Modelling breast cancer heterogeneity: Effects of stromal compartments on sub-clonal decision-making

乳腺癌的诊断和治疗已经取得了重大进展，然而，肿瘤的异质性仍然是设计有效治疗方法的一个相当大的挑战。同一肿瘤内不同遗传和表型特征的癌细胞亚群的共存会导致负面的治疗影响、最终的复发风险和患者较差的生存。尽管这对治疗效果有明显的影响，但癌症的异质性问题常常被忽视。到目前为止，由于缺乏足够的模型或纵向数据，异质性方面在癌症研究中的实施非常有限。到目前为止，这个问题要么是通过分析小鼠模型中的患者来源的异种移植物(PDX)来解决的，要么是通过应用计算生物学从多区域活检中重建癌症的进化及其异质性。BC的死亡本质上是由于转移到远处和继发肿瘤，这些肿瘤的细胞和基因特征与原始肿瘤的大量数据不同。不幸的是，目前还没有对转移瘤构成的详细纵向评估。然而，最近，人们对同一肿瘤内或患者之间异质性的重要性的认识正在稳步增长，并被认为是未来诊断和治疗的主要因素。在我们的实验室中，我们目前正在开发一种异质性的时空分辨3D模型，我们应用该模型来更好地了解BC的发生和发展。我们特别感兴趣的是了解多克隆竞争环境中已知和假定的BC驱动程序的层次结构。因此，我们开发了一种多基因-多荧光团结构，允许我们诱导特定的癌症信号并实时追踪不同的谱系。我们目前的模型研究了一组25个不同多基因组合的基因，这些组合可以在大多数BC病例中找到，而不受其分子亚型的影响。根据cBio门户数据库，该小组涵盖了大约86%的BC病例的基因签名。然而，由于构造独特的生物砖建筑体系，面板可以很容易地扩展或更换。在建立了BC异质性和肿瘤演化模型的基础上，本研究重点研究了异质性肿瘤与微环境的相互作用，以及基质细胞，特别是内皮细胞、成纤维细胞和滞留或需要的巨噬细胞在肿瘤发生和发展中的作用。我们假设不同的亚克隆群体之间以及与微环境本身的特定和明显的相互作用取决于其组成。关于BC的异质性，目的是在体外将RAFT系统与多尺度3D模型相结合或在小鼠体内使用活体成像来研究在没有和存在肿瘤微环境的情况下多克隆肿瘤的相互作用。最终，这项研究将1)揭示BC驱动因素的层次结构及其可能的变化，取决于微环境成分；2)参考现有的计算异质性和肿瘤进展模型，并开发一种捕捉这种复杂串扰的模型；3)分析体内亚克隆相互作用以及与微环境的相互作用，以及后来对次生生态位的亚克隆偏好。综上所述，这项研究将检验这样一种假设，即在不同的肿瘤中，细胞轮廓和行为的亚克隆变化将导致不同的生存和进展计划，并且肿瘤发展中的这些优势将随着周围微环境的组成而改变。

Significant progress has been made in the diagnosis and treatment of breast cancer (BC); however, tumour heterogeneity remains a considerable challenge in the design of effective therapies. The coexistence of subpopulations of cancer cells within one tumour with different genetic and phenotypic characteristic leads to negative treatment impact, ultimate recurrence risk and poor patient survival. Despite this identified impact on treatment efficacy the subject of cancer heterogeneity is often neglected. To this date, the implementation of the heterogeneity aspect into cancer studies is very limited due to the lack of adequate models or longitudinal data. So far, this issue is either tackled by analysing Patient-Derived Xenografts (PDXs) in mouse models or through application of computational biology for the reconstruction of cancer evolution and its heterogeneity from multiregional biopsies. BC mortality essentially results from metastases to distant sites and secondary tumours, which cellular and genetic profiles differ from the original tumour bulk data. Unfortunately, detailed longitudinal assessments of metastases composition are currently not available. Nevertheless, lately, the understanding of the importance of heterogeneity within the same tumour or between patients is steadily growing and is recognised to be a major factor in future diagnosis and treatment. In our laboratory, we currently are developing a spatio-temporal resolving in vitro 3D model for heterogeneity that we apply to understand the onset and progression of BC better. We are specifically interested in understanding the hierarchy of known and putative BC drivers in a multiclonal competing environment. Therefore, we develop a multigene-multifluorophore construct that allows us to induce specific cancer signatures and to trace different lineages in real-time. Our current model investigates a panel of 25 genes in different multigene-combinations which can be found in most BC cases independent of their molecular subtype. This panel covers genetic signatures of about 86% of all BC cases according to the cBioPortal data bank. Nevertheless, due to the constructs unique bio brick building system, the panel can easily be extended or exchanged. Having developed a model for BC heterogeneity and tumour evolution this study focuses on the interaction of the heterogeneous tumour mass with the microenvironment and the contribution of stromal cells, specifically endothelial cells, fibroblasts and resident or requited macrophages on tumour development and progression.We hypothesis that different sub-clonal populations will have specific and distinct interactions with each other and with the microenvironment itself depending on its composition. Relating to heterogeneity in BC the aim is to study the interaction of a multiclonal tumour in absence and presence of tumour microenvironment in vitro combining the RAFT system with a multiscale 3D model or in vivo using intravital imaging in mice. Ultimately, the study will 1) reveal the hierarchy of BC drivers and its possible variation depending on microenvironmental components; 2) consult existing computational heterogeneity and tumour progression models and develop one that captures this complex crosstalk; 3) analyse in vivo sub-clonal interactions with each other and with the microenvironment and later on sub-clonal preference for a secondary niche. Taking together, the study will test the hypothesis that sub-clonal variation in cellular profiles and behaviour in a heterogeneous tumour will lead to different survival and progression programmes and that these advantages in tumour development will change with the composition of the surrounding microenvironment.