浸润型肝细胞癌临床诊疗策略及分子病理特征的研究

发布时间：2018-04-30 15:15

本文选题：巴塞罗那分期 + 香港分期　；参考：《南京大学》2017年博士论文

【摘要】：肝细胞癌(hepatocellular carcinoma,HCC)是全球第五大恶性肿瘤,目前其发病率和死亡率仍高居不下。长期以来,形态各异的肝细胞癌与其生物学行为和预后的关系一直是学者们关注的热点。目前认为,HCC的大体形态与其分子病理学特征有着密切的联系。2011年,美国Johns Hopkins大学医学院的研究团队首先提出了呈浸润性生长的HCC亚型(称为浸润型肝细胞癌,infiltrative hepatocellular carcinoma,iHCC),其临床特征主要包括:①多数合并乙型肝炎病毒(hepatitis B virus,HBV)感染;②具有与传统HCC不同的影像学特点,病灶边界模糊不清,动脉期强化不明显,极易发生脉管侵犯等;③大多数有血清甲胎蛋白(alpha-fetoprotein,AFP)显著升高。iHCC的概念一经提出,立刻引起国际上临床学者的重视。目前普遍认为,iHCC呈浸润性生长方式,肿瘤生长迅速、浸润范围广,很快发生脉管癌栓、肝内播散及全身转移,是HCC中恶性程度最高的临床亚型。但是,由于iHCC的高侵袭转移特征,绝大多数病例被发现和确诊时已到了晚期,失去了根治性治疗的机会。因此,完整系统的临床-影像-病理资料收集非常困难,限制了相关研究的深入。本研究团队自2013年开始高度关注iHCC的相关临床和基础研究,经过不断积累获得以下研究结果:①由于HBV感染是HCC主要发病因素,我国iHCC的发病率高于其他亚洲和欧美国家,我们统计的结果占HCC的30%,显著高于欧美和日本的5～10%;②在不同大体分型的HCC中,iHCC的预后最差;③经过大量临床病例的观察,我们在国内外首先报道了在早-中期HCC中亦存在iHCC亚型。因此,本课题着重于iHCC的临床和基础研究。主要包括以下三个方面:①选择适合浸润型肝细胞癌诊疗决策和预后分析的临床分期;②通过分析手术切除iHCC病人的临床特征及预后等信息,探索合适的治疗策略;③通过全外显子测序技术,探索iHCC的关键特异性基因,探讨其分子病理学特征。第一部分选择适合浸润型肝细胞癌诊疗决策和预后分析的临床分期目的:比较香港分期(Hong Kong Liver Cancer,HKLC)和巴塞罗那分期(Barcelona Clinic Liver Cancer,BCLC)在HBV相关性HCC病人中的预测预后能力。方法:668例HCC病人纳入研究,应用赤池信息量准则(Akaike information criterion,AIC)、一致性指数(concordance-index,c-index)和 ROC 曲线下面积(area under the receiver operating characteristic curve,AUC)比较两种分期的预后预测能力。通过单因素及多因素分析确定与生存相关的独立危险因素。结果:与生存相关的独立危险因素包括Child-Pugh评分、乳酸脱氢酶(lactate dehydrogenase,LDH)、白蛋白(albumin,ALB)、肿瘤部位、肿瘤数目、肿瘤大小和血管侵犯。HKLC分期的1、3、5年AUC分别为0.740,0.695和0.615,BCLC分期的1、3、5年AUC分别为0.622,0.569和0.548,提示HKLC分期的区分能力优于BCLC分期。HKLC分期对生存的预测效能上亦优于BCLC分期(HKLC 分期:AIC = 4709.480,c-index=0.805;BCLC 分期:AIC = 4852.708,c-index=0.717)。结论:HBV相关性HCC病人中,HKLC分期相比于BCLC分期来说预测预后能力更强,可能更适合以HBV感染为主要病因的中国HCC人群的预后分析以及治疗决策。第二部分手术切除浸润型肝细胞癌的临床特征分析目的:探讨接受手术切除的浸润型肝细胞癌(infiltrative hepatocellular carcinoma,iHCC)病人的临床特征及预后情况。方法:回顾性分析2003年1月至2012年12月在南京大学医学院附属鼓楼医院肝胆胰外科行肝切除术的47名iHCC病人。通过单因素及多因素分析确定与生存及复发相关的独立危险因素。运用卡方检验、t检验或Mann-Whitney 检验分析微血管侵犯(microvascular invasion,MVI)与相关临床指标之间的关系。应用Kaplan-Meier曲线进行生存分析。结果:该组病人的中位生存时间为27.37个月,1年无复发生存率为61.7%。AFP并不是iHCC病人的特征性指标。解剖性肝切除与较高的无复发生存期显著相关(P=0.007),存在MVI的病人无复发生存期较低(P0.001)。血清LDH水平较高的病人总生存期(P=0.003)及无复发生存期(P=0.020)均较低。病人的MVI与血清谷草转氨酶(aspartate aminotransferase,AST)、谷氨酰转肽酶(gamma glutamyl transpeptidase,GGT)以及LDH水平有关。亚组分析提示在高水平LDH的病人中,低级别MVI的病人有较高的无复发生存期(P=0.019)。结论:iHCC病人具有较高的MVI发生率,对于早中期的病人来说,解剖性肝切除仍能使病人获益。MVI分级可以用来区分出预后更差的一部分病人,尤其是具有较高血清LDH水平的病人。第三部分浸润型肝细胞癌分子病理学特点的探索目的:通过与单发结节型(single nodular,SN)HCC进行比较,探索浸润型肝细胞癌(infiltrative hepatocellular carcinoma,iHCC)的特异性基因表达谱。方法:对6例肿瘤/相对正常的HCC组织样本(3例iHCC、3例SN HCC)进行全外显子测序。随后在30例HCC组织样本(15例iHCC、15例SN HCC)中采用Sanger测序及RT-PCR技术进行结果验证。结果:在对相应组织样本进行全外显子测序、Sanger测序及生物信息学分析后,结果提示iHCC与SN HCC展现出显著不同的基因谱。尤其在iHCC中发现了一个较为典型的生长因子受体酪氨酸激酶突变基因FGFR3,全外显子测序发现了FGFR3的一个非同义突变位点c.G285T(p.Q95H),后续Sanger测序发现了另外五个突变位点(c.G938A:p.G313D,c.G1291A:p.A431T,c.C1355G:p.T452R,c.C1377T:p.L459L and c.A1445T:p.E482V)。免疫组化实验证实了 FGFR3 在 iHCC中存在较高的蛋白表达。结论:FGFR3可能是具有早期复发特点的iHCC的潜在候选癌基因及治疗分子靶标。
