Investigating the function of subclonal cooperation in breast cancer progression

项目摘要/摘要 肿瘤内遗传异质性存在于绝大多数实体癌中，并与肿瘤相关。 攻击性强，治疗反应差。肿瘤异质性是否促进肿瘤生长和 存活率和驱动这些表型变化的潜在机制是开放的研究领域。至 为了探索这一根本的知识鸿沟，我建立了一个研究亚克隆合作与肿瘤的模型 分别使用过表达MYC的等基因MCF10A细胞系在体外和体内的异质性 (MYC)或致癌的HRASG12V(HRAS)。MYC和致癌RAS是已知的两个典型癌基因 以不同的途径传递信号，尽管有关MYC和HRAS相互作用的研究主要探索 细胞内在的合作机制。因此，我将使用MYC和HRAS作为亚克隆的样本 合作研究它们在不同细胞群体中的协作性。在使用MYC和 HRAS模型，我做了令人兴奋的观察，表达MYC或HRAS的单个亚克隆相互合作 非细胞自主：当亚克隆在体外共同生长和肿瘤生长时，细胞增殖增加 当亚克隆被联合注射到小鼠体内时，速度更快。重要的是，这些数据表明靶向克隆 相互依赖可能是改善治疗策略所必需的。我们假设肿瘤亚克隆 合作诱导新的肿瘤特性促进肿瘤生长，并潜在地促进肿瘤转移和 抗药性。 这项拟议研究的三个实验目标将严格检验我的假设，研究 细胞培养及同种异种移植中的亚克隆协作现象 模特们。实验目标1将测试亚克隆合作是否需要更快的肿瘤启动和 增殖，通过在肿瘤进展过程中的不同时间点解构异质肿瘤来实现。 实验目标2将通过定义外切体介导的蛋白质来探索肿瘤异质性是如何发挥作用的 转移有助于亚克隆协作。最后，在实验目标3中，我将测试亚克隆 相互作用促进人-乳腺癌原位小鼠乳腺肿瘤的治疗耐药 衍生的细胞系。此外，我将通过对亚克隆异质性的表征来验证我们的发现 在现成的PDX模型库中存在的特定癌基因的亚克隆表达。一起， 这些数据将提高我们对亚克隆合作在肿瘤进展中的理解，提供关键的见解 未来的工作目标是有效地解构和治疗复杂的、异质的肿瘤。

Project Summary/Abstract Intratumor genetic heterogeneity is present in a vast majority of solid cancers and correlates with tumor aggressiveness and poor therapeutic response. Whether tumor heterogeneity promotes tumor growth and survival, and the potential mechanisms driving these phenotypic changes, are open areas of investigation. To explore this fundamental gap in knowledge, I generated a model for studying subclonal cooperation and tumor heterogeneity in vitro and in vivo, respectively, using isogenic MCF10A cell lines that either overexpress MYC (MYC) or oncogenic HRASG12V (HRAS). MYC and oncogenic RAS are two canonical oncogenes that are known to signal in distinct pathways, though studies focused on MYC and HRAS interactions have primarily explored cell intrinsic mechanisms of cooperation. Therefore, I will use MYC and HRAS as an exemplar for subclonal cooperation to study their cooperativity in distinct cell populations. In preliminary studies using the MYC and HRAS model, I made the exciting observation that individual subclones expressing MYC or HRAS cooperate non-cell autonomously: cell proliferation increases when subclones are grown together in vitro and tumor growth is more rapid when subclones are co-injected into mice. Importantly, these data suggest targeting clonal interdependence may be necessary to improve therapeutic strategies. We hypothesize tumor subclones cooperate to elicit emergent tumor properties promoting tumor growth, and, potentially, tumor metastasis and drug resistance. The three Experimental Aims of this proposed research will rigorously test my hypothesis, studying the phenomenon of subclonal cooperation in cell culture and in orthotopic and patient derived xenograft (PDX) models. Experimental Aim 1 will test if subclonal cooperation is necessary for more rapid tumor initiation and proliferation, by deconstructing heterogenous tumors at distinct timepoints during tumor progression. Experimental Aim 2 will explore how tumor heterogeneity is functional by defining how exosome mediated protein transfer contributes to subclonal cooperation. Finally, in Experimental Aim 3, I will test whether subclonal interactions promote therapeutic resistance in orthotopic mouse mammary tumors generated using human- derived cell lines. Additionally, I will validate our findings of subclonal heterogeneity by characterizing the subclonal expression of specific oncogenes present in in a library of readily available PDX models. Together, these data will improve our understanding of subclonal cooperation in tumor progression, providing key insights for future work aimed at efficiently deconstructing and treating complex, heterogenous tumors.