RNAi及其在肝癌治疗中的作用

RNA干扰(RNAi)是指生物体内利用双链RNA诱导同源靶基因的mRNA特异性降解,导致转录后基因沉默的现象.近年来,RNA干扰技术在医学研究中的广泛应用取得显著的基因沉默效果,RNAi以其高特异性、高效性等显著优势将成为研究基因功能的新手段.此文综述该技术的发现、定义、机制、特征等及其在肝癌治疗中的应用.     

原发性肝癌基因治疗的研究进展

针对原发性肝癌的传统治疗方法疗效欠佳、预后较差，基因治疗成为新的研究方向和热点。新的基因治疗策略不断涌现，载体系统不断更新，导人方法不断优化，动物模型不断改进，本文就以上方面进行综述。     

Inflammatory microenvironment and expression of chemokines in hepatocellular carcinoma

AIM:To study the inflammatory microenvironment and expression of chemokines in hepatocellular carcinoma(HCC) in nude mice.METHODS:CBRH-7919 HCC cells were injected into the subcutaneous region of nude mice.Beginning two weeks after the challenge,tumor growth was measured every week for six weeks.The stromal microenvironment and inflammatory cell infiltration was assessed by immunohistochemistry in paired tumor and adjacent peritumoral samples,and macrophage phenotype was assessed using double-stain immunohistochemistry incorporating expression of an intracellular enzyme.A chemokine PCR array,comprised of 98 genes,was used to screen differential gene expressions,which were validated by Western blotting.Additionally,expression of identified chemokines was knocked-down by RNA interference,and the effect on tumor growth was assessed.RESULTS:Inflammatory cell infiltrates are a key feature of adjacent peritumoral tissues with increased macrophage,neutrophil,and T cell(specifically helperand activated subsets)infiltration.Macrophages within adjacent peritumoral tissues express inducible nitric oxide synthase,suggestive of a proinflammatory phenotype.Fifty-one genes were identified in tumor tissues during the progression period,including 50that were overexpressed(including CXCL1,CXCL2 and CXCL3)and three that were underexpressed(CXCR1,Ifg and Actb).RNA interference of CXCL1 in the CBRH-7919 cells decreased the growth of tumors in nude mice and inhibited expression of CXCL2,CXCL3and interleukin-1βprotein.CONCLUSION:These findings suggest that CXCL1plays a critical role in tumor growth and may serve as a potential molecular target for use in HCC therapy.     

Molecular targeted therapy for hepatocellular carcinoma:Current and future

Hepatocellular carcinoma(HCC)is one of the most frequent tumors worldwide.The majority of HCC cases occur in patients with chronic liver disease.Despite regular surveillance to detect small HCC in these patients,HCC is often diagnosed at an advanced stage.Because HCC is highly resistant to conventional systemic therapies,the prognosis for advanced HCC patients remains poor.The introduction of sorafenib as the standard systemic therapy has unveiled a new direction for future research regarding HCC treatment.However,given the limited efficacy of the drug,a need exists to look beyond sorafenib.Many molecular targeted agents that inhibit different pathways involved in hepatocarcinogenesis are under various phases of clinical development,and novel targets are being assessed in HCC.This review aims to summarize the efforts to target molecular components of the signaling pathways that are responsible for the development and progression of HCC and to discuss perspectives on the future direction of research.     

Surgical resection of a solitary para-aortic lymph node metastasis from hepatocellular carcinoma

Lymph node (LN) metastases from hepatocellular carcinoma (HCC) are considered uncommon. We describe the surgical resection of a solitary para-aortic LN metastasis from HCC. A 65-year-old Japanese man with B-type liver cirrhosis was admitted for the evaluation of a liver tumor. He had already undergone radiofrequency ablation, transcatheter arterial chemoembolization, and percutaneous ethanol injection therapy for HCC. Despite treatment, viable regions remained in segments 4 and 8. We performed a right paramedian sectionectomy with partial resection of the left paramedian section of the liver. Six months later, serum concentrations of alpha-fetoprotein (189 ng/mL) and PIVKA-2 (507 mAU/mL) increased. Enhanced computed tomography of the abdomen revealed a tumor (20 mm in diameter) on the right side of the abdominal aorta. Fluorine-18 fluorodeoxyglucose positron emission tomography revealed an increased standard uptake value. There was no evidence of recurrence in other regions. Esophagogastroduodenoscopy and colonoscopy revealed no malignant tumor in the gastrointestinal tract. Para-aortic LN metastasis from HCC was thus diagnosed. We performed lymphadenectomy. Histopathological examination revealed that the tumor was largely necrotic, with poorly differentiated HCC on its surface, which confirmed the suspected diagnosis. After 6 mo tumor marker levels were normal, with no evidence of recurrence. Our experience suggests that a solitary para-aortic LN metastasis from HCC can be treated surgically.     

