醣科學於大腸直腸癌的角色

Abstract

異常的細胞表面分子醣化現象跟細胞的惡性變化有很大的關連。腫瘤關連抗原， T 抗原 (Galbeta1, 3GalNAc) ，是一個癌胎兒碳水化合物抗原，在許多癌症上有表現。而 T抗原的表現會增加癌症轉移的機會，同時也和大腸直腸癌的不良預後相關。因此，T抗原可能可以是癌症疫苗治療的標的之一。然而，對於醣基因是怎樣調控T抗原的合成與表現，以及這些醣基因的功能，卻鮮少被人了解。為了瞭解在大腸直腸癌中，那些醣基因調控 T抗原的表現，我們首先從醣基轉移蛋白質的角度來了解。我們發現醣基轉移酵素beta1,4-N-acetylgalactosaminyltransferase III (beta4GalNAc-T3) 在生體外，可以合成 N,N'-diacetyllactosediamine, GalNAcbeta1,4GlcNAc。因為，我們研究beta4GalNAc-T3 在大腸直腸癌的表現，以及這樣的表現對 HCT116 大腸直腸癌細胞株的影響。 Real-time reverse transcription-PCR 的結果顯示， beta4GalNAc-T3 在 72.5% (n = 40) 的大腸直腸癌檢體中的表現量，相較於正常黏膜組織是增加的。beta4GalNAc-T3過度表現會增加細胞與細胞外間質的黏附，移行，無定錨細胞生長，以及大腸直腸癌細胞的侵襲。此外， beta4GalNAc-T3的過度表現還會增加裸鼠生體內腫瘤的生長以及轉移，並且減少裸鼠的存活。 beta4GalNAc-T3 的過度表現會增加局部黏附磷酸酶的酪胺酸磷酸化。同時也會增加樁蛋白質的磷酸化。這樣的結果顯示，過度表現的beta4GalNAc-T3 可能在癌症惡性化過程中扮演著重要的角色，而這樣的影響是經由整合素以及分裂素活化磷酸酶訊息傳導路徑而來。 在另外一個層次， Cosmc 是一個伴侶蛋白質分子，是T合成素形成，活化過程中所需要的蛋白質分子之一。所以可以催化T抗原的生成。然而，在大腸直腸癌中，過度表現的 Cosmc會造成怎樣的變化還不是很清楚。我們發現，利用 real-time PCR 顯示，過度表現的 Cosmc mRNA 跟腫瘤的期數有著正相關。在HCT116大腸直腸癌細胞株中，過度表現的可以顯著增加HCT116大腸直腸癌細胞株的T抗原表現。同時，也會增加細胞生長，移行，以及侵襲能力。這些都跟局部黏附磷酸酶，ERK， 以及Akt的磷酸化增加有關。我們同時發現 Cosmc 的過度表現可以增加生體內腫瘤生長以及減少SCID老鼠的存活。整體來看，這樣的結果顯示，Cosmc的過度表現會增加大腸直腸癌細胞的惡性表現，這樣的改變是經過 MEK/ERK以及 PI3K/Akt 的活化訊息傳導路徑。所以 Cosmc可能可以作為大腸直腸癌治療上的標的及指標。

Aberrant glycosylation of cell surface molecules is associated with malignant transformation. Tumor-associated T antigen (Galbeta1, 3GalNAc) is an oncofetal carbohydrate antigen and expression of T antigen is associated with enhanced metastatic potential and poor prognosis in colorectal cancer。Therefore T antigen could be a potential target for vaccine of many cancers. However, the glycogenes responsible for regulating T antigen expression and their biological functions remain largely unknown. To identify the glycogenes that can regulate T antigen expression in colorectal adenocarcinomas. We found that The enzyme beta1,4-N-acetylgalactosaminyltransferase III (beta4GalNAc-T3) exhibits in vitro activity of synthesizing N,N'-diacetyllactosediamine, GalNAcbeta1,4GlcNAc. Here, we investigate the expression of beta4GalNAc-T3 in primary colon tumors and the effects of its overexpression on HCT116 colon cancer cells. Real-time reverse transcription-PCR showed that the expression of beta4GalNAc-T3 was up-regulated in 72.5% (n = 40) of primary colon tumors compared with their normal counterparts. beta4GalNAc-T3 overexpression resulted in enhanced cell-extracellular matrix adhesion, migration, anchorage-independent cell growth, and invasion of colon cancer cells. Moreover, beta4GalNAc-T3 overexpression increased tumor growth and metastasis and decreased survival of tumor-bearing nude mice. beta4GalNAc-T3 overexpression showed increased tyrosine phosphorylation of focal adhesion kinase and paxillin Y118 as well as increased extracellular signal-regulated kinase phosphorylation. These results suggest that up-regulation of beta4GalNAc-T3 may play a critical role in promoting tumor malignancy and that integrin and mitogen-activated protein kinase signaling pathways could be involved in the underlying mechanism. In another level, Cosmc is a molecular chaperone required for the formation of an active T-synthase, which catalyzes the synthesis of T antigen. However, the expression and role of Cosmc in colorectal cancer are still unclear. Here, real-time PCR showed that overexpression of Cosmc mRNA in colorectal tumors compared with paired non-tumorous tissues was associated with increased American Joint Committee on Cancer (AJCC) tumor stage. Forced expression of Cosmc in HCT116 cells significantly increased T antigen expression and enhanced cell growth, migration, and invasion, which was associated with increased phosphorylation of focal adhesion kinase (FAK), ERK, and Akt. These Cosmc-enhanced malignant phenotypes were significantly suppressed by specific inhibitor of MEK or PI3K. We also found that Cosmc overexpression increased tumor growth and decreased survival of tumor-bearing SCID mice. Taken together, these results suggest that Cosmc promotes malignant phenotypes of colon cancer cells mainly via activation of MEK/ERK and PI3K/Akt signaling pathways, and that Cosmc may serve as a potential target for colorectal cancer treatment.