Payload-delivering engineered γδ T cells display enhanced cytotoxicity, persistence, and efficacy in preclinical models of osteosarcoma

# γδ T cells engineered to secrete immune-active payloads display enhanced persistence and efficacy in preclinical models of osteosarcoma [https://doi.org/10.5061/dryad.q2bvq83t1](https://doi.org/10.5061/dryad.q2bvq83t1) 2 datasets are provided, one pertaining to the main figures (Dataset 1) and one to the supplementary. figures (Dataset 2). The datasets comprise tabulated data for all of the figures shown. ## Description of the data and file structure Data is shown in tabulated Excel files, with each tab pertaining to a panel in the figures **Data File S1** 1B: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with Unmodified gdT cells or OPS-gdT cells, as determined by flow cytometry 1C: Absolute numbers of gdT cells ± engineering at different days of expansion in the conditions indicated in the columns 1D-E: Differences in expression or indicated markers between unmodified-gdT cells or 14G2a stIL15-OPS-gdT cells expressed as Earth Movers Distance values, derived from mass cytometric analysis. Calculation of Earth Movers Distance for this and subsequent figures was done using a custom Python script which is available at: [https://doi.org/10.5281/zenodo.10993245.](https://doi.org/10.5281/zenodo.10993245.) 1F: Fold change of SUP-T1-wt or SUP-T1-GD2 tumour cell numbers in co-culture with gdT cells ± engineering. Numbers shown are fold changes in target cell number, relative to baseline. 2B: % human Vd2 gdT cells in mouse blood as determined by flow cytometry 2C: Bioluminescence of subcutaneous SUP-T1-GD2 tumours in NSG mice treated with gdT cells ± engineering, over a 15 day period 2D: Survival of mice bearing subcutaneous SUP-T1-GD2 tumours treated with gdT cells ± engineering 3C: DREMI scores for the interaction between transgene and pERK in CAR-abT cells and 14G2a-stIL15-OPS-gdT cells in the presence or absence of antigen-postivie targets. DREMI scores derived from mass cytometric signalling analysis and calculated using simpledremi ([https://dpeerlab.github.io/dpeerlab-website/dremi-download.html](https://dpeerlab.github.io/dpeerlab-website/dremi-download.html)) 3E: DREMI scores for the interaction between transgene and key signalling markers in CAR-abT cells, calculated from mass cytometric data 3F: DREMI scores for the influence of transgene expression on key markers in 14G2a-stIL15-OPS-gdT cells, calculated from mass cytometric data 3H: Earth movers distance for the differences between transduced and non-transduced abT or gdT cells, with corresponding DREMI scores for the influence of transgene expression on key markers in 14G2a-stIL15-OPS-gdT or CAR-abT cells, calculated from mass cytometric data 4A: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with Unmodified gdT cells in the presence of supernatant from unmodified gdT supernatant or 14G2a-OPS-gdT supernatant, as determined by flow cytometry 4B: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with fresh NK cells in the presence of supernatant from unmodified gdT supernatant, 14G2a-OPS-gdT supernatant, 14G2a-stIL15-OPS-gdT supernatant or 1ug/ml ch14.18 as determined by flow cytometry 4C: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with macrophages in the presence of supernatant from unmodified gdT supernatant, 14G2a-OPS-gdT supernatant, 14G2a-stIL15-OPS-gdT supernatant or 1ug/ml ch14.18 as determined by flow cytometry and expressed as % death relative to the mean of the tumour alone control. 5B: Dead cell % for SUP-T1-GD2 targets co-cultured with unmodified gdT cells in the presence of different concentrations of ch14.18, as determined by flow cytometry 5C: Dead cell % for SUP-T1-GD2 targets co-cultured with unmodified gdT cells in the presence of 1 ng/ml ch14.18, as determined by flow cytometry 5D: Relative fluorescence intensity for chemokine receptor expression in gdT cells ± engineering, as determined by flow cytometry and compared to FMO controls. 6B: Differences in marker expression between unmodified gdT cell monocultures and 14G2a-stIL15-OPS-gdT cells ± co-culture with osteosarcoma cells, expressed as earth movers distance derived from mass cytometric data 6C: Differences in marker expression between osteosarcoma monocultures and those in co-culture with unmodified gdT or 14G2a-stIL15-OPS-gdT cells, expressed as earth movers distance derived from mass cytometric data 6D: Killing of luciferase-transduced osteosarcoma cells by gdT cells ± engineering ± ch14.18, expressed as luminescence relative to targets alone 6E: Killing of luciferase-transduced osteosarcoma cells by eingeered gdT cells ± zoledronic acid, expressed as luminescence relative to targets alone 7A: Tumour growth of osteosarcoma in NSG mice treated with the indicated interventions, expressed as total bioluminescent flux over time 7B: Event free survival of NSG mice treated with the indicated interventions. An event is defined as