Development of a New Human Model of Lung Squamous Cell Carcinoma Progression

肺鳞状细胞癌（LUSC）是一种毁灭性的疾病，约占英国所有癌症死亡率的5%，与吸烟密切相关，缺乏靶向治疗。检测早期LUSC为治疗成功提供了最佳机会，因为这些患者有资格接受根治性手术（60%的5年总生存率）。然而，75%的LUSC患者被诊断为晚期疾病，并接受化疗和最近的免疫治疗，但治疗获益有限（5年总生存率为6%）。这些令人沮丧的数字突出了改善LUSC药物的必要性。这只能通过开发更具创新性和雄心勃勃的研究模型来实现，这些模型反映了LUSC的遗传复杂性，并概括了从癌前阶段到浸润性癌症的进展。不幸的是，驱动这种进展的机制仍然不清楚，这阻碍了开发这些急需的患者相关模型的努力。LUSC的基因组很复杂，患者之间存在差异，但仔细观察显示，大多数患者都发生了一些遗传改变。此外，低频率的遗传改变通常针对相同的细胞过程，并在肿瘤中按照周期性的时间顺序出现，从而产生一系列“遗传阶段”。这些共性可以用来设计更相关的LUSC模型。然而，在小鼠中进行这种建模工作将需要大量的动物，因为涉及的基因数量和患者中发现的遗传多样性。我们癌症生物学家必须努力避免这种小鼠生命的代价，并在尽可能多的转化应用中找到替代动物的策略，以取代LUSC研究中的动物。人类基底细胞（HBCs）是LUSC的起源细胞，可以很容易地在体外培养，更有可能更好地反映人类细胞生物学。因此，HBCs是一种有吸引力的和通用的系统，以研究这种疾病，并取代小鼠在翻译LUSC研究。在这项工作中，我们打算设计HBC以重现LUSC患者中发现的遗传阶段和多样性。在遵循这一策略，我的目标是建立一个模型的LUSC进展，最好的概括谱的癌前病变和侵袭性LUSC阶段，以取代mouse models.The第一个目标的这个项目是基因工程HBCs复制一系列的遗传阶段中观察到的患者。改变的选择将基于其在重要LUSC途径中的高频率和已知作用。第二个目标是分析每个遗传阶段的细胞变化，包括侵袭性，细胞增殖和组织结构的变化。最后一个目标是研究基因表达、免疫调节因子和肿瘤内异质性的变化。在完成验证阶段后，我们的目标是获得一个全面的数据集，提供驱动LUSC进展的生物学变化的最完整的知识，以及一个易于处理的LUSC模型，以扩大我们的临床前研究机会。这种建模策略最具创新性的方面是a）使用人类细胞而不是小鼠模型，B）旨在将肿瘤演变纳入癌症建模策略的遗传阶段重演，以及c）强调癌前阶段。这些功能将扩大我们的能力，以阶段依赖性的方式研究LUSC生物学的特定领域，这是使用现有临床前模型无法实现的，同时避免了小鼠和人类之间肺生物学的重要差异。在完成模型的验证后，我们打算专注于癌前阶段的生物学，新的癌症脆弱性和免疫微环境中的阶段依赖性变化。这些研究领域将加速开发新的早期检测方法，新的治疗方式，并改善现有的方法。

Lung squamous cell carcinoma (LUSC) is a devastating disease which accounts for approximately 5% of all cancer mortality in the UK, is strongly associated with smoking and lacks targeted therapies. Detecting early stage LUSC provides the best chance of therapeutic success as these patients are eligible for curative surgery (60% 5-year overall survival). However, 75% of LUSC patients are diagnosed with advanced disease and are treated with chemotherapy and, more recently, immunotherapy but with limited therapeutic benefit (6% 5-year overall survival).These dismal figures highlight the need to improve LUSC medicine. This can only be achieved by developing more innovative and ambitious research models that reflect the genetic complexity of LUSC and recapitulate progression from premalignant stages to invasive cancer. Unfortunately, the mechanisms driving this progression remain unclear and this hinders efforts to develop these much-needed patient-relevant models.The genomes of LUSC are complex and differ among patients, but a closer look shows that some genetic alterations occur in most patients. Additionally, low-frequency genetic alterations often target the same cellular processes and arise in the tumour following a recurrent temporal order that gives rise to a series of 'genetic stages'. These commonalities can be exploited to design more relevant models of LUSC. However, carrying out this modelling effort in mice would require large cohorts of animals due to the number of genes involved and the genetic diversity found in patients. We cancer biologists must try to avoid this cost in mouse lives and find alternative strategies to replace animals in LUSC research in as many translational applications as possible.Human basal cells (HBCs) are the cell-of-origin of LUSC, can be easily cultured in-vitro and are more likely to reflect better the human cell biology. Therefore, HBCs are an attractive and versatile system to investigate this disease and replace mice in translational LUSC studies. In this work, we intend to engineer HBCs to reproduce the genetic stages and the diversity found in LUSC patients. In following this strategy, I aim to build a model of LUSC progression that best recapitulates the spectrum of premalignant and invasive LUSC stages to replace mouse models.The first objective of this project is to genetically-engineer HBCs to reproduce the series of genetic stages observed in patients. The selection of alterations will be based on their high frequency and known role in important LUSC pathways. The second objective will be to analyse the cellular changes in each genetic stage including invasiveness, cell proliferation and changes to tissue architecture. The last objective will be to investigate changes in gene expression, immuno-modulatory factors, and intra-tumour heterogeneity. On completing the validation stage, we aim to have acquired a comprehensive dataset providing the most complete knowledge of the biological changes driving LUSC progression as well as a tractable LUSC model to expand our preclinical research opportunities. The most innovative aspects of this modelling strategy are a) the use of human cells instead of mouse models, b) the recapitulation of genetic stages that aims to incorporate tumour evolution into cancer modelling strategies and c) the emphasis on premalignant stages. These features will broaden our capabilities to investigate specific areas of LUSC biology in a stage-dependent manner that cannot be undertaken using existing preclinical models, while avoiding the important differences in lung biology between mouse and human. After completing the validation of the model, we intend to focus on the biology of premalignant stages, new cancer vulnerabilities and stage-dependent changes in the immune microenvironment. These research areas will accelerate the development of new early detection methods, novel therapeutic modalities, and the improvement of existing ones.