NEW YORK (GenomeWeb) – Stanford University researchers last week published a study on a relatively new circulating tumor cell capture technology from Vortex Biosciences, reporting that the system offers a promising method for isolating CTCs with high enough purity to allow successful whole-genome amplification and targeted sequencing.
Published in NPJ Genomic Medicine last week, the study, conducted with funding from Vortex, explored the firm's VTX-1 system as a tool to capture CTCs from colorectal cancer patients ahead of whole genome amplification and targeted next-generation sequencing.
Researchers optimized a DNA extraction, amplification and sequencing workflow, which they then tested on both tissue and blood samples from patients with metastatic colorectal cancer.
Numerous technologies have emerged for the isolation tumor or other rare cells from blood samples, and more recently, a divide has emerged between affinity technologies — which use specific surface markers or other molecules to tag CTCs and pull them out of the larger pool of circulating cells — and label-free methods that take advantage of size-based filtration, or other microfluidic techniques to concentrate CTCs from the overall background.
Vortex's approach falls under the latter category. The VTX-1 uses a microfluidic chip to capture CTCs in what the company describes as "micro-scale vortices" based on physical properties like cell size and deformability. CTCs are trapped in these microscopic currents and eddies, while red and white blood cells flow past.
Steve Crouse, Vortex's chief commercial officer, said that because the system uses only fluidic properties to isolate CTCs and has a unidirectional flow (as opposed to other microfluidic systems that involve filtering), it is truly fully automated, from the input of a blood tube directly to the output of a concentrated pool of CTCs.
While CTCs have been a subject of research attention for some time, the thrust of non-invasive cancer analysis has shifted more recently to circulating cell-free DNA due to the challenges posed by sequencing DNA from rare cells. The Stanford authors argued, however, that with the right methodology, CTCs might also be harnessed as a surrogate for conventional tissue biopsies.
A central issue, they wrote is purity. While technologies exist that can pick out individual CTCs from concentrated pools, they are cumbersome. But if you try to perform NGS on pooled cells you need a really good enrichment platform with high capture efficiency and purity.
Crouse said that Vortex believes, based on the published results from studies of other systems, that it's technology surpasses competitors in terms of purity — enough so that it makes molecular profiling using CTCs much more feasible.
"All systems capture some white blood cells, but our technology really minimizes that," he argued. "From a typical blood tube we get less than 80 white blood cells per [milliliter]," he added.
However, the firm has not conducted head-to-head studies with other microfluidic CTC platforms — like Angle PLC's Parsortix — that would prove superiority for VTX-1 in reducing contamination by white blood cells.
In the recently published Genome Medicine study, investigators from Stanford and from Vortex applied a finalized extraction, amplification, and sequencing protocol to CTCs that were isolated from blood samples from three colorectal cancer patients using the VTX platform, comparing them to healthy donors.
The cancer patients showed a range of CTC numbers — from 50 per ml in the first patient to only about two per ml in the third. Illustrating the limitations of simply counting CTCs in order to detect or quantify cancer, the two healthy control donors also had cells that were captured by the Vortex system: 11 and 16 per tube respectively.
Sequencing results revealed that there were no somatic mutations in the CTC-like cells isolated from the control subjects. However, germline mutations in two genes did show up in both the tumor-like cells captured by the Vortex system and the white blood cells of one healthy control.
The first CRC patient showed three mutations initially — in MLH1, MSH2, and ATM. But the MLH1 variant was also present in the patient's white blood cells and in a liver metastasis biopsy, indicating that it was likely a SNP rather than a cancer-associated mutation.
The second patient showed no mutations in white blood cells, but had an ATM mutation detected in CTCs and another two variants — in APC and TP52 in a matched liver tissue sample.
Results for the third CRC subject were similar. One mutation was detected in CTCs and two additional mutations in tumor tissue.
Researchers reported that they are now studying a much larger cohort of patients to further evaluate the concordance between CTCs and tumor tissue NGS results.
According to Crouse, while the new study illustrates how Vortex's platform offers sample purity that can negate the need for picking out individual CTCs to sequence, researchers that want to study individual cells can also use the system to do that.
He argued that Vortex's technology also has advantages over others like Parsortix in this arena, because its unidirectional flow allows a very flexible output.
"We have several adaptors that allow you to output to a slide chamber, an Eppendorf tube, to micro-wells, even to a petri dish," he said. "So with Eppendorf tubes for example, we can transfer that to a range of technologies for single cell isolating, whether DEPArray or single cells pickers."
According to Crouse the firm has customers using both of these approaches in their labs. He declined to detail the firm's installed base after launching the platform at the beginning of this year, but said that groups the firm has published with include Stanford, UCLA, and UC Berkeley.