Hepatocellular Carcinoma Cells Up-regulate PVRL1, Stabilizing PVR and Inhibiting the Cytotoxic T-Cell Response via TIGIT to Mediate Tumor Resistance to PD1 Inhibitors in Mice
Hepatocellular Carcinoma Cells Up-regulate PVRL1, Stabilizing PVR and Inhibiting the Cytotoxic T-Cell Response via TIGIT to Mediate Tumor Resistance to PD1 Inhibitors in Mice
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DOI：
10.1053/j.gastro.2020.03.074
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发表时间：
2020-08-01
期刊：
影响因子：
29.4
通讯作者：
Wong, Carmen Chak-Lui
中科院分区：
文献类型：
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作者：
Chiu, David Kung-Chun;Yuen, Vincent Wai-Hin;Wong, Carmen Chak-Lui
背景与目的：免疫检查点抑制剂在治疗某些肝细胞癌（HCC）中是有效的，但这些肿瘤并不总是对程序性细胞死亡1（PDCD 1，也称为PD 1）抑制剂有反应。我们研究了小鼠肝肿瘤对浸润性T细胞的抵抗机制。方法：对小鼠给予成簇的规律间隔的短回文重复序列-Cas9（CRISPR-Cas9）和转座子载体的水动力尾静脉注射以破坏Trp 53并过表达C-Myc（Trp 53（KO）/C-Myc（OE）小鼠）。用短发夹RNA敲除Hepa 1 -6细胞中的Pvrl 1和Pvrl 3。将Hepa 1 -6细胞注射到C57 BL/6小鼠的肝脏中;在注射癌细胞之前，向一些小鼠腹膜内注射针对PD 1、具有IG和ITIM结构域的T细胞免疫受体（TIGIT）或CD 8的抗体。从小鼠中收集肝组织，并通过组织学、免疫组织化学和定量实时聚合酶链反应进行分析;使用标记物检测T细胞和其他淋巴细胞，通过质谱细胞术分析肿瘤。我们从香港接受手术的患者中获得了HCC和非肿瘤性肝组织和临床数据，并通过免疫组织化学分析了组织。结果：Trp 53（KO）/C-Myc（OE）小鼠在3-5周内发生肝肿瘤;注射抗PD 1并没有减缓肿瘤的发展。与未给予抗体的小鼠相比，给予抗PD 1的小鼠的肿瘤具有更多数量的记忆性CD 8（+）T细胞（CD 44（+）CD 62 L（-）KLRG 1（int））和表达PD 1、淋巴细胞活化3（LAG 3）和TIGIT的T细胞。患者的HCC组织中PVRL 1信使RNA和蛋白水平高于非肿瘤组织。PVRL 1增加与无病生存期缩短相关。Hepa 1 -6细胞中Pvrl 1的敲低导致它们在小鼠中形成更小的肿瘤，被更多表达抑制蛋白TIGIT的CD 8（+）T细胞浸润;在CD 8（+）T细胞耗尽的小鼠中没有观察到这些效应。在Hepa 1 -6细胞中，PVRL 1稳定细胞表面PVR，其与CD 8（+）T细胞上的TIGIT相互作用; Pvrll的敲低降低PVR的细胞表面水平，但不降低Pvr信使RNA的水平。在Trp 53（KO）/C-Myc（OE）小鼠和具有从Hepa 1 -6细胞生长的肿瘤的小鼠中，注射抗PD 1和抗TIGIT的组合显著降低肿瘤生长，增加肿瘤中细胞毒性T细胞与调节性T细胞的比率，并延长存活。结论：由HCC细胞上调的PVRL 1稳定细胞表面PVR，其与CD 8（+）效应记忆T细胞上的抑制分子TIGIT相互作用。这抑制了抗肿瘤免疫反应。PVRL 1/TIGIT的抑制剂，沿着与抗PD 1一起可能被开发用于治疗HCC。
BACKGROUND & AIMS: Immune checkpoint inhibitors are effective in the treatment of some hepatocellular carcinomas (HCCs), but these tumors do not always respond to inhibitors of programmed cell death 1 (PDCD1, also called PD1). We investigated mechanisms of resistance of liver tumors in mice to infiltrating T cells. METHODS: Mice were given hydrodynamic tail vein injections of clustered regularly interspaced short palindromic repeats-Cas9 (CRISPR-Cas9) and transposon vectors to disrupt Trp53 and overexpress C-Myc (Trp53(KO)/C-Myc(OE) mice). Pvrl1 and Pvrl3 were knocked down in Hepa1-6 cells by using short hairpin RNAs. Hepa1-6 cells were injected into livers of C57BL/6 mice; some mice were given intraperitoneal injections of antibodies against PD1, T-cell immunoreceptor with Ig and ITIM domains (TIGIT), or CD8 before the cancer cells were injected. Liver tissues were collected from mice and analyzed by histology, immunohistochemistry, and quantitative real-time polymerase chain reaction; tumors were analyzed by mass cytometry using markers to detect T cells and other lymphocytes. We obtained HCC and nontumorous liver tissues and clinical data from patients who underwent surgery in Hong Kong and analyzed the tissues by immunohistochemistry. RESULTS: Trp53(KO)/C-Myc(OE) mice developed liver tumors in 3-5 weeks; injections of anti-PD1 did not slow tumor development. Tumors from mice given anti-PD1 had larger numbers of memory CD8(+) T cells (CD44(+)CD62L(-)KLRG1(int)) and T cells that expressed PD1, lymphocyte activating 3 (LAG3), and TIGIT compared with mice not given the antibody. HCC tissues from patients had higher levels of PVRL1 messenger RNA and protein than nontumorous tissues. Increased PVRL1 was associated with shorter times of disease-free survival. Knockdown of Pvrl1 in Hepa1-6 cells caused them to form smaller tumors in mice, infiltrated by higher numbers of CD8(+) T cells that expressed the inhibitory protein TIGIT; these effects were not observed in mice with depletion of CD8(+) T cells. In Hepa1-6 cells, PVRL1 stabilized cell surface PVR, which interacted with TIGIT on CD8(+) T cells; knockdown of Pvrl1 reduced cell-surface levels of PVR but not levels of Pvr messenger RNA. In Trp53(KO)/C-Myc(OE) mice and mice with tumors grown from Hepa1-6 cells, injection of the combination of anti-PD1 and anti-TIGIT significantly reduced tumor growth, increased the ratio of cytotoxic to regulatory T cells in tumors, and prolonged survival. CONCLUSIONS: PVRL1, which is up-regulated by HCC cells, stabilizes cell surface PVR, which interacts with TIGIT, an inhibitory molecule on CD8(+) effector memory T cells. This suppresses the ant-tumor immune response. Inhibitors of PVRL1/TIGIT, along with anti-PD1 might be developed for treatment of HCC.