MULTIMODAL PARTICLES FOR BIOLOGICAL DETECTION AND THERAPY

Kanaka Hettiarachchi,1 Paul A. Dayton,2 and Abraham P. Lee1 1Department of Biomedical Engineering, University of California, Irvine, USA 2Joint Department of Biomedical Engineering, University of North Carolina –

North Carolina State University, USA

ABSTRACT A novel micron-sized hybrid particle complex, consisting of an Acoustically

Active Liposphere (AAL) linked to superparamagnetic iron oxide (SPIO) particles, that can be useful for multimodal imaging with ultrasound (US) and magnetic resonance imaging (MRI), as well as a drug delivery system (DDS) is reported. Microfluidics technology is utilized for the precise control of particle composition, size, and polydispersity. We show the use of low frequency radio waves for heating the SPIO component, melting the lipid shell and releasing the Doxorubicin (DOX) drug contents within minutes of application. KEYWORDS: Drug Delivery, Ultrasound, MRI, Microbubbles, Microfluidics

INTRODUCTION

Contrast agents (CAs), as stabilized gas microbubbles for targeted ultrasound (US) imaging or magnetic nanoparticles for magnetic resonance imaging (MRI), are valuable tools for diagnosis of the presence and extent of disease [1]. Lipid-shelled microbubbles as acoustically-active lipospheres (AAL) [2] or metal nanoparticles [3] show promise as novel drug delivery systems (DDS). We envisioned combining the utility of the AAL DDS with superparamagnetic iron oxide (SPIO)-based CAs to create a hybrid particle complex (Fig. 1a) that can be useful for multimodal imaging with US and MRI, as well as therapy in living systems.

THEORY

Microfluidic flow-focusing techniques have already shown to be powerful technologies for the generation of highly controlled microbubbles [4]. A narrow size distribution is important since parameters such as resonant frequency (important for imaging), drug content, and biodistribution are affected by the vehicle diameter.

Figure 1. (a) Schematic of an acoustically active microbubble liposphere

attached to superparamagnetic iron oxide particles and (b) microfluidic flow-focusing device design with main functional area and orifice.

978-0-9798064-1-4/µTAS2008/$20©2008CBMS 1765

Twelfth International Conference on Miniaturized Systems for Chemistry and Life SciencesOctober 12 - 16, 2008, San Diego, California, USA

Magnetic separation has been applied to many aspects of biomedical and biological research [5]. The physical principles of magnetophoresis are derived from the magnetic force exerted on a superparamagnetic particle at a distance by a magnetic field gradient. For a one-dimensional magnetic field gradient, the total magnetic force Ftspm balances the viscous (stokes) drag FD for an SPIO-AAL complex moving in a fluid due to a magnetic field gradient, as represented by the following eqn. (1):

tspm

spmspmspmaD FB

VNRF −=∇−=−= 2

0216

μχ

ηνπ (1)

where Ra is the radius of the AAL, η is the viscosity of the aqueous medium, v is the velocity of the AAL, Nspm is the number of SPIO particles conjugated to an AAL, Vspm is the volume of the SPIO particle, χspm is the net magnetic susceptibility of a SPIO particle in aqueous solution, B is the magnetic field, and μ0 is the permeability of free space. The procedure for catabolism of tumors involves dispersing magnetic particles throughout the target tissue, and then applying an AC magnetic field of sufficient strength and frequency to cause the particles to heat [5]. This thermal triggering can also be used to release a drug or payload from a DDS.

