Microfluidic formulation, cryoprotection and long-term stability of paclitaxel-loaded π electron-stabilized polymeric micelles

Journal of Controlled Release, vol. 375, pp. 614–626

Abstract

Controlled manufacturing and long-term stability are key challenges in the development and translation of nanomedicines. This is exemplified by the mRNA-nanoparticle vaccines against COVID-19, which require (ultra-)cold temperatures for storage and shipment. Various cryogenic protocols have been explored to prolong nanomedicine shelf-life. However, freezing typically induces high mechanical stress on nanoparticles, resulting in aggregation or destabilization, thereby limiting their performance and application. Hence, evaluating the impact of freezing and storing on nanoparticle properties already early-on during preclinical development is crucial. In the present study, we used prototypic π electron-stabilized polymeric micelles based on mPEG-b-p(HPMAm-Bz) block copolymers to macro- and microscopically study the effect of different cryoprotective excipients on nanoformulation properties like size and size distribution, as well as on freezing-induced aggregation phenomena via in-situ freezing microscopy. We show that sucrose, unlike trehalose, efficiently cryoprotected paclitaxel-loaded micelles, and we exemplify the impact of formulation composition for efficient cryoprotection. We finally establish microfluidic mixing to formulate paclitaxel-loaded micelles with sucrose as a cryoprotective excipient in a single production step and demonstrate their stability for 6 months at -20 °C. The pharmaceutical properties and preclinical performance (in terms of tolerability and tumor growth inhibition in a patient-derived triple-negative breast cancer xenograft mouse model) of paclitaxel-loaded micelles were successfully cryopreserved. Together, our efforts promote future pharmaceutical development and translation of π electron-stabilized polymeric micelles, and they illustrate the importance of considering manufacturing and storage stability issues early-on during nanomedicine development.

Authors 22

1. Rahaf Mihyar Aachen Institute for Experimental Molecular Imaging

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

2. Armin Azadkhah Shalmani Aachen Institute for Experimental Molecular Imaging

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

3. Viktor Wildt Aachen Institute for Experimental Molecular Imaging

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

4. Maryam Sheybanifard Aachen Institute for Experimental Molecular Imaging

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

5. Alec Wang Aachen Institute for Experimental Molecular Imaging

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

6. Jan‐Niklas May Aachen Institute for Experimental Molecular Imaging

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

7. Saba Shahzad

   Forschungszentrum Jülich · Ernst Ruska Centre

   Affiliation as printed

   Ernst Ruska-Centre for Microscopy and Spectroscopy with Electrons (ER - C -3): Structural Biology, Forschungszentrum Jülich, Wilhelm-Johnen-Straße, 52428 Jülich, Germany

8. Eva Miriam Buhl Aachen Institute of Pathology Electron Microscopy Facility

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Electron Microscopy Facility, Institute of Pathology, RWTH Aachen University Hospital, Pauwelsstrasse 30, 52074 Aachen, Germany

9. Stephan Rütten Aachen Institute of Pathology Electron Microscopy Facility

   RWTH Aachen University · Universitätsklinikum Aachen

   Affiliation as printed

   Electron Microscopy Facility, Institute of Pathology, RWTH Aachen University Hospital, Pauwelsstrasse 30, 52074 Aachen, Germany

10. Diana Behrens
    Affiliation as printed

    Experimental Pharmacology and Oncology GmbH, Robert-Roessle-Str. 10, 13125 Berlin, Germany

11. Wolfgang Walther
    Affiliation as printed

    Experimental Pharmacology and Oncology GmbH, Robert-Roessle-Str. 10, 13125 Berlin, Germany

12. Mattia Tiboni

    University of Urbino

    Affiliation as printed

    University of Urbino Carlo Bo, Department of Biomolecular Sciences, Via Ca' le Suore 2/4, 61029, Urbino (PU), Italy

13. Luca Casettari

    University of Urbino

    Affiliation as printed

    University of Urbino Carlo Bo, Department of Biomolecular Sciences, Via Ca' le Suore 2/4, 61029, Urbino (PU), Italy

14. Johannes F. Buyel

    BOKU University

    Affiliation as printed

    Institute of Bioprocess Science and Engineering (IBSE), University of Natural Resources and Life Sciences (BOKU), Department of Biotechnology, Muthgasse 18, 1190 Vienna, Austria

15. Cristianne J.F. Rijcken

    Cristal Therapeutics (Netherlands)

    Affiliation as printed

    Cristal Therapeutics, Oxfordlaan 55, 6229 EV Maastricht, the Netherlands

16. Wim E. Hennink

    Utrecht University

    Affiliation as printed

    Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences, Faculty of Science, Utrecht University, 3508 TB Utrecht, the Netherlands

17. Saskia Freifrau von Stillfried Aachen Institute of Pathology

    RWTH Aachen University · Universitätsklinikum Aachen

    Affiliation as printed

    Institute of Pathology, RWTH Aachen University Hospital, Pauwelsstrasse 30, 52074 Aachen, Germany

18. Fabian M. Kiessling Aachen Institute for Experimental Molecular Imaging

    RWTH Aachen University · Universitätsklinikum Aachen

    Affiliation as printed

    Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

19. Yang Mei Shi Aachen Institute for Experimental Molecular Imaging

    RWTH Aachen University · Universitätsklinikum Aachen

    Affiliation as printed

    Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

20. Josbert M. Metselaar Aachen Institute for Experimental Molecular Imaging

    RWTH Aachen University · Universitätsklinikum Aachen

    Affiliation as printed

    Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany

21. Twan Lammers Aachen Institute for Experimental Molecular Imaging

    RWTH Aachen University · Universitätsklinikum Aachen

    Affiliation as printed

    Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany. Electronic address: tlammers@ukaachen.de

22. Quim Peña Aachen Institute for Experimental Molecular Imaging

    RWTH Aachen University · Universitätsklinikum Aachen

    Affiliation as printed

    Institute for Experimental Molecular Imaging, RWTH Aachen University Hospital, Forckenbeckstrasse 55, 52074 Aachen, Germany. Electronic address: jpena@ukaachen.de