title: Real-time polymerase chain reaction monitoring of epithelial cell adhesion molecule-induced T-cell stimulation in patients with lung cancer and healthy individuals using LightCycler technology.
abstract: The epithelial cell adhesion molecule is strongly expressed in carcinomas of diverse histologic origin and has attracted attention as a target for immunotherapy. We have previously demonstrated that the epithelial cell adhesion molecule-derived human leukocyte antigen-A*0201-restricted nonamer peptide ILYENNVIT(184-192) can be efficiently recognized by peptide-specific cytotoxic T lymphocytes (CTL). Using this peptide (Ep-2) we could readily expand precursor CTL and demonstrate their cytotoxicity by standard chromium-release assay. We now sought to evaluate the immune reactivity of Ep-2-specific CTL expanded from the peripheral blood mononuclear cells by real-time quantitative polymerase chain reaction assay (LightCycler system) measuring the change in interferon-gamma mRNA levels upon stimulation. Cytotoxic T lymphocyte response against the Ep-2 peptide in two patients with epithelial cell adhesion molecule-expressing lung cancer and three healthy donors was compared with the chromium-release assay. In vitro expanded epithelial cell adhesion molecule-specific CTL precursors that exhibited significantly elevated interferon-gamma mRNA levels also lysed Ep-2 peptide-pulsed target cells. Our study indicates that quantitative polymerase chain reaction may be a supplement to chromium-release assay for monitoring in vitro expanded CTL precursor reactivity.
authors: Division of Oncology, Department of Internal Medicine, University Hospital Zurich, Rämistrasse 100, CH-8091 Zurich, Switzerland. andreas.trojan@dim.usz.ch
authors: Andreas
authors: A
authors: Trojan
authors: Mirjana
authors: M
authors: Urosevic
authors: Reinhard
authors: R
authors: Dummer
authors: Frank O
authors: FO
authors: Nestle
authors: Rolf A
authors: RA
authors: Stahel
mesh_terms: D000951:Antigens, Neoplasm / Q000276:immunology*; D016827:CD8 Antigens / Q000235:genetics; D015815:Cell Adhesion Molecules / Q000276:immunology*; D002460:Cell Line; D003602:Cytotoxicity, Immunologic; D000071858:Epithelial Cell Adhesion Molecule; D006801:Humans; D007371:Interferon-gamma / Q000235:genetics; D008175:Lung Neoplasms / Q000276:immunology*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012333:RNA, Messenger / Q000032:analysis; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
chemical_list: D000951:Antigens, Neoplasm; D016827:CD8 Antigens; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D012333:RNA, Messenger; D007371:Interferon-gamma