Developing a chick embryo model to aid in development of personalised therapies for malignant pleural mesothelioma

间皮瘤是一种侵袭性的、基本上无法治愈的肺内膜癌，主要由环境暴露于石棉引起。迫切需要新的治疗方法或新的治疗方法。我们现在可以从一个病人癌症的小样本中读取有关基因变化的详细信息，然后可以用来决定最有效的抗癌药物，作为“精准医疗”给个别病人。最近的研究揭示了间皮瘤中发生的遗传变化的类型和频率，这可能有助于预测新的治疗方法。在许多癌症中，基因变化开启了“致癌基因”，加速了癌细胞一分为二的速度，推动了肿瘤的生长。许多癌症治疗使用直接阻断致癌基因活性的药物来防止这种不受控制的肿瘤生长。然而，间皮瘤是不寻常的，因为没有常见的致癌基因突变。相反，基因变化主要发生在“肿瘤抑制”基因上，使通常会起到抑制细胞分裂从而阻止肿瘤生长的蛋白质失效。这对寻找治疗间皮瘤的方法提出了一个困难的挑战，因为我们需要充分了解每个特定的肿瘤抑制基因突变是如何改变间皮瘤细胞的癌变行为的，以便找到我们可能能够用药物靶向的致命弱点。最终，我们还需要开发出最好的实验室模型来测试这些药物，然后才能将它们用于间皮瘤患者。在一半以上的间皮瘤中发现了肿瘤抑制因子BAP1的致残性突变。正常情况下，BAP1控制细胞内其他蛋白质的产生和破坏。因此，在没有BAP1的间皮瘤中，许多不同蛋白质的数量可能会发生变化，从而影响癌症行为。使用BAP1基因编辑突变的细胞，我们发现了许多这些蛋白质的变化。我们发现BAP1突变不仅影响改变癌细胞生长的蛋白质，还影响控制癌细胞如何移动、进入血管和在体内扩散的蛋白质。我们目前正在评估哪些蛋白质使间皮瘤细胞对特定的抗癌药物更敏感。然而，我们需要在能够提供人类间皮瘤生长和扩散的良好复制品的模型中测试这些药物。为此，我们将开发一种鸡胚胎间皮瘤模型，作为目前使用的小鼠模型的替代品。根据《动物科学程序法》，鸡胚胎模型被列为不受保护的动物，因此是一种取代动物试验的有用技术。与鼠标模型相比，它有许多额外的优势，包括成本效益、可访问性和速度。这是研究肿瘤细胞生长和扩散的一个很好的模型，因为它们可以很容易地移植到“绒毛膜尿囊膜”上。这是一个可接近的表面，位于鸡胚的外面，直接在蛋壳下面，有良好的血管供应。几天之内，一个小肿瘤就会形成，它可以扩散到细胞膜上，有可能进入血管，扩散到特定的器官。重要的是，新的药物治疗可以很容易地在鸡胚胎模型中进行测试，肿瘤细胞随着时间的推移进行成像，以评估它们的生存和行为。我们将使用鸡胚胎模型来培养间皮瘤细胞，有和没有BAP1突变，并评估我们的候选药物的治疗反应。成功的结果将建议将新药纳入间皮瘤患者的精准医学试验。在项目期间，我们将制定第一个标准操作程序，以在该模型中产生和监测间皮瘤肿瘤。我们将把这些方案和关键试剂提供给间皮瘤研究界，鼓励广泛更换小鼠模型。

Mesothelioma is an aggressive and largely untreatable cancer of the lung lining, mainly caused by environmental exposure to asbestos. New treatments, or new approaches to treatment, are urgently required. We can now read detailed information about genetic changes from a small sample of a patient's cancer, which can then be used to make decisions about the most effective anti-cancer drugs to give to an individual patient as "precision medicine". Recent studies have revealed the type and frequency of genetic changes that occur in mesothelioma, which may help in predicting new treatments.In many cancers, genetic changes switch on "oncogenes", which accelerate the speed with which cancer cells divide into two, driving tumour growth. Many cancer treatments use drugs that directly block the activity of oncogenes to prevent this uncontrolled tumour growth. However, mesothelioma is unusual, as there are no common oncogene mutations. Instead, genetic changes mostly occur in "tumour suppressor" genes, disabling proteins that would normally apply a brake to slow down dividing cells and so prevent tumour growth. This presents a difficult challenge for finding ways to treat mesothelioma, as we need to fully understand how each specific tumour suppressor mutation alters the cancerous behaviour of mesothelioma cells, in order to find an Achilles' heel that we might be able to target with drugs. Ultimately, we also need to develop the best laboratory models in which to test the drugs, before they can be given to mesothelioma patients. Disabling mutations of the tumour suppressor BAP1 are found in more than half of all mesotheliomas. Normally, BAP1 controls the production and destruction of other proteins within the cell. Therefore, in mesothelioma without BAP1, there are potentially changes in the amounts of many different proteins that could affect cancerous behaviour. Using cells with gene-edited mutations of BAP1, we identified many of these protein changes. We found that BAP1 mutation not only affects proteins that alter the growth of cancer cells, but also proteins that control how they move, gain access to blood vessels, and spread around the body. We are currently evaluating which of these proteins make mesothelioma cells more sensitive to specific anti-cancer drugs. However, we need to test these drugs in models that can provide a good replica of human mesothelioma growth and spread.To do this, we will develop a chick embryo model of mesothelioma, as a replacement for currently used mouse models. The chick embryo model is classified as non-protected under the Animals Scientific Procedures Act, and so is a useful technique to replace testing in animals. It has many additional advantages over mouse models, including cost effectiveness, accessibility and speed. It is an excellent model to study the growth and spread of tumour cells, as they can be easily engrafted onto the "chorioallantoic membrane". This is an accessible surface, located outside the chick embryo directly beneath the eggshell, with a good supply of blood vessels. Within a few days, a small tumour develops, which can spread across and into the membrane, potentially accessing blood vessels to spread to specific organs. Importantly, new drug treatments can be readily tested in the chick embryo model, and the tumour cells imaged over time to assess their survival and behaviour. We will use the chick embryo model to grow mesothelioma cells, with and without BAP1 mutation, and evaluate therapeutic responses to our candidate drugs. Successful outcomes will suggest new drugs for inclusion in precision medicine trials in mesothelioma patients. During the project, we will develop the first standard operating procedures to generate and monitor mesothelioma tumours in this model. We will make these protocols, and key reagents, available to the mesothelioma research community, encouraging widespread replacement of murine models.