Speaker:
Corinne Molyneux, University College London
Abstract:
Colorectal cancer (CRC) displays high levels of nongenetic phenotypic plasticity, allowing cancer cells to dynamically shift between different functions, enabling therapy evasion. This is particularly evident in poor prognosis, stromal-rich CRC tumours. These tumours also have an elevated presence of revival cancer stem cells (revCSCs). In the microenvironment of these tumours cancer-associated fibroblasts (CAFs) act as dynamic regulators driving the reversible acquisition of YAP-dependant, slow-cycling, chemoresistant features that define the revCSC state. The soluble factors that are regulating this transition were however undefined.
To resolve this question, we developed a high-throughput single-cell CRISPR assay to systematically perturb the CAF secretome when in direct co-culture with patient derived organoids (PDO). Individual components of the secretome were targeted by synthetic gRNAs in Cas9 expressing CAFs before co-culture. The perturbed co-cultures were then assessed via multiplexed mass cytometry, capturing cell fate markers, post-translational modification (PTM) signalling, cell-cycle activity, and apoptosis in both PDOs and CAFs at single-cell resolution.
Using this platform we have successfully targeted 202 ligands in our PDO+CAF model and quantified the role of each in the revCSC transition, unveiling the primary effector of stromal-induced CRC plasticity changes. These findings demonstrate that CRC phenotypic plasticity is regulated by stromal-derived signals, providing new mechanistic insight into therapy failure and describing a potential therapeutic intervention strategy to overcome plasticity driven chemoresistance.
