Dissecting the role of extra centrosomes-induced exosome secretion in PDAC microenvironment

尽管取得了进展，但癌症仍然是医学界的一个严峻挑战。胰腺癌尤其如此，它的5年存活率令人沮丧，只有5%。这种高度侵袭性的行为部分是由纤维化引起的，纤维化形成了肿瘤周围的屏障，阻碍了药物的输送和疗效。这种屏障是由肿瘤引起的被称为星状细胞的细胞群的变化引起的。因此，抗击胰腺癌纤维化被认为是治疗干预的一个重要领域。我们最近发现，携带一种名为中心体的细胞器放大的癌细胞群体能够激活星状细胞。中心体放大是一种存在于癌细胞中的异常现象，在19世纪末被描述为肿瘤发生的可能原因。我们现在知道，这些异常是人类肿瘤的共同特征，可以积极地促进肿瘤的形成。我们实验室发表的工作首次发现了这些异常在影响周围细胞方面的一种新功能(Arnandis等人，2018年)。这项工作表明，中心体放大的癌细胞通过沟通和改变肿瘤内的其他细胞，在肿瘤发生中具有比先前预期更广泛的作用。特别是，我们新的未发表的数据表明，具有额外中心体的癌细胞和胰腺星状细胞之间的这种联系导致了它们的激活，这可能有助于PDAC的纤维化。这是通过释放带有额外中心体的细胞的小泡来调节的，外体充满了指示星状细胞变得激活的因子。因此，了解具有额外中心体的细胞如何激活星状细胞有助于理解纤维化，并指导治疗胰腺癌患者的治疗策略。在这项研究提案中，我们的目标是确定中心体扩增如何诱导外切体的释放，进而这些小泡如何激活星状细胞。我们将结合使用多种方法，包括活细胞成像，以确定导致外切体释放的机制。此外，使用成熟的方法来鉴定这些外切体中包含的因子，我们建议揭示负责星状细胞激活的分子。我们将使用3D培养模型来模拟肿瘤和肿瘤中星状细胞之间的相互作用，来测试阻断已识别的因子是否可以防止星状细胞的激活和纤维化。此外，还将研究外切体在星状细胞中诱导的信号变化，以了解激活的机制。这可能对确定胰腺肿瘤中星状细胞激活和纤维化的特征很重要，并将提供另一个机会来确定阻止纤维化的潜在治疗靶点。最后，我们将使用小鼠模型在体内测试我们的发现。这将提供相关的临床前模型，以评估我们的发现和测试对治疗的反应的重要性。因此，我们的工作不仅有助于首次了解中心体放大在肿瘤微环境中的作用，而且可能在开发针对纤维化的治疗策略以治疗胰腺癌患者方面产生重大影响。

Despite progress, cancer is still a serious challenge in medicine. This is particularly true for pancreatic cancer, which has a dismal 5-year survival rate of <5%. This highly aggressive behaviour is cause in part caused by fibrosis, which forms a barrier around the tumour that hinders drug delivery and efficacy. This barrier is cause by changes in a cell population, named stellate cells, induced by the tumour. Combating fibrosis in pancreatic cancer is thus considered to be an important area for therapeutic intervention. We recently found that a population of cancer cells that carries amplification of a cellular organelle called centrosome is able to activate stellate cells. Centrosome amplification, an aberration that exists in cancer cells, was described in later 1800s as a possible cause of tumourigenesis. We now know that these abnormalities are a common feature of human tumours and can actively contribute to tumour formation. Published work from our lab was the first to identify a novel function of these abnormalities in influencing the surrounding cells (Arnandis et al., 2018). This work demonstrates that cancer cells with amplified centrosomes have a broader role in tumourigenesis than previously anticipated by communicating and changing other cells within tumours. In particular our new unpublished data shows that this communication between cancer cells with extra centrosomes and pancreatic stellate cells leads to their activation, which could contribute to fibrosis in PDAC. This is mediated via the release of small vesicles from cells with extra centrosomes, exosomes, that are packed with factors that will instruct stellate cells to become activated. Thus, understanding how cells with extra centrosomes activate stellate cells could help understanding fibrosis and lead to therapeutic strategies to treat pancreatic cancer patients.In this research proposal we aim to determine how centrosome amplification induces the release of exosomes and in turn how these vesicles activate stellate cells. We will use a combination of approaches, including live cell imaging, to identify the mechanisms responsible for exosome release. Furthermore, using well-established methods to identify the factors contained in these exosomes, we propose to uncover the molecules responsible for stellate cell activation. We will test if blocking the identified factors prevents stellate activation and fibrosis using 3-D culture models that mimic the interaction between cancer and stellate cells in tumours. Changes in signalling induced by exosomes in stellate cells will also be studied to understand the mechanisms of activate. This could be important to identify signatures of stellate cell activation and fibrosis in pancreatic tumours and will provide another opportunity to identify potential therapeutic targets to block fibrosis. Finally, we will use mouse models to test our findings in vivo. This will provide relevant pre-clinical models to assess the importance of our findings and test response to therapy. Thus, our work will not only contribute to understand for the first time the role of centrosomes amplification in tumour microenvironment but could also have a major impact in developing therapeutic strategies that target fibrosis to treat patients that suffer from pancreatic cancer.