口腔癌係屬頭頸癌亞型，其惡性腫瘤好發於口腔區域，包括臉頰、舌頭、嘴唇及口底部，其中有超過九成之口腔癌病患癌細胞型態為鱗狀上皮細胞癌。近十年來，口腔癌病患術後之總體五年存活率並無顯著改善，不過，若於口腔癌早期發現病徵並及時予以治療之病患術後總體存活率皆有八成以上，其顯示出早期診斷對於提升口腔癌病患預後結果之必要性，而透過專一性生物標記物來進行早期診斷對於改善口腔癌病變之病患亦日益重要。 研究證明 microRNAs 表現失調與口腔癌病變有著密不可分的關係，而人體口腔內唾液也被認為是能夠直接映照出人體身體狀況與即時反映其口腔環境之生物標記物。 此研究中，我們發現唾液中 miR-X 及 miR-Y 的表現異常上升，不只與口腔癌病患癌症進程相關，亦與病患不良之臨床預後結果有關。其中，若給予 miR-X 或 miR-Y 類似物異位表現，會有效增加其口腔癌細胞株生長及爬行能力，係指出不論 miR-X 抑或 miR-Y 在口腔癌細胞株中皆扮演著 onco-miR 之角色。 為了研究 miR-X 與 miR-Y 於調節口腔癌病變之機制，我們首先以一系列 miR-target 預測系統去篩選 miR-X/-Y 可能與其進行標的作用之六個假定性標的基因。 TCGA 資料庫中亦顯示其六個預測標的基因與 miR-X/-Y 皆有負相關性表現之現象，然而在我們的實驗中發現唯有 PIK3C2B 之3’-UTR 不只具備 miR-X/-Y 之標的區域，其本身表現亦會被 miR-X/-Y 所影響並下調其 mRNA 表現。 PIK3C2B 不只扮演著腫瘤抑制者的角色，同時我們亦證明其 mRNA 表現量下調會與口腔癌癌症進程以及口腔癌病患較差臨床預後表現有關。 除此之外，於口腔癌細胞株中與先天性免疫相關之Toll-like receptors (TLRs) 活化不只會降低 miR-X/-Y 表現，亦會提升 PIK3C2B 之 mRNA 表現量。 雖然截至目前為止之實驗未能解釋其餘 microRNAs 對於口腔癌病患癌病變之影響，本篇研究中所發現之 miR-X 與 miR-Y 皆有機會作為具有潛力並有發展前瞻性之非侵略性唾液生物標記物，以用於偵測其具有高度惡化風險之口腔癌病患相關癌症病變進程。

Oral cancer, a subtype of head and neck cancer, is a malignant tumor that develops in the oral cavity, including cheeks, tongue, lips, and floor of the mouth. More than 90% of this cancer type is squamous cell carcinoma. The overall 5-year survival rate for oral cancer remains unchanged in the past few decades. Since the overall 5-year survival rate for early-stage oral cancer can be higher than 80%, early diagnosis of oral carcinogenesis through identification of specific biomarkers is thus essential for improving prognosis for oral cancer patients. Several studies have demonstrated a close relation of microRNAs deregulation with oral carcinogenesis, and that saliva can directly mirror systemic health and reflect oral environmental conditions. In this study, we found that the increase of two salivary miRs, miR-X and miR-Y, were not only associated with the progression of clinical staging but also reduced patient clinical outcome. Ectopic expression of miR-X or miR-Y mimics significantly increased oral cancer cell proliferation and migration, indicating that both functions as onco-miR in oral cancer cells. To identify the action mechanism whereby miR-X and/or miR-Y mediated oral carcinogenesis, we first used web-based miR-target prediction tools and identified six putative target genes for both miRs. Despite a negative correlation of the respective expression of six putative targets with that of miR-X or miR-Y in the TCGA database for head and neck cancer, we found only one of them, PIK3C2B bearing one miR-targeting site in the 3’-untranslated region, was the bona fide target for both miR-X and miR-Y. Consistent with being a tumor suppressor, PIK3C2B was down-regulated with disease progression and the reduction was also associated with the reduced clinical outcome. The activation of Toll-like receptors (TLRs), key players for innate immunity, repressed the inverse expression of miR-X/-Y with PIK3C2B in oral cancer cells. Although we cannot rule the involvement of additional miRs in oral carcinogenesis, the miRs identified in this study should hold great promise of being developed into noninvasive salivary biomarkers for detecting individuals at high risk of developing oral malignancy.

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