PolyI:C–Induced, TLR3/RIP3-Dependent Necroptosis Backs Up Immune Effector–Mediated Tumor Elimination In Vivo
PolyI:C–Induced, TLR3/RIP3-Dependent Necroptosis Backs Up Immune Effector–Mediated Tumor Elimination In Vivo
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DOI：
10.1158/2326-6066.cir-14-0219
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发表时间：
2015-04
影响因子：
10.1
通讯作者：
R. Takemura;Hiromi Takaki;S. Okada;H. Shime;T. Akazawa;H. Oshiumi;M. Matsumoto;T. Teshima;T. Seya
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文献类型：
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作者：
R. Takemura;Hiromi Takaki;S. Okada;H. Shime;T. Akazawa;H. Oshiumi;M. Matsumoto;T. Teshima;T. Seya
Takemura及其同事表明，在不存在NK细胞和CTL的情况下，polyI：C+zVAD诱导CT 26肿瘤坏死性凋亡和消退，并且针对肿瘤的半胱天冬酶抑制剂通过调节肿瘤微环境和树突状细胞介导的抗肿瘤反应来增强RNA辅助免疫疗法的功效。双链RNA直接作用于成纤维细胞和骨髓谱系，诱导坏死性凋亡，如TNFα。在这里，我们研究了这种类型的细胞死亡是否发生在癌细胞中，以响应聚肌胞苷酸（polyI：C）和泛半胱天冬酶抑制剂z-Val-Ala-Asp氟甲基酮（zVAD）。我们发现结肠癌细胞系CT 26对坏死性凋亡高度敏感，如用膜联蛋白V/碘化丙啶染色所示。CT 26细胞具有RNA传感器TLR 3和MDA 5，它们通过干扰素（IFN）诱导途径上调，并分别通过TICAM-1或MAVS衔接子与受体相互作用蛋白激酶（RIP）1/3激活相关。尽管单独外源性添加polyI：C在CT 26细胞中仅轻微诱导坏死性凋亡，但polyI：C和zVAD的联合方案在体外诱导约50%的CT 26坏死性凋亡，而无TNFα或I型IFN的继发作用。CT 26坏死性凋亡依赖于肿瘤细胞中的TLR 3-TICAM-1-RIP 3轴产生活性氧，而不依赖于MDA 5、MAVS或半胱天冬酶/炎性小体激活。然而，在施用polyI：C + zVAD的CT 26肿瘤植入的Balb/c小鼠模型中，RNA衍生的坏死性凋亡在体内几乎没有再现。仅当将polyI：C（100 μg）和zVAD（1 mg）腹膜内注射到细胞毒性T淋巴细胞和自然杀伤细胞耗尽的荷瘤小鼠中时，才显示CT 26肿瘤的显著缩小。结果用没有淋巴细胞的免疫受损小鼠证实。尽管坏死性凋亡诱导的肿瘤生长迟缓在机制上显得复杂，并且依赖于polyI：C和zVAD的注射途径，但针对肿瘤细胞的抗胱天蛋白酶试剂将通过调节杀肿瘤微环境和树突状细胞诱导的抗肿瘤免疫系统的形成，使RNA佐剂免疫治疗更有效。Cancer Immunol Res; 3（8）; 902-14.©2015 AACR.
Takemura and colleagues show that polyI:C+zVAD induced CT26 tumor necroptosis and regression in the absence of NK cells and CTLs, and caspase inhibitors directed to the tumor enhanced the efficacy of RNA adjuvant immunotherapy by modulating the tumor microenvironment and dendritic cell–mediated antitumor response. Double-stranded RNA directly acts on fibroblast and myeloid lineages to induce necroptosis as in TNFα. Here, we investigated whether this type of cell death occurred in cancer cells in response to polyinosinic–polycytidylic acid (polyI:C) and the pan-caspase inhibitor z-Val-Ala-Asp fluromethyl ketone (zVAD). We found that the colon cancer cell line CT26 is highly susceptible to necroptosis, as revealed by staining with annexin V/propidium iodide. CT26 cells possess RNA sensors, TLR3 and MDA5, which are upregulated by interferon (IFN)-inducing pathways and linked to receptor-interacting protein kinase (RIP) 1/3 activation via TICAM-1 or MAVS adaptor, respectively. Although exogenously added polyI:C alone marginally induced necroptosis in CT26 cells, a combined regimen of polyI:C and zVAD induced approximately 50% CT26 necroptosis in vitro without secondary effects of TNFα or type I IFNs. CT26 necroptosis depended on the TLR3–TICAM-1–RIP3 axis in the tumor cells to produce reactive oxygen species, but not on MDA5, MAVS, or the caspases/inflammasome activation. However, the RNA-derived necroptosis was barely reproduced in vivo in a CT26 tumor–implanted Balb/c mouse model with administration of polyI:C + zVAD. Significant shrinkage of CT26 tumors was revealed only when polyI:C (100 μg) was injected intraperitoneally and zVAD (1 mg) subcutaneously into tumor-bearing mice that were depleted of cytotoxic T lymphocytes and natural killer cells. The results were confirmed with immune-compromised mice with no lymphocytes. Although necroptosis-induced tumor growth retardation appears mechanistically complicated and dependent on the injection routes of polyI:C and zVAD, anti-caspase reagent directed to tumor cells will make RNA adjuvant immunotherapy more effective by modulating the formation of the tumoricidal microenvironment and dendritic cell–inducing antitumor immune system. Cancer Immunol Res; 3(8); 902–14. ©2015 AACR.