【摘要】：背景:全球每年肺癌的發(fā)病率居首位,同時也是第一位常見的腫瘤死亡原因。迄今為止,手術(shù)切除被認為是治療肺癌的主要手段,然而,大部分的肺癌病人就診時已屬于進展期,單純手術(shù)療效有限,因此化療在肺癌治療中占有極其重要的地位�；熞驯蛔C實可以提高治療效果、延長生存時間。紫杉醇(Paclitaxel,Ptx),作為化療領(lǐng)域中里程碑式的藥物,在肺癌治療中也作為一線方案的重要組成部分被列入NCCN肺癌臨床實踐指南中。盡管如此,但其單藥或者聯(lián)合化療的有效率均小于50%。此外,中藥單體小分子化合物漢防己甲素(Tetrandrine,Tet)也在我們前期研究中被證實具有潛在抗腫瘤效應(yīng)。因此如何減少化療藥物的毒副作用且能夠發(fā)現(xiàn)新型的潛在藥物來增加化療效果成為抗腫瘤藥物研發(fā)中的一個非常重要的問題。納米載藥體系不僅具有改善水溶性較差藥物的水相溶解度,增加其穩(wěn)定性,從而獲得控制釋放等傳統(tǒng)藥物輸送體系所具有的作用外,由于應(yīng)用了不同的設(shè)計理念,在抗腫瘤藥物輸送中又具有許多獨特的應(yīng)用優(yōu)勢。目的:本研究構(gòu)建了兩種不同的納米載藥體系。首先采用自行合成的聚吡咯烷酮-聚己內(nèi)脂(PVP-PCL)兩親嵌段共聚物,用納米沉淀法制備了漢防己甲素載藥納米微球(Tet-NPs)。同時,我們基于Carriar fre的概念合成了Ptx-SA小分子化合物,通過其自組裝構(gòu)建了負載紫杉醇的Ptx-SA載藥納米纖維。在隨后的研究中考察了兩種載藥納米體系的穩(wěn)定性、緩釋效應(yīng)以及體外與體內(nèi)協(xié)同抑瘤活性來考察載藥納米體系在肺癌治療中的優(yōu)勢,為這兩種新型的納米給藥體系的繼續(xù)研發(fā)提供實踐基礎(chǔ)。方法:用化學(xué)合成法合成了 PVP-PCL二嵌段共聚物以及Ptx-SA小分子化合物。用原子力顯微鏡和透射電子顯微鏡觀察兩種載藥體系的形貌,并用動態(tài)光散射儀和Zeta電位測定儀檢測載藥納米體系的粒徑大小和表面電位。同時采用高效液相色譜(HPLC)測定載藥納米體系的載藥效率,并且考察兩種載藥體系的體外緩釋特性。在體外細胞及體內(nèi)動物模型上研究載藥納米微球的抗腫瘤效果及其可能的機制。結(jié)果:第一,本文中所構(gòu)建的Tet-NPs在掃描電子顯微鏡和透射電子顯微鏡的觀察下呈現(xiàn)為較規(guī)則的表面光滑的球形,粒徑比較均一,在120nm左右。其載藥量為18.2%左右。Tet-NPs具有良好的穩(wěn)定性以及體外緩釋特征。且?guī)в袩晒馐聚櫟腡et-NPs能夠在與細胞共孵育的短時間內(nèi)有效的進入細胞。Tet-NPs的誘導(dǎo)凋亡的效果顯著強于Tet裸藥。同等劑量下Tet-NPs比Tet裸藥能夠更為有效的調(diào)控凋亡相關(guān)蛋白的表達。此外,Tet-NPs比同等劑量下的裸藥能夠更為顯著抑制肺癌細胞的侵襲和轉(zhuǎn)移,其機制可能為通過調(diào)控MMP相關(guān)蛋白的表達。第二,Ptx-SA在掃描電子顯微鏡和透射電子顯微鏡的觀察下呈現(xiàn)為直徑為30-40nm,長約1-3um的纖維狀結(jié)構(gòu)。隨后的體外釋放實驗證實了該載藥納米纖維良好的體外緩釋性。細胞攝取實驗發(fā)現(xiàn)Ptx-SA載藥纖維具有良好的親細胞性。Ptx-SA與Ptx裸藥所顯示的細胞毒效應(yīng)的效果都具有濃度依賴性,且Ptx-SA的效果強于Ptx裸藥。Western blot結(jié)果顯示同等劑量下Ptx-SA比Ptx裸藥能夠更為有效的調(diào)控Akt蛋白及凋亡相關(guān)蛋白的表達。在體內(nèi)肺癌模型上也證實了Ptx-SA比Ptx裸藥更為有效的抑瘤效果。結(jié)論:本文中所構(gòu)建的兩種載藥納米體系在體外以及體內(nèi)均顯示出了良好的抗腫瘤效果,并且在體內(nèi)外的療效評價中證實了相比于裸藥的優(yōu)越性�？梢�,制備新型的納米載藥體系,是抗腫瘤藥物發(fā)展的新方向,具有良好的實用價值和發(fā)展前景。
[Abstract]:Background: The incidence of lung cancer in the world is the first and the first common cause of tumor death. So far, surgical resection is thought to be the main means to treat lung cancer. However, most of the patients with lung cancer have been in the stage of progression and the simple operation is limited, so the chemotherapy plays a very important role in the treatment of lung cancer. The chemotherapy has been proved to improve the treatment effect and prolong the survival time. Paclitaxel (Ptx), as a landmark drug in the field of chemotherapy, is also included in the NCCN lung cancer clinical practice guide as an important part of the first-line protocol in the treatment of lung cancer. Nevertheless, the effective rate of either single or combination chemotherapy is less than 50%. In addition, Tetrandrine (Tet), a small-molecule compound of the traditional Chinese medicine, is also proved to have a potential anti-tumor effect in our earlier studies. Therefore, how to reduce the toxic and side effect of the chemotherapy medicine and to find the new potential medicine to increase the effect of the chemotherapy becomes a very important problem in the research and development of the anti-tumor medicine. The nano-carrier system not only has the functions of improving the water phase solubility of the water-soluble poor drug, increasing the stability of the water phase, and thus obtaining the traditional drug delivery system such as controlled release, and has a plurality of unique application advantages in the anti-tumor drug delivery. Objective: In this study, two different nano-carrier systems were constructed. The nano-microball (Tet-NPs) of tetrandrine (Tetrandrine) was prepared by using the self-synthetic polydioxanone-polycaprolactone (PVP-PCL). At