Engineering Circadian Biology into Induced Pluripotent Stem Cell Organ-on-a-Chip Models

Lead Research Organisation: Queen Mary University of London


Organ-on-a chip (OOAC) devices are an emerging tool for studying how organs function and for testing new and repurposed drugs in a laboratory environment. Each chip contains hollow channels containing combinations of human cells that interact with each other to mimic the functioning of our organs. Mechanical forces can be applied to replicate the physical forces that cells experience in our tissues and organs, for example fluid flow and stretch for the heart and blood vessels. Ultimately OOAC systems may be used to reduce the use of animals in research and ultimately to replace routine use of animals altogether.

All tissues in the body have an internal biological circadian 'clock' which co-ordinates cellular processes in our tissues and organs according to the time of day or night, creating cycles of biological function every 24h. Importantly drug effectiveness and toxicity often varies according to the time of administration, which means that there may be times of day when a drug is most effective and produces the least side effects. However, for the majority of treatments, the influence of biological time on therapeutic effectiveness has not been studied or exploited. This is partly because the early stages of a drug development, involving simple cell systems and then animal models do not replicate the human circadian clock. Accordingly, the first time a novel drug encounters a relevant human circadian clock is during phase I clinical trials many years into the development process. This results in costly late-stage failure of drug discovery programmes and the unnecessary use of animals in earlier stages of development.

Currently circadian rhythms are not incorporated into OOAC technology and there is no means of establishing the biological time of cells within these devices, limiting the therapeutic and commercial potential of the technology. The overall aim of this project is, therefore, to incorporate the circadian clock into OOAC systems, allowing the cellular 'time of day' to be monitored and for drugs to be tested at different time points that reflect day and night for our organs.

In this project we will develop cell lines from Induced Pluripotent Stem Cells (iPSCs) that are able to 'report' the time of the circadian clock. This involves using the gene editing technique CRISPR to link a luminescent tag to key genes that control the clock. Cellular circadian rhythms can then be induced and monitored non-invasively. iPSCs can be differentiated into all cell types in the body allowing us to develop a library of novel human iPSC-derived differentiated clock reporter cells. In this project we will turn the iPSCs into endothelial cells which line our blood vessels and skeletal muscle cells.

The cells will be grown in OOAC devices and stimuli will be provided that 'set the time' of the clock resulting in the expression of clock genes, measured by luminescence, that will oscillate over a 24hr period. We will use a variety of stimuli to 'set the time' of the clock including chemical factors, such as growth factors and glucocorticoids, but also mechanical stimuli, such as fluid flow and stretch which are integral to OOAC devices.

Finally we will look at whether known drugs have a different effect on the cells depending on when it is administered during the 24hr circadian rhythm. The ability to incorporate biological time into OOAC systems will make the systems better able to replicate the functioning of organs in our bodies and enhance their ability to be used as laboratory systems to replace the routine use of animals in research.

Technical Summary

Organ-on-a-chip (OOAC) technologies are important emerging tools for evaluating the efficacy and safety of novel therapies and may ultimately reduce the requirement for animals in research. There is increasing interest in the use of induced pluripotent stem cell (IPSC)-derived differentiated cell models as validated OOAC systems for drug discovery, which would allow personalised systems to be developed. Physiological processes are co-ordinated by circadian clocks, that endow tissues with oscillations in gene expression with a period of ~24hr. Over 80% of proteins that are druggable targets are circadian and likely to benefit from timed administration. However, OOAC technologies do not currently incorporate circadian rhythmicity, which limits their physiological relevance and value as platforms for ex-vivo therapeutic evaluation.

Overall, this project will generate iPSC-OOAC technologies with circadian physiology, as a novel and powerful technology for use as disease models and to evaluate investigative therapies with the potential to reduce the use of animals in research and ultimately to replace routine use of animals.

Project aim: To create human iPSC-derived OOAC model systems expressing robust and sustained circadian rhythmicity achieved via the following objectives:
1. Modify human iPSCs using CRIPSR genome editing so that, upon differentiation using established protocols, they emit bioluminescence that reports circadian clock function.
2. Define the circadian mechanical timing cues that are necessary and sufficient to sustain robust circadian physiology within OOAC technology.
3. Determine the optimal time of day for the administration of exemplar therapeutic agents, as proof of concept of the technology application.


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