Lipid bilayer partitioning and stability of camptothecin drugs

Abstract

The intense intrinsic fluorescence emissions from several clinically relevant camptothecin drugs have been exploited in order to determine (1) the structural basis of drug binding to lipid bilayers, (2) the lipid bilayer stability of each drug's alpha-hydroxylactone moiety, a pharmacophore which is essential for antitumor activity, and (3) the site of drug binding in the bilayer. Equilibrium affinities of camptothecin and related congeners for small unilamellar vesicles composed of electroneutral dimyristoylphosphatidylcholine (DMPC) or negatively-charged dimyristoylphosphatidylglycerol (DMPG) were determined using the method of fluorescence anisotropy titration. Experiments were conducted in phosphate-buffered saline (PBS) at 37 degrees C and overall association constants (K values) were determined. Of the seven compounds studied, the new compound 9-chloro-10,11-methylenedioxy-(20S)-camptothecin (CMC) was found to display the highest membrane affinities (KDMPC = 400 M-1, KDMPG = 320 M-1), followed by 10,11-methylenedioxy-camptothecin and camptothecin, which exhibited KDMPC and KDMPG values of 100 M-1 or greater. Topotecan displayed markedly reduced binding to lipid bilayers (KDMPC = 10 M-1, KDMPG = 50 M-1). HPLC assays were subsequently employed to assess the relative stabilities of the lactone ring of membrane-bound drugs. Our results clearly indicate that lipid bilayer interactions stabilize the lactone moiety of camptothecin drugs. In comparison to half-lives in PBS (37 degrees C) of 17 and 19 min for camptothecin and CMC, respectively, DMPC- or DMPG-bound drugs were found to be stable even for periods up to 72 h. Iodide quenching data indicate that membrane-bound camptothecin intercalates between the lipid acyl chains, in a protected environment well removed from the aqueous interface. In this manner lipid bilayer interactions stabilize the lactone ring structure of camptothecins and thereby conserve the biologically active form of each medication.

Liposomal Irinotecan

“…Previous lipid-based techniques used to encapsulate camptothecins have exploited their ability to partition into the lipid bilayer. This has been shown to confer protection to the lactone species of irinotecan (44,45). Therapeutically, this approach is limited by low drug loading efficiencies and the rapid exchange of membrane-localized drug from the liposomes to endogenous membranes after i.v.…”

Determination of intraliposomal pH and its effect on membrane partitioning and passive loading of a hydrophobic camptothecin, DB-67

“…Previously, liposomal formulation strategies for neutral camptothecins have focused on the benefits of membrane binding of the lactone to improve the stability of camptothecins toward hydrolysis (Burke et al, 1993;Peikov et al, 2005;Zhang et al, 2004). We have recently investigated the potential of a novel pH gradient strategy for improving the liposomal retention of neutral camptothecins such as DB-67 by encapsulating the drug in rigid gel phase liposomes at high pH (Joguparthi and Anderson, 2007;Joguparthi et al, 2007).…”

Determination of the Protein-Binding Properties of Camptothecins by Means of Optical Spectroscopy Methods

“…Using method described in [12], the double-reciprocal plots were drawn (C and D) and the association constants of drugs binding to albumin were calculated (Table III). The uorescence anisotropy of 9-amino-CPT was measured at pH 3 because of low uorescence of this compound at pH 7.4 [15]. Unfortunately, the addition of albumin causes a further uorescence quenching which makes determination of a nity constant of 9-amino-CPT by the anisotropy measurements impossible.…”