Fig. 1: An antigen discovery and prioritization process identifies PHOX2B as a target for immunotherapy.
From: RETRACTED ARTICLE: Cross-HLA targeting of intracellular oncoproteins with peptide-centric CARs
a, Summary of tumour-antigen discovery and CAR-engineering workflow: (1) integrated genomics and immunopeptidomics process; (2) target validation; (3) CAR engineering; and (4) cross-HLA tumour killing. b, Computational filtering of 9,117 peptide instances identified by immunopeptidomics in primary tumours (1% FDR) resulted in 56 neuroblastoma-specific peptides (33 unique peptides) derived from 29 distinct proteins. c, Primary neuroblastoma tumour immunopeptidome compared with 190 normal tissues. Each point on the x-axis represents one of 5,832 unique peptides identified in primary tumours, with the proportion of neuroblastoma tumours presenting a given peptide annotated above axis in dark blue and the proportion of normal tissue expressing the identical peptide below the axis in light blue. The green horizontal bar indicates 1,492 peptides not previously observed in the normal tissue immunopeptidome. Parent genes from neuroblastoma-specific peptides resulted in the top two GO enrichment terms: noradrenergic neuron differentiation and sympathetic nervous system development. The arrow denotes 351 recurring peptides presented in neuroblastoma that were not previously detected in normal tissues. d, Five antigens differentially expressed in PDX and primary tumours and further prioritized for analysis, HLA allele frequency, relative peptide abundance (percentile rank annotated below pMHC), predicted pMHC-binding affinity, and relevance to neuroblastoma tumourigenesis. e, PHOX2B expression in RNA-seq data from 153 neuroblastoma tumours versus 1,641 normal tissues in GTEx. PHOX2B expression is restricted to tumours, compared with the immunotherapy target HER2 and the neuroblastoma chemotherapy target TOP1 (note differences in the y-axis scales). FPKM, fragments per kilobase of transcript per million mapped reads. f, Crystal structure of PHOX2B 9mer QYNPIRTTF (red) refolded with HLA-A*24:02 (grey). g, Chromatin immunoprecipitation followed by DNA sequencing (ChIP-seq) of neuroblastoma tissue shows binding of all CRC proteins at the PHOX2B locus and association with a H3K27Ac super-enhancer mark. h, RNA-seq of fetal tissue shows that PHOX2B is expressed in early development and downregulated before birth across seven tissues. Created with BioRender.com.
