Rabbit derived VL single-domains as promising scaffolds to generate antibody-drug conjugates

Area of Science:

• Oncology
• Biotechnology
• Immunotherapy

Background:

• Antibody-drug conjugates (ADCs) are a rapidly advancing therapeutic class in oncology.
• Significant engineering challenges remain for current ADC development, necessitating more stable and effective designs.
• Rabbit light chains possess a unique disulfide bridge absent in humans and mice, offering a novel conjugation site.

Purpose of the Study:

• To explore rabbit-derived VL-single-domain antibody scaffolds (sdAbs) for site-selective payload conjugation.
• To develop a new generation of stable and homogenous ADCs.
• To validate a platform for novel ADC development using a canine lymphoma model.

Main Methods:

• Generated and screened a rabbit sdAb library targeting canine non-Hodgkin lymphoma (cNHL) using in vitro and in vivo phage display.
• Identified and selected a specific VL-sdAb, C5, with high affinity for cNHL cells and promising in vivo tumor uptake.
• Performed site-selective conjugation of SN-38 payload to C5 via its exposed Cys80 to create C5-DAB-SN-38.

Main Results:

• Identified several highly specific VL-sdAbs, including C5, demonstrating potent in vitro targeting of cNHL cells.
• C5 exhibited promising in vivo tumor uptake, supporting its potential for ADC development.
• The resulting ADC, C5-DAB-SN-38, showed potent cytotoxicity against cNHL cells and inhibited DNA-TopoI activity.

Conclusions:

• Validated a novel platform for ADC development utilizing rabbit VL-sdAb scaffolds.
• Demonstrated the potential of site-selective conjugation at Cys80 for creating stable and homogenous ADCs.
• The canine lymphoma model serves as a valuable preclinical framework for translating novel ADC therapeutics for cancer treatment.