Elucidating the dependencies of tumor initiating and drug-resistant niches in human malignancies by genome- wide molecualr profiling of single cells

 描述(由申请人提供)：从易治疗的肿瘤发展到难治的、耐药的形式，无论是在肿瘤生物学机制的基本阐明方面，还是就其翻译和临床意义而言，都可能是最艰巨的挑战。即使不是所有的肿瘤都会自发发展到转移性耐药阶段，我们识别进展风险更大的患者的能力也是极其有限的。对于注定要进展的肿瘤，这一挑战提出了两种截然不同但高度互补的观点。从宏观上看，进展的发生是因为对于特定的恶性肿瘤尚不存在药理上可操作的机制，或者由于遗传和表观遗传机制而产生耐药性。例如，虽然70%的HER2+乳腺癌最初对曲妥珠单抗有反应，但其中70%最终将复发为曲妥珠单抗耐药肿瘤。同样令人沮丧的结果也反映在大多数靶向治疗中。然而，在显微镜下，耐药性的出现根源于癌症的极端异质性，既跨个体(肿瘤间)，更重要的是跨个体肿瘤细胞(肿瘤内)。这项建议的目标是开发一种新的方法学框架，整合实验和计算方法，系统地阐明肿瘤异质性驱动肿瘤进展和出现耐药的机制。它将特别专注于在单细胞水平上研究肿瘤内的异质性，以确定有助于亚克隆扩张和耐药生态位出现的独立分子事件的范围。这些研究产生的方法学进步将得到广泛传播，并将适用于对任何有适当数据可用的人类恶性肿瘤的无偏见分析。这些方法将产生假设，产生全面的高可能性分子机制曲目，将得到实验验证。在这一更广泛的背景下，拟议研究的主要重点将是扩展成功应用于多细胞样本的基于网络的方法，以在单细胞水平上研究肿瘤异质性对进展和耐药性的影响。我们将集中于三个异质性来源：(A)遗传上不同的肿瘤亚克隆，(B)表观重编程但相同基因的肿瘤亚群，以及(C)正常细胞(例如，间质或免疫系统相关)，它们的存在调节肿瘤细胞的行为和对治疗剂的反应。我们的假设是，阐明单细胞中与肿瘤相关的机制对于开发更好的预防、诊断和治疗这种疾病的策略至关重要。

 DESCRIPTION (provided by applicant): Tumor progression from a tractable to an intractable, drug-resistant form represents perhaps the most formidable challenge both in terms of basic elucidation of tumor biology mechanisms and in terms of its translational and clinical implications. Even though not all tumors will spontaneously progress to a metastatic drug-resistant stage; our ability to identify the patients at greater risk of progression is extremely limited. For tumors destined to progress, the challenge presents two distinct, yet highly complementary perspectives. Macroscopically, progression occurs because either pharmacologically actionable mechanisms do not yet exist for a specific malignancy or because drug resistance ensues, due to genetic and epigenetic mechanisms. For instance, while 70% of HER2+ breast adenocarcinomas initially respond to trastuzumab, 70% of these will eventually relapse to trastuzumab-resistant tumors. The same dismal outcome is reflected across most targeted therapeutics. Microscopically, however, emergence of drug resistance is rooted in the exceedingly heterogeneous nature of cancer, both across individuals (inter-tumor) and, more importantly, across individual tumor cells (intra-tumor). The goal of this proposal is the development of a novel methodological framework integrating both experimental and computational approaches to systematically elucidate the mechanisms by which tumor heterogeneity drives tumor progression and emergence of drug resistance. It will focus specifically on the study of intra-tumor heterogeneity at the single cell level to identify the rane of independent molecular events contributing to sub-clonal expansion and emergence of drug-resistant niches. The methodological advances resulting from these studies will be broadly disseminated and will be applicable to the unbiased analysis of any human malignancy for which appropriate data is available. These methodologies will be hypothesis generating, producing comprehensive repertoire of high-likelihood molecular mechanisms that will be experimentally validated. Within this broader context, the primary focus of the proposed research will be on extending network-based methodologies, which were successfully applied to multicellular samples, to study the impact of tumor heterogeneity on progression and drug resistance, at the single cell level. We will focus on three sources of heterogeneity: (a) genetically distinct tumor subclones, (b) epigenetically reprogrammed yet isogenic tumor sub-populations, and (c) normal cells (e.g. stromal or immune system related), whose presence modulates tumor cell behavior and response to therapeutic agents. Our hypothesis is that elucidating tumor-related mechanisms in single cells is critical to the development of better strategies for prevention, diagnosis, and treatment of the disease.