肺癌之研究： I. 結合人類間葉幹細胞和蛋白質疫苗治療小鼠肉瘤細胞引起之肺臟轉移 II. 生物冷光造影活體監測人類肺癌細胞之骨轉移潛能

Lung cancer studies： I. Combination of human mesenchymal stem cells and fusion protein vaccine for tumor therapy in murine sarcoma-derived lung metastases model II. In vivo monitoring bone metastatic potential of human lung cancer cells by bioluminescen

魏鴻健(Hon-Jian Wei)

指導教授 ： 鄧文炳

臺北醫學大學/口腔學院/生物醫學材料研究所/碩士(2006年)

摘要

近年來，無論在全世界或台灣，癌症的發生率及死亡率都居高不下，而其中由肺癌所造成的癌症死亡仍然佔了很高的比例。傳統對於肺癌的治療，主要是以放射線治療、化學治療和手術切除為主，而治療後發生轉移以及預後不佳的問題，為致死的主要原因。而癌症轉移對於臨床上更是一個棘手的問題，臨床對於癌症轉移有偵測及診斷上的困難。骨骼為一個易受癌症轉移的組織，而由癌症所發生的骨轉移，會帶給病患很多痛苦，如骨痛、骨折或骨髓壓迫等，更甚者危及生命。目前在臨床上，分子造影技術已被廣泛的應用在許多疾病的診斷上，包括了癌症、神經性疾病、心血管疾病等。 本研究主要針對肺癌進行兩部分的實驗。第一部分是利用幹細胞具有活體趨癌的特性，以人類間葉幹細胞為媒介追蹤由小鼠肉瘤細胞引起之肺臟轉移，並結合蛋白質疫苗針對幹細胞趨癌所誘發體內免疫來抑制腫瘤之生長。經過實驗發現Kp-hMSCs的確能於活體內趨向肺臟腫瘤，而使用Kp-hMSCs和PE(ΔIII)-E7-KDEL3合併進行治療的老鼠，其肺部所產生的腫瘤結節數比對照組少 (P<0.05)，顯示結合此兩種方式的確可以對肺部腫瘤達到抑制的效果。第二部實驗以非侵入性的方式利用高敏感度及專一性的生物冷光造影，監測轉植luciferase基因的人類肺癌細胞，其於活體內轉移的情形。生物冷光影像顯示H441GL細胞存在於肺部以及後肢的位置，在經過組織切片分析的佐證後，可以證明我們所使用的H441GL肺癌細胞，可以經由循環系統轉移到骨頭中，表示的確具有骨轉移的能力。 結合幹細胞和蛋白質疫苗進行癌症治療，改善了疫苗療法只針對其獨特目標的限制，可提供未來臨床上對於不同癌症治療的應用性。而結合報導基因和報導探針策略的非侵入式的分子基因造影，由於其高專一性和敏感性的表現，可以準確的偵測癌症於活體內的生物活動，更可用來偵測初期癌症及微小的癌症轉移，基於專一性分子基因造影的積極發展，期望將來能廣泛應用於癌症診斷、治療，以及抗癌藥物的開發與測試。

關鍵字

肺癌 ； 分子基因造影 ； 蛋白質疫苗 ； 轉移 ； 第一型泡疹病毒胸腺嘧啶激? ； 螢火蟲生物冷光素

並列摘要

Recently, the incidence and mortality of cancer remains very high in the world, in which large proportion of cancer mortality is caused by lung cancer. Traditional treatments of cancer are radiotherapy, chemotherapy and surgery; however unfortunately, primary cancers often metastasize after treatment resulting in poor-prognosis. Metastases of the cancer have become a troublesome problem for clinic therapy and are hard to detect and to diagnose. Bone is a common tissue of several cancer metastases. When bone metastasis occurs, it always brings agonies to patients including bone pain, pathologic fracture, nerve compression, and also leads to death while being more malignant. Current molecular imaging technologies have been widely used for various diseases diagnosis on clinic, including cancer, neuropathic disease, and cardiovascular disease. In this study we focused on the lung cancer from two parts. For first aspect, previous studies have shown that the adoptively transferred stem cell have characteristics to target and to migrate to tumor lesion in vivo. Therefore, human mesenchymal stem cells (hMSCs) were used to target murine sarcoma-derived lung metastases and viewed as a “Trojan Horse”. Fusion protein vaccine was also used to recognize the hMSCs and then leaded to the inhibition of tumor growth. In our study, the Kp-hMSCs showed homing to lung tumor in vivo, and mice treated with Kp-hMSCs and PE(ΔIII)-E7-KDEL3 showed less lung nodules than the control group (p<0.05). The results show that combination of hMSCs and fusion protein vaccine can efficiently suppress the growth of lung tumor. In the second aspect, highly sensitive and specific bioluminescence imaging was used to non-invasively monitor the metastases of luciferase-transduced human lung cancer cell in vivo. The bioluminescence imaging results show that H441GL cells exist in lung and metastasize to hindlimb in later time point. After histological analyses, it demonstrates that H441GL lung cancer cells can metastasize to skeleton through the circulation and shows the bone metastatic potential. Combination of stem cells and protein vaccine for cancer therapy could improve the limitation of vaccine therapy that aim at only a unique organ. It provides an alternative clinical application of different cancer therapy in the future. Because of the reporter gene/reporter probe strategies used in highly specific and sensitive expressions of molecular gene imaging, it could accurately detect biological process of cancer in living object and could be used for diagnosis of cancer in early stage and microscopic tumor metastases. Base on the great development of specific molecular gene imaging, we expect that it can be facilitated widely application of cancer diagnosis, cancer therapy, and the development and test of the anticancer drug.

並列關鍵字

Lung cancer ； Molecular gene imaging ； Protein vaccine ； metastases ； Herpes simplex virus type 1 thymidine kinase ； Firefly luciferase

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