Herein we describe a nucleic acid functionalized nanocapsule in which nucleic acid ligands are assembled and disassembled in the presence of enzymes. The particles are fully degradable in response to esterases due to an embedded ester cross-linker in the particle's core. During synthesis the nanocapsules can be loaded with hydrophobic small molecules and post self-assembly undergo covalent cross-linking using copper catalyzed click chemistry. They can then be functionalized with thiolated DNA through stepwise thiolyne chemistry using UV light irradiation. Additionally, the capsule is compatible with enzyme mediated functionalization of a therapeutic mRNA-cleaving DNAzyme at the particle's surface. The resulting particle is highly stable, monodisperse in size, and maximizes the therapeutic potential of both the particles interior and exterior.
Aberrant glycosylation has been linked to many different cancer types. In breast cancer metastasis to the brain the blood brain barrier, a region of the brain that regulates the entrance of ions, diseases, toxins, etc., fails to block breast cancer cells from crossing. Here we present a study of identifying and quantifying the glycosylation of six breast and brain cancer cell lines using hydrophilic interaction liquid chromatography (HILIC) and electrostatic repulsion liquid chromatography (ERLIC) enrichments and LC-MS/MS analysis. Qualitative and quantitative analyses of N-linked glycosylation were performed by both enrichment techniques for individual and complementary comparison. Potential cancer glycopeptide biomarkers were identified and confirmed by chemometric and statistical evaluations. A total of 497 glycopeptides were characterized of which 401 were common glycopeptides (80.6% overlap) identified from both enrichment techniques. HILIC enrichment yielded 320 statistically significant glycopeptides in 231BR relative to the other cell lines out of 494 unique glycopeptides, and sequential HILIC-ERLIC enrichment yielded 212 statistically significant glycopeptides in 231BR compared to the other cell lines out of 404 unique glycopeptides. The results provide the first comprehensive glycopeptide listing for these six cell lines.
Herein, we describe the characterization of a novel self-assembling and intracellular disassembling nanomaterial for nucleic acid delivery and targeted gene knockdown. By using a recently developed nucleic acid nanocapsule (NAN) formed from surfactants and conjugated DNAzyme (DNz) ligands, it is shown that DNz-NAN can enable cellular uptake of the DNAzyme and result in 60 % knockdown of a target gene without the use of transfection agents. The DNAzyme also exhibits activity without chemical modification, which we attribute to the underlying nanocapsule design and release of hydrophobically modified nucleic acids as a result of enzymatically triggered disassembly of the NAN. Fluorescence-based experiments indicate that the surfactant-conjugated DNAzymes are better able to access a fluorescent mRNA target within a mock lipid bilayer system than the free DNAzyme, highlighting the advantage of the hydrophobic surfactant modification to the nucleic acid ligands. In vitro characterization of DNz-NAN's substrate-cleavage kinetics, stability in biological serum, and persistence of knockdown against a proinflammatory transcription factor, GATA-3, are presented.
Using
a recently developed nucleic acid delivery platform, we demonstrate
the effective delivery of metallodrug [AuIIIBr2(SSC-Inp-OEt)] (AP228; Inp = isonipecotic moiety), a hydrophobic,
low solubility gold complex cytotoxic to cancer cells. It is shown
that AP228 is delivered more effectively into HeLa cells using micellular
surfactant assemblies compared to that of a more polar derivative
[AuIIIBr2(SSC-Inp-GlcN1)] (AP209; GlcN1 = (α,β)-d-glucosamino moiety). When AP228 is codelivered with siRNA
targeting Bcl-2, a key regulator of apoptosis, the overall cytotoxic
therapeutic effects of the drug are maximized. The optimized delivery
and distribution of the compound is monitored by both fluorescence
microscopy and inductively coupled plasma mass spectrometry. We show
that codelivery of the AP228 and Bcl-2 targeting siRNA results in
a substantial increase in drug efficacy, wherein the cytotoxic therapeutic
effects of the drug are maximized, reducing the IC50 from
760 nM to 11 nM. This hybrid small molecule drug and therapeutic nucleic
acid delivery vehicle is shown to enable both the improved solubility
and uptake of the gold(III) metallodrugs and the delivery of chemically
unmodified siRNA, resulting in enhanced cytotoxic effects.
Enhanced stability and dual-responsive behavior were imparted to ZIF-8 MOFs via encapsulation in DNA-surfactant conjugates and a peptide crosslinked micelle.
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