Department of Blood Transfusion, The Affiliated Hospital of Qingdao University, Qingdao, China.
Department of Clinical Laboratory, The Affiliated Hospital of Qingdao University, Qingdao, China.
J Chemother. 2023 Nov;35(7):653-661. doi: 10.1080/1120009X.2023.2182573. Epub 2023 Feb 27.
Multidrug resistance (MDR) is a major obstacle to the efficacy of hepatocellular carcinoma (HCC) chemotherapy. Previous studies have identified that low FZD3 predicted decreased survival after intraperitoneal versus intravenous-only chemotherapy in ovarian cancer. This study aimed to identify a potential target in HCC chemotherapy. The FZD3 expression variant in HCC cell lines was detected by RT-qPCR and western blotting. The FZD3 expression in the early recurrent HCC group (RE group) and the non-early recurrent HCC group (non-RE group) was measured by RT-qPCR. Then, the 50% inhibitory concentrations (IC50) in HCC cell lines were studied by MTT assay. TOP/FOP FLASH luciferase assay was performed to measure TCF-binding activities. We found that FZD3 was upregulated in three HCC cell lines, and the FZD3 expression was significantly higher in the RE group than in the non-RE group (P = 0.0344). A positive correlation between FZD3 and MDR1 was observed in HCC tissues (R = 0.6368, P = 0.0001). Then, we found that FZD3 knockdown significantly altered Huh-7 cell chemotherapeutic sensitivity to cisplatin [50.43 µM in the FZD3 siRNA (siFZD3) group vs 98.59 µM in the siRNA negative control (siNC) group; P = 0.007] or doxorubicin (7.43 µM in the siFZD3 group vs 14.93 µM in the siNC group; P = 0.017). TOP/FOP FLASH luciferase assay showed FZD3 could inhibit Wnt/β-catenin signaling in HCC cells. Moreover, FZD3 expression knockdown in SNU-449 and Huh-7 cells markedly reduced β-catenin and phosho-β-catenin (S37) protein expression, and Cyclin D1, c-myc and MDR1 were significantly decreased. This is the first study to describe the significantly increased FZD3 expression in patients with early recurrent HCC. FZD3 knockdown led to increased sensitivity to chemotherapy by Wnt/β-catenin signaling inhibition in HCC cell lines. Our study suggests FZD3 as a potential target for reversing chemoresistance in HCC.
多药耐药性 (MDR) 是肝癌 (HCC) 化疗疗效的主要障碍。先前的研究已经确定，在卵巢癌中，FZD3 低表达预示着腹腔内与静脉内化疗相比生存率降低。本研究旨在确定 HCC 化疗中的一个潜在靶点。通过 RT-qPCR 和 Western blot 检测 HCC 细胞系中的 FZD3 表达变异。通过 RT-qPCR 测量早期复发性 HCC 组 (RE 组) 和非早期复发性 HCC 组 (非-RE 组) 中的 FZD3 表达。然后，通过 MTT 测定法研究 HCC 细胞系中的 50%抑制浓度 (IC50)。进行 TOP/FOP FLASH 荧光素酶测定以测量 TCF 结合活性。我们发现，在三种 HCC 细胞系中 FZD3 上调，并且在 RE 组中 FZD3 表达明显高于非-RE 组 (P=0.0344)。在 HCC 组织中观察到 FZD3 与 MDR1 之间存在正相关 (R=0.6368，P=0.0001)。然后，我们发现 FZD3 敲低显着改变了 Huh-7 细胞对顺铂的化疗敏感性[FZD3 siRNA (siFZD3) 组为 50.43μM，siRNA 阴性对照 (siNC) 组为 98.59μM；P=0.007]或阿霉素 (siFZD3 组为 7.43μM，siNC 组为 14.93μM；P=0.017)。TOP/FOP FLASH 荧光素酶测定显示，FZD3 可以抑制 HCC 细胞中的 Wnt/β-catenin 信号通路。此外，在 SNU-449 和 Huh-7 细胞中敲低 FZD3 表达显着降低了 β-catenin 和磷酸化-β-catenin(S37)蛋白表达，Cyclin D1、c-myc 和 MDR1 明显降低。这是第一项描述早期复发性 HCC 患者中 FZD3 表达显著增加的研究。FZD3 敲低通过抑制 HCC 细胞系中的 Wnt/β-catenin 信号通路导致对化疗的敏感性增加。我们的研究表明 FZD3 是逆转 HCC 化疗耐药性的潜在靶点。
