Carbotti Grazia, Petretto Andrea, Naschberger Elisabeth, Stürzl Michael, Martini Stefania, Mingari Maria Cristina, Filaci Gilberto, Ferrini Silvano, Fabbi Marina
IRCCS Ospedale Policlinico San Martino, Biotherapies Unit, Largo R. Benzi 10, 16132 Genoa, Italy.
Core Facilities-Clinical Proteomics and Metabolomics, IRCCS Istituto Giannina Gaslini, Via Gerolamo Gaslini 5, 16147 Genoa, Italy.
Cancers (Basel). 2020 Feb 19;12(2):488. doi: 10.3390/cancers12020488.
We showed that IL-27 shares several effects with IFN-γ in human cancer cells. To identify novel extracellular mediators, potentially involved in epithelial ovarian cancer (EOC) biology, we analyzed the effect of IL-27 or IFN-γ on the secretome of cultured EOC cells by mass-spectrometry (nano-UHPLC-MS/MS). IL-27 and IFN-γ modulate the release of a limited fraction of proteins among those induced in the whole cell. We focused our attention on GBP1, a guanylate-binding protein and GTPase, which mediates several biological activities of IFNs. Cytokine treatment induced GBP1, 2, and 5 expressions in EOC cells, but only GBP1 was secreted. ELISA and immunoblotting showed that cytokine-stimulated EOC cells release full-length GBP1 in vitro, through non-classical pathways, not involving microvesicles. Importantly, full-length GBP1 accumulates in the ascites of most EOC patients and ex-vivo EOC cells show constitutive tyrosine-phosphorylated STAT1/3 proteins and GBP1 expression, supporting a role for Signal Transducer And Activator Of Transcription (STAT)-activating cytokines in vivo. High GBP1 gene expression correlates with better overall survival in the TCGA (The Cancer Genome Atlas) dataset of EOC. In addition, GBP1 transfection partially reduced EOC cell viability in an MTT assay. Our data show for the first time that cytokine-stimulated tumor cells release soluble GBP1 in vitro and in vivo and suggest that GBP1 may have anti-tumor effects in EOC.
我们发现白细胞介素-27（IL-27）在人类癌细胞中与干扰素-γ（IFN-γ）具有多种共同作用。为了鉴定可能参与上皮性卵巢癌（EOC）生物学过程的新型细胞外介质，我们通过质谱分析（纳升超高效液相色谱-串联质谱，nano-UHPLC-MS/MS）研究了IL-27或IFN-γ对培养的EOC细胞分泌组的影响。IL-27和IFN-γ调节全细胞诱导产生的蛋白质中有限部分的释放。我们将注意力集中在鸟苷酸结合蛋白1（GBP1）上，它是一种鸟苷酸结合蛋白和GTP酶，介导IFN的多种生物学活性。细胞因子处理可诱导EOC细胞中GBP1、GBP2和GBP5的表达，但只有GBP1被分泌。酶联免疫吸附测定（ELISA）和免疫印迹表明，细胞因子刺激的EOC细胞在体外通过不涉及微泡的非经典途径释放全长GBP1。重要的是，全长GBP1在大多数EOC患者的腹水中积累，体外培养的EOC细胞显示组成型酪氨酸磷酸化的信号转导和转录激活因子（STAT）1/3蛋白以及GBP1表达，支持体内STAT激活细胞因子的作用。在EOC的癌症基因组图谱（TCGA）数据集中，高GBP1基因表达与更好的总生存期相关。此外，在MTT试验中，GBP1转染部分降低了EOC细胞活力。我们的数据首次表明，细胞因子刺激的肿瘤细胞在体外和体内释放可溶性GBP1，并提示GBP1可能在EOC中具有抗肿瘤作用。
