Optimization of liposomal retention of vincristine
Creators: name: Boman, Nancy L.
Publisher: name: University of British Columbia
Year: 2009

description: Vincristme is a widely used antineoplastic agent. It is a cell cycle specific drug arresting cell growth during metaphase. Since tumors contain dividing cells distributed throughout the cell cycle, it is important to achieve as long an exposure time as possible. Encapsulation of drugs within liposomes can prolong circulation time in vivo as well as reduce toxic side effects. This thesis examines the effect of liposomal encapsulation on reducing toxicity as well as improving antineoplastic activity of vincristine by altering drug pharmacokinetic and biodistribution behaviour. The first focus of investigation demonstrates that encapsulation of vincristine within liposomes greatly reduces soft-tissue toxicity of the drug. Subcutaneous injections of liposomal vincristine are shown to demonstrate minimal toxic effects whereas similar injections of free vincristine result in gross necrosis and ulceration. Free drug is rapidly cleared from the area of injection. Liposomal drug remains at the area of injection much longer, but remains trapped within the liposomes. Slow release rates presumably prevent it from exerting cytotoxic effects. The next topic concerns improving the retention of vincristine within liposomes. The influence of lipid composition, internal pH and internal buffering capacity on the retention properties of vincristine loaded into LUVs in response to transmembrane pH gradients has been assessed. It is shown that increasing the (saturated) acyl chain length of the phosphatidyicholine molecule, increasing the internal buffering capacity, and decreasing the internal pH all result in increased drug retention. Further, a study of the pH dependence on the rates of accumulation indicate that uptake proceeds via the neutral form of the vincristine molecule. This uptake is associated with an activation energy of 37 kcallmol for DSPCICho1 LUVs. It is shown that the major improvement in drug retention in vitro is achieved by employing low initial internal pH values, where 90% retention is obtained over 24 h for an initial internal pH of 2. Improved retention over the same system with an internal pH of 4 in viva was also observed where a drug-to-lipid ratio approximately 4-fold greater at 24 h was maintained. The third area of investigation concerns the incorporation of cationic lipids to further improve vincristine retention within liposomes. The influence of both the incorporation of 10 mol% cationic lipid (AL-l, stearylamine, or sphingosine) into DSPC/Chol (55:45; mol:mol) vesicles and lowering the internal pH to pH 2.0 on the circulation life-time and antitumor activity of liposomal vincristine systems (drug-to-lipid ratio of 0.1:1) has been examined. With an internal pH of 2.0, the incorporation of 10 mol% cationic lipid is shown to significantly increase drug retention within the liposomes without affecting lipid clearance times. The resulting increase in plasma drug concentration seen by the incorporation of 10 mol% sphingosine results in a significant increase in therapeutic activity against the P388 lymphocytic leukemia cell line in viva. The final area of investigation examines two different methods for increasing the circulation longevity of vincristine encapsulated in liposomes. The first involves incorporation of the ganglioside GM₁, which acts to increase the circulation longevity of liposomal carriers, while the second approach relies on modification of the vincristine encapsulation procedure to enhance drug retention. It is shown that these approaches are synergistic and increase the circulation half-life of vincristine from approximately one hour to greater than 12 hours. This results in a dramatic improvement in the therapeutic activity of liposomal vincristirie as measured using a murine P388 lymphocytic leukemia model. At doses above 2 mgfkg the optimized liposomal vincristine formulation cures greater than 50% of mice bearing the P388 tumor, whereas free vincristine results in no cures. The optimized formulation is also shown to significantly increase solid tumor uptake of vincristine within the Lewis Lung tumor model and result in improved therapeutic activity.