Single-cell derived tumor organoids display diversity in HLA class I peptide presentation

Abstract

Tumor heterogeneity is a major cause of therapeutic resistance. Immunotherapy may exploit alternative vulnerabilities of drug-resistant cells, where tumor-specific human leukocyte antigen (HLA) peptide ligands are promising leads to invoke targeted anti-tumor responses. Here, we investigate the variability in HLA class I peptide presentation between different clonal cells of the same colorectal cancer patient, using an organoid system. While clone-specific differences in HLA peptide presentation were observed, broad inter-clone variability was even more prevalent (15–25%). By coupling organoid proteomics and HLA peptide ligandomics, we also found that tumor-specific ligands from DNA damage control and tumor suppressor source proteins were prominently presented by tumor cells, coinciding likely with the silencing of such cytoprotective functions. Collectively, these data illustrate the heterogeneous HLA peptide presentation landscape even within one individual, and hint that a multi-peptide vaccination approach against highly conserved tumor suppressors may be a viable option in patients with low tumor-mutational burden.

A facile immunopeptidomics workflow for capturing the HLA-I ligandome with PEAKS XPro

“…To demonstrate the utility of our immunopeptidomics workflow we re-analyzed the ligandome data from Demmers et al . (6) and demonstrate how it can capture the diversity in HLA-I ligands presented on different organoids from the same tumour. The genetic diversity among clones of the same tumour is an important driver of tumour evolution and acquiring resistance against chemotoxic stress.…”

“…Immunopeptidomics is a valuable method for characterizing the antigens displayed on the surface of tumour cells by HLA-I molecules. As described in (6), by sampling different organoids from the same tumour of a patient and defining inter-clonal differences in HLA-I ligands, more effective immunotherapy drugs can be developed.…”

“…The workflow relies on de novo sequencing to retrieve peptide sequences directly from the MS data and is combined with a homology-based database search to identify missense mutations. Using highly stringent parameters in our MS analysis, and predicting HLA-I binding peptides with NetMHCpan-4.1, we improve HLA-I ligand identification with high confidence and characterize diverse sets of peptides across colorectal cancer tumour clones from a previously published dataset (6).…”

Organoids in cancer research: a review for pathologist‐scientists

“…Mutational signatures in the organoid sequencing data suggested distinct mutagenic processes in normal colon cells and colon cancer cells, and differences in drug response were identified even in closely related clones from the same cancer, highlighting functional as well as genetic heterogeneity using the organoid model. In addition to genetic heterogeneity, Demmers et al [24] assessed proteomic heterogeneity between organoids derived from single cells of human colorectal cancer specimens. Although there was some heterogeneity between cancer clones, there were far greater proteomic differences between cancer and normal organoids.…”

“…Although there was some heterogeneity between cancer clones, there were far greater proteomic differences between cancer and normal organoids. In addition, this study analyzed heterogeneity in peptide ligands binding HLA between these single cell‐derived organoids [24]. Although the majority of HLA ligands were shared between tumor and normal organoids, indicating that the majority of ligands were the result of routine protein turnover, almost one‐quarter of ligands showed >two‐fold variation between organoids derived from the same tumor.…”