Automated high throughput oncology drug screening in zebrafish using robotics
PURPOSE: Cancer is a leading cause of deaths world-wide. For a large number of cancer patients there is no satisfactory treatment regime available. Moreover, many of the treatment strategies currently in use in the clinic bring with them a vast amount of short- and long-term side effects. There is an urgent need for alternative therapeutic strategies, ideally tailored to each individual patient. With this project, we will validate newly developed injection and imaging robots designed to automate oncology drug screening in zebrafish. After determining optimal injection conditions, we will perform a proof-of-concept drug screen for glioblastoma and diffuse intrinsic pontine glioma (DIPG) cancers with 25 compounds, using immortalised lines as well as primary patient material. Additionally, we will study how these brain tumor cells interact with their microenvironment and vice versa. In particular, we are interested in the induction of seizures by tumor cells, as well as the effect of seizures on tumor cell proliferation.
BENEFIT: Zebrafish xenograft models offer a promising solution for drug discovery owing to their small size, low cost of care and rapid development of optically transparent larvae. They enable screening of compounds against human tumours in the context of a living organism. However, currently the application of these models is limited due to labour intensive workflows by experienced researchers, which cause incompatibility with the high-throughput demands of drug screening. The robots to be validated within this project will optimize sample handling and automate organ-specific injection and imaging, enabling high-throughput drug screening with zebrafish larvae up to 12 dpf. Additionally, we will validate this pipeline for implantation of primary patient material. When succesful, the injected larvae may act as a personalised avatar for cancer patients in which to test personalised treatment regimes.
REPLACEMENT, REDUCTION AND REFINEMENT: By choosing zebrafish, we replace rodents with a lower sentient vertabrate. In the early stages of development of the new robotic technology, zebrafish will not be kept beyond 120hpf. We will refine our protocols based on <120hpf outcomes, to reduce the numbers of older animals recuired for statistical significance.
DISTRESS: Animals will be monitored throughout the experiments and euthanized prior to reaching defined humane endpoints. As described by NORECOPA, animals are expected to experience moderate discomfort due to to the presence of localized tumors. Any pharmacological treatment will be supplied via immersion in a diluted compound/E3 mix, and is estimated to lead to a mild discomfort.
NUMBER OF ANIMALS: In this project, there will be a maximum of 21485 larvae used, up to 12 dpf.
BENEFIT: Zebrafish xenograft models offer a promising solution for drug discovery owing to their small size, low cost of care and rapid development of optically transparent larvae. They enable screening of compounds against human tumours in the context of a living organism. However, currently the application of these models is limited due to labour intensive workflows by experienced researchers, which cause incompatibility with the high-throughput demands of drug screening. The robots to be validated within this project will optimize sample handling and automate organ-specific injection and imaging, enabling high-throughput drug screening with zebrafish larvae up to 12 dpf. Additionally, we will validate this pipeline for implantation of primary patient material. When succesful, the injected larvae may act as a personalised avatar for cancer patients in which to test personalised treatment regimes.
REPLACEMENT, REDUCTION AND REFINEMENT: By choosing zebrafish, we replace rodents with a lower sentient vertabrate. In the early stages of development of the new robotic technology, zebrafish will not be kept beyond 120hpf. We will refine our protocols based on <120hpf outcomes, to reduce the numbers of older animals recuired for statistical significance.
DISTRESS: Animals will be monitored throughout the experiments and euthanized prior to reaching defined humane endpoints. As described by NORECOPA, animals are expected to experience moderate discomfort due to to the presence of localized tumors. Any pharmacological treatment will be supplied via immersion in a diluted compound/E3 mix, and is estimated to lead to a mild discomfort.
NUMBER OF ANIMALS: In this project, there will be a maximum of 21485 larvae used, up to 12 dpf.