ABSTRACT
In this study, we present a targeted drug delivery system to improve intravesical therapy of bladder diseases. The drug delivery system consists of wheat germ agglutinin (WGA) to facilitate specific interaction with the surface of bladder cells and α-poly-(L)-glutamic acid (PGA) as polymeric backbone to increase the number of drug molecules per targeting moiety. Additionally, fluorescein cadaverine was coupled to PGA to visualise and track the delivery system. Using 5637 single cells and cell monolayers, the optimised F-PGA-WGA delivery system, with an approximate molecular weight of 670kDa, could convince with its promising cytoadhesive as well as cytoinvasive potential. Using the competitive inhibitor N, N', Nâ³-triacetylchitotriose a specificity of the carbohydrate-mediated interaction between the cell and the delivery system of up to 98% was determined. F-PGA alone did not show any interaction with the cells. Moreover, a high drug loading of 77 molecules of the model drug Dansylcadaverine per backbone was achieved. Microscopic analysis further confirmed binding and uptake of the cytoadhesive polymer even after additional loading with the model drug. Combining the auspicious targeting properties of WGA with the high drug loading possibilities of the backbone might finally lead to an enhanced efficacy when used for intravesical therapy.
Subject(s)
Cadaverine/analogs & derivatives , Polyglutamic Acid/chemistry , Urinary Bladder Diseases/drug therapy , Wheat Germ Agglutinins/chemistry , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacokinetics , Biocompatible Materials , Biological Transport , Cadaverine/administration & dosage , Cadaverine/chemistry , Cadaverine/pharmacokinetics , Cell Line, Tumor , Drug Delivery Systems , Humans , Urothelium/cytologyABSTRACT
The rates of acid hydrolysis of N-benzoyl-cadaverine (1), mono-N-(tert-butoxy)carbonyl cadaverine (2), and benzaldoxime (3) with binding motifs for cucurbit[6]uril (1,2) and cucurbit[7]uril (1,3) were investigated in the absence and presence of these hosts. Significant rate enhancements (up to a factor of ca. 300 for the hydrolysis of 3) were observed. Competitive inhibition due to encapsulation of added cadaverine and the successful use of sub-stoichiometric amounts of macrocycle confirmed the function of cucurbiturils in promoting acid hydrolysis.
Subject(s)
Cadaverine/pharmacokinetics , Macrocyclic Compounds/chemistry , Bridged-Ring Compounds/chemistry , Cadaverine/chemistry , Catalysis , Hydrolysis , Imidazoles/chemistry , Molecular StructureABSTRACT
A chemotherapeutic vitamin D analogue, EB1089, kills tumor cells via a caspase-independent pathway that results in chromatin condensation and DNA fragmentation. Employing transmission- and immunoelectronmicroscopy as well as detection of autophagosome-associated LC3-beta protein in the vacuolar structures, we show here that EB1089 also induces massive autophagy in MCF-7 cells. Interestingly, inhibition of autophagy effectively hindered apoptosis-like nuclear changes and cell death in response to EB1089. Furthermore, restoration of normal levels of beclin 1, an autophagy-inducing tumor suppressor gene that is monoallelically deleted in MCF-7 cells, greatly enhanced the EB1089-induced nuclear changes and cell death. Thus, EB1089 triggers nuclear apoptosis via a pathway involving Beclin 1-dependent autophagy. Surprisingly, tumor cells depleted for Beclin 1 failed to proliferate suggesting that even though the monoallelic depletion of beclin 1 in human cancer cells suppresses EB1089-induced autophagic death, one intact beclin 1 allele is essential for tumor cell proliferation.
Subject(s)
Antineoplastic Agents/pharmacology , Breast Neoplasms/drug therapy , Calcitriol/analogs & derivatives , Proteins/physiology , Adenine/analogs & derivatives , Adenine/pharmacology , Apoptosis Regulatory Proteins , Autophagy/drug effects , Beclin-1 , Breast Neoplasms/genetics , Breast Neoplasms/metabolism , Breast Neoplasms/pathology , Cadaverine/analogs & derivatives , Cadaverine/pharmacokinetics , Calcitriol/antagonists & inhibitors , Calcitriol/pharmacology , Cathepsins/metabolism , Cell Death/drug effects , Cell Death/physiology , Cell Growth Processes/drug effects , Cell Growth Processes/physiology , Cell Line, Tumor , Cryoelectron Microscopy , Drug Interactions , Genes, Tumor Suppressor , HeLa Cells , Humans , Immunoblotting , Membrane Proteins , Proteins/genetics , Proteins/metabolism , RNA, Small Interfering/genetics , Transfection , Tumor Cells, Cultured , Tumor Necrosis Factor-alpha/pharmacologyABSTRACT
A method for investigating cell-surface enzymatic oxidative deamination of amino acids and amines in seawater was developed. This technique used synthetic fluorescent Lucifer Yellow derivatives of the amino acid lysine and the amine cadaverine as molecular probes to investigate oxidation pathways and rates. The probes were chemically stable under the conditions used and did not adsorb to container surfaces. The oxidative deamination of the fluorescent probes added to phytoplankton cultures and the subsequent production of their fluorescent oxidation products could be selectively detected by HPLC at 250 pM levels. This approach allows selective investigation of cell-surface enzymatic oxidation since neither transport of the probes across the cell membrane nor chemical transformation of the probes occurs. Bacteria were also capable of oxidizing the fluorescent amino acid probe.
