Temperature dependence of glucocorticoid binding in sensitive and refractory murine leukaemia cells.
The validity of in vitro assays in predicting the susceptibility of leukaemic cells to glucocorticoid-mediated lysis was evaluated in a panel of six murine leukaemia cell lines. In this panel susceptibility to glucocorticoids ranged from highly sensitive to fully resistant. The panel was screened for specific 3H-dexamethasone binding in whole cells and for activation of cytosolic receptors in cell lysates. Specific binding of 3H-dexamethasone was strongly affected by the incubation temperature. In all cell lines, rapid and reversible changes were observed in the stability of agonist-receptor association with a transition temperature of 28 degrees C. Below this temperature, intracellular receptors were found to be in a stable-binding, high-affinity configuration, masking differences in receptor status among the various cell lines. When assayed at 37 degrees C, refractory and fully resistant cells revealed nonsaturating, low-affinity binding of steroid. Saturating, high-affinity binding was, however, restored in these cells by the drug meta-iodobenzylguanidine with concomitant sensitization to dexamethasone-induced lysis. Contrary to observations with intact cells, heat-induced agonist-receptor dissociation in cytosols caused irreversible loss of (re)binding capacity. Activation of cytosolic receptors only recognized fully resistant cell lines as being deficient in the transformation of liganded receptors into a DNA-binding configuration. The assay, however, could not discriminate between three cell lines with highest but varying degrees of sensitivity because of maximal activation. The results indicate that non-physiological temperature and cell disruption strongly and differentially affect steroid binding and receptor activation, respectively. The observations may account for the poor correlation between conventional predictive assays and steroid responsiveness in clinical leukaemia.
van den Berg
J D
JD
The Netherlands Cancer Institute/Antoni van Leeuwenhoekhuis, Division of Experimental Therapy, Amsterdam.
Smets
L A
LA
van den Elshout
M M
MM
van Geel
I P
IP
Janssen
M
M
D019797:3-Iodobenzylguanidine; D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D003600:Cytosol / Q000378:metabolism; D004273:DNA, Neoplasm / Q000378:metabolism; D003907:Dexamethasone / Q000378:metabolism; D004351:Drug Resistance; D004354:Drug Screening Assays, Antitumor; D004355:Drug Stability; D005938:Glucocorticoids / Q000378:metabolism* / Q000494:pharmacology; D007462:Iodobenzenes / Q000494:pharmacology; D007939:Leukemia L1210 / Q000378:metabolism*; D051379:Mice; D010641:Phenotype; D011965:Receptors, Glucocorticoid / Q000378:metabolism* / Q000502:physiology; D013696:Temperature; D014316:Tritium; D014407:Tumor Cells, Cultured
D000970:Antineoplastic Agents; D004273:DNA, Neoplasm; D005938:Glucocorticoids; D007462:Iodobenzenes; D011965:Receptors, Glucocorticoid; D014316:Tritium; D019797:3-Iodobenzylguanidine; D003907:Dexamethasone