Comprehensive cell contact tracing (C3T)

In multicellular organisms from worms to humans, cell-cell communication is essential for proper development and homeostasis. Cells are equipped with a plethora of receptors, which relay physical interactions at the cell surface into the cell nucleus, resulting in a molecular and subsequently often phenotypic change of the cell. Interactions with cells in their surroundings also critically influence the behavior of diseased cells (including cancer), and these cell s in turn are known to influence their environment. Although of utmost importance, it has remained difficult to investigate cell-cell interactions and specifically resulting cellular changes at scale and in a natural setting. To overcome this hurdle, we will leverage recent developments in the fields of synthetic biology, microfluidic devices, in vivo imaging, and computational biology, and use these technologies for comprehensive cell contact tracing (C 3T). Working with zebrafish as an experimentally amenable model system, C3T will use cells that have been genetically engineered such that selected cell contacts elicit a response that enables a molecular recording of these interactions. A query cell will carry an activator for this receptor. Thus, contacts with this query cell (and only with this cell) will be engraved in interacting cells. This makes it possible to explore cell contacts throughout the entire zebrafish organism under the microscope or using high-throughput molecular assays. In our pilot project, we will use C3T to follow the history of cell interactions of tumor cells through the body on their way to forming metastases. As metastasis remains the leading cause of cancer- related deaths, but the process is intrinsically difficult to study outside the endpoints (i.e., the primary tumor and the metastasis), this proof-of-principle application may open up an entirely new way of channeling research into transient phases of this process (mobilization, extravasation, etc.). If successful, our approach will also be transferrable to other model organisms and research questions.

In multicellular organisms from worms to humans, cell-cell communication is essential for proper development and homeostasis. Cells are equipped with a plethora of receptors, which relay physical interactions at the cell surface into the cell nucleus, resulting in a molecular and subsequently often phenotypic change of the cell. Interactions with cells in their surroundings also critically influence the behavior of diseased cells (including cancer), and these cells in turn are known to influence their environment. Although of utmost importance, it has remained difficult to investigate cell-cell interactions and specifically resulting cellular changes at scale and in a natural setting. To overcome this hurdle, we attempted to implement a system for comprehensive cell contact tracing (C3T). We worked with zebrafish as an experimentally amenable model and generated new strains that enabled molecular recording of cell-cell interactions between "sender cells" (for which we used human cancer cell lines) and "receiver cells" (all zebrafish cells). Interactions between these cells were to elicit the expression of fluorescent proteins that would enable making them visible under the microscope and for detailed molecular follow-up studies. Initial attempts to implement the C3T system using published methods yielded unsatisfactory results in zebrafish: either cell contacts did not produce a reliable response or the fluorescence, which was meant to mark only contacted cells, was found to be not specific enough. In a laborious and time-consuming effort, we continued to test and evaluate different alternative genetic constructs by re-engineering components of the system and making use of latest breakthroughs in the community, yet with limited success. In the final phase of the project, we eventually obtained promising results with a new prototype system, which now forms the foundation for further experimentation. While our project was ongoing, the field advanced considerably with multiple new systems and applications published in scientific literature. This illustrates the great interest and potential of the technology, which is not limited to the tracing of cell contacts but might even allow for activation of contact-specific cellular responses. This might be an important enabling technology for future generations of interventive medicine.