【作者】 沈徐宁；

【导师】 梁廷波；

【作者基本信息】 浙江大学 ， 外科学， 2010， 硕士

【摘要】 背景肝癌的早期诊断相当困难,并且缺乏有效的治疗方法,因而多数患者预后不佳。西妥昔单抗(Cetuximab,一种表皮生长因子受体抑制剂)是一种在其他癌症治疗中非常有效的药物,如直肠癌。但是在肝癌患者的临床试验中却疗效欠佳。信号转导与转录激活子(Signal transducers and activators of transcription,简称Stat)家族是一组重要的调控细胞增殖和生长,侵袭,转移和血管生成的细胞因子,尤其是其中的Stat3。先前的研究已经表明,肝癌组织中Stat3的表达较正常肝脏组织高,可作为肝癌化疗药物的敏感性决定因素。而关于Cetuximab在肝癌患者中耐药机理的研究至今未见报道。材料和方法本实验选用Hep G2和SK-HEP 1两种肝癌细胞株,通过MTT法比较二者对于Cetuximab的耐受程度。同时采用NSC 74859作为Stat3通路的特异性阻断剂,研究阻断或不阻断Stat3通路下两种细胞株对于Cetuximab的耐药情况。采用SiRNA转染使细胞株丧失或获得Stat3的表达,通过RT-PCR和Western blot检测EGFR, Stat3和p-Stat3在不同用药组的表达水平。结果我们在研究中发现,Hep G2细胞株比SK-HEP 1细胞株对单独使用Cetuximab更敏感。p-Stat3蛋白在SK-HEP 1中的表达明显高于在Hep G2中的表达。用NSC74859或者Stat3的SiRNA阻断p-Stat3的表达可明显提高Cetuximab对两种细胞株的生长抑制作用,尤其是SK-HEP 1.而转染了Stat3基因的Hep G2细胞株较对照组则丧失了对Cetuximab敏感性。结论我们的研究首次发现,通过NSC 74859抑制p-Stat3的表达可以增强肝癌细胞株对于Cetuximab的敏感性,这表明将NSC 74859与Cetuximab联合应用将可能是未来一种有效的肝癌治疗方法。更多还原

【Abstract】 BackgroundHCC is a troublesome tumor with a poor prognosis due to late diagnoses and a lack of effective treatment options. Cetuximab(an Epidermal growth factor receptor inhibitor) is an effective drug in other cancers such as rectal cancer which shown only modest efficacy in clinical trials of HCC. Signal transducers and activators of transcription (STAT) family of transcriptional factors were originally identified as the critical mediators of cytokine responses and subsequently implicated in diverse external stimuli-initiated cellular signaling. Stat3 is a member of STAT family which has been considered as an oncogene based on its ability to transactivate a number of genes whose products are involved in cell proliferation and survival, invasion, metastasis and angiogenesis. Previous studies have shown that the level of Stat3 in hepatic tumor tissue was higher than in normal tissue and expression of Stat3 may be as a determinant of sensitivity of HCC to antitumor drugs. But the relationship of Stat3 and the mechanism of resistance to Cetuximab in liver cancer patients has not been reported.Materials and MethodsWe compared the tolerance for Cetuximab between two HCC cell lines, Hep G2 and SK-HEP 1 by MTT. We used the Stat3 pathway specific blocker NSC 74859 to block the Stat3 pathway. And we compared the percentage of resistance to Cetuximab of the two cell lines with or without NSC 74859. Hep G2 and SK-HEP 1 cells were transfected with Nagative SiRNA, Stat3 SiRNA and wild type stat3. EGFR, Stat3, and p-Stat3 expression in different levels of treatment group were estimated by RT-PCR and Western blot.ResultIn our study, we found that Hep G2 was more sensitive to cetuximab than Sk-Hep-1. We also found that the expression of p-stat3 in Sk-Hep-1 cell line was higher than that in Hep G2 cell line. Blocking p-stat3 using NSC 74859 or an small interfere RNA(stat3 SiRNA) boost the cetuximab’s effect in both cell lines especially in Sk-Hep-1 while transfection stat3 cDNA plasmid into Hep G2 lost the sensitivity to cetuximab compared to control Hep G2.ConclusionOur study first found that p-stat3 inhibition by NSC74859 mediate sensitivity to cetuximab, suggesting that the combined NSC74859 with cetuximab would be an effective therapy for HCC.更多还原