RainDance Targeted Sequencing and Digital PCR: ‘A Potential Game-Changer’
As Director of the West Midlands Regional Genetics Laboratory at Birmingham Women’s Hospital in England — the largest National Health Service genetics diagnostic laboratory, with 150 staff members serving some 5.5 million people — Mike Griffiths is responsible for betting on technologies that will improve monitoring, diagnosing, and understanding disease.
Griffiths has placed his latest bet on RainDance Technologies, which offers platforms in droplet PCR targeted sequencing and in droplet digital PCR. Both serve unmet needs in Griffiths’ diagnostic laboratory, and that’s why he has chosen to add them to his research technology portfolio. Ultimately, he says, these new instruments could dramatically change the way Griffiths’ team is able to care for patients across a range of diseases.
The genetics lab Griffiths oversees is an integrated, comprehensive facility that runs all types of diagnostic genetic services, from chromosome analysis to next-generation sequencing. Under Griffiths’ tenure, the lab has built a growing research division to evaluate technology platforms for potential use in diagnostics. “We are primarily a diagnostic laboratory service within the NHS, but in order to be a leading-edge diagnostics service you have to have a strong research arm as well,” says Griffiths, who has a research appointment as a Professor in the School of Cancer Sciences at the University of Birmingham.
The genetics lab, which also serves as a technology hub for the Cancer Research UK Stratified Medicine Programme, runs some 50,000 samples annually. Cancer diagnostics make up about 40 percent of the lab’s work (the majority of this falling into hemato-oncology), and the other 60 percent comes under the umbrella of the lab’s Germline Genetics program. Molecular pathology represents a growing area within the Cancer Genetics program, Griffiths notes.
The lab’s mandate makes it ideally suited to RainDance’s picodroplet PCR-based platforms. Griffiths’ team first acquired the RDT 1000™, an instrument in the company’s targeted sequencing line that allows users to prepare many genes at once for analysis on a next-gen sequencer. The West Midlands group designed a 54-gene panel for myeloid leukemia consisting of 1,200 amplicons and is running that on the RDT 1000 to prepare libraries for the Illumina MiSeq. “The way the technology works gives us relatively even amplification of each amplicon compared to other amplicon amplification methods,” Griffiths says, noting that this allows multiplexing more patients into a single run on the MiSeq than would be possible with other approaches. Part of his research team’s evaluation of any technology includes considering how it could be rolled out to the diagnostic lab, so expense is a real factor. Using the RainDance technology to reduce the amount of sequencing needed “keeps the costs down, and the cost of these tests will be very important in the context of the NHS,” Griffiths adds.
Deploying the RDT 1000 to analyze mutations in a tumor sample serves as a good illustration. “When you’re looking at tumor DNA you’ve still got variable amounts of DNA from normal cells in the background, and also potential subclonal heterogeneity, so the mutation might not be there in all the tumor,” Griffiths says, noting that this dramatically increases the read depth needed to ensure a comprehensive profile of the tumor’s mutations. Using other amplicon or capture enrichment methods may result in more variation in the amount of each target region to be sequenced. As a consequence, for example, in order to achieve a minimum read depth of 300x, you might need to generate an average read depth of 1,000x or more. But with more uniform amplification, you might only need to go as high as 500x. “The more consistent yield from the RainDance approach should provide the more even amplification required,” Griffiths says. In this scenario, his team would be able to sequence perhaps twice as many patients in a single MiSeq run than they would with a method that generates more variable amplification.
Griffiths’ lab was also the first to order the RainDrop™, the new droplet digital PCR platform from RainDance. He sees the technologies complementing each other and potentially giving his lab the ideal feedback loop for patients who need cancer monitoring based on mutations in the tumor; the platforms may ultimately have applications in prenatal diagnostics. The RDT 1000 targeted sequencing tool (or higher-throughput, multi-sample ThunderStorm™ targeted sequencing tool) can be used to assess mutations, and then the RainDrop digital PCR tool can be used as a highly sensitive detector of disease residue within cell-free DNA in serum or plasma. “That’s the vision: that we do genomic profiling of a tumor, find multiple mutations, and those mutations then are all potential targets for therapy or treatment stratification. In addition, we can use the mutations as biomarkers that we may ultimately be able to turn into assays for the RainDrop to monitor residual disease in each of these patients,” Griffiths says.
For instance, scientists could analyze a tumor’s mutation profile with targeted sequencing, and then use droplet digital PCR to screen the patient’s serum for those mutations before and after the tumor is surgically removed to ensure surgery is complete. “That also leads to the potential for longer-term surveillance for recurrence with just a blood test,” he adds.
This potential is based on the remarkable sensitivity of the RainDrop platform. With 5 million to 10 million picodroplets in each experiment, Griffiths says, the RainDrop offers at least one or two orders of magnitude of improvement over the sensitivity of RT-qPCR or other current standards. “There’s more statistical power in the assay when you’re looking for low-level detection,” he says. Griffiths points to the absolute quantification of RainDance’s digital PCR approach as another benefit: digital PCR results need no standard curve or external scale, and therefore are less likely to suffer from variation between labs seen in some RT-qPCR assays.
Ultimately, Griffiths says this level of sensitivity could allow RainDance’s digital PCR platform to replace numerous other tests — such as the thousands of BCR-ABL1 and JAK2 assays his lab runs each year — and rapidly expand the portfolio of quantifiable residual disease markers. The RainDrop platform might also open the doors for non-invasive prenatal diagnosis of single gene disorders on circulating fetal DNA in the maternal plasma. With this opportunity to monitor patients for a range of biomarkers, the RainDance technology “becomes a potential game-changer,” Griffiths says. “The possibilities just start growing.”
— Meredith Salisbury