Journal: Journal of Personalized Medicine
Article Title: αβ-T Cells Engineered to Express γδ-T Cell Receptors Can Kill Neuroblastoma Organoids Independent of MHC-I Expression
doi: 10.3390/jpm11090923
Figure Lengend Snippet: Neuroblastoma tumor organoids are killed by TEG002 in the presence of Pamidronate, independent of MHC-I expression. ( A ) Expression of HLA-a,b,c of all six organoids, measured by flow cytometry. Green indicates “No Activation” and red indicates “Activation” of TEG002. ( B ) Detailed dynamics of AMC691B killing by TEG002 over time. Representative graphs of one effector donor, which was tested in three different E:T ratios, are shown. Red line indicates the maximal cell lysis induced by the lysis control. ( C ) Killing of all four organoids by TEG002 over time at a 1:1 E:T ratio. AMC691T, AMC691B, and NB067 were tested with effectors from two different donors and NB129 with effectors from one donor. Graphs show mean of all experiments. ( D ) Bargraph of cytolysis at 48 h. Only the 1:1 E:T ratio is shown here. ( E ) Expression of HLA-a,b,c on four selected organoids in co-culture setting, measured by flow cytometry. Median Fluorescence Intensity (MFI) of HLA-a,b,c was used. Flow cytometry was performed only on organoids in co-culture with TEG002, with the addition of PAM at E:T 1:1. A Friedman test and a Dunn’s post hoc test, in comparison to T = 0, were performed to determine significant upregulation. ( F ) Cytolysis of AMC691T and AMC691B by TEG002 and untransduced αβ-T cells in the presence of 10μg/mL MHC-I blocking antibody (clone W6/32, Bio-Techne, ) or isotype control. Lysis control = Puromycin (1 μg/mL); PAM = Pamidronate; E:T = Effector to Target; all data are normalized to the target only condition; lysis control at 24 h.
Article Snippet: For two organoids, the cytotoxicity and MHC-I independent recognition was tested in the presence of 10 μg/mL MHC-I blocking antibody (clone W6.32, Cat. NB100-64775, BioTechne, Minneapolis, MN, USA) or 10 μg/mL IgG isotype control (Cat. 31903, ThermoFisher, Waltham, MA, USA).
Techniques: Expressing, Flow Cytometry, Activation Assay, Lysis, Co-Culture Assay, Fluorescence, Comparison, Blocking Assay
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