Knockdown of survivin gene expression by RNAi induces apoptosis in human hepatocellular carcinoma cell line SMMC-7721

AIM: To investigate the survivin gene expression in human hepatocellular carcinoma cell line SMMC-7721 and the effects of survivin gene RNA interference (RNAi) on cell apoptosis and biological behaviors of SMMC-7721 cells. METHODS: Eukaryotic expression vector of survivin gene RNAi and recombinant plasmid pSuppressorNeo-survivin (pSuNeo-SW), were constructed by ligating into the vector, pSupperssorNeo (pSuNeo) digested with restriction enzymes Xba I and Sail and the designed double-chain RNAi primers. A cell model of SMMC-7721 after treatment with RNAi was prepared by transfecting SMMC-7721 cells with the lipofectin transfection method. Strept-avidin-biotin-complex (SABC) immunohistochemical staining and RT-PCR were used to detect survivin gene expressions in SMMC-7721 cells. Flow cytometry was used for the cell cycle analysis. Transmission electron microscopy was performed to determine whether RNAi induced cell apoptosis, and the method of measuring the cell growth curve was utilized to study the growth of SMMC-7721 cells before and after treatment with RNAi. RESULTS: The eukaryotic expression vector of survivin gene RNAi and pSuNeo-SW, were constructed successfully. The expression level of survivin gene in SMMC-7721 cells was observed. After the treatment of RNAi, the expression of survivin gene in SMMC-7721 cells was almost absent, apoptosis index was increased by 15.6%, and the number of cells was decreased in G2/M phase and the cell growth was inhibited. CONCLUSION: RNAi can exert a knockdown of survivin gene expression in SMMC-7721 cells, and induce apoptosis and inhibit the growth of carcinoma cells.     

Emerging role of ~(18)F-fluorodeoxyglucose positron emission tomography for guiding management of hepatocellular carcinoma

Hepatocellular carcinoma(HCC) is one of major causes of cancer mortality worldwide. For decades, ~(18)F-fluorodeoxyglucose(FDG) positron emission tomography(PET) has been widely used for staging, predicting prognosis, and detecting cancer recurrence in various types of malignant diseases. Due to low sensitivity of FDG PET for detecting intrahepatic HCC lesions, the clinical value of FDG PET in HCC patients has been limited. However, recent studies with diverse analytic methods have shown that FDG PET has promising role in aiding management of HCC patients. In this review, we will discuss the clinical role of FDG PET for staging, predicting prognosis, and evaluating treatment response in HCC. Further, we will focus on recent clinical studies regarding implication of volumetric FDG PET parameters, the significance of FDG uptake in HCC for selecting treatment and predicting treatment response, and the use of radiomics of FDG PET in HCC.     

MT1E在肝癌形成过程中的表达及其在肝癌细胞中的作用

目的: 探讨MT1E基因在肝癌形成过程中不同阶段的表达及其在肝癌细胞中的生物学功能.方法: DEN诱发大鼠肝癌形成模型,分别观察造模4、8、16、20 wk后肝脏组织病理形态学的改变以及MT1E基因表达的差异;针对MT1E基因mRNA序列设计2个siRNA靶点,分别经质粒重组后电转入SMMC-7721肝癌细胞株中,实时荧光定量PCR检测MT1E基因的表达,MTT法检测细胞生长增殖.结果: 大鼠肝癌造模4、8 wk时肝组织主要表现为炎性病变,而到16 wk后呈现典型的增生病变,20 wk后已全部发展为肝细胞癌. MT1E基因在造模16 wk后表达明显增加,与对照组比较差异显著(芯片读数: 11524 vs 5462). 成功筛选到MT1E基因RNA干扰的一个有效靶序列,电转染72 h后该基因的表达量较空白对照组与阴性对照组明显降低(0.38 vs 1.00,0.93,P<0.01). 与阴性对照组比较,有效干扰靶点电转染144 h后细胞的生长增殖得到了明显抑制(0.1700±0.0313 vs 0.5748±0.0480,P<0.01).结论: 成功应用D E N诱发大鼠肝癌形成,MT1E基因在肝癌形成的后期表达明显升高,其高表达与肿瘤细胞的恶性增殖有关.     

