肺腺癌不同驱动基因相关性研究

发布时间：2018-03-20 10:51

本文选题：肺腺癌　切入点：间变性淋巴瘤激酶　出处：《北京协和医学院》2017年博士论文　论文类型：学位论文

【摘要】：恶性肿瘤严重危害人们的身心健康,随着经济的发展和生活水平的提高,肿瘤的发病率呈逐年上升的态势,预计到2030年,全球肿瘤的发病和死亡人数将分别达到2640万和1700万。肺癌是世界范围内发病率及死亡率均居于首位的恶性肿瘤。非小细胞肺癌(NSCLC)约占肺癌的85%,其中以肺腺癌最为常见。肿瘤的异质性问题是目前肿瘤研究的热点,为肿瘤精确诊断及个体化治疗带来挑战。肺腺癌是一组不仅在组织学分类,而且在分子遗传学方面均表现高度异质性的恶性肿瘤。目前肺腺癌中超过一半的肿瘤驱动基因被发现,而针对这些驱动基因的个体化靶向治疗已成为晚期NSCLC标准化治疗。间变性淋巴瘤激酶(ALK)基因是继表皮生长因子受体(EGFR)基因突变之后第二个酪氨酸激酶抑制剂(TKI)作用靶点。小分子抑制剂克唑替尼可特异性靶向抑制ALK蛋白。对驱动基因突变状态检测是临床靶向药物治疗的依据。目前常见的ALK基因融合状态的检测方法有荧光原位杂交法、逆转录聚合酶链式反应法及Ventana ALK D5F3免疫组织化学染色法。在本研究中我们利用Ventana ALK D5F3免疫组织化学染色法检测ALK融合蛋白表达,收集了较大样本量的ALK阳性病例进行临床病理因素分析;其次以往研究认为同一肿瘤中不同驱动基因突变是相互排斥的,但目前已有联合突变NSCLC病例的报道,所以本研究同时对ALK阳性肺腺癌进行其他驱动基因如EGFR基因、KRAS基因、BRAF基因、HER2基因突变检测,以进一步了解肺腺癌此类病例的临床病理特征及意义。肿瘤异质性问题是现在研究的热点,为肿瘤的诊断及个体化治疗带来挑战。在很多恶性实体瘤中发现肿瘤遗传学异质性,例如原发肿瘤与其转移灶间分子表达不一致现象。肺腺癌具有高度遗传学异质性,关于其靶向癌基因的异质性问题的研究主要是在EGFR基因异质性的相关研究报道,而关于ALK是否具有异质性的研究报道很少。本研究旨在通过检测肺腺癌原发灶肿瘤与淋巴结转移灶间ALK基因融合的表达,探讨肺腺癌中ALK表达是否具有差异性,以便为肺腺癌患者的临床诊断和个体化治疗提供一定的参考。第一部分:我们共收集通过Ventana ALK D5F3全自动免疫组织化学染色法检测ALK阳性肺腺癌106例。所有病例术前均未接受任何治疗。首先采用Sanger 一代测序法对106例ALK阳性肺腺癌进行其他相关驱动基因EGFR、KRAS、BRAF、HER2突变检测,结果显示,106例ALK阳性肺腺癌中5例发生EGFR突变,而其他相关驱动基因未见改变。5例EGFR突变病例其中3例是第19号外显子缺失突变,2例是第21号外显子L858R点突变。此外随机选取同时期90例ALK D5F3免疫组织化学染色阴性肺腺癌病例做阴性对照,探讨ALK阳性肺腺癌临床病理因素特征。结果显示ALK阳性肺腺癌更常见于年轻患者,更容易发生淋巴结转移及处于高临床分期。此外,我们发现ALK阳性患者虽然较多见不吸烟者,但是并无统计学意义。女性患者易见在本研究中也未能被体现。组织学亚型上ALK阳性更常见于以微乳头为主型及实体为主型肺腺癌。第二部分:收集37例ALK D5F3阳性肺腺癌及其配对转移淋巴结和16例ALK D5F3阴性肺腺癌病例及其配对转移淋巴结,共53个配对肺腺癌原发肿瘤及其转移淋巴结肿瘤组织进行ALK表达差异性研究。结果显示肺腺癌原发肿瘤与淋巴结转移灶间ALK表达符合率为100%。此外我们对ALK阳性肺腺癌肿瘤组织免疫组织化学染色切片进行镜下观察,发现ALK融合蛋白表达是弥漫性的,存在于每一个肿瘤细胞中,无差异性着色。因此我们认为:ALK基因融合在肺腺癌中表达是一致性的,且在肿瘤进展过程中一旦发生即为稳定的分子改变。
[Abstract]:Malignant tumor which seriously endanger people's physical and mental health, with the development of economy and the improvement of living standards, the incidence of cancer is rising year by year, is expected to 2030, the global incidence of tumor and the death toll will reach 26 million 400 thousand and 17 million. Lung cancer incidence and mortality in the world are in the first place of malignant tumor. Small cell lung cancer (NSCLC) accounts for about 85% of the lung cancer, lung adenocarcinoma is the most common problem. Tumor heterogeneity is currently the focus of cancer research, a challenge for accurate diagnosis and individualized treatment of tumor. Lung cancer is a group of not only the classification in the organization, and are highly heterogeneous malignant the tumor in molecular genetics. At present, more than half of lung adenocarcinoma tumor driven gene was found, and the individual target for these to drive gene therapy has become the standard treatment of advanced NSCLC anaplastic. Lymphoma kinase (ALK) gene is the epidermal growth factor receptor (EGFR) gene mutation after second tyrosine kinase inhibitors (TKI) target. Small molecule inhibitors for specific targeting of G Nikete inhibition of ALK protein. The gene mutation driven state detection is clinical targeted therapy on the basis of the current detection method of ALK gene. The common fusion status of fluorescence in situ hybridization, reverse transcription polymerase chain reaction and Ventana ALK D5F3 immunohistochemical staining. In this study we use Ventana ALK D5F3 immunohistochemical staining were used to detect the expression of ALK fusion protein, ALK collection of positive cases with big sample clinical pathological factors analysis; secondly, the past on the same tumor in different driving