Piriya Luangwattananun1,2, Mutita Junking3,4, Jatuporn Sujjitjoon1,2, Yupanun Wutti-In1,5, Naravat Poungvarin6, Chanitra Thuwajit1,7, Pa-Thai Yenchitsomanus8,9. 1. Siriraj Center of Research Excellence for Cancer Immunotherapy (SiCORE-CIT), Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. 2. Division of Molecular Medicine, Research Department, Faculty of Medicine Siriraj Hospital, Mahidol, University, Bangkok, 10700, Thailand. 3. Siriraj Center of Research Excellence for Cancer Immunotherapy (SiCORE-CIT), Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. mutita.jun@mahidol.ac.th. 4. Division of Molecular Medicine, Research Department, Faculty of Medicine Siriraj Hospital, Mahidol, University, Bangkok, 10700, Thailand. mutita.jun@mahidol.ac.th. 5. Graduate Program in Immunology, Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. 6. Department of Clinical Pathology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. 7. Department of Immunology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. 8. Siriraj Center of Research Excellence for Cancer Immunotherapy (SiCORE-CIT), Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, 10700, Thailand. pathai.yen@mahidol.edu. 9. Division of Molecular Medicine, Research Department, Faculty of Medicine Siriraj Hospital, Mahidol, University, Bangkok, 10700, Thailand. pathai.yen@mahidol.edu.
Abstract
PURPOSE: Treatment of breast cancer (BC) by standard methods is effective in the early stage, but ineffective in the advanced stage of disease. To develop an adoptive T cell therapy for advanced and severe BC, we generated fourth-generation chimeric antigen receptor (CAR) T cells targeting folate receptor alpha antigen (FRα) expressed on BC cells, and preclinically evaluated their anti-BC activities. METHODS: The fourth-generation FRα-CAR T cells containing extracellular FRα-specific single-chain variable fragment (scFv) and three intracellular costimulatory domains (CD28, 4-1BB, and CD27) linked to CD3ζ were generated using a lentiviral system, and then were evaluated for their anti-BC activities in two-dimensional and three-dimensional (spheroid) cultures. RESULTS: When our fourth-generation FRα-CAR T cells were cocultured with FRα-expressing MDA-MB-231 BC cell line at an effector to target ratio of 20:1, these CAR T cells specifically lysed 88.7 ± 10.6% of the target cells. Interestingly, the cytotoxic lysis of FRα-CAR T cells was more pronounced in target cells with higher surface FRα expression. This specific cytotoxicity of the CAR T cells was not observed when cocultured with FRα-negative MCF10A normal breast-like cell line at the same ratio (34.3 ± 4.7%). When they were cocultured with MDA-MD-231 spheroid, the FRα-CAR T cells exhibited antitumor activity marked with spheroid size reduction and breakage. CONCLUSION: This proof-of-concept study thus shows the feasibility of using these fourth-generation FRα-CAR T cells for adoptive T cell therapy in BC.
PURPOSE: Treatment of breast cancer (BC) by standard methods is effective in the early stage, but ineffective in the advanced stage of disease. To develop an adoptive T cell therapy for advanced and severe BC, we generated fourth-generation chimeric antigen receptor (CAR) T cells targeting folate receptor alpha antigen (FRα) expressed on BC cells, and preclinically evaluated their anti-BC activities. METHODS: The fourth-generation FRα-CAR T cells containing extracellular FRα-specific single-chain variable fragment (scFv) and three intracellular costimulatory domains (CD28, 4-1BB, and CD27) linked to CD3ζ were generated using a lentiviral system, and then were evaluated for their anti-BC activities in two-dimensional and three-dimensional (spheroid) cultures. RESULTS: When our fourth-generation FRα-CAR T cells were cocultured with FRα-expressing MDA-MB-231 BC cell line at an effector to target ratio of 20:1, these CAR T cells specifically lysed 88.7 ± 10.6% of the target cells. Interestingly, the cytotoxic lysis of FRα-CAR T cells was more pronounced in target cells with higher surface FRα expression. This specific cytotoxicity of the CAR T cells was not observed when cocultured with FRα-negative MCF10A normal breast-like cell line at the same ratio (34.3 ± 4.7%). When they were cocultured with MDA-MD-231 spheroid, the FRα-CAR T cells exhibited antitumor activity marked with spheroid size reduction and breakage. CONCLUSION: This proof-of-concept study thus shows the feasibility of using these fourth-generation FRα-CAR T cells for adoptive T cell therapy in BC.
Entities:
Keywords:
Adoptive cell therapy; Breast cancer; Chimeric antigen receptor (CAR) T cell; Folate receptor alpha (FRα); Immunotherapy
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