Investigating multifactorial beta-catenin activation in hepatocellular cancers

摘要 肝细胞癌（HCC）是一种致命的恶性肿瘤，治疗选择有限，缺乏分子- 靶向治疗。Wnt/β-catenin级联反应的激活在HCC的发生发展中起重要作用 发病机制编码β-连环蛋白的基因CTNNB 1的突变干扰其降解，导致 其功能获得（GOF）和活化，并且涉及所有HCC的20-35%。相互排斥的群体 另外约8%的HCC是AXIN 1中具有功能丧失（LOF）突变的HCC，AXIN 1编码 对于β-连环蛋白降解至关重要的支架蛋白AXIN 1。我们以前的研究也表明，β- 单独的连环蛋白激活不足以促进HCC的发展。基于CTNNB 1的伴随存在 在约11%的人HCC中存在LOF突变和c-MET激活，在AXIN 1和c-MET中存在LOF突变。 在约4%的人HCC中MET一起活化，我们建立了两种小鼠HCC模型，c-Met/β-catenin和 c-Met/sgAxin 1，使用睡美人转座子/转座酶和水动力尾静脉注射（SB-HDTVI）。 这些模型概括了基于基因表达研究的各自的人类HCC子集。有趣的是， 使用这些小鼠模型和人HCC样本，我们发现AXIN 1 LOF突变型HCC不 显示典型肝特异性β-连环蛋白靶基因如谷氨酰胺合成酶（Gs）和Tbx 3的激活， 这在CTNNB 1突变型HCC中是明显的。相比之下，Hippo级联在L0 F突变中失活， AXIN 1突变型，但CTNNB 1突变型HCC中没有。基于上述观察，我们的总体假设 尽管β-catenin是常见的下游效应子，但CTNNB 1和AXIN 1的突变导致 HCC的不同分子亚型，这两类肿瘤的发展需要不同的分子参与。 信号通路我们提出以下三个具体目标来解决我们高度相关的假设。在 目的1，我们计划确定c-Met/sgAxin 1是否需要Wnt/β-catenin的配体依赖性激活 在小鼠中诱导HCC形成。在目标2中，我们将研究Gs依赖性和非依赖性代谢， 小鼠肝癌发展中的信号级联反应。在目标3中，我们将描述 Ctnnb 1 GOF和Axin 1 LOF突变型HCC中的Hippo级联反应。总之，我们的研究将阐明不同的 由于两种不同的机制，β-连环蛋白激活诱导的信号通路，以及我们如何有效地 根据基因变异来治疗这些肿瘤。该提案代表了一种持续的、富有成效的合作 加州大学旧金山分校的陈欣博士和匹兹堡大学的保罗·蒙加博士之间的对话。结果可能会铺平一条 肝癌精准医疗的方法。

Abstract Hepatocellular carcinoma (HCC) is a deadly malignancy with limited treatment options, and lacks molecular- targeted therapies. Activation of Wnt/β-catenin cascade has been shown to play a major role during HCC pathogenesis. Mutations in CTNNB1, the gene encoding for β-catenin, interfere with its degradation leading to its gain-of function (GOF) and activation, and are implicated in 20-35% of all HCCs. A mutually exclusive group of additional around 8% of HCCs is the one with the loss-of-function (LOF) mutations in AXIN1, which encodes for a scaffolding protein AXIN1, essential for β-catenin degradation. Our previous studies also showed that β- catenin activation alone is insufficient for HCC development. Based on the concomitant presence of CTNNB1 mutations and c-MET activation in ~11% of human HCC, and the presence of LOF mutations in AXIN1 and c- MET activation together in ~4% of human HCC, we established two murine HCC models, c-Met/β-catenin and c-Met/sgAxin1, using sleeping beauty transposon/transposase and hydrodynamic tail vein injection (SB-HDTVI). These models recapitulate the respective human HCC subsets based on gene expression studies. Intriguingly, using these mouse models and human HCC samples, we discovered that AXIN1 LOF mutant HCC does not show activation of canonical liver-specific β-catenin target genes such as glutamine synthetase (Gs) and Tbx3, which was evident in CTNNB1-mutant HCCs. In contrast, Hippo cascade is inactivated in LOF mutations in AXIN1 mutant, but not in CTNNB1-mutant HCCs. Based on the above observations, our overarching hypothesis is that despite β-catenin being the common downstream effector, mutations in CTNNB1 and AXIN1 lead to distinct molecular subtypes of HCC, and tumor development in these two classes requires participation of distinct signaling pathways. We propose the following three specific aims to address our highly relevant hypothesis. In Aim 1, we plan to define whether ligand dependent activation of Wnt/β-catenin is required for c-Met/sgAxin1 induced HCC formation in mice. In Aim 2, we will investigate Gs dependent and independent metabolic and signaling cascades in mouse HCC development. And in Aim 3, we will characterize the functional contribution of Hippo cascade in Ctnnb1 GOF and Axin1 LOF mutant HCCs. Altogether, our studies will elucidate the distinct signaling pathways induced by β-catenin activation due to two distinct mechanisms, and how we may effectively target these tumors based on genetic variations. The proposal represents an ongoing & productive collaboration between Dr. Xin Chen from UCSF and Dr. Paul Monga from University of Pittsburgh. The results may pave a way for precision medicine in HCC.