Development of a human antibody fragment directed against the alpha folate receptor as a promising molecule for...

Area of Science:

• Oncology
• Immunology
• Biotechnology

Background:

• Alpha folate receptor (FRα) is a promising therapeutic target for non-small-cell lung cancer (NSCLC) due to its overexpression on tumor cells and limited presence in normal tissues.
• Developing targeted therapies requires specific agents that can effectively bind to cancer cells while minimizing off-target effects.

Purpose of the Study:

• To develop and characterize a novel human variable domain of a heavy-chain (VH) antibody fragment targeting FRα.
• To evaluate the binding affinity, specificity, and internalization potential of the developed VH fragment for potential NSCLC targeted therapy.

Main Methods:

• Phage display technology was employed to enrich and select a human VH antibody fragment (3A102 VH) specific for FRα.
• Enzyme-linked immunosorbent assay (ELISA) was used to confirm FRα binding specificity and cross-reactivity with FRβ.
• Soluble protein expression in *E. coli*, affinity determination (K_aff), cell-based ELISA, and confocal microscopy were utilized to assess binding and internalization.

Main Results:

• The selected 3A102 VH clone specifically bound to FRα and showed cross-reactivity with FRβ.
• Soluble 3A102 VH exhibited high affinity for FRα (K_aff ~7.77 × 10⁷ M^-1) and specifically bound to FRα-expressing NSCLC cells and primary cancer cells.
• Confocal microscopy confirmed that soluble 3A102 VH was internalized into FRα-expressing cells, indicating its potential as a drug delivery vehicle.

Conclusions:

• The study successfully developed a human VH antibody fragment (3A102 VH) with high affinity and specificity for FRα.
• The FRα-targeting VH fragment demonstrates potential for targeted drug delivery in NSCLC and other FRα-positive cancers.
• Phage display is an effective strategy for generating VH antibody fragments suitable for targeted cancer therapies.