Generation and validation of structurally defined antibody-siRNA conjugates

Area of Science:

• Biotechnology
• Molecular Biology
• Immunology

Background:

• Gene silencing via RNA interference (RNAi) shows therapeutic potential for diseases, including cancer.
• Efficient and selective delivery of small interfering RNAs (siRNAs) to cancer cells is a major hurdle for therapeutic application.
• Existing antibody-based siRNA delivery methods face limitations such as non-specific conjugation or complex site-specific strategies.

Purpose of the Study:

• To develop a novel, efficient, and site-specific method for generating antibody-siRNA conjugates (ARCs) for targeted cancer therapy.
• To create structurally defined ARCs using engineered dual variable domain (DVD) antibodies and chemically stabilized siRNA.
• To demonstrate the proof-of-concept for DVD-ARCs in delivering siRNA to multiple myeloma cells for gene knockdown.

Main Methods:

• Engineered dual variable domain (DVD) antibodies with a unique lysine residue were utilized for site-specific conjugation.
• A β-lactam linker-functionalized siRNA was conjugated to the DVD antibody, forming antibody-siRNA conjugates (ARCs) in a 1:2 ratio.
• DVD-ARCs targeting BCMA on multiple myeloma cells were generated to deliver siRNA against CTNNB1.

Main Results:

• A straightforward method for generating structurally defined, site-specific antibody-siRNA conjugates (ARCs) was established.
• The conjugation process was efficient and did not impair the antibody's antigen-binding affinity.
• Low concentrations (10 nM) of BCMA-targeting DVD-ARCs effectively reduced CTNNB1 mRNA and protein levels in multiple myeloma cells.

Conclusions:

• The developed ARC platform offers an efficient and easy-to-assemble approach for site-specific antibody-siRNA conjugation.
• This technology enables precise delivery of RNA interference therapeutics, holding promise for targeted cancer treatments.
• The proof-of-concept study demonstrates the potential of DVD-ARCs for selective gene silencing in cancer cells, such as multiple myeloma.