Restoring Tumor Immunogenicity with Dendritic Cell Reprogramming [human RNA-Seq]

Decreased antigen presentation contributes to the ability of cancer cells to evade the immune system. We used the minimal gene regulatory network of type 1 conventional dendritic cells (cDC1) to reprogram cancer cells into professional antigen- presenting cells (tumor-APCs). Enforced expression of the transcription factors PU.1, IRF8, and BATF3 (PIB) was sufficient to induce cDC1 phenotype in 36 cell lines derived from human and mouse hematological and solid tumors. Within 9 days of reprogramming, tumor-APCs acquired transcriptional and epigenetic programs associated with cDC1 cells. Reprogramming restored the expression of antigen presentation complexes and costimulatory molecules on the surface of tumor cells, allowing the presentation of endogenous tumor antigens on MHC-I, and facilitating targeted killing by CD8+ T cells. Functionally, tumor-APCs engulfed and processed proteins and dead cells, secreted inflammatory cytokines, and cross- presented antigens to naïve CD8+ T cells. Human primary tumor cells could also be reprogrammed to increase their capability to present antigen and to activate patient-specific tumor-infiltrating lymphocytes. In addition to acquiring improved antigen presentation, tumor-APCs had impaired tumorigenicity in vitro and in vivo. Injection of in vitro generated melanoma-derived tumor-APCs into subcutaneous melanoma tumors delayed tumor growth and increased survival in mice. Antitumor immunity elicited by tumor-APCs was synergistic with immune checkpoint inhibitors. Our approach serves as a platform for the development of immunotherapies that endow cancer cells with the capability to process and present endogenous tumor antigens. Overall design: Population mRNAseq profiling of 17 human cancer cell lines and human embryonic fibroblasts (HEFs) after transduction with polycystronic vectors encoding PU.1, IRF8 and BATF3 at day 3, 5, 7 and 9. mRNA profilling of eGFP-transduced human cell lines and freshly isolated peripheral blood Dendritic cells type 1 (HLA-DR+CD11C+CD141+), Dendritic cells type 2 (HLA-DR+CD11C+CD141-CD1C+) and plasmacytoid dendritic cells (HLA-DR+CD11C-CD123+) were used as controls.

抗原呈递功能缺陷是癌细胞逃避免疫监视的关键机制之一。本研究借助1型常规树突状细胞（type 1 conventional dendritic cells, cDC1）的最小基因调控网络，将癌细胞重编程为专职抗原呈递细胞（professional antigen-presenting cells, tumor-APCs）。仅过表达转录因子PU.1、IRF8与BATF3（简称PIB），即可在源自人类与小鼠的血液系统及实体瘤的36株细胞系中诱导出cDC1表型。在重编程启动后的9天内，肿瘤-APCs即可获得与cDC1细胞高度相似的转录组与表观遗传程序。重编程恢复了癌细胞表面抗原呈递复合物与共刺激分子的表达，使得内源性肿瘤抗原可通过主要组织相容性复合体I类（MHC-I）进行呈递，并有效促进CD8阳性T细胞（CD8+ T cells）对癌细胞的靶向杀伤。功能层面，肿瘤-APCs可吞噬并加工蛋白质与死细胞，分泌炎性细胞因子，同时能将抗原交叉呈递至初始CD8阳性T细胞。人类原代肿瘤细胞亦可被成功重编程，以增强其抗原呈递能力并激活患者自身的肿瘤浸润淋巴细胞（tumor-infiltrating lymphocytes）。除获得增强的抗原呈递功能外，肿瘤-APCs在体外与体内的致瘤能力均显著受损。将体外构建的黑色素瘤来源肿瘤-APCs注射至小鼠皮下黑色素瘤病灶中，可延缓肿瘤生长并延长小鼠生存期。肿瘤-APCs诱导的抗肿瘤免疫可与免疫检查点抑制剂（immune checkpoint inhibitors）产生协同增效作用。本研究方法可作为一种通用平台，用于开发可赋予癌细胞加工并呈递内源性肿瘤抗原能力的免疫治疗策略。整体实验设计：对17株人类癌细胞系及人类胚胎成纤维细胞（human embryonic fibroblasts, HEFs）进行编码PU.1、IRF8与BATF3的多顺反子载体（polycistronic vectors）转染后，于转染后第3、5、7、9天分别进行群体mRNA测序（mRNAseq）分析。以转染eGFP载体的人类细胞系，以及新鲜分离的外周血1型树突状细胞（peripheral blood type 1 dendritic cells, HLA-DR+CD11C+CD141+）、2型树突状细胞（peripheral blood type 2 dendritic cells, HLA-DR+CD11C+CD141-CD1C+）与浆细胞样树突状细胞（plasmacytoid dendritic cells, HLA-DR+CD11C-CD123+）作为实验对照。