Publications
2023
Liang, Kai Ling; Roels, Juliette; Lavaert, Marieke; Putteman, Tom; Boehme, Lena; Tilleman, Laurentijn; Velghe, Imke; Pegoretti, Valentina; de Walle, Inge Van; Sontag, Stephanie; Vandewalle, Jolien; Vandekerckhove, Bart; Leclercq, Georges; Vlierberghe, Pieter Van; Libert, Claude; Nieuwerburgh, Filip Van; Fischer, Roman; Kontermann, Roland E.; Pfizenmaier, Klaus; Doody, Gina; Zenke, Martin; Taghon, Tom
Intrathymic dendritic cell-biased precursors promote human T cell lineage specification through IRF8-driven transmembrane TNF. Journal Article
In: Nature immunology, vol. 24, iss. 3, pp. 474–486, 2023, ISSN: 1529-2916.
@article{Liang2023,
title = {Intrathymic dendritic cell-biased precursors promote human T cell lineage specification through IRF8-driven transmembrane TNF.},
author = {Kai Ling Liang and Juliette Roels and Marieke Lavaert and Tom Putteman and Lena Boehme and Laurentijn Tilleman and Imke Velghe and Valentina Pegoretti and Inge Van de Walle and Stephanie Sontag and Jolien Vandewalle and Bart Vandekerckhove and Georges Leclercq and Pieter Van Vlierberghe and Claude Libert and Filip Van Nieuwerburgh and Roman Fischer and Roland E. Kontermann and Klaus Pfizenmaier and Gina Doody and Martin Zenke and Tom Taghon},
doi = {10.1038/s41590-022-01417-6},
issn = {1529-2916},
year = {2023},
date = {2023-03-01},
journal = {Nature immunology},
volume = {24},
issue = {3},
pages = {474--486},
abstract = {The cross-talk between thymocytes and thymic stromal cells is fundamental for T cell development. In humans, intrathymic development of dendritic cells (DCs) is evident but its physiological significance is unknown. Here we showed that DC-biased precursors depended on the expression of the transcription factor IRF8 to express the membrane-bound precursor form of the cytokine TNF (tmTNF) to promote differentiation of thymus seeding hematopoietic progenitors into T-lineage specified precursors through activation of the TNF receptor (TNFR)-2 instead of TNFR1. In vitro recapitulation of TNFR2 signaling by providing low-density tmTNF or a selective TNFR2 agonist enhanced the generation of human T cell precursors. Our study shows that, in addition to mediating thymocyte selection and maturation, DCs function as hematopoietic stromal support for the early stages of human T cell development and provide proof of concept that selective targeting of TNFR2 can enhance the in vitro generation of T cell precursors for clinical application.},
keywords = {},
pubstate = {ppublish},
tppubtype = {article}
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The cross-talk between thymocytes and thymic stromal cells is fundamental for T cell development. In humans, intrathymic development of dendritic cells (DCs) is evident but its physiological significance is unknown. Here we showed that DC-biased precursors depended on the expression of the transcription factor IRF8 to express the membrane-bound precursor form of the cytokine TNF (tmTNF) to promote differentiation of thymus seeding hematopoietic progenitors into T-lineage specified precursors through activation of the TNF receptor (TNFR)-2 instead of TNFR1. In vitro recapitulation of TNFR2 signaling by providing low-density tmTNF or a selective TNFR2 agonist enhanced the generation of human T cell precursors. Our study shows that, in addition to mediating thymocyte selection and maturation, DCs function as hematopoietic stromal support for the early stages of human T cell development and provide proof of concept that selective targeting of TNFR2 can enhance the in vitro generation of T cell precursors for clinical application.
2021
Xiong, Ranhua; Hua, Dawei; Hoeck, Jelter Van; Berdecka, Dominika; Léger, Laurens; Munter, Stijn De; Fraire, Juan C.; Raes, Laurens; Harizaj, Aranit; Sauvage, Félix; Goetgeluk, Glenn; Pille, Melissa; Aalders, Jeffrey; Belza, Joke; Acker, Thibaut Van; Bolea-Fernandez, Eduardo; Si, Ting; Vanhaecke, Frank; Vos, Winnok H. De; Vandekerckhove, Bart; Hengel, Jolanda; Raemdonck, Koen; Huang, Chaobo; Smedt, Stefaan C. De; Braeckmans, Kevin
Photothermal nanofibres enable safe engineering of therapeutic cells. Journal Article
In: Nature nanotechnology, vol. 16, iss. 11, pp. 1281–1291, 2021, ISSN: 1748-3395.
