Abstract

8-Chloro-cyclic AMP (8-Cl-cAMP) produces growth-inhibitory and differentiating activity in the promyelocytic leukemia cell line HL-60. Adriamycin (ADR)-resistant HL-60 (HL-60/AR) cells exhibit the multidrug-resistant phenotype but do not express the mdr1 gene product P-glycoprotein. To explore potential signaling processes that may be involved in this atypical form of drug resistance, 8-Cl-cAMP was used as a modulator of the cAMP second messenger signal transduction pathway. Treatment for 48 hr with a 10% inhibitory concentration of 8-Cl-cAMP potentiated ADR cytotoxicity 14-fold in HL-60/AR cells but not in the parental cell line. 8-Cl-cAMP was stable to hydrolysis in the medium after 48 hr and was present intracellularly predominantly as phosphorylated metabolites (70%) and the parent compound (30%). No difference occurred in ADR accumulation in HL-60/AR cells after treatment with 8-Cl-cAMP. Accompanying the 8-Cl-cAMP-mediated increase in ADR cytotoxicity in HL-60/AR cells was a reduction in the cytosolic type I cAMP-dependent protein kinase (PKA) and disappearance of the nuclear PKA holoenzyme. Coincident with these changes in drug-resistant cells was a marked reduction in the DNA-binding activity of the cAMP response element-binding protein to levels equivalent to those in sensitive cells. This effect appears to result from reduced phosphorylation of the cAMP response element-binding protein. These results suggest that the potentiation by 8-Cl-cAMP of ADR cytotoxicity in HL-60/AR cells occurs through down-regulation of nuclear type I PKA and cAMP response element-binding factors whose activities are regulated by PKA.