title: Reduced human leukocyte antigen expression in advanced-stage Ewing sarcoma: implications for immune recognition.
abstract: Ewing sarcoma (EWS) is a tumour most commonly arising in bone, although on occasion in soft tissue, with a poor prognosis in patients with refractory or relapsed disease, despite multimodal therapy. Immunotherapeutic strategies based on tumour-reactive T and/or natural killer cells may improve the treatment of advanced-stage EWS. Since cellular immune recognition critically depends on human leukocyte antigen (HLA) expression, knowledge about HLA expression in EWS is crucial in the design of cellular immunotherapeutic strategies. Constitutive and IFNgamma-induced HLA class I expression was analysed in EWS cell lines (n = 6) by flow cytometry, using antibodies against both monomorphic and allele-specific antigens. Expression of antigen processing pathway components and beta-2 microglobulin (beta2m) was assessed by western blot. Expression of class II transactivator (CIITA), and its contribution to HLA class II expression, was evaluated by qRT-PCR, transduction assays, and flow cytometry. beta2m/HLA class I and class II expression was validated in EWS tumours (n = 67) by immunofluorescence. Complete or partial absence of HLA class I expression was observed in 79% of EWS tumours. Lung metastases consistently lacked HLA class I and sequential tumours demonstrated a tendency towards decreased expression upon disease progression. Together with absent or low constitutive expression levels of specific HLA class I loci and alleles, and differential induction of identical alleles by IFNgamma in different cell lines, these results may reflect the existence of an immune escape mechanism. Inducible expression of TAP-1/-2, tapasin, LMP-2/-7, and the beta2m/HLA class I complex by IFNgamma suggests that regulatory mechanisms are mainly responsible for heterogeneity in constitutive class I expression. EWSs lack IFNgamma-inducible HLA class II, due to lack of functional CIITA. The majority of EWS tumours, particularly if advanced-stage, exhibit complete or partial absence of both classes of HLA. This knowledge will be instrumental in the design of cellular immunotherapeutic strategies for advanced-stage EWS.
authors.affiliation: Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands.
authors.forename: Dagmar
authors.initials: D
authors.lastname: Berghuis
authors.forename: Alfons S K
authors.initials: AS
authors.lastname: de Hooge
authors.forename: Susy J
authors.initials: SJ
authors.lastname: Santos
authors.forename: Danielle
authors.initials: D
authors.lastname: Horst
authors.forename: Emmanuel J
authors.initials: EJ
authors.lastname: Wiertz
authors.forename: Marja C
authors.initials: MC
authors.lastname: van Eggermond
authors.forename: Peter J
authors.initials: PJ
authors.lastname: van den Elsen
authors.forename: Antonie H M
authors.initials: AH
authors.lastname: Taminiau
authors.forename: Laura
authors.initials: L
authors.lastname: Ottaviano
authors.forename: Karl-Ludwig
authors.initials: KL
authors.lastname: Schaefer
authors.forename: Uta
authors.initials: U
authors.lastname: Dirksen
authors.forename: Erik
authors.initials: E
authors.lastname: Hooijberg
authors.forename: Arend
authors.initials: A
authors.lastname: Mulder
authors.forename: Cornelis J M
authors.initials: CJ
authors.lastname: Melief
authors.forename: R Maarten
authors.initials: RM
authors.lastname: Egeler
authors.forename: Marco W
authors.initials: MW
authors.lastname: Schilham
authors.forename: Ekaterina S
authors.initials: ES
authors.lastname: Jordanova
authors.forename: Pancras C W
authors.initials: PC
authors.lastname: Hogendoorn
authors.forename: Arjan C
authors.initials: AC
authors.lastname: Lankester
mesh_terms: D000293:Adolescent; D017951:Antigen Presentation; D015415:Biomarkers / Q000032:analysis; D015153:Blotting, Western; D001859:Bone Neoplasms / Q000276:immunology* / Q000401:mortality / Q000556:secondary; D045744:Cell Line, Tumor; D005434:Flow Cytometry; D006684:HLA-DR Antigens / Q000235:genetics; D015395:Histocompatibility Antigens Class I / Q000032:analysis*; D000949:Histocompatibility Antigens Class II / Q000235:genetics; D006801:Humans; D007114:Immunization; D007150:Immunohistochemistry; D007371:Interferon-gamma / Q000494:pharmacology; D053208:Kaplan-Meier Estimate; D009687:Nuclear Proteins / Q000235:genetics; D012512:Sarcoma, Ewing / Q000276:immunology* / Q000401:mortality / Q000556:secondary; D018709:Statistics, Nonparametric; D015534:Trans-Activators / Q000235:genetics; D014407:Tumor Cells, Cultured; D019139:Tumor Escape; D001613:beta 2-Microglobulin / Q000032:analysis
chemical_list: D015415:Biomarkers; D006684:HLA-DR Antigens; D015395:Histocompatibility Antigens Class I; D000949:Histocompatibility Antigens Class II; C083728:MHC class II transactivator protein; D009687:Nuclear Proteins; D015534:Trans-Activators; D001613:beta 2-Microglobulin; D007371:Interferon-gamma