Journal: Oncoimmunology
Article Title: Tumor-infiltrating lymphocytes-derived CD8 + clonotypes infiltrate the tumor tissue and mediate tumor regression in glioblastoma
doi: 10.1080/2162402X.2025.2559784
Figure Lengend Snippet: Treatment schedule and TIL product characteristics. (A) Treatment schedule indicating TIL infusion and cyclophosphamide administration. A partial resection of the GBM tumor in the patient’s occipital lobe was performed 6 weeks prior to TIL-1 infusion (day −43) to obtain tissue for TIL manufacturing. The patient was treated with 60 mg/kg cyclophosphamide (CTX) one day prior to TIL infusion on either occasion (TIL-1 and TIL-2 infusions). Eight hours after TIL infusion, IL-2 (60,000 IU/kg) was administered i.V., followed by anti-TNF-α (soluble TNF receptor, etanercept, 25 mg) s.C. And anti-IL-6 receptor (IL-6 R, tocilizumab, 4 mg/kg) i.V. To prevent systemic hyperinflammatory reactions 24 and 72 hours after TIL-1 and TIL-2 infusion, respectively. Serial sampling was performed to assess tumor mutation burden, transcriptomics and TIL persistence/infiltration analysis. (B) Flow cytometric analysis of T-cell phenotypes (CD4 + and CD8 + ) and contaminants evaluation (CD56 + NK-cells and tregs) in manufactured TIL products. (C) CD107a induction in CD4 + /CD8 + T-cells post-PMA stimulation and IFN-γ production in TIL products after 24-hours stimulation with OKT3 were used to gauge TIL functionality prior to infusion. (D) Cytotoxic potential of the TIL products as assessed by standard chromium release. Cr 51 -labeled autologous tumor cell line (ATCL) or control tumor cell lines (targets) were incubated with tumor-infiltrating lymphocytes (TILs, effectors) at varying effector-to-target (E:T) ratios, and cytotoxic activity was calculated based on Cr 51 release into the supernatant. Controls included allogeneic GBM cell lines (U-373, DBTRG05), Daudi B-lymphoma cell line and autologous EBV-transformed B-cell line. (E) Cold target inhibition assays using a constant E:T ratio of 90:1, varying cell numbers of cold tumor cells were incubated with hot tumor cells to titrate the blocking of TIL activity. Highest blocking was commensurate in the presence of higher numbers of cold tumor cells (up to almost 100% at 90:1 and approximately 95% at 30:1) co-incubated with hot tumor cells. A set ratio of cold:hot tumor cell was used (90:1) to gauge TIL activity at varying cell numbers of TILs.
Article Snippet: Allogeneic GBM cell lines U-373 (ATCC no: HTB-17) and DBTRG05 (ATCC no: CRL-2020), Daudi B-lymphoma cell line (ATCC no: CRL-213) and the autologous EBV-transformed B cell line served as controls.
Techniques: Sampling, Mutagenesis, Labeling, Control, Incubation, Activity Assay, Transformation Assay, Inhibition, Blocking Assay
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