本文選題：乳腺癌　切入點：MTH1　出處：《北京協(xié)和醫(yī)學(xué)院》2015年博士論文　論文類型：學(xué)位論文

【摘要】：第一部分MTH1在人乳腺癌組織中的表達目的：分析MTH1蛋白、mRNA在人乳腺癌組織的表達情況以及其表達和不同臨床病理特征的關(guān)系。方法：從普通標(biāo)本庫中隨機選取Luminal型、Her-2過表達型、Basal-like型浸潤性乳腺癌標(biāo)本、非癌乳腺組織標(biāo)本各10例,行免疫組織化學(xué)檢測MTHl蛋白的表達；從液氮標(biāo)本庫隨機選取Luminal型、Her-2過表達型、Basal-like型浸潤性乳腺癌標(biāo)本以及其相對應(yīng)的非癌乳腺組織標(biāo)本各10例行熒光定量PCR檢測MTH1 mRNA的表達。結(jié)果：乳腺癌組織中MTH1蛋白強陽/中陽表達率為93.3%,非癌乳腺組織MTH1蛋白強陽/中陽表達率為10%,其余為弱陽/陰性表達。乳腺癌組織與非癌乳腺組織中MTH1蛋白的表達強度存在顯著差異(P0.001)。MTH1蛋白的表達在乳腺癌不同分子亞型、不同年齡、不同腫瘤大小、不同淋巴結(jié)狀況等亞組之間無明顯組間差異(P0.05)。乳腺癌組織中MTHl mRNA的相對表達水平是非癌乳腺組織中MTHl mRNA表達水平的3.15(±1.67)倍,差異顯著(P=0.003)。 MTH1 mRNA的表達在乳腺癌不同分子亞型、不同年齡、不同腫瘤大小、不同淋巴結(jié)狀況等亞組之間無明顯組間差異(P0.05)。結(jié)論：在蛋白和mRNA水平,乳腺癌組織中MTH1的表達顯著高于非癌乳腺組織。乳腺癌組織中MTH1的高表達不因分子亞型、年齡、腫瘤大小、淋巴結(jié)狀況等臨床病理特征而出現(xiàn)顯著改變。第二部分MTH1途徑對不同亞型乳腺癌細胞系的體外抑制作用研究目的：研究MTH1途徑抑制劑對不同亞型的乳腺癌細胞系體外生長的影響方法：選取不同乳腺癌亞型有代表性的細胞系,Luminal型乳腺癌細胞系MCF-7、 Basal-like型乳腺癌細胞系MDA-MB-231以及Her-2過表達型乳腺癌細胞系MDA-MB-453進行MTH1的Western Blot檢測確定其表達是否陽性。采用CCK-8細胞增殖實驗測定不同濃度的MTH1抑制劑對各細胞系體外增殖的影響,采用克隆形成實驗測定不同濃度的MTH1抑制劑對各細胞系克隆生存能力的影響。結(jié)果：不同亞型乳腺癌細胞系的MTH1蛋白均有較強的表達。CCK-8細胞增殖實驗顯示,隨著MTH1抑制劑藥物濃度的上升,乳腺癌細胞系的活力呈下降趨勢。Luminal型乳腺癌細胞系MCF-7及Basal-l ike型乳腺癌細胞系MDA-MB-231在試驗藥物濃度范圍內(nèi)均取得了IC50值,分別為11.91umol/l和12.86umol/l:Her-2過表達型細胞系MDA-MB-453在試驗藥物濃度范圍內(nèi)未取得IC50值。克隆形成實驗顯示,不同亞型的乳腺癌細胞系的克隆形成均受到了明顯的抑制并且取得了IC50值。Luminal型細胞系MCF-7.Basal-like型乳腺癌細胞系MDA-MB-231、Her-2過表達型細胞系MDA-MB-453的IC50值分別為9.78umol/l、6.96umol/l和8.97umol/l。結(jié)論：細胞增殖實驗及克隆形成實驗顯示,在體外條件下,對MTH1途徑進行干預(yù),能夠顯著的抑制Lumirlal型乳腺癌細胞系MCF-7、Basal-l ike型乳腺癌細胞系MDA-MB-231以及Her-2過表達型乳腺癌細胞系MDA-MB-453的生長。第三部分MTH1途徑對荷人不同亞型乳腺癌裸鼠的抑瘤作用研究目的：研究MTH1途徑抑制劑對荷人不同亞型乳腺癌裸鼠體內(nèi)移植瘤生長的影響方法：選取不同乳腺癌亞型的有代表性的細胞系,Luminal型乳腺癌細胞系MCF-7、Basal-like型乳腺癌細胞系MDA-MB-231以及Her-2過表達型乳腺癌細胞系MDA-MB-453接種于裸鼠建立裸鼠荷人不同乳腺癌亞型移植瘤模型。對荷瘤裸鼠及正常裸鼠進行MTH1抑制劑干預(yù),繪制腫瘤生長曲線及裸鼠體重曲線,并進行血液和生化學(xué)檢測,測定MTH1抑制劑對不同亞型乳腺癌裸鼠移植瘤生長的影響,以及MTH1抑制劑的安全性。結(jié)果：藥物抑制實驗顯示,MTH1抑制劑療程結(jié)束后,不同亞型乳腺癌細胞系移植瘤的最終體積分別為MCF-7(給藥組 vs.對照組,60.1 mm3 vs.820.8 mm3, P0.05), MDA-MB-231(給藥組 vs.對照組,30.1 mm3 vs.313.5 mm3, P0.05), MDA-MB-453(給藥組 vs.對照組,0 mm3 vs.5.2 mm3, P0.05),差異顯著。給藥組裸鼠的最終體重與給藥前初始體重分別為：MCF-7(最終 vs.給藥前,16.81g vs.17.44g,P0.05), MDA-MB-231(最終 vs.給藥前,17.42g vs.17.68g, P0.05), MDA-MB-453(最終vs. 給藥前,17.23g vs.17.69g, P0.05)。單純給藥組裸鼠的最終體重與給藥前體重變化為0.73(±0.34)g,無瘤無藥組裸鼠相應(yīng)的體重變化為2.26(±0.29)g,二者差異顯著(P0.05)。單純給藥組與無瘤無藥組裸鼠主要血常規(guī)和肝腎功指標(biāo)無明顯差別(P0.05)。結(jié)論：在裸鼠體內(nèi),對MTH1途徑進行干預(yù),能夠顯著的抑制Luminal型乳腺癌細胞系MCF-7、Basa1-like型乳腺癌細胞系MDA-MB-231以及Her-2過表達型乳腺癌細胞系MDA-MB-453移植瘤的生長。裸鼠的生長體重和血液學(xué)檢查提示,短期應(yīng)用MTH1抑制劑是安全的。
[Abstract]:The first part of the expression of MTH1 in human breast carcinoma: analysis of MTH1 protein, the expression of mRNA in human breast cancer tissues and the relationship between its expression and different clinical pathological features. Methods: the samples were randomly selected from the general library of Luminal, over expression of Her-2, Basal-like type of invasive breast cancer from non cancer breast tissue samples from 10 patients, the expression of MTHl protein