From In Silico to Experimental Validation: Tailoring Peptide Substrates for a Serine Protease

Area of Science:

• Biochemistry
• Nanotechnology
• Oncology

Background:

• Smart nanocarriers offer targeted drug delivery for cancer treatment.
• Enzymatic cleavage of nanocapsules enables selective drug release near tumor cells.
• Hepsin, a type II transmembrane serine protease, is overexpressed in prostate cancer.

Purpose of the Study:

• To develop and optimize peptide substrates cleavable by hepsin for targeted drug delivery.
• To identify novel hepsin substrates with enhanced cleavage efficiency.
• To create hepsin-cleavable nanocarriers resistant to plasma degradation.

Main Methods:

• In silico screening of combinatorial peptide libraries against hepsin's binding cavity.
• In vitro enzymatic assays to verify substrate cleavage efficiency.
• Introduction of d-amino acids to enhance peptide stability.

Main Results:

• Identification of multiple hepsin substrates with high in silico docking scores.
• In vitro validation confirmed hepsin cleavage of designed substrates, surpassing known substrates.
• Optimized peptides with d-amino acids showed hepsin specificity and resistance to plasma/serum degradation.

Conclusions:

• A methodology for designing and optimizing hepsin-specific peptide substrates was established.
• These substrates are effective for developing smart nanocarriers for targeted prostate cancer therapy.
• The d-amino acid modification provides stability, crucial for in vivo drug delivery applications.