Non-invasive imaging to reduce and refine the use of animals and monitor their welfare during the course of experimentations in oncology

尽管包括我们在内的国际社会作出了努力，但在实验动物模型中忠实和可靠地模拟人类癌症仍然是提高我们对这种疾病的理解和制定/评估更有效的治疗策略的基本要求。人们普遍承认，当癌细胞位于它们起源的器官中时，原位模型比异位模型更有信息量和可靠性(当癌细胞被迫在它们通常不会使用的地方生长时，比如在皮下)。然而，分析原位模型中的肿瘤进展是非常具有挑战性的，因为它们不能用肉眼看到。因此，需要在每个时间点处死一批动物，以分析解剖后的器官(在本项目中为骨髓和肺)。活体成像技术提供了令人兴奋的可能性，可以随着时间的推移进行非侵入性和纵向研究，以了解活动物在很长一段时间内的生物过程动力学。基本的想法是使用成像方法来量化肿瘤的数量及其随着时间的推移在动物体内的分布，以一种模仿人类患者诊断的方式。在这种情况下，没有必要牺牲受试者来获得这些测量结果，然后就可以评估癌细胞随时间的演变。因此，活体成像可以减少回答生物学问题所需的动物数量和/或新治疗的有效性。此外，不同的成像技术可以组合在同一动物身上，以便在同一时间回答不同的问题，而不是做不同的实验。从患者的样本中异种移植人急性髓系白血病(AML)启动细胞(AML-ICs)是了解人类白血病的发展和评估治疗反应的一个非常强大的模型。然而，为了随着时间的推移收集结果，有必要要么牺牲一些动物，要么进行骨髓活检，这是一种耐受性很好但具有侵入性的程序。我们可以使用生物发光(BLI)来追踪骨髓中的基因工程细胞系，但这不能用于患者的样本。据我们所知，到目前为止，还没有报道过非侵入性方法来监测原发人类AML的时间。我们建议在这里测试不同的策略来实现这一目标，这将允许减少完成该领域研究所需的动物数量。此外，能够客观地量化“肿瘤负荷”将使人们能够同时评估动物的健康状况，类似于为患者所做的事情。因此，研究人员将有能力改进他们的实验条件，并在获得所需答案后立即停止实验。我们还使用了基因工程小鼠模型，其中特定基因发生了突变，将诱导肺癌的自发显现，与人类一种特定类型的癌症非常相似。我们可以在小鼠身上使用X射线计算机断层扫描(CT)来检测肺部肿瘤，就像在人类身上做的那样，但我们正在努力开发更敏感的技术，并收集更多仅有结节大小的信息。因此，我们需要测试不同类型的探针(荧光标记的或与放射性示踪剂结合的)，这些探针可以用于通过现代活体成像技术(近红外荧光成像和切伦科夫发光成像)检测白血病/肺癌的存在和进展。我们还想测量生理参数(心率、呼吸频率、氧合)，以客观地评估动物在研究过程中的福利。总之，这些不同的信息将使我们能够减少对动物的需求，并改进实验中使用动物的方式

Despite the efforts of the international community, including ours, faithfully and reliably mimicking human cancers in experimental animal models remains a fundamental requirement to improve our understanding of the disease and develop / assess more effective therapeutic strategies. It is generally admitted that orthotopic models, when cancer cells are located in the organ they originate from, are more informative and reliable than ectopic models (when the cancer cells are forced to grow in place they would normally not use, like under the skin). However, it is very challenging to analyse the tumour progression in orthotopic models since they cannot be seen with naked eyes. As a consequence, a batch of animals needs to be sacrificed at each time point for analysing the organs after dissection (bone marrow and lungs in the case of this project). In vivo imaging technologies offer exciting possibilities to conduct noninvasive and longitudinal studies over time to understand the dynamics of biological processes in live animals over long period of time. The basic idea is to use imaging approaches to quantify the amount of tumours and their distribution within the body of the animal over time, in a way that mimics diagnosis for human patients. In that case there is no need to sacrifice the subject to obtain these measurements and it is then possible to assess the evolution of cancer cells over time. Therefore in vivo imaging allows to reduce the number of animals needed to answer a biological question and / or the efficacy of a new treatment. Furthermore, different imaging techniques can be combined on the same animal in order to answer different questions at the same time instead of doing different experiments.Xenotransplantation of human acute myeloid leukaemia (AML) initiating cells (AML-ICs) from samples coming directly from patients is a very powerful model to understand the development of human leukaemia and assess response to treatments. However to collect results over time, it is necessary either to sacrifice some animals, or to perform bone marrow biopsies, a well tolerated but sill invasive procedure. We can use bioluminescence (BLI) to track genetically engineered cell lines in the bone marrow, but this cannot be used for patients' samples. To our knowledge, no noninvasive approach has been reported so far to monitor primary human AML over time. We propose here to tests different strategies to achieve this goal, which would allow to decrease the number of animals needed to complete the studies in that field. Furthermore, being able to objectively quantify the "tumour load" would allow one to assess at the same time the health status of the animal, similarly to what is done for patients. Consequently, researchers would have the capability to refine their experimental conditions and stop the experiment as soon as they have obtained the answers they need.We also use genetically engineered mouse models where a specific gene has been mutated and will induce the spontaneous apparition of a lung cancer, closely mimicking a specific type of human cancer. We can use x-ray computed tomography (CT) in mice to detect lung tumours, like it is done in humans, but we are trying to develop more sensitive techniques and also to collect more information that just the size of the nodules.Therefore we need to test different types of probes (fluorescently labelled or coupled with radioactive tracers), which could be used to detect by modern in vivo imaging technologies (Near-infrared fluorescence imaging and Cerenkov luminescence imaging) the presence and the progression of the leukaemia / lung tumours. We also want to measure physiologic parameters (heart rate, breathing rate, oxygenation) to assess objectively the welfare of the animals during the course of the study.Altogether, these different information would allow us to reduce the need for animals and refine the way they are used during the experiments