EXPERIMENTAL

The AALs were produced by a PDMS flow-focusing device (Fig. 1b), forcing a central stream of nitrogen gas and three (stabilizer, lipid, and oil) double-sided liquid sheath flows through a narrow 20 micron orifice. Attachment to magnetic iron oxide particles was achieved at the outlet reservoir. The outermost sheath flow stream contains a 10% Poloxamer 188 solution that enhances the stability of the AAL shell. The middle sheath flow stream contains a 10% aqueous glycerol/propylene glycol mixture with the stabilizing lipids DSPC and DSPE-PEG2000-Biotin at a 9:1 molar ratio and concentration of 0.5 mg/mL DSPC. The inner sheath flow stream consists of triacetin oil, premixed with Oil Blue N dye for easier visualization. Doxorubicin (DOX) is used as the model antitumor drug due to the intrinsic fluorescence properties of the molecule, and is mixed with triacetin at a concentration of 1mmol/L. Irregularly shaped superparamagnetic iron oxide particles covered with a Streptavidin coating for attachment to biotin groups on the AAL shell are used to create complexes. Nitrogen gas is supplied from a pressurized tank via flexible tubing and delivered into the chamber using a micro flow meter. The continuous liquid phase mixtures are pumped at constant flow rates using digitally controlled syringe pumps. An inverted microscope and high-speed camera are used to capture images and record movies. A function generator and power amplifier are used to drive a standard AC electromagnet, which energizes the iron oxide particles. Sample temperature is measured with a laser infrared interferometer. RESULTS AND DISCUSSION

At specific flow rates (QP188=10µL/min, Qlipid=20µL/min, Qoil=0.5µL/min, Pgas=1.5psi) we can produce stable AALs with attached iron oxide particles (Fig. 2a).

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Twelfth International Conference on Miniaturized Systems for Chemistry and Life SciencesOctober 12 - 16, 2008, San Diego, California, USA

Figure 2. (a) Brightfield image of 15 micron AAL with attached SPIO particles.

(b) Melted DSPC hybrid vehicles at 55°C. (c) Local DOX fluorescence. The velocity v of the hybrid particle complex in the aqueous medium can be

calculated from eqn. (1). The experimentally calculated complex velocity is 0.12mm/s, but can be orders of magnitude different according to theory, depending on the number of SPIO attached to AALs, irregularity in SPIO shape, solution viscosity changes due to oil, and strength of the external magnetic field. Application of alternating magnetic fields at a maximum strength of 775 Gauss increases sample temperature to 55°C in 10 minutes by way of heating the iron oxide particles, melting the lipids (Fig. 2b) and releasing DOX (Fig. 2c). CONCLUSIONS

We have developed a method for synthesizing iron oxide linked microbubble lipospheres and have demonstrated their value as a DDS. The next step is to conduct multimodal imaging studies with US and MRI, and determine in-vivo stability. With these multimodal hybrid particle complexes, diagnostics and therapeutics will no longer have to be sequential elements in patient care.

ACKNOWLEDGEMENTS

We thank the National Institutes of Health (R03 EB006846) for financial support. REFERENCES [1] A. M. Morawski, G. A. Lanza, and S. A. Wickline, “Targeted contrast agents

for magnetic resonance imaging and ultrasound,” Current Opinion in Biotechnology, Vol. 16, pp. 89-92 (2005).

[2] E. C. Unger, T. P. McCreery, R. H. Sweitzer, V. E. Caldwell and Y. Wu, “Acoustically active lipospheres containing paclitaxel: a new therapeutic ultrasound contrast agent,” Invest Radiol, Vol. 33, pp. 886-892 (1998).

[3] A. K. Gupta and S. Wells, “Surface-modified superparamagnetic nanoparticles for drug delivery,” Ieee T Nanobiosci, Vol. 3, pp. 66-73 (2004).

[4] K. Hettiarachchi, E. Talu, M. L. Longo, P. A. Dayton, and A. P. Lee, “On-chip generation of microbubbles as a practical technology for manufacturing contrast agents for ultrasonic imaging,” Lab Chip, Vol. 7, pp. 463-468 (2007).

[5] Q. A. Pankhurst, J. Connolly, S. K. Jones and J. Dobson, “Applications of magnetic nanoparticles in biomedicine,” J Phys D Appl Phys, Vol. 36, pp. R167-R181 (2003).

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