High-risk neuroblastoma is an aggressive and invasive paediatric malignancy, with few actionable somatic mutations. As such, intense multi-agent chemotherapy remains the standard-of-care. Failure to effectively activate apoptosis, or the ability to evade apoptosis, has been established as a key mechanism of chemoresistance in neuroblastoma. Despite this, there is little understanding of the apoptotic mechanism-of-action of individual standard-of-care chemotherapeutic agents, let alone their combined mechanism of action. Here we apply a network-wide, systems level approach to identify drug-specific apoptotic signalling axes which will inform patient-specific, synergistic drug combinations.
A functional genomics screen was performed on a high content cellomics platform with a siRNA library of 200 apoptotic genes with current standard-of-care chemotherapy and preclinical drugs. Multi-dimensional analysis of this dataset elegantly demonstrated that synergy between any two chemotherapy drugs is proportional to the magnitude of divergence in apoptotic signalling between individual drugs. Identified drug-specific apoptotic signalling nodes were validated using genetically engineered stable cell lines harbouring fluorescent biosensors and/or CRISPR-Cas9 mediated endogenously tagged proteins on our high content cellomics platform. These tools have allowed us to perform high-throughput kinetic live cell analysis at the single cell resolution and will help establish how synergy arises due to differential apoptotic signalling at the single cell level.
The application of our systems biology approach to rationalise mechanism-based drug selection will address fundamental questions about the network level functioning of apoptotic signalling pathways which has clinically relevant implications. Our data has demonstrated that it is differences in single agent drug-induced apoptotic signalling that will give rise to synergistic drug combinations. This is contrary to the current dogma of utilising drugs with different molecular targets in combination chemotherapy. This research will inform the development of precision medicine approaches with the aim to improve patient outcomes for high-risk neuroblastoma.