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Title ARTIFICIAL CIRCULAR MIRNA SPONGES AS A NOVEL MIRNA INTERFERENCE TECHNOLOGY |
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Type Young Investigator Award |
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Theme ACC Asia & SCS 32nd Annual Scientific Meeting |
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Topic Basic Science |
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Main Author Annadoray Lavenniah1 2 |
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Presenting Author Annadoray Lavenniah1 2 |
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Co-Author Matthew Ackers-Johnson1 2 Yiqing Li1 2 Tuan Luu Danh Anh1 2 Tingsen Benson Lim1 2 Roger Foo1 2 |
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Department / Institution / Country Human Genetics 3 / Genome Institute of Singapore / Singapore1 Cardiovascular Research Institute / National University of Singapore / Singapore2 |
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Objective(s) Circular RNAs (circRNAs) are an emerging class of noncoding RNAs with a circular structure that confers nucleolytic resistance. Recently, circRNAs have been discovered to sequester microRNAs (miRs) and repress their activity. We hypothesise that artificial circRNA sponges (circmiRs) can be constructed to target miRs therapeutically, with low dosage requirement and extended half-lives compared to current alternatives. |
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Material and Method CircmiRs were expressed using plasmid vectors in which inverted repeats flanked a linear miR sponge sequence. In vitro testing of circmiRs was performed using luciferase rescue, RNA pull-down and stability assays. Adeno-associated viral vectors were used to deliver circmiRs to cardiomyocytes in vivo in a transverse aortic constriction (TAC) mouse pressure overload model. Cardiac function was monitored by echocardiography. T7 in vitro transcription was used to synthesise exogenous circmiRs for comparison with antagomiRs. |
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Result(s) A circmiR sponge was constructed and optimised to target known pro-hypertrophic miRs-132 and -212. CircmiR expression competitively inhibited miR-132 and -212 activity in luciferase rescue assays. circmiRs were more stable than linear miR sponges. Direct sequestration of target miR-132 and -212 was confirmed by RNA pull down. In vivo, circmiR delivery preserved cardiac function and suppressed induction of stress genes up to 4 weeks post-TAC. Finally, artificial circmiRs were successfully synthesised at high yields and showed greater efficacy than antagomiRs in inhibiting miR function. |
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Conclusion circmiRs were successfully expressed and showed inhibition of target miR-132 and -212 activity in vitro. Delivery into a TAC mouse model attenuated hypertrophic characteristics suggesting the potential efficacy of circmiR as a novel therapeutic tool in vivo. Follow-up studies are ongoing to validate the molecular inhibition of miR-132 and -212 in vivo, and to confirm the in vivo efficacy of in vitro synthesised circmiRs in comparison to current linear miRNA interference technologies. |