CSAG1基因在乳腺癌及分子分型中的差异表达研究及临床意义
发布时间：2018-07-31 09:11
【摘要】：乳腺癌来源于乳腺上皮组织,为女性常见的一种恶性肿瘤,近年来我国乳腺癌发病率逐年上升并朝低龄化趋势发展,已成为当今社会的重大公共卫生问题。虽然随着医学的发展,早期诊断和治疗降低了乳腺癌患者的死亡率,但就目前而言,乳腺癌仍是恶性肿瘤中死亡率较高的疾病。乳腺癌属于高度异质性疾病,即使在病理类型、组织学分级、临床分期及治疗均相同的情况下,患者的生存预后也存在极大差别。Perou和Sorlie等人将乳腺癌根据分子分型,分为Basal-like、Luminal A、Luminal B、HER2过表达、Normal等五型,不同分型的乳腺癌患者其治疗及预后差异明显。因此,详细的分子分型能有效的反映乳腺癌的生物学特征,并指导临床选择治疗方案及判断预后。软骨肉瘤相关基因1(Chondrosarcoma associated gene 1,CSAG1),定位于人染色体Xq28,研究表明CSAG1在正常睾丸组织中表达,同时也有报道显示CSAG1是在黑色素瘤细胞株、白血病K562细胞以及恶性神经鞘瘤组织中高表达的基因,研究表明CSAG1与肿瘤发生、发展有着密切的联系,但有关乳腺癌中CSAG1基因的表达情况和预后价值的相关研究还未见报道。目的:探索CSAG1在不同乳腺癌分子分型中的表达及作用,为临床乳腺癌治疗和预后判断提供必要的理论依据。方法:1.通过免疫组织化学方法对100例乳腺癌福尔马林固定石蜡包埋(Formalin Fixed and Paraffin Embedded,FFPE)组织样本结合ER、PR、HER2、Ki-67的表达情况进行分子分型。2.采用RT-PCR方法检测CSAG1基因在不同分子分型的乳腺癌临床患者FFPE组织样本及乳腺癌细胞系中mRNA表达水平。3.通过免疫组织化学方法检测CSAG1蛋白在不同分子分型的乳腺癌临床患者FFPE组织样本中表达水平。4.利用卡方检验及Spearman等级相关分析CSAG1与临床病理参数及乳腺癌经典标志物之间的相对表达水平及相关性。5.运用GraphPad Prism 5统计软件及cbioportal生物信息学分析软件进行生存分析。结果:1.根据2011年st.gallen共识,并结合术后病理免疫组织化学染色er、pr、her2、ki-67表达情况将100例乳腺癌进行分子分型,分别筛选出luminala亚型18例,luminalb亚型29例,her2过表达亚型4例,basal-like亚型35例,normal(non-basal-like)亚型14例。2.乳腺癌分子分型与临床病理参数的关系分析结果显示:在luminala亚型乳腺癌中组织学分级i-ii级所占比例最高,组间具有统计学差异(p0.0001)。3.在临床乳腺癌患者ffpe组织样本中rt-pcr结果显示:csag1基因在乳腺癌分子分型luminala亚型中的表达量高于其他乳腺癌分子亚型,组间差异具有统计学意义(p0.05)。4.在不同乳腺癌细胞系中rt-pcr结果显示:csag1基因在t47d(luminala型乳腺癌,er和/或pr+,her2-,ki-6714%)中较其他乳腺癌细胞系(zr75.1、hdq-p1、cal-120、cal-51、mda-mb-436、mda-mb-231)及正常乳腺上皮细胞系(hbl-100)显著高表达,差异具有统计学意义(p0.0001)。5.在临床乳腺癌患者ffpe组织样本中免疫组化结果显示:csag1蛋白表达阳性率在乳腺癌分子分型luminala亚型中高于其他乳腺癌分子亚型,组间差异具有统计学意义(p0.05)。6.csag1蛋白表达与临床病理参数之间的关系分析结果显示:csag1蛋白表达在组织学分级的i-ii级中较iii级中相对表达水平高,差异具有统计学意义(p0.05);在临床病理组织学类型、tnm分期、年龄、肿瘤直径、淋巴结转移分组中相对表达水平均无明显差别。7.csag1与临床病理参数相关性分析利用spearman等级相关结果显示:csag1蛋白表达与临床病理组织学分级呈负相关,差异具有统计学意义(p0.05);而与年龄、肿瘤直径、淋巴结转移、临床病理组织学类型及tnm分期均无相关性。8.csag1蛋白表达与乳腺癌经典标志物之间的关系分析结果显示:csag1蛋白表达在er及ki-67阳性组中较er及ki-67阴性组中相对表达水平稍高,在her2阴性组中较her2阳性组中相对表达水平高,而在pr阳性与阴性组中表达水平无明显差别。9.csag1与乳腺癌经典标志物相关性分析利用spearman等级相关结果显示:csag1蛋白表达与er、pr表达呈正相关(r=0.237、0.173),与ki-67增殖指数呈负相关(r=-0.251),差异均具有统计学意义(p0.05);而与her2蛋白表达无相关性。10.与临床乳腺癌患者生存相关性分析结果显示:csag1蛋白表达阳性的乳腺癌患者较CSAG1蛋白表达阴性的乳腺癌患者后期的生存稍好,但差异无统计学意义。结论:1.CSAG1在乳腺癌分子分型Luminal A亚型中高表达,提示CSAG1高表达在乳腺癌中筛选出Luminal A亚型有一定作用。2.高表达CSAG1的乳腺癌患者其临床病理组织学分级低、生存期稍高,预后相对较好。3.CSAG1表达与ER、PR表达呈正相关,可作为辅助临床乳腺癌患者行内分泌治疗及预后判断的指标之一。4.CSAG1表达与Ki-67增殖指数呈负相关,可辅助Ki-67进行分子分型并指导临床进行预后分析。
[Abstract]:Breast cancer is a common malignant tumor of women. In recent years, the incidence of breast cancer in China has risen year by year and has been developing towards a low age. It has become a major public health problem in today's society. Although with the development of medicine, early diagnosis and treatment have reduced the mortality of breast cancer patients. Breast cancer is still a high mortality disease in malignant tumors. Breast cancer is a highly heterogeneous disease. Even in pathological type, histological grade, clinical stage and treatment, the survival prognosis of the patients is greatly different,.Perou and Sorlie and others classify breast cancer according to the molecular classification, divided into Basal-like, Luminal A, L Uminal B, HER2 overexpression, Normal and other types of type five, different types of breast cancer patients have significant differences in their treatment and prognosis. Therefore, detailed molecular typing can effectively reflect the biological characteristics of breast cancer and guide clinical selection and prognosis. Chondrosarcoma related groups are located in 1 (Chondrosarcoma associated gene 1, CSAG1). Human chromosome Xq28, studies show that CSAG1 is expressed in normal testicular tissue, and there are also reports that CSAG1 is a high expression gene in melanoma cell lines, leukemia K562 cells and malignant neurilemmoma. Research shows that CSAG1 has a close relationship with the development of tumor, but the expression of CSAG1 gene in breast cancer The study of the status and prognostic value has not been reported. Objective: To explore the expression and role of CSAG1 in the molecular classification of different breast cancer, and to provide the necessary theoretical basis for the treatment and prognosis of clinical breast cancer. Methods: 1. Formalin Fixed and Paraff (Formalin Fixed and Paraff) was fixed by immunohistochemistry. In