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Liposomal tacrolimus and intestinal drug concentration
Liposomal Tacrolimus and Intestinal Drug Concentration S. Dutta, M. Mezei, T.D.G. Lee, and V.C. McAlister
T
ACROLIMUS (TAC) has allowed routinely successful intestinal transplantation, although the therapeutic window remains problematic.1 Liposomal tacrolimus (LTAC) may allow increased efficacy without nephrotoxicity,2 but effectiveness may depend on adequate graft penetration. Improved canine liver graft survival with L-TAC is associated with higher drug levels in the graft.3 To study the effect of liposomal encapsulation on biodisposition in the intestine, we made a multilayered liposome with 3H-labeled tacrolimus and 14C-labeled liposomes. MATERIALS AND METHODS Liposomes were prepared by a solvent evaporation method. Briefly, 5% phospholipon 90-H, loralan-CH cholesterol, 0.02% tacrolimus, 300 mCi 3H-tacrolimus, and 100 mCi 14C-L-3-phosphatidyl (N-methyl-14C) choline,1-2-dipalmitoyl (14C-DPPC) were dissolved in 20 mL chloroform/methanol (2:1 v/v), and dried by evaporation under vacuum. Ninety-five milliliters of preheated 0.9% NaCl (55°C) were added, and the mixture was hand-shaken for 1 minute at 55°C. The dissolved liposomes were subsequently placed in a preheated Environ Shaker (55°C) for 20 minutes, after which they were slowly brought back to room temperature with constant shaking. Microscopic examination determined the liposomes to be multilamellar. The control solution was prepared by adding 300 mCi 3Htacrolimus to commercial tacrolimus (Prograf), and diluted to a final concentration of 0.02% tacrolimus in 0.9% NaCl. Female CD-1 mice (6 to 8 weeks old) were injected intravenously (tail vein) with 0.5 mg/kg L-TAC and TAC. Mice were anesthetized with sodium pentobarbitol at 2, 6, 12, 24, or 96 hours, and blood was collected by cardiac puncture. Blood was separated into plasma, white blood cell, and red blood cell fractions using lympholyte-M.
Fig 2. Tissue concentration of tacrolimus. Liver, kidney, spleen, and small intestine were removed, and tissue levels of TAC and L-TAC were assessed at 24 and 96 hours.
RESULTS AND DISCUSSION
Liposomal encapsulation did not significantly alter the plasma kinetics of tacrolimus (Fig 1). It did, however, alter the organ penetration of the drug (Fig 2). Intestinal drug concentration and exposure were increased fourfold (Fig 3). The ratio of the specific activity of 3H-tacrolimus to that of 14C-liposome in the L-TAC preparation was 88:1 before
Fig 1. Plasma concentration of intravenous tacrolimus.
From the Department of Surgery and College of Pharmacy, Dalhousie University, Halifax, Nova Scotia, Canada. Supported by a grant from Fujisawa, Canada. Address reprint requests to V.C. McAlister, Suite 8815 Victoria Wing, QEII Health Sciences Centre, Halifax, Nova Scotia, Canada B3H 2Y9.
© 1998 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
0041-1345/98/$19.00 PII S0041-1345(98)00774-X
Transplantation Proceedings, 30, 2651–2652 (1998)
2651
2652
DUTTA, MEZEI, LEE ET AL
Fig 3. Concentration of tacrolimus in small intestine. Fig 4. Ratio of liposome to tacrolimus in small intestine.
administration. This ratio was maintained by L-TAC found in the small intestine at 24 and 96 hours (Fig 4). These findings suggest that liposomal encapsulation significantly increases penetration and retention of drug in the small intestine (and other organs), and that most drugs found there are liposomally bound. Liposomal encapsulation may improve the effect of tacrolimus in transplant patients.