[Abstract]:Hepatocellular carcinoma (HCC) is the fifth largest malignant tumor in the world, and its morbidity and mortality still remain high. For a long time, the relationship between the different forms of hepatocellular carcinoma and its biological behavior and prognosis has been the focus of attention of scholars. At present, it is believed that the general morphology of HCC is closely related to its molecular pathological characteristics. In.2011, the research team of the medical school of Johns Hopkins University in the United States first proposed an infiltrative HCC subtype (called invasive hepatocellular carcinoma, infiltrative hepatocellular carcinoma, iHCC). Its clinical features mainly include: (1) most of the combination of hepatitis B virus (hepatitis B virus, HBV) infection; and (2) with traditional Chinese Medicine C has different imaging features, the boundary of the focus is unclear, the arterial phase is not obvious, and the vascular invasion is very easy to occur. (3) the concept of alpha-fetoprotein (AFP), which has a significant increase of.IHCC, is put forward immediately. Rapid, extensive infiltration, rapid vascular tumor thrombus, intrahepatic dissemination and whole body metastasis are the most malignant clinical subtypes in HCC. However, due to the high invasion and metastasis of iHCC, most cases have been found and confirmed at the late stage and lost the opportunity for radical treatment. The research team has been paying high attention to the related clinical and basic research of iHCC since 2013. The research team has accumulated the following results after continuous accumulation: (1) because HBV infection is the main factor of HCC, the incidence of iHCC in China is higher than that of other Asian and European countries, and our statistical results account for HC. The 30% of C was significantly higher than 5 to 10% in Europe and the United States and Japan; (2) the prognosis of iHCC was the worst in the HCC of different general classification. After a large number of clinical cases, we first reported the existence of iHCC subtype in the early and mid-term HCC. Therefore, this topic focuses on the clinical and basic research of iHCC, including the following aspects: (1) To select the clinical stages for the diagnosis and treatment of invasive hepatocellular carcinoma (HHC), and to explore the clinical features and prognosis of iHCC patients, explore the appropriate therapeutic strategies, and explore the key specific genes of iHCC through exon sequencing technology and explore its molecular pathological features. Clinical stages for the diagnosis and treatment of invasive hepatocellular carcinoma (Hong Kong Liver Cancer, HKLC) and Barcelona staging (Barcelona Clinic Liver Cancer, BCLC) in HBV associated HCC patients. Methods: 668 patients were included in the study. Ation criterion, AIC), the conformance index (concordance-index, c-index) and the area under the ROC curve (area under the receiver operating characteristic) compared the predictive ability of the two stages. Independent risk factors associated with survival were determined by single factor and multifactor analysis. Results: independent risk associated with survival Factors including Child-Pugh score, lactate dehydrogenase (LDH), albumin (albumin, ALB), tumor site, tumor number, tumor size, and vascular invasion.HKLC stages were 0.740,0.695 and 0.615, respectively, and 1,3,5 years of BCLC staging were different and 0.548. .HKLC staging was also superior to BCLC staging for survival (HKLC staging: AIC = 4709.480, c-index=0.805; BCLC staging: AIC = 4852.708, c-index=0.717). Conclusion: in HBV associated HCC patients, HKLC staging is more likely to predict prognosis than BCLC stages, and may be more suitable for the Chinese population with the main cause of infection. Prognostic