Exploring lipid nanoparticle design spaces using self-regulating microfluidic machines and multiplexed in vivo biodistribution

bioRxiv (Cold Spring Harbor Laboratory)

Abstract

Abstract Delivering therapeutic mRNA relies on lipid nanoparticles (LNPs). Finding optimal process parameters for new lipid combinations in LNP formulations remains a challenge. In our work, we used an automated, self-regulating microfluidic platform that actively changes process parameters to tune LNP formulations for preset, desired quality standards. We tested new LNPs by swapping poly(ethylene glycol) (PEG) lipopolymers for alternatives based on poly(2-methyl-2-oxazoline) (PMeOx) and poly(2-ethyl-2-oxazoline) (PEtOx). For each lipopolymer variant, the platform independently identified optimal production conditions in four or fewer iterative cycles, yielding particles of the preset size and high mRNA encapsulation. Small-angle X-ray scattering revealed that smaller LNPs modified with PEtOx had more structural surface variety and higher mRNA loading efficiency. When multiplexing these formulations in mice, the PEtOx-containing LNPs accumulated more in bone marrow compared to those with PEG, indicating trends that the chemistry of the lipopolymer affects the biodistribution of the resulting LNPs. By combining automated formulation and in vivo multiplexed testing, our approach provides a practical way to rapidly plan, formulate, and evaluate large pharmaceutical design spaces, to select excipients and process parameters yielding optimal biological performance of LNPs. Abstract Figure

Authors 14

1. Josef Kehrein

   University of Würzburg

   Affiliation as printed

   Institute for Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, 97074 Würzburg, Germany

2. Elena Reus

   University of Würzburg

   Affiliation as printed

   Institute for Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, 97074 Würzburg, Germany

3. Caroline T. Holick

   Friedrich Schiller University Jena

   Affiliation as printed

   Jena Center for Soft Matter (JCSM), Friedrich Schiller University, Philosophenweg 7, 07743 Jena, Germany

   Laboratory of Organic and Macromolecular Chemistry (IOMC), Friedrich Schiller University, Humboldtstraße 10, 07743 Jena, Germany

4. Sebastian Mummel
   Affiliation as printed

   NGP Polymers GmbH, Botzstraße 5, 07743 Jena, Germany

5. Ilayda Ates

   Bayer (Germany)

   Affiliation as printed

   Pharmaceutical Sciences, Research & Development, Bayer Pharmaceuticals, Müllerstraße 178, 13353 Berlin, Germany

6. Markus Hafke

   Bayer (Germany)

   Affiliation as printed

   Pharmaceutical Sciences, Research & Development, Bayer Pharmaceuticals, Müllerstraße 178, 13353 Berlin, Germany

7. Jonas Käsbach Aachen Chair of Bioprocess Engineering

   RWTH Aachen University

   Affiliation as printed

   Chair of Bioprocess Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074 Aachen, Germany

8. Christine Weber

   Friedrich Schiller University Jena

   Affiliation as printed

   Jena Center for Soft Matter (JCSM), Friedrich Schiller University, Philosophenweg 7, 07743 Jena, Germany

   Laboratory of Organic and Macromolecular Chemistry (IOMC), Friedrich Schiller University, Humboldtstraße 10, 07743 Jena, Germany

9. Tessa C. Lühmann

   University of Würzburg

   Affiliation as printed

   Institute for Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, 97074 Würzburg, Germany

10. Stephanie Schubert

    Friedrich Schiller University Jena

    Affiliation as printed

    Jena Center for Soft Matter (JCSM), Friedrich Schiller University, Philosophenweg 7, 07743 Jena, Germany

    Laboratory of Organic and Macromolecular Chemistry (IOMC), Friedrich Schiller University, Humboldtstraße 10, 07743 Jena, Germany

    NGP Polymers GmbH, Botzstraße 5, 07743 Jena, Germany

11. Jørgen Magnus Aachen Chair of Bioprocess Engineering

    RWTH Aachen University

    Affiliation as printed

    Chair of Bioprocess Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074 Aachen, Germany

12. Florian A. Mann

    Bayer (Germany)

    Affiliation as printed

    Pharmaceutical Sciences, Research & Development, Bayer Pharmaceuticals, Müllerstraße 178, 13353 Berlin, Germany

13. Ulrich S. Schubert

    Friedrich Schiller University Jena

    Affiliation as printed

    Jena Center for Soft Matter (JCSM), Friedrich Schiller University, Philosophenweg 7, 07743 Jena, Germany

    Laboratory of Organic and Macromolecular Chemistry (IOMC), Friedrich Schiller University, Humboldtstraße 10, 07743 Jena, Germany

14. Lorenz Meinel corresponding

    University of Würzburg · Helmholtz Institute for RNA-based Infection Research

    Affiliation as printed

    Helmholtz Institute for RNA-based Infection Research (HIRI), Josef-Schneider-Strasse 2, 97080 Würzburg, Germany

    Institute for Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, 97074 Würzburg, Germany