SYK(S)对肝癌侵袭转移的影响及其机制研究

Our preliminary study indicated that two isoforms of SYK gene, the full length form SYK(L) and short form SYK(S), had opposite biologic functions and prognostic value in human hepatocellular carcinoma (HCC). SYK(L) acts as a tumor suppressor which could inhibited proliferation and xenograft growth, and the presence of SYK(L) expression was associated with better outcome among HCC patients after resection of primary tumor. While SYK(S) plays the opposite role as oncogene whose expression level was associated with malignant proliferation and recurrence of HCC. However, the role of SYK(S) in invasion and metastasis of HCC have been rarely reported in literature. In this project, multiple methods including stable cell lines constructed with retroviral packaging system, gene transfection, RNA interference, Transwell assay and immunofluorescence staining will be used to explore whether SYK(S) has effects on invasion and metastasis in HCC cells and its related molecular mechanism. Moreover, metastatic assay in a nude mice model with orthotopic HCCs, in which SYK(S) is retrovirally overexpressed or knocked-down, will be used to investigate the effects of SYK(S) on liver invasion and lung metastasis in vivo. In addition, with the detection of the expression of SYK(S) and related proteins involved in its signal pathway in at least 120 HCC simples, we will explore whether the expression pattern of single SYK(S) or with other biomarkers could predict the recurrence among patients with HCC after resection. This study would help to elucidate the mechanism of invasion and metastasis, and offer new evidences of targeted therapies for HCC.

我们的前期研究发现：肝癌SYK基因两种异构体的生物学功能与临床预后价值相反，其中全长型SYK(L)具有抑癌基因功能，能够抑制肝癌细胞增殖和裸鼠体内成瘤等，是肝癌患者术后生存较好的预测指标；而缩短型SYK(S)具有潜在癌基因功能，其表达水平与肝癌恶性增殖及复发转移密切相关，但具体作用机制尚不明确。本项目拟采用构建病毒感染的稳定细胞株、基因转染、RNA干扰、Transwell小室、细胞免疫荧光等方法，探讨SYK(S)对肝癌侵袭转移的影响及其相关机制；其次建立SYK(S)过表达或敲低的裸鼠肝脏原位移植瘤模型，观察SYK(S)对肿瘤原位侵袭及远处肺转移的影响；最后检测至少120例肝癌标本中SYK(S)与其信号通路中的相关蛋白的表达，分析SYK(S)单独或联合其信号通路相关蛋白在肝癌术后复发转移中的预测价值。本研究结果将有助于阐明肝癌侵袭转移的机制，以及为肝癌靶向治疗提供新的理论依据。

全长型的脾酪氨酸激酶SYK亦称之为SYK(L)，由于在转录过程中的选择性拼接，翻译产生一种在Interdomain B功能域上较SYK(L)减少23个氨基酸的异构体蛋白SYK(S)；恶性实体肿瘤中SYK(L)和SYK(S)两种异构体的生物学功能存在较大的差异。已有研究表明SYK可作为肝癌的预后预测指标，但SYK两种异构体SYK(L)/(S)在肝癌中的生物学功能未明。本研究主要研究肝癌中SYK两种异构体的生物学功能、及其可能的潜在临床价值。我们的研究结果提示：SYK(L)在约38%的肝癌组织中表达下调，而在大约40%肝癌组织中可检测到其异构体SYK(S)的异常表达，但在无肝硬化的正常肝组织中却未检测到SYK(S)表达。肿瘤中SYK(S)表达水平，与肝癌侵袭转移有关的临床病理因素呈显著相关性，包括多发肿瘤（P = 0.003）和肿瘤血管侵犯（P = 0.001）等。152例肝癌组织检测发现仅SYK(S)表达与EMT相关，进一步的功能学实验提示SYK(S)可促进肝癌生长、抑制肿瘤细胞凋亡，并通过ERK信号通路诱导肿瘤发生EMT；而SYK(L)与其异构体SYK(S)的上述功能相反。与肿瘤中表达SYK(L-/S-)或SYK(L+/S+)的肝癌患者相比较，肿瘤表达SYK(L+/S-)者有更长期的总体生存率和无肿瘤复发率（P < 0.001）。因此，SYK(S)促进肝癌的侵袭能力，而SYK(L)抑制肿瘤侵袭转移。针对肿瘤中SYK(L)下调或其异构体SYK(S)异常表达的肝癌患者，需加强术后辅助治疗，以延缓肿瘤的复发转移。