Time-resolved phenotypic profiling maps EGFR and c-Met trafficking signatures

Area of Science:

• Cell Biology
• Molecular Oncology
• Biochemistry

Background:

• Receptor tyrosine kinases (RTKs), including epidermal growth factor receptor (EGFR) and hepatocyte growth factor receptor (c-Met/MET), are crucial for cell signaling.
• Their signaling relies on endocytic routing, and impaired down-regulation contributes to tumor progression and therapy resistance.
• Understanding how RTK trafficking states influence signaling and lipid programs is vital, especially in cancer contexts.

Purpose of the Study:

• To compare the coupling of signaling and lipid programs to routing states between EGFR and c-Met under matched conditions.
• To investigate how late receptor persistence phenotypes relate to specific lipid trafficking and signaling biases.
• To establish a comparative imaging framework for mapping RTK trafficking states in cancer.

Main Methods:

• Utilized a chemically diverse annotated inhibitor library for targeted drug screening.
• Employed time-resolved, pathway-focused high-content imaging in A549 cells.
• Compared co-varying phenotypes across matched EGFR and c-Met assay systems to analyze receptor persistence and associated cellular responses.

Main Results:

• Late receptor persistence phenotypes were heterogeneous and dispersed across a fused phenotypic atlas.
• EGFR persistence correlated with reduced phosphatidylinositol 4,5-bisphosphate readouts and signaling biases.
• c-Met persistence was associated with increased perinuclear phosphatidylinositol 4-phosphate heterogeneity and altered transferrin-recycling readouts, demonstrating receptor-selective lipid-trafficking coupling.

Conclusions:

• A comparative imaging framework was established for mapping RTK trafficking states.
• Phosphoinositide-linked features are nominated as candidate readouts for trafficking rewiring in cancer.
• These findings provide insights into RTK signaling dynamics and potential therapeutic strategies targeting cancer progression and resistance.