Exploitation of RAS signaling to develop therapy and early detection strategies for PDA

描述（由申请人提供）：我们解决的治疗问题是如何抑制IPMN（导管内乳头状粘液性肿瘤）、PanIN（胰腺上皮内瘤变）和PDA（胰腺导管腺癌）的发展和进展。生物学问题是确定启动和传播IPMN，PanIN和PDA的关键信号通路。该领域的一个中心争论是PDA是否起始于胰腺祖细胞或实质腺泡细胞向祖细胞样导管细胞的去分化，在称为腺泡至导管化生（ADM）的过程中。腺泡细胞身份的转录因子主调节子（MR）的功能对于该过程至关重要，但是通过受体酪氨酸激酶（RTK）和G蛋白偶联受体（GPCR）信号传导对其活性的翻译后调节完全未被探索。G蛋白信号传导（RGS）蛋白的调节剂整合来自RTK和GPCR的多种细胞内信号以反馈调节Gi和Gq蛋白信号传导。我们已经开发了一种GFP报告基因（Rgs 16：GFP），使我们能够在IPMN、PanIN和PDA的体内小鼠模型中从早期阶段监测瘤形成和肿瘤形成。将Rgs 16：GFP报告基因与IPMN、PanIN和PDA的小鼠模型相结合，可以深入了解这些肿瘤的发展，并为该过程的抑制剂提供一种新的快速体内筛选方法。我们发现了抑制IPMN向浸润性癌进展的RGS基因。基于使用这些模型的初步分析，我们发现，IPMN和PanIN开始在离散的导管祖细胞在胰腺的腺泡或内分泌隔室的压力。这种解释挑战了目前青睐的范式，即PDA启动与细胞经历ADM，而我们提出，慢性ADM刺激增殖的预先存在的祖细胞在导管。我们发现RTK Axl在胰腺导管中表达，并且在原代培养的PDA祖细胞中与Rgs 16：GFP高度表达。我们的假设是，在基质中表达的Axl-配体Gas 6在上皮祖细胞中引起Axl信号传导，并有助于KrasG 12 D的激活、MR水平和/或活性的翻译后控制、PDA的起始和进展。在我们抑制PDA进展的新疗法的初步体内筛选中，我们确定了阻断Gas 6诱导的Axl活化的华法林与标准护理吉西他滨和Abraxane（GA）组合显著降低PDA进展。该提议的具体目的是1）鉴定最佳药物组合，包括Axl信号传导抑制剂，以抑制PDA。2)鉴定血浆中IPMN、PanIN和PDA的早期标志物。3)确定PDA中的Gas 6-Axl功能。所提出的项目如果成功，将通过以下方式对癌症产生重大影响：i）鉴定PDA的早期血液标志物，以及可能的其他上皮癌，ii）引入新型、通用和快速的体内筛选技术以发现抑制PDA进展的新型药物组合，iii）提供新型遗传工具以确定PDA是否在胰管中的祖细胞中起始，Iv）鉴定先前未知的MR转录因子的调节机制。

DESCRIPTION (provided by applicant): The therapeutic problem we address is how to inhibit the development and progression of IPMN (intraductal papillary mucinous neoplasm), PanIN (Pancreatic Intraepithelial Neoplasia) and PDA (pancreatic ductal adenocarcinoma). The biological problem is to identify key signaling pathways that initiate and propagate IPMN, PanIN and PDA. A central debate in the field is whether PDA initiates in pancreatic progenitor cells or with dedifferentiation of parenchymal acinar cells to progenitor-like ductal cells, in a process termed acinar to ductal metaplasia (ADM). The function of the transcription factor Master Regulators (MRs) of acinar cell identity is critical to this process but post-translational regulaton of their activity through receptor tyrosine kinase (RTK) and G protein coupled receptor (GPCR) signaling is completely unexplored. Regulators of G protein signaling (RGS) proteins integrate multiple intracellular signals from RTKs and GPCRs to feedback regulate Gi and Gq protein signaling. We have developed a GFP reporter (Rgs16:GFP) that allows us to monitor neoplasia and tumor formation from early stages in in vivo mouse models of IPMN, PanIN and PDA. Combining the Rgs16:GFP reporter with mouse models for IPMN, PanIN and PDA is providing insights into development of these tumors and a novel and rapid in vivo screen for inhibitors of this process. We discovered RGS genes that suppress progression of IPMN to invasive carcinoma. Based on preliminary analyses using these models, we find that IPMN and PanIN initiate in discrete ductal progenitor cells in response to stress in the acinar or endocrine compartments of the pancreas. This interpretation challenges the currently favored paradigm that PDA initiates with cells that underwent ADM whereas we propose that chronic ADM stimulates proliferation of pre-existing progenitor cells in ducts. We find that the RTK Axl is expressed in pancreatic ducts and is highly expressed with Rgs16:GFP in PDA progenitor cells in primary culture. Our hypothesis is that the Axl-ligand Gas6 expressed in stroma evokes Axl signaling in epithelial progenitor cells and contributes to activation of KrasG12D, post-translational control of MR levels and/or activity, PDA initiation and progression. In our preliminary in vivo screens of new therapeutics to inhibit PDA progression, we determined that warfarin, which blocks Gas6 induced activation of Axl, combined with the standard of care Gemcitabine and Abraxane (GA), significantly reduced PDA progression. The Specific Aims of this proposal are 1) Identify optimal drug combinations, including inhibitors of Axl signaling, to inhibit PDA. 2) Identify early markers of IPMN, PanIN and PDA in plasma. 3) Determine Gas6-Axl function in PDA. The proposed project, if successful, will have a major impact on cancer by i) identifying early blood markers of PDA, and perhaps other epithelial cancers, ii) introducing a novel, general and rapid in vivo screening technique to find novel drug combinations that inhibit PDA progression, iii) providing novel genetic tools to determine if PDA initiates in progenitor cells in the pancreatic duct, Iv) identify previously unknown regulatory mechanisms of the MR transcription factors.