Synthesis and Biological Evaluation of Novel Folic Acid Receptor-Targeted, β-Cyclodextrin-Based Drug Complexes for Cancer Treatment

Drug targeting is an active area of research and nano-scaled drug delivery systems hold tremendous potential for the treatment of neoplasms. In this study, a novel cyclodextrin (CD)-based nanoparticle drug delivery system has been assembled and characterized for the therapy of folate receptor-positive [FR(+)] cancer. Water-soluble folic acid (FA)-conjugated CD carriers (FACDs) were successfully synthesized and their structures were confirmed by 1D/2D nuclear magnetic resonance (NMR), matrix-assisted laser desorption ionization time-of-flight mass spectrometer (MALDI-TOF-MS), high performance liquid chromatography (HPLC), Fourier transform infrared spectroscopy (FTIR), and circular dichroism. Drug complexes of adamatane (Ada) and cytotoxic doxorubicin (Dox) with FACD were readily obtained by mixed solvent precipitation. The average size of FACD-Ada-Dox was 1.5–2.5 nm. The host-guest association constant Ka was 1,639 M−1 as determined by induced circular dichroism and the hydrophilicity of the FACDs was greatly enhanced compared to unmodified CD. Cellular uptake and FR binding competitive experiments demonstrated an efficient and preferentially targeted delivery of Dox into FR-positive tumor cells and a sustained drug release profile was seen in vitro. The delivery of Dox into FR(+) cancer cells via endocytosis was observed by confocal microscopy and drug uptake of the targeted nanoparticles was 8-fold greater than that of non-targeted drug complexes. Our docking results suggest that FA, FACD and FACD-Ada-Dox could bind human hedgehog interacting protein that contains a FR domain. Mouse cardiomyocytes as well as fibroblast treated with FACD-Ada-Dox had significantly lower levels of reactive oxygen species, with increased content of glutathione and glutathione peroxidase activity, indicating a reduced potential for Dox-induced cardiotoxicity. These results indicate that the targeted drug complex possesses high drug association and sustained drug release properties with good biocompatibility and physiological stability. The novel FA-conjugated β-CD based drug complex might be promising as an anti-tumor treatment for FR(+) cancer.

Testing the Efficacy of Panax ginseng Extract as a Novel Anti-Quorum Sensing Agent

Quorum sensing (QS) is the phenomenon by which microorganisms regulate gene expression in response to cell-population density. These microorganisms synthesize and secrete small molecules known as autoinducers that increase in concentration as a function of cell density. Once the cell detects the minimal threshold concentration of an autoinducer, the gene expression is altered accordingly. Although the cellular circuitry responding to QS is relatively conserved, the cellular processes regulated by QS, for example are conjugation, motility, sporulation, biofilm formation and production of virulence factors importamt for infection. Since many pathogens have developed resistance to available antibiotics, novel therapies are required to further treat these pathogens. Inhibitors of QS are potential therapeutic candidates since they would inhibit virulence without selecting for antibiotic resistant strains. Previous studies have shown that the Chinese herb Panax ginseng contains compounds that inhibit QS activity. To further characterize this activity, a highly sensitive quantitative liquid assay was developed using a Chromobacterium violaceum AHL mutant, CV026 as a biomonitor strain. After confirmation that P. ginseng aqueous extracts had anti-QS activity, the extract was fractionated on a reverse phase C18 Sep Pak column. Most anti-QS activity was present in the flow through, and compounds eluted with ten percent acetonitrile in water antibacterial activity. We thus conclude that P. ginseng has anti-QS activity and the active compound is water soluble. Compounds from P. ginseng, could be used as a lead structure to design compound with higher anti-QS activity for therapeutic treatments.

Contribution of lipophilic secondary metabolites to the toxicity of strains of freshwater cyanobacterial harmful algal blooms, identified using the zebrafish (Danio rerio) embryo as a model for vertebrate development

Cyanobacteria ("blue-green algae") are known to produce a diverse repertoire of biologically active secondary metabolites. When associated with so-called "harmful algal blooms", particularly in freshwater systems, a number of these metabolites have been associated—as "toxins", or commonly "cyanotoxins"—with human and animal health concerns. In addition to the known water-soluble toxins from these genera (i.e. microcystins, cylindrospermopsin, and saxitoxins), our studies have shown that there are metabolites within the lipophilic extracts of these strains that inhibit vertebrate development in zebrafish embryos. Following these studies, the zebrafish embryo model was implemented in the bioassay-guided purification of four isolates of cyanobacterial harmful algal blooms, namely Aphanizomenon, two isolates of Cylindrospermopsis, and Microcystis, in order to identify and chemically characterize the bioactive lipophilic metabolites in these isolates. ^ We have recently isolated a group of polymethoxy-1-alkenes (PMAs), as potential toxins, based on the bioactivity observed in the zebrafish embryos. Although PMAs have been previously isolated from diverse cyanobacteria, they have not previously been associated with relevant toxicity. These compounds seem to be widespread across the different genera of cyanobacteria, and, according to our studies, suggested to be derived from the polyketide biosynthetic pathway which is a common synthetic route for cyanobacterial and other algal toxins. Thus, it can be argued that these metabolites are perhaps important contributors to the toxicity of cyanobacterial blooms. In addition to the PMAs, a set of bioactive glycosidic carotenoids were also isolated because of their inhibition of zebrafish embryonic development. These pigmented organic molecules are found in many photosynthetic organisms, including cyanobacteria, and they have been largely associated with the prevention of photooxidative damage. This is the first indication of these compounds as toxic metabolites and the hypothesized mode of action is via their biotransformation to retinoids, some of which are known to be teratogenic. Additional fractions within all four isolates have been shown to contain other uncharacterized lipophilic toxic metabolites. This apparent repertoire of lipophilic compounds may contribute to the toxicity of these cyanobacterial harmful algal blooms, which were previously attributed primarily to the presence of the known water-soluble toxins.^