本文選題：PD-L1 + 免疫治療��； 參考：《南方醫(yī)科大學(xué)》2017年碩士論文

【摘要】：研究背景及目的肺癌是最常見(jiàn)的腫瘤,同時(shí)也是國(guó)內(nèi)外癌癥死因之首。表皮生長(zhǎng)因子受體酪氨酸激酶抑制劑(EGFR-TKIs)能有效的延長(zhǎng)部分肺癌患者的生存期。然而EGFR-T790M 突變、MET 擴(kuò)增、HGF 分泌、HER2 擴(kuò)增、IGF-1R 的活化、EMT、PTEN缺失等機(jī)制使患者對(duì)EGFR-TKIs產(chǎn)生耐藥。因此,尋找有效的治療方法對(duì)于肺癌患者尤其是EGFR-TKIs耐藥的患者而言意義重大。靶向PD-1/PD-L1通路的免疫檢查點(diǎn)治療已在多種惡性腫瘤的治療中取得重大突破,2017年NCCN指南推薦抗PD-1/PD-L1療法用于高表達(dá)PD-L1的肺癌患者。既往研究報(bào)道EGFR-TKIs可降低EGFR突變肺癌細(xì)胞PD-L1的表達(dá),然而肺癌患者發(fā)生EGFR-TKIs耐藥后腫瘤PD-L1的表達(dá)情況卻鮮有報(bào)道,EGFR-TKIs耐藥患者能否從抗PD-L1療法中獲益也有待明確。本研究通過(guò)TCGA數(shù)據(jù)庫(kù)明確EGFR-TKIs敏感及耐藥腫瘤及腫瘤微環(huán)境的差異,通過(guò)體內(nèi)外實(shí)驗(yàn)證明EGFR-TKIs耐藥可上調(diào)肺癌PD-L1的表達(dá),并探究?jī)?nèi)在調(diào)控機(jī)制,同時(shí)通過(guò)體內(nèi)外實(shí)驗(yàn)評(píng)估了抗PD-L1療法對(duì)EGFR-TKIs耐藥肺癌的療效,為EGFR-TKI耐藥患者提供新的治療方法。方法使用吉非替尼耐藥MET擴(kuò)增的肺癌細(xì)胞PC-9R、肝細(xì)胞生長(zhǎng)因子HGF刺激后的肺癌細(xì)胞、轉(zhuǎn)染EGFR-19Del和/或EGFR-T790M突變質(zhì)粒的293FT細(xì)胞,分別建立MMET擴(kuò)增、HGF和EGFR-T790M突變的EGFR-TKIs耐藥細(xì)胞模型。用Western Blot、流式細(xì)胞技術(shù)和RT-qPCR等技術(shù)比較前述細(xì)胞PD-L1的表達(dá),并探究調(diào)控機(jī)制。將前述模型與T細(xì)胞共培養(yǎng)并用LDH細(xì)胞毒性實(shí)驗(yàn)比較腫瘤的免疫逃逸能力。PD-L1基因敲除的肺癌細(xì)胞及其陰性對(duì)照建立皮下瘤模型并用T細(xì)胞治療比較腫瘤的免疫逃逸能力。分析TCGA數(shù)據(jù)庫(kù)中肺腺癌相關(guān)的數(shù)據(jù),比較EGFR-TKIs敏感及耐藥肺癌的腫瘤突變負(fù)荷、PD-L1等免疫抑制分子及的表達(dá)、腫瘤微環(huán)境中腫瘤浸潤(rùn)淋巴細(xì)胞、細(xì)胞毒活性的改變。結(jié)果體外實(shí)驗(yàn)結(jié)果表明前述三種機(jī)制均可上調(diào)腫瘤PD-L1的表達(dá),PI3K/Akt、MAPK、NF-kappa B信號(hào)通路參與EGFR-T790M突變誘導(dǎo)的PD-L1的表達(dá);PI3K/Akt信號(hào)通路、MAPK信號(hào)通路而非NF-kappa B信號(hào)通路參與HGF和MET擴(kuò)增誘導(dǎo)的PD-L1的表達(dá);體內(nèi)外實(shí)驗(yàn)結(jié)果表明MET擴(kuò)增、HGF、EGFR-T790M突變上調(diào)腫瘤細(xì)胞PD-L1的表達(dá)降低T淋巴細(xì)胞的殺傷效果,敲除PD-L1基因降低PD-L1表達(dá)或抗PD-L1單克隆抗體可恢復(fù)人T淋巴細(xì)胞的殺傷能力。TCGA數(shù)據(jù)庫(kù)中肺腺癌相關(guān)的數(shù)據(jù)提示:較EGFR-TKIs敏感的腫瘤而言,EGFR-TKIs耐藥肺癌的腫瘤突變負(fù)荷高,PD-L1、PD-1等免疫抑制分子的表達(dá)增加。結(jié)論(1)相比EGFR-TKIs敏感肺癌而言,EGFR-TKIs耐藥肺癌腫瘤突變負(fù)荷更高,腫瘤免疫抑制作用更強(qiáng)。(2)MMET擴(kuò)增通過(guò)P13K/Akt和MAPK信號(hào)通路上調(diào)腫瘤細(xì)胞PD-L1的表達(dá)促進(jìn)非小細(xì)胞肺癌的免疫逃逸(3)HGF通過(guò)PI3K/Akt和MAPK信號(hào)通路誘導(dǎo)PD-L1的表達(dá)促進(jìn)非小細(xì)胞肺癌免疫逃逸(4)EGFR-T790M 突變通過(guò) PI3K/Akt、MAPK 和 NF-kappa B信號(hào)通路誘導(dǎo)PD-L1的表達(dá)促進(jìn)肺癌的免疫逃逸(5)EGFR-TKIs耐藥細(xì)胞及皮下瘤通過(guò)上調(diào)PD-L1降低T細(xì)胞的殺傷能力,抗PD-L1療法可恢復(fù)T細(xì)胞的殺傷能力。
[Abstract]:Background and objective lung cancer is the most common tumor, and it is also the leading cause of cancer death at home and abroad. Epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKIs) can effectively prolong the survival period of some lung cancer patients. However, EGFR-T790M mutation, MET amplification, HGF secretion, HER2 amplification, IGF-1R activation, EMT, PTEN deletion and other mechanisms make Patients are resistant to EGFR-TKIs. Therefore, finding effective treatments is of great significance for patients with lung cancer, especially for EGFR-TKIs resistant patients. Immunologic checkpoints targeting PD-1/PD-L1 pathway have made significant breakthroughs in the treatment of multiple malignant tumors. In 2017, NCCN refers to the recommendation of anti PD-1/PD-L1 therapy for high expression of PD-L1. EGFR-TKIs can reduce the expression of PD-L1 in EGFR mutant lung cancer cells. However, the expression of PD-L1 is rarely reported in patients with lung cancer after EGFR-TKIs