腫瘤胚胎基因（Oncofetal genes）是指在胚胎發育表現量較高之基因，而隨著年齡增加表現量也會隨之減少或不表現。當患有惡性腫瘤時，腫瘤胚胎基因會有再度表現之情形，在臨床上可以作為癌症診斷以及偵測復發之風險。為了找到新穎的腫瘤胚胎基因，實驗室在先前建立了一套生物資訊分析系統，利用表達序列標幟（EST）資料庫尋找可能為腫瘤胚胎之基因。由此生物資訊系統我們找到了29個已知基因以及44個未知基因，其中在這29個已知基因之中有5個基因已被證實為腫瘤胚胎基因。而實驗室也進一步去研究這44個未知基因，目前證實至少有3個基因為腫瘤胚胎基因，包括Lin28B、LRRC16B以及U41。此外Lin28B經實驗證實為一個良好的循環腫瘤胚胎幹細胞標記基因（Circulating oncofetal cancer-stem-cell-like marker），此基因在正常人週邊血並不表現，因此可用以偵測週邊血中的腫瘤幹細胞（Cancer stem cell），並且針對病患進行預後風險及復發機率之評估。接著我們發現這29個已知基因之中富含著Wnt訊息傳遞路徑之標靶基因（Wnt target gene），透過實驗證實未知基因U41為Wnt訊息傳遞路徑之標靶基因。接著實驗室進一步去設立生物資訊之篩選方法，希望能找到更多Wnt訊息傳遞路徑之標靶基因。經過生物資訊篩選我們找到了50個未知基因，實驗室先前已證實至少有2個基因為Wnt訊息傳遞路徑之標靶基因，包括RMI2以及ZNF496所以我們第一個實驗目的是希望在實驗室所建立尋找新穎oncofetal gene及Wnt target gene的生物資訊方法分別所找出的44個未知基因及50個未知基因中，尋找其他和Lin28B有相同特性且不會在周邊血表現並且有腫瘤胚胎幹細胞特性之新穎標記分子（oncofetal cancer stem cell marker）。利用RT-PCR去觀察這些基因在肝癌細胞株（HCC cell lines）、正常人週邊血（Normal peripheral blood）、誘導性多功能幹細胞（induced pluripotent stem cell）以及正常人各部位組織中的表現。我們發現有4個基因（SVOP、MDFI、MARCH4、BCAN）擁有腫瘤胚胎特性，且在正常人之週邊血中不會有表現。接著我們利用The Cancer Genome Altas（TCGA） data去探討這4個基因與癌症之間的關係，發現這4個基因的表現會造成許多不同癌症之死亡率有顯著的差異，代表這4個基因可能是值得研究的標誌基因。接著我們利用QRT-PCR做進一步的研究，發現SVOP、MDFI、MARCH4以及BCAN在周邊血會有微量的表現；在肝癌腫瘤-非腫瘤組織配對的檢體中，在腫瘤組織以及非腫瘤組織的表現量並沒有顯著的差異。這些結果不如我們的預期，因此這4個基因無法做為偵測循環肝癌腫瘤幹細胞（Circulating HCC cancer stem cells）之腫瘤胚胎標記基因，但我們推測這4個基因在其他癌症中還是具有研究的潛力。第二個實驗目的是要繼續研究先前實驗室利用生物資訊方式並且經過實驗證實為新的Wnt訊息傳遞路徑之標靶基因－Zinc Finger Protein 496（ZNF496）。在先前的研究指出在許多癌症中，常發現Wnt訊息傳遞路徑不正常的活化，其中也包括肝癌，而肝癌為全球最常見的惡性腫瘤之一。我們的目的是為了研究ZNF496在肝癌細胞中所扮演的角色及功能，在先前的實驗結果顯示過表達ZNF496會促使幹細胞特性、上皮－間質轉化（EMT）及細胞增生。在我的研究結果發現，過表達ZNF496會促使參與在細胞週期（cell cycle）中的基因表達量上升、細胞轉移性（cell migration）及侵襲性（cell invasion）增加。

Oncofetal genes typically are present only during fetal development, down-regulated in adults, but re-expressed in certain types of cancers. Our laboratory performed bioinformatics analyses using expressed sequence tags (EST) database and found 29 known genes and 44 unknown genes. We further studied the 44 then unknown genes and discovered the novel oncofetal gene Lin28B. We used Lin28B as a marker to detect circulating tumor cells and were able to split early stage HCC into 2 recurrence curves, in which patients with detectable Lin28B positive cells in circulation behaved more like late stage HCC. Our first aim in this study is to find novel oncofetal cancer stem cell markers other than Lin28B which are not expressed in the normal peripheral blood cells. Four genes (SVOP, MDFI, MARCH4, and BCAN) emerged which showed good oncofetal patterns and were not expressed in normal peripheral blood cells as tested by RT-PCR. When we used QRT-PCR for further investigation, it was found that these four genes were expressed in normal peripheral blood cells at high-enough background levels. These results were not measuring up to our criteria as oncofetal markers. We modified the bioinformatics algorithms in hope to enrich WNT target genes. The algorithms resulted in 50 unknown genes, we found ZNF496 is a novel WNT target gene. The second aim of this study is to continue the study of Zinc Finger Protein 496 (ZNF496). Previous studies indicated that ZNF496 can promote EMT, stemness, and cell proliferation. In my studies, we found ZNF496 can promote cell migration and invasion.

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