Identification and analysis of stereoselective drug interactions with low-density lipoprotein by high-performance affinity chromatography

Abstract

Columns containing immobilized low density lipoprotein (LDL) were prepared for the analysis of drug interactions with this agent by high-performance affinity chromatography (HPAC). R/S-Propranolol was used as a model drug for this study. The LDL columns gave reproducible binding to propranolol over 60 h of continuous use in the presence of pH 7.4, 0.067 M potassium phosphate buffer. Experiments conducted with this type of column through frontal analysis indicated that two types of interactions were occurring between R-propranolol and LDL, while only a single type of interaction was observed between S-propranolol and LDL. The first type of interaction, which was seen for both enantiomers, involved non-saturable binding; this interaction had an overall affinity (nKa) of 1.9 (± 0.1) × 105 M-1 for R-propranolol and 2.7 (± 0.2) × 105 M-1 for S-propranolol at 37 °C. The second type of interaction was observed only for R-propranolol and involved saturable binding that had an association equilibrium constant (Ka) of 5.2 (± 2.3) × 105 M-1 at 37 °C. Similar differences in binding behavior were found for the two enantiomers at 20 °C and 27 °C. This is the first known example of stereoselective binding of drugs by LDL or other lipoproteins. This work also illustrates the ability of HPAC to be used as a tool for characterizing mixed-mode interactions that involve LDL and related binding agents.

Analysis of drug interactions with very low density lipoprotein by high-performance affinity chromatography

“…Lipoproteins like VLDL are also known to interact with and transport several types of basic or neutral and hydrophobic drugs in the bloodstream [7,23-30]. For instance, it has recently been proposed in work with HDL and LDL that several binding mechanisms, including both saturable interactions with apolipoproteins and non-saturable interactions with the hydrophobic core, may occur between drugs and these lipoproteins [10,14,31]. …”

“…These techniques have included equilibrium dialysis and capillary electrophoresis (CE) using high-performance frontal analysis [14-17]; however, only equilibrium dialysis has been used in studying drug interactions with VLDL [14]. High-performance affinity chromatography (HPAC) is an alternative to these techniques and has been used in various studies to examine the interactions of drugs with proteins and other binding agents [3-6,10,31-38]. …”

“…Evaluation of the subsequent elution profiles can provide data on the equilibrium constants, binding mechanism and number of interaction sites that are present between the drug and immobilized agent. Advantages of this approach include its speed, precision, ease of automation, and good correlation versus solution-phase results and reference methods [3-6,10,31-38]. …”

“…HPAC has been utilized in previous reports to examine the binding of drugs such as propranolol with HDL and LDL [10,31]. The use of HPAC to perform binding studies of drugs with HDL and LDL has led to the confirmation of a non-saturable binding mechanism, as has been proposed in work with other techniques [14-17], as well as the identification of high affinity binding between propranolol and some apolipoproteins [10,31].…”

“…The use of HPAC to perform binding studies of drugs with HDL and LDL has led to the confirmation of a non-saturable binding mechanism, as has been proposed in work with other techniques [14-17], as well as the identification of high affinity binding between propranolol and some apolipoproteins [10,31]. In the case of LDL, this latter process has been found to be stereoselective, with the R- enantiomer but not the S -enantiomer of propranolol undergoing this type of interaction [10]. …”