resistance, and it is also unclear whether EGFR-TKIs resistant patients can benefit from the anti PD-L1 therapy. This study uses a TCGA database to clarify EGFR-TKIs sensitivity and tolerance. The difference between drug tumor and tumor microenvironment shows that EGFR-TKIs resistance can up regulate the expression of PD-L1 in lung cancer and explore the internal regulation mechanism, and evaluate the efficacy of anti PD-L1 therapy on EGFR-TKIs resistant lung cancer in vivo and in vitro, and provide a new treatment for EGFR-TKI resistant patients. Methods use gefitinib resistance. MET expanded lung cancer cells PC-9R, lung cancer cells stimulated by hepatocyte growth factor HGF, transfected 293FT cells with EGFR-19Del and / or EGFR-T790M mutant plasmids, respectively, to establish MMET amplification, HGF and EGFR-T790M mutant EGFR-TKIs resistant cell models, compared with Western Blot, flow cytometry, and other techniques. The previous model was co cultured with T cells and used LDH cytotoxicity test to compare the tumor immune escape ability.PD-L1 knockout lung cancer cells and their negative controls to establish a subcutaneous tumor model and compare the immune escape ability of the tumor with T cells. Analysis of the data related to lung adenocarcinoma in the TCGA database and compared E GFR-TKIs sensitive and drug-resistant lung cancer mutation load, PD-L1 and other immunosuppressive molecules and expression, tumor infiltrating lymphocytes, cytotoxic activity changes in tumor microenvironment. Results in vitro results show that the above three mechanisms can up-regulate the expression of tumor PD-L1, PI3K/Akt, MAPK, NF-kappa B signaling pathway involved in EGFR-T790M mutation The expression of induced PD-L1; PI3K/Akt signaling pathway, MAPK signaling pathway and not NF-kappa B signaling pathway involved in HGF and MET induced PD-L1 expression; in vitro and in vivo experimental results showed that MET amplification, HGF, EGFR-T790M mutation up regulation of tumor cell PD-L1 expression reduced the killing effect of lymphocyte. D-L1 monoclonal antibodies can restore human T lymphocyte killing ability in.TCGA database associated with lung adenocarcinoma data: compared with EGFR-TKIs sensitive tumors, EGFR-TKIs resistant lung cancer has high mutation load, and the expression of PD-L1, PD-1 and other immunosuppressive molecules increases. Conclusion (1) compared with EGFR-TKIs sensitive lung cancer, EGFR-TKIs resistant lung cancer The tumor mutation load is higher and the tumor immune inhibition is stronger. (2) MMET amplification through P13K/Akt and MAPK signaling pathway up-regulated the expression of PD-L1 in tumor cells promoting the immune escape of non small cell lung cancer (3) HGF induced PD-L1 expression through PI3K/Akt and MAPK signaling pathway to promote non small cell lung cancer immune escape (4) EGFR-T790M mutation through PI3K. /Akt, MAPK and NF-kappa B expression induced by PD-L1 signaling pathway to promote lung cancer immune escape (5) and EGFR-TKIs resistant cell subcutaneous tumor decreased T cell killing ability through upregulation of PD-L1, anti PD-L1 therapy can restore T cell killing ability.
【學(xué)位授予單位】：南方醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R734.2

