
CTCs also harboured immunotherapy resistance signatures, specifically loss-of-function STK11 and STAT5B mutations, that were completely undetected in matched ctDNA
Why This Matters?
This study makes clear that single-cell CTC genomics and ctDNA are not interchangeable; they are complementary windows into a patient’s evolving tumour biology. For the substantial subset of CRC patients whose ctDNA returns negative, or who cannot provide tissue for biopsy, sCTC profiling offers a path to actionable insight that would otherwise be missed entirely.
As CRC tumours evolve under treatment pressure, capturing therapy-resistance signatures at single-cell resolution gives oncologists a more complete, more honest picture of what they’re actually treating, and what might be quietly working against them.
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