Identification of novel HLA-restricted preferentially expressed antigen in melanoma peptides to facilitate off-the-shelf tumor-associated antigen-specific T-cell therapies

Published:April 05, 2021DOI:


      Background aims

      Preferentially expressed antigen in melanoma (PRAME) is a cancer/testis antigen that is overexpressed in many human malignancies and poorly expressed or absent in healthy tissues, making it a good target for anti-cancer immunotherapy. Development of an effective off-the-shelf adoptive T-cell therapy for patients with relapsed or refractory solid tumors and hematological malignancies expressing PRAME antigen requires the identification of major histocompatibility complex (MHC) class I and II PRAME antigens recognized by the tumor-associated antigen (TAA) T-cell product. The authors therefore set out to extend the repertoire of HLA-restricted PRAME peptide epitopes beyond the few already characterized.


      Peptide libraries of 125 overlapping 15-mer peptides spanning the entire PRAME protein sequence were used to identify HLA class I- and II-restricted epitopes. The authors also determined the HLA restriction of the identified epitopes.


      PRAME-specific T-cell products were successfully generated from peripheral blood mononuclear cells of 12 healthy donors. Ex vivo-expanded T cells were polyclonal, consisting of both CD4+ and CD8+ T cells, which elicited anti-tumor activity in vitro. Nine MHC class I-restricted PRAME epitopes were identified (seven novel and two previously described). The authors also characterized 16 individual 15-mer peptide sequences confirmed as CD4-restricted epitopes.


      TAA T cells derived from healthy donors recognize a broad range of CD4+ and CD8+ HLA-restricted PRAME epitopes, which could be used to select suitable donors for generating off-the-shelf TAA-specific T cells.

      Key Words

      To read this article in full you will need to make a payment

      Purchase one-time access:

      Academic and Personal
      One-time access price info
      • For academic or personal research use, select 'Academic and Personal'
      • For corporate R&D use, select 'Corporate R&D Professionals'


      Subscribe to Cytotherapy
      Already a print subscriber? Claim online access
      Already an online subscriber? Sign in
      Institutional Access: Sign in to ScienceDirect


