Spatiotemporal characterization of extrachromosomal DNA transcription and its key regulators in cancer cells

Extrachromosomal circular DNA (ecDNA) present in cancer cells stochastically amplifies oncogenes and drug resistance genes independent from the core genome and thereby contributes to tumor adaptability and heterogeneity.

While ecDNA prevalence, diversity, and biological traits have been studied, we lack insights into their expression regulation.

It is unclear how it differs from chromosomal genes and how it is linked to the non-random and dynamic nuclear localization patterns observed for many ecDNA.

Moreover, we are unaware of the proteins that participate in interactions with nuclear compartments to control ecDNA transcription. Most studies using sequencing and FISH methods fall short in describing these dynamic processes.

However, understanding their implications is vital for designing therapies that target ecDNA expression as a whole instead of targeting individual oncogenes.

To advance our knowledge, we need tools to map ecDNA transcription at high temporal and spatial resolution and systematic methods to screen for associated proteins.

The aim of this project is to identify crucial overarching mechanisms and players of ecDNA transcription through an interdisciplinary approach.

The Medema group has devised protocols to induce extrachromosomal amplification of drug resistance genes in diverse cellular backgrounds and genetically tag them for imaging and perturbation.

The present study will build on this work to for the first time reveal the spatiotemporal co-regulation of ecDNA transcription and localization using innovative imaging reporters for measurements in single live cells and use unbiased proteomic profiling to map factors that interact with ecDNA and regulate their expression.

Besides addressing outstanding questions in the field, our study will create unique cell models and datasets as a basis for future studies.

Our results will furthermore inform the design of more effective treatments for the many cancer patients affected in Europe and worldwide.