Unveiling the influence of CAR-negative T-cells: enhancing efficacy and ensuring safety in CAR-T therapies
Background Despite the promising results of CAR-T cell therapy, still a significant proportion of patients experience treatment failure due to absence of response, progression or treatment-related toxicities. Our research aims to investigate how the CAR-T cell product characteristics and particularl...
| Autores: | , , , , , , , , |
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| Tipo de recurso: | artículo |
| Estado: | Versión publicada |
| Fecha de publicación: | 2025 |
| País: | España |
| Institución: | Universidad de Sevilla (US) |
| Repositorio: | idUS. Depósito de Investigación de la Universidad de Sevilla |
| OAI Identifier: | oai:dnet:idus________::f3fa9aecb0b5d769d8d09645e8c8e69f |
| Acceso en línea: | https://hdl.handle.net/11441/183806 |
| Access Level: | acceso abierto |
| Palabra clave: | CAR-T FACS-sorting CAR-negative RNA-seq ICANS CRS IL-6 |
| Sumario: | Background Despite the promising results of CAR-T cell therapy, still a significant proportion of patients experience treatment failure due to absence of response, progression or treatment-related toxicities. Our research aims to investigate how the CAR-T cell product characteristics and particularly the percentage of CAR-negative cells present in the infusion product affects its efficacy and safety. Methods CAR-positive (CAR-pos) and CAR-negative (CAR-neg) cells were analyzed during in vitro expansion with IL-2 or IL-7/IL-15. FACS-sorted populations were assessed for cytotoxic effect and secretion of cytokines. Transcriptomic analyses were conducted for CAR-pos and CAR-neg populations. Results A significant reduction in the percentage of transduced (CAR-pos) T cells was observed throughout the in vitro cell culture, where CAR-neg T cells displayed greater expansion along the procedure compared to CAR-pos T cells, using either IL-2 or IL-7/IL-15. CAR-pos T cells exhibited increased expression of activation and exhaustion markers. Comparative analyses of CAR-pos and CAR-neg showed an increased cytotoxic effect in purified samples with 100% CAR-pos T cells compared to non-purified product. Quantification of cytokine levels revealed a significant decrease in IL-6 in purified compared to non-purified products. RNA sequencing analysis revealed significant differential gene expression profile between CAR-pos and CAR-neg cells, the prior exhibiting an upregulation of genes associated with immune checkpoint regulation, cytokine-cytokine receptor interaction and Th1/Th2 and Th17 cell differentiation pathways. Conclusion CAR-neg T cells exhibit a significantly greater expansion compared to CAR-pos T cells alongside CAR-T cell production. The percentage of CAR-neg cells influences the specific cytotoxicity of the product and on the cytokine pattern, which might influence both efficacy and toxicity and, therefore, the number of CAR-neg T cellswithin the infusion product must be considered in the clinical setting. Purification of CAR-pos T cells could potentially optimize product characteristics. |
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