Embedded, FFPE) tissue samples combined with the expression of ER, PR, HER2, Ki-67 to carry out molecular typing.2. using RT-PCR method to detect CSAG1 gene in the FFPE tissue samples of breast cancer patients with different molecular typing and the level of mRNA expression in breast cancer cell lines. Expression level of.4. in FFPE tissue samples of breast cancer patients using chi square test and Spearman level correlation analysis the relative expression level between CSAG1 and clinicopathological parameters and classic markers of breast cancer and correlation.5. using GraphPad Prism 5 statistical software and cbioportal bioinformatics analysis software for survival analysis. Results: 1. According to the 2011 St.Gallen consensus, combined with postoperative pathological immunohistochemical staining of Er, PR, HER2 and Ki-67, 100 cases of breast cancer were classified, 18 cases of luminala subtype, 29 cases of luminalb subtype, 4 cases of HER2 overexpression subtype, 35 cases of basal-like subtype, 14 case of normal (non-basal-like) subtype of.2. breast cancer molecular classification, respectively. The analysis of the relationship with the clinicopathological parameters showed that the proportion of I-II levels in the luminala subtype of breast cancer was the highest, and there was a statistically significant difference between the groups (P0.0001).3. in the FFPE tissue samples of the patients with clinical breast cancer, the RT-PCR results showed that the expression of the CSAG1 gene in the subtype luminala subtype of breast cancer was higher than that of the other milk. The differences in the subtypes of adenocarcinoma molecules were statistically significant (P0.05).4. in different breast cancer cell lines, RT-PCR results showed that the CSAG1 gene was in T47D (luminala type breast cancer, ER and / or pr+, her2-, ki-6714%) and the other breast cancer cell lines (zr75.1, hdq-p1, pr+, etc.) and normal mammary epithelial cell lines Significant high expression, the difference was statistically significant (P0.0001).5. in the FFPE tissue samples of patients with clinical breast cancer, the immunohistochemical results showed that the positive rate of CSAG1 protein expression in the luminala subtype of breast cancer was higher than that of other breast cancer subtypes, and the difference between the groups was statistically significant (P0.05) the expression of.6.csag1 protein and clinical disease. The analysis of the relationship between the parameters showed that the expression of CSAG1 protein was higher in the I-II level of the histological grade than in the III level, and the difference was statistically significant (P0.05). There was no significant difference in the relative expression level between the clinical histopathology type, the TNM stage, the age, the tumor diameter and the lymph node metastasis group, and the relative expression level was.7.csag1 and clinical. The correlation analysis of pathological parameters showed that the expression of CSAG1 protein was negatively correlated with the clinical histopathological classification, and the difference was statistically significant (P0.05), but there was no correlation with age, tumor diameter, lymph node metastasis, clinical histopathology type and TNM stage, and the expression of.8.csag1 protein and the classic breast cancer markers were not related. The analysis of the relationship between objects showed that the expression of CSAG1 protein in the positive group of ER and Ki-67 was slightly higher than that in the negative group of ER and Ki-67 negative group, and the relative expression level in the negative group of HER2 negative group was higher than that in the HER2 positive group, but there was no significant difference between the expression level of the positive and negative groups of PR and the correlation analysis of the classic markers of breast cancer in the positive and negative groups of pr. The results of Spearman correlation showed that the expression of CSAG1 protein was positively correlated with Er, PR expression (r=0.237,0.173), and negatively correlated with Ki-67 proliferation index (r=-0.251), and the difference was statistically significant (P0.05), while the correlation analysis between.10. and clinical breast cancer patients