Randy A. Patton

Daunorubicin‐Loaded DNA Origami Nanostructures Circumvent Drug‐Resistance Mechanisms in a Leukemia Model

Many cancers show primary or acquired drug resistance due to the overexpression of efflux pumps. A novel mechanism to circumvent this is to integrate drugs, such as anthracycline antibiotics, with nanoparticle delivery vehicles that can bypass intrinsic tumor drug-resistance mechanisms. DNA nanoparticles serve as an efficient binding platform for intercalating drugs (e.g., anthracyclines doxorubicin and daunorubicin, which are widely used to treat acute leukemias) and enable precise structure design and chemical modifications, for example, for incorporating targeting capabilities. Here, DNA nanostructures are utilized to circumvent daunorubicin drug resistance at clinically relevant doses in a leukemia cell line model. The fabrication of a rod-like DNA origami drug carrier is reported that can be controllably loaded with daunorubicin. It is further directly verified that nanostructure-mediated daunorubicin delivery leads to increased drug entry and retention in cells relative to free daunorubicin at equal concentrations, which yields significantly enhanced drug efficacy. Our results indicate that DNA origami nanostructures can circumvent efflux-pump-mediated drug resistance in leukemia cells at clinically relevant drug concentrations and provide a robust DNA nanostructure design that could be implemented in a wide range of cellular applications due to its remarkably fast self-assembly (≈5 min) and excellent stability in cell culture conditions.

Engineering Cell Surface Function with DNA Origami

A specific and reversible method is reported to engineer cell-membrane function by embedding DNA-origami nanodevices onto the cell surface. Robust membrane functionalization across epithelial, mesenchymal, and nonadherent immune cells is achieved with DNA nanoplatforms that enable functions including the construction of higher-order DNA assemblies at the cell surface and programed cell-cell adhesion between homotypic and heterotypic cells via sequence-specific DNA hybridization. It is anticipated that integration of DNA-origami nanodevices can transform the cell membrane into an engineered material that can mimic, manipulate, and measure biophysical and biochemical function within the plasma membrane of living cells.

Towards High-Repetition Rate Rayleigh and Raman Scattering Imaging in Turbulent Jets and Flames

In this paper we will describe recent advances made in our laboratory in the development of high-repetition-rate Rayleigh and Raman scattering imaging capabilities. High-repetition-rate 1D Raman and 2D Rayleigh scattering imaging capabilities are being developed to image the time-varying mixture fraction and temperature fields in turbulent non-reacting and reacting flows. Initial results using a custom pulse-burst laser system at Ohio State University have demonstrated the ability to capture ten sequential 2D Rayleigh scattering images at a repetition rate of 10 kHz in both turbulent non-reacting jets and non-premixed jet-flames with pulse energies approaching 200 mJ at 532 nm. This paper will also describe the development and pending application of a new higher-energy, long-duration, next-generation burst-mode laser system and the use of higher resolution cameras for high-speed Raman/Rayleigh imaging.

Cation-Activated Avidity for Rapid Reconfiguration of DNA Nanodevices

The ability to design and control DNA nanodevices with programmed conformational changes has established a foundation for molecular-scale robotics with applications in nanomanufacturing, drug delivery, and controlling enzymatic reactions. The most commonly used approach for actuating these devices, DNA binding and strand displacement, allows devices to respond to molecules in solution, but this approach is limited to response times of minutes or greater. Recent advances have enabled electrical and magnetic control of DNA structures with sub-second response times, but these methods utilize external components with additional fabrication requirements. Here, we present a simple and broadly applicable actuation method based on the avidity of many weak base-pairing interactions that respond to changes in local ionic conditions to drive large-scale conformational transitions in devices on sub-second time scales. To demonstrate such ion-mediated actuation, we modified a DNA origami hinge with short, weakly complementary single-stranded DNA overhangs, whose hybridization is sensitive to cation concentrations in solution. We triggered conformational changes with several different types of ions including mono-, di-, and trivalent ions and also illustrated the ability to engineer the actuation response with design parameters such as number and length of DNA overhangs and hinge torsional stiffness. We developed a statistical mechanical model that agrees with experimental data, enabling effective interpretation and future design of ion-induced actuation. Single-molecule Förster resonance energy-transfer measurements revealed that closing and opening transitions occur on the millisecond time scale, and these transitions can be repeated with time resolution on the scale of one second. Our results advance capabilities for rapid control of DNA nanodevices, expand the range of triggering mechanisms, and demonstrate DNA nanomachines with tunable analog responses to the local environment.

Developement of Gas-Phase Mixing Measurements in Turbulent Spray Flows Using Filtered Rayleigh Scattering

In this paper we will describe recent advances made in our laboratory towards the development, validation, and application of a filtered Rayleigh Scattering technique for measurements of gas-phase concentration fields in the presence of liquid-phase droplets. Preliminary studies of the maximum level of droplet scattering attenuation attainable utilizing an injection-seeded, Q-Switched Nd:YAG laser in conjunction with a molecular iodine filter will be presented. These results demonstrate the correlation of the spectral purity of the laser with the effectiveness of the molecular filter in suppressing particle (Mie) scattering. Improvements in spectral purity and the maximum Mie scattering rejection as a result of implementing an external etalon into the optical path of the laser output will be demonstrated. Initial validation studies of the FRS technique also are presented in which gas-phase information is extracted from a droplet-laden turbulent jet. Comparisons of suppression capabilities with and without the external etalon are presented, allowing for an assessment gains offered in implementing the etalon. Finally, we present initial results from an evaporating spray jet of acetone into air.

NO PLIF Imaging in the CUBRC 48" Shock Tunnel

Nitric Oxide Planar Laser-Induced Fluorescence (NO PLIF) imaging is demonstrated at a 10 kHz repetition rate in the Calspan-University at Buffalo Research Center’s (CUBRC) 48-inch Mach 9 hypervelocity shock tunnel using a pulse burst laser–based high frame rate imaging system. Sequences of up to ten images are obtained internal to a supersonic combustor model, located within the shock tunnel, during a single ~10millisecond duration run of the ground test facility. This represents over an order of magnitude improvement in data rate from previous PLIF-based diagnostic approaches. Comparison with a preliminary CFD simulation shows good overall qualitative agreement between the prediction of the mean NO density field and the observed PLIF image intensity, averaged over forty individual images obtained during several facility runs.

Towards Simultaneous High-Speed Mixture Fraction and Velocity Imaging in Turbulent Jets

In this paper we will describe recent advances made in our laboratory in the development of both high-speed mixture fraction ([) imaging and simultaneous mixture fraction and velocity imaging in turbulent nonreacting jets. High-repetition rate mixture fraction imaging has been developed in turbulent jets using planar Rayleigh scattering imaging, with the goal of extending this measurement capability to include simultaneous high-speed planar velocity measurements using particle imaging velocimetry (PIV). In this respect, conventional Rayleigh scattering imaging will be replaced with Filtered Rayleigh scattering (FRS), so that [ [ [ [ can be measured in the presence of the PIV seed particles without interference. In this paper, we will report a successful demonstration of high-speed (10 kHz) mixture fraction imaging using Rayleigh scattering and progress towards simultaneous mixture fraction and velocity imaging using FRS/Mie scattering at low (10 Hz) repetition rates. Future work entails merging the two independent work paths to enable a simultaneous high-speed mixture fraction/velocity measurement capability.