85P - True single-circulating tumor cell genomics reveals enriched therapy-resistance signatures in advanced colorectal cancer patients

Background

Plasma-ctDNA (circulating tumor DNA) has emerged as a novel biomarker to detect genomic alterations and longitudinal monitoring of CRC patients, and in nearly 30 % of patients show - no mutations detected, potentially missing companion therapy. Single circulating tumor cells genomics can be more sensitive to detect actionable targets. We show CGP of live sCTCs and paired ctDNA from an advanced CRC population.

Methods

Retrospectively, live sCTCs and CTC clusters were isolated in 6 patients with stage 4 CRC patients using OncoRADAR technology. Whole genomes of sCTCs were amplified, target enriched with hybridization capture using OncoIndx, a comprehensive 1080 gene panel to obtain sequencing libraries, and sequenced on Illumina NextSeq2000 platform in paired-end mode (depth 500x). Raw sequence alignment and variant calling were performed using iCare software. Paired ctDNA samples were processed similarly, but sequenced at high depth (5500x).

Results

A total of 22 sCTCs were isolated including 4 CTC clusters. Combined mutational landscape showed a presence of 142 clinically relevant mutations including 65 missense (55.77%), 25 nonsense (17.60%), 16 frameshift (11.26%), 7 indels (4.6%), 10 splice variants (7.4%) and 19 structural variants (13.4%). NRAS was the most frequently mutated gene occurring in 52% of samples followed by SMO (47.6%), TAP1 (42.85%), and TP53 (42.5%). Paired ctDNA showed TP53 (66%), KRAS (50%), and TAP1 (33.33%) as the most frequently mutated genes. At the individual gene level, 40% concordance was observed between sCTC and ctDNA. The genomic profile of sCTC was particularly enriched with mutations in proliferative and stemness maintenance signaling (NRAS:p.A146T, SMO:p.V392G,) suggesting potential for therapy evasion pathways. CTC showed a higher accumulation of immunotherapy resistance signatures (loss of function STK11 and STAT5B mutations), undetected in paired ctDNA samples.

Conclusions

The genomic profile of sCTCs exhibited enriched mutations in proliferative and stemness maintenance signaling. Therapy resistance signatures were prevalent in sCTCs compared to ctDNA, and may offer clinical insights for patients unable to provide tissue biopsy or show negative ctDNA.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

OneCell Dx.

Funding

OneCell Dx.

Disclosure

All authors have declared no conflicts of interest.