Inhibition of ATR opposes glioblastoma invasion through disruption of cytoskeletal networks and integrin internalisation via macropinocytosis

Background

Glioblastomas have highly infiltrative growth patterns that contribute to recurrence and poor survival. Despite infiltration being a critical therapeutic target, no clinically useful therapies exist which counter glioblastoma invasion. Here, we report that inhibition of Ataxia telangiectasia and Rad 3 related kinase (ATR) reduces invasion of glioblastoma cells through dysregulation of cytoskeletal networks and subsequent integrin trafficking.

Methods

Glioblastoma motility and invasion were assessed in vitro and in vivo in response to ATR inhibition (ATRi) and ATR over-expression using timelapse microscopy, two orthotopic glioblastoma models and intravital imaging. Disruption to cytoskeleton networks and endocytic processing were investigated via high throughput, super resolution imaging and intravital imaging.

Results

High ATR expression was associated with significantly poorer survival in clinical datasets whilst histological, protein expression and spatial transcriptomics using glioblastoma tumour specimens revealed higher ATR expression at infiltrative margins. Pharmacological inhibition with two different compounds and RNAi targeting of ATR opposed invasion of glioblastoma, whereas overexpression of ATR drove migration. Subsequent investigation revealed that cytoskeletal dysregulation reduced macropinocytotic internalisation of integrins at growth cone-like structures, resulting in a tumour microtube retraction defect. The biological relevance and translational potential of these findings was confirmed using two orthotopic in vivo models of glioblastoma and intravital imaging.

Conclusion

We demonstrate a novel role for ATR in determining invasion in glioblastoma cells and propose that pharmacological targeting of ATR could have far reaching clinical benefits beyond radiosensitisation.