Novel vector for efficient siRNA delivery to lymphoblasts and melanoma based on genipin-spermine nanocarriers protected with hybrid erythrocyte membrane coating

Efficient delivery of small interfering RNA (siRNA) remains a significant challenge in gene therapy because of the instability, poor cellular uptake, and immunogenicity of the carriers. In this study, we developed a hybrid delivery system combining genipin-spermine-glycine nanoparticles (G10S5) with erythrocyte membrane vesicles (EMVs) doped with DPPC and DSPE-PEG2000. G10S5 nanoparticles offer robust siRNA complexation and biocompatibility but may suffer from rapid clearance and immune detection. By camouflaging G10S5-siRNA polyplexes with hybrid EMVs, we aimed to increase their cell uptake and delivery efficiency. Physicochemical characterization via DLS, FTIR, TEM, cryo-EM, and AFM confirmed successful coating and favorable nanoscale morphology. Solvatochromic fluorescence analysis via the fluorescence of G10S5 indicated efficient coating. The optimized formulations at a phosphate-to‑nitrogen (P/N) ratio of 1:12 exhibited excellent RNase A resistance, strong siRNA binding, and storage stability. Compared with uncoated controls, in vitro assays demonstrated significantly enhanced cellular uptake of hybrid-coated G10S5-siRNA, with distinct internalization mechanisms. Gene silencing efficiency was validated by targeting tdTomato in tdTomato-expressing B16F10 cells, which showed effective knockdown with minimal cytotoxicity. Further validation was achieved in lymphoblastoid cell lines by targeting FARSA that has recently been implicated in C9orf72 mutation mechanism in lymphoblastoid lines. Our findings establish hybrid membrane-camouflaged G10S5 nanoparticles as promising siRNA delivery platforms, addressing the limitations of conventional carriers by leveraging their natural membrane properties and polymeric versatility. This strategy opens new avenues for the development of biomimetic, nonviral nucleic acid therapeutics. • Genipin-spermine nanoparticles show strong siRNA binding and RNase resistance. • Novel hybrid coating enhances G10S5-siRNA biocompatibility and cellular uptake. • Coating confirmed via DLS, TEM, cryo-EM, AFM, and solvatochromic fluorescence. • tdTomato knockdown in B16F10 cells confirms gene silencing effectiveness • FARSA silencing in lymphoblastoid cells validates therapeutic potential.