Refining in vivo studies of cancer metastasis with next-generation explant-in-chip perfusion models

利用下一代芯片外植体灌注模型完善癌症转移的体内研究

• 批准号: NC/X001210/1
• 负责人: Darryl Overby
• 金额: $ 66.59万
• 依托单位: Imperial College London
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=NC%2FX001210%2F1
• 关键词: Refining; vivo; studies; cancer; metastasis

Our overall goal is to create and validate a novel "explant-in-chip" perfusion device to study the early stages of metastasis, specifically cancer cell arrest, extravasation, and early colonisation within multiple secondary tissues of interest (brain, lung, liver). We estimate that at least 12,000 mice are used annually for metastasis research, and our device promises to reduce this number by providing a user-friendly approach to investigate the early stages of metastasis using ex vivo tissue, rather than performing in vivo trials for each experimental condition.The explants are several millimetres in size, which is too large to be supported by oxygen and nutrient diffusion alone, but large enough to encompass a significant portion of the tissue microenvironment and tissue-specific microvasculature architectures. To achieve effective oxygen and nutrient delivery, explants are perfused with nutrient-containing medium after being placed within a specially designed microchannel that achieves self-sealing around the explant. Sealing is necessary so that when a pressure-drop is applied across the explant, flow is forced to pass through the explant (per-fusion) as opposed to around the explant (peri-fusion). Our preliminary data demonstrate that perfusion preserves explant viability for at least 6 days. By including labelled cancer cells in the perfusate, the cells enter the vasculature and distribute throughout the microvascular network within the explant to mimic the hematogenous route of tumour cell dissemination in vivo.This approach is important from a 3Rs perspective because multiple explants can be isolated from an individual animal and studied within our device. This reduces the experimental unit from the individual animal to the individual explant. Because many explants can be isolated from a single mouse and exposed to a range of different treatment conditions, our device reduces animal numbers and maximises the scientific value of each animal used for research. Because ex vivo approaches can be used to screen and triage many experimental conditions, the approach allows for more refined follow-on in vivo studies once the optimal drugs or drug concentrations have been identified in ex vivo screens.Our device captures and preserves the native tissue microenvironment which has a critical role on early metastasis, while also providing a route for vascular infusion that is typically unavailable in other ex vivo models. Our approach also provides a means to visualise the spatiotemporal dynamics of early metastasis within the native tissue microenvironment, which in vivo would require intravital microscopy and implantation of an optical window.We aim to validate our device by comparing against gold standard in vivo models of experimental metastasis. We also aim to disseminate our technology for maximum impact on the 3Rs by making the design user-friendly and by hosting workshops to facilitate other laboratories using our device.By the end of this project, we will have developed and validated a user-friendly device that will provide single cell resolution of the early spatiotemporal events involved in metastatic dissemination, alleviating the need for technologically challenging and invasive procedures in mice.

我们的总体目标是创建并验证一种新的“外植体芯片”灌注装置，以研究转移的早期阶段，特别是癌细胞在多个次要组织（脑、肺、肝）中的阻滞、外渗和早期定植。我们估计每年至少有12,000只小鼠用于转移研究，我们的设备有望通过提供一种用户友好的方法来研究体外组织转移的早期阶段，而不是针对每种实验条件进行体内试验，从而减少这一数字。外植体的尺寸为几毫米，这太大了，无法单独由氧气和营养扩散支撑，但足够大，可以包含组织微环境和组织特异性微血管结构的很大一部分。为了实现有效的氧气和营养输送，外植体被放置在一个特殊设计的微通道中，在外植体周围实现自密封，然后灌注含有营养的培养基。密封是必要的，这样当在植体上施加压降时，流体被迫通过植体（预融合），而不是围绕植体（预融合）。我们的初步数据表明，灌注可使外植体存活至少6天。通过在灌注液中加入标记的癌细胞，细胞进入脉管系统并分布在外植体内的微血管网络中，以模拟肿瘤细胞在体内传播的血液途径。从3r的角度来看，这种方法很重要，因为可以从单个动物中分离出多个外植体，并在我们的设备中进行研究。这将实验单位从单个动物减少到单个外植体。由于许多外植体可以从一只老鼠身上分离出来，并暴露在一系列不同的处理条件下，我们的设备减少了动物数量，并最大限度地提高了用于研究的每只动物的科学价值。由于离体方法可用于筛选和分类许多实验条件，一旦在离体筛选中确定了最佳药物或药物浓度，该方法允许更精细的后续体内研究。我们的设备捕获并保存了在早期转移中起关键作用的原生组织微环境，同时也提供了其他离体模型通常无法获得的血管输注途径。我们的方法还提供了一种在原生组织微环境中可视化早期转移的时空动态的方法，这在体内需要活体显微镜和植入光学窗口。我们的目标是通过与体内实验转移模型的金标准进行比较来验证我们的装置。我们亦致力推广我们的技术，使设计更易于使用，并举办工作坊，方便其他实验室使用我们的设备，以最大限度地影响3r。到本项目结束时，我们将开发并验证一种用户友好的设备，该设备将提供涉及转移性传播的早期时空事件的单细胞分辨率，减轻对技术挑战性和侵入性手术的需要。