Positron-emission tomography for hepatocellular carcinoma:Current status and future prospects

Hepatocellular carcinoma(HCC) is one of the leading causes of cancer mortality worldwide. Various imaging modalities provide important information about HCC for its clinical management. Since positron-emission tomography(PET) or PET-computed tomography was introduced to the oncologic setting, it has played crucial roles in detecting, distinguishing, accurately staging, and evaluating local, residual, and recurrent HCC. PET imaging visualizes tissue metabolic information that is closely associated with treatment. Dynamic PET imaging and dual-tracer have emerged as complementary techniques that aid in various aspects of HCC diagnosis. The advent of new radiotracers and the development of immuno-PET and PET-magnetic resonance imaging have improved the ability to detect lesions and have made great progress in treatment surveillance. The current PET diagnostic capabilities for HCC and the supplementary techniques are reviewed herein.     

靶向cyclinE小干扰RNA对肝癌HepG2细胞增殖和凋亡的影响

目的:以cyclinE基因编码区为靶位,构建表达小干扰RNA(siRNA)的质粒载体,观察转染后对HepG2细胞的影响.方法:针对cyclinE基因序列构建表达siRNA的真核表达载体pSilencer3.1-H1hygro,利用脂质体Metafectene转染CyclinE基因高表达的肝癌细胞株HepG2.流式细胞仪检测细胞周期及凋亡率,MTT法检测细胞增殖活性,RT-PCR和Western blot法观察转染后细胞cyclinE基因表达.结果:成功构建了表达siRNA的真核质粒载体,转染后cyclinE基因mRNA及蛋白表达水平分别下降了79%和65%.结论:靶向cyclinE基因的siRNA可有效沉默HepG2细胞高表达的cyclinE基因,从而抑制肝癌细胞的增殖并促进凋亡.     

Survivin与肝癌关系的研究进展

Survivin是一种凋亡抑制蛋白,具有调节细胞分化和抑制凋亡的双重功能.因其在人类大多数肿瘤中高表达,故成为近年来肿瘤基因治疗研究热点.此文就Survivin在肝癌中的研究进展作一综述.     

Positron emission tomography/computer tomography： Challenge to conventional imaging modalities in evaluating primary and metastatic liver malignancies

Computer tomography (CT) and magnetic resonance imaging (MRI),as conventional imaging modalities,are the preferred methodology for tumor,nodal and systemic metastasis (TNM) staging. However,all the noninvasive techniques in current use are not sufficiently able to identify primary tumors and even unable to define the extent of metastatic spread. In addition,relying exclusively on macromorphological characteristics to make a conclusion runs the risk of misdiagnosis due mainly to the intrinsic limitations of the imaging modalities themselves. Solely based on the macromorphological characteristics of cancer,one cannot give an appropriate assessment of the biological characteristics of tumors. Currently,positron emission tomography/computer tomography (PET/CT) are more and more widely available and their application with 18F-fluorodeoxyglucose (18F-FDG) in oncology has become one of the standard imaging modalities in diagnosing and staging of tumors,and monitoring the therapeutic efficacy in hepatic malignancies. Recently,investigators have measured glucose utilization in liver tumors using 18F-FDG,PET and PET/CT in order to establish diagnosis of tumors,assess their biologic characteristics and predict therapeutic effects on hepatic malignancies. PET/ CT with 18F-FDG as a radiotracer may further enhance the hepatic malignancy diagnostic algorithm by accurate diagnosis,staging,restaging and evaluating its biological characteristics,which can benefit the patients suffering from hepatic metastases,hepatocellular carcinoma and cholangiocarcinoma.     

Polymorphisms of folate metabolism genes in patients with cirrhosis and hepatocellular carcinoma