        • Boon K.
        • et al.
        Comparison of medulloblastoma and normal neural transcriptomes identifies a restricted set of activated genes.
        Oncogene. 2003; 22: 7687-7694
        • Epping M.
        • et al.
        PRAME expression and clinical outcome of breast cancer.
        British Journal of Cancer. 2008; 99: 398-403
        • Ikeda H.
        • et al.
        Characterization of an antigen that is recognized on a melanoma showing partial HLA loss by CTL expressing an NK inhibitory receptor.
        Immunity. 1997; 6: 199-208
        • Oberthuer A.
        • et al.
        The tumor-associated antigen PRAME is universally expressed in high-stage neuroblastoma and associated with poor outcome.
        Clinical Cancer Research. 2004; 10: 4307-4313
        • Steinbach D.
        • et al.
        PRAME gene expression in childhood acute lymphoblastic leukemia.
        Cancer Genetics and Cytogenetics. 2002; 138: 89-91
        • Szczepanski M.
        • et al.
        PRAME expression in head and neck cancer correlates with markers of poor prognosis and might help in selecting candidates for retinoid chemoprevention in pre-malignant lesions.
        Oral Oncology. 2013; 49: 144-151
        • Tan P.
        • et al.
        Expression and prognostic relevance of PRAME in primary osteosarcoma.
        Biochemical and Biophysical Research Communications. 2012; 419: 801-808
        • van Baren N.
        • et al.
        PRAME, a gene encoding an antigen recognized on a human melanoma by cytolytic T cells, is expressed in acute leukaemia cells.
        British Journal of Haematology. 1998; 102: 1376-1379
        • Yang L.
        • et al.
        PRAME Gene Copy Number Variation Is Related to Its Expression in Multiple Myeloma.
        DNA and Cell Biology. 2017; 36: 1099-1107
        • Epping M.T.
        • et al.
        The human tumor antigen repressor of retinoic acid PRAME is a dominant receptor signaling.
        Cell. 2005; 122: 835-847
        • Yin B.
        PRAME: from diagnostic marker and tumor antigen to promising target of RNAi therapy in leukemic cells.
        Leukemia Research. 2011; 35: 1159-1160
        • Griffioen M.
        • et al.
        Detection and functional analysis of CD8+ T cells specific for PRAME: a target for T-cell therapy.
        Clin Cancer Res. 2006; 12: 3130-3136
        • Matko S.
        • et al.
        PRAME peptide-specific CD8(+) T cells represent the predominant response against leukemia-associated antigens in healthy individuals.
        Eur J Immunol. 2018; 48: 1400-1411
        • Rezvani K.
        • et al.
        Functional leukemia-associated antigen-specific memory CD8+ T cells exist in healthy individuals and in patients with chronic myelogenous leukemia before and after stem cell transplantation.
        Blood. 2003; 102: 2892-2900
        • Rezvani K.
        • et al.
        Ex vivo characterization of polyclonal memory CD8(+) T-cell responses to PRAME-specific peptides in patients with acute lymphoblastic leukemia and acute and chronic myeloid leukemia.
        Blood. 2009; 113: 2245-2255
        • Chapuis A.G.
        • et al.
        Transferred WT1-Reactive CD8(+) T Cells Can Mediate Antileukemic Activity and Persist in Post-Transplant Patients.
        Science Translational Medicine. 2013; 5: 174ra27
        • Chapuis A.G.
        • et al.
        Transferred melanoma-specific CD8(+) T cells persist, mediate tumor regression, and acquire central memory phenotype.
        Proceedings of the National Academy of Sciences of the United States of America. 2012; 109: 4592-4597
        • Hont A.B.
        • et al.
        Immunotherapy of Relapsed and Refractory Solid Tumors With Ex Vivo Expanded Multi-Tumor Associated Antigen Specific Cytotoxic T Lymphocytes: A Phase I Study.
        Journal of Clinical Oncology. 2019; 37: 2349-2359
        • Williams K.M.
        • et al.
        Complete Remissions Post Infusion of Multiple Tumor Antigen Specific T Cells for the Treatment of High Risk Leukemia and Lymphoma Patients after HCT.
        Blood. 2017; 130: 1
        • Bach P.B.
        • Giralt S.A.
        • Saltz L.B.
        FDA Approval of Tisagenlecleucel Promise and Complexities of a $475 000 Cancer Drug.
        JAMA. 2017; 318: 1861-1862
        • Roddie C.
        • et al.
        Manufacturing chimeric antigen receptor T cells: issues and challenges.
        Cytotherapy. 2019; 21: 327-340
        • Leen A.M.
        • et al.
        Multicenter study of banked third-party virus-specific T cells to treat severe viral infections after hematopoietic stem cell transplantation.
        Blood. 2013; 121: 5113-5123
        • Leen A.M.
        • et al.
        Monoculture-derived T lymphocytes specific for multiple viruses expand and produce clinically relevant effects in immunocompromised individuals.
        Nat Med. 2006; 12: 1160-1166
        • Withers B.
        • et al.
        Third-Party Donor Virus-Specific T Cells Are Efficacious in the Treatment of Refractory Viral Infection Following Allogeneic HSCT, but May Not Persist Post-Infusion.
        Blood. 2015; 126: 623
        • Kessler J.H.
        • et al.
