PRECISE: accessible sample-to-answer RT-PCR detection of hepatitis C infection from whole blood

Columbia University Health Sciences, New York NY

Investigators

Abstract

PROJECT SUMMARY This proposal is in response to NOSI NOT-AI-23-001, which calls for new point-of-care HCV diagnostics. Most of global HCV burden is concentrated in low- and middle-income countries (LMICs), but the U.S. has witnessed a steady increase in infection rates in the past decade, with an estimated 140,000 new cases annually. Whereas direct-acting antivirals (DAAs) can over 95% of HCV-infected individuals, and are becoming more accessible, about 40% of infected people are unaware of their status. The current two-step diagnostic process requires an initial antibody screen followed by an expensive and time-consuming RNA test to confirm active infection. This cumbersome workflow requires multiple office visits for patients, resulting in delays in treatment initiation and significant patient follow-up loss. Our lab has been developing plasmonic-based thermocycling into a clinically useful method for fast multiplexed RT-PCR at the POC. In this approach, heating is achieved not externally via the Peltier effect, but rather internally via infrared excitation of nanoparticles; as a result, the heating is rapid, and powered with low- power robust optical components. We have gathered substantial preliminary data to validate the premise of this approach to work on clinical specimens (saliva and nasal swabs); the results (published in Nature Nanotechnology, 2022) showed the ability, within 25 minutes from sample to result, to detect SARS-CoV-2 with high sensitivity and specificity. Moreover, even in the presence of the plasmonic nanoparticles, we showed that real-time qPCR could be performed with accurate quantitative determinations of cycle threshold (Ct) values. This capability for sample-to-result analysis is faster than traditional PCR approaches using Peltier heating, and the instrumentation using readily available optics renders the approach suitable for community testing sites where rapid and accurate results are sought. This proposal integrates plasmonic PCR with plasma filtration (starting with whole blood) and magnetic bead-based nucleic-acid extraction (a technique we recently published called PRECISE, Lab Chip, 2024). The proposed combined method, PRECISE plasmonic PCR, takes advantage of special nanoparticles to facilitate a seamless sample-to-result workflow for the end user, suitable for the target community health settings to facilitate HCV diagnosis in a single patient visit to enable on-the-spot treatment. The target metrics would be the best in class among HCV nucleic-acid tests, meeting all of the criteria in the target product profile recommended by the NOSI for an “optimal” test except for the 15-minute turnaround time (our target is 30 minutes), and surpassing all “minimal” targets. The target metrics include cost considerations.