Abstract 4354: Harnessing VLPs and nanodiscs to unlock antibody discovery for challenging membrane proteins

Abstract Multi-pass transmembrane proteins (MP-TMPs), including G protein-coupled receptors (GPCRs), ion channels, and transporters, are vital regulators of cellular communication and signaling. They represent over 60% of current drug targets, yet their structural complexity and dependence on lipid environments make recombinant production and antibody discovery extremely difficult. Traditional soluble formats fail to preserve their conformational integrity, limiting immunogenicity and screening relevance. To address these limitations, KACTUS developed a versatile membrane-protein display platform integrating virus-like particles (VLPs) and nanodiscs to stabilize full-length MP-TMPs in their native conformations. VLPs, composed of self-assembling structural proteins lacking genetic material, provide a natural lipid envelope that supports proper folding and high-density surface display. Using this system, KACTUS successfully presented the full-length STEAP1 protein (a prostate-cancer-associated antigen) and demonstrated specific binding to the therapeutic antibody Vandortuzumab with an EC50 of 89.8 ng/mL. Complementing this, SMA-based nanodiscs produced in mammalian cells offer detergent-free solubilization of membrane proteins within native lipid bilayers. The A2A adenosine receptor (A2AR) nanodiscs retained structural integrity and exhibited strong binding to anti-A2AR monoclonal antibodies by ELISA (EC50 = 0.15 µg/mL) and SPR (K_D = 0.32 nM). In addition, KACTUS developed CXCR4 VLP immunogens that generated functional antisera and monoclonal antibodies capable of ligand-blocking comparable to the commercial CXCR4 antibody Ulocuplumab. Further extending this approach, biotinylated VLPs and nanodiscs enabled efficient phage-display antibody panning, as validated with GPRC5D constructs (EC50 = 7.4 ng/mL for VLPs; 0.28 µg/mL for nanodiscs). KACTUS also established the first TCR-CD3 complex nanodisc, preserving all native subunits (TCRα/β, CD3ε/γ/δ/ζ) for accurate CD3 antibody screening and binding analysis (SP34 K_D = 0.1 nM). Together, these results demonstrate that the KACTUS VLP and nanodisc platforms deliver native-like, bioactive MP-TMPs suitable for antibody generation, ligand binding, and structural interrogation. This integrated technology provides a powerful solution to overcome long-standing challenges in membrane-protein immunization and screening, paving the way for next-generation therapeutic antibody discovery. Citation Format: Hannah Flaherty Duncan, Shicheng Chen, Jia Cui, Tingxu Chen. Harnessing VLPs and nanodiscs to unlock antibody discovery for challenging membrane proteins [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 4354.