A Potent Single-Domain Antibody Targeting LAG-3 for Efficient Tumor Immunotherapy

Area of Science:

• Immunology
• Oncology
• Biotechnology

Background:

• Lymphocyte activation gene-3 (LAG-3) is a key immune checkpoint receptor negatively regulating T-cell responses.
• Existing LAG-3-blocking monoclonal antibodies have limitations, driving the need for improved antibody formats.
• Single-domain antibodies (sdAbs) offer potential for enhanced biological and pharmacological properties in immunotherapy.

Purpose of the Study:

• To generate and characterize novel single-domain antibodies (sdAbs) targeting human LAG-3.
• To evaluate the efficacy of a lead sdAb candidate, 2H-G7, in preclinical models of cancer immunotherapy.

Main Methods:

• Phage display technology was used to generate a panel of sdAbs against human LAG-3.
• High-affinity sdAb 2H-G7 was selected and characterized for its binding kinetics and ability to block LAG-3 ligand interactions.
• Functional validation involved assessing T-cell activation markers (IL-2, CD69) in human PBMCs.
• Epitope mapping was performed to determine the binding site of 2H-G7.
• Efficacy was evaluated in a xenogeneic mouse model of non-small-cell lung cancer (NSCLC).

Main Results:

• A high-affinity sdAb, 2H-G7, was identified, binding LAG-3 with nanomolar affinity.
• 2H-G7 effectively blocked the interactions of LAG-3 with its ligands, FGL1 and MHC-II.
• 2H-G7 demonstrated restoration of T-cell function, evidenced by increased IL-2 secretion and CD69 expression.
• Epitope mapping revealed 2H-G7 binds to the D1D2 domains of LAG-3, distinct from relatlimab.
• The 2H-G7-Fc fusion protein showed superior tumor growth inhibition in an NSCLC xenograft model compared to relatlimab.

Conclusions:

• 2H-G7 is a potent LAG-3-blocking sdAb with high affinity and functional efficacy.
• The distinct epitope and superior preclinical efficacy suggest 2H-G7 as a promising candidate for next-generation LAG-3 immunotherapies.
• This sdAb format offers a potential advantage over conventional monoclonal antibodies for cancer treatment.