Direct Visualization and Control of DNA Replication Kinetics in Single Cells and Cancer Tissues by X-MARK

ASPIRE Award (2021-Present)

Bennie Lemmens, PhD (Principal) and Jiri Bartek, MD, PhD, Karolinska Institutet (Sweden)

Uncontrolled cellular proliferation is one of the hallmarks of cancer. Because of this, cancer cells must replicate their DNA at extremely high rates and bypass the normal checkpoints which regulate cell turnover in normal tissue. Due to the complexity of DNA replication, it has largely been studied using model systems or unicellular organisms. Therefore, much remains to be learned about this process in human cells in more biologically relevant systems. Such systems would allow researchers to ask how the process and kinetics of DNA replication differ between normal cells and tumor cells, and how these differences can be leveraged into developing new therapeutic strategies for treating cancer.

Here Bennie Lemmens and Jiri Bartek will develop an ultrasensitive and versatile technology coined X-MARK (eXpansion Microscopy Assay for Replication Kinetics) to study DNA replication and cancer cell biology with unprecedented precision in time and space. Dr. Lemmens recently established state-of-the-art microscopy and DNA labelling techniques to study fundamental principles of human cell division, while Dr. Bartek has developed unique cancer models and DNA fiber technologies to define how hasty DNA replication drives genomic instability and therapy resistance. The team will detect DNA replication kinetics in single cells at nanoscale resolution by physically enlarging human genomes in their native cellular context. They will do so by combining three distinct technologies: direct DNA nucleotide labeling, super-resolution microscopy, and 3D tissue expansion. By developing these techniques and applying them to cancer systems, they will look gain a better understanding of DNA replication in cancer cells at an unprecedented spatial and temporal resolution.