Research Project
10000005
Abstract In response to PA-15-065, the broad long-term goal of this project is to develop a portable radiation biodosimeter to be used at the point-of-care (POC) for assessing precise radiation exposure and susceptibility for acute radiation syndrome (ARS) within the first 7 days of exposure starting with ~250 µL of blood collected from a finger stick and a turnaround time (TAT) of less than 40 minutes. The device measures changes in circulating microRNA (miRNA), small RNA molecules 18?24 nucleotides long that are released into the circulatory system by cells throughout the body, as biomarkers. These molecules can provide a means to assess the systemic health of the individual via a minimally invasive blood draw of a small amount of blood. Exposure to radiation has been shown to alter expression specific miRNA sequences, thereby establishing that these biomarkers could be used as a predictive assay for ARS. To measure changes in these miRNA radiation biomarkers, we will develop a microRNA-based Radiation Biodosimeter (MiRAD) system, which combines (1) POC isolation of circulating miRNA from whole blood, (2) detection of miRNA abundance using a battery-operated, portable RT- qPCR system adapted for use at the POC, and (3) specialized data processing algorithms to estimate exposed dose based on the miRNA profile of a panel of key markers. In Phase I, we have demonstrated in non-human primate (NHP) and pilot clinical studies that circulating concentrations of specific miRNA biomarkers are sensitive to exposure to whole body ionizing radiation in a dose- and time-dependent manner. A small amount of whole blood (200?500 µL), taken in the field, is processed using a novel sample preparation kit we developed and commercialized (www.chromologic.com/mir-clear), that performs (a) chemo-mechanical isolation of plasma, and (b) extraction of circulating microRNA (miRNA) without the need for power, laboratory equipment, or a technical user. Specific miRNA radiation biomarkers are analyzed by the MiRAD PCR assay using commercial, off-the- shelf, battery-operated PCR machines. Based on the profile of the miRNA biomarker panel, MiRAD estimates the amount of absorbed radiation by the patient. During Phase I, we developed a robust analytical method and identified a panel of miRNA biomarkers capable of accurate estimation of exposed dose and asymptomatically predict the onset of hematopoiesis and ARS using mice, non-human primate (NHP), and clinical samples. Moreover, we found that key miRNA radiation biomarkers were present in sufficient abundance, and changed in response to radiation dose (0?6.5 Gy) with sufficient magnitude (up to 25-fold), to allow isolation from < 200 µL of plasma with our proprietary POC sample processing kit and quantification by COTS qPCR.
