A SUSTAINED INCREASE IN THE ENDOGENOUS LEVEL OF CAMP REDUCES THE RETINOID CONCENTRATION REQUIRED FOR APL CELL MATURATION TO NEAR PHYSIOLOGICAL LEVELS

cAMP-dependent signal transduction co-operates with retinoids to induc e acute promyelocytic leukaemia (APL) cell maturation. The rationale o f this work was to determine whether signal cross-talk could be used t o decrease the pharmacological doses of retinoids in the treatment of APL. When only the basal level of adenylate-cyclase (AC) activity is p resent in NB4 cells, up to 1 mu M concentration of all-trans retinoic acid (RA) is required for full maturation (100%). In these conditions, with only 10 nM RA less than 20% of cells will differentiate. Althoug h the use of membrane receptor agonists to activate AC has been proved to synergize with RA treatment, these agents were never as potent as cell permeant cAMP analogues. Analogues have disadvantages since cleav age by serum and cellular phosphodiesterases generates metabolites whi ch interfere in cellular response. In the present study, we observed c ell maturation by engrafting an autonomous Bordetella pertussis AC whi ch steadily delivers natural cAMP into the cell. The enzyme alone had no effect on cell maturation. Importantly, cell maturation was increas ed in a dose-dependent manner when the bacterial AC (1 ng/ml to 1 mu g /ml) was used to potentiate the effects of low doses RA (10 nM). More than 50% of cells matured with only 10 nM of RA and 200 ng/ml of B. pe rtussis AC. The maturation response was significantly increased when l ower amounts of enzyme were repetitively added to the culture to compe nsate for enzymatic decay. These results indicate that a sustained AC activity enhanced cell maturation. We were able to reduce to 3 nM the RA requirement, provided that a minimal amount (20 ng/ml) of B. pertus sis AC was added every 12 h in culture. Membrane signalling maintainin g high the level of cAMP substantially improved the efficacy of APL ce ll maturation by retinoids. Therefore, therapeutic benefits are expect ed by lowering the concentration of RA towards physiological (nanomola r) levels, thus reducing the side-effects of the drug. cAMP-elevating drugs that act on a post-cyclase target (cyclic-nucleotide phosphodies terases) or cell-targeted drug carriers (cAMP and RA loaded liposomes) should be evaluated as maturation therapies combining the activation of multiple signalling pathways.