Cyclosporine A transfer between high- and low-density lipoproteins: Independent from lipid transfer protein I-facilitated transfer of lipoprotein-coated phospholipids because of high affinity of cyclosporine A for the protein component of lipoproteins

The objectives of this study were to determine if lipid transfer protein I (LTP I)-facilitated phospholipid (PC) transfer activity regulates the plasm a lipoprotein distribution of cyclosporine (CSA) and if the association of CSA with high-density lipoproteins (HDL) is due to the high protein and/or alterations in coat lipid content of HDL. To assess if LTP I-facilitated PC transfer activity regulates the plasma lipoprotein distribution of CSA, C- 14-PC- or H-3-CSA-enriched HDL or low-density lipoproteins (LDL) were incub ated in T150 buffer [pH 7.4, containing a C-14-PC- or H-3-CSA-free lipoprot ein counterpart +/- exogenous LTP I (1.0 mug protein/mL)] or in delipidated human plasma that contained 1.0 mug protein/mL of endogenous LTP I in the presence or absence of a monoclonal antibody TP1 (30 mug protein/mL) direct ed against LTP I for 90 min at 37 degreesC. To assess the influence of HDL subfraction lipid composition and structure on the plasma distribution of C SA, CSA at 1000 ng of drug/mL of plasma was incubated in human plasma pretr eated for 24 h with a lecithin:cholesterol acyltransferase (LCAT) inhibitor , dithionitrobenzoate (DTNB; 3 mM). To assess the binding of CSA to apolipo proteins AI, AII, and B, increasing concentrations of CSA were added to a c onstant concentration of either apolipoprotein AI, AII, or B. Equilibrium d ialysis was used to determine free and bound fractions and Scatchard plot a nalysis was used to determine binding coefficients. To assess the influence of hydrophobic core lipid volume on the plasma distribution of CSA, CSA wa s incubated in plasma from patients with well-characterized dyslipidemias. The hydrophobic core lipid volume (CE + TG) within each lipoprotein subfrac tion was correlated to the amount of CSA recovered in each plasma sample fr om the different human subjects. The percent transfer of PC from LDL to HDL was different than the percent transfer of CSA in T150 buffer or human pla sma source. In the presence of TP1, only PC transfer from LDL to HDL decrea sed. For plasma incubated with CSA and separated into HDL2 and HDL3, 35-50% of drug originally incubated was recovered in the HDL3 fraction, with the remaining drug being found within the other fractions. When CSA was incubat ed in pla-sma pretreated with DTNB, the percentage of CSA recovered in the HDL3 and HDL2 fractions was not significantly different compared with that in the HDL3 and HDL2 fractions from untreated control plasma. CSA distribut ion into HDL inversely correlated with the hydrophobic core lipid volume of HDL, whereas distribution into LDL and triglyceride-rich lipoproteins dire ctly correlated with their respective hydrophobic core lipid volumes. We fu rther observed that CSA has high binding affinity and multiple binding site s with apolipoproteins AI (k(d) = 188.9 nM; n = 2), AII (k(d) = 184.7 nM; n = 2), and B (k(d) = 191 nM; n = 3). These findings suggest that the transf er of CSA between different lipoprotein particles is not influenced by LTP I-facilitated PC transfer activity probably because of the high affinity of CSA for the protein components of HDL and LDL. (C) 2001 Wiley-Liss, Inc. a nd the American Pharmaceutical Association.