tumour total flux doubling. **Data file S2** S1C: Transduction efficiency of gdT cells as determined by % GFP expression using flow cytometry. Vd2 % in the specified preparations (also detemrined by flow cytometry) is also shown S1E: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with Unmodified gdT cells, as determined by flow cytometry S1F: Median Fluorescence Intensity Standard curve generation for determination of SFP concentration by flow cytometry, with curresponding SFP yield from engineered gdT cells S2B: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with Unmodified gdT cells, 14G2a-OPS gdT cells or 14G2a CAR-gdT cells, as determined by flow cytometry S2C: Relative tumour cell count for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with Unmodified gdT cells, 14G2a-OPS gdT cells or 14G2a CAR-gdT cells, as determined by flow cytometry and relative to baseline cell numbers S2D: Median fluorecence intensity of 4 markers associated with exhaustion on 14G2a-OPS gdT cells or 14G2a CAR-gdT cells, determined by flow cytometry S3B: % GFP expression over time on unmodified or engineered Vd2+CD3+ cells in presence or absence of IL-2 as determined by flow cytometry S3D: Concentration of IL-15 in supernatants from unmodified or engineered gdT cells, as determined by IL-15 ELISA S3E-I: Differences in expression or indicated markers between unmodified-gdT cells or stIL15-gdT cells expressed as Earth Movers Distance values, derived from mass cytometric analysis S4A: Frequency of CD3+Vd2+ cells within live cell populations in the blood of NSG mice at 7 and 14 days after injection of indicated cell type S4B: Concentration of IL-15 in supernatants from unmodified or engineered gdT cells, as determined by IL-15 ELISA S4D: Concentration of SFP in supernatants from unmodified or engineered gdT cells, as determined by flow cytometry S4F: Fold change in unmodfiied or engineered Vd2 numbers during expansion S4G: Percentage of Vd2+CD3+ cells at D0 and D12 of expansion, comparin unmodified and engineered gdT cells S4H: Absolute Vd2+CD3+ cell counts for cultures of engineered gdT cells S5A-C: Differences in expression or indicated markers between unmodified-gdT cells or 14G2a-stIL15-OPS-gdT cells expressed as Earth Movers Distance values, derived from mass cytometric analysis S5E: Numbers of engineered or unmodfieid Vd2+ cells in co-cultures with SUP-T1-WT or SUP-T1-GD2, expressed relative to cell numbers at D0 S5G: Killing of Kelly neuroblastoma cells by unmodified or engineered gdT cells, expressed relative to target only S6C-D: Subcutaneous tumour volumes (mm3) for SUP-T1-GD2 tumours in NSG mice treated with unmodified or engineered gdT cells S8C: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with stem-cell derived neutrophils, in the presence of supernatant from 14G2a-OPS gdT cells or unmodified gdT cells, as determined by flow cytometry S9B: Dead cell % for SUP-T1-wt targets or SUP-T1-GD2 targets co-cultured with unmodified gdT cells with varying CD16 expression (expressed as % of cells CD16+) in the presence of 1ug/ml ch14.18 antibody S9C: % Expression of CD16 on CD3+Vd2+ cells ± engineering on D12-14 of expansion, as determined by flow cytometry S9D: CD45RA/CD27 expression on CD3+Vd2+ cells ± engineering on D12 of expansion, as determined by flow cytometry S10: Dead cell % for SUP-T1-GD2 targets co-cultured with unmodified gdT cells or engineered gdT cells which were prepared in a G-REX culture system S12A: Differences in marker expression between unmodified gdT cell monocultures and 14G2a-stIL15-OPS-gdT cells ± co-culture with PDOS25 osteosarcoma cells, expressed as earth movers distance derived from mass cytometric data S12A: Differences in marker expression between unmodified gdT cell monocultures and 14G2a-stIL15-OPS-gdT cells ± co-culture with PDOS19 osteosarcoma cells, expressed as earth movers distance derived from mass cytometric data S13: Killing of luciferase-transduced osteosarcoma cells by NK cells ± supernatasnt from engineered gdT cells, expressed as luminescence relative to targets alone S14A: Pre-treatment bioluminescent total flux for NSG mice with intratibial xenografts of luciferase expressing PDOS25, divided by treatment group S15B: Transduction efficiency of 14G2a-CAR abT cells as determined by flow cytometry S15C: Killing of luciferase-transduced PDOS25 osteosarcoma cells or Kelly neuroblastoma cells by 14G2a CAR abT cells or unmodified abT cells, expressed as luminescence relative to targets alone S15D: Expression of GD2 on surviving cells after co-culture with 14G2a CAR abT cells or unmodified abT cells, expressed relative to starting levels of GD2 expression S16A: Event free survival of NSG mice treated with the indicated interventions. An event is defined as tumour total flux doubling.

搜集汇总

背景与挑战

背景概述

该数据集支持一项关于工程化γδ T细胞免疫疗法治疗骨肉瘤的研究。数据集包含两个Excel文件，提供了研究中所有主图和补充图的表格化实验数据，主要涉及工程化γδ T细胞（stIL15-OPS-γδ T细胞）在体外和体内模型中的细胞毒性、持久性、旁观者免疫激活及抗肿瘤疗效的评估结果。这些数据验证了该工程化细胞平台结合直接细胞裂解和旁观者激活以促进肿瘤控制的潜力。