Figure S5 from Immunoproteasome Activation Expands the MHC Class I Immunopeptidome, Unmasks Neoantigens, and Enhances T-cell Anti-Myeloma Activity
Year: 2026
Type: other
Abstract: Figure S5. Effect of Compound A on pan-MHC-I expression. a. ARH77 and U266 cells were treated with Compound A and probed with pan-MHC-I or MHC-I subtype allele-specific antibodies after which the samples were analyzed by flow cytometry. Pretreatment with Compound A significantly increased pan-MHC class-I HLA-ABC and HLA-B surface presentation in all MMCLs assayed. b. ARH77 and U266 cells were pre-treated with Compound A for 48 h after which they were washed and co-cultured with healthy CD8+ cytotoxic T-cells for 18 h at the E:T ratios of 1:1, 2:1and 3:1. Compound A increased CTL-mediated cytotoxicity of tumor cells which was measured by gating for CD138, Annexin-V and PI positive cells at E:T 2:1 and 3:1. Pretreatment of MMCLs and T-cells alone with Compound A resulted in ∼5-10% cell death which was enhanced further when MM cells were co-cultured at E:T ratios of 2:1 and 3:1. T-cells alone also did not show significant cell death when pre-treated with Compound A. There was <5% MM cell death detected with or without Compound A treatment. Similar results were observed with CD8+ T-cells alone. c. Effect of Compound A on the viability of ARH77 and U266 cells alone or combined with ICIs and CD8+ T-cells. MM cells and CD8+ T-cells were separately treated with or without Compound A and the indicated agents. Following co-culture, the relative percent of indicated viable cells was quantitated by annexin-V and propidium iodide staining and flow cytometry. c. Effect of Compound A on T-cell exhaustion markers. CD8+ T-cells were treated with compound A for 24 h after which they were assessed for surface expression of PD-1, TIM-3, CTLA-4, and LAG-3. Compound A induced a marked decrease in PD-1 surface expression, while no significant change was observed in TIM-3, CTLA-4, and LAG-3.
Language: English
Subfield: Immunology
Field: Immunology and Microbiology
Domain: Life Sciences
Open Access status: gold
OpenAlex ID: W7164993768