Journal: bioRxiv

Article Title: Sensitivity to Vγ9Vδ2TCR T cells is imprinted after single mutations during early oncogenesis

doi: 10.1101/2024.11.19.624272

Figure Lengend Snippet: (A) MCF10a cells were transduced with different ErbB2 variants and co-cultured with either Vγ9Vδ2TCR or HER2-CAR transduced T cells. Tumor cells were pre-treated with the PI3K kinase inhibitor Pictilisib at 2uM overnight. After an overnight co-culture, supernatant was used to determine IFNγ production by the Vγ9Vδ2TCR T cells. (B) Furthermore, tumor cells were isolated and stained for BTN2A1 via TCR tetramer staining and (C) BTN3A cell surface expression. (D) Protein expression of HER2, phosphorylated AKT (pAKT) and total AKT in MCF10a mutant lines cultured for 24h in full culture medium (F) or medium without additional growth factors (S) . (E) Multiple tumor cell lines were co-cultured with Vγ9Vδ2TCR T-cells after pre-treatment with either the PI3K kinase inhibitor, AKT inhibitor or MEK inhibitor. After an overnight co-culture, supernatant was used to determine IFNγ production by the Vγ9Vδ2TCR T cells. (F) Multiple tumor cell lines were pre-treated with either PI3K kinase inhibitor, AKT inhibitor, mTOR inhibitor Rapamycin or mTOR inhibitor Torin1 and subsequently co-cultured with Vγ9Vδ2TCR T cells. After an overnight co-culture, supernatant was used to determine IFNγ production by the Vγ9Vδ2TCR T cells. (G) Heatmap illustrating the expression of genes from the ‘PI3K_and_AKT_family’ gene list, in the WT-AKPS model, with or without PAM. (H) Heatmap of Pearson’s correlation of the expression of genes from the ‘PI3K_and_AKT_family’ and BTNx genes in the WT-AKPS model, with or without PAM.

Article Snippet: Retroviral pBabe-Puro plasmids were purchased from Addgene: ERBB2 (#40978), ERBB2::S310F (#40991), and ERBB2:: A775_G775insV, G776C (V777E) (#40979).

Techniques: Transduction, Cell Culture, Co-Culture Assay, Isolation, Staining, Expressing, Mutagenesis

Journal: bioRxiv

Article Title: Sensitivity to Vγ9Vδ2TCR T cells is imprinted after single mutations during early oncogenesis

doi: 10.1101/2024.11.19.624272

Figure Lengend Snippet: (A) MCF10a cells were transduced with different ErbB2 variants and co-cultured with either Vγ9Vδ2TCR or HER2-CAR transduced T cells. Tumor cells were pre-treated with the PI3K kinase inhibitor Pictilisib at 2uM overnight. After an overnight co-culture, supernatant was used to determine IFNγ production by the Vγ9Vδ2TCR T cells. (B) Furthermore, tumor cells were isolated and stained for BTN2A1 via TCR tetramer staining and (C) BTN3A cell surface expression. (D) Protein expression of HER2, phosphorylated AKT (pAKT) and total AKT in MCF10a mutant lines cultured for 24h in full culture medium (F) or medium without additional growth factors (S) . (E) Multiple tumor cell lines were co-cultured with Vγ9Vδ2TCR T-cells after pre-treatment with either the PI3K kinase inhibitor, AKT inhibitor or MEK inhibitor. After an overnight co-culture, supernatant was used to determine IFNγ production by the Vγ9Vδ2TCR T cells. (F) Multiple tumor cell lines were pre-treated with either PI3K kinase inhibitor, AKT inhibitor, mTOR inhibitor Rapamycin or mTOR inhibitor Torin1 and subsequently co-cultured with Vγ9Vδ2TCR T cells. After an overnight co-culture, supernatant was used to determine IFNγ production by the Vγ9Vδ2TCR T cells. (G) Heatmap illustrating the expression of genes from the ‘PI3K_and_AKT_family’ gene list, in the WT-AKPS model, with or without PAM. (H) Heatmap of Pearson’s correlation of the expression of genes from the ‘PI3K_and_AKT_family’ and BTNx genes in the WT-AKPS model, with or without PAM.

Article Snippet: pBABE-Puro retroviral vector (EV) and pBABE-Puro vectors carrying ERBB2 wild-type and mutants were co-transfected independently with pUMVC (Addgene #8449) and VSV-G (Addgene #8454) retroviral packaging plasmids into HEK-293T cells using PEI-transfection.

Techniques: Transduction, Cell Culture, Co-Culture Assay, Isolation, Staining, Expressing, Mutagenesis