YB-1促進蕾莎瓦抗藥性肝癌細胞之表皮細胞間質化

Abstract

肝癌在全世界中是一常見的癌症型態。在疾病晚期的案例中，蕾莎瓦被認為是一個較好的治療選擇，然而，蕾莎瓦的抗藥性對於臨床的有效應用，仍然是一重要的阻礙原因。根據我們先前實驗室所建立的定量磷酸化蛋白體學結果，並結合目前癌症基因體圖譜資料分析，我們找到一個轉錄因子，YB-1，其絲胺酸102磷酸化位點在蕾莎瓦抗藥性的肝癌細胞 (HuH-7R) 中具有顯著上升的表達量。隨著蕾莎瓦的治療，發現會活化PI3K和AKT訊息傳遞途徑，並增加YB-1的磷酸化表現量。在功能性分析中，抑制YB-1的表現會減少細胞的爬行和侵襲能力。在分子層次，抑制YB-1進而也會抑制Snail, Twist1, Zeb1, MMP-2 和 Vimentin的表現量，這些結果都顯示著YB-1在HuH-7R細胞之表皮細胞間質轉化過程的可能重要角色。除此之外，YB-1會透過Cdc42途徑的活化，影響F-actin細胞骨架的重新排列，並導致HuH-7R細胞型態外觀上的改變。致突變實驗分析中，我們在HuH-7R細胞中將YB-1的絲胺酸102磷酸化位點進行點突變，發現其將抑制細胞的爬行及侵襲能力。綜合以上的研究結果，在HuH-7R細胞中我們發現蕾莎瓦活化了EGFR/PI3K/AKT途徑，進而促進YB-1的磷酸化並增強了HCC的轉移能力。對於這條特定的致病機制有更深入的了解，將有助於我們去研發新的抗癌藥物及新穎的肝癌治療策略以克服抗藥性等問題。

Hepatocellular carcinoma is one of the most common cancer types worldwide. In cases of advanced-stage disease, sorafenib is considered the treatment of choice. However, resistance to sorafenib remains a major obstacle for effective clinical application. Based on integrated phosphoproteomic and The Cancer Genome Atlas (TCGA) data, we identified a transcription factor, Y-box binding protein-1 (YB-1), with elevated phosphorylation of Ser102 in sorafenib-resistant HuH-7R cells. Phosphoinositide-3-kinase (PI3K) and protein kinase B (AKT) were activated by sorafenib, which, in turn, increased the phosphorylation level of YB-1. In functional analyses, knockdown of YB-1 led to decreased cell migration and invasion in vitro. At the molecular level, inhibition of YB-1 induced suppression of zinc-finger protein SNAI1 (Snail), twist-related protein 1 (Twist1), zinc-finger E-box-binding homeobox 1 (Zeb1), matrix metalloproteinase-2 (MMP-2) and vimentin levels, implying a role of YB-1 in the epithelial-mesenchymal transition (EMT) process in HuH-7R cells. Additionally, YB-1 contributes to morphological alterations resulting from F-actin rearrangement through Cdc42 activation. Mutation analyses revealed that phosphorylation at S102 affects the migratory and invasive potential of HuH-7R cells. Our findings suggest that sorafenib promotes YB-1 phosphorylation through effect from the EGFR/PI3K/AKT pathway, leading to significant enhancement of hepatocellular carcinoma (HCC) cell metastasis. Elucidation of the specific mechanisms of action of YB-1 may aid in the development of effective strategies to suppress metastasis and overcome resistance.