Arenobufagin induces cell apoptosis by modulating the cell cycle regulator claspin and the JNK pathway in nasopharyngeal carcinoma cells

The high recurrence rate and incidence of distant metastasis of nasopharyngeal carcinoma (NPC) result in poor prognosis. It is necessary to identify natural compounds that can complement combination radiation therapy. Arenobufagin is commonly used for heart diseases and liver cancer, but its effectiveness in NPC is unclear. The effect of arenobufagin-induced apoptosis was measured by a cell viability assay, tumorigenic assay, fluorescence assay, and Western blot assay through NPC-039 and NPC-BM cell lines. The protease array, Western blot assay, and transient transfection were used to investigate the underlying mechanism of arenobufagin-induced apoptosis. An NPC xenograft model was established to explore the antitumor activity of arenobufagin <i>in vivo</i>. Our findings indicated that arenobufagin exerted cytotoxic effects on NPC cells, inhibiting proliferation through apoptosis activation. Downregulation of claspin was confirmed in arenobufagin-induced apoptosis. Combined treatment with arenobufagin and mitogen-activated protein kinase inhibitors demonstrated that arenobufagin induced NPC apoptosis through the c-Jun N-terminal kinases (JNK) pathway inhibition. Furthermore, arenobufagin suppressed NPC tumor proliferation in vivo. Our results revealed the antitumor effect of arenobufagin in vitro and in vivo. Arenobufagin may have clinical utility in treating NPC due to its suppression of claspin and inhibition of the JNK pathway.

鼻咽癌（nasopharyngeal carcinoma, NPC）具有较高的复发率与远处转移发生率，患者预后不佳，因此亟需筛选可辅助联合放疗的天然化合物。阿瑞蟾毒配基（arenobufagin）通常被用于治疗心脏疾病与肝癌，但其针对鼻咽癌的疗效尚不明确。本研究通过NPC-039与NPC-BM细胞系，采用细胞活力检测、成瘤实验、荧光检测及蛋白质印迹（Western blot）实验，评估阿瑞蟾毒配基诱导的细胞凋亡效应；通过蛋白酶阵列、蛋白质印迹实验与瞬时转染技术，探究阿瑞蟾毒配基诱导细胞凋亡的潜在分子机制；此外构建鼻咽癌异种移植瘤模型，以体内（in vivo）实验探究阿瑞蟾毒配基的抗肿瘤活性。研究结果显示，阿瑞蟾毒配基对鼻咽癌细胞具有细胞毒性作用，可通过激活细胞凋亡途径抑制癌细胞增殖；实验证实阿瑞蟾毒配基诱导的凋亡过程中存在claspin的下调表达。联合使用阿瑞蟾毒配基与丝裂原活化蛋白激酶抑制剂的实验表明，阿瑞蟾毒配基通过抑制c-Jun氨基末端激酶（c-Jun N-terminal kinases, JNK）通路诱导鼻咽癌细胞凋亡。此外，阿瑞蟾毒配基在体内可抑制鼻咽癌肿瘤增殖。本研究明确了阿瑞蟾毒配基在体外与体内的抗肿瘤效应。鉴于其可下调claspin并抑制JNK通路，阿瑞蟾毒配基有望在鼻咽癌的临床治疗中具备应用价值。