analysis and treatment decisions. Clinical features of second parts of surgical excision of infiltrating hepatocellular carcinoma (infiltrative hepatocellular carcinoma, iHCC): the clinical features and prognosis of patients undergoing surgical excision (carcinoma, iHCC). Methods: retrospective analysis from January 2003 to December 2012 at the Nanjing University medicine 47 iHCC patients who underwent hepatectomy of hepatobiliary and pancreatic surgery affiliated to the College Affiliated Drum hospital. Independent risk factors associated with survival and recurrence were determined by single factor and multifactor analysis. The relationship between microvascular invasion (microvascular invasion, MVI) and related clinical indicators was analyzed by chi square test, t test or Mann-Whitney test. The application of K Survival analysis of the aplan-Meier curve. Results: the median survival time of the patients was 27.37 months, and the 1 year non recurrent survival rate was 61.7%.AFP, not the characteristic index of the iHCC patients. The anatomical hepatectomy was significantly correlated with the higher recurrence free survival (P=0.007), and the non recurrent survival time in the patients with MVI was lower (P0.001). The serum LDH level was low (P0.001). Higher patient total survival (P=0.003) and non recurrent survival (P=0.020) were lower. The patient's MVI was associated with serum aspartate aminotransferase (AST), glutamyl transpeptidase (gamma glutamyl transpeptidase, GGT), and LDH levels. Subgroup analysis suggested that patients with low level LDH were higher in the high level LDH patients. No recurrence of survival (P=0.019). Conclusion: iHCC patients have a high incidence of MVI. For patients with early and middle stages, anatomical hepatectomy can still benefit the patients with.MVI classification, which can be used to distinguish some patients with worse prognosis, especially in patients with higher serum LDH levels. Third part of the molecular pathology of infiltrating hepatocellular carcinoma. Objective: To explore the specific gene expression profiles of invasive hepatocellular carcinoma (infiltrative hepatocellular carcinoma, iHCC) by comparing with single nodular (SN) HCC. Methods: 6 cases of tumor / relatively normal HCC tissue samples (3 iHCC, 3 SN HCC) were sequenced in 30 cases. The fabric samples (15 iHCC, 15 SN HCC) were verified by Sanger sequencing and RT-PCR technology. Results: after the whole exon sequencing, Sanger sequencing and bioinformatics analysis of the corresponding tissue samples, the results showed that the iHCC and SN HCC showed significant different gene spectrum. In particular, a more typical growth factor was found in iHCC. Subreceptor tyrosine kinase mutation gene FGFR3, exon sequencing found a non synonymous mutation site of FGFR3 c.G285T (p.Q95H), followed by Sanger sequencing found five other mutation sites (c.G938A:p.G313D, c.G1291A:p.A431T, c.C1355G:p.T452R, c.C1377T:p.L459L and c.A1445T:p.E482V). Immunohistochemistry test confirmed FGFR3 in Conclusion: FGFR3 may be a potential candidate gene and therapeutic molecular target for iHCC with early recurrence characteristics. Conclusion: iHCC is a potential candidate gene for early relapse.

【学位授予单位】：南京大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R735.7

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