Abstract
The majority of oncogenic drivers are intracellular proteins, thus constraining their immunotherapeutic targeting to mutated peptides (neoantigens) presented by individual human leukocyte antigen (HLA) allotypes1. However, most cancers have a modest mutational burden that is insufficient to generate responses using neoantigen-based therapies2,3. Neuroblastoma is a paediatric cancer that harbours few mutations and is instead driven by epigenetically deregulated transcriptional networks4. Here we show that the neuroblastoma immunopeptidome is enriched with peptides derived from proteins that are essential for tumourigenesis and focus on targeting the unmutated peptide QYNPIRTTF, discovered on HLA-A*24:02, which is derived from the neuroblastoma dependency gene and master transcriptional regulator PHOX2B. To target QYNPIRTTF, we developed peptide-centric chimeric antigen receptors (CARs) using a counter-panning strategy with predicted potentially cross-reactive peptides. We further hypothesized that peptide-centric CARs could recognize peptides on additional HLA allotypes when presented in a similar manner. Informed by computational modelling, we showed that PHOX2B peptide-centric CARs also recognize QYNPIRTTF presented by HLA-A*23:01 and the highly divergent HLA-B*14:02. Finally, we demonstrated potent and specific killing of neuroblastoma cells expressing these HLAs in vitro and complete tumour regression in mice. These data suggest that peptide-centric CARs have the potential to vastly expand the pool of immunotherapeutic targets to include non-immunogenic intracellular oncoproteins and widen the population of patients who would benefit from such therapy by breaking conventional HLA restriction.
Highlights
The curative potential of chimeric antigen receptors (CARs) T cell-based cancer immunotherapies has been established in leukaemias, but their application in solid tumours has been limited by a paucity of known tumour-specific membrane proteins[5]
We show that the neuroblastoma immunopeptidome is enriched with peptides derived from proteins that are essential for tumourigenesis and focus on targeting the unmutated peptide QYNPIRTTF, discovered on human leukocyte antigen (HLA)-A*24:02, which is derived from the neuroblastoma dependency gene and master transcriptional regulator PHOX2B
These tumours are low in mutational burden[15] and major histocompatibility complex (MHC) expression[16], making neuroblastoma both a challenging tumour to target with MHC-based immunotherapies and an ideal model for addressing the major problems currently hindering the wider advancement of cancer immunotherapies
Summary
We have presented a process for identifying tumour-specific antigens derived from non-mutated oncoproteins, engineering PC-CARs against these tumour self-peptides and screening for cross-reactivity against MHC and the normal immunopeptidome. These methods have resulted in PC-CARs that can induce potent tumour killing across multiple HLA alleles in neuroblastoma and provide a roadmap for addressing the major challenges of therapeutic targeting of intracellular oncoproteins. We expect that the methods presented here will facilitate the discovery of tumour-specific targets in other human cancers with high unmet need and envision a library of scFv-based synthetic immunotherapies that provides population-scale coverage of HLA genotypes for both neoantigens and self-antigens. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/
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