本文選題：前列腺癌　切入點：Sox5　出處：《天津醫(yī)科大學》2016年博士論文　論文類型：學位論文

【摘要】：前列腺癌在我國已成為泌尿系統(tǒng)中發(fā)病率最高的腫瘤,目前的治療方法有手術、放射治療和化療、免疫治療及激素治療等,藥物治療主要是針對雄激素剝奪治療(Androgen deprivation therapy,ADT),主要有比卡魯胺、MDV3100、醋酸阿比特龍等抑制雄激素產(chǎn)生或者抑制其功能,即便有著越來越多的藥物可供選擇,絕大多數(shù)患者在經(jīng)過ADT治療后的一到兩年內(nèi)疾病繼續(xù)惡化,發(fā)展為去勢抵抗性前列腺癌(castration resistance prostate cancer,CRPC),而且前列腺癌轉移發(fā)生率高,許多患者診斷時已經(jīng)伴隨骨轉移,骨轉移不僅影響患者預后,其帶來的并發(fā)癥如骨痛、病理性骨折、神經(jīng)壓迫癥狀等還嚴重降低患者生活質量。深入研究CRPC的形成以及腫瘤轉移機制,對于患者的治療至關重要。在肝癌、垂體瘤等腫瘤的研究中發(fā)現(xiàn)Sox5與增殖轉移密切相關,對于前列腺癌患者腫瘤的原位組織標本行免疫組化檢測發(fā)現(xiàn)Sox5表達增高患者的往往伴隨著轉移的發(fā)生,獲取淋巴結轉移灶的標本Sox5的表達也較高,而且Sox5表達升高的患者間質標志物表達也高,Sox5與N-cad的表達呈現(xiàn)顯著正相關。對前列腺癌細胞系22Rv1、PC3敲減Sox5,劃痕實驗及遷移實驗均提示細胞的遷移能力明顯下降,蛋白電泳檢測上皮標志物增加,間質標志物減少,提示Sox5與上皮間質轉化(epithelial mesenchymal transition,EMT)的發(fā)生相關,而且敲減Sox5后細胞的增殖能力也減低。利用22Rv1細胞系敲減Sox5進行小鼠前列腺原位種瘤,發(fā)現(xiàn)敲減Sox5后的腫瘤發(fā)生淋巴結等轉移相對較少,而且皮下種瘤的瘤體體積也較小,提示Sox5與前列腺癌進展相關。對LNCa P細胞應用TGF-β進行EMT誘導時發(fā)現(xiàn)Sox5的表達也增加,在應用TGF-β的同時敲減Sox5可以逆轉EMT的發(fā)生,在TGF-β信號通路中Smad2/3發(fā)揮著重要作用,利用JASPAR在線預測軟件發(fā)現(xiàn)Smad3可以結合Sox5的promotoer區(qū)域,利用Smad3行CHIP以及Luciferase實驗驗證了這一直接調節(jié)作用。根據(jù)文獻報道及JASPAR預測Sox5可以調節(jié)TWIST1的表達,敲減Sox5后檢測TWIST1的表達下降,提示Sox5調節(jié)Twist1的表達進而調控EMT,利用Sox5行CHIP以及Luciferase實驗驗證了這一直接調節(jié)作用。為了更好研究CRPC的形成,我們利用實驗室已有的去雄激素血清長期培養(yǎng)LNCa P細胞系建立去雄激素抵抗的細胞系CR-LN,在長期去雄培養(yǎng)以后,細胞的Sox5的表達增加,細胞的增殖能力回升,而且細胞發(fā)生EMT的轉變,對CR-LN敲減Sox5后,細胞的間質標志物表達下降,但是對CR-LN進行TGF-β誘導時未發(fā)生明顯EMT,提示CR-LN的TGF-β信號通路的自身活化。對CR-LN敲減Sox5后行遷移實驗發(fā)現(xiàn)細胞的遷移能力下降,MTT實驗發(fā)現(xiàn)細胞的活力也下降,球形成實驗表明細胞的自我更新能力下降,提示Sox5對于CRPC的形成也發(fā)揮著一定作用。上述結果提示,在持續(xù)性雄激素剝奪治療前列腺癌過程中,由于AR對于Sox5的抑制作用解除,導致Sox5的表達升高,腫瘤細胞的增殖能力也逐漸恢復,而且細胞可能受腫瘤微環(huán)境中TGF-β等細胞因子作用發(fā)生EMT,促進腫瘤的轉移。對于CRPC的治療,阻斷Sox5及其下游可能是新的治療策略。
[Abstract]:Prostate cancer in China has become the highest incidence of cancer in the urinary system, the current treatment methods include surgery, radiotherapy and chemotherapy, immune therapy and hormone therapy, drug treatment is mainly for androgen deprivation therapy (Androgen deprivation, therapy, ADT), the main bicalutamide, MDV3100, etc. abbitt acetic acid inhibits androgen Liuzhou to produce or inhibit its function, even if there are more and more drugs are available, most of the patients after ADT treatment after one to two years the disease continues to deteriorate, the development of adenocarcinoma before castration resistant (castration resistance prostate cancer column, CRPC), and the high rate of metastasis of prostate cancer, many patients have been diagnosed with bone metastasis, bone metastasis not only affects the prognosis of the patients, the complications such as bone ache, pathological fracture, nerve compression symptoms also seriously reduce the quality of life of patients . to research the mechanism of the CRPC formation and tumor metastasis, is essential for the treatment of patients. In the liver of pituitary tumor and tumor was found in Sox5 and the proliferation of tumor metastasis for patients with prostate cancer in situ tissue specimens for immunohistochemical detection showed that the expression of Sox5 was increased in patients with often accompanied with metastasis, access the expression of metastatic lymph node specimens of Sox5 