TY - JOUR
T1 - Investigation of pH-Responsiveness inside Lipid Nanoparticles for Parenteral mRNA Application Using Small-Angle X-ray Scattering
AU - Uebbing, Lukas
AU - Ziller, Antje
AU - Siewert, Christian
AU - Schroer, Martin A.
AU - Blanchet, Clement E.
AU - Svergun, Dmitri I.
AU - Ramishetti, Srinivas
AU - Peer, Dan
AU - Sahin, Ugur
AU - Haas, Heinrich
AU - Langguth, Peter
N1 - Funding Information:
This research was funded by the “Deutsche Forschungsgemeinschaft DFG” as part of the collaborative research center (CRC) 1066. The DFG had no involvement in study design, collection, analysis, interpretation of data, writing of the report, and in the decision to submit the article for publication. DLin-MC3-DMA was a gift from the Dan Peer Group (Tel Aviv University, Israel).
Publisher Copyright:
© 2020 American Chemical Society.
Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
PY - 2020/11/10
Y1 - 2020/11/10
N2 - Messenger ribonucleic acid (mRNA)-based nanomedicines have shown to be a promising new lead in a broad field of potential applications such as tumor immunotherapy. Of these nanomedicines, lipid-based mRNA nanoparticles comprising ionizable lipids are gaining increasing attention as versatile technologies for fine-tuning toward a given application, with proven potential for successful development up to clinical practice. Still, several hurdles have to be overcome to obtain a drug product that shows adequate mRNA delivery and clinical efficacy. In this study, pH-induced changes in internal molecular organization and overall physicochemical characteristics of lipoplexes comprising ionizable lipids were investigated using small-angle X-ray scattering and supplementary techniques. These changes were determined for different types of ionizable lipids, present at various molar fractions and N/P ratios inside the phospholipid membranes. The investigated systems showed a lamellar organization, allowing an accurate determination of pH-dependent structural changes. The differences in the pH responsiveness of the systems comprising different ionizable lipids and mRNA fractions could be clearly revealed from their structural evolution. Measurements of the degree of ionization and pH-dependent mRNA loading into the systems by fluorescence assays supported the findings from the structural investigation. Our approach allows for direct in situ determination of the structural response of the lipoplex systems to changes of the environmental pH similar to that observed for endosomal uptake. These data therefore provide valuable complementary information for understanding and fine-tuning of tailored mRNA delivery systems toward improved cellular uptake and endosomal processing.
AB - Messenger ribonucleic acid (mRNA)-based nanomedicines have shown to be a promising new lead in a broad field of potential applications such as tumor immunotherapy. Of these nanomedicines, lipid-based mRNA nanoparticles comprising ionizable lipids are gaining increasing attention as versatile technologies for fine-tuning toward a given application, with proven potential for successful development up to clinical practice. Still, several hurdles have to be overcome to obtain a drug product that shows adequate mRNA delivery and clinical efficacy. In this study, pH-induced changes in internal molecular organization and overall physicochemical characteristics of lipoplexes comprising ionizable lipids were investigated using small-angle X-ray scattering and supplementary techniques. These changes were determined for different types of ionizable lipids, present at various molar fractions and N/P ratios inside the phospholipid membranes. The investigated systems showed a lamellar organization, allowing an accurate determination of pH-dependent structural changes. The differences in the pH responsiveness of the systems comprising different ionizable lipids and mRNA fractions could be clearly revealed from their structural evolution. Measurements of the degree of ionization and pH-dependent mRNA loading into the systems by fluorescence assays supported the findings from the structural investigation. Our approach allows for direct in situ determination of the structural response of the lipoplex systems to changes of the environmental pH similar to that observed for endosomal uptake. These data therefore provide valuable complementary information for understanding and fine-tuning of tailored mRNA delivery systems toward improved cellular uptake and endosomal processing.
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U2 - 10.1021/acs.langmuir.0c02446
DO - 10.1021/acs.langmuir.0c02446
M3 - Article
C2 - 33108188
AN - SCOPUS:85095853177
VL - 36
SP - 13331
EP - 13341
JO - Langmuir
JF - Langmuir
SN - 0743-7463
IS - 44
ER -