Codon specific readthrough as a mechanism of BRCA2 restoration in acquired PARP inhibitor and chemotherapy resistance

Area of Science:

• Oncology
• Molecular Biology
• Genetics

Background:

• BRCA2 mutations are key in certain cancers (ovarian, breast, prostate, pancreatic) and influence treatment sensitivity.
• Therapy resistance in these cancers is incompletely understood, though secondary mutations restoring BRCA2 are known.
• Understanding resistance mechanisms is crucial for improving cancer treatment strategies.

Purpose of the Study:

• To investigate novel mechanisms of drug resistance in BRCA2-mutated cancers.
• To identify causes of resistance to poly(ADP-ribose) polymerase inhibitors (PARPis) and cisplatin beyond secondary mutations.
• To elucidate the role of translational readthrough in acquired therapy resistance.

Main Methods:

• Utilized PARPi selection on BRCA2-mutated ovarian cancer cells (PEO1).
• Employed immunoblotting and tandem mass spectrometry to detect full-length BRCA2 protein.
• Conducted reporter assays to assess UAG-selective readthrough.
• Performed custom proteomic data searching and multi-omic analysis of cellular machinery.

Main Results:

• Identified translational readthrough of a premature termination codon (PTC) as a resistance mechanism.
• Detected low-level full-length BRCA2 protein in resistant cells despite the persistent BRCA2 mutation.
• Demonstrated UAG-selective readthrough in resistant clones and identified broader stop codon readthrough in other proteins.
• Multi-omic analysis revealed alterations in nonsense-mediated decay and termination pathways favoring readthrough.

Conclusions:

• Translational readthrough of PTCs is a novel mechanism driving drug resistance in BRCA2-mutated cancers.
• This mechanism allows for the production of functional BRCA2 protein, conferring resistance to PARPis and cisplatin.
• Targeting or understanding these readthrough events could offer new therapeutic avenues for resistant cancers.