showed no correlation with HER2 protein expression: the positive expression of CSAG1 protein was positive. The survival of breast cancer patients is better than that of CSAG1 protein negative breast cancer patients, but the difference is not statistically significant. Conclusion: 1.CSAG1 is highly expressed in the Luminal A subtype of breast cancer, suggesting that the high expression of CSAG1 in breast cancer can be selected as the Luminal A subtype, and the clinical disease of the breast cancer patients with the high expression of.2. and CSAG1 is certain. The histological grade is low, the survival time is slightly higher, the prognosis is relatively good, the expression of.3.CSAG1 is positively correlated with the expression of ER and PR. It can be used as one of the indicators of endocrine therapy and prognosis judgment in adjuvant clinical breast cancer patients,.4.CSAG1 expression is negatively correlated with the Ki-67 proliferation index, which can assist Ki-67 to enter the molecular classification and guide the clinical prognosis analysis.
【学位授予单位】：中国人民解放军军事医学科学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.9
[Abstract]:Breast cancer is a common malignant tumor of women. In recent years, the incidence of breast cancer in China has risen year by year and has been developing towards a low age. It has become a major public health problem in today's society. Although with the development of medicine, early diagnosis and treatment have reduced the mortality of breast cancer patients. Breast cancer is still a high mortality disease in malignant tumors. Breast cancer is a highly heterogeneous disease. Even in pathological type, histological grade, clinical stage and treatment, the survival prognosis of the patients is greatly different,.Perou and Sorlie and others classify breast cancer according to the molecular classification, divided into Basal-like, Luminal A, L Uminal B, HER2 overexpression, Normal and other types of type five, different types of breast cancer patients have significant differences in their treatment and prognosis. Therefore, detailed molecular typing can effectively reflect the biological characteristics of breast cancer and guide clinical selection and prognosis. Chondrosarcoma related groups are located in 1 (Chondrosarcoma associated gene 1, CSAG1). Human chromosome Xq28, studies show that CSAG1 is expressed in normal testicular tissue, and there are also reports that CSAG1 is a high expression gene in melanoma cell lines, leukemia K562 cells and malignant neurilemmoma. Research shows that CSAG1 has a close relationship with the development of tumor, but the expression of CSAG1 gene in breast cancer The study of the status and prognostic value has not been reported. Objective: To explore the expression and role of CSAG1 in the molecular classification of different breast cancer, and to provide the necessary theoretical basis for the treatment and prognosis of clinical breast cancer. Methods: 1. Formalin Fixed and Paraff (Formalin Fixed and Paraff) was fixed by immunohistochemistry. In Embedded, FFPE) tissue samples combined with the expression of ER, PR, HER2, Ki-67 to carry out molecular typing.2. using RT-PCR method to detect CSAG1 gene in the FFPE tissue samples of breast cancer patients with different molecular typing and the level of mRNA expression in breast cancer cell lines. Expression level of.4. in FFPE tissue samples of breast cancer patients using chi square test and Spearman level correlation analysis the relative expression level between CSAG1 and clinicopathological parameters and classic markers of breast cancer and correlation.5. using GraphPad Prism 5 statistical software and cbioportal bioinformatics analysis software for survival analysis. Results: 1. According to the 2011 St.Gallen consensus, combined with postoperative pathological immunohistochemical staining of Er, PR, HER2 and Ki-67, 100 cases of breast cancer were classified, 18 cases of luminala subtype, 29 cases of luminalb subtype, 4 cases of HER2 overexpression subtype, 35 cases of basal-like subtype, 14 case of normal (non-basal-like) subtype of.2. breast cancer molecular classification, respectively. The analysis of the relationship with the clinicopathological parameters showed that the