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研究生： 趙鈺瑛
Chao, Yu-Ying
論文名稱： 初級纖毛降低胰臟導管腺癌對於順一雙氨雙氯鉑的化療敏感性
Primary cilia alleviate the chemosensitivity of pancreatic ductal adenocarcinoma to cisplatin
指導教授： 王家義
Wang, Chia-Yih
學位類別： 碩士
Master
系所名稱： 醫學院 - 細胞生物與解剖學研究所
Institute of Cell Biology and Anatomy
論文出版年： 2020
畢業學年度： 108
語文別： 英文
論文頁數： 43
中文關鍵詞： 化療敏感度 、初級纖毛 、中心粒衛星 、DNA損傷反應
外文關鍵詞： Chemosensitivity, primary cilia, centriolar satellite, ATM, ATR
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    • 胰臟導管腺癌是是最致命的癌症之一，胰臟導管腺癌的臨床治療效果較差主要是因為其診斷發現的時間通常已經到達晚期以及對化療的耐藥性。因此，增加胰臟導管腺癌對化療的敏感度對於提升存活率是很重要的議題。初級纖毛廣泛存在於大部分人體細胞以維持發育以及分化的過程。中心粒衛星是散佈在中心體周圍的帶電胞器，參與調控初級纖毛的生成。在初級纖毛生成的過程中，動力蛋白/動力蛋白介導的中心粒衛星會向著中心體的基部聚集，這一過程被稱為中心粒衛星重塑，並維持初級纖毛的形成。在腫瘤發生過程中，胰腺導管細胞的初級纖毛會逐漸減少，代表胰臟導管腺癌細胞會有初級纖毛缺失的現象。有趣的是，具有化療抗藥性的癌細胞卻可以使初級纖毛再生，代表著這些纖毛可能部分地有助於發展癌症的幹細胞特性或是抗藥性，也意味著初級纖毛對腫瘤的發生和化療抗藥性扮演重要的角色。在我們的研究中，我們發現當胰臟導管腺癌經過順鉑處理後，會誘導中心粒衛星重塑以生成初級纖毛。我們透過抑制初級纖毛提高了胰臟導管腺癌對順鉑的化療敏感性，這也代表順鉑誘導的初級纖毛會降低胰臟導管腺癌的化療敏感性。除此之外，我們也發現動力蛋白/動力蛋白運輸的中斷或中心粒衛星重要蛋白PCM1的耗竭會抑制初級纖毛的形成，這證明了了中心粒衛星重塑在順鉑誘導的初級纖毛形成中扮演相當重要的角色。此外，我們也探討參與其中的分子機制。當順鉑處理後，會誘發ATM、ATR與DNA-PK蛋白激酶的活化。但是，只有ATM和ATR的失活或是降解才會抑制順鉑誘導的中心粒衛星重塑和初級纖毛生成。因此，我們的研究發現了初級纖毛對於化學敏感性的調控中扮演重要的角色，並揭示了參與其中的分子機制。

    Pancreatic ductal adenocarcinomas (PDAC) is one of the most lethal cancers. The poor clinical outcome of PDAC is mainly due to late diagnosis and chemotherapeutic resistance. Thus, accelerating chemosensitivity of PDAC is an important issue to improve clinical survival rate. Primary cilia grow in most cells of human body for maintaining development and differentiation. Centriolar satellites, electron dense granules scatter around the centrosome, regulate primary cilia formation. During the growth of primary cilia, dynein/dynactin-mediated centriolar satellites cluster toward the base of centrosome, a process known as centriolar satellite remodeling, for maintaining primary cilia formation. Gradually reduced primary cilia of pancreatic ductal cells during progression of tumorigenesis suggesting PDAC cells are devoid of cilia. Interestingly, some chemoresistant cancer cells can regrow primary cilia, and these cilia, at least in part, contribute to cancer stemness and chemoresistance, implying primary cilia has distinct effects on tumorigenesis and chemoresistance. Here we showed that PDAC could grow primary cilia via centriolar satellite remodeling upon cisplatin treatment. Genetic and pharmacological inhibition of primary cilia improved chemosensitivity of PDAC to cisplatin, suggesting cisplatin-induced primary cilia reduced chemosensitivity of PDAC. Disruption of dynein/dynactin transportation or depletion of centriolar satellite scaffold PCM1 reduced cilia formation supporting the important role of centriolar satellite remodeling in cisplatin-induced primary cilia formation. Then, the molecular signaling were examined. Ataxia telangiectasia mutated (ATM), ATM and RAD3-related (ATR), and DNA-dependent protein kinase were activated by cisplatin treatment; however, only depletion or inactivation of ATM and ATR alleviated cisplatin-induced centriolar satellite remodeling and growth of primary cilia. Thus, our data provide a novel role of primary cilia in chemosensitivity and uncover the underlying molecular mechanism.

    論文考試合格證明書 I 中文摘要 II Abstract III Contents V Abbreviations VII Introduction 1 A. Pancreatic cancer: symptoms and therapeutic strategy 1 B. Primary cilia 2 C. DNA damage response 4 Material and Method 6 Results 9 A. Cisplatin induces primary cilia formation in PANC-1 cells. 9 B. Cisplatin promotes centriolar satellites remodeling in PANC-1 cells. 11 C. Cisplatin induces primary cilia formation and centriolar satellites remodeling via ATM/ATR signaling in cisplatin-insensitive cells. 12 Discussion 15 Reference 17 Figures 21 Fig. 1. Cisplatin induces ciliogenesis in PANC-1 cells. 21 Fig. 2. Cisplatin-induced primary cilia contain intact ciliary components. 23 Fig. 3. CP110 is removed from basal body upon primary cilia formation. 24 Fig. 4. Depletion of IFT88 inhibits CPT-induced primary cilia formation. 25 Fig. 5. Depletion of ciliary protein, IFT88, sensitizes cells to cisplatin. 26 Fig. 6. Roscovitine inhibits cisplatin-induced primary cilia. 27 Fig. 7. Roscovitine enhances cisplatin chemosensitivity. 28 Fig. 8. Cisplatin promotes centriolar satellites remodeling in PANC-1 cells. 29 Fig. 9. Depletion of centriolar satellite scaffold protein, PCM1, inhibits CPT-induced ciliogenesis. 30 Fig. 10. Sodium orthovanadate inhibits CPT-induced centriolar satellites remodeling and further suppress CPT-induced primary cilia. 31 Fig. 11. Sodium orthovanadate enhances cisplatin chemosensitivity. 32 Fig. 12. Depletion of p150glued inhibits CPT-induced centriolar satellites remodeling and suppress CPT-induced ciliogenesis. 34 Fig. 13. Cisplatin induces DNA damage response. 35 Fig. 14. Cisplatin induces DNA damage response via ATM, ATR and DNA-PK activation. 36 Fig. 15. ATM and ATR induce primary cilia formation upon cisplatin treatment. 37 Fig. 16. Depletion of ATM or ATR inhibit CPT-induced centriolar satellites remodeling. 39 Fig. 17. Cisplatin activates DNA damage response effectors, including AKT, p53, CHEK1 and CHEK2. 40 Fig. 18. CHEK1 and p53 do not participate in CPT-induced primary cilia formation. 41 Fig. 19. DNA damage response effectors do not participate in CPT-induced primary cilia formation. 42 Fig. 20. A summary model shows that cisplatin activates ATM/ATR activation induces ciliogenesis via centriolar satellites remodeling, further regulates cisplatin chemosensitivity. 43

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