Studies on the impact of amine-containing compounds on lysosomes

Abstract

Lysosomes accumulate weakly basic amine-containing drugs (i.e., lysosomotropic amines) through an ion trapping-type mechanism and are capable of disrupting lysosomal structure and function. A common finding following treatment with hydrophobic amine-containing drugs is a cellular lipidosis marked by the lysosomal accumulation of lipids. The impact of this drug-induced lipidosis on the lysosomal accumulation of weakly basic amine-containing drugs is unknown. In this work, hydrophobic (logP 2) amine-containing drugs (imipramine, haloperidol, risperidone, chlorpromazine, lidocaine, bupivacaine, amiodarone, verapamil, propranolol) were found to cause a significant increase in the cellular accumulation of the lysosomotropic amine, LysoTracker Red (LTR). Imipramine was further found to specifically increase the cellular accumulation of amine-containing substrates of lysosomal ion trapping (LysoTracker Green, daunorubicin, methylamine, propranolol) but not compounds that don't accumulate by ion trapping in lysosomes, suggesting that the drug-induced lipidosis specifically increases the cellular accumulation of drugs that accumulate in lysosomes. Imipramine-induced accumulation of LTR occurred in a time- and temperature-dependent manner in agreement with an energy-dependent intracellular remodeling process that has been reported to coincide with induction of the cellular lipidosis. Prevention of the imipramine-induced lipidosis by treatment with 0.1% hydroxypropyl-ß-cyclodextrin or growth in lipoprotein-depleted media prevented the increased accumulation of LTR, suggesting the hyperaccumulation of LTR is secondary to the drug-induced lipidosis. Determination of lysosome specific accumulation of LTR in cells with or without an intact lysosomal pH gradient revealed that imipramine caused an approximately three-fold increase in the lysosome-specfic uptake of LTR. Previous reports have suggested that drugs accumulate following a cellular lipidosis through enhanced hydrophobic binding sites. However, these results are consistent with an increased aqueous volume of lysosomes resulting in increased drug accumulation. Theoretical calculations of lysosomal volume changes following imipramine treatment reveal an approximately 4-fold increase in apparent lysosomal volume. Therefore, drug-induced lipidosis is found to increase the cellular accumulation of amine-containing drugs by causing a marked increase in the lysosomal volume of cells. These findings have implications towards understanding how drugs can interact at the intracellular level. The ability of one drug to affect the distribution properties of a second is further hypothesized to result in drug-drug interactions.