@article{Xiong2021,
title = {Photothermal nanofibres enable safe engineering of therapeutic cells.},
author = {Ranhua Xiong and Dawei Hua and Jelter Van Hoeck and Dominika Berdecka and Laurens Léger and Stijn De Munter and Juan C. Fraire and Laurens Raes and Aranit Harizaj and Félix Sauvage and Glenn Goetgeluk and Melissa Pille and Jeffrey Aalders and Joke Belza and Thibaut Van Acker and Eduardo Bolea-Fernandez and Ting Si and Frank Vanhaecke and Winnok H. De Vos and Bart Vandekerckhove and Jolanda Hengel and Koen Raemdonck and Chaobo Huang and Stefaan C. De Smedt and Kevin Braeckmans},
doi = {10.1038/s41565-021-00976-3},
issn = {1748-3395},
year = {2021},
date = {2021-11-01},
journal = {Nature nanotechnology},
volume = {16},
issue = {11},
pages = {1281--1291},
abstract = {Nanoparticle-sensitized photoporation is an upcoming approach for the intracellular delivery of biologics, combining high efficiency and throughput with excellent cell viability. However, as it relies on close contact between nanoparticles and cells, its translation towards clinical applications is hampered by safety and regulatory concerns. Here we show that light-sensitive iron oxide nanoparticles embedded in biocompatible electrospun nanofibres induce membrane permeabilization by photothermal effects without direct cellular contact with the nanoparticles. The photothermal nanofibres have been successfully used to deliver effector molecules, including CRISPR-Cas9 ribonucleoprotein complexes and short interfering RNA, to adherent and suspension cells, including embryonic stem cells and hard-to-transfect T cells, without affecting cell proliferation or phenotype. In vivo experiments furthermore demonstrated successful tumour regression in mice treated with chimeric antibody receptor T cells in which the expression of programmed cell death protein 1 (PD1) is downregulated after nanofibre photoporation with short interfering RNA to PD1. In conclusion, cell membrane permeabilization with photothermal nanofibres is a promising concept towards the safe and more efficient production of engineered cells for therapeutic applications, including stem cell or adoptive T cell therapy.},
keywords = {},
pubstate = {ppublish},
tppubtype = {article}
}
Nanoparticle-sensitized photoporation is an upcoming approach for the intracellular delivery of biologics, combining high efficiency and throughput with excellent cell viability. However, as it relies on close contact between nanoparticles and cells, its translation towards clinical applications is hampered by safety and regulatory concerns. Here we show that light-sensitive iron oxide nanoparticles embedded in biocompatible electrospun nanofibres induce membrane permeabilization by photothermal effects without direct cellular contact with the nanoparticles. The photothermal nanofibres have been successfully used to deliver effector molecules, including CRISPR-Cas9 ribonucleoprotein complexes and short interfering RNA, to adherent and suspension cells, including embryonic stem cells and hard-to-transfect T cells, without affecting cell proliferation or phenotype. In vivo experiments furthermore demonstrated successful tumour regression in mice treated with chimeric antibody receptor T cells in which the expression of programmed cell death protein 1 (PD1) is downregulated after nanofibre photoporation with short interfering RNA to PD1. In conclusion, cell membrane permeabilization with photothermal nanofibres is a promising concept towards the safe and more efficient production of engineered cells for therapeutic applications, including stem cell or adoptive T cell therapy.
Raes, Laurens; Smedt, Stefaan C. De; Raemdonck, Koen; Braeckmans, Kevin
Non-viral transfection technologies for next-generation therapeutic T cell engineering. Journal Article
In: Biotechnology advances, vol. 49, pp. 107760, 2021, ISSN: 1873-1899.
