Bispecific targeting system based on DARPins-modified light-chain ferritin for cancer chemotherapy

Area of Science:

• Oncology
• Biotechnology
• Molecular Biology

Background:

• Cancer's heterogeneity poses challenges for targeted therapies.
• Dual-receptor targeting can enhance therapeutic efficacy.
• Human epidermal growth factor receptor 2 (HER2) and epithelial cell adhesion molecule (EpCAM) are overexpressed in epithelial carcinomas.

Purpose of the Study:

• To create a bispecific fusion protein for simultaneous targeting of HER2 and EpCAM.
• To develop a potent protein-drug conjugate for enhanced cancer therapy.
• To evaluate the efficacy and specificity of the novel therapeutic agent.

Main Methods:

• Constructed a hybrid protein (DARP_9-29-FTLsh-EC1) using a truncated ferritin L-subunit scaffold and Designed Ankyrin Repeat Proteins (DARPins).
• Conjugated the hybrid protein to monomethyl auristatin E (MMAE) to form a protein-drug conjugate (DARP_9-29-FTLsh-EC1/MMAE).
• Utilized flow cytometry, confocal microscopy, in vitro cytotoxicity assays, and a mouse xenograft model for evaluation.

Main Results:

• The DARP_9-29-FTLsh-EC1 fusion protein demonstrated specific binding and internalization into HER2/EpCAM-expressing cells.
• The protein-drug conjugate exhibited HER2/EpCAM-specific cytotoxicity in vitro, correlating with receptor density.
• In vivo studies confirmed selective accumulation in tumors and significant tumor reduction in a mouse xenograft model.

Conclusions:

• Bispecific targeting using DARPins and a ferritin scaffold offers a promising strategy for precise anticancer therapy.
• The developed protein-drug conjugate shows potential for overcoming tumor heterogeneity and improving therapeutic outcomes.
• This approach opens new avenues for developing effective targeted cancer treatments.