Generating PDTO-specific T cells

Co-culture of PDTO and autologous immune cells will be based on the protocol described by Cattaneo et al. [21 (link)] Briefly, PDTO specific T cells will be induced through serial co-culture with dissociated PDTO. PBMC (~ 10.10⁶ cells) will be thawed and set to resting condition with IL-2 (150 U/mL) overnight. Meanwhile, PDTO will be treated with IFNγ (200 ng/mL) for 24 hours to favor antigen presentation and will be dissociated to produce Antigen Presenting Tumor Cells (~ 0.5.10⁶ cells APTC). PBMC and APTC will be next co-cultured (20:1 ratio) in a CD28 (5 μg/mL) coated culture plate for successive periods of 7 days to induce clonal expansion of PDTO specific T cells. T cells will be then evaluated for their tumor reactivity through the detection of activation (CD137) and functional (CD107a, IFNγ) markers by flow cytometry (Cytoflex, Beckman Coulter) and will be cryopreserved for later use for the evaluation or response to treatments.

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Perréard M., Florent R., Divoux J., Grellard J.M., Lequesne J., Briand M., Clarisse B., Rousseau N., Lebreton E., Dubois B., Harter V., Lasne-Cardon A., Drouet J., Johnson A., Le Page A.L., Bazille C., Jeanne C., Figeac M., Goardon N., Vaur D., Micault E., Humbert M., Thariat J., Babin E., Poulain L., Weiswald L.B, & Bastit V. (2023). ORGAVADS: establishment of tumor organoids from head and neck squamous cell carcinoma to assess their response to innovative therapies. BMC Cancer, 23, 223.

Publication 2023

Cytoflex

Antigen presentation Antigen presenting cells Cd137 Cells Clonal cells Flow cytometry Ifnγ T cells Tumor

Corresponding Organization :

Other organizations : Cancers et Préventions, Centre François Baclesse, Université de Lille, Laboratoire de Physique Corpusculaire de Caen, École Nationale Supérieure d'Ingénieurs de Caen

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Variable analysis

independent variables

Serial co-culture with dissociated PDTO
Treatment of PDTO with IFNγ (200 ng/mL) for 24 hours

dependent variables

Induction of PDTO-specific T cells
Tumor reactivity of T cells (measured through the detection of activation (CD137) and functional (CD107a, IFNγ) markers by flow cytometry)

control variables

PBMC (~ 10.10^6 cells) resting condition with IL-2 (150 U/mL) overnight
Co-culture of PBMC and Antigen Presenting Tumor Cells (APTC, ~ 0.5.10^6 cells) in a CD28 (5 μg/mL) coated culture plate

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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