Universal Biotin-PEG-Linked Gold Nanoparticle Probes for the Simultaneous Detection of Nucleic Acids and Proteins.

Novel biotin-polyethylene glycol (biotin-PEG) gold nanoparticle probes have been synthesized and used as universal constructs for the detection of protein (prostate-specific antigen, PSA) and nucleic acid targets (microRNAs) from a single sample. Microarray assays based upon these probes enabled sensitive detection of biomarker targets (50 fM for nucleic acid targets and 1 pg/µL for the PSA target). Ways of detecting biomarkers, including nucleic acids and proteins, are necessary for the clinical diagnosis of many diseases, but currently available diagnostic platforms rely primarily on the independent detection of proteins or nucleic acids. In addition to the economic benefits associated with the use of a single platform to detect both classes of analytes, studies have shown that the simultaneous identification of multiple classes of biomarkers in the same sample could be useful for the detection and management of early stage diseases, especially when sample amounts are limited. Therefore, these new probes and the assays based upon them open the door for high-sensitivity combination-target assays for studying and tracking biological pathways and diseases.

Biodegradable DNA-Brush Block Copolymer Spherical Nucleic Acids Enable Transfection Agent-Free Intracellular Gene Regulation.

By grafting multiple DNA strands onto one terminus of a polyester chain, a DNA-brush block copolymer that can assemble into micelle structure is constructed. These micelle spherical nucleic acids have a density of nucleic acids that is substantively higher than linear DNA block copolymer structures, which makes them effective cellular transfection and intracellular gene regulation agents.

The Sequence-Specific Cellular Uptake of Spherical Nucleic Acid Nanoparticle Conjugates.

The sequence-dependent cellular uptake of spherical nucleic acid nanoparticle conjugates (SNAs) is investigated. This process occurs by interaction with class A scavenger receptors (SR-A) and caveolae-mediated endocytosis. It is known that linear poly(guanine) (poly G) is a natural ligand for SR-A, and it has been proposed that interaction of poly G with SR-A is dependent on the formation of G-quadruplexes. Since G-rich oligonucleotides are known to interact strongly with SR-A, it is hypothesized that SNAs with higher G contents would be able to enter cells in larger amounts than SNAs composed of other nucleotides, and as such, cellular internalization of SNAs is measured as a function of constituent oligonucleotide sequence. Indeed, SNAs with enriched G content show the highest cellular uptake. Using this hypothesis, a small molecule (camptothecin) is chemically conjugated with SNAs to create drug-SNA conjugates and it is observed that poly G SNAs deliver the most camptothecin to cells and have the highest cytotoxicity in cancer cells. Our data elucidate important design considerations for enhancing the intracellular delivery of spherical nucleic acids.

Biocompatible infinite-coordination-polymer nanoparticle-nucleic-acid conjugates for antisense gene regulation.

Herein, we report the synthesis of DNA-functionalized infinite-coordination-polymer (ICP) nanoparticles as biocompatible gene-regulation agents. ICP nanoparticles were synthesized from ferric nitrate and a ditopic 3-hydroxy-4-pyridinone (HOPO) ligand bearing a pendant azide. Addition of Fe(III) to a solution of the ligand produced nanoparticles, which were colloidally unstable in the presence of salts. Conjugation of DNA to the Fe(III)-HOPO ICP particles by copper-free click chemistry afforded colloidally stable nucleic-acid nanoconstructs. The DNA-ICP particles, when cross-linked through sequence-specific hybridization, exhibited narrow, highly cooperative melting transitions consistent with dense DNA surface loading. The ability of the DNA-ICP particles to enter cells and alter protein expression was also evaluated. Our results indicate that these novel particles carry nucleic acids into mammalian cells without the need for transfection agents and are capable of efficient gene knockdown.