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Sequencing Patient's Circulating Tumor Cells Reveals Possible Treatment Approaches

NEW YORK (GenomeWeb) – Analyzing the genomes of as few as five circulating tumor cells, a Complete Genomics-led team of researchers was able to identify potentially actionable mutations, according to a new study.

Using immunomagnetic enrichment and fluorescence-activated cell-sorting, the team isolated 34 circulating tumor cells from a patient with metastatic breast cancer at two time points and sequenced those cells' genomes using long-fragment reads. As they reported in Cancer Research today, the researchers identified amplifications and deletions that were common across the set of CTCs as well as mutations present only in a portion of cells, including alterations that suggested possible treatment approaches.

When they narrowed their focus to sets of just five CTCs at a time, the researchers were still able to pick out the spectrum of mutations and uncover potentially clinically actionable mutations.

"That our sequencing method could detect the most important somatic mutations from just five CTCs in a noninvasive liquid biopsy is important, demonstrating cost-effectiveness and utility in clinical settings," Brock Peters, senior director of research at Complete Genomics, which is owned by BGI, said in a statement.

The researchers collected two blood samples from a 61-year-old woman with ER-positive/HER2-negative metastatic breast cancer. After isolating 34 CTCs, and five normal cells, they underwent long-fragment read whole-genome sequencing. The genomes of the CTCs were analyzed as more than 3,000 barcoded subgenomic compartments, and each cell was sequenced to about 23X coverage.

All the CTCs had amplifications in chromosome 1 and deletions in chromosomes 13 and 16, the researchers reported, noting that those deleted regions harbor the tumor suppressor genes BRCA2, CDH1, and RB1. None of these alterations were present in normal cells.

Within all CTCs, the researchers uncovered 2,766 somatic SNVs, along with 543 indels and multi-base substitutions. From these variants, Peters and his colleagues were able to tease out apparent driver mutations. In particular, they noticed a frameshift mutation in the tumor suppressor gene CDH1 that was present in all CTCs. They suggested that this mutation, in conjunction with the loss of one copy of chr16q — where this gene is located — likely led to the complete loss of the E-cadherin gene product, which the researchers said is a frequent event in lobular breast cancer.

But the cells were also highly heterogeneous, the researchers reported, noting that there were nearly 17,000 somatic mutations that only appeared in subsets of CTCs.

The mutation patterns the researchers captured resembled those of breast cancer data in the Cancer Genome Atlas, also suggesting that CTCs could reveal a tumor's tissue of origin.

The mutations the researchers uncovered within the CTCs also suggested potential treatment options. For instance, likely activating mutations in PDPK1, MAPK8, and PDGFRA indicated that this patient might have benefited from kinase inhibitor therapy. At the same time, the authors uncovered a number of defects in DNA repair genes. The patient, though, died in 2014.

Peters and his colleagues also analyzed the CTCs in sets of five, as in a clinical setting, 34 CTCs might not be available or be too expensive to sequence. They estimated that sequencing and analyzing five CTCs might cost $3,000 within the next few years, in line with current diagnostic tests in oncology.

Analysis of just this portion of CTCs, though, largely reflected their overall findings. They were still about to identify complex structural rearrangements, amplifications of chr1q, and deletions of chr13 and chr16q, as well as the same spectrum of somatic SNVs.

"From 34 cells, we accurately detected mutations present in as few as 12 percent of CTCs, established the tissue of origin, and identified potential personalized combination therapies for this patient's highly heterogeneous disease," Peters said.