detected by immunohistochemistry; specimens were randomly selected from liquid nitrogen Luminal, Her-2 over expression of type of breast cancer specimens and to detect the expression of MTH1 mRNA in breast cancer tissue samples which correspond to 10 patients each fluorescent quantitative PCR Basal-like. Results: invasive breast cancer in Qiangyang / Zhongyang MTH1 protein expression rate was 93.3%, non cancerous breast tissue MTH1 protein positive expression rate was 10% in Zhongyang, the expression was weak positive / negative. MTH1 breast cancer and non cancerous breast tissue There are significant differences in the expression intensity of protein (P0.001) expression of.MTH1 protein in different molecular subtypes of breast cancer, different age, different tumor size, there was no difference between the different lymph node status subgroup (P0.05). The relative expression of MTHl in breast cancer tissue mRNA level is the level of MTHl mRNA expression in non cancer breast tissues 3.15 (+ 1.67) times, significant difference (P=0.003). The expression of MTH1 mRNA in different molecular subtypes of breast cancer, different age, different tumor size, there was no difference between the different lymph node status subgroup (P0.05). Conclusion: the mRNA and protein level, the expression of MTH1 in breast cancer is significantly higher than that in breast cancer tissue. High expression of MTH1 in breast cancer by molecular subtype, age, tumor size, lymph node status and clinicopathological characteristics had significant change. The second part of the MTH1 pathway in different subtypes of breast cancer Objective to study inhibitory effect of cell lines in vitro: breast cancer cells of MTH1 pathway inhibitors on different subtypes: different subtypes of breast cancer cell lines representative, Luminal type of breast cancer cell line MCF-7 Western Blot detection of Basal-like breast cancer cell line MDA-MB-231 and Her-2 over expression of type MDA-MB-453 breast cancer cell line MTH1 to determine whether the expression is positive. The effects of MTH1 inhibitors in different concentration were determined by CCK-8 cell proliferation assay in vitro proliferation of different cell lines, the effect of clone formation assay of MTH1 inhibitors with different concentrations on the viability of the cell clones. Results: different subtypes of breast cancer cell lines MTH1 protein had strong expression of.CCK-8 cell proliferation experiment, with increasing inhibitor concentration MTH1, breast cancer cell vitality Decreased.Luminal breast cancer cell lines MCF-7 and Basal-l type Ike breast cancer cell line MDA-MB-231 in the experimental drug concentration range were obtained in IC50, respectively 11.91umol/l and 12.86umol/l:Her-2 over expression of type MDA-MB-453 cells in the experimental drug concentration range did not obtain IC50 value. Clone formation assay showed that the cloned human breast cancer cell line different subtypes of formation were inhibited