SCY1-Like 3在肝癌中的功能特征和临床相关性

Liver cancer (HCC) is the fifth most common malignancy worldwide and the second leading cause of cancer fatalities in Southeast Asia and Hong Kong. Patients with this cancer type have a poor prognosis because the cancer is often detected at an advanced stage. The advanced stage of the disease is largely due to the presence of frequent metastases. To improve the prognosis of patients with HCC, an understanding of the molecular mechanism of metastasis in HCC is urgently needed. Despite recent advances in biomedical technologies, the molecular mechanism that underlies cancer metastasis is unclear. In our pilot study, we employed a cDNA microarray to compare the genetic profiles of our newly established pair of corresponding primary and metastatic HCC cell lines (namely, PLC/PRF/5-PT and PLC/PRF/5-LM, respectively), which were both derived from the same parental cell line (PLC/PRF/5); but differed in terms of their metastatic potential. This analysis showed that SCY1-Like 3 (S. Cerevisiae) (SCYL3) was elevated in the metastatic cell line (PLC/PRF/5-LM) compared with its primary counterpart (PLC/PRF/5-PT). Further analysis using an anti-SCYL3 antibody confirmed an elevated level of SCYL3 in PLC-LM cells by Western blot analysis. SCYL3 is a pseudokinase and is also designated as PACE-1 (Protein-associating with the carboxyl-terminal domain of Ezrin), as it physically interacts with the C-terminal domain of Ezrin. Thus far, only one report shows the possible functional role of SCYL3, and therefore, the clinical relevance and functional role of SCYL3 in cancer remain uncharacterized. After an analysis of the Cancer Genome Atlas (TCGA) data, we demonstrated that SCYL3 is overexpressed in liver, colon and breast cancers. The overexpression of SCYL3 in HCC was further confirmed at both the mRNA and protein levels based on our in-house RNA sequencing data, qPCR and protein analyses. The observation that SCYL3 was increased in the more metastatic HCC cell line together with the observed increase in SCYL3 in HCC clinical samples suggested that the overexpression of SCYL3 may be associated with HCC tumor growth and metastasis. Our pilot study demonstrated for the first time that alterations in SCYL3 affected several critical phenotypes of HCC including proliferation, metastasis and chemoresistance. Further preliminary data showed that SCYL3 physically interacts with and phosphorylates Ezrin at Threonine 567 through its association with another kinase, ROCK2. Because the phosphorylation of Ezrin is critical in the functional regulation of Ezrin, it has been hypothesized that the effect of the SCYL3/ROCK2 complex in the regulation of the pathogenesis of HCC occurs through the modulation of Ezrin activity. In this proposal, we aim to evaluate the clinical significance of SCYL3 in HCC, to delineate the biological characteristics of SCYL3 in HCC, and to characterize the mechanism that underlies its function. The new knowledge gained in this study will provide insight into the role of SCYL3 in the progression of HCC, highlight a novel target for molecular therapy against HCC metastasis, and advance our understanding of HCC metastasis.

肝癌是威胁中国人群的主要恶性肿瘤，患者预后较差。晚期肝癌大部分伴有远处转移。为改善肝癌患者预后，迫切需要了解肝癌转移的分子机制。我们新建立了一对原发和转移肝癌细胞系。分析发现SCY1-Like3(SCYL3)在转移肝癌细胞系中高于原发肝癌细胞系。SCYL3在癌症中的临床相关性与功能仍然未知。qPCR和蛋白分析证实SCYL3在肝癌中的mRNA和蛋白水平过表达。我们初步研究首次证实SCYL3影响肝癌的增殖、转移和化疗耐药。进一步的初步数据显示SCYL3与Ezrin具有物理相关作用，并在另一个激酶—ROCK2—的相互作用下对Ezrin的苏氨酸567位点磷酸化。我们推测SCYL3/ROCK2复合物通过调节Ezrin活性参与肝癌的发病机制。本研究将洞察SCYL3在肝癌发生发展中的作用，寻找转移性肝脏治疗新靶点，深入理解肝癌转移的分子机制。

背景与目的：SCY1 样假激酶 3 (SCYL3) 被确定为 ezrin 的结合伙伴，表明它与转移有关。然而，SCYL3 在癌症中的临床相关性和功能作用仍未得到表征。在这项研究中，我们旨在阐明 SCYL3 在肝细胞癌 (HCC) 进展中的作用。..研究手段：在公开数据集中和内部 HCC 队列的 qPCR 分析中评估了 SCYL3 在 HCC 中的临床意义。在 SCYL3 敲低/过表达 HCC 细胞中检查了 SCYL3 的功能意义和机制后果。使用睡美人转座子系统在 Tp53 KO/c-Myc OE 小鼠中评估体内肿瘤进展。通过共免疫沉淀、蛋白质印迹分析和免疫荧光染色研究了潜在的下游通路。..结果：SCYL3在HCC中经常过表达；它优先在转移性人类 HCC 肿瘤中表达，并且与较差的患者存活率相关。在 HCC 细胞中抑制 SCYL3 可减弱细胞增殖和迁移以及体内转移。有趣的是，内源性 SCYL3 过表达增加了 Tp53 KO/c-Myc OE 小鼠的肿瘤发展和转移。机制研究表明，SCYL3 通过其 C 末端结构域物理结合并调节 ROCK2（Rho 激酶 2）的稳定性和反式激活活性，从而导致肌动蛋白应力纤维和粘着斑的形成增加。..结论：这些发现表明 SCYL3 在促进 HCC 的进展中起着关键作用，并且对开发新的治疗策略以应对转移性 HCC 具有重要意义。..影响和意义：SCYL3 最初被报道为转移相关基因 ezrin 的结合伙伴。迄今为止，SCYL3 在癌症中的临床相关性和功能作用仍未得到表征。在此，我们揭示了它在肝癌进展中的关键作用。我们表明它物理结合并调节 ROCK2 的稳定性和反式激活活性，从而导致 HCC 肿瘤进展。我们的数据提供了 SCYL3 介导的 ROCK2 蛋白稳定性在 HCC 细胞生长和转移中起关键作用的机制见解。靶向SCYL3/ROCK2信号级联可能是治疗HCC患者的一种新的治疗策略。