Engineering an inducible leukemia- associated fusion protein enables large- scale ex vivo production of functional human phagocytes

• 成果类型: Article
• 署名作者: Windisch, Roland; Soliman, Sarah; Hoffmann, Adrian; Chen-Wichmann, Linping; Danese, Anna; Vosberg, Sebastian; Bravo, Jimena; Lutz, Sebastian; Kellner, Christian; Fischer, Alexander; Gebhard, Claudia; Monte, Enric Redondo; Hartmann, Luise; Schneider, Stephanie; Beier, Fabian; Strobl, Carolin Dorothea; Weigert, Oliver; Peipp, Matthias; Schuendeln, Michael; Stricker, Stefan H.; Rehli, Michael; Bernhagen, Juergen; Humpe, Andreas; Klump, Hannes; Brendel, Christian; Krause, Daniela S.; Greif, Philipp A.; Wichmann, Christian
• 署名单位: University of Munich; University of Munich; University of Munich; University of Munich; University of Munich; Helmholtz Association; German Cancer Research Center (DKFZ); Helmholtz Association; German Cancer Research Center (DKFZ); Medical University of Graz; Octapharma; University of Regensburg; University of Munich; University of Munich; RWTH Aachen University; RWTH Aachen University Hospital; University of Kiel; University of Duisburg Essen; University of Duisburg Essen; University of Munich; University of Duisburg Essen; RWTH Aachen University; RWTH Aachen University Hospital; Harvard University; Harvard Medical School; Harvard University Medical Affiliates; Dana-Farber Cancer Institute; Boston Children's Hospital; Goethe University Frankfurt; Goethe University Frankfurt Hospital
• 刊物名称: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
• ISSN/ISSBN: 0027-11365
• DOI: 10.1073/pnas.2312499121
• 发表日期: 2024-06-18
• 关键词: hematopoietic-stem-cell; telomere length; mll; expansion; macrophages; regulators; differentiation; translocations; dimerization; activation

摘要

Ex vivo expansion of human CD34+ hematopoietic stem and progenitor cells remains a challenge due to rapid differentiation after detachment from the bone marrow niche. In this study, we assessed the capacity of an inducible fusion protein to enable sustained ex vivo proliferation of hematopoietic precursors and their capacity to differentiate into functional phagocytes. We fused the coding sequences of an FK506- Binding Protein 12 (FKBP12)- derived destabilization domain (DD) to the myeloid/lymphoid lineage leukemia/eleven nineteen leukemia (MLL- ENL) fusion gene to generate the fusion protein DD- MLL- ENL and retrovirally expressed the protein switch in human CD34+ progenitors. Using Shield1, a chemical inhibitor of DD fusion protein degradation, we established large- scale and long- term expansion of late monocytic precursors. Upon Shield1 removal, the cells lost self- renewal capacity and spontaneously differentiated, even after 2.5 y of continuous ex vivo expansion. In the absence of Shield1, stimulation with IFN-gamma , LPS, and GM- CSF triggered terminal differentiation. Gene expression analysis of the obtained phagocytes revealed marked similarity with na & iuml;ve monocytes. In functional assays, the novel phagocytes migrated toward CCL2, attached to VCAM- 1 under shear stress, produced reactive oxygen species, and engulfed bacterial particles, cellular particles, and apoptotic cells. Finally, we demonstrated Fc gamma receptor recognition and phagocytosis of opsonized lymphoma cells in an antibody- dependent manner. Overall, we have established an engineered protein that, as a single factor, is useful for large- scale ex vivo production of human phagocytes. Such adjustable proteins have the potential to be applied as molecular tools to produce functional immune cells for experimental cell- based approaches.