Abstract Body (Do not enter title and authors here): Background: Hutchinson-Gilford Progeria Syndrome (HGPS) is caused by a mutation in LaminA (progerin), and is characterized by accelerated aging and death from coronary or carotid disease in the mid-teens. We have shown that vascular smooth muscle cells (VSMCs) and endothelial cells (ECs) derived from HGPS children manifest many of the hallmarks of aging including telomere erosion, reduced proliferation, impaired function, DNA damage and senescence markers, altered cellular and nuclear morphology, and an aberrant transcriptional profile. These hallmarks of aging are substantially reversed by treatment with telomerase (hTERT) linear RNA,with greater benefit in HGPS cells than the current therapy, lonafarnib. However, linear RNA has a short half-life, which necessitates frequent administration. By contrast, circular (circ) RNA is more stable than linear RNA, and with an internal ribosome entry site can be translated into protein. We hypothesized the hTERT circRNA would provide for longer duration of telomerase expression, and would have a greater benefit for HGPS ECs.
HGPS ECs were treated with hTERT linear or circ RNAs. A single treatment with 1 µg/ml hTERT circRNA reversed multiple stigmata of senescence . However, at day 28 post treatment, the benefit of hTERT circRNA exceeded that of hTERT linear RNA in all measured variables. hTERT circRNA provided for greater recovery of telomere length as determined by quantitative fluorescence in situ hybridization; induced a three-fold greater reduction in beta-gal positive cells and morphologically aberrant nuclei. In HGPS ECs, hTERT circRNA provided for a 2-3 fold greater reduction of senescentfactors, inflammatory cytokines and DNA damage signals, including Progerin, p16, p21, IL-1B, IL-6, IL-8 MCP-1 and gH2A.X. In addition, hTERT circRNA to a greater degree restored NO production, promoted cell proliferation, enhanced angiogenesis and improved LDL uptake . Importantly, mitochondrial functions, as evaluated by the oxidative stress marker (MitoSOX) and mitochondrial membrane potential marker (JC-1 staining), were restored more completely by hTERT circRNA.
Conclusion: hTERT circRNA is more effective than hTERT linear RNA in rejuvenating senescent ECs, possibly because of its longer half-life. The novel hTERT circRNA is a promising therapy for HGPS and other disorders associated with accelerated vascular aging.