Nanoengineered 3D culture substrate enables superior persistence and polyclonal engraftment of genetically engineered hematopoietic stem cells
Ex vivo culture of hematopoietic stem and progenitor cells (HSPCs) is required for gene therapy applications but inadvertently triggers detrimental cellular responses, potentially threatening clinical success. In this study, we employ nichoids, biocompatible 3D culture substrates with cell-scale resolution, to provide HSPCs with mechanical support during ex vivo manipulation. This innovative 3D system improves HSPC multi-lineage differentiation and engraftment capacity by leveraging mechanobiological control over nuclear morphology, cytoskeleton organization, metabolism, and DNA integrity. Notably, 3D culture enables efficient genetic engineering across multiple platforms, including long-range gene editing, base- and prime-editing, and lentiviral-mediated gene addition. Moreover, this scaffold increases the clonal output and persistence of genetically engineered cells in xenotransplantation experiments, including a clinical protocol for lentiviral gene addition in Wiskott-Aldrich syndrome. Overall, we propose a transformative approach to enhance the efficacy and safety of emerging and established hematopoietic stem cell-based gene therapy applications.
Authors
- Bernhard Gentner (ORCID: https://orcid.org/0000-0001-6024-4718)
- Edoardo Carsana
- Martina Fiumara (ORCID: https://orcid.org/0009-0007-5934-9273)
- Fabrizio Benedicenti (ORCID: https://orcid.org/0000-0002-6378-125X)
- Sergio Arévalo (ORCID: https://orcid.org/0000-0002-2637-7100)
- Teresa Tavella
- Dafne Barozzi (ORCID: https://orcid.org/0009-0002-7377-3461)
- Samuele Ferrari (ORCID: https://orcid.org/0000-0003-1122-1231)
- Kety Giannetti (ORCID: https://orcid.org/0000-0001-7989-2520)
- Federica Cugnata (ORCID: https://orcid.org/0000-0002-1886-569X)
- Andrea Brendolan (ORCID: https://orcid.org/0000-0003-0109-6879)
- Giulio Cerullo (ORCID: https://orcid.org/0000-0002-9534-2702)
- Cristina Scielzo (ORCID: https://orcid.org/0000-0001-7988-2824)
- Claudio Conci (ORCID: https://orcid.org/0000-0002-2335-9046)
- Federico Midena
- Manuela Teresa Raimondi (ORCID: https://orcid.org/0000-0003-2585-7206)
- Erika Zonari (ORCID: https://orcid.org/0000-0002-7915-3995)
- Lucrezia della Volpe (ORCID: https://orcid.org/0000-0002-9087-1293)
- Laura Alessandrini
- Francesco Gazzo (ORCID: https://orcid.org/0009-0000-4626-1386)
- Luigi Naldini
- Matteo Barcella (ORCID: https://orcid.org/0000-0001-7350-3192)
- Giacomo Farina
- Leonardo Cherubin
- Di Micco Raffaella
- Martina Franchino
- Emanuela Jacchetti
- Marina Radrizzani
- Ivan Merelli
- Federico Fraschetta
- Alessandro Aiuti
- Chiara Brombin
- Eugenio Montini
- Roberta Vacca
- Roberto Osellame
- Francesca Ferrua
Institutions
- Vita-Salute San Raffaele University (IT)
- The San Raffaele Telethon Institute for Gene Therapy (IT)
- Center for Biomolecular Nanotechnologies (IT)
- Istituti di Ricovero e Cura a Carattere Scientifico (IT)
- National Research Council (IT)
- Istituto di Ricovero e Cura a Carattere Scientifico San Raffaele (IT)
- Politecnico di Milano (IT)
Publication Details
- Journal
- Cell stem cell
- Published
- 2026-01-08
- DOI
- https://doi.org/10.1016/j.stem.2025.12.016
- Citations
- 1
- Primary Topic
- Virus-based gene therapy research
- Type
- article
- Field-Weighted Citation Impact
- 30.93