Human stem cell models of de novo tumorigenesis to replace the use of animals for the study of cancer initiation

该项目旨在开发一种新的复杂人类细胞模型来研究乳腺癌三阴性乳腺癌（TNBC），同时减少研究所需动物的使用。TNBC是最具侵袭性的乳腺肿瘤亚型，通常表现为高分级、高转移潜力和预后差。TNBC仍然是临床上最具挑战性的乳腺癌类型，因为没有有效的靶向治疗方法。TNBC的异质性进一步阻碍了进展，因为TNBC也不是一种单一的疾病，而是根据影响患者预后的特定分子途径特征和突变景观进一步细分为不同的分子和临床亚型。迄今为止，鉴定可用于早期发现和治疗TNBC患者的TNBC起始生物标志物仍然是一个未满足的临床需求。这些问题的科学和临床重要性已经在乳腺癌研究中广泛使用动物。基因工程小鼠模型（GEMM）和带有基因修饰细胞的异种移植小鼠模型目前代表了研究癌症起始的金标准，因为它们允许分析由突变驱动的早期致癌事件，并密切模仿人类肿瘤的组织病理学和分子特征。因此，大量的乳腺癌研究使用了这些模型（在过去的五年里每年约有1220项研究），因此每年估计需要24000多只动物。我们打算在本研究项目中使用的方法提供了一种新的方法来研究TNBC开始时特定基因突变的功能，这将取代GEMM和异种移植模型的使用。我们的系统包括将与TNBC相关的遗传变化引入乳腺细胞的正常诱导多能干细胞中，以在体外3D乳腺类器官模型中重建癌症的发展。这项技术不涉及使用动物来扩展细胞，也不使用动物产品来创建3D模型。事实上，该模型将使用一种合成水凝胶，它将提供一种优越的基质，而不是来自动物肿瘤，以研究肿瘤细胞与其环境的相互作用。重要的是，这种先进的细胞模型提供了优于其他癌症患者来源的培养细胞系统的优势，因为它允许模拟癌症起始，这对于针对早期疾病的治疗策略的发展至关重要。我们估计，这种方法将减少动物模型的使用，至少300只小鼠/年，全球约7000只小鼠/年。合成水凝胶的使用还将减少使用动物源性基质进行类器官分化研究，在我们的实验室中估计每年减少约40只小鼠。该项目还将为开发其他类型癌症的类似模型提供基础，从而在癌症研究中全面取代动物研究方面产生重大影响和遗产。重要的是，创建这个独特的模型将提供关于这种疾病的新知识，以及它在不同TNBC患者中的差异，从而开发精确的治疗方法，提高患者的生存率。

This project aims to develop a novel complex human cell model to study breast Triple Negative Breast Cancer (TNBC), whilst reducing the use of animals needed for the research. TNBC represents the most aggressive breast tumour subtype, typically presenting with high grade, high metastatic potential, and poor prognosis. TNBC remains the most clinical challenging type of breast cancer as no effective targeted therapies are available. Progress is further hindered by TNBC heterogeneity as TNBC is also a not a single disease, but further subdivided into distinct molecular and clinical subtypes based on specific molecular pathway signatures and mutational landscapes that impact on patient outcome. To date, the identification of biomarkers of TNBC initiation that can be used for early detection and treatment of TNBC patients remains an unmet clinical need.The scientific and clinical importance of these questions has seen a wide use of animals in breast cancer research. Genetically engineered mouse models (GEMM) and xenotransplantation mouse model with genetically modified cells currently represent the gold standard for studying cancer initiation as they allow the analysis of early oncogenic events driven by mutations and closely mimic the histopathological and molecular characteristics of human tumours. Consequently, a large number of breast cancer studies use these models (~1,220 studies/year in the last five years), thus requiring an estimated number of more than 24,000 animals per year. The approach we intend to use in this research project provides a novel way to investigate the function of specific genetic mutations in the initiation of TNBC, which will replace the use of GEMM and xenograft models. Our system involves introducing genetic changes associated with TNBC into normal induced pluripotent stem cells made for mammary cells to recreate the development of cancer in an in vitro 3D breast organoid model. This technology does not involve using animals to expand the cells and it does not use animal products to create the 3D model. Indeed, the model will use a synthetic hydrogel that will provide a superior matrix not derived from animal tumours to study interaction of tumour cells with their environment. Importantly, this advanced cell model provides advantages over other cancer patient-derived cultured cell systems as it allows modelling cancer initiation which is critical for the development of therapeutic strategies to target early-stage disease. We estimate that this approach will reduce animal model use by at least ~300 mice/year locally and ~7,000 mice /year globally. The use of the synthetic hydrogel will also reduce the use of animal-derived matrix for differentiation studies using organoids, with an estimated reduction of ~40 mice/year in our laboratory. The project will also provide the foundations to develop similar models for other types of cancer, therefore creating a significant impact and legacy on the overall replacement of animals in cancer research. Importantly, creating this unique model will provide new knowledge of the disease and how it differs in different TNBC patients for the development of precise therapies and increased patients' survival.