the same time, we synthesized the Ptx-SA small molecule compound based on the concept of Carriar fre, and the Ptx-SA drug-loaded nano-fiber loaded with paclitaxel was constructed by self-assembly. In the following study, the stability, sustained-release effect of the two drug-containing nano-systems, and the in vitro and in vivo co-inhibition of the tumor-inhibiting activity were investigated in order to study the advantages of the drug-containing nano-system in the treatment of lung cancer, and provide the practical basis for the continuation and development of the two new drug delivery systems. Methods: PVP-PCL diblock copolymer and Ptx-SA small molecular compound were synthesized by chemical synthesis. The morphology of two drug-carrying systems was observed by atomic force microscope and transmission electron microscope, and the particle size and surface potential of the drug-containing nano-system were detected by dynamic light-scattering instrument and Zeta potential tester. The drug-carrying efficiency of the drug-containing nano-system was determined by high performance liquid chromatography (HPLC), and the in vitro release characteristics of the two drug-carrying systems were investigated. The anti-tumor effect and possible mechanism of the drug-loaded nanospheres were studied in vitro and in vivo animal models. Results: First, the Tet-NPs constructed in this paper showed a smooth spherical surface with a uniform particle size of about 120nm under the observation of a scanning electron microscope and a transmission electron microscope. The drug loading is about 18. 2%. Tet-NPs have good stability and in vitro slow-release characteristics. and a Tet-NPs with a fluorescent trace can effectively enter the cells within a short period of time with the cell co-incubation. The effect of Tet-NPs on the induction of apoptosis was stronger than that of Tet. Tet-NPs at the same dose can be used to control the expression of apoptosis-related protein more effectively than the Tet naked drug. In addition, Tet-NPs can significantly inhibit the invasion and metastasis of lung cancer cells than those of the same dose, and the mechanism may be to regulate the expression of MMP-related proteins. Second, Ptx-SA is present in a fibrous structure having a diameter of 30-40nm and a length of about 1-3um under the observation of a scanning electron microscope and a transmission electron microscope. The subsequent in vitro release experiment confirmed the good in vitro release of the drug-containing nanofibres. The cell uptake experiments showed that the Ptx-SA drug-containing fiber had good cell affinity. The effect of Ptx-SA with the cytotoxic effect shown by the Ptx naked drug has a concentration dependence, and the effect of the Ptx-SA is stronger than that of the Ptx naked drug. Western blot showed that Ptx-SA could more effectively control the expression of Akt protein and apoptosis-related protein at the same dose. In vivo lung cancer model, a more effective tumor-inhibiting effect of Ptx-SA than Ptx bare drug was also confirmed. Conclusion: The two drug-loading nano-systems constructed in this paper show good anti-tumor effect in vitro and in vivo. It can be seen that the preparation of a novel nano-carrier system is a new direction for the development of anti-tumor drugs, and has good practical value and development prospect.
【學(xué)位授予單位】：南京醫(yī)科大學(xué)
【學(xué)位級別】：博士
【學(xué)位授予年份】：2017
【分類號】：R734.2

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相關(guān)期刊論文 前2條

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本文編號：2335514