Journal: EMBO Molecular Medicine

Article Title: A therapeutic regimen using neoantigen-specific TCR-T cells for HLA-A*2402-positive solid tumors

doi: 10.1038/s44321-024-00184-1

Figure Lengend Snippet: ( A , B ) Tcr-T1 cells were generated as mentioned in the methods. Schematic for TCR-T cell generation, purification, and in vitro expansion. ( A ) Representative flow cytometric images indicated the electroporation efficiency and purification of Tcr-T1 cells ( B ). ( C ) HLA-A*2402-positive MKN45-Luci cells were treated with Tcr-T1 cells, with or without Pep-4 pulsing. 24 h later, cell viability was tested via CCK-8 assay. ( D ) Human synovial sarcoma cell line SW982 was modified with stable expression of the HLA-A*2402 molecule to generate target cells (SW982/2402). Then these target cells, SW982 and SW982/2402, were transfected with vectors expressing SYT-SSX fusion mutated peptide containing/deleting Pep-4 sequence (SYT-SSX ΔPep4, SYT-SSX, SYT-SSX ΔPep4/2402, and SYT-SSX/2402). Target cells were incubated with Tcr-T1 cells at ratio 0:1, 0.2:1, 1:1, and 5:1 (T-cell/SW982). Cell viability was determined by using CCK-8. ( E ) SW982 cells overexpressing HLA-A*2402 and SYT-SSX fusion mutated peptide containing Pep-4 sequence (SYT-SSX/2402) were incubated with Tcr-T1 cells ( E : T = 1:1). Cell viability was determined by CCK-8. IFN-γ secretion was examined using a CBA assay ( n = 3 independent replication). ( F ) Cytotoxicity of Tcr-T1 against HLA-A*2402-positive MKN45 was determined as indicated ( n = 3 independent replication). ( G – I ) Treatment scheme ( G , up). SYT-SSX/2402 and SYT-SSX ΔPep4/2402 cells (1 × 10 7 cells/mouse) were injected subcutaneously in nude mice ( n = 6). When tumor volume reached to ~100 mm 3 , Tcr-T1 cells, and UTD T cells (5 × 10 6 cells/mouse) were intravenously injected. Mice were sacrificed at the endpoint. Representative photographs of tumor tissues isolated from each group of mice ( G , bottom). Tumor volumes of each group were recorded ( H ). The overall survival of each group was recorded. ( I ) Data are presented as mean ± SEM. Statistical significance was calculated using the Student’s t test ( C , E ), one-way ANOVA ( C ), two-way ANOVA ( D – F , H ), and Kaplan–Meier survival analysis ( I ). ns not significant, * P < 0.05; ** P < 0.01; *** P < 0.001. .

Article Snippet: Human synovial sarcoma cell line SW982 (HLA-A24-, SYT/SSX fusion-negative), human gastric adenocarcinoma cell line MKN45 (HLA-A24 + , SYT/SSX fusion-negative), human breast cancer cell line MDA-MB-231 (HLA-A02 + , SYT/SSX fusion-negative) and human transporter-associated protein (TAP)-deficient cell line T2 was originally purchased from ATCC, China Center for Type Culture Collection (CCTCC) and the typical cell culture collection of the Committee of the Chinese Academy of Sciences Library.

Techniques: Generated, Purification, In Vitro, Electroporation, CCK-8 Assay, Modification, Expressing, Transfection, Sequencing, Incubation, Injection, Isolation

Journal: EMBO Molecular Medicine

Article Title: A therapeutic regimen using neoantigen-specific TCR-T cells for HLA-A*2402-positive solid tumors

doi: 10.1038/s44321-024-00184-1

Figure Lengend Snippet: Reagents and tools table

Article Snippet: Human synovial sarcoma cell line SW982 (HLA-A24-, SYT/SSX fusion-negative), human gastric adenocarcinoma cell line MKN45 (HLA-A24 + , SYT/SSX fusion-negative), human breast cancer cell line MDA-MB-231 (HLA-A02 + , SYT/SSX fusion-negative) and human transporter-associated protein (TAP)-deficient cell line T2 was originally purchased from ATCC, China Center for Type Culture Collection (CCTCC) and the typical cell culture collection of the Committee of the Chinese Academy of Sciences Library.

Techniques: Cell Culture, Recombinant, Bicinchoninic Acid Protein Assay, Software