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

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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.