title: Expression and structural analysis of glucocorticoid receptor isoform gamma in human leukaemia cells using an isoform-specific real-time polymerase chain reaction approach.
abstract: Glucocorticoids are broadly used for chemotherapy in childhood acute lymphoblastic leukaemia (ALL). The intracellular effects of glucocorticoids are mediated through the glucocorticoid receptor. The human glucocorticoid receptor gamma isoform (hGR-gamma) differs from the main isoform (hGR-alpha) by an additional amino acid within the DNA binding domain of the receptor protein. This may decrease hGR-alpha-mediated transcriptional activation. The importance of hGR-gamma expression in childhood ALL is unknown. To evaluate hGR-gamma mRNA expression levels, a real-time polymerase chain reaction (PCR)-based approach, allowing the selective amplification of hGR-gamma, was developed and optimized. We were able to demonstrate target selectivity of hGR-gamma amplification using sequence-specific primers. Studying the structure of the 3' end of hGR-gamma, a combination of this isoform with other hGR isoforms could be demonstrated. Using analysis of hGR-gamma-specific amplification in comparison with the expression of hGR-total (all isoforms) in leukaemic blasts from patients with either a good response to prednisone (PGR) or poor-prednisone response (PPR) in vivo, relative hGR-gamma expression was observed to be lower in cells from patients with PGR compared with PPR, in particular after 10 h of dexamethasone stimulation. These data were correlated with cell survival, demonstrating a more pronounced induction of apoptosis in cells from patients with PGR as compared with PPR.
authors.affiliation: Department of Paediatric Haematology and Oncology, Hannover Medical School, Hannover, Germany. Berger.Carmela@mh-hannover.de
authors.forename: Carmela
authors.initials: C
authors.lastname: Beger
authors.forename: Katrin
authors.initials: K
authors.lastname: Gerdes
authors.forename: Melchior
authors.initials: M
authors.lastname: Lauten
authors.forename: Wim J E
authors.initials: WJ
authors.lastname: Tissing
authors.forename: Ivonne
authors.initials: I
authors.lastname: Fernandez-Munoz
authors.forename: Martin
authors.initials: M
authors.lastname: Schrappe
authors.forename: Karl
authors.initials: K
authors.lastname: Welte
mesh_terms: D017209:Apoptosis; D001483:Base Sequence; D016022:Case-Control Studies; D002648:Child; D017931:DNA Primers; D003907:Dexamethasone; D005938:Glucocorticoids / Q000627:therapeutic use; D006801:Humans; D008969:Molecular Sequence Data; D054198:Precursor Cell Lymphoblastic Leukemia-Lymphoma / Q000235:genetics*; D011241:Prednisone / Q000627:therapeutic use; D020033:Protein Isoforms / Q000235:genetics*; D011965:Receptors, Glucocorticoid / Q000235:genetics*; D012074:Remission Induction; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D017931:DNA Primers; D005938:Glucocorticoids; D020033:Protein Isoforms; D011965:Receptors, Glucocorticoid; D003907:Dexamethasone; D011241:Prednisone