@article{Raes2021,
title = {Non-viral transfection technologies for next-generation therapeutic T cell engineering.},
author = {Laurens Raes and Stefaan C. De Smedt and Koen Raemdonck and Kevin Braeckmans},
doi = {10.1016/j.biotechadv.2021.107760},
issn = {1873-1899},
year = {2021},
date = {2021-01-01},
journal = {Biotechnology advances},
volume = {49},
pages = {107760},
abstract = {Genetically engineered T cells have sparked interest in advanced cancer treatment, reaching a milestone in 2017 with two FDA-approvals for CD19-directed chimeric antigen receptor (CAR) T cell therapeutics. It is becoming clear that the next generation of CAR T cell therapies will demand more complex engineering strategies and combinations thereof, including the use of revolutionary gene editing approaches. To date, manufacturing of CAR T cells mostly relies on γ-retroviral or lentiviral vectors, but their use is associated with several drawbacks, including safety issues, high manufacturing cost and vector capacity constraints. Non-viral approaches, including membrane permeabilization and carrier-based techniques, have therefore gained a lot of interest to replace viral transductions in the manufacturing of T cell therapeutics. This review provides an in-depth discussion on the avid search for alternatives to viral vectors, discusses key considerations for T cell engineering technologies, and provides an overview of the emerging spectrum of non-viral transfection technologies for T cells. Strengths and weaknesses of each technology will be discussed in relation to T cell engineering. Altogether, this work emphasizes the potential of non-viral transfection approaches to advance the next-generation of genetically engineered T cells.},
keywords = {},
pubstate = {ppublish},
tppubtype = {article}
}
Genetically engineered T cells have sparked interest in advanced cancer treatment, reaching a milestone in 2017 with two FDA-approvals for CD19-directed chimeric antigen receptor (CAR) T cell therapeutics. It is becoming clear that the next generation of CAR T cell therapies will demand more complex engineering strategies and combinations thereof, including the use of revolutionary gene editing approaches. To date, manufacturing of CAR T cells mostly relies on γ-retroviral or lentiviral vectors, but their use is associated with several drawbacks, including safety issues, high manufacturing cost and vector capacity constraints. Non-viral approaches, including membrane permeabilization and carrier-based techniques, have therefore gained a lot of interest to replace viral transductions in the manufacturing of T cell therapeutics. This review provides an in-depth discussion on the avid search for alternatives to viral vectors, discusses key considerations for T cell engineering technologies, and provides an overview of the emerging spectrum of non-viral transfection technologies for T cells. Strengths and weaknesses of each technology will be discussed in relation to T cell engineering. Altogether, this work emphasizes the potential of non-viral transfection approaches to advance the next-generation of genetically engineered T cells.
Harizaj, Aranit; Wels, Mike; Raes, Laurens; Stremersch, Stephan; Goetgeluk, Glenn; Brans, Toon; Vandekerckhove, Bart; Sauvage, Félix; Smedt, Stefaan De; Lentacker, Ine; Braeckmans, Kevin
Photoporation with biodegradable polydopamine nanosensitizers enables safe and efficient delivery of mRNA in human T cells Journal Article
In: ADVANCED FUNCTIONAL MATERIALS, vol. 31, no. 28, pp. 12, 2021, ISSN: 1616-301X.
@article{8717064,
title = {Photoporation with biodegradable polydopamine nanosensitizers enables safe and efficient delivery of mRNA in human T cells},
author = {Aranit Harizaj and Mike Wels and Laurens Raes and Stephan Stremersch and Glenn Goetgeluk and Toon Brans and Bart Vandekerckhove and Félix Sauvage and Stefaan De Smedt and Ine Lentacker and Kevin Braeckmans},
url = {http://dx.doi.org/10.1002/adfm.202102472},
issn = {1616-301X},
year = {2021},
date = {2021-01-01},
journal = {ADVANCED FUNCTIONAL MATERIALS},
volume = {31},
number = {28},
pages = {12},
abstract = {Safe and efficient production of chimeric antigen receptor (CAR)-T cells is of crucial importance for cell-based cancer immunotherapy. Physical transfection methods have quickly gained in importance to transfect T-cells, being readily compatible with different cell types and a broad variety of cargo molecules. In particular, nanoparticle-sensitized photoporation has been introduced in recent years as a gentle yet efficient method to transiently permeabilize cells, allowing subsequent entry of external cargo molecules into the cells. Gold nanoparticles (AuNPs) have been used the most as photothermal sensitizers because they can easily form laser-induced vapor nanobubbles, a photothermal phenomenon that is shown to be particularly efficient for permeabilizing cells. However, as AuNPs are not biodegradable, clinical translation is hampered. Here, for the first time, it is reported on the possibility to form laser-induced vapor nanobubbles from biocompatible polymeric nanoparticles. Compared to electroporation as the most used physical transfection method for T cells, 2.5 times more living mRNA transfected human T cells are obtained via photoporation sensitized by polydopamine nanoparticles. Together, it shows that photoporation is a viable approach for efficiently producing therapeutic engineered T-cells at a throughput easily exceeding 10(5) cells per second.},
keywords = {},
pubstate = {published},
tppubtype = {article}
}
Safe and efficient production of chimeric antigen receptor (CAR)-T cells is of crucial importance for cell-based cancer immunotherapy. Physical transfection methods have quickly gained in importance to transfect T-cells, being readily compatible with different cell types and a broad variety of cargo molecules. In particular, nanoparticle-sensitized photoporation has been introduced in recent years as a gentle yet efficient method to transiently permeabilize cells, allowing subsequent entry of external cargo molecules into the cells. Gold nanoparticles (AuNPs) have been used the most as photothermal sensitizers because they can easily form laser-induced vapor nanobubbles, a photothermal phenomenon that is shown to be particularly efficient for permeabilizing cells. However, as AuNPs are not biodegradable, clinical translation is hampered. Here, for the first time, it is reported on the possibility to form laser-induced vapor nanobubbles from biocompatible polymeric nanoparticles. Compared to electroporation as the most used physical transfection method for T cells, 2.5 times more living mRNA transfected human T cells are obtained via photoporation sensitized by polydopamine nanoparticles. Together, it shows that photoporation is a viable approach for efficiently producing therapeutic engineered T-cells at a throughput easily exceeding 10(5) cells per second.
