PROJECT SUMMARY
Stroke is currently the leading cause of permanent disability and fifth leading cause of death in the United States.
Early and accurate recognition reduces mortality and morbidity by expediting access to neurocritical care.
Unfortunately, recognition of stroke during triage is dependent on symptom-based assessments that are often
unreliable, and up to 35% of patients are misdiagnosed at initial clinician contact. Thus, the development of
accurate biomarker-based screening tools that could be used to rapidly detect stroke in emergency medicine
settings could substantially reduce rates of mistriage, enable earlier access to intervention, and improve patient
outcomes. Numerous prior investigations have attempted to use blood measures of various proteins released from
damaged neural tissue as an indicator of stroke. Unfortunately, it is now known that many of the proteins targeted
in these prior studies exhibit a limited degree of enrichment in brain tissue, curtailing their diagnostic specificity.
Furthermore, they typically circulate at too low of levels to enable reliable detection using the immunoassay
platforms available for rapid blood testing in emergency medicine settings, blocking their path to clinical use.
Circular RNAs (circRNAs) are a family of circularly continuous non-coding transcripts that are resistant to RNase
degradation; they accumulate in long-lived cells such as neurons, and recent work has demonstrated that there
are variants which are truly found only in the brain. In addition to the potential for improved diagnostic specificity,
targeting brain-specific circRNAs released from damaged neural tissue could have significant translational
advantages over targeting the proteins studied for blood-based stroke recognition in the past. The amplification
methods used to measure nucleic acids are thousands of times more sensitive than the immunoassay techniques
used to measure proteins, which could dramatically increase the odds of detection. Furthermore, because these
circRNAs would be cell-free, they could be directly assayed in serum without upstream RNA isolation; this could
allow for rapid direct detection using qRT-PCR on existing hardware found in clinical labs, or even methods such
as loop-mediated isothermal amplification (LAMP) at the point-of-care. Despite these tremendous advantages,
brain-originating circRNAs have never been investigated as a potential source of biomarkers for stroke recognition.
In the work proposed here, we aim to use digital droplet PCR to evaluate the absolute circulating levels of five
brain-specific circRNAs in 500 patients with various cerebrovascular and non-cerebrovascular pathologies who
present with stroke-like symptoms at hospital admission, as well as develop a set of clinically viable custom rapid
qRT-PCR and LAMP assays targeting said circRNAs and evaluate their sensitivity and specificity for stroke. This
novel, innovative, and highly translational project addresses an area of dire clinical need; we fully expect the
assays generated as part of this work to establish the immediate technical and molecular foundation for the
downstream development of a pair of rapid stroke triage diagnostics: one that could be deployed for stat testing in
the clinical laboratory on existing hardware, and another that could be deployed at the point-of-care.