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 王志剛,竇科峰;腫瘤免疫逃逸與轉(zhuǎn)化生長(zhǎng)因子-β關(guān)系的研究進(jìn)展[J];中華實(shí)驗(yàn)外科雜志;2005年04期

2 張?jiān)?,陳劍平 ,張彩;腫瘤免疫逃逸與腫瘤免疫治療的思考[J];實(shí)用醫(yī)學(xué)雜志;2005年04期

3 王萍;郝鈺;;中醫(yī)藥調(diào)節(jié)腫瘤免疫逃逸機(jī)制研究進(jìn)展[J];中國(guó)中醫(yī)藥信息雜志;2007年10期

4 呂程;蔣洪昆;劉宇;潘柳吟;;T淋巴細(xì)胞亞群檢測(cè)在腫瘤免疫逃逸中的作用[J];中國(guó)民族民間醫(yī)藥;2011年02期

5 寧波;腫瘤免疫與腫瘤免疫逃逸機(jī)制[J];中國(guó)實(shí)用婦科與產(chǎn)科雜志;2000年06期

6 張煊,曹雪濤;腫瘤免疫逃逸機(jī)制研究新進(jìn)展[J];中國(guó)腫瘤生物治療雜志;2001年01期

7 傅云峰,呂衛(wèi)國(guó);白介素2受體及其與腫瘤免疫逃逸[J];上海免疫學(xué)雜志;2002年02期

8 丁紅仙;;腫瘤免疫逃逸機(jī)制的研究進(jìn)展[J];海峽藥學(xué);2008年07期

9 宋彥;朱喜科;;腫瘤免疫逃逸機(jī)制的研究新進(jìn)展[J];現(xiàn)代腫瘤醫(yī)學(xué);2011年06期

10 范濟(jì)輝;陳越;頊志兵;;中醫(yī)藥調(diào)節(jié)腫瘤免疫逃逸機(jī)制研究的進(jìn)展[J];山西中醫(yī)學(xué)院學(xué)報(bào);2013年01期

相關(guān)會(huì)議論文 前5條

1 宋英暉;崔蓮仙;何維;;封閉內(nèi)源性IL-15的表達(dá)可促進(jìn)MHCI類分子的表達(dá)[A];第七屆全國(guó)腫瘤生物治療學(xué)術(shù)會(huì)議論文集[C];2001年

2 韓巖梅;李和權(quán);張明剛;郭秋莉;鮑嫣;徐勝;陳朱波;曹雪濤;;髓系抑制性細(xì)胞促進(jìn)腫瘤免疫逃逸機(jī)制的初步研究[A];中國(guó)免疫學(xué)會(huì)第五屆全國(guó)代表大會(huì)暨學(xué)術(shù)會(huì)議論文摘要[C];2006年

3 李慧君;趙瑋t，

本文編號(hào)：2045424