CETSA interaction proteomics define specific RNA-modification pathways as key components of fluorouracil-based cancer...

Area of Science:

• Molecular Biology
• Cancer Pharmacology
• Proteomics

Background:

• Understanding cancer drug mechanisms of action (MoA) is crucial for optimizing treatment efficacy.
• The antimetabolite 5-fluorouracil (5-FU) is a widely used chemotherapy agent, but its complete MoA is not fully elucidated.
• Existing knowledge primarily links 5-FU to thymidylate synthase (TYMS) inhibition and DNA damage pathways.

Purpose of the Study:

• To investigate the comprehensive protein interaction landscape induced by 5-fluorouracil (5-FU) using a high-resolution proteome-wide cellular thermal shift assay (CETSA).
• To identify novel cellular pathways and proteins involved in the mechanism of action of 5-FU.
• To explore differences in protein responses in drug-resistant cancer models.

Main Methods:

• Application of a high-resolution proteome-wide cellular thermal shift assay (CETSA).
• Analysis of protein interaction changes upon treatment with 5-fluorouracil (5-FU) and related nucleosides.
• Comparison of protein responses between sensitive and resistant cancer cell models.

Main Results:

• Confirmed known effects of 5-FU on thymidylate synthase (TYMS) and pyrimidine metabolism enzymes.
• Identified a broad range of previously unrecognized protein interactions, particularly in RNA modification and processing pathways.
• Observed attenuated protein responses in a resistant cell model, highlighting key components of 5-FU's MoA.

Conclusions:

• 5-fluorouracil (5-FU) exerts its effects through a wider network of protein interactions than previously understood, significantly impacting RNA biology.
• The abrogation of thymidylate synthase (TYMS) inhibition is not essential for cell proliferation, suggesting alternative mechanisms of action for 5-FU.
• These findings provide a more comprehensive understanding of 5-FU's MoA, potentially informing the development of more effective cancer therapies.