ABSTRACT
New technologies are needed to deliver medicines safely and effectively. Polymeric nanoparticulate carriers are one such technology under investigation. We examined the intracellular trafficking of doxorubicin-bound block copolymers quantitatively and by imaging doxorubicin-derived fluorescence using confocal microscopy. The polymers were internalized by endocytosis and distributed in endosomal/lysosomal compartments and the endoplasmic reticulum; unlike free doxorubicin, the polymers were not found in the nucleus. Moreover, the ATP-binding cassette protein B1 (ABCB1) transporter may be involved in the efflux of the polymer from cells. This drug delivery system is attractive because the endogenous transport system is used for the uptake and delivery of the artificial drug carrier to the target as well as for its efflux from cells to medium. Our results show that a drug delivery system strategy targeting this endogenous transport pathway may be useful for affecting specific molecular targets.
Subject(s)
Antibiotics, Antineoplastic/metabolism , Doxorubicin/metabolism , Drug Carriers , Polyethylene Glycols/metabolism , Uterine Cervical Neoplasms/metabolism , ATP Binding Cassette Transporter, Subfamily B , ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , ATP Binding Cassette Transporter, Subfamily B, Member 1/metabolism , Antibiotics, Antineoplastic/chemistry , Biological Transport , Cell Survival/drug effects , Chemistry, Pharmaceutical , Doxorubicin/chemistry , Drug Compounding , Endocytosis , Endoplasmic Reticulum/metabolism , Endosomes/metabolism , Female , HeLa Cells , Humans , Hydrolysis , Hydrophobic and Hydrophilic Interactions , Lysosomes/metabolism , Micelles , Microscopy, Confocal , Polyethylene Glycols/chemistry , Polyethylene Glycols/toxicity , RNA Interference , Technology, Pharmaceutical/methods , Time Factors , Transfection , Uterine Cervical Neoplasms/geneticsABSTRACT
Doxorubicin, a highly effective anticancer drug, produces severe side effect such as cardiotoxicity, which is mainly caused by its metabolite, doxorubicinol. While in vitro studies by measuring cellular concentration of doxorubicin have been reported, there have been no reports on measuring cellular concentration of the metabolites. In this report, we developed a sensitive and high-throughput method for measuring cellular concentrations of doxorubicin and its metabolites by ultra-high-performance liquid chromatography. The method achieved more than 96% recovery of doxorubicin and its metabolites from cell homogenates. Using simple separation conditions, doxorubicin and its three main metabolites, and the internal standard, were separated within 3 min. The method has a limit of quantification of 17.4 pg (32.0 fmol) injected doxorubicin. This high sensitivity enables the detection and intracellular quantification of doxorubicin and its metabolite, doxorubicinol, in cell homogenates, and its use will facilitate studies of the relationship between doxorubicin pharmacokinetics and therapeutic outcome.
Subject(s)
Chromatography, High Pressure Liquid/methods , Doxorubicin , Doxorubicin/analogs & derivatives , Doxorubicin/analysis , Doxorubicin/chemistry , HT29 Cells , HeLa Cells , Humans , Intracellular Space , Linear Models , Sensitivity and SpecificityABSTRACT
Biomolecules, especially large polymeric molecules such as enzymes and antibodies, mediate various biological functions, including biochemical reactions and molecular recognition, with high reactivity, efficiency, selectivity and accuracy. Many researchers have investigated methods to take advantage of these characteristics in analytical devices. One way to accomplish this is to immobilize biomolecules in the devices. For example, biomolecules have been immobilized by means of silica sol-gel technology and used for basic research in the food and pharmaceutical industries. Proteins encapsulated by this method retain their structure and biological activity for a prolonged period. This review describes methodologies for immobilization of biomolecules and the applications of sol-gel technology to analytical devices, especially flow-through systems.
