For every mutant KRAS gene circulating in plasma, there are as many as 10,000 wild-type copies. At the annual Advances in Genome Biology and Technology meeting held in Marco Island, Fla., in February, Boreal Genomics' Andre Marziali described how his company's OnTarget platform can selectively deplete wild-type alleles and enrich their mutant counterparts, even those that differ by a single base. The allele-enrichment system will enable researchers and clinicians to detect cancer-related mutations directly in plasma, he said at the time.
"Current mutation detection systems based on qPCR or next-generation sequencing are limited by enzymatic errors in PCR amplification from wild-type templates," Marziali now adds.
Using the firm's allele-enrichment system, which is based on a highly specific hybridization-based sequence enrichment technique, Marziali says he and his colleagues "took a commercial qPCR assay for BRAF mutation and made it work about 200-times better. For two molecules that differ by one nucleotide, we can enrich a million-fold now." Another benefit is that "we don't have to trade off yield for specificity," he adds. "We're getting yields in the 60 to 80 percent range with million-fold enrichments, and that's without PCR, so we're not generating any new strands that might have errors."
Boreal CEO Nitin Sood says a key benefit of the company's technology "is that we can knock down the wild-type template, eliminating the false positives that arise because of PCR-generated errors." And because of the non-invasive nature of the plasma-based testing, he adds that researchers and clinicians might use OnTarget to monitor "mutations that drive clinical decisions" over time. Sood says his company intends to produce pre-packaged panels for this system that will target actionable cancer mutations, like those for drug-resistance.
"Even with next-generation sequencing — deep sequencing — you'll hit a wall simply because of the error rates generated by PCR during preparation," Sood says. "Because we knock down wild type, we will improve the sensitivity of [sequencing]-based assays. We are going to be complementary to clinical sequencing."
Marziali adds that "any hope of cancer-monitoring by a non-invasive test in plasma requires [the ability] to enrich for the mutant."
Sood says Boreal will begin placing systems with early-access customers in April, and expects to have a full commercial launch of OnTarget by the end of year.