Abstract A031: Discovery and characterization of a novel, immunoproteasome activator that modulates the immunopeptidome, increases MHC class I antigenic presentation and enhances antitumor immunity

Abstract

Abstract The transformative success of cancer immunotherapy has revolutionized cancer treatment and can induce long lasting responses in patients with cancers from a wide range of histologies. However, only a fraction of patients respond to immunotherapy and cancer cells escape immunosurveillance by downregulating antigen presentation through mechanisms that remain elusive. Proteasomes play a central role in the immune response by generating peptides from intracellular antigens which are presented on the cell surface for recognition by CD8+ cytotoxic T lymphocytes (CTLs). Immune cells contain a highly specialized variant of proteasomes, known as immunoproteasomes, in which the three constitutive catalytic subunits are replaced by cytokine-inducible, homologous catalytic subunits. Immunoproteasomes are specially adapted for a role in MHC class I antigen processing and presentation to CD8+ T-cells. Here, we performed a high-through screen (HTS) to detect novel molecular entities that activated proteasomal peptide-hydrolyzing activity. We identified a curated panel of molecules which not only enhanced proteasomal hydrolysis of the cell-permeable substrate LLVY-R110 but also enhanced the presentation of MHC class I antigens. E.G7-Ova is a mouse lymphoma cell line derived by electroporation of EL4 cells (C57BL/6, H-2b, T lymphoma) with chicken ovalbumin (OVA) cDNA. Proteasomes degrade OVA to generate the peptide SIINFEKL that is presented by MHC-class I molecules. Studies demonstrated that the hits identified in the screen increased SIINFEKL presentation on E.G7-Ova cells. The lead compound identified in the HTS, compound A, dramatically enhanced expression of the proteasome activator PA28, which is expressed by PSME1 and PSME2. Treatment of multiple myeloma (MM) cells with compound A enhanced the hydrolysis of Ac-ANW-MCA, which is selectively cleaved by the immunoproteasome beta5-associated catalytic subunits. Treatment of MM cells with compound A followed by mass spectrometry (MS) indicated that the relative presentation of individual antigenic peptides bound to MHC class I molecules on tumor cells was dramatically increased. Novel immunoproteasome-selective therapeutics have the potential to generate neoantigens and upregulate cspecific antigens on tumor cells to enhance CD8+ T-cell-mediated control of human cancers. Citation Format: James J. Driscoll, Priyanka S. Rana, James J. Ignatz-Hoover. Discovery and characterization of a novel, immunoproteasome activator that modulates the immunopeptidome, increases MHC class I antigenic presentation and enhances antitumor immunity [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2023 Oct 1-4; Toronto, Ontario, Canada. Philadelphia (PA): AACR; Cancer Immunol Res 2023;11(12 Suppl):Abstract nr A031.