Autophagy and Epithelial Cell Fate During Anoikis and 3D Morphogenesis

描述（由申请人提供）：自噬是一个受到严格调控的溶酶体降解过程，在这个过程中细胞在各种形式的胁迫下消化自身的内容物。在真核细胞中，自噬的主要功能是在营养耗尽或应激时作为一种保护反应。总的来说，关于这一基本过程如何影响癌症的发生、进展和对治疗的反应，还有很多有待研究。在本提案中，我们将验证自噬通过两种机制影响上皮癌发展的假设：1)增强上皮细胞在ECM脱离诱导的应激和凋亡（anoikis）中的生存能力；2)限制增殖。这一假设是基于一种独特的3D上皮培养系统的初步研究，该系统研究了癌性损伤如何影响腺体结构中正常中空腔的形成。在该模型中，管腔的形成涉及缺乏与细胞外基质（ECM）直接接触的中心细胞的选择性死亡。最近，我们发现了高水平的自噬在管腔形成过程中，表明这一关键过程也有助于上皮细胞的命运。在随后的工作中，我们首次证实自噬在ECM剥夺期间被强烈诱导（anoikis）。基于这些初步数据，在Aim 1中，我们将评估自噬是否通过减轻动脉粥样硬化和三维管腔形成过程中ECM脱离的压力来促进细胞存活。此外，我们已经获得了自噬调节分子Beclin/ATG6（一种已知的肿瘤抑制因子）在3D培养中抑制细胞增殖的初步证据。在目标2中。我们将研究其他自噬调节剂（称为ATGs）是否在生长因子戒断和3D形态发生过程中抑制增殖，以明确确定自噬可以抑制上皮细胞增殖，并确定能够抑制肿瘤细胞扩张的新自噬调节剂。在目标3。我们将描述调节自噬如何影响癌基因表达细胞的存活和增殖，以确定如何利用自噬来治疗已知癌症途径中存在扰动的肿瘤细胞。总的来说，提出的实验将为自噬在人类癌症的发生和发展过程中的精确作用提供独特的信息。最终，这些研究可能会导致新的治疗策略，通过控制自噬来防止癌细胞不受控制的扩张。

DESCRIPTION (provided by applicant): Autophagy is a tightly regulated lysosomal degradation process in which a cell digests its own contents under various forms of duress. In eukaryotic cells, autophagy primarily functions as a protective response during nutrient depletion or stress. Overall, much remains to be learned about how this fundamental process affects cancer initiation, progression and response to therapy. In this proposal, we will test the hypothesis that autophagy influences epithelial cancer development through two mechanisms-1) enhancing the ability of epithelial cells to survive ECM detachment-induced stress and apoptosis (anoikis) and 2) limiting proliferation. This hypothesis is based on initial studies using a unique 3D epithelial culture system to examine how oncogenic insults affect the formation of the normally hollow lumen in glandular structures. In this model, lumen formation involves the selective death of central cells lacking direct contact with extracellular matrix (ECM). Recently, we have discovered high levels of autophagy during lumen formation, indicating that this critical process also contributes to epithelial cell fate. In subsequent work, we are the first to establish that autophagy is strongly induced during ECM deprivation (anoikis). Based on this preliminary data, in Aim 1, we will evaluate whether autophagy promotes cell survival via mitigating the stresses of ECM detachment during anoikis and 3D lumen formation. In addition, we have obtained preliminary evidence that the autophagy regulatory molecule Beclin/ATG6 (a known tumor suppressor) inhibits cell proliferation in 3D culture. In Aim 2. we will interrogate whether other autophagy regulators (called ATGs) inhibit proliferation during growth factor withdrawal and 3D morphogenesis in order to definitively establish that autophagy can restrain epithelial proliferation and to identify new autophagy regulators that are able to inhibit neoplastic cell expansion. In Aim 3. we will delineate how modulating autophagy influences survival and proliferation in oncogene-expressing cells in order to determine how autophagy can be exploited to therapeutically target tumor cells harboring perturbations in known cancer pathways. Overall, the proposed experiments will provide unique information on the precise roles of autophagy during the development and progression of human carcinomas. Ultimately, these studies may lead to new treatment strategies to prevent the uncontrolled expansion of cancer cells via manipulating autophagy.