Photoacoustic (PA) imaging is continuing to be applied for physiological imaging and more recently for molecular imaging of living subjects. Owing to its high spatial resolution in deep tissues, PA ...imaging holds great potential for biomedical applications and molecular diagnostics. There is however a lack of probes for targeted PA imaging, especially in the area of enzyme-activatable probes. Here we introduce a molecular probe, which upon proteolytic processing is retained at the site of enzyme activity and provides PA contrast. The probe oligomerizes via a condensation reaction and accumulates in cells and tumors that express the protease. We demonstrate that this probe reports furin and furin-like activity in cells and tumor models by generating a significantly higher PA signal relative to furin-deficient and nontarget controls. This probe could report enzyme activity in living subjects at depths significantly greater than fluorescence imaging probes and has potential for molecular imaging in deep tumors.
The dynamic interactions between complex molecular structures underlie a wide range of sophisticated behaviors in biological systems. In building artificial molecular machines out of DNA, an ...outstanding challenge is to develop mechanisms that can control the kinetics of interacting DNA nanostructures and that can compose the interactions together to carry out system-level functions. Here we show a mechanism of DNA tile displacement that follows the principles of toehold binding and branch migration similar to DNA strand displacement, but occurs at a larger scale between interacting DNA origami structures. Utilizing this mechanism, we show controlled reaction kinetics over five orders of magnitude and programmed cascades of reactions in multi-structure systems. Furthermore, we demonstrate the generality of tile displacement for occurring at any location in an array in any order, illustrated as a tic-tac-toe game. Our results suggest that tile displacement is a simple-yet-powerful mechanism that opens up the possibility for complex structural components in artificial molecular machines to undergo information-based reconfiguration in response to their environments.
The original version of this Article omitted a reference to previous work in 'Stojanovic, M. N. & Stefanovic, D. A deoxyribozyme-based molecular automaton. Nat. Biotechnol. 21, 1069-1074 (2003)'. ...This has been added as reference 42. The following has been added after the third sentence of the fifth paragraph of the Discussion: 'Integration could also allow more sophisticated information processing, for example as shown by the classic deoxyribozyme-based automaton that plays tic-tac-toe
, to direct structural reconfiguration (Supplementary Discussion)'. This has been corrected in the PDF and HTML versions of the Article.
Directed self-assembly of small molecules in living systems could enable a myriad of applications in biology and medicine, and already this has been used widely to synthesize supramolecules and ...nano/microstructures in solution and in living cells. However, controlling the self-assembly of synthetic small molecules in living animals is challenging because of the complex and dynamic in vivo physiological environment. Here we employ an optimized first-order bioorthogonal cyclization reaction to control the self-assembly of a fluorescent small molecule, and demonstrate its in vivo applicability by imaging caspase-3/7 activity in human tumour xenograft mouse models of chemotherapy. The fluorescent nanoparticles assembled in situ were imaged successfully in both apoptotic cells and tumour tissues using three-dimensional structured illumination microscopy. This strategy combines the advantages offered by small molecules with those of nanomaterials and should find widespread use for non-invasive imaging of enzyme activity in vivo.
The electronic and optical properties of colloidal quantum dots, including the wavelengths of light that they can absorb and emit, depend on the size of the quantum dots. These properties have been ...exploited in a number of applications including optical detection, solar energy harvesting and biological research. Here, we report the self-assembly of quantum dot complexes using cadmium telluride nanocrystals capped with specific sequences of DNA. Quantum dots with between one and five DNA-based binding sites are synthesized and then used as building blocks to create a variety of rationally designed assemblies, including cross-shaped complexes containing three different types of dots. The structure of the complexes is confirmed with transmission electron microscopy, and photophysical studies are used to quantify energy transfer among the constituent components. Through changes in pH, the conformation of the complexes can also be reversibly switched, turning on and off the transfer of energy between the constituent quantum dots.
A major drawback with current cancer therapy is the prevalence of unrequired dose‐limiting toxicity to non‐cancerous tissues and organs, which is further compounded by a limited ability to rapidly ...and easily monitor drug delivery, pharmacodynamics and therapeutic response. In this report, the design and characterization of novel multifunctional “theranostic” nanoparticles (TNPs) is described for enzyme‐specific drug activation at tumor sites and simultaneous in vivo magnetic resonance imaging (MRI) of drug delivery. TNPs are synthesized by conjugation of FDA‐approved iron oxide nanoparticles ferumoxytol to an MMP‐activatable peptide conjugate of azademethylcolchicine (ICT), creating CLIO‐ICTs (TNPs). Significant cell death is observed in TNP‐treated MMP‐14 positive MMTV‐PyMT breast cancer cells in vitro, but not MMP‐14 negative fibroblasts or cells treated with ferumoxytol alone. Intravenous administration of TNPs to MMTV‐PyMT tumor‐bearing mice and subsequent MRI demonstrates significant tumor selective accumulation of the TNP, an observation confirmed by histopathology. Treatment with CLIO‐ICTs induces a significant antitumor effect and tumor necrosis, a response not observed with ferumoxytol. Furthermore, no toxicity or cell death is observed in normal tissues following treatment with CLIO‐ICTs, ICT, or ferumoxytol. These findings demonstrate proof of concept for a new nanotemplate that integrates tumor specificity, drug delivery and in vivo imaging into a single TNP entity through attachment of enzyme‐activated prodrugs onto magnetic nanoparticles. This novel approach holds the potential to significantly improve targeted cancer therapies, and ultimately enable personalized therapy regimens.
