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Little is known about if and how circular RNAs (circRNAs) are involved in skeletal muscle atrophy. Here a conserved circular RNA Damage‐specific DNA binding protein 1 (circDdb1), derived from the host gene encoding Damage‐specific DNA binding protein 1 (DDB1), as a mechanism of muscle atrophy is identified. circDdb1 expression is markedly increased in a variety of muscle atrophy types in vivo and in vitro, and human aging muscle. Both in vivo and in vitro, ectopic expression of circDdb1 causes muscle atrophy. In contrast, multiple forms of muscle atrophy caused by dexamethasone, tumor necrosis factor‐alpha (TNF‐α), or angiotensin II (Ang II) in myotube cells, as well as by denervation, angiotensin II, and immobility in mice, are prevented by circDdb1 inhibition. Eukaryotic initiation factor 4A3 (EIF4A3) is identified as a regulator of circDdb1 expression in muscle atrophy, whereas circDdb1 encodes a novel protein, circDdb1‐867aa. circDdb1‐867aa binds with and increases the phosphorylation level of eukaryotic elongation factor 2 (eEF2) at Thr56 to reduce protein translation and promote muscle atrophy. In summary, these findings establish circDdb1 as a shared regulator of muscle atrophy across multiple diseases and a potential therapeutic target. [ABSTRACT FROM AUTHOR]

Aims Cardiotoxicity leading to heart failure (HF) is a growing problem in many cancer survivors. As specific treatment strategies are not available, RNA discovery pipelines were employed and a new and powerful circular RNA (circRNA)-based therapy was developed for the treatment of doxorubicin-induced HF. Methods and results The circRNA sequencing was applied and the highly species-conserved circRNA insulin receptor (Circ-INSR) was identified, which participates in HF processes, including those provoked by cardiotoxic anti-cancer treatments. Chemotherapy-provoked cardiotoxicity leads to the down-regulation of Circ-INSR in rodents and patients, which mechanistically contributes to cardiomyocyte cell death, cardiac dysfunction, and mitochondrial damage. In contrast, Circ-INSR overexpression prevented doxorubicin-mediated cardiotoxicity in both rodent and human cardiomyocytes in vitro and in a mouse model of chronic doxorubicin cardiotoxicity. Breast cancer type 1 susceptibility protein (Brca1) was identified as a regulator of Circ-INSR expression. Detailed transcriptomic and proteomic analyses revealed that Circ-INSR regulates apoptotic and metabolic pathways in cardiomyocytes. Circ-INSR physically interacts with the single-stranded DNA-binding protein (SSBP1) mediating its cardioprotective effects under doxorubicin stress. Importantly, in vitro transcribed and circularized Circ-INSR mimics also protected against doxorubicin-induced cardiotoxicity. Conclusion Circ-INSR is a highly conserved non-coding RNA which is down-regulated during cardiotoxicity and cardiac remodelling. Adeno-associated virus and circRNA mimics-based Circ-INSR overexpression prevent and reverse doxorubicin-mediated cardiomyocyte death and improve cardiac function. The results of this study highlight a novel and translationally important Circ-INSR-based therapeutic approach for doxorubicin-induced cardiac dysfunction. [ABSTRACT FROM AUTHOR]

This chapter serves as a guide for researchers embarking on circular RNA-based translational studies. It provides a foundation for the successful encapsulation of circular RNA into lipid nanoparticles (LNPs) and facilitates progress in this emerging field. Crucial scientific methods and techniques involved in the formulation process, particle characterization, and downstream processing of circ-LNPs are covered. The production of in vitro transcribed circular RNA-containing LNPs based on a commercially available lipid mix is provided, in addition to the fundamentals for successful encapsulation based on lipid mixes composed of single components. Furthermore, the transfection and validation protocols for the identification of a functional and potentially therapeutic circRNA candidate for initial in vitro verification, before subsequent LNP studies, are explained., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)