title: Frameshift-derived neoantigens constitute immunotherapeutic targets for patients with microsatellite-instable haematological malignancies: frameshift peptides for treating MSI+ blood cancers.
abstract: OBJECTIVE
  Microsatellite instability (MSI) resulting from loss of functional DNA mismatch repair was recently found in various haematological disorders. In coding sequences, MSI leads to frameshift mutations (FSMs) and the production of C-terminally altered proteins which are foreign to the immune system. Here, we wondered whether these frame-shifted peptide (FSP) sequences represent tumour-specific antigens also for MSI(+) leukaemia and lymphomas (L/L).
  
  
  METHODS
  A total of 33 coding region microsatellites were examined in MSI(+) L/L cell lines for the presence of FSMs. Thereafter, recognition of MSI(+) cells by established FSP-specific CD8(+) T cell lines was quantified using interferon (IFN)-γ enzyme-linked immunospot (ELISpot) assays. In each experiment, MSI(+) L/L cell lines and T2 targets exogenously loaded with the cognate peptide (=internal control) were employed. Supplementary, lytic activity towards tumour cells was analysed by standard chromium release assay ((51)Cr).
  
  
  RESULTS
  Mutational profiling of 33 coding microsatellite loci in nine MSI(+) L/L cell lines revealed instability in at least nine microsatellites. In each cell line, a distinct mutational profile was observed. Only three of the 33 loci were stable. FSP-specific and human leukocyte antigen-A2 (HLA-A2)-restricted T cells specifically recognised MSI(+) L/L cells endogenously expressing TGFβRII(-1), Caspase 5 (-1) and MSH3 (-1) in ELISpot assays. Moreover, specific killing of Caspase 5 (-1) and MSH3 (-1) expressing L/L cell lines was achieved in functional cytotoxicity assays.
  
  
  CONCLUSIONS
  Data presented here expand the importance of FSPs as shared and general tumour-specific antigens. Consequently, they open new avenues for specific immunotherapies not only for solid but also for MSI(+) haematological malignancies.
authors.affiliation: Division of Molecular Oncology and Immunotherapy, Department of General Surgery, University of Rostock, Rostock, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Maletzki
authors.affiliation: Division of Molecular Oncology and Immunotherapy, Department of General Surgery, University of Rostock, Rostock, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Schmidt
authors.affiliation: German Collection of Microorganisms and Cell Cultures (DSMZ), Braunschweig, Germany.
authors.forename: Wilhelm G
authors.initials: WG
authors.lastname: Dirks
authors.affiliation: Cellular Immunotherapy, Department of Internal Medicine V, University Clinic Heidelberg, Heidelberg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Schmitt
authors.affiliation: Division of Molecular Oncology and Immunotherapy, Department of General Surgery, University of Rostock, Rostock, Germany. Electronic address: michael.linnebacher@med.uni-rostock.de.
authors.forename: Michael
authors.initials: M
authors.lastname: Linnebacher
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D000951:Antigens, Neoplasm / Q000235:genetics* / Q000276:immunology*; D045744:Cell Line, Tumor; D016368:Frameshift Mutation; D019337:Hematologic Neoplasms / Q000188:drug therapy / Q000235:genetics* / Q000276:immunology*; D006801:Humans; D053842:Microsatellite Instability; D000070957:MutL Protein Homolog 1; D051718:MutS Homolog 2 Protein / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D009687:Nuclear Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D010455:Peptides / Q000235:genetics / Q000276:immunology / Q000627:therapeutic use; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D000951:Antigens, Neoplasm; C501486:MLH1 protein, human; D009687:Nuclear Proteins; D010455:Peptides; C497172:MSH2 protein, human; D000070957:MutL Protein Homolog 1; D051718:MutS Homolog 2 Protein