THE IMMUNOSUPPRESSIVE DRUGS CYCLOSPORINE-A AND FK506 INHIBIT CALCINEURIN PHOSPHATASE-ACTIVITY AND GENE-TRANSCRIPTION MEDIATED THROUGH THE CAMP-RESPONSIVE ELEMENT IN A NONIMMUNE CELL-LINE

Cyclosporin A and the macrolide tacrolimus (FK506) are powerful immuno suppressive drugs that in T cells inhibit the calcium/calmodulin-depen dent phosphatase calcineurin thereby preventing the activation of T-ce ll-specific transcription factors, such as NF-AT, involved in lymphoki ne gene expression. While this may explain, at least in part, the mech anism of cyclosporin A/FK506 immunosuppression, additional mechanisms have to be invoked in order to explain the pharmacological properties and toxic effects of these drugs, such as nephrotoxicity and neurotoxi city. We have studied the effects of cyclosporin A and FK506 on calcin eurin phosphatase activity and gene transcription mediated by the cAMP -responsive element (CRE), a binding site of the ubiquitous transcript ion factor CREB. A reporter gene was placed under the transcriptional control of the CRE of the rat glucagon gene and transiently transfecte d into the glucagon-expressing cell line alphaTC2. Cyclosporin A and F K506 inhibited depolarization-induced gene transcription in a concentr ation-dependent manner (IC50 Of about 1 nM and 30 nM for FK506 and cyc losporin A, respectively). Both cyclosporin A and FK506 inhibited calc ineurin phosphatase activity at drug concentrations that inhibited gen e transcription. The FK506 analogue rapamycin had no effect on calcine urin activity and gene transcription, but excess concentrations of rap amycin prevented the effects of FK506 on both calcineurin activity and gene transcription. These results support the notion that the interac tion of drug-immunophilin complexes with calcineurin may be the molecu lar basis of cyclosporin A/FK506-induced inhibition of CREB/CRE-mediat ed gene transcription. The ability to interfere with CREB/CRE-mediated gene transcription represents a novel mechanism of cyclosporin A/FK50 6 action which may underlie pharmacological effects and toxic manifest ations of these potent immunuosuppressive drugs.