The design and characterization of novel multifunctional “theranostic” nanoparticles (TNPs) is described for enzyme‐specific drug activation at tumor sites and simultaneous in vivo magnetic resonance imaging (MRI) of drug delivery. TNPs consist of iron oxide core for MR imaging, MMP‐14 cleavable peptide linker for specific activation in tumors, and a prodrug that is non‐toxic unless activated.
Bright, photostable luminescent labels are powerful tools for the in vitro and in vivo imaging of biological events. Semiconductor nanocrystals have emerged as attractive alternatives to commonly ...used organic lumophores because of their high quantum yields and the spectral tunability that can be achieved through synthetic control. Although conventional synthetic methods generally yield high-quality nanocrystals with excellent optical properties for biological imaging, ligand exchange and biological conjugation are necessary to make nanocrystals biocompatible and biospecific. These steps can substantially deteriorate the optical characteristics of these nanocrystals. Moreover, the complexity of multistep nanocrystal synthesis, typically requiring inert and anhydrous conditions, prohibits many end users of these lumiphores from generating their own custom materials. We sought to streamline semiconductor nanocrystal synthesis and develop synthetic routes that would be accessible to scientists from all disciplines. In search of such an approach, we turned to nucleic acids as a programmable and versatile ligand set and found that these biomolecules are indeed appropriate for biocompatible semiconductor nanocrystals preparation. In this Account, we summarize our work on nucleic acids-programmed nanocrystal synthesis that has resulted in the successful development of a one-step synthesis of biofunctionalized nanocrystals in aqueous solution. We first discuss results obtained with nucleotide-capped cadmium and lead chalcogenide-based nanocrystals that served to guide further investigation of polynucleotide-assisted synthesis. We investigated the roles of individual nucleobases and their structures in passivation of the surfaces of nanocrystals and modulating morphology and optical characteristics. The nucleic acid structures and sequences and the reaction conditions greatly influence the nanocrystals’ optical properties and morphologies. Moreover, studies using live cells reveal low toxicity and rapid uptake of DNA-passivated CdS nanocrystals, demonstrating their suitability for bioimaging. Finally, we describe a new approach that leads to the production of biofunctionalized, DNA-capped nanocrystals in a single step. Chimeric DNA molecules enable this strategy, providing both a domain for nanocrystal passivation and a domain for biomolecule recognition. Nanocrystals synthesized using this approach possess good spectral characteristics as well as high specificity to cognate DNA, protein, and cancer cell targets. Overall, this approach could make nanocrystal lumiphores more readily accessible to researchers working in the biological sciences.
The presence of tumor-associated macrophages (TAM) in breast cancer correlates strongly with poor outcome. The purpose of this study was to develop a clinically applicable, noninvasive diagnostic ...assay for selective targeting and visualization of TAMs in breast cancer, based on magnetic resonanceI and clinically applicable iron oxide nanoparticles.
F4/80-negative mammary carcinoma cells and F4/80-positive TAMs were incubated with iron oxide nanoparticles and were compared with respect to magnetic resonance signal changes and iron uptake. MMTV-PyMT transgenic mice harboring mammary carcinomas underwent nanoparticle-enhanced magnetic resonance imaging (MRI) up to 1 hour and 24 hours after injection. The tumor enhancement on MRIs was correlated with the presence and location of TAMs and nanoparticles by confocal microscopy.
In vitro studies revealed that iron oxide nanoparticles are preferentially phagocytosed by TAMs but not by malignant tumor cells. In vivo, all tumors showed an initial contrast agent perfusion on immediate postcontrast MRIs with gradual transendothelial leakage into the tumor interstitium. Twenty-four hours after injection, all tumors showed a persistent signal decline on MRIs. TAM depletion via αCSF1 monoclonal antibodies led to significant inhibition of tumor nanoparticle enhancement. Detection of iron using 3,3'-diaminobenzidine-enhanced Prussian Blue staining, combined with immunodetection of CD68, localized iron oxide nanoparticles to TAMs, showing that the signal effects on delayed MRIs were largely due to TAM-mediated uptake of contrast agent.
These data indicate that tumor enhancement with clinically applicable iron oxide nanoparticles may serve as a new biomarker for long-term prognosis, related treatment decisions, and the evaluation of new immune-targeted therapies.
Non-invasive detection of caspase-3/7 activity
has provided invaluable predictive information regarding tumor therapeutic efficacy and anti-tumor drug selection. Although a number of caspase-3/7 ...targeted fluorescence and positron emission tomography (PET) imaging probes have been developed, there is still a lack of gadolinium (Gd)-based magnetic resonance imaging (MRI) probes that enable high spatial resolution detection of caspase-3/7 activity
. Here we employ a self-assembly approach and develop a caspase-3/7 activatable Gd-based MRI probe for monitoring tumor apoptosis in mice. Upon reduction and caspase-3/7 activation, the caspase-sensitive nano-aggregation MR probe (C-SNAM:
) undergoes biocompatible intramolecular cyclization and subsequent self-assembly into Gd-nanoparticles (GdNPs). This results in enhanced
relaxivity-19.0 (post-activation) vs. 10.2 mM
s
(pre-activation) at 1 T in solution-and prolonged accumulation in chemotherapy-induced apoptotic cells and tumors that express active caspase-3/7. We demonstrate that C-SNAM reports caspase-3/7 activity by generating a significantly brighter
-weighted MR signal compared to non-treated tumors following intravenous administration of C-SNAM, providing great potential for high-resolution imaging of tumor apoptosis
.
15N-labeled rosette nanotubes were synthesized and investigated using high-field solid-state NMR spectroscopy, X-ray diffraction, atomic force microscopy, and electron microscopy. The results ...established the H-bond network involved in the self-assembly of the nanostructure as well as bound water molecules in the nanotube’s channel.
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