Research ArticleMATERIALS SCIENCE

Functionalized lipid-like nanoparticles for in vivo mRNA delivery and base editing

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Science Advances  21 Aug 2020:
Vol. 6, no. 34, eabc2315
DOI: 10.1126/sciadv.abc2315
  • Fig. 1 Design and screenings of FTT LLNs.

    (A) Chemical structures of the FTT derivatives. FTT1 to FTT10 compounds share the same core structure and have three different types of lipid side chains, which are presented as gray (carbon chain), blue (branched ester chain), and green (linear ester chain), respectively. (B) Size and polydispersity index (PDI) of FLuc mRNA–encapsulated FTT1 to FTT10 LLNs for in vivo screenings. (C) In vivo mRNA delivery efficiency of FTT1 to FTT10 LLNs, represented as fold of increase in the luminescence intensity of FTT LLNs to that of TT3 LLNs in the livers of mice (n = 2).

  • Fig. 2 Characterization of FTT5 LLNs.

    (A) Size distribution of FTT5 LLNs measured by DLS. (B) mRNA encapsulation efficiency (EE) and zeta potential (ZP) of FTT5 LLNs. (C) Cryo-TEM image of an FTT5 LLN. Scale bar, 50 nm. (D and E) Biodegradation of FTT5 and FTT9 LLNs in the liver and blood of mice within 48 hours, respectively (n = 3). (F) Fold of changes in the cellular uptake of FTT5 LLNs with the treatment of different endocytic inhibitors, EIPA, MβCD, and CPZ, as compared with control group (n = 3; two-tailed Student’s t test; *P < 0.05; **P < 0.01). (G) Confocal microscopy images on the endosomal escape of FTT5 LLNs. Diffusion of calcein was observed in the cytosol of cells in the presence of FTT5 LLNs containing Alexa Fluor 647–labeled RNA, suggesting that FTT5 LLNs mediated a rupture of endosomal membranes and thus led to a release of the RNAs from endosomes. Scale bars, 10 μm.

  • Fig. 3 In vivo hFVIII mRNA delivery using FTT5-LLNs.

    (A) hFVIII protein level in WT mice 6 and 12 hours after intravenous administration of FTT5-hFVIII mRNA LLNs at an mRNA dose of 2 mg/kg (n = 3). (B) hFVIII protein level and (C) activity in hemophilia A (HA) mice 6 and 12 hours after intravenous administration of FTT5-hFVIII mRNA LLNs at an mRNA dose of 2 mg/kg (n = 3). (D) Histopathological images from hemophilia A mice treated with FTT5-hFVIII mRNA LLNs and untreated hemophilia A mice as a control. Scale bars, 100 μm.

  • Fig. 4 In vivo base editing of PCSK9 gene using FTT5 LLNs.

    (A) In vivo dose-response curve of base editing of the PCSK9 gene (n = 5). (B) Representative targeted deep sequencing data from a mouse treated at a dose of 0.125 mg/kg.

Supplementary Materials

  • Supplementary Materials

    Functionalized lipid-like nanoparticles for in vivo mRNA delivery and base editing

    Xinfu Zhang, Weiyu Zhao, Giang N. Nguyen, Chengxiang Zhang, Chunxi Zeng, Jingyue Yan, Shi Du, Xucheng Hou, Wenqing Li, Justin Jiang, Binbin Deng, David W. McComb, Robert Dorkin, Aalok Shah, Luis Barrera, Francine Gregoire, Manmohan Singh, Delai Chen, Denise E. Sabatino, Yizhou Dong

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    • Synthesis of FTT compounds
    • Figs. S1 to S7
    • Table S1
    • References

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