Encapsulation, metabolism and release of 2-fluoro-ara-AMP from human erythrocytes

Abstract

2-Fluoro-ara-AMP (fludarabine phosphate) is a purine analogue with anti-neoplastic activity in lymphoproliferative malignancies. Fludarabine phosphate activity and toxicity is schedule-dependent; multiple daily administrations (for five days) are more effective than single dose. We have encapsulated fludarabine phosphate in human erythrocytes and found that it is slowly released as fludarabine for more than four days. Encapsulated fludarabine phosphate does not affect erythrocyte metabolism and is rapidly converted by erythrocyte enzymes both to fludarabine with a K m of 0.4 mM and a V max of 20 nmol/min per g hemoglobin and to fludarabine diphosphate and triphosphate. The apparent K m for fludarabine monophosphate in the phosphorylation reaction was 0.4 mM and the V max 40 nmol/min per g hemoglobin. In the phosphorylation of 2-fluoro-ara-AMP to the di- and triphosphate derivatives, ATP was the phosphate donor with apparent K m of 0.12 and 1.0 mM, respectively. During incubations of 2-fluoro-ara-AMP-loaded erythrocytes at 37°C fludarabine was found in equilibrium between the erythrocyte and the culture medium suggesting that permeation of the erythrocyte membrane is not rate-limiting. Thus, fludarabine phosphate-loaded erythrocytes might be used as a slow-delivery system for fludarabine administration in the treatment of lymphoid malignancies.