Article ; Online: Identification and characterisation of novel CAR-T cells to target IL13Rα2 positive human glioma in vitro and in vivo.
Clinical and translational medicine
2024 Volume 14, Issue 5, Page(s) e1664
Abstract: Background: Previously, we discovered that human solid tumours, but not normal human tissues, preferentially overexpress interleukin-13Receptor alpha2, a high binding receptor for IL-13. To develop novel anti-cancer approaches, we constructed a chimeric ...
| Abstract | Background: Previously, we discovered that human solid tumours, but not normal human tissues, preferentially overexpress interleukin-13Receptor alpha2, a high binding receptor for IL-13. To develop novel anti-cancer approaches, we constructed a chimeric antigen receptor construct using a high binding and codon optimised scFv-IL-13Rα2 fragment fused with CD3ζ and co-stimulatory cytoplasmic domains of CD28 and 4-1BB. Methods: We developed a scFv clone, designated 14-1, by biopanning the bound scFv phages using huIL-13Rα2Fc chimeric protein and compared its binding with our previously published clone 4-1. We performed bioinformatic analyses for complementary determining regions (CDR) framework and residue analyses of the light and heavy chains. This construct was packaged with helper plasmids to produce CAR-lentivirus and transduced human Jurkat T or activated T cells from peripheral blood mononuclear cells (PBMCs) to produce CAR-T cells and tested for their quality attributes in vitro and in vivo. Serum enzymes including body weight from non-tumour bearing mice were tested for assessing general toxicity of CAR-T cells. Results: The binding of 14-1 clone is to IL-13Rα2Fc-chimeric protein is ∼5 times higher than our previous clone 4-1. The 14-1-CAR-T cells grew exponentially in the presence of cytokines and maintained phenotype and biological attributes such as cell viability, potency, migration and T cell activation. Clone 14-1 migrated to IL-13Rα2Fc and cell free supernatants only from IL-13Rα2+ve confluent glioma tumour cells in a chemotaxis assay. scFv-IL-13Rα2-CAR-T cells specifically killed IL-13Rα2+ve but not IL-13Rα2-ve tumour cells in vitro and selectively caused significant release of IFN-γ only from IL-13Rα2+ve co-cultures. These CAR-T cells regressed IL-13Rα2+ve glioma xenografts in vivo without any general toxicity. In contrast, the IL-13Rα2 gene knocked-down U251 and U87 xenografts failed to respond to the CAR-T therapy. Conclusion: Taken together, we conclude that the novel scFv-IL-13Rα2 CAR-T cell therapy may offer an effective therapeutic option after designing a careful pre-clinical and clinical study. |
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| MeSH term(s) | Humans ; Interleukin-13 Receptor alpha2 Subunit/metabolism ; Interleukin-13 Receptor alpha2 Subunit/genetics ; Mice ; Glioma/immunology ; Glioma/therapy ; Glioma/genetics ; Glioma/pathology ; Glioma/metabolism ; Animals ; Immunotherapy, Adoptive/methods ; Disease Models, Animal ; Receptors, Chimeric Antigen/metabolism ; Receptors, Chimeric Antigen/immunology |
| Chemical Substances | Interleukin-13 Receptor alpha2 Subunit ; IL13RA2 protein, human ; Receptors, Chimeric Antigen |
| Language | English |
| Publishing date | 2024-04-27 |
| Publishing country | United States |
| Document type | Journal Article ; Research Support, Non-U.S. Gov't |
| ZDB-ID | 2697013-2 |
| ISSN | 2001-1326 ; 2001-1326 |
| ISSN (online) | 2001-1326 |
| ISSN | 2001-1326 |
| DOI | 10.1002/ctm2.1664 |
| Database | MEDical Literature Analysis and Retrieval System OnLINE |
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