Targeted inducible delivery of immunoactivating cytokines reprograms glioblastoma microenvironment and inhibits growth in mouse models [scRNA-seq]

We exploited a lentiviral vector-based platform to engineer hematopoietic stem cells ex vivo with the aim of releasing, via their tumor-infiltrating monocyte/macrophage progeny, IFN-a or IL-12 at the tumor site in a spatial and temporal regulated manner. Taking advantage of a syngeneic GBM mouse model, we showed that inducible release of IFN-a within the TME achieved robust tumor inhibition up to eradication and outperformed systemic treatment with the recombinant protein in terms of efficacy, tolerability and specificity. Single-cell RNAseq of the tumor immune infiltrate revealed reprogramming of the immune microenvironment towards a pro-inflammatory and anti-tumoral state associated with loss of a macrophage subpopulation predicting poor prognosis in human GBM. Overall design: Single-cell RNA-seq of CD45+ tumor-infiltrating (glioblastoma) mice cells belonging to control and gene therapy treated (IFN-a and IFN-a-DHFR) groups.

本研究基于慢病毒载体（lentiviral vector）平台，对造血干细胞（hematopoietic stem cells）进行体外工程化改造，旨在通过其肿瘤浸润性单核细胞/巨噬细胞子代，在肿瘤部位以时空调控的方式释放干扰素α（IFN-α）或白细胞介素12（IL-12）。本研究利用同基因胶质母细胞瘤（GBM）小鼠模型，证实于肿瘤微环境（tumor microenvironment, TME）中诱导性释放干扰素α，可实现强效肿瘤抑制乃至肿瘤根除，且在疗效、耐受性与特异性方面均优于重组蛋白的全身给药治疗。对肿瘤免疫浸润细胞开展单细胞RNA测序（single-cell RNA sequencing）分析后发现，免疫微环境被重编程为促炎抗肿瘤状态，该状态与一类巨噬细胞亚群的缺失密切相关；而此类巨噬细胞亚群可作为人类胶质母细胞瘤预后不良的预测标志物。整体实验设计：对对照组与基因治疗处理组（IFN-α组与IFN-α-DHFR组）的CD45阳性肿瘤浸润性（胶质母细胞瘤）小鼠细胞进行单细胞RNA测序。