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標題: Melanoma antigen-A在犬正常及間質來源腫瘤組織的表現
Expression of Melanoma Antigen-A in Canine Normal and Mesenchymal Neoplastic Tissues
作者: 陳音竹
Chen, Yin-Chu
關鍵字: cancer/testis antigen;腫瘤睪丸抗原;MAGE-A;canine melanoma;黑色素細胞瘤抗原-A;狗黑色素細胞瘤
出版社: 獸醫學系暨研究所
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黑色素細胞瘤抗原-A (Melanoma antigen-A, MAGE-A)於1991年被發現,其包含12個相關抗原為MAGE-A1到MAGE-A12,這些抗原在人類組織中會少量且侷限的表現在正常組織中,如睪丸、胎盤、及胎兒性腺,在腫瘤方面,會被表現在多種腫瘤細胞上,此種抗原之表現特性適合作為免疫療法之標的抗原。然而,相關於狗MAGE-A的研究相當稀少,因此,本實驗目的為確立人類MAGE-A抗體6C1使用於犬組織的免疫組織化學染色流程、調查MAGE-A在犬正常組織及腫瘤組織之表現、以犬口腔黑色素瘤做一延伸,探討其表現與各種臨床特性、存活時間的相關性,並偵測MAGE-A亞型的表現。本實驗所用的抗體可偵測MAGE-A1, -A2, -A3, -A6, -A10, -A11, 及 -A12,在確立犬組織所適用的免疫組織化學染色流程後,總共偵測了18個正常組織、28個黑色素細胞瘤、32個口鼻腔惡性腫瘤(含口腔黑色素瘤)、40個圓形細胞瘤及37個軟組織肉瘤。此外,為了得知MAGE-A亞型在犬組織中的表現,聚合酵素鏈鎖反應中的引子可作用於MAGE-A基因中高度保留之區域,可藉由增幅出此高度保留片段,定序後經BLAST軟體比對,便可分型。免疫組織化學染色的結果顯示,所有正常組織包括皮膚、皮膚附屬器、齒齦、肌肉、脂肪組織、結締組織、唾液腺、淋巴結、小腸黏膜、乳腺、肝臟、軟骨、輸卵管、子宮內膜、大腦及小腦皆呈陰性,而睪丸的生殖細胞及卵巢的卵母細胞除外。於犬腫瘤中MAGE-A的表現會隨腫瘤類別不同而有差異,黑色素細胞瘤有75.00% (21/28) 的表現率,口鼻腔惡性腫瘤68.75% (22/32)的表現率,圓型細胞瘤52.50% (21/40)的表現率,軟組織肉瘤則有40.50% (15/37)的表現率。此外,分析一隻正常睪丸及九隻黑色素細胞瘤的犬隻所表現的MAGE-A亞型,正常睪丸的MAGE-A序列與人的MAGE-A2 (NM_005361)及狗的MAGE-A10 (XM_549339)最為相似,相似度分別為68.6%及91.4%,本實驗中十隻狗的MAGE-A序列彼此之相似度為71.7-95.3%。根據此抗原在犬正常組織中的侷限表現,且廣泛表現在多種腫瘤中的特性,以MAGE-A為標的抗原的免疫療法用來治療腫瘤已轉移或是腫瘤難以完全切除的犬隻,將有可能為一有治療潛力的輔助療法。

Melanoma antigen-A, MAGE-A, which contained 12 members from A1 to A12 was discovered since 1991. These antigens expressed in various types of tumors but limited expressed in normal human placentas and testicles. However, researches about MAGE-A in dogs were rare. The aims of this study were: first, to establish an immunohistochemistry protocol with MAGE-A antibody 6C1 for canine tissues. Second, to investigate the expression of MAGE-A in canine normal and neoplastic tissues. Third, concerning about canine melanoma, to investigate the expression of MAGE-A subtypes and the correlation between MAGE-A expression rate, clinical characteristics and surviving. Immunohistochemical staining with an antibody 6C1 against MAGE-A1, -A2, -A3, -A6, -A10, -A11, and -A12 was used to detect the expression of MAGE-A in 18 normal tissues, 26 melanomas, 5 nasal malignant tumors, 6 oral malignant tumors, 40 round cell tumors, and 37 soft tissue sarcomas. Besides, according to the highly conserved MAGE-A genes, a pair of primers were used to amplify the specific sequences of each subtype by PCR. The expression of each subtype was affirmed by DNA sequencing. According to our results, normal tissues including skin, skin adnexa, gingiva, muscle, adipose tissue, connective tissue, salivary gland, lymph node, intestine mucosa, mammary gland, liver, cartilage, oviduct, endometrium, cerebellum, and cerebrum showed the negative immunoreaction except for testicular germ cells and ovarian oocytes. The expression rate varied with the tumor type, 75.00% (21/28) of melanomas, 52.50% (21/40) of round cell tumors, and 40.50% (15/37) of soft tissue sarcomas showed the positive immunoreaction. Additionally, MAGE-A sequences from one normal testis and nine melanomas were analyzed. MAGE-A sequence from normal canine testis showed 68.6% identity with human MAGE-A2 (NM_005361) and 91.4% identity with canine MAGE-A10 (XM_549339). MAGE-A in canine testicle shared 71.7-95.3% of identities with canine melanoma sequences generated in this study. Based on its limited expression in canine normal tissues and widely expression in various types of canine mesenchymal tumors, MAGE-A might be used for antigen-specific cancer immunotherapy which was regarded as a promising treatment for metastasis cancer, or tumors which were difficult to be excised completely.
其他識別: U0005-1808201016462200
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