AIM To evaluated the association of the risk factors and polymorphisms in MTHFR C677 T, MTHFR A1298 C, MTR A2756 G and MTRR A66 G genes.METHODS Patients with cirrhosis(n = 116), hepatocellular carcinoma(HCC)(n = 71) and controls(n = 356) were included. Polymerase chain reaction followed by enzymatic digestion and allelic discrimination technique real-time PCR techniques were used for analysis. MINITAB-14.0and SNPstats were utilized for statistical analysis. RESULTS Showed that age ≥ 46 years(OR = 10.31; 95%CI: 5.66-18.76; P 0.001) and smoking(OR = 0.47; 95%CI: 0.28-0.78; P = 0.003) were associated with cirrhosis. Age ≥ 46 years(OR = 16.36; 95%CI: 6.68-40.05; P 0.001) and alcohol habit(OR = 2.01; 95%CI: 1.03-3.89; P = 0.039) were associated with HCC. MTHFR A1298 C in codominant model(OR = 3.37; 95%CI: 1.52-7.50; P = 0.014), recessive model(OR = 3.04; 95%CI: 1.43-6.47; P = 0.0051) and additive model(OR = 1.71; 95%CI: 1.16-2.52; P = 0.0072) was associated with HCC, as well as MTR A2756 G in the additive model(OR = 1.68; 95%CI: 1.01-2.77; P = 0.047), and MTRR A66 G in the codominant model(OR = 3.26; 95%CI: 1.54-6.87; P 0.001), dominant model(OR = 2.55; 95%CI: 1.24-5.25; P = 0.007) and overdominant model(OR = 3.05; 95%CI: 1.66-5.62; P 0.001). MTR A2756 G in the additive model(OR = 1.54; 95%CI: 1.02-2.33; P = 0.042) and smokers who presented at least one polymorphic allele for MTRR A66G(OR = 1.71; 95%CI: 0.77-3.82; P = 0.0051) showed increased risk for cirrhosis. There was no association between clinical parameters and polymorphisms. CONCLUSION Age ≥ 46 years, alcohol habit and MTR A2756 G, MTHFR A1298 C and MTRR A66 G polymorphisms are associated with an increased risk of HCC development; age ≥ 46 years, tobacco habit and the MTR A2756 G polymorphism are associated with cirrhosis.     

单克隆抗体介导的肝癌靶向治疗

单克隆抗体（单抗）介导的肝细胞癌（HCC）靶向治疗近来发展迅速。多种单抗如抗HCC、抗血管内皮生长因子（VEGF）、抗表皮生长因子受体（EGFR）和抗肿瘤坏死因子α（TNF-α）等抗体所介导的HCC放射免疫治疗,抗体介导酶前体药物治疗,免疫脂质体治疗,HCC发生相关信号途径干预等,已在HCC治疗研究中取得了明显进展,部分已经进入临床研究。本文就近年来单抗介导的HCC靶向治疗进行综述。     

Progress in systemic therapy of advanced hepatocellular carcinoma

Primary liver cancer, mainly consisting of hepatocellular carcinoma (HCC), is one of common malignancies worldwide, and prevalent among the Chinese population. A diagnosis of early stage HCC has proven to be very difficult because of its insidious feature in onset and development. At the time of diagnosis, most HCC cases are locally advanced and/or distant metastatic, which results in difficulty to be treated and poor prognosis. For advanced HCC, systemic therapy is frequently adopted as an important palliative method. In recent years, clinical studies and observations have often reported about systemic anti-cancer therapy of advanced HCC, including molecular target therapy, systemic chemotherapy and immunotherapy. In this article, we review these treatment modalities to provide a reference for clinicians.     

原发性肝癌伴门脉癌栓综合诊治的研究进展

原发性肝细胞癌（HCC）是较常见的消化道恶性肿瘤之一，其死亡率居我国恶性肿瘤的第二位，而伴有门脉癌栓（PVTT）的肝癌因易发生肝内及远处转移，被认为是预后差的主要指标之一，生存期为2．4～2．7个月。虽然HCC合并门脉癌栓是预后不良的标志，但在肝功能及一般情况允许的情况下，仍可尝试多种治疗方法以提高此类患者的生存率，综合治疗在该病的治疗中起着重要的作用。     

肝细胞癌的分子靶向治疗

分子靶向治疗是近10年来肿瘤治疗领域的重大突破,在肺癌、恶性胃肠间质细胞瘤、恶性淋巴瘤、肠癌、乳腺癌等肿瘤的治疗中已获得很大的成功,尤其HCC的分子靶向治疗在小分子靶向治疗上取得了令人瞩目的新进展,特别是多靶点药物索拉非尼在HCC的靶向治疗中取得的成功,使HCC分子靶向全身治疗有了新的标准。     

肝细胞癌相关分子靶向治疗研究进展

外科手术仍然是治疗肝细胞癌(HCC)最主要的手段,但外科手术的低切除率和高复发率,迫使人们深入研究HCC的发病、转移和侵袭过程中的分子机制,来开发更有效的早期诊断和治疗手段。通过复习相关文献,系统总结了部分HCC相关的主要信号通路及对应的分子靶向治疗研究新进展,包括PI3K/AKT/m TOR信号通路、RAS/RAF/MEK/ERK信号通路、VEGF/VEGFR、PDGFR、FGFR信号通路等,并对HCC分子靶向治疗的研究方向进行了展望。