Poly(sarcosine) and Poly(2-ethyl-2-oxazoline) Tethered To mRNA Provide Stealth Properties To mRNA Polyplexes

The immunogenicity of poly(ethylene glycol) (PEG) in nanomedicines gives rise to concerns regarding safety and efficacy, driving research and development of PEG alternatives. This study investigates the potential of PEG alternatives, including poly(sarcosine) (pSar) and poly(2-ethyl-2-oxazoline) (pEtOx), in polyplexes designed to deliver mRNA (mRNA). The density of these polymers on the polyplex surface was precisely controlled by mRNA engineering, which involved mRNA hybridizationto RNA oligonucleotides (OligoRNAs) conjugated to pSar, pEtOx, or PEG. By co-formulating linear poly(ethylene imine) (LPEI) and mRNA tethered with varying numbers of these OligoRNAs, polyplexes coated with varying densities of pSar, pEtOx, or PEG and hydrodynamic diameters below 100 nm were fabricated. The ζ-potential exhibited a shift towards a neutral charge following surface modification with these polymers.These polymers enhanced the resistance of polyplexes to aggregation following albumin treatment. Upon delivery to cultured cells, these polymers decreased the efficiency of polyplex cellular uptake and protein expression from the delivered mRNA. After systemic administration into mice, polyplexes without these polymers accumulated in the lungs, presumably due to the induction of blood cell agglomeration. In contrast, polyplexes containing PEG, pSar, or pEtOx alleviated blood cell agglomeration and enabled selective protein expression in the spleen. These results highlight the potential of pSar and pEtOx as promising PEG alternatives conferring stealth properties to mRNA polyplexes. Notably, this study also uncovers discrepancies among these polymers with respect to their stealth properties in physicochemical characterization, cultured cell experiments and animal studies. These findings provide a substantial rationale for further research into PEG alternatives.