Department of Pharmacology, Vanderbilt University, Nashville, TN, 37232, USA.
Vanderbilt University School of Medicine, Nashville, TN, USA.
BMC Cancer. 2020 Oct 7;20(1):970. doi: 10.1186/s12885-020-07450-8.
New treatment options for ovarian cancer are urgently required. Tumor-associated macrophages (TAMs) are an attractive target for therapy; repolarizing TAMs from M2 (pro-tumor) to M1 (anti-tumor) phenotypes represents an important therapeutic goal. We have previously shown that upregulated NF-kappaB (NF-κB) signaling in macrophages promotes M1 polarization, but effects in the context of ovarian cancer are unknown. Therefore, we aimed to investigate the therapeutic potential of increasing macrophage NF-κB activity in immunocompetent mouse models of ovarian cancer.
We have generated a transgenic mouse model, termed IKFM, which allows doxycycline-inducible overexpression of a constitutively active form of IKK2 (cIKK2) specifically within macrophages. The IKFM model was used to evaluate effects of increasing macrophage NF-κB activity in syngeneic murine TBR5 and ID8-Luc models of ovarian cancer in two temporal windows: 1) in established tumors, and 2) during tumor implantation and early tumor growth. Tumor weight, ascites volume, ascites supernatant and cells, and solid tumor were collected at sacrifice. Populations of macrophages and T cells within solid tumor and/or ascites were analyzed by immunofluorescent staining and qPCR, and soluble factors in ascitic fluid were analyzed by ELISA. Comparisons of control versus IKFM groups were performed by 2-tailed Mann-Whitney test, and a P-value < 0.05 was considered statistically significant.
Increased expression of the cIKK2 transgene in TAMs from IKFM mice was confirmed at the mRNA and protein levels. Tumors from IKFM mice, regardless of the timing of doxycycline (dox) administration, demonstrated greater necrosis and immune infiltration than control tumors. Analysis of IKFM ascites and tumors showed sustained shifts in macrophage populations away from the M2 and towards the anti-tumor M1 phenotype. There were also increased tumor-infiltrating CD3/CD8 T cells in IKFM mice, accompanied by higher levels of CXCL9, a T cell activating factor secreted by macrophages, in IKFM ascitic fluid.
In syngeneic ovarian cancer models, increased canonical NF-κB signaling in macrophages promoted anti-tumor TAM phenotypes and increased cytotoxic T cell infiltration, which was sufficient to limit tumor progression. This may present a novel translational approach for ovarian cancer treatment, with the potential to increase responses to T cell-directed therapy in future studies.
迫切需要新的卵巢癌治疗选择。肿瘤相关巨噬细胞（TAMs）是治疗的一个有吸引力的靶点；将 TAMs 从 M2（促肿瘤）重编程为 M1（抗肿瘤）表型是一个重要的治疗目标。我们之前已经表明，巨噬细胞中 NF-κB（NF-κB）信号的上调促进了 M1 极化，但在卵巢癌的背景下尚不清楚。因此，我们旨在研究在卵巢癌免疫活性小鼠模型中增加巨噬细胞 NF-κB 活性的治疗潜力。
我们构建了一种转基因小鼠模型，称为 IKFM，该模型允许在巨噬细胞中特异性诱导表达组成型激活形式的 IKK2（cIKK2）。该 IKFM 模型用于评估在两个时间窗口内增加巨噬细胞 NF-κB 活性对同源小鼠 TBR5 和 ID8-Luc 卵巢癌模型的影响：1）在已建立的肿瘤中，和 2）在肿瘤植入和早期肿瘤生长期间。在处死时收集肿瘤重量、腹水体积、腹水上清液和细胞以及实体瘤。通过免疫荧光染色和 qPCR 分析实体瘤和/或腹水中的巨噬细胞和 T 细胞群体，并通过 ELISA 分析腹水可溶性因子。通过 2 尾 Mann-Whitney 检验比较对照与 IKFM 组，P 值<0.05 被认为具有统计学意义。
在 IKFM 小鼠的 TAMs 中转基因 cIKK2 的表达在 mRNA 和蛋白水平均得到证实。无论给予多西环素（dox）的时间如何，来自 IKFM 小鼠的肿瘤均显示出比对照肿瘤更大的坏死和免疫浸润。对 IKFM 腹水和肿瘤的分析表明，巨噬细胞群从 M2 向抗肿瘤 M1 表型的持续转变。在 IKFM 小鼠中也有更多的肿瘤浸润性 CD3/CD8 T 细胞，同时 IKFM 腹水中巨噬细胞分泌的 T 细胞激活因子 CXCL9 水平也更高。
在同源卵巢癌模型中，巨噬细胞中经典 NF-κB 信号的增加促进了抗肿瘤 TAM 表型和细胞毒性 T 细胞浸润，这足以限制肿瘤进展。这可能为卵巢癌治疗提供一种新的转化方法，在未来的研究中有可能增加对 T 细胞导向治疗的反应。
