Abstract A075: Molecular glues activate immunoproteasomes to amplify and expand the MHC-class-I immunopeptidome that promotes TCR-engineered T-cell cytotoxic activity against multiple myeloma

Abstract The proteasome is a cornerstone of adaptive immunity and is required for the efficient processing of antigenic peptides that are presented by MHC class I molecules to cytotoxic T lymphocytes. Proteasomes are macromolecular proteolytic machines and central components of the ubiquitin-proteasome system (UPS) that is responsible for degrading short-lived, misfolded, or damaged proteins to maintain proteostasis in eukaryotic cells. Proteasome-dependent protein degradation enables CD8+ T-cells to identify pathological cells that are synthesizing abnormal, mutant or viral proteins. Cancer cells have evolved elaborate mechanisms to subvert the antigen presentation machinery (APM) and avoid immunosurveillance mechanisms and elimination by CD8+ T-cells. Since the loss of MHC I antigen presentation is common in cancers and allows these cells to evade immunosurveillance, we hypothesized that defects in MHC class I antigen presentation could be reversed to reestablish immune control and to restore responses to immunotherapy. A multitude of endogenous proteins have been identified that activate proteasome catalytic activity and proteasome stimulation is recognized as a promising approach to augment antigen presentation. We reasoned that novel molecules that stimulate proteasomal endopeptidase activity would enhance the processing of intracellular proteins to amplify MHC class I antigen presentation and promote the antimyeloma activity of TCR-engineered T-cells. Immunoproteasomes are highly specialized forms of the proteasome that exhibit distinct catalytic activities to expand the immunopeptidome repertoire that is presented on tumor cells. Here, we report a novel molecule, designated Compound A, that “glues” the endogenous activator PA28alpha/beta to immunoproteasomes and enhances the presentation of MHC class I-bound peptides on multiple myeloma (MM) patient tumor cells. Immunoproteasomes isolated from MM cells treated with Compound A preferentially cleaved substrates that harbored a hydrophobic or basic residue at the C-terminus. Treatment with Compound A also promoted the co-immunoprecipitation of PA28alpha/beta with immunoproteasomes and the formation of a stable PA28alpha/beta-immunoproteasome complex. Mass spectrometry (MS) and peptide sequencing indicated that treatment of patient CD138+ cells with Compound A increased the proportion of immunopeptides that terminated in leucine or lysine residues, increased the presentation of both canonical and de novo peptides and enhanced the presentation of certain neoantigens by over 100-fold relative to untreated cells. Importantly, Compound A treatment of T-cells that had been engineered to express a T-cell receptor (TCR) that recognized the NY-ESO-1 antigen significantly promoted anti-myeloma activity. Immunoproteasome activation represents a powerful, paradigm-shifting strategy to amplify public and private neoantigens as targets for broadly applicable and personalized TCR-driven T-cell therapies that can be used to treat or ameliorate cancers, neurodegenerative disorders, infectious diseases and aging. Citation Format: James J. Driscoll, Nida Mubin, James J. Ignatz-Hoover. Molecular glues activate immunoproteasomes to amplify and expand the MHC-class-I immunopeptidome that promotes TCR-engineered T-cell cytotoxic activity against multiple myeloma [abstract]. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr A075.