obviously and achieved IC50 values of type MCF-7.Basal-like breast cancer cell line MDA-MB-231 and.Luminal cell lines, expression of Her-2 MDA-MB-453 IC50 cell line values were respectively 9.78umol/l, 6.96umol/l and 8.97umol/l. conclusion: the cell proliferation and clone formation assay, in vitro, intervention on the way to MTH1, can significantly inhibit type Lumirlal breast cancer cell line MCF-7, Basal-l type Ike breast cancer cell line MDA-MB-231 and Her Over expression of -2 in breast cancer cell line MDA-MB-453 growth. Objective to study on anti tumor effect of MTH1 pathway on third different subtypes of breast cancer bearing nude mice: Study of MTH1 pathway inhibitors growth in different subtypes of breast cancer xenograft in nude mice bearing human breast cancer: different subtypes of representative the cell line, Luminal type of breast cancer cell line MCF-7 Basal-like breast cancer cell line MDA-MB-231 and Her-2 in nude mice xenografts in different subtypes of breast cancer xenograft model of breast cancer cell line MDA-MB-453 was expression. For MTH1 inhibitor intervention on nude mice and normal mice, the tumor growth curve and the weight of nude mice the curve, and the blood and Biochemistry test, determination of the effects of MTH1 inhibitors on the growth of different subtypes of breast cancer in nude mice, and the safety of MTH1 inhibitors. Results: drug Inhibition experiment showed that after MTH1 inhibitor treatment, the final volume of different subtypes of breast cancer cell line xenografts were MCF-7 (drug group vs. control group, 60.1 mm3 vs.820.8 mm3, P0.05), MDA-MB-231 (drug group vs. control group, 30.1 mm3 vs.313.5 mm3, P0.05), MDA-MB-453 (group of vs. the control group, 0 mm3 vs.5.2 mm3, P0.05), the difference was significant. To the final weight of nude mice and the medicine group before administration of initial weight were: MCF-7 (vs. 16.81g vs.17.44g, before administration, P0.05 (MDA-MB-231), the final vs. before administration, 17.42g vs.17.68g, P0.05 MDA-MB-453 (vs.), the final delivery before 17.23g, vs.17.69g, P0.05). Only to the final weight of nude mice and medicine group before administration of weight change was 0.73 (+ 0.34) g, the changes of body weight of tumor free drug free mice corresponding to 2.26 (+ 0.29) g, two had significant difference (P0.05). The simple drug group and no tumor free the main blood group of nude mice There was no significant difference between the normal and kidney function index (P0.05). Conclusion: in vivo intervention on MTH1 pathway, inhibited the Luminal breast cancer cell line MCF-7, Basa1-like type of breast cancer cell line MDA-MB-231 and Her-2 overexpression breast cancer cell line MDA-MB-453 on the growth of transplanted tumor growth in nude mice body weight and blood. Examination indicated that short-term use of MTH1 inhibitors is safe.

【學(xué)位授予單位】：北京協(xié)和醫(yī)學(xué)院
【學(xué)位級別】：博士
【學(xué)位授予年份】：2015
【分類號】：R737.9

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