Intracellular disposition of fludarabine triphosphate in human natural killer cells

Purpose: Fludarabine is a key component of several reduced-intensity conditioning regimens for hematopoietic cell transplantation (HCT). Shortly after reduced-intensity conditioning, the percent of donor natural killer (NK) cells has been associated with progression-free survival. Insufficient suppression of the recipient's NK cells by fludarabine may lead to lower donor chimerism; however, the effect of fludarabine upon NK cells is poorly understood. Thus, in purified human NK cells we evaluated the uptake and activation of fludarabine to its active metabolite, fludarabine triphosphate (F-ara-ATP), and assessed the degree of interindividual variability in F-ara-ATP accumulation. Methods: Intracellular F-ara-ATP was measured in purified NK cells isolated from healthy volunteers (n = 6) after ex vivo exposure to fludarabine. Gene expression levels of the relevant transporters and enzymes involved in fludarabine uptake and activation were also measured in these cells. Results: F-ara-ATP accumulation (mean ± SD) was 6.00 ± 3.67 pmol/1 × 10⁶ cells/4 h, comparable to average levels previously observed in CD4⁺ and CD8⁺ T-lymphocytes. We observed considerable variability in F-ara-ATP accumulation and mRNA expression of transporters and enzymes relevant to F-ara-ATP accumulation in NK cells from different healthy volunteers. Conclusions: Human NK cells have the ability to form F-ara-ATP intracellularly and large interindividual variability was observed in healthy volunteers. Further studies are needed to evaluate whether F-ara-ATP accumulation in NK cells are associated with apoptosis and clinical outcomes.