gene mutations are mutually exclusive, but there are combined mutations of NSCLC cases were reported, so this study with ALK Other positive lung cancer driving genes such as EGFR gene, KRAS gene, BRAF gene, HER2 gene mutation detection, to further understand the clinical pathological characteristics and significance of lung cancer cases. The problem of tumor heterogeneity is a hot research now, a challenge for tumor diagnosis and individualized treatment of tumor genetic heterogeneity. In many malignant tumors, such as expression of primary tumor and metastatic molecular inconsistency. Lung cancer has a high degree of genetic heterogeneity, research on the heterogeneity of cancer gene to its target is mainly based on EGFR related research reports heterogeneity, and reports about whether ALK has the heterogeneity rarely. This study is to detect lung cancer primary tumor and lymph node metastasis or expression of ALK gene fusion, to investigate whether there is difference of the expression of ALK in lung adenocarcinoma, for lung adenocarcinoma To provide a reference for clinical diagnosis and treatment of individual cancer patients. The first part: We collected by Ventana ALK D5F3 automatic immunohistochemistry method to detect 106 cases of ALK positive lung cancer. All patients had not received any treatment. The first generation Sanger sequencing method for driving other related gene EGFR in 106 cases, ALK positive lung cancer KRAS, BRAF, mutation detection, HER2 results showed that ALK was positive in 106 cases of lung adenocarcinoma in 5 cases of EGFR mutation, and other related genes did not alter the.5 drive with EGFR mutations, 3 cases of the exon nineteenth deletion mutation, 2 cases of exon twenty-first L858R point mutation. In addition the random select the same period 90 cases of ALK negative immunostaining of D5F3 in lung adenocarcinoma cases negative control, positive factors to investigate the clinicopathologic features of lung adenocarcinoma ALK. The results showed that ALK positive lung cancer is more common in young Patients are more prone to lymph node metastasis and clinical stage is high. In addition, we found that ALK positive patients were more common although nonsmokers, but there was no statistically significant. Female patients were more likely to see in this study also failed to be reflected. The histological subtypes of ALK positive are more common in with micropapillary type and solid type adenocarcinoma of the lung. The second part: collected 37 cases of lymph node metastasis and 16 cases of lymph node metastasis of lung adenocarcinoma ALK D5F3 negative cases and matched positive lung adenocarcinoma ALK D5F3 and paired, a total of 53 pairs of lung adenocarcinoma primary tumor and metastatic lymph node tumor tissue ALK expression difference. The results showed adenocarcinoma of the lung cancer primary tumor expression with the rate of 100%. in addition to our ALK positive lung adenocarcinoma immunohistochemical staining sections were observed under microscope and lymph node metastasis or ALK, found that ALK fusion protein expression is diffuse There is no difference coloration in every tumor cell. Therefore, we think that the expression of ALK fusion is consistent in lung adenocarcinoma, and it is stable in the process of tumor progression.

【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2017
【分类号】：R734.2

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