Role of CD44 in adaptive plasticity of pancreatic cancer

 描述(由申请人提供)： 适应性可塑性是一个用来解释癌细胞如何通过对环境做出表型变化来获得选择性生长和生存能力的短语。适应性可塑性的例证是上皮向间充质转化(EMT)，这是一种使癌细胞迁移和侵袭并增加生存特性的过程。这一过程可以逆转为间充质到上皮的转变(MET)，据信这有利于肿瘤在转移部位的种植和癌细胞的生长。癌细胞中的EMT与跨膜蛋白CD44从表达CD44变异异构体(CD44v)到表达CD44标准型(CD44s)的异构体转换有关。在这一背景下，我们发现吉西他滨处理BxPC3胰腺导管腺癌(PDAC)细胞株可诱导EMT和高水平的CD44s表达，并降低CD44v亚型的表达。这表明，化疗的不良反应可能会推动EMT，增强侵袭性，使肿瘤细胞对化疗更具抗药性。此外，我们还利用流式细胞术从PDAC细胞系(CFPAC-1和AsPC1)中筛选出高表达CD44s的细胞亚群。在这些细胞中，CD44s的敲除导致它们接受MET，侵袭性更小，对化疗更敏感。这些研究表明，化疗或其他环境压力可能会诱导CD44s表型，从而提高生存率和侵袭性；然而，化疗后肿瘤的最佳再生长或微转移种植后的生长可能需要重新转换为CD44v表型并下调CD44s的表达。我们推测CD44s及其异构体是胰腺癌细胞适应性可塑性的关键调节因子。我们进一步假设，了解适应性可塑性的分子基础将为改进治疗提供新的策略。这些研究代表了我们实验室的新数据，并建立在20年来仅专注于胰腺癌生物学和治疗的研究基础上。为了解决这些假设，我们提出了以下三个目标。目的1.确定CD44s和CD44v在PDAC中的功能相关性。这一目标将使用CD44s和CD44v亚型表达受到调控的等基因匹配细胞系模型来实现。对CD44v和CD44s表达的调节作用将从表型、侵袭力、化疗反应、肿瘤种植和生长等方面进行评估。目的：探讨S调节CD44亚型转换和CD44表达的机制。这一目标将确定EMT相关转录抑制因子Snail和ZEB1在抑制CD44v亚型表达所需的ESRP1中的潜在作用，并进一步确定导致CD44s转录增加的机制。目的3.确定抗CD44抗体是否能阻止CD44亚型转换或表达水平，以及这是否能改善化疗反应，减少转移。这些研究将提供与适应性可塑性相关的重要新见解，并确定这些知识是否可以用于对抗化疗诱导的表型转换的不良影响。抑制或逆转向EMT表型的转换可能被证明是提高对化疗的反应和患者生存的重要策略。

 DESCRIPTION (provided by applicant): Adaptive plasticity is a phrase used to explain how cancer cells gain selective growth and survival capabilities through phenotypic changes in response to their environment. Adaptive plasticity is exemplified by epithelial to mesenchymal transition (EMT) a process, which enables cancer cells to become migratory and invasive as well as having an increase in survival properties. This process can be reversed to a mesenchymal to epithelial transition (MET), which is believed to favor tumor engraftment and growth of cancer cells at metastatic sites. EMT in cancer cells is linked with isoform switching of the transmembrane protein CD44 from expressing the CD44 variant isoform (CD44v) to expressing CD44 standard form (CD44s). In this context we found that treatment of BxPC3 pancreatic ductal adenocarcinoma (PDAC) cell line with gemcitabine induces EMT and a high level of expression of CD44s with a decrease in expression of CD44v isoforms. This suggests that an adverse effect of chemotherapy may drive EMT, enhancing invasiveness and rendering tumor cells more resistant to chemotherapy. Moreover, we were able to use flow cytometry to select out from a PDAC cell lines (CFPAC-1 and AsPC1) a subpopulation of cells expressing high levels of CD44s. Knockdown of CD44s, in these cells, caused them to undergo MET, to be less invasive and to become more responsive to chemotherapy. These studies suggest that chemotherapy or other environmental stresses may induce a CD44s phenotype that promotes survival and is more invasive; however optimal tumor regrowth following chemotherapy or growth after engraftment of micrometastases may require re-switching to a CD44v phenotype and down regulation of CD44s expression. We hypothesize that CD44s and its variant isoforms are key regulators of adaptive plasticity in pancreatic cancer cells. We further hypothesize that understanding the molecular basis for adaptive plasticity will provide new strategies for improving therapy. These studies represent new data from our laboratory and build on 20 years of research solely focused on the biology and treatment of pancreatic cancer. To address these hypotheses we propose the following three objectives. Objective 1. Determine the functional relevance of CD44s and CD44v in PDAC. This objective will be addressed using isogenic matched cell line models in which expression of CD44s and CD44v isoforms are regulated. The effects on modulating expression of CD44v and CD44s will be assessed for phenotype, invasiveness, response to chemotherapy, tumor engraftment and growth. Objective 2. Determine the mechanism(s) that regulates CD44 isoform switching and CD44 expression. This objective will determine the potential role of EMT related transcriptional repressors Snail and Zeb1 in suppressing ESRP1 that is required for expression of CD44v isoforms and further identify the mechanisms that lead to the increased transcription of CD44s. Objective 3. Determine whether anti-CD44 antibody prevents CD44 isotype switching or expression level and whether this improves response to chemotherapy and reduces metastasis. These studies will provide important new insights related to adaptive plasticity and determine whether this knowledge can be used to counter act adverse effects of chemotherapy induced phenotypic switching. Inhibiting or reversing the switch towards an EMT phenotype may prove to be an important strategy for increasing the response to chemotherapy and patient survival.