靶向HERV-K修饰的嵌合抗原受体修饰T细胞通过下调Ras抑制乳...

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Chimeric antigen receptor T cells targeting HERV-K inhibit breast cancer and its metastasis through downregulation of Ras.  

靶向HERV-K修饰的嵌合抗原受体修饰T细胞通过下调Ras抑制乳腺癌及其转移

摘要：

我们此前曾报道，人类内源性逆转录酶病毒k（HERV-K）外壳（ENV）蛋白是一种用来制作癌症疫苗的肿瘤相关抗原（TAA），其抗体（单克隆抗体）具有抗肿瘤的抗肿瘤活性。在这个研究中，我们使用抗HERV-K 单克隆抗体来生产靶向HERV-K 外壳蛋白的CAR-T细胞（K-CAR-T细胞）。K-CAR T细胞从9名乳腺癌（BC）患者体内和12位正常献血者的外周单个核细胞(PBMCs)中提取，均能对乳腺癌细胞表现出抑制并具有明显的细胞毒性，而对正常的MCF 10A乳腺细胞没有影响。当使用控制T细胞，或者使用shRNA将HERV-K的表达敲除，K-CAR T细胞对癌症的抑制效果就大大降低。多种细胞因子的分泌，包括干扰素γ，TNF-α和IL-2的使用能够增加K-CARs抑制乳腺癌细胞的效果。异种移植MDA-MB-231 或者MDA-MB-435.eB1 乳腺癌细胞的模型中，可以观察到肿瘤生长速度和细胞质量明显减少。此外，K-CAR使得HERV-K在小鼠肿瘤表面表达下调，并且会伴随p53的上调以及MDM2和p-ERK的下调。重要的是，使用K-CAR处理的肿瘤组织中，HERV-K壳蛋白的表达与Ras的表达有关。我们的结果暗示了HERV-K壳蛋白是一种肿瘤蛋白，可能在与p53和Ras信号途径相关的肿瘤发生过程中扮演了很重要的角色。抗HERV-K治疗，包括了K-CAR T细胞治疗方式，显示了免疫治疗乳腺癌的潜能。

Abstract

We have previously reported that human endogenous retrovirus-K (HERV-K) envelope (env) protein is a tumor-associated antigen (TAA) for cancer vaccines, and that its antibodies (mAbs) possess antitumor activity against cancer. In this study, a chimeric antigen receptor (CAR) specific for HERV-K env protein (K-CAR) was generated using anti-HERV-K mAb. K-CAR T cells from peripheral blood mononuclear cells (PBMCs) of 9 breast cancer (BC) patients and 12 normal donors were able to inhibit growth of, and to exhibit significant cytotoxicity toward, BC cells but not MCF-10A normal breast cells. The antitumor effects in cancer cells were significantly reduced when control T cells were used, or the expression of HERV-K was knocked down by an shRNA. Secretion of multiple cytokines, including IFNγ, TNF-α, and IL-2, was significantly enhanced in culture media of BC cells treated with K-CARs. Significantly reduced tumor growth and tumor weight was observed in xenograft models bearing MDA-MB-231 or MDA-MB-435.eB1 BC cells. Importantly, the K-CAR prevented tumor metastasis to other organs.Furthermore, downregulation of HERV-K expression in tumors of mice treated with K-CAR correlated with upregulation of p53 and downregulation of MDM2 and p-ERK. Importantly, the expression of HERV-K env protein in metastatic tumor tissues treated with K-CAR T cells correlated with the expression of Ras. Our results indicate that HERV-K env protein is an oncoprotein and may play an important role in tumorigenesis related to p53 and Ras signaling pathways. Anti-HERV-K treatment, including K-CAR treatment, shows potential for immunotherapy of BC.

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