proportion of I-II levels in the luminala subtype of breast cancer was the highest, and there was a statistically significant difference between the groups (P0.0001).3. in the FFPE tissue samples of the patients with clinical breast cancer, the RT-PCR results showed that the expression of the CSAG1 gene in the subtype luminala subtype of breast cancer was higher than that of the other milk. The differences in the subtypes of adenocarcinoma molecules were statistically significant (P0.05).4. in different breast cancer cell lines, RT-PCR results showed that the CSAG1 gene was in T47D (luminala type breast cancer, ER and / or pr+, her2-, ki-6714%) and the other breast cancer cell lines (zr75.1, hdq-p1, pr+, etc.) and normal mammary epithelial cell lines Significant high expression, the difference was statistically significant (P0.0001).5. in the FFPE tissue samples of patients with clinical breast cancer, the immunohistochemical results showed that the positive rate of CSAG1 protein expression in the luminala subtype of breast cancer was higher than that of other breast cancer subtypes, and the difference between the groups was statistically significant (P0.05) the expression of.6.csag1 protein and clinical disease. The analysis of the relationship between the parameters showed that the expression of CSAG1 protein was higher in the I-II level of the histological grade than in the III level, and the difference was statistically significant (P0.05). There was no significant difference in the relative expression level between the clinical histopathology type, the TNM stage, the age, the tumor diameter and the lymph node metastasis group, and the relative expression level was.7.csag1 and clinical. The correlation analysis of pathological parameters showed that the expression of CSAG1 protein was negatively correlated with the clinical histopathological classification, and the difference was statistically significant (P0.05), but there was no correlation with age, tumor diameter, lymph node metastasis, clinical histopathology type and TNM stage, and the expression of.8.csag1 protein and the classic breast cancer markers were not related. The analysis of the relationship between objects showed that the expression of CSAG1 protein in the positive group of ER and Ki-67 was slightly higher than that in the negative group of ER and Ki-67 negative group, and the relative expression level in the negative group of HER2 negative group was higher than that in the HER2 positive group, but there was no significant difference between the expression level of the positive and negative groups of PR and the correlation analysis of the classic markers of breast cancer in the positive and negative groups of pr. The results of Spearman correlation showed that the expression of CSAG1 protein was positively correlated with Er, PR expression (r=0.237,0.173), and negatively correlated with Ki-67 proliferation index (r=-0.251), and the difference was statistically significant (P0.05), while the correlation analysis between.10. and clinical breast cancer patients showed no correlation with HER2 protein expression: the positive expression of CSAG1 protein was positive. The survival of breast cancer patients is better than that of CSAG1 protein negative breast cancer patients, but the difference is not statistically significant. Conclusion: 1.CSAG1 is highly expressed in the Luminal A subtype of breast cancer, suggesting that the high expression of CSAG1 in breast cancer can be selected as the Luminal A subtype, and the clinical disease of the breast cancer patients with the high expression of.2. and CSAG1 is certain. The histological grade is low, the survival time is slightly higher, the prognosis is relatively good, the expression of.3.CSAG1 is positively correlated with the expression of ER and PR. It can be used as one of the indicators of endocrine therapy and prognosis judgment in adjuvant clinical breast cancer patients,.4.CSAG1 expression is negatively correlated with the Ki-67 proliferation index, which can assist Ki-67 to enter the molecular classification and guide the clinical prognosis analysis.
【学位授予单位】：中国人民解放军军事医学科学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.9
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