O2164: Ecirp Scavenger, MOP3, Is a Novel Therapeutic to Protect Against Neonatal Sepsis-induced Intestinal Inflammation and Injury

Background: Sepsis is characterized by a dysregulated host response to infection. It can lead to life-threatening organ injury and is associated with a 10% mortality in infants. Previously, milk fat globule-EGF factor VIII (MFG-E8) has been shown to be beneficial in inflammatory states, particularly in models of neonatal inflammation and systemic sepsis. On the other hand, the new damage-associated molecular pattern, extracellular cold-inducible RNA binding protein (eCIRP), worsens inflammation and is elevated in neonatal sepsis. Although previous studies have investigated MFG-E8-based therapeutics in neonatal inflammation, it is unknown whether eCIRP-antagonism will effectively prevent devastating inflammation and organ injury. Furthermore, given that there is no effective drug to target intestinal injury in sepsis, the aim of this study was to determine the therapeutic potential of a novel, MFG-E8-derived oligopeptide (MOP3) designed to clear eCIRP and protect against intestinal inflammation and injury sepsis.

Methods: Computational modeling was used to determine the likely domain of MFG-E8’s binding to eCIRP. The known amino acid (AA) sequence of this domain was used to develop and test potential therapeutic peptides. An MFG-E8 derived oligopeptide, MOP3, was developed based on 15 AA from MFG-E8 discoidin 2 domain (mimicking MFG-E8’s C-terminal) and tagged with the 3 AA, RGD, (mimicking MFG-E8’s binding capacity for the integrin receptor at the N-terminal). Thus, the peptide MOP3 was designed to bind eCIRP and link eCIRP to phagocytes through RGD-based binding for uptake. In a preclinical model of neonatal sepsis, mouse pups (5 days old) were intraperitoneally injected with cecal slurry (CS) consisting of enteric contents from adult mice. At the time of CS injection, pups were treated with either MOP3 (10 mg/kg, intraperitoneal injection) or vehicle. After 10 hours, intestines were collected and analyzed for injury and by qPCR, histopathologic analysis (assessed by Chiu’s score), apoptosis (assessed by TUNEL assay), and gap junctional protein expression, ZO-1 (assessed by immunofluorescence staining).

Results: MOP3 treatment protected septic neonatal mice from intestinal inflammation by reducing mRNA expression of IL-6, IL-1β, TNFα, and CXCL2 by 56.7%, 82.3%, 30.3%, and 75.2%, respectively (Table). MOP3 also protected against histopathologic injury by significantly reducing Chiu’s score by 51.9% and reducing the number of apoptotic cells in the intestine by 44.0% (Table 1, Figure 1A, B). Finally, MOP3 treatment was associated with the preservation of intestinal tight junction protein expression, ZO-1, as demonstrated by immunofluorescence staining (Figure 1C).

Conclusion: Through the specific design of binding and clearing eCIRP from circulation, MOP3 effectively protects against sepsis-induced intestinal inflammation and injury in a pre-clinical model of neonatal sepsis. Given the advantages of synthetic peptides as therapeutics, MOP3 provides a promising new treatment to target intestinal injury and improve outcomes in neonatal sepsis.