Analysis of Intratumoral Crosstalk in Clonal Populations of OvarianTumor Cells

项目摘要 控制癌症的最重大挑战之一是肿瘤细胞内的高度异质性。 单个肿瘤的肿瘤细胞。虽然肿瘤细胞异质性已经在许多水平上被描述， 对创造异质性的过程的理解正在出现，异质性的程度 肿瘤细胞亚群的功能活性不同， 影响肿瘤发生和发展知之甚少。调查这些问题的主要障碍 缺乏维持培养中人肿瘤异质性的方法， 从单个肿瘤中繁殖不同的克隆亚群。与我们的合作伙伴一起，我们 开发了克服这些障碍的方法，使分离、培养和 表征卵巢肿瘤细胞的克隆群体，然后跟踪条形码化的、表位标记的克隆 从移植克隆的混合物产生的肿瘤异种移植物内的群体。使用这些方法 在一项初步研究中，我们获得了克隆间协作显著影响肿瘤进展的证据-- 最有趣的是，我们发现不能启动肿瘤形成的癌细胞克隆群 它们本身可以强烈地促进原发性肿瘤的扩张和特定器官部位的转移 当与“肿瘤起始”克隆混合时。这支持了一个假设，即克隆亚群在一个 肿瘤之间相互配合，促进肿瘤的扩张和转移。在本建议中，我们描述 计划使用一种系统的方法在人类卵巢肿瘤中测试这一假设，方法是（1）分离和 描述了大量克隆种群之间遗传和表型变异的程度， （2）比较单个克隆群体与这些群体的混合物在卵巢肿瘤中的活性， 为了研究肿瘤内串扰的存在和性质，（3）跟踪肿瘤内定位， 克隆种群随时间的变化，以及（4）阐明负责表型产生的机制， 肿瘤内串扰。

Project Summary One of the most significant challenges in controlling cancer is the high degree of heterogeneity within the neoplastic cells of individual tumors. While tumor cell heterogeneity has been described at many levels and an understanding of the processes that create heterogeneity is emerging, the extent to which the heterogeneous subpopulations of tumor cells vary in their functional activities and whether interactions between subpopulation influence tumor initiation and progression is poorly understood. Major barriers to investigating these questions have been the lack of methodologies to maintain the heterogeneity of human tumors in culture and to propagate distinct clonal subpopulations from individual tumors. Together with our collaborators, we have developed methodologies that overcome these barriers and make it feasible to isolate, culture, and characterize clonal populations of ovarian tumor cells and then track barcoded, epitope-tagged clonal populations within tumor xenografts generated from mixtures of transplanted clones. Using these approaches in a pilot study, we obtained evidence for interclonal collaborations that significantly affect tumor progression -- most interestingly, we found that clonal populations of cancer cells which are unable to initiate tumor formation on their own, can strongly promote the expansion of primary tumors and metastasis at specific organ sites when mixed with 'tumor-initiating' clones. This supports the hypothesis that clonal subpopulations within a tumor cooperate with one another to promote tumor expansion and metastasis. In this proposal, we describe plans to use a systematic approach to test this hypothesis in human ovarian tumors by (1) isolating and characterizing the extent of genetic and phenotypic variation among a large set of clonal populations derived from ovarian tumors, (2) comparing the activities of single clonal populations to mixtures of such populations in order to investigate the existence and nature of intratumoral cross talk, (3) tracking intratumoral localization of clonal populations over time, and (4) elucidating the mechanisms responsible for phenotypes generated by intratumoral crosstalk.