卵巢癌是一種致命的婦科癌症，由於卵巢癌早期診斷的困難，因此患者被發現罹癌時通常已是癌症晚期，而造成治療上的難題，並且伴隨著預後不佳以及存活率較差的情況。根據實驗室先前的研究發現，在高分化級別漿液性卵巢癌 (High grade serous ovarian cancer, HGSOC) 中delta-like 1 homolog (DLK1) 的表現量會較高，而藉由基因工程knockdown DLK1後，會降低HGSC的惡性程度，因此指出DLK1具有做為早期卵巢癌診斷的生物指標。而實驗室先前利用phage display技術篩選出四支DLK1 single-chain fragment variable (scFv) 抗體，作為發展治療卵巢癌的目標。在此研究中，我們將scFv anti-DLK1片段克隆到pET-28a，生產四支scFv DLK1，抗體，分別作用在高分化級別漿液性卵巢癌細胞SKOV3及低度分化漿液性卵巢癌細胞MPSC1上，發現scFv anti-DLK1#1，具有最佳抑制SKOV3和MPSC1的侵略能力和菌落形成能力，接著挑選scFv anti-DLK1 #1和#2克隆到帶有全人類抗體骨架的載體中，利用Expi293細胞生產、製作成全人類IgG抗體，而發現anti-DLK1 #1和#2具有抑制SKOV3細胞侵略、菌落形成和自我新生的能力，且也使用兩支DLK1抗體開發專用的ELISA檢驗組，根據實驗結果顯示，希望未來將此抗體應用於診斷及治療卵巢癌。

Ovarian cancer is one of the most lethal gynecological malignancies. Due to the difficult detected in early stage. Usually, when patients were diagnosed at late stage in ovarian cancer, leading to unfavorable prognosis and poor survival rate. According to our previous research, delta-like 1 homolog (DLK1) has high expression in high-grade serous ovarian cancer (HGSOC) and with gene knockdown DLK1 has reduced malignant in HGSOC. Therefore, DLK1 can be a potential therapeutic target in ovarian cancer. Our previous phage display screening have identified four single-chain fragment variable (scFv) anti-DLK1 with phage display technique, that as an object developed to therapeutic ovarian cancer. We subcloned scFv anti-DLK1 fragments into pET-28a vector then purified 4 different scFv anit-DLK1 proteins, which were employed for treatment in high-grade serous ovarian cancer and low-grade serous ovarian cancer are SKOV3 and MPSC1, respectively. The scFv anti-DLK1#1 was most effective in inhibiting the invasion and colony formation of SKOV3 and MPSC1 cells. Subsequently, scFv anti-DLK1 #1 and #2 were sucloned into human IgG vectors then tranfected into Expi293 cells for generation of fully human IgG antibody. The anti-DLK1#1 and #2 remained capable of inhibitingthe invasion, colony formation and self-renewal in SKOV3 cells. The two fully human anti-DLK1 were develop ELISA assay kit. These findings suggest that fully humanized anti-DLK1 may be applicable to the diagnosis and treatment of ovarian cancer.

INDEX
論文審定書 i
致謝 ii
中文摘要 iii
ABSTRACT iv
FIGURES AND LEGENDS INDEX vi
INTRODUCTION 1
MATERIALS AND METHODS 7
RESULTS 15
DISCUSSION 20
FIGURES AND LEGENDS 23
APPENDIX 54
REFERENCES 56

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