Publication:
Design, synthesis, characterization, and evaluation of 3D synthetic scaffolds for T and CAR-T cells culture and co-cultures

dc.contributor.advisor Torres Lugo, Madeline
dc.contributor.author Lizana Vasquez, Gaby del Rocio
dc.contributor.college College of Engineering
dc.contributor.committee Domenech Garcia, Maribella
dc.contributor.committee Torres Garcia, Wandaliz
dc.contributor.committee Latorre Esteves, Magda
dc.contributor.department Department of Chemical Engineering
dc.contributor.representative Huertas Miranda, Javier A.
dc.date.accessioned 2023-12-05T18:25:34Z
dc.date.available 2023-12-05T18:25:34Z
dc.date.issued 2023-11-16
dc.description.abstract During the manufacturing process of CAR-T cell products, there are many challenges that researchers are currently addressing. One of them is the need to enhance the expansion of T and CAR-T cells, and another is to evaluate the potency of CAR-T cells against tumor spheroids. Therefore, this research aims to design, characterize, and evaluate a thermosensitive terpolymer capable of encapsulating and harvesting cells while reducing mechanical manipulation and allowing microscopic monitoring. For this purpose, three different monomers, including N-isopropylacrylamide, boronic acid, and poly(ethylene glycol), were selected to form a synthetic-based hydrogel scaffold. Different techniques such as the Sol-Gel transition, proton nuclear magnetic resonance (1H-NMR), and Fourier-transformed infrared spectroscopy (FT-IR) were used to characterize the terpolymers resulting from various combinations of monomers. The FT-IR and 1H-NMR spectra confirmed the success of the polymerization and its reproducibility in manufacturing, while the Sol-Gel transition allowed confirming the formation of the hydrogel at 37°C. The feasibility of the most promising terpolymers for 3D in vitro cell culture was evaluated with various cell types, including adherent cancer cell lines SKOV-3, U87-GBM, and patient-derived GSC-GBM. In addition, Jurkat, CD4+ T, Pan-T, and CAR-T suspension cell lines were successfully encapsulated, cultured, and expanded within terpolymer scaffolds. In addition, co-culture experiments of activated T cells with encapsulated U87, and anti-GD2 CAR-T cells with encapsulated GSCs, were performed. Co-culture experiment results provided evidence that T cell migration through the terpolymer matrix to reach the encapsulated spheroids was possible and further analyses of T cell potency could be performed after harvesting cells from terpolymer scaffolds. In conclusion, obtained results indicated that the terpolymer material could be successfully used for cell culture applications and it has the potential to be used in cell potency assays development as well as cell manufacture.
dc.description.abstract Durante el proceso de fabricación de productos de células CAR-T, existen muchos desafíos que los investigadores están abordando actualmente. Uno de ellos es la necesidad de potenciar la expansión de las células T y CAR-T, y otro es evaluar la potencia de las células CAR-T frente a esferoides tumorales. Por lo tanto, esta investigación tiene como objetivo diseñar, caracterizar y evaluar un terpolímero termosensible capaz de encapsular y recolectar células mientras reduce la manipulación mecánica y permite el monitoreo microscópico. Para este propósito, se seleccionaron tres monómeros diferentes, N-isopropilacrilamida, ácido borónico y poli(etilenglicol), para formar un andamio de hidrogel sintético. Se utilizaron diferentes técnicas como la transición Sol-Gel, la resonancia magnética nuclear de protones (1H-NMR) y la espectroscopia infrarroja transformada de Fourier (FT-IR) para caracterizar los terpolímeros resultantes de varias combinaciones de monómeros. Los espectros FT-IR y 1H-NMR confirmaron el éxito de la polimerización y su reproducibilidad en la fabricación, mientras que la transición Sol-Gel permitió confirmar la formación de hidrogel a 37°C. La viabilidad de los terpolímeros más prometedores para el cultivo celular 3D in vitro se evaluaron con varios tipos de células, incluidas las líneas celulares de cáncer adherente SKOV-3, U87-GBM y derivadas de pacientes GSC-GBM. Además, las líneas celulares en suspensión Jurkat, CD4+ T, Pan-T y CAR-T se encapsularon, cultivaron y expandieron con éxito dentro de andamios de terpolímero. También se realizaron experimentos de co-cultivo de células T activadas con U87 encapsulado y células CAR-T anti-GD2 con GSC encapsuladas. Los resultados del co-cultivo proporcionaron evidencia de que era posible la migración de células T a través de la matriz de terpolímero para alcanzar los esferoides encapsulados y que se podía realizar análisis adicionales de la potencia de las células T después de recogerlas de los andamios de terpolímero. En conclusión, los resultados obtenidos indicaron que el material de terpolímero podría usarse con éxito para aplicaciones de cultivo celular y tiene el potencial para usarse en el desarrollo de ensayos de potencia celular, así como en la fabricación de células.
dc.description.graduationSemester Fall
dc.description.graduationYear 2023
dc.description.note This dissertation contains portions published by the author in the Journal of Polymers, and submitted to the Journal of Bioactive Materials and the Journal of Materials Chemistry B.
dc.description.sponsorship Engineering Research Center on Cell Manufacturing Technologies (CMaT); National Science Foundation (NSF) grant number EEC-1648035
dc.identifier.uri https://hdl.handle.net/20.500.11801/3581
dc.language.iso en
dc.rights Attribution-NonCommercial-NoDerivatives 4.0 International *
dc.rights.holder (c) 2023 Gaby R. Lizana Vasquez
dc.rights.uri http://creativecommons.org/licenses/by-nc-nd/4.0/ *
dc.subject Biomaterials
dc.subject Hydrogels
dc.subject Cell manufacturing
dc.subject Polymer synthesis and characterization
dc.subject Immunotherapy
dc.subject.lcsh Biomedical materials
dc.subject.lcsh Cell coculture
dc.subject.lcsh Manufacturing cells
dc.subject.lcsh Cellular therapy
dc.subject.lcsh Synthetic hard materials
dc.subject.lcsh Scaffold proteins
dc.subject.lcsh Nanogels
dc.title Design, synthesis, characterization, and evaluation of 3D synthetic scaffolds for T and CAR-T cells culture and co-cultures
dc.type Dissertation
dspace.entity.type Publication
thesis.degree.discipline Chemical Engineering
thesis.degree.level Ph.D.
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