Characterization of liposomal tacrolimus in lung surfactant-like phospholipids and evaluation of its immunosuppressive activity.

Abstract

Tacrolimus (FK506) is a hydrophobic immunosuppressive agent that rapidly penetrates the plasmatic membrane and inhibits the signal transduction cascade of T lymphocytes. The objective of this study was the characterization of liposomal FK506 with surfactant-like phospholipids to be administered intratracheally after lung transplantation or in inflammatory lung diseases. We evaluated the optimal incorporation of FK506 in dipalmitoylphosphatidylcholine (DPPC) and DPPC/1-palmitoyl-2-oleoylphosphatidylglycerol (POPG) monolayers and bilayers and the effects of FK506 on the physical properties of DPPC and DPPC/POPG (8:2 w/w) vesicles. In addition, we assessed the immunosuppressive effects of surfactant-like phospholipid vesicles containing different amounts of FK506 on T-cell proliferation and interleukin 2 production. From surface pressure measurements of FK506/DPPC and FK506/DPPC/POPG mixed monolayers, we determined that FK506 was embedded into these monolayers up to an FK506 concentration of about 0.4 mol %. Beyond this concentration, FK506 was not quantitatively incorporated into the monolayer, suggesting possible concentration-dependent aggregation of tacrolimus. The incorporation of FK506 into DPPC monolayers, at concentrations <or= 5 microM, occurred with a partition coefficient of (3.9 +/- 0.3) x 10(3) at the bilayer equivalence pressure. FK506 was incorporated in DPPC bilayers up to an FK506 concentration of about 0.7-1 mol %, which was about double that obtained via the monolayer technique. FK506 hardly affected the transition enthalpy, the T(m), and cooperativity of the phase transition of DPPC and DPPC/POPG vesicles as determined by differential scanning calorimetry and steady-state 1,6-diphenyl-1,3,5-hexatriene anisotropy. Finally, this study provides evidence that liposomal FK506 retains the immunosuppressive efficacy of the drug.