IFN? modulates the immunopeptidome of triple negative breast cancer cells by enhancing and diversifying antigen processing and presentation

To identify potential T-cell targets for Triple-Negative Breast Cancer (TNBC) vaccination, we examined the effect of the pro-inflammatory cytokine interferon-? (IFN?) on the transcriptome, proteome and immunopeptidome of the TNBC cell line MDA-MB-231. Using high resolution mass spectrometry, we identified a total of 84,131 peptides from 9,647 source proteins presented by human leukocyte antigen (HLA)-I and HLA-II alleles. Treatment with IFN? resulted in a remarkable remoulding of the immunopeptidome, with only a 34% overlap between untreated and treated cells across the HLA-I immunopeptidome, and expression of HLA-II only on treated cells. IFN? increased the overall number, diversity and abundance of the immunopeptidome, as well as the proportion of coverage of source antigens. The suite of peptides displayed under conditions of IFN? treatment included many known tumour associated antigens, with the HLA-II repertoire sampling 265 breast cancer associated antigens absent from those sampled by HLA-I. Quantitative analysis of the transcriptome (10,248 transcripts) and proteome (6783 proteins) of these cells revealed 229 proteins and transcripts were commonly differentially expressed, most of which involved in downstream targets of IFN? signalling including components of the antigen processing machinery such as tapasin and HLA. However, these changes in protein expression did not explain the dramatic modulation of the immunopeptidome following IFN? treatment. These results demonstrate the high degree of plasticity in the immunopeptidome TNBC cells following cytokine stimulation and provide evidence that under pro-inflammatory conditions a greater variety of HLA-I and HLA-II vaccine targets are unveiled to the immune system. This has important implications for the development of personalised cancer vaccination strategies. Overall design: To study the effects of IFN? on the transcriptome of TNBC cells in three replicates