Single-Cell Heterogeneity of EGFR Pathway Is Linked to Unique Signatures of Drug Response and Malignancy in Patient Derived Glioblastoma Stem Cells.

Purpose: In glioblastoma, the therapeutically intractable and resistant phenotypes can be derived from glioma stem cells, which often have different underlying mechanisms from non-stem glioma cells. Aberrant signaling across the EGFR-PTEN-AKT-mTOR pathways have been shown as common drivers of glioblastoma. Revealing the inter and intra-cellular heterogeneity within glioma stem cell populations in relations to signaling patterns through these pathways may be key to precision diagnostic and therapeutic targeting of these cells.

Materials and methods: Single cell parallel proteomic heterogeneity profiling of the EGFR-PTEN-AKT-mTOR pathways was conducted in a panel of fifteen glioma stem cell models derived from patient glioblastoma biopsies.

Results: The analysis included 59,464 data points from 14,866 cells and identified forty-nine molecularly distinct signaling phenotypes. High content bioinformatics resolved two unique patient clusters diverging on EGFR expression and AKT/TORC1 activation. Phenotypic validation indicated drug responsive phenotypes to EGFR blocking in the high EGFR expressing cluster with lower tumor initiating potential in comparison to the AKT/TORC1 activated cluster. High EGFR expression trended with improved patient prognosis while AKT/TORC1 activated samples trended with poorer patient outcomes. Genetic heterogeneity was observed in both clusters with proneural, classical and mesenchymal subtypes observed.

Conclusion: Quantitative single cell heterogeneity profiling reveals divergent EGFR-PTEN-AKT-mTOR pathways of patient derived glioma stem cells, which would inform future research and personalized therapeutic strategies.