An engineered miRNA PS-OMe miR130 inhibits acute lung injury by targeting eCIRP in sepsis

Abstract

Background Sepsis is caused by the dysregulated immune response due to an initial infection and results in significant morbidity and mortality in humans. Extracellular cold inducible RNA binding protein (eCIRP) is a novel mediator identified in sepsis. We have previously discovered that microRNA 130b-3p inhibits eCIRP mediated inflammation. As RNA mimics are very unstable in vivo, we hypothesize that an engineered miRNA 130b-3p mimic named PS-OMe miR130, improves stability of the miRNA by protection from nuclease activity. We further hypothesize that PS-OMe miR130 reduces not only eCIRP-mediated inflammation and but also acute lung injury in a murine model of polymicrobial sepsis. Methods Single stranded PS-OMe miR130 was synthesized and the binding affinity to eCIRP was evaluated using surface plasmon resonance (SPR) and computational modeling. Macrophages were treated with PS-OMe miR130 with and without eCIRP and cell supernatant analyzed for cytokines. In vitro stability and the in vivo half-life of PS-OMe miR130 were also assessed. The effect of PS-Ome miR130 on eCIRP’s binding to TLR4 was evaluated by SPR analysis and modeling. Finally, the effect of PS-OMe miR130 on inflammation and injury was assessed in a murine model of sepsis. Results We demonstrate via SPR and computational modeling that PS-OMe miR130 has a strong binding affinity to eCIRP. This engineered miRNA decreases eCIRP induced TNF-α and IL-6 proteins, and it is highly stable in vitro and has a long in vivo half-life. We further demonstrate that PS-OMe miR130 blocks eCIRP binding to its receptor TLR4. Finally, we show that PS-OMe miR130 inhibits inflammation and lung injury, and improves survival in murine sepsis. Conclusion PS-OMe miR130 can be developed as a novel therapeutic by inhibiting eCIRP-mediated inflammation and acute lung injury in sepsis.

Novel Ps-Ome Mirna130b-3p Reduces Inflammation and Injury and Improves Survival After Renal Ischemia-Reperfusion Injury

“…However, given that any miRNA is unstable in vivo because of its susceptibility to degradation by nucleases, miRNA 130b-3p was chemically engineered in an in effort to improve its stability and thereby increase its potential efficacy (41). Recently, this modified miRNA, which we termed PS-OME miR130, has shown to reduce inflammation and tissue injury in mice, which underwent either hepatic I/R or cecal ligation puncture sepsis (19,20). It has been shown that using surface plasmon resonance and three-dimensional computational modeling, PS-OME miR130 retained its binding affinity to eCIRP and that the binding was comparable to that which was observed with the unmodified miRNA 130b-3p (19).…”

“…Recently, this modified miRNA, which we termed PS-OME miR130, has shown to reduce inflammation and tissue injury in mice, which underwent either hepatic I/R or cecal ligation puncture sepsis (19,20). It has been shown that using surface plasmon resonance and three-dimensional computational modeling, PS-OME miR130 retained its binding affinity to eCIRP and that the binding was comparable to that which was observed with the unmodified miRNA 130b-3p (19). Importantly, studies on the half-life of the unmodified mimic were unsuccessful because it was undetectable in serum, whereas the modified miRNA was found to have a half-life of 277.2 min (18,19).…”

“…Immediately after the closure, mice were randomly allocated to be administered intravenously, via retro-orbital injection, of PBS (vehicle) or 12.5 nmol of PS-OME miR130 (treatment) in both the 24-h experiment and the 10-day survival experiment. The dosing was based on previous studies showing an effective reduction in inflammatory markers in both unmodified and modified miRNA 130b-3p mimics in other injury models (11,19,20). Analgesia was provided by delayed subcutaneous injection of buprenorphine (50 μg/kg), and resuscitation was provided by 500 μL bolus of normal saline.…”

“…An miRNA 130b-3p mimic was then synthesized, and its effectiveness in reducing inflammation was experimentally confirmed (11). To improve the in vivo miRNA stability, we have chemically engineered the miRNA 130b-3p mimic and designated as PS-OME miR130 (19,20). The selection of modifications was focused on using well-studied miRNA modification strategies and the presence of significant nuclease protective attributes.…”

“…Given that our compound acts specifically on an extracellular target, the focus is on preventing degradation without impeding its eCIRP specific binding affinity (34). In fact, this modified miRNA was shown to successfully reduce tissue injury and inflammation in rodent models of sepsis and hepatic I/R injury (19,20). Previously, we have shown that renal I/R results in elevated serum and tissue levels of eCIRP and additionally that, in CIRP knockout mice, there is an attenuated inflammation and injury response (35).…”