title: A PR1-human leukocyte antigen-A2 tetramer can be used to isolate low-frequency cytotoxic T lymphocytes from healthy donors that selectively lyse chronic myelogenous leukemia.
abstract: We previously showed (E. Clave et al., J. Immunother., 22: 1-6, 1999; J. Molldrem et al., Blood, 88: 2450-2457, 1996) that PR1, a human-lymphocyte-antigen (HLA)-A2.1-restricted peptide from proteinase 3, could be used to elicit CTLs from normal individuals. These CTLs showed HLA-restricted cytotoxicity and colony inhibition of myeloid leukemia cells that overexpress proteinase 3. In this study, we constructed a phycoerythrin-labeled PR1-HLA-A2 tetramer to identify PR1-specific CTLs by flow cytometry. No peripheral blood lymphocytes from three HLA-2.1+ donors stained with the tetramer, but, after 20 days in culture with weekly PR1 stimulation, 2-8% became tetramer+. Tetramer staining identified up to 40-fold more PR1-specific CTLs than were identified by limiting dilution analysis and correlated better with lysis of PR1-coated T2 cells (R2 = 0.95 versus R2 = 0.76). Tetramer+ CTLs were memory phenotype (91% CD45RO+), and most (58% CD95+) were activated. Tetramer-sorted allogeneic CTLs produced 83% lysis of HLA-A2.1+ chronic myelogenous leukemia (CML) blasts at an E:T ratio of 2.5:1, compared with 23% lysis by nonsorted CTLs, with no background lysis of HLA-A2.1+ normal cells. Cytoplasmic proteinase-3 expression was one log greater in CML blasts than in normal granulocytes. These results show that a PR1-HLA-A2 tetramer can be used to identify and select CTLs from normal donors that preferentially lyse CML cells, which could be used for leukemia-specific adoptive immunotherapy.
authors.affiliation: Blood and Marrow Transplant Department, University of Texas M.D. Anderson Cancer Center, Houston 77030-4095, USA. jmolldre@notes.mdacc.tmc.edu
authors.forename: J J
authors.initials: JJ
authors.lastname: Molldrem
authors.forename: P P
authors.initials: PP
authors.lastname: Lee
authors.forename: C
authors.initials: C
authors.lastname: Wang
authors.forename: R E
authors.initials: RE
authors.lastname: Champlin
authors.forename: M M
authors.initials: MM
authors.lastname: Davis
mesh_terms: D005434:Flow Cytometry; D015789:HLA-A2 Antigen / Q000276:immunology*; D006801:Humans; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive / Q000276:immunology*; D053146:Myeloblastin; D010446:Peptide Fragments / Q000276:immunology*; D010641:Phenotype; D012697:Serine Endopeptidases / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
chemical_list: D015789:HLA-A2 Antigen; D010446:Peptide Fragments; D012697:Serine Endopeptidases; D053146:Myeloblastin