Endogenous H3.3K27M derived peptide restricted to HLA-A∗02:01 is insufficient for immune-targeting in diffuse midline glioma

Stacie S Wang; Kirti Pandey; Katherine A Watson; Rebecca C Abbott; Nicole A Mifsud; Fiona M Gracey; Sri H Ramarathinam; Ryan S Cross; Anthony W Purcell; Misty R Jenkins

doi:10.1016/j.omto.2023.08.005

Endogenous H3.3K27M derived peptide restricted to HLA-A∗02:01 is insufficient for immune-targeting in diffuse midline glioma

Mol Ther Oncolytics. 2023 Aug 15:30:167-180. doi: 10.1016/j.omto.2023.08.005. eCollection 2023 Sep 21.

Authors

Stacie S Wang^{1

2

3}, Kirti Pandey⁴, Katherine A Watson¹, Rebecca C Abbott^{1

3}, Nicole A Mifsud⁴, Fiona M Gracey⁵, Sri H Ramarathinam⁴, Ryan S Cross¹, Anthony W Purcell⁴, Misty R Jenkins^{1

3

6}

Affiliations

¹ The Walter and Eliza Hall Institute of Medical Research, Immunology Division, Parkville, VIC 3052, Australia.
² Murdoch Children's Research Institute, Parkville, VIC 3052, Australia.
³ The University of Melbourne, Department of Medical Biology, Parkville, VIC 3052, Australia.
⁴ Department of Biochemistry and Molecular Biology and Infection and Immunity Program, Biomedicine Discovery Institute, Monash University, Clayton, VIC 3800, Australia.
⁵ Myrio Therapeutics, 6-16 Joseph St, Blackburn North, Melbourne, VIC 3130, Australia.
⁶ La Trobe University, La Trobe Institute for Molecular Science, Bundoora, VIC, Australia.

Abstract

Diffuse midline glioma (DMG) is a childhood brain tumor with an extremely poor prognosis. Chimeric antigen receptor (CAR) T cell therapy has recently demonstrated some success in DMG, but there may a need to target multiple tumor-specific targets to avoid antigen escape. We developed a second-generation CAR targeting an HLA-A∗02:01 restricted histone 3K27M epitope in DMG, the target of previous peptide vaccination and T cell receptor-mimics. These CAR T cells demonstrated specific, titratable, binding to cells pulsed with the H3.3K27M peptide. However, we were unable to observe scFv binding, CAR T cell activation, or cytotoxic function against H3.3K27M⁺ patient-derived models. Despite using sensitive immunopeptidomics, we could not detect the H3.3K27M_26-35-HLA-A∗02:01 peptide on these patient-derived models. Interestingly, other non-mutated peptides from DMG were detected bound to HLA-A∗02:01 and other class I molecules, including a novel HLA-A3-restricted peptide encompassing the K27M mutation and overlapping with the H3 K27M_26-35-HLA-A∗02:01 peptide. These results suggest that targeting the H3 K27M_26-35 mutation in context of HLA-A∗02:01 may not be a feasible immunotherapy strategy because of its lack of presentation. These findings should inform future investigations and clinical trials in DMG.

Keywords: CAR T cells; DIPG; DMG; H3.3K27M; T cell cytotoxicity; immunopeptidomics; immunotherapy; pediatric brain tumor; peptide-MHC; proteomics.