2. Academic Validation
  3. Engineered nucleocytosolic vehicles for loading of programmable editors

Engineered nucleocytosolic vehicles for loading of programmable editors

  • Cell. 2025 May 15;188(10):2637-2655.e31. doi: 10.1016/j.cell.2025.03.015.
Julian Geilenkeuser 1,  Niklas Armbrust 1,  Emily Steinmaßl 1,  Samuel W Du 2,  Sebastian Schmidt 3,  Eva Maria Hildegard Binder 1,  Yuchun Li 1,  Niklas Wilhelm Warsing 1,  Stephanie Victoria Wendel 1,  Florian von der Linde 1,  Elisa Marie Schiele 1,  Xiya Niu 4,  Luisa Stroppel 1,  Oleksandr Berezin 1,  Tobias Heinrich Santl 1,  Tanja Orschmann 5,  Keith Nelson 1,  Christoph Gruber 6,  Grazyna Palczewska 7,  Carolline Rodrigues Menezes 2,  Eleonora Risaliti 2,  Zachary J Engfer 2,  Naile Koleci 8,  Andrea Schmidts 8,  Arie Geerlof 9,  Krzysztof Palczewski 10,  Gil Gregor Westmeyer 11,  Dong-Jiunn Jeffery Truong 12
Affiliations

Affiliations

  • 1 Institute for Synthetic Biomedicine, Helmholtz Munich, Neuherberg, Germany; Department of Bioscience, TUM School of Natural Sciences, Technical University of Munich, Munich, Germany.
  • 2 Gavin Herbert Eye Institute - Center for Translational Vision Research, Department of Ophthalmology, University of California, Irvine, Irvine, CA, USA; Department of Physiology & Biophysics, University of California, Irvine, Irvine, CA, USA.
  • 3 Institute for Synthetic Biomedicine, Helmholtz Munich, Neuherberg, Germany; Department of Bioscience, TUM School of Natural Sciences, Technical University of Munich, Munich, Germany; Institute of Developmental Genetics, Helmholtz Munich, Neuherberg, Germany.
  • 4 Institute for Synthetic Biomedicine, Helmholtz Munich, Neuherberg, Germany.
  • 5 Institute for Synthetic Biomedicine, Helmholtz Munich, Neuherberg, Germany; Institute of Developmental Genetics, Helmholtz Munich, Neuherberg, Germany.
  • 6 Institute of Developmental Genetics, Helmholtz Munich, Neuherberg, Germany.
  • 7 Gavin Herbert Eye Institute - Center for Translational Vision Research, Department of Ophthalmology, University of California, Irvine, Irvine, CA, USA.
  • 8 Department of Medicine III: Hematology/Oncology, Klinikum rechts der Isar of the Technical University of Munich, TUM School of Medicine and Health, Munich, Germany.
  • 9 Institute of Structural Biology, Helmholtz Munich, Neuherberg, Germany.
  • 10 Gavin Herbert Eye Institute - Center for Translational Vision Research, Department of Ophthalmology, University of California, Irvine, Irvine, CA, USA; Department of Physiology & Biophysics, University of California, Irvine, Irvine, CA, USA; Department of Chemistry, University of California, Irvine, Irvine, CA, USA; Department of Molecular Biology and Biochemistry, University of California, Irvine, Irvine, CA, USA.
  • 11 Institute for Synthetic Biomedicine, Helmholtz Munich, Neuherberg, Germany; Department of Bioscience, TUM School of Natural Sciences, Technical University of Munich, Munich, Germany. Electronic address: gil.westmeyer@tum.de.
  • 12 Institute for Synthetic Biomedicine, Helmholtz Munich, Neuherberg, Germany; Department of Bioscience, TUM School of Natural Sciences, Technical University of Munich, Munich, Germany. Electronic address: jeffery.truong@helmholtz-munich.de.
PMID: 40209705 DOI: 10.1016/j.cell.2025.03.015
Abstract

Advanced gene editing methods have accelerated biomedical discovery and hold great therapeutic promise, but safe and efficient delivery of gene editors remains challenging. In this study, we present a virus-like particle (VLP) system featuring nucleocytosolic shuttling vehicles that retrieve pre-assembled Cas-effectors via aptamer-tagged guide RNAs. This approach ensures preferential loading of fully assembled editor ribonucleoproteins (RNPs) and enhances the efficacy of prime editing, base editing, trans-activators, and nuclease activity coupled to homology-directed repair in multiple immortalized, primary, stem cell, and stem-cell-derived cell types. We also achieve additional protection of inherently unstable prime editing guide RNAs (pegRNAs) by shielding the 3'-exposed end with Csy4/Cas6f, further enhancing editing performance. Furthermore, we identify a minimal set of packaging and budding modules that can serve as a platform for bottom-up engineering of enveloped delivery vehicles. Notably, our system demonstrates superior per-VLP editing efficiency in primary T lymphocytes and two mouse models of inherited retinal disease, highlighting its therapeutic potential.

Keywords

VLPs; base editing; cell therapy; gene delivery; gene therapy; genome editing; in vivo delivery; prime editing; virus-like particles.

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