Highly Selective PTK2 Proteolysis Targeting Chimeras to Probe Focal Adhesion Kinase Scaffolding Functions

Area of Science:

• Oncology
• Molecular Biology
• Biochemistry

Background:

• Focal adhesion tyrosine kinase (PTK2) is frequently overexpressed in human hepatocellular carcinoma (HCC).
• Previous studies suggest PTK2 inhibition or depletion reduces HCC tumorigenicity.
• The clinical utility of targeting PTK2 in HCC remains unconfirmed.

Purpose of the Study:

• To develop and characterize novel PTK2 proteolysis-targeting chimeras (PROTACs).
• To investigate the efficacy of PTK2 degradation using PROTACs in HCC cell lines.
• To assess whether PTK2 degradation by PROTACs recapitulates known anti-tumorigenic effects.

Main Methods:

• Design and synthesis of two selective PTK2 PROTACs employing von Hippel-Lindau and cereblon ligands.
• Evaluation of PTK2 degradation efficiency using a cereblon-based PROTAC (BI-3663) in 11 HCC cell lines.
• Assessment of cellular proliferation and phenotypic changes following PTK2 degradation.

Main Results:

• BI-3663 demonstrated potent PTK2 degradation with a median DC50 of 30 nM, achieving >80% degradation across tested HCC cell lines.
• Despite effective PTK2 protein depletion, the PROTACs did not induce the previously reported anti-proliferative effects.
• The study highlights a potential disconnect between PTK2 degradation and its functional consequences in HCC.

Conclusions:

• Novel PTK2-targeting PROTACs are effective tools for inducing protein degradation in HCC.
• Effective PTK2 degradation via PROTACs does not necessarily phenocopy the effects of genetic or pharmacological PTK2 inhibition.
• These compounds serve as valuable research tools for further investigating PTK2 biology and degradation pathways.