CSAG1基因在乳腺癌及分子分型中的差異表達研究及臨床意義
發(fā)布時間：2018-07-31 09:11
【摘要】：乳腺癌來源于乳腺上皮組織,為女性常見的一種惡性腫瘤,近年來我國乳腺癌發(fā)病率逐年上升并朝低齡化趨勢發(fā)展,已成為當今社會的重大公共衛(wèi)生問題。雖然隨著醫(yī)學(xué)的發(fā)展,早期診斷和治療降低了乳腺癌患者的死亡率,但就目前而言,乳腺癌仍是惡性腫瘤中死亡率較高的疾病。乳腺癌屬于高度異質(zhì)性疾病,即使在病理類型、組織學(xué)分級、臨床分期及治療均相同的情況下,患者的生存預(yù)后也存在極大差別。Perou和Sorlie等人將乳腺癌根據(jù)分子分型,分為Basal-like、Luminal A、Luminal B、HER2過表達、Normal等五型,不同分型的乳腺癌患者其治療及預(yù)后差異明顯。因此,詳細的分子分型能有效的反映乳腺癌的生物學(xué)特征,并指導(dǎo)臨床選擇治療方案及判斷預(yù)后。軟骨肉瘤相關(guān)基因1(Chondrosarcoma associated gene 1,CSAG1),定位于人染色體Xq28,研究表明CSAG1在正常睪丸組織中表達,同時也有報道顯示CSAG1是在黑色素瘤細胞株、白血病K562細胞以及惡性神經(jīng)鞘瘤組織中高表達的基因,研究表明CSAG1與腫瘤發(fā)生、發(fā)展有著密切的聯(lián)系,但有關(guān)乳腺癌中CSAG1基因的表達情況和預(yù)后價值的相關(guān)研究還未見報道。目的:探索CSAG1在不同乳腺癌分子分型中的表達及作用,為臨床乳腺癌治療和預(yù)后判斷提供必要的理論依據(jù)。方法:1.通過免疫組織化學(xué)方法對100例乳腺癌福爾馬林固定石蠟包埋(Formalin Fixed and Paraffin Embedded,FFPE)組織樣本結(jié)合ER、PR、HER2、Ki-67的表達情況進行分子分型。2.采用RT-PCR方法檢測CSAG1基因在不同分子分型的乳腺癌臨床患者FFPE組織樣本及乳腺癌細胞系中mRNA表達水平。3.通過免疫組織化學(xué)方法檢測CSAG1蛋白在不同分子分型的乳腺癌臨床患者FFPE組織樣本中表達水平。4.利用卡方檢驗及Spearman等級相關(guān)分析CSAG1與臨床病理參數(shù)及乳腺癌經(jīng)典標志物之間的相對表達水平及相關(guān)性。5.運用GraphPad Prism 5統(tǒng)計軟件及cbioportal生物信息學(xué)分析軟件進行生存分析。結(jié)果:1.根據(jù)2011年st.gallen共識,并結(jié)合術(shù)后病理免疫組織化學(xué)染色er、pr、her2、ki-67表達情況將100例乳腺癌進行分子分型,分別篩選出luminala亞型18例,luminalb亞型29例,her2過表達亞型4例,basal-like亞型35例,normal(non-basal-like)亞型14例。2.乳腺癌分子分型與臨床病理參數(shù)的關(guān)系分析結(jié)果顯示:在luminala亞型乳腺癌中組織學(xué)分級i-ii級所占比例最高,組間具有統(tǒng)計學(xué)差異(p0.0001)。3.在臨床乳腺癌患者ffpe組織樣本中rt-pcr結(jié)果顯示:csag1基因在乳腺癌分子分型luminala亞型中的表達量高于其他乳腺癌分子亞型,組間差異具有統(tǒng)計學(xué)意義(p0.05)。4.在不同乳腺癌細胞系中rt-pcr結(jié)果顯示:csag1基因在t47d(luminala型乳腺癌,er和/或pr+,her2-,ki-6714%)中較其他乳腺癌細胞系(zr75.1、hdq-p1、cal-120、cal-51、mda-mb-436、mda-mb-231)及正常乳腺上皮細胞系(hbl-100)顯著高表達,差異具有統(tǒng)計學(xué)意義(p0.0001)。5.在臨床乳腺癌患者ffpe組織樣本中免疫組化結(jié)果顯示:csag1蛋白表達陽性率在乳腺癌分子分型luminala亞型中高于其他乳腺癌分子亞型,組間差異具有統(tǒng)計學(xué)意義(p0.05)。6.csag1蛋白表達與臨床病理參數(shù)之間的關(guān)系分析結(jié)果顯示:csag1蛋白表達在組織學(xué)分級的i-ii級中較iii級中相對表達水平高,差異具有統(tǒng)計學(xué)意義(p0.05);在臨床病理組織學(xué)類型、tnm分期、年齡、腫瘤直徑、淋巴結(jié)轉(zhuǎn)移分組中相對表達水平均無明顯差別。7.csag1與臨床病理參數(shù)相關(guān)性分析利用spearman等級相關(guān)結(jié)果顯示:csag1蛋白表達與臨床病理組織學(xué)分級呈負相關(guān),差異具有統(tǒng)計學(xué)意義(p0.05);而與年齡、腫瘤直徑、淋巴結(jié)轉(zhuǎn)移、臨床病理組織學(xué)類型及tnm分期均無相關(guān)性。8.csag1蛋白表達與乳腺癌經(jīng)典標志物之間的關(guān)系分析結(jié)果顯示:csag1蛋白表達在er及ki-67陽性組中較er及ki-67陰性組中相對表達水平稍高,在her2陰性組中較her2陽性組中相對表達水平高,而在pr陽性與陰性組中表達水平無明顯差別。9.csag1與乳腺癌經(jīng)典標志物相關(guān)性分析利用spearman等級相關(guān)結(jié)果顯示:csag1蛋白表達與er、pr表達呈正相關(guān)(r=0.237、0.173),與ki-67增殖指數(shù)呈負相關(guān)(r=-0.251),差異均具有統(tǒng)計學(xué)意義(p0.05);而與her2蛋白表達無相關(guān)性。10.與臨床乳腺癌患者生存相關(guān)性分析結(jié)果顯示:csag1蛋白表達陽性的乳腺癌患者較CSAG1蛋白表達陰性的乳腺癌患者后期的生存稍好,但差異無統(tǒng)計學(xué)意義。結(jié)論:1.CSAG1在乳腺癌分子分型Luminal A亞型中高表達,提示CSAG1高表達在乳腺癌中篩選出Luminal A亞型有一定作用。2.高表達CSAG1的乳腺癌患者其臨床病理組織學(xué)分級低、生存期稍高,預(yù)后相對較好。3.CSAG1表達與ER、PR表達呈正相關(guān),可作為輔助臨床乳腺癌患者行內(nèi)分泌治療及預(yù)后判斷的指標之一。4.CSAG1表達與Ki-67增殖指數(shù)呈負相關(guān),可輔助Ki-67進行分子分型并指導(dǎo)臨床進行預(yù)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.
【學(xué)位授予單位】：中國人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：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.
【學(xué)位授予單位】：中國人民解放軍軍事醫(yī)學(xué)科學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R737.9
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