Fig. 1: Immunopeptidome analysis reveals a VGLL1-derived peptide expressed by two PDAC patient-derived organoid lines.

a Experimental strategy to identify PDAC tumor-specific, HLA class I-bound peptides from 39 tumor specimens derived from 35 M.D. Anderson PDAC patients. b Bioinformatics screening strategy to identify potentially targetable TAAs from amongst the eluted PDAC-associated peptides. Peptide-encoding genes were assessed for PDAC tumor RNAseq expression compared with transcript expression in 53 GTex Portal normal tissues. Excluding testis, normal tissues were separated into four categories (non-essential, caution, hazard, and danger tissues) that reflected the potential toxicities expected from off-tumor killing activity against different tissues (listed in Table S2). All peptide-encoding genes were filtered successively using four corresponding expression thresholds of increasing stringency (30, 10, 3, and 1 TPM, indicated by green dotted lines) to eliminate candidate TAAs most likely to elicit autoimmune toxicity in the context of CTL therapy (red dotted lines). Screening of high-confidence peptides isolated from tumor organoid cell lines of PDAC patients MP015 and MP081 is depicted, showing that few eluted peptides met these stringent criteria. c Mass spectra of an HLA-A*0101-restricted VGLL1-derived peptide isolated from two different PDAC organoid cell lines, MP015 and MP081 (top 2 panels). The patient-derived peptides co-eluted with and matched the MS fragmentation spectra of the synthetic isotope-labeled VGLL1 peptide LSELETPGKY (containing a ¹³C/¹⁵N-labeled lysine residue), with the labeled y⁺ fragment ion series (shown in blue) demonstrating an expected shift of 8 atomic mass units, unlike the b⁺ fragment ions (bottom panel). IP immunoprecipitation, TPM transcripts per million, TAA tumor-associated antigen.