García-Salum Tamara, Villablanca Andrea, Matthäus Franziska, Tittarelli Andrés, Baeza Mauricio, Pereda Cristián, Gleisner M Alejandra, González Fermín E, López Mercedes N, Hoheisel Jörg D, Norgauer Johannes, Gebicke-Haerter Peter J, Salazar-Onfray Flavio
Disciplinary Program of Immunology, Institute of Biomedical Sciences, Faculty of Medicine, Universidad de Chile, 8380453 Santiago, Chile.
Millennium Institute on Immunology and Immunotherapy, Faculty of Medicine, Universidad de Chile, 8380453 Santiago, Chile.
Oncotarget. 2018 Mar 30;9(24):17014-17027. doi: 10.18632/oncotarget.24795.
We previously showed that autologous dendritic cells (DCs) loaded with an allogeneic heat shock (HS)-conditioned melanoma cell-derived lysate, called TRIMEL, induce T-cell-mediated immune responses in stage IV melanoma patients. Importantly, a positive delayed-type hypersensitivity (DTH) reaction against TRIMEL after vaccination, correlated with patients prolonged survival. Furthermore, we observed that DTH reaction was associated with a differential response pattern reflected in the presence of distinct cell subpopulations in peripheral blood. Detected variations in patient responses encouraged molecular studies aimed to identify gene expression profiles induced after vaccination in treated patients, allowing the identification of new molecular predictive markers.
Gene expression patterns were analyzed by microarrays during vaccination, and some of them confirmed by quantitative real-time reverse transcriptase PCR (qRT-PCR) in the total leukocyte population of a representative group of responder and non-responder patients. New candidates for biomarkers with predictive value were identified using bioinformatics, molecular analysis, and flow cytometry.
Seventeen genes overexpressed in responder patients after vaccination respect to non-responders were identified after a mathematical analysis, from which ten were linked to immune responses and five related to cell cycle control and signal transduction. In immunological responder patients, increased protein levels of the chemokine receptor CXCR4 and the Fc-receptor CD32 were observed on cell membranes of CD8+ T and B cells and the monocyte population, respectively, confirming gene expression results.
Our study contributes to finding new molecular markers associated with clinical outcome and better understanding of clinically relevant immunological responses induced by anti-tumor DC-vaccines.
我们之前的研究表明，负载同种异体热休克（HS）预处理的黑色素瘤细胞裂解物（称为TRIMEL）的自体树突状细胞（DC）可在IV期黑色素瘤患者中诱导T细胞介导的免疫反应。重要的是，接种疫苗后针对TRIMEL的阳性迟发型超敏反应（DTH）与患者的生存期延长相关。此外，我们观察到DTH反应与外周血中不同细胞亚群的存在所反映的差异反应模式相关。患者反应的检测差异促使进行分子研究，旨在确定治疗患者接种疫苗后诱导的基因表达谱，从而识别新的分子预测标志物。
在接种疫苗期间通过微阵列分析基因表达模式，并在一组有反应和无反应患者的代表性总白细胞群体中通过定量实时逆转录聚合酶链反应（qRT-PCR）对其中一些进行确认。使用生物信息学、分子分析和流式细胞术鉴定具有预测价值的生物标志物新候选物。
经过数学分析，确定了17个在接种疫苗后有反应的患者中相对于无反应患者过度表达的基因，其中10个与免疫反应相关，5个与细胞周期控制和信号转导相关。在免疫反应患者中，分别在CD8 + T细胞、B细胞和单核细胞群体的细胞膜上观察到趋化因子受体CXCR4和Fc受体CD32的蛋白质水平升高, 证实了基因表达结果。
我们的研究有助于发现与临床结果相关的新分子标志物，并更好地理解抗肿瘤DC疫苗诱导的临床相关免疫反应。
