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標題: 建立老鼠腦內黑色素瘤模型與其免疫治療之應用
Establishment of the Intracranial Melanoma Mice Model and Immunotherapeutic Application
作者: 林宛頻
Lin, Wan-Pin
關鍵字: intracranial melanoma mice model;老鼠腦內黑色素瘤;dendritic cells immunotherapy therapy;imiquimod cream;樹突細胞免疫治療;樂得美乳膏
出版社: 獸醫學系暨研究所
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惡性神經膠質瘤是最常見的原發性腦瘤,約佔所有腦瘤的40-50 %,其死亡率極高。而目前臨床腦瘤的治療仍以侵入性的外科手術為第一選擇,再輔以術後的放射線治療及化學治療(Temodal),或是加碼刀放射性手術(γ-knife radiosurgery)。但因腫瘤極易擴散且常發生於腦內的功能區上,使得手術時切除困難,其預後不良又極易容易復發,所以發展新的治療方向是重要的,期能更精確的針對腫瘤細胞進行治療將傷害降到最低。而因發現時期的不同,病人的存活期平均短則數星期、長則一年不等。在腫瘤的微環境(Microenvironment)中,目前已證實腫瘤細胞會分泌一些細胞激素,如介白素10(Interleukin-10)、轉化生長因子(Transforming growth factor-β)等抑制病人的免疫力。透過樹突疫苗注射後,可使干擾素(Interferon-γ)大量分泌,活化毒殺性T細胞,已知可用來治療周邊的惡性黑色素瘤。而常見的神經膠質瘤中,其星狀細胞癌(Astrocytoma)與黑色素細胞癌皆由神經外胚層所發展而來的,且小鼠皮膚黑色素細胞癌與人類膠質瘤均會表現共同的胜肰gp100與酪氨酸酶相關蛋白2(Tyrosinase-related protein 2, TRP-2),加上臨床上用於皮膚表面塗抹治療尖銳濕疣患者的樂得美乳膏(Imiquimod cream),可以透過類鐸受體-7 (Toll-like receptor 7, TLR-7)來活化刺激患者本身的免疫發炎反應,已成功用於治療皮膚的腫瘤,但未有相關文獻證實可用於治療其它部位的腫瘤。本實驗希望藉由建立成功的小鼠腦內黑色素瘤模式後,進一步模擬腦內膠質瘤免疫治療的可行性,大膽假設周邊低侵入性的注射經腫瘤抗原激活的樹突細胞(Dendritic cell)與周邊淋巴附近皮膚塗抹Imiquimod,可以藉由樹突細胞抗原呈現之特性調節身體的免疫反應,激活自身淋巴系統中之T細胞並認識腫瘤,並進一步減緩腦內腫瘤增生或甚至消滅腫瘤,達到人體治療的舒適性與延長存活的目標。實驗結果發現於顱內注射點(以Bregma為原點,往後1.04 ± 0.06 mm、往右2.10 ± 0.18 mm、深度2.79 ± 0.48 mm),注射1000顆/2μl的黑色素瘤細胞可以100%成功建立小鼠顱內黑色素瘤的模型。在以gp100-DC或imiquimod周邊的治療方式,雖不能完全治癒或避免小鼠的死亡,但確實可以延長小鼠的存活天數1至2天,而以抗原呈現樹突細胞和imiquimod合併治療時,動物的個體療效差異卻很大。腦內的腫瘤於周邊的gp100-DC或imiquimod治療後,腫瘤冷光影像有減小的趨勢,在H&E染色下可明顯見到腦內黑色素瘤與正常組織交界處的炎症細胞(Glial cell)浸潤較多,且腫瘤的有絲分裂較不明顯。在脾臟的共軛焦免疫螢光雙重染色的觀察中,發現第14天可見治療後有不等程度的活化態巨噬細胞(CD163與CD63陽性)增加,帶有CD63(melanoma相關抗原ME491)陽性的CD4輔助性T細胞與CD8毒殺性T細胞數量表現也有不同程度的調節。雖然在腦內的染色尚未見到這些細胞明顯的表現,但可以確信的是採行周邊樹突細胞的免疫治療確實可以活化體內自身的免疫細胞,也間接的證實免疫治療的方式可能也改變了腦內的微環境進而影響腫瘤的大小。未來進一步在腦內免疫細胞與微環境改變的研究,以及免疫適期的調整配合下,多點注射經腫瘤抗原激活的樹突細胞與其他免疫調節藥物的併用治療,將會是治療腦內黑色素瘤或腦內膠質瘤的一大利器。
其他識別: U0005-2306201120173800
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