        Efficient identification of novel HLA-A*0201-presented cytotoxic T lymphocyte epitopes in the widely expressed tumor antigen PRAME by proteasome-mediated digestion analysis.
        Journal of Experimental Medicine. 2001; 193: 73-88
        • Kessler J.H.
        • Melief C.J.M.
        Identification of T-cell epitopes for cancer immunotherapy.
        Leukemia. 2007; 21: 1859-1874
        • Quintarelli C.
        • et al.
        High-avidity cytotoxic T lymphocytes specific for a new PRAME-derived peptide can target leukemic and leukemic-precursor cells.
        Blood. 2011; 117: 3353-3362
        • Weber G.
        • et al.
        Generation of multi-leukemia antigen-specific T cells to enhance the graft-versus-leukemia effect after allogeneic stem cell transplant.
        Leukemia. 2013; 27: 1538-1547
        • Hanley P.J.
        • et al.
        Functionally active virus-specific T cells that target CMV, adenovirus, and EBV can be expanded from naive T-cell populations in cord blood and will target a range of viral epitopes.
        Blood. 2009; 114: 1958-1967
        • Epping M.T.
        • Bernards R.
        A causal role for the human tumor antigen preferentially expressed antigen of melanoma in cancer.
        Cancer Research. 2006; 66: 10639-10642
        • Toledo S.R.C.
        • et al.
        Insights on PRAME and osteosarcoma by means of gene expression profiling.
        Journal of Orthopaedic Science. 2011; 16: 458-466
        • Tzannou I.
        • et al.
        Off-the-Shelf Virus-Specific T Cells to Treat BK Virus, Human Herpesvirus 6, Cytomegalovirus, Epstein-Barr Virus, and Adenovirus Infections After Allogeneic Hematopoietic Stem-Cell Transplantation.
        Journal of Clinical Oncology. 2017; 35: 3547-3557
        • Haque T.
        • et al.
        Allogeneic cytotoxic T-cell therapy for EBV-positive posttransplantation lymphoproliferative disease: results of a phase 2 multicenter clinical trial.
        Blood. 2007; 110: 1123-1131
        • Keller M.D.
        • Bollard C.M.
        Virus-specific T-cell therapies for patients with primary immune deficiency.
        Blood. 2020; 135: 620-628
        • O'Reilly R.J.
        • et al.
        Virus-specific T-cell banks for ‘off the shelf’ adoptive therapy of refractory infections.
        Bone Marrow Transplantation. 2016; 51: 1163-1172
        • Quintarelli C.
        • et al.
        Cytotoxic T lymphocytes directed to the preferentially expressed antigen of melanoma (PRAME) target chronic myeloid leukemia.
        Blood. 2008; 112: 1876-1885
        • Dome J.S.
        • et al.
        Advances in Wilms Tumor Treatment and Biology: Progress Through International Collaboration.
        Journal of Clinical Oncology. 2015; 33: 2999-3007
        • Mullen E.A.
        • et al.
        Impact of Surveillance Imaging Modality on Survival After Recurrence in Patients With Favorable-Histology Wilms Tumor: A Report From the Children's Oncology Group.
        Journal of Clinical Oncology. 2018; 36JCO1800076
        • Ha T.C.
        • et al.
        An international strategy to determine the role of high dose therapy in recurrent Wilms’ tumour.
        European Journal of Cancer. 2013; 49: 194-210
        • Hanley P.J.
        • et al.
        CMV-specific T cells generated from naïve T cells recognize atypical epitopes and may be protective in vivo.
        Sci Transl Med. 2015; 7: 285ra63
        • Le Gal F.A.
        • et al.
        Tissue homing and persistence of defined antigen-specific CD8(+) tumor-reactive T-cell clones in long-term melanoma survivors.
        Journal of Investigative Dermatology. 2007; 127: 622-629
        • Abdelaal H.M.
        • Cartwright E.K.
        • Skinner P.J.
        Detection of Antigen-Specific T Cells Using In Situ MHC Tetramer Staining.
        International Journal of Molecular Sciences. 2019; 20: 11
        • Shao H.W.
        • et al.
        Identification of peptide-specific TCR genes by in vitro peptide stimulation and CDR3 length polymorphism analysis.
        Cancer Letters. 2015; 363: 83-91
        • Rego R.T.
        • Morris E.C.
        • Lowdell M.W.
        T-cell receptor gene-modified cells: past promises, present methodologies and future challenges.
        Cytotherapy. 2019; 21: 341-357
        • Oka Y.
        • et al.
        Wilms' Tumor Gene 1 (WT1) Peptide Vaccine Therapy for Hematological Malignancies: From CTL Epitope Identification to Recent Progress in Clinical Studies Including a Cure-Oriented Strategy.
        Oncology Research and Treatment. 2017; 40: 682-690
        • Kawahara M.
        • et al.
        Identification of HLA class I-restricted tumor-associated antigens in adult T cell leukemia cells by mass spectrometric analysis.
        Experimental Hematology. 2006; 34: 1496-1504
        • Castellino F.
        • Germain R.N.
        Cooperation between CD4(+) and CD8(+) T cells: when, where, and how.
        Annual Review of Immunology. 2006; 24: 519-540
        • Schoenberger S.P.
        • et al.
        T-cell help for cytotoxic T lymphocytes is mediated by CD40-CD40L interactions.
        Nature. 1998; 393: 480-483
        • Maiers M.
        • Gragert L.
        • Klitz W.
        High resolution HLA alleles and haplotypes in the US population.
        Human Immunology. 2006; 67: S16