Identifying therapeutic options for intrahepatic cholangiocarcinoma

在美国，每年大约有10,000名新诊断的胆管细胞癌患者，他们的5年 存活率不到10%。肝内胆管细胞来源和分子特征的异质性 (ICC)强调对生物标记物的需求，以对肿瘤进行分层并产生靶向治疗。除胆管上皮细胞外 肝细胞(Hcs)被认为是人类ICC的细胞来源。这一点从快速的HC驱动中可见一斑 激活的AKT(MyrAKT)和Notch胞内域(NICD)共表达诱导小鼠ICC HCS采用睡美人转座系统。 我们对大量ICC患者队列的初步分析表明，AKT-NICD的遗传特征 (AN)驱动的小鼠ICC与约30%的人类ICC显著相关，支持该ICC的临床相关性 模特。Sox9或YAP的缺失延迟了ICC的形成，而是诱导了AN驱动的SOX9-/YAP+或SOX9+/YAP- ICC，也已在人类ICC中发现。这些数据表明，Notch独立地调节 Sox9和YAP在HC驱动的ICC中。重要的是，我们发现YAP和Sox9的共同抑制完全阻止了Notch- 依存性ICC形成。然而，YAP和SOX9驱动肿瘤发生的机制以及它们的 与AKT信号的相互作用仍然知之甚少。根据我们的初步观察，我们的中心假设 HC驱动的ICC肿瘤生长依赖于由两个主要下游驱动的转录和表观遗传改变 Notch信号的效应因子，SOX9和YAP，以及AKT信号。 在目标1中，我们的目标是测试YAP和SOX9共同抑制晚期ICCs的治疗效果，以更好地解决 临床上需要进行晚期治疗。我们将采用遗传性AN-ICC模型和基于肝毒素的ICC模型。 利用创新的可诱导基因调控系统诱导同时、有条件和可诱导的Sox9和YAP 晚期ICC中的压抑。在目标2中，我们提出了三个亚目的来描述一个潜在的分子机制。 介导HC驱动的ICC形成。首先，使用芯片序列和生物信息学工具，我们将识别独特的和 在HC衍生的ICC形成过程中，由Sox9和YAP调节的重叠下游靶点。第二，我们将澄清 NICD-YAP/TEAD-DNMT1表观遗传轴通过调节NICD-YAP/TEAD-DNMT1在HC驱动的ICC发育中的病理作用 AXIS在体内系统中使用药理和遗传工具并研究其对ICC肿瘤甲基组的影响。 为了实现这两个子目标，我们将确定Sox9和YAP的下游效应，以实现更具选择性和更安全的治疗 目标锁定在国际刑事法院。第三，我们的目标是阐明AKT如何介导HC到ICC的转化，这一转化仍然很差 明白了。 这项研究的成功实施将1)为考虑SOX9和YAP联合 抑制作为精准药物治疗的一个有吸引力的候选者，理论上被认为是最有效的方法 攻克致命性肿瘤；2)揭示AKT、SOX9和YAP的关键下游效应因子，有助于开发 为国际刑事法院提供更有效的治疗选择。

In the US, approximately 10,000 patients are newly diagnosed annually with cholangiocarcinoma, and their 5-year survival rate is less than 10%. Heterogeneity in cellular origins and molecular signatures of intrahepatic cholangiocarcinoma (ICC) highlight the demand for biomarkers to stratify tumors and generate targeted therapies. Besides biliary epithelial cells (BECs), hepatocytes (HCs) have been considered as a cellular origin of human ICC. This is evident by rapid HC-driven ICC induction through the co-expression of activated AKT (myrAKT) and Notch intracellular domain (NICD) in mouse HCs using sleeping beauty transposon system. Our preliminary analysis using large ICC patient cohorts demonstrates that the genetic signature of AKT-NICD (AN)-driven murine ICC correlates significantly with ~30% of human ICC, supporting the clinical relevance of this ICC model. Deletion of either Sox9 or Yap delayed ICC formation, and instead induced AN-driven SOX9-/YAP+ or SOX9+/YAP- ICC respectively, which also have been identified in human ICC. These data indicate that Notch independently regulates Sox9 and Yap in HC-driven ICC. Importantly, we found that co-repression of Yap and Sox9 completely prevents Notch- dependent ICC formation. However, the mechanisms of tumorigenesis driven by YAP and SOX9, as well as their interactions with AKT signaling, remain poorly understood. Based on our preliminary observations, our central hypothesis is that HC-driven ICC tumor growth depends on transcriptional and epigenetic alterations driven by two major downstream effectors of Notch signaling, SOX9 and YAP, alongside AKT signaling. In aim 1, we aim to test therapeutic effect of co-repression of YAP and SOX9 in advanced ICCs to better address the clinical need for late-stage therapy. We will employ genetic AN-ICC model as well as liver toxin-based ICC model using innovative inducible gene modulation systems to induce simultaneous, conditional and inducible Sox9 and YAP repression in advanced ICC. In aim 2, we are proposing 3 subaims to delineate the molecular mechanisms underlying AN- mediated HC-driven ICC formation. First, using ChIP-seq and bioinformatic tools, we will identify both the unique and the overlapping downstream targets regulated by Sox9 and Yap during HC-derived ICC formation. Second, we will elucidate the pathologic role of the NICD-YAP/TEAD-DNMT1 epigenetic axis in HC-driven ICC development by modulating this axis with pharmacological and genetic tools in the in vivo system and studying its effects on the methylome of ICC tumors. Pursuing these 2 subaims, we will identify downstream effectors of Sox9 and Yap for more selective and safer therapeutic targeting in ICC. Third, we aim to elucidate how AKT mediates HC-to-ICC transformation, which remains poorly understood. The successful execution of this study will 1) provide an essential evidence for considering SOX9 and YAP co- inhibition as an attractive candidate for precision medicine therapy, which is theorized to be the most effective approach to conquer lethal tumor, and 2) reveal the key downstream effectors for AKT, SOX9 and YAP which will help to develop more potent therapeutic options for ICC.