NEW YORK ─ Italian researchers are developing a photonic biosensor testing platform that they believe could prove especially useful for clinicians prescribing colorectal cancer therapies by eliminating time-consuming steps associated with traditional liquid biopsy testing technologies.
The test uses surface plasmon resonance imaging (SPRI) to detect circulating DNA RAS mutations in patients' blood that are associated with resistance to colorectal cancer treatments.
In a preliminary study, published recently in Biosensors and Bioelectronics, the group described the development of an ultrasensitive nanoparticle-enhanced plasmonic biosensor for detecting RAS single nucleotide variants in plasma.
The technology has the potential for implementation as a laboratory-developed test within about three years, Giuseppe Spoto, a professor of chemical sciences at the University of Catania and corresponding author on the study, said in an interview.
SPRI is an optical detection technique for analyzing biomolecular interactions that the researchers adopted for its potential to detect nucleic acid mutations and protein biomarkers associated with many cancer types, including lung, liver, and colorectal cancers.
The test uses a combination of peptide nucleic acid (PNA) probes and gold nanoparticles to enable high sensitivity without special sample preparation and amplification, according to the researchers.
The PNA probes are synthetic substitutes for oligonucleotide probes, short stretches of single-stranded DNA frequently used in genetic testing to detect the presence of complementary, target nucleic acid sequences.
In the SPRI platform, the PNA probes are immobilized on the sensor surface, and plasma, after being separated from blood, flows over the surface and the probes hybridize to complementary RAS sequences.
The platform also uses gold nanoparticles functionalized with oligonucleotides whose sequences are complementary to a separate portion of the target RAS sequence, to enhance plasmon detection.
Surface plasmon resonance imaging is deployed to detect interactions between the probes and the target DNA to distinguish patients whose tumors have specific RAS mutations from patients with normal DNA.
The results of the study on a relatively limited number of samples would need to be successfully validated in a broader population of patients prior to the launch of a commercial test. However, the assay has so far detected DNA mutations with high sensitivity from a single drop of blood, requiring "at least an order of magnitude" less volume than is needed to run tests using traditional liquid biopsy detection technologies, such as PCR and next-generation sequencing, Spoto said.
Such a reduction in volume could prove vital for patients who are fragile and for whom frequent blood draws are risky, he noted.
In the recent study, the researchers observed an area under the curve (AUC) of 0.947 with 100 percent sensitivity and 83.33 percent specificity in tissue samples obtained from 20 colorectal cancer patients and 10 healthy patients, according to Spoto.
In addition, in plasma samples spiked with mutant DNA, the group saw attomolar-level analytical sensitivity levels that are suitable to meet most clinical needs, he added.
Andreas Weinhäusel, a researcher of molecular diagnostics technologies at the AIT Austrian Institute of Technology in Vienna who was not involved with the study, said in an interview that it demonstrates the direct detection of mutations without prior amplification "not only for simple tests but for the very challenging analysis of cfDNA from plasma."
Weinhäusel said that the development of the technology for commercial diagnostic testing applications appears feasible. The development of "an SPRI-measuring system together with the microfluidic measuring chips for implementation for a wide range of molecular biological analysis applications, right up to routine clinical laboratories, can safely be implemented at short notice with today's established manufacturing options," he said.
Aline Martayan, director of the NGS lab in the clinical pathology unit at IRCCS Regina Elena National Cancer Institute who was also not involved with the study, said that the technology could help eliminate an anticipated bottleneck in testing if regulatory authorities approve tests for the detection of RAS mutations in the blood of colorectal cancer patients.
"When liquid biopsy will hopefully become routine, I foresee the potential for a major bottleneck, both pre-analytical and analytical," said Martayan. "With each patient being monitored for resistance multiple times during treatment and throughout disease progression … our routines will likely grow exponentially. If plasmonic detection technology holds its promise, it may … drastically simplify upstream blood processing and downstream all-RAS testing."
Plasmonic detection may lead to "critical advances abating labor, turnaround times, and ultimately cost, while enhancing throughput at the same time," she added.
Spoto said that the researchers are working toward validating the results of the study in a broader patient population and developing the biosensor for clinical use. He and his colleagues are sketching plans to commercialize the platform and seeking partnerships with in vitro diagnostic companies that may be interested in using a laboratory-developed test as part of a testing service in a certified laboratory.
The development of the plasmonic detection technology stems from an EU-sponsored research project launched in 2017 to develop and validate a plasmonic testing platform that integrates DNA, microRNA, and protein-based tumor autoantibody detection from blood.
A multinational consortium involving research organizations, companies, and the Regina Elena Italian National Cancer Institute collaborated to develop an industrial prototype test for cancer diagnosis and monitoring. The current project aims to develop one of the first commercial applications for the prototype, Spoto said.
The promise of liquid biopsy testing for colorectal cancer is stimulating investment, regulatory approvals, and new companies.
For example, InterVenn Biosciences announced Monday it has raised $34 million in a Series B financing round to commercialize its ovarian cancer diagnostic. The funding will also be used to accelerate development for its immune-oncology treatment response and colorectal cancer indications.
Last week, Clinical Genomics announced that its Colvera liquid biopsy test for patients with colorectal cancer is now covered under Medicare Administrative Contractor Novitas Solution's revised local coverage determination.
Genetron Health said recently that its eye is firmly on liquid biopsy testing, especially for early cancer detection and monitoring. The firm has been conducting internal research targeting colorectal cancer, with data expected in 2021.
Colorectal cancer is also a prime target for many potential competitors, including Exact Sciences, Freenome, Guardant Health, and Singlera.
Spoto said that the Italian researchers are continuing development work on the assay and its underlying platform, including investigating the detection of new mutations associated with undisclosed medical conditions using different configurations of the PNA probes.
Additional validation of the blood-based assay in real-world colorectal cancer patient samples is ongoing through a collaboration with the Regina Elena National Cancer Institute, which on a daily basis provides PCR- and NGS-based liquid biopsy testing for patients with colorectal and other cancers.
The researchers are also in discussions with companies that are part of their European consortium and other firms outside Europe that may be interested in taking the technology to market, Spoto said. The group is seeking opportunities to license its technology to companies with experience in taking diagnostic testing technology through regulatory approvals, he added.
Weinhäusel said he believes the plasmonic technology and process developed by the Italian research team can eventually be applied broadly to detect mutations in cfDNA for many cancer diagnostic applications. However, there needs to be a willingness to invest in the technology by funding agencies and diagnostic industry companies for that to happen, he noted.
Martayan added that the major challenge for use of photonic technologies, such as plasmonics, for liquid biopsy-based diagnostic testing will be to generate enough of an impact to "overturn a market dominated by conventional approaches," including NGS.
"Photonics experts will have to make a cogent case that their techniques are sensitive, specific, reliable, and robust in a routine clinical pathology lab," she said. "But the advantages are so many and the idea of diagnosing cancer with light is so appealing that these technologies are among the best candidates for next next-generation Dx."