Oral Cancer Initiating Cells: Characterization

描述(申请人提供)：尽管强化治疗通常结合了手术、放疗和化疗，但口腔鳞状细胞癌(OSCC)的长期存活率只有15%-50%。因此，口腔鳞状细胞癌的药物治疗/化疗方法亟需改进。目前的一种理论是，传统疗法之所以失败，是因为它没有充分治疗癌症起始细胞(CIC)，也就是癌症干细胞(CSCs)。本实验室建立了4-硝基喹啉氧化物(4-NQO)口腔癌小鼠致癌模型，是目前应用最广泛的口腔鳞癌研究动物模型。当我们在饮用水中提供4-NQO(一种致癌物质和吸烟引起的肿瘤损害的替代品)时，小鼠的口腔内会出现类似于人类的病变，包括增生、异型增生、白斑、乳头状瘤和侵袭性鳞状细胞癌(SCCs)；此外，在这个小鼠模型中，OSCC的分子标记与许多人类OSCC中的分子标记相同。在这里，我们建议在这个4-NQO口腔癌变模型中使用细胞谱系追踪的方法来检验这一假设，即参与染色质调节的某些多梳蛋白，特别是Bmi1，也在口腔中可能的CICs中发挥主要作用。在添加三苯氧胺时，表达Bmi1的细胞将被永久标记，方法是使用具有三苯氧胺调节的、由Bmi1启动子驱动的Creer()融合蛋白基因的转基因小鼠，并将它们与rosa26报告转基因小鼠杂交。这些小鼠将在致癌过程中被跟踪，以确定表达Bmi1的细胞后代以及随时间发展的口腔鳞状细胞中Bmi1+标记细胞的表达。我们还将通过在口腔癌发生过程中通过使用多西环素调控的表达载体，在口腔上皮细胞中过度表达Bmi1，并以一种受调控的方式来表征Bmi1在口腔鳞癌中的功能。这些目标的完成将为我们提供许多关于Bmi1基因的新信息，该基因被认为是人类口腔鳞状细胞癌形成CICs所必需的关键基因。此外，这项拟议研究中使用和进一步发展的技术将为我们提供有用的、强大的工具来鉴定和研究口腔鳞状细胞癌中的CICs，包括它们的自我更新能力、它们的分化能力和它们的表型流动性。这种知识对于发现新的治疗方法是必不可少的。 并在人类口腔鳞状细胞中筛选靶向CICs的药物。

DESCRIPTION (provided by applicant): Despite intensive treatment that generally combines surgery, radiation, and chemotherapy, oral squamous cell carcinomas (OSCCs) have a long-term survival rate of only 15-50%. Thus, there is a great need for improvements in pharmacologic treatments/chemotherapeutics for OSCCs. One current theory is that conventional treatment fails because it does not adequately treat cancer-initiating cells (CICs), also called cancer stem cells (CSCs). Our laboratory developed the 4-NQO (4-nitroquinoline oxide) carcinogenesis model of oral cancer for mice, now the most widely used murine model for the study of the development of OSCC. When we provide 4-NQO, a carcinogen and a surrogate for the neoplastic lesions caused by smoking, in the drinking water, mice develop lesions in their oral cavities that mimic those in humans, including hyperplasia, dysplasia, leukoplakia, papilloma, and invasive squamous cell carcinomas (SCCs); moreover, the molecular markers of OSCC in this murine model are the same as many of those in human OSCCs. Here we propose to use a cell lineage-tracing approach in this 4-NQO oral carcinogenesis model to test the hypothesis that certain Polycomb proteins which are involved in chromatin regulation, specifically Bmi1, also play a major role in putative CICs in the oral cavity. Bmi1-expressing cells will be permanently marked at the time of tamoxifen addition by using transgenic mice that have a tamoxifen-regulated, creER(TAM) fusion protein gene driven by the Bmi1 promoter, and crossing them with Rosa26 "confetti" reporter transgenic mice. These mice will be followed during the carcinogenesis process to determine the Bmi1-expressing cell progeny and the expression of Bmi1+ marked cells in OSCCs that develop over time. We will also characterize the functions of Bmi1 in OSCC by over- expressing Bmi1, specifically in the oral cavity epithelium and in a regulated manner, through the use of a doxycycline-regulated expression vector in mice during oral cavity carcinogenesis. Completion of these aims will provide us with much new information about the Bmi1 gene, which is thought to be a key gene required for formation of CICs in human OSCCs. Moreover, the techniques used and further developed in this proposed research will provide us with useful, powerful tools with which to identify and study CICs in OSCC, including their ability to self-renew, their abilityto differentiate, and their phenotypic fluidity. This knowledge is essential to discover new therapies and to screen for drugs that target CICs in human OSCCs.