title: Integrated Whole Genome and Transcriptome Analysis Identified a Therapeutic Minor Histocompatibility Antigen in a Splice Variant of ITGB2.
abstract: OBJECTIVE
  In HLA-matched allogeneic hematopoietic stem cell transplantation (alloSCT), donor T cells recognizing minor histocompatibility antigens (MiHAs) can mediate desired antitumor immunity as well as undesired side effects. MiHAs with hematopoiesis-restricted expression are relevant targets to augment antitumor immunity after alloSCT without side effects. To identify therapeutic MiHAs, we analyzed the in vivo immune response in a patient with strong antitumor immunity after alloSCT.
  
  
  METHODS
  T-cell clones recognizing patient, but not donor, hematopoietic cells were selected for MiHA discovery by whole genome association scanning. RNA-sequence data from the GEUVADIS project were analyzed to investigate alternative transcripts, and expression patterns were determined by microarray analysis and qPCR. T-cell reactivity was measured by cytokine release and cytotoxicity.
  
  
  RESULTS
  T-cell clones were isolated for two HLA-B*15:01-restricted MiHA. LB-GLE1-1V is encoded by a nonsynonymous SNP in exon 6 of GLE1 For the other MiHAs, an associating SNP in intron 3 of ITGB2 was found, but no SNP disparity was present in the normal gene transcript between patient and donor. RNA-sequence analysis identified an alternative ITGB2 transcript containing part of intron 3. qPCR demonstrated that this transcript is restricted to hematopoietic cells and SNP-positive individuals. In silico translation revealed LB-ITGB2-1 as HLA-B*15:01-binding peptide, which was validated as hematopoietic MiHA by T-cell experiments.
  
  
  CONCLUSIONS
  Whole genome and transcriptome analysis identified LB-ITGB2-1 as MiHAs encoded by an alternative transcript. Our data support the therapeutic relevance of LB-ITGB2-1 and illustrate the value of RNA-sequence analysis for discovery of immune targets encoded by alternative transcripts. Clin Cancer Res; 22(16); 4185-96. ©2016 AACR.
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Margot J
authors.initials: MJ
authors.lastname: Pont
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Dyantha I
authors.initials: DI
authors.lastname: van der Lee
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Edith D
authors.initials: ED
authors.lastname: van der Meijden
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Cornelis A M
authors.initials: CA
authors.lastname: van Bergen
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Michel G D
authors.initials: MG
authors.lastname: Kester
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Maria W
authors.initials: MW
authors.lastname: Honders
authors.affiliation: Department of Human Genetics, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Martijn
authors.initials: M
authors.lastname: Vermaat
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Matthias
authors.initials: M
authors.lastname: Eefting
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Erik W A
authors.initials: EW
authors.lastname: Marijt
authors.affiliation: Department of Medical Statistics and Bioinformatics, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Szymon M
authors.initials: SM
authors.lastname: Kielbasa
authors.affiliation: Department of Human Genetics, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: Peter A C 't
authors.initials: PA
authors.lastname: Hoen
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands.
authors.forename: J H Frederik
authors.initials: JH
authors.lastname: Falkenburg
authors.affiliation: Department of Hematology, Leiden University Medical Center, Leiden, the Netherlands. m.griffioen@lumc.nl.
authors.forename: Marieke
authors.initials: M
authors.lastname: Griffioen
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D001483:Base Sequence; D018984:Epitopes, T-Lymphocyte / Q000235:genetics / Q000276:immunology; D005260:Female; D020869:Gene Expression Profiling; D059910:HLA-B15 Antigen / Q000235:genetics / Q000276:immunology; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D039661:Integrin beta3 / Q000737:chemistry / Q000235:genetics*; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000235:genetics / Q000276:immunology; D015778:Minor Histocompatibility Antigens / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D010455:Peptides / Q000235:genetics / Q000276:immunology; D013601:T-Lymphocytes; D014184:Transplantation, Homologous; D000073336:Whole Genome Sequencing
chemical_list: D018984:Epitopes, T-Lymphocyte; D059910:HLA-B15 Antigen; C061248:ITGB3 protein, human; D039661:Integrin beta3; D015778:Minor Histocompatibility Antigens; D010455:Peptides