Gjb4 為一新穎之肺癌生物標記並透過活化 Src 以促進 轉移以及抗藥性

Abstract

大部分肺癌病患在初次確診時都已經是晚期且已經發生轉移（metastasis)。轉移是造成癌症病患復發以及死亡的主要原因，並且復發腫瘤通常展現出抗藥性。在這裡我們藉由流式細胞儀篩選出與血清成份有高度結合以及高轉移能力的細胞株，並發現該細胞株會高表現間隙蛋白30.3 (Connexin 30.3, Gjb4) 。高表現 Gjb4 或抑制 Gjb4 表現會分別增加或減少癌細胞在老鼠模型中的轉移。在臨床檢體中我們發現肺癌組織相較於正常組織會高表現Gjb4 (p=0.0026)，並發現使用Gjb4在血液樣本的白細胞層 (blood buffy coat)的表現可以鑑別出一到三期 (p=0.002814) 以及第四期 (p<0.0001) 的肺癌病患。我們也發現 Gjb4 的高表現與肺癌病患的較差預後 (poor prognosis) (p=1.4e-4) 和復發 (recurrence) (p=1.9e-12) 有關。 使用同源老鼠模型，我們發現 Gjb4 會促進腫瘤生長。高分子量的血清樣品中以類胰島素生長因子1 (Insulin-like Growth Factor-1, IGF-1)為主要的生長因子，並能夠透過PKC路徑誘導Gjb4表現上升。我們也發現 Gjb4 會透過MET活化Src，並且通過Src的活化來促進腫瘤球形成能力(sphere-forming ability)以及軟膠集落形成 （soft-agar colony formation）。此外，我們也發現了 Gjb4 透過活化 Src 來增強對於抗癌藥物健擇 （gemcitabine） 和癌妥滅 (etoposide) 的抗藥性 （drug resistance)。抑制Gjb4的表現和Src抑制劑柏萊膜 (Dasatinib)同時使用能夠進一步降低健擇 （gemcitabine）的毒殺癌細胞的存活率。總結，我們的研究顯示出Gjb4具有潛力成為一個作為肺癌診斷 (diagnostic) 和評估預後 (prognostic) 的新穎生物標記 (biomarker)， 並且Gjb4也可以作為一個潛在的標靶來改善肺癌病患的治療。

Most lung cancer patients are diagnosed late with metastasis, which is the major cause of cancer-related death and recurrent tumors that often exhibit chemoresistance. In the present study, we initially identified gap junction beta-4 protein (Gjb4) to be overexpressed in highly metastatic cancer cells selected by their enhanced binding to serum components. Overexpression or knockdown of Gjb4 increased or decreased lung metastasis of syngeneic mice, respectively. We found that Gjb4 expression was higher in lung tumors than normal tissues (p=0.0026), and Gjb4 levels in blood buffy coat samples showed significant performance in diagnosing stage I-III (p=0.002814) and stage IV (p<0.0001) lung cancer. Moreover, high Gjb4 expression levels were correlated with poor prognosis (p=1.4e-4) and recurrence (p=1.9e-12). Using syngeneic mouse model, we observed that Gjb4 was able to promote tumor growth. High molecular weight serum fraction containing the major growth factor component IGF1 was able to induce Gjb4 via PKC pathway. Gjb4 activated Src signaling via MET, and overexpression of Gjb4 enhanced sphere-forming ability and anchorage-independent growth, which were reversed by inhibition of Src. In addition, we demonstrated that Gjb4-mediated Src activation enhanced chemoresistance of cancer cells toward gemcitabine and etoposide. The combination of Gjb4 knockdown, gemcitabine, and dasatinib further enhanced the inhibition of cancer cell viability. Together, our study has identified Gjb4 as a potential novel diagnostic and prognostic biomarker for lung cancer. Targeting Gjb4 may be exploited as a modality for improving lung cancer therapy.