Ral GTPases as novel regulators of acinar cell plasticity in the pancreas – implications for cancer development and treatment

胰腺癌是最致命的癌症之一，5年生存率为10%。胰腺导管腺癌（PDAC）发展的第一步是腺泡细胞向导管细胞的转分化（腺泡-导管化生，ADM），然后进一步发展为肿瘤，最后形成浸润性癌。我们发现RALGAP复合物（RALA和RALB的负调控信号）的缺失使腺泡细胞对ADM敏感，并通过致癌KRASG12D和PDAC的发展显著增加了它们对致癌转化的易感性。RALGAP缺乏还会阻止急性胰腺炎后腺泡细胞室的再生，而急性胰腺炎ADM通常是短暂的，这表明RALGAP/RAL活性的失调是癌症发展的一个危险因素。在PDAC样品和细胞系中，RAL GTPase活性经常上调。然而，目前还没有研究表明RAL GTPase在PDAC中的作用，并且到目前为止，还没有研究表明RAL GTPase的功能与腺泡可塑性有关。本建议的目的是对ADM过程中RAL GTPase的功能进行全面分析。我们希望充分表征RALGAPβ丢失的表型，其中RAL GTPase过度活跃，关于ADM及其对急性和慢性胰腺炎炎症反应的影响，我们将阐明在胰腺炎症过程中何时以及如何调节RAL GTPase活性。此外，我们将在各自的体内模型中研究RALA和/或RALB基因消融对急性和慢性炎症期间ADM和肿瘤发展的影响。在这里，我们将能够表征，在腺泡可塑性的哪个阶段RAL GTPases是关键的，以及是否存在RAL异构体特异性依赖。由于胰腺癌仍然难以治疗，因此对确定新的治疗方案有很大的研究兴趣。我们将测试使用小分子BQU57抑制RAL是否可以预防或恢复ADM，从而作为早期干预策略。最后，我们希望揭示由RAL驱动的控制腺泡可塑性的信号通路和细胞过程。我们已经建立了3D类器官和细胞培养系统，可以在离体环境中分析ADM，我们将结合RNA测序和磷酸化蛋白质组学方法来识别RAL GTPases下游的关键介质和过程。我们的研究结果将提高我们对胰腺炎后再生过程和PDAC发展起始事件的理解，我们希望这将促进发现早期检测的标志物以及早期干预的策略。

Pancreatic cancer represents one of the deadliest cancers with a 5-year survival rate of 10 %. The initial step in pancreatic ductal adenocarcinoma (PDAC) development is the transdifferentiation of acinar to ductal cells (acinar-to-ductal metaplasia, ADM), which then further progress to form neoplasias and finally invasive carcinoma. We found that loss of RALGAP complexes, negative regulators of signaling by the small RAL GTPases RALA and RALB, sensitizes acinar cells to ADM and drastically increases their susceptibility to oncogenic transformation through oncogenic KRASG12D and PDAC development. RALGAP deficiency also prevents regeneration of the acinar cell compartment after acute pancreatitis, where ADM is normally transient, suggesting deregulation of RALGAP/RAL activity as a risk factor for cancer development. RAL GTPase activity is frequently upregulated in PDAC samples and cell lines. However, no in vivo studies have addressed the role of RAL GTPases in PDAC and RAL GTPase function has until now not been connected to acinar plasticity. The aim of this proposal is to provide a comprehensive analysis of RAL GTPase function(s) in the ADM process. We wish to fully characterize the phenotype of RALGAPβ loss, where RAL GTPases are hyperactive, in respect to ADM and its impact on the inflammatory response upon acute and chronic pancreatitis and we will elucidate when and how RAL GTPase activity is regulated during inflammatory processes in the pancreas. Furthermore, we will investigate the effects of genetic ablation of RALA and/or RALB on ADM and tumor development during acute and chronic inflammation in respective in vivo models. Here, we will be able to characterize, for which phase during acinar plasticity RAL GTPases are critical and whether RAL isoform-specific dependencies exist. As pancreatic cancer remains difficult to treat, there is great research interest in identifying new treatment options. We will test if RAL inhibition using the small molecule BQU57 can prevent or revert ADM and thus serve as an early intervention strategy. Finally, we wish to unravel the signaling pathways and cellular processes that are driven by RAL to control acinar plasticity. We have established 3D organoid and cell culture systems that allow analysis of ADM in ex vivo settings, which we will combine with RNA sequencing and phosphoproteomics approaches to identify critical mediators and processes downstream of RAL GTPases. Our findings will improve our understanding of the regeneration processes after pancreatitis and the initiating events in PDAC development, which we hope will promote the discovery of markers for early detection as well as strategies for early intervention.