WDR4驅動PML降解以形成免疫抑制及促進細胞轉移的肺腫瘤微環境之機制研究

Abstract

腫瘤微環境對於腫瘤生長與轉移，扮演著相當重要之角色。然而，癌細胞如何調控微環境中細胞與非細胞物質之機制，至今仍未完全明瞭。 PML為一腫瘤抑制蛋白，許多研究指出PML具有多方面抑制腫瘤生長及特性的相關能力，並且在許多不同種類的腫瘤中，其表現量有降低的情形；但是目前PML對於腫瘤微環境調控之探討，仍然相當不足。在本篇論文中，我們發現WDR4可作為受質辨識器(substrate adaptor)，並與Cullin4、Roc1和DDB1蛋白形成連接酶(E3 ligase)，將PML進行多次泛素化 (polyubiquitination)，並且降解PML。臨床上的證據也顯示出在肺癌中，WDR4/PML的作用路徑有高度活化之現象，並且此路徑的高度活化與病人不良的預後有相關性。接著我們想研究WDR4/PML路徑在肺癌進程中的生物意義，因此我們利用基因微陣列分析發現，WDR4/PML可誘導一群細胞表面蛋白或是分泌蛋白之表現，它們分別是CD73、uPAR及SAA2。藉由在生物體外(in vitro)及動物實驗，我們發現WDR4/PML可透過這些誘導出的下游基因，促進肺癌細胞移動、侵襲和轉移之能力。此外，我們利用異體移植動物實驗和基因轉殖小鼠模式，觀察到WDR4/PML會增加入侵腫瘤內的調控性T細胞 (Tregs)與腫瘤相關巨噬細胞 (M2-like macrophages)之數目，但降低細胞毒殺性T細胞(CD8+ cytotoxic T lymphocytes)之數目；因此WDR4/PML作用路徑，可以創造出一個免疫抑制的腫瘤微環境；然而，此一現象卻可以被CD73抑制劑所抑制。整體來說，我們發現WDR4是一個新興致癌蛋白，可以透過多次泛素化以降解PML，此調控會創造出免疫抑制及促進癌細胞轉移之腫瘤微環境；此研究也顯示未來在治療PML不正常降解之肺癌時，也許可以使用免疫調控之方式，以達到治療效果。

The tumor microenvironment plays an important role in tumor growth and metastasis. However, the mechanism by which tumor cells regulate the cell and non-cell constituents of surrounding stroma remains incompletely understood. PML is a pleiotropic tumor suppressor but its role in tumor microenvironment regulation is poorly characterized. PML protein is frequently downregulated in many cancer types, including lung cancer. Here, we identify a novel PML ubiquitination/destruction pathway mediated by ubiquitin ligase CRL4WDR4. Clinically, this PML destruction pathway is hyperactivated in lung cancer and correlates with poor prognosis. The WDR4/PML axis induces a set of cell surface or secreted factors, including CD73, uPAR, and SAA2, which elicit paracrine effects to stimulate migration, invasion, and metastasis in multiple lung cancer models. Furthermore, in both xenograft and genetically engineered mouse models, the WDR4/PML axis elevates intratumoral Tregs and M2-like macrophages and reduces CD8+ T cells to promote lung tumor growth and these immunosuppressive effects are all reversed by CD73 blockade. Our study identifies WDR4 as a novel oncoprotein which negatively regulates PML via ubiquitination to promote lung cancer progression by fostering an immunosuppressive and pro-metastatic tumor microenvironment and suggests a potential of immune-modulatory approaches for treating lung cancer with aberrant PML degradation.