REFERENCES 1. McAlister VC, Grant DR: In Grant DR, Wood RFM (eds): Small Bowel Transplantation. Edward Arnold; London: 1994, p 121 2. Smeesters C, Giroux L, Vinet B, et al: Cana J Surg 31:34, 1988 3. Ko S, Nakajima Y, Kanehiro H, et al: Transplantation 59:1384, 1995
T
ACROLIMUS (TAC) has allowed routinely successful intestinal transplantation, although the therapeutic window remains problematic.1 Liposomal tacrolimus (LTAC) may allow increased efficacy without nephrotoxicity,2 but effectiveness may depend on adequate graft penetration. Improved canine liver graft survival with L-TAC is associated with higher drug levels in the graft.3 To study the effect of liposomal encapsulation on biodisposition in the intestine, we made a multilayered liposome with 3H-labeled tacrolimus and 14C-labeled liposomes. MATERIALS AND METHODS Liposomes were prepared by a solvent evaporation method. Briefly, 5% phospholipon 90-H, loralan-CH cholesterol, 0.02% tacrolimus, 300 mCi 3H-tacrolimus, and 100 mCi 14C-L-3-phosphatidyl (N-methyl-14C) choline,1-2-dipalmitoyl (14C-DPPC) were dissolved in 20 mL chloroform/methanol (2:1 v/v), and dried by evaporation under vacuum. Ninety-five milliliters of preheated 0.9% NaCl (55°C) were added, and the mixture was hand-shaken for 1 minute at 55°C. The dissolved liposomes were subsequently placed in a preheated Environ Shaker (55°C) for 20 minutes, after which they were slowly brought back to room temperature with constant shaking. Microscopic examination determined the liposomes to be multilamellar. The control solution was prepared by adding 300 mCi 3Htacrolimus to commercial tacrolimus (Prograf), and diluted to a final concentration of 0.02% tacrolimus in 0.9% NaCl. Female CD-1 mice (6 to 8 weeks old) were injected intravenously (tail vein) with 0.5 mg/kg L-TAC and TAC. Mice were anesthetized with sodium pentobarbitol at 2, 6, 12, 24, or 96 hours, and blood was collected by cardiac puncture. Blood was separated into plasma, white blood cell, and red blood cell fractions using lympholyte-M.
Fig 2. Tissue concentration of tacrolimus. Liver, kidney, spleen, and small intestine were removed, and tissue levels of TAC and L-TAC were assessed at 24 and 96 hours.
RESULTS AND DISCUSSION
Liposomal encapsulation did not significantly alter the plasma kinetics of tacrolimus (Fig 1). It did, however, alter the organ penetration of the drug (Fig 2). Intestinal drug concentration and exposure were increased fourfold (Fig 3). The ratio of the specific activity of 3H-tacrolimus to that of 14C-liposome in the L-TAC preparation was 88:1 before
Fig 1. Plasma concentration of intravenous tacrolimus.
From the Department of Surgery and College of Pharmacy, Dalhousie University, Halifax, Nova Scotia, Canada. Supported by a grant from Fujisawa, Canada. Address reprint requests to V.C. McAlister, Suite 8815 Victoria Wing, QEII Health Sciences Centre, Halifax, Nova Scotia, Canada B3H 2Y9.
© 1998 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
0041-1345/98/$19.00 PII S0041-1345(98)00774-X
Transplantation Proceedings, 30, 2651–2652 (1998)
2651
2652
DUTTA, MEZEI, LEE ET AL
Fig 3. Concentration of tacrolimus in small intestine. Fig 4. Ratio of liposome to tacrolimus in small intestine.
administration. This ratio was maintained by L-TAC found in the small intestine at 24 and 96 hours (Fig 4). These findings suggest that liposomal encapsulation significantly increases penetration and retention of drug in the small intestine (and other organs), and that most drugs found there are liposomally bound. Liposomal encapsulation may improve the effect of tacrolimus in transplant patients.
REFERENCES 1. McAlister VC, Grant DR: In Grant DR, Wood RFM (eds): Small Bowel Transplantation. Edward Arnold; London: 1994, p 121 2. Smeesters C, Giroux L, Vinet B, et al: Cana J Surg 31:34, 1988 3. Ko S, Nakajima Y, Kanehiro H, et al: Transplantation 59:1384, 1995