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Angle Liquid Biopsy Cell Capture Might Also Be Useful in Tissue, Study Finds


NEW YORK (GenomeWeb) – As investigators continue to work to adapt techniques originally developed for analyzing tissue samples to liquid biopsy, a small study published late last year has suggested that the reverse might also be true in some cases — that tools developed for analysis of blood and other body fluids can aid in the pathological and molecular analysis of tissue.

Investigators from Germany's Fraunhofer Institute for Toxicology and Experimental Medicine reported in the International Journal of Cancer that the Angle's Parsortix system, a device thus far marketed mainly for isolation of circulating tumor cells from blood samples, helped them better detect the presence of cancer cells in samples of lymph node tissue in melanoma patients. The application of the CTC technology could also make it easier to perform genomic analyses of cancer cells that spread to the lymph nodes than using tissue prepared as FFPE specimens.

Angle continues to work to highlight wide applicability of its system not just in cancer liquid biopsy, but for detecting rare cells in body fluids in other contexts like prenatal testing, but CEO Andrew Newland said in an interview that the Fraunhofer study is "the first time that anybody has used it [in tissue analysis]."

Bernhard Polzer, the study's lead author and head of Fraunhofer's department of personalized tumor therapy, said this week that the group's initial results suggest that the method could help improve patient outcomes by ensuring better detection of metastasis, and providing more opportunity for genomic analysis and targeted therapy in the adjuvant setting.

He and his colleagues are already extending the approach to other cancer types and planning a prospective follow-up.

Examining lymph nodes is a crucial and standardized part of the care of melanoma patients, as it is used to guide decisions about drugs and other therapies in addition to surgical removal. "This can make quite a big difference to the outcome for the patient really," Newland argued. "Obviously it's important to make the right decision whether to do chemotherapy or not." And the same is true to various degrees for several other cancers, including head and neck tumors and lung cancer.

In current practice, clinicians remove one or more lymph nodes that are closest to the site of a melanoma or other cancer, fix the tissue, embed it in paraffin wax, slice thin sections, and stain them for microscopic analysis for evidence of cancer cells.

Polzer and his colleagues have been working for several years to try to improve the ability to detect the presence of cancer cells in lymph nodes, called "disseminated cancer cells" in this context. And they were successful initially, with a method of taking the lymph node tissue and — instead of fixing and sectioning it — "disaggregating" it and smearing the resulting cell suspension onto slides.

The concept made sense, Polzer said, because lymph node tissue is relatively "loose" compared to other structures. "With no-so-forceful mechanical disaggregation you can get a single-cell suspension," he explained.

In a 2014 study, the group performed a parallel analysis — splitting lymph nodes into two and evaluating each patient's node using both FFPE analysis and the disaggregation method. The group found that their "quantitative immunocytology" approach was better at predicting melanoma outcomes than standard FFPE histology analysis.

As important as this improvement was in assessing lymph nodes for signs of cancer, Polzer and colleagues have argued that the disaggregation of samples can make it possible to do genomic or other testing, even in cases where the amount of cancer cells present is too minute for the same to be done with FFPE samples.

The problem that the researchers hoped to address using Angle's CTC platform was that their initial approach was too time consuming and expensive. "You get the single cell suspension and you have several million cells … which are sedimented on slides, stained, and you search through many, many slides," Polzer said. "But we usually could look at just two million cells per patient and had to stop there because the effort was just too much."

Before experimenting with Parsortix, the team tried other CTC platforms to enrich for tumor cells and reduce the number of slides needed, including antibody-based systems and membrane-based platforms, but they weren't able to make them work.

Membrane filtration, for example, couldn't handle the cell suspension from lymph node tissue without getting clogged, Polzer said.

Investigators in the current study didn't evaluate any other non-membrane microfluidic CTC platforms, though others have been developed for many of the same applications as Parsortix — Vortex Biosciences' VTX-1, for example.

Newland said that Angle now has about 200 Parsortix systems in use, with about a third of those in Angle labs or Angle-supported labs, and the rest disseminated more widely in the cancer research community, where the system grabbed the attention of the Fraunhofer group.

In their paper in the IJC, the researchers performed a variety of experiments examining the applicability of Parsortix to lymph node samples, testing various protocols to make it possible to not just separate disseminated cancer cells, but to make sure they could be removed from the device and molecularly profiled.

As a proof of concept, the team applied the resulting optimized workflow to samples from three melanoma patients, reporting a "high recovery [rate for DCCS] with harvest rates above 60 percent."

In addition, the group looked at the integrity of the transcriptomes and genomes of individual, isolated cancer cells after passage through the Parsortix instrument, and said they could detect expression of melanoma-associated transcripts and assess BRAF and NRAS mutational status.

"As the approach also allows isolation of viable cells, future studies may exploit the possibility for propagation of patient-derived cell models for preclinical and clinical studies in personalized medicine," the team added.

Newland said that independent German research labs have been experimenting with the Fraunhofer team's lymph node disaggregation approach in other cancers, including a team in Dusseldorf looking at head and neck cancers and esophageal cancer and a group in Tubingen also working in melanoma.

So far these groups have been manually examining unenriched slides, andPolzer and colleagues are the only team that has adopted Parsortix. But the recently published results could persuade others to use the system.

For their part, the Fraunhofer researchers are now piloting their approach in prostate and bladder cancer and are experimenting with other non-liquid applications of liquid biopsy, for example in disaggregating heterogeneous tumor tissue and sorting clean cancer cell subpopulations to allow for "really clean," or subspecific genetic analyses, Polzer said.

According to Newland, Angle has no plans at this time to pursue regulatory approval of its system for the lymph-node application being pursued by the Fraunhofer team, though with more evidence on the table that could change.

Polzer added that while the value of molecular profiling is becoming widely accepted in the pathology community, it might be challenging to get the clinic to widely embrace the team's lymph node disaggregation and Parsortix-based cell enrichment approach, even as the team collects more evidence.

"Many pathologists are conservative and really feel like seeing tissue architecture is important," he said. "Some pathologists we work with are very eager to do this, but it would be false to say that everyone is like this."

The team intends to do a prospective study to try to demonstrate that the improvement in the ability to find disseminated cancer cells, as well as to perform genetic testing for actionable biomarkers like BRAF also leads to improved outcomes for patients over standard pathology practice.