were higher, and the expression of Sox5 increased in patients with interstitial marker expression is also high, showing a significant positive correlation with the expression of Sox5 N-cad. 22Rv1 on prostate cancer cell line, PC3 knockdown of Sox5, scratch test and migration experiments indicate that the ability of cell migration was significantly decreased, protein electrophoresis detection of epithelial markers increased, interstitial marker decreased, suggesting that Sox5 and epithelial mesenchymal transition (epithelial mesenchymal, transition, EMT) related to the occurrence, and knock in Sox5 cells by increasing Colonization ability also decreased. Knockdown of Sox5 mouse prostate tumor cell lines by in situ 22Rv1, found that knockdown of Sox5 after tumor lymph node metastasis is relatively small, and the tumor volume of subcutaneous tumor is small, suggesting that progress of Sox5 associated with prostate cancer. The LNCa P cells with TGF- beta induced by EMT to find the expression of Sox5 also increased at the same time, application of TGF- beta knockdown of Sox5 can reverse the occurrence of EMT, in the TGF- beta Smad2/3 pathway plays an important role in predicting software found Smad3 can bind to promotoer region of Sox5 by JASPAR online, using Smad3 CHIP and Luciferase experimental results verify the direct role the expression of JASPAR. According to the literatures and prediction of Sox5 can regulate TWIST1, knockdown of Sox5 expression after detection of TWIST1 decreased, suggesting that Sox5 regulates the expression and regulation of EMT by Twist1, Sox5 CHIP and Lucifera Se experimental results verify the direct role. In order to form a better study of CRPC, we use the serum androgen deprivation long-term cultures of LNCa P cell line to establish androgen resistant cell line CR-LN, after long-term training increased the expression of emasculation, Sox5 rebound, cell proliferation, and cell EMT change of CR-LN knockdown Sox5 cells, interstitial marker expression decreased, but the CR-LN TGF- did not change significantly when induced by EMT, suggesting that TGF- beta CR-LN signaling pathway activation. The knockdown of CR-LN on Sox5 after migration experiments showed decreased cell migration, MTT experiments showed that the cell vitality the ball drops, forming assay showed that cell self-renewal capacity decreased, suggesting that Sox5 for the formation of CRPC also plays a role. The above results suggest that, in the continuous androgen deprivation therapy for prostate In the process of cancer, due to AR for the inhibition of Sox5 release, resulting in increased expression of Sox5, tumor cell proliferation also gradually restored, and TGF- beta cells may be the tumor microenvironment in the role of cytokines EMT promote tumor metastasis. For the treatment of CRPC, blocking Sox5 and its downstream treatment strategies may be new.

【學位授予單位】：天津醫(yī)科大學
【學位級別】：博士
【學位授予年份】：2016
【分類號】：R737.25

【參考文獻】

相關期刊論文 前1條

1 Wanqing Chen;Rongshou Zheng;Hongmei Zeng;Siwei Zhang;Jie He;;Annual report on status of cancer in China, 2011[J];Chinese Journal of Cancer Research;2015年01期

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本文編號：1642787