2020
Lavaert, Marieke; Liang, Kai Ling; Vandamme, Niels; Park, Jong-Eun; Roels, Juliette; Kowalczyk, Monica S.; Li, Bo; Ashenberg, Orr; Tabaka, Marcin; Dionne, Danielle; Tickle, Timothy L.; Slyper, Michal; Rozenblatt-Rosen, Orit; Vandekerckhove, Bart; Leclercq, Georges; Regev, Aviv; Vlierberghe, Pieter Van; Guilliams, Martin; Teichmann, Sarah A.; Saeys, Yvan; Taghon, Tom
Integrated scRNA-Seq Identifies Human Postnatal Thymus Seeding Progenitors and Regulatory Dynamics of Differentiating Immature Thymocytes. Journal Article
In: Immunity, vol. 52, iss. 6, pp. 1088–1104.e6, 2020, ISSN: 1097-4180.
@article{Lavaert2020,
title = {Integrated scRNA-Seq Identifies Human Postnatal Thymus Seeding Progenitors and Regulatory Dynamics of Differentiating Immature Thymocytes.},
author = {Marieke Lavaert and Kai Ling Liang and Niels Vandamme and Jong-Eun Park and Juliette Roels and Monica S. Kowalczyk and Bo Li and Orr Ashenberg and Marcin Tabaka and Danielle Dionne and Timothy L. Tickle and Michal Slyper and Orit Rozenblatt-Rosen and Bart Vandekerckhove and Georges Leclercq and Aviv Regev and Pieter Van Vlierberghe and Martin Guilliams and Sarah A. Teichmann and Yvan Saeys and Tom Taghon},
doi = {10.1016/j.immuni.2020.03.019},
issn = {1097-4180},
year = {2020},
date = {2020-06-01},
journal = {Immunity},
volume = {52},
issue = {6},
pages = {1088--1104.e6},
abstract = {During postnatal life, thymopoiesis depends on the continuous colonization of the thymus by bone-marrow-derived hematopoietic progenitors that migrate through the bloodstream. The current understanding of the nature of thymic immigrants is largely based on data from pre-clinical models. Here, we employed single-cell RNA sequencing (scRNA-seq) to examine the immature postnatal thymocyte population in humans. Integration of bone marrow and peripheral blood precursor datasets identified two putative thymus seeding progenitors that varied in expression of CD7; CD10; and the homing receptors CCR7, CCR9, and ITGB7. Whereas both precursors supported T cell development, only one contributed to intrathymic dendritic cell (DC) differentiation, predominantly of plasmacytoid dendritic cells. Trajectory inference delineated the transcriptional dynamics underlying early human T lineage development, enabling prediction of transcription factor (TF) modules that drive stage-specific steps of human T cell development. This comprehensive dataset defines the expression signature of immature human thymocytes and provides a resource for the further study of human thymopoiesis.},
keywords = {},
pubstate = {ppublish},
tppubtype = {article}
}
During postnatal life, thymopoiesis depends on the continuous colonization of the thymus by bone-marrow-derived hematopoietic progenitors that migrate through the bloodstream. The current understanding of the nature of thymic immigrants is largely based on data from pre-clinical models. Here, we employed single-cell RNA sequencing (scRNA-seq) to examine the immature postnatal thymocyte population in humans. Integration of bone marrow and peripheral blood precursor datasets identified two putative thymus seeding progenitors that varied in expression of CD7; CD10; and the homing receptors CCR7, CCR9, and ITGB7. Whereas both precursors supported T cell development, only one contributed to intrathymic dendritic cell (DC) differentiation, predominantly of plasmacytoid dendritic cells. Trajectory inference delineated the transcriptional dynamics underlying early human T lineage development, enabling prediction of transcription factor (TF) modules that drive stage-specific steps of human T cell development. This comprehensive dataset defines the expression signature of immature human thymocytes and provides a resource for the further study of human thymopoiesis.