Supported by NIH R01 HL74082, the Hartwell Foundation, and the Focused Ultrasound Surgery Foundation.

1. Microbubble Production
<ol>
	<li>To prepare albumin microbubbles ( MBs), place a 1% solution of serum albumin in normal saline in a flask with a blanket of gas (octafluoropropane) above the aqueous phase. Briefly sonicate the solution (30 sec) with an ultrasound disintegrator equipped with an extended &frac12;" titanium probe. This formulation is similar to Optison (GE Heathcare), which is provided in a concentration range of 0.5- 1.2 x 109 MBs/ml. Determine mean MB diameter with a Multisizer Coulter ...

<ol>
	<li>Chappell, J., Song, J., Burke, C., Klibanov, A., Price, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeted+delivery+of+nanoparticles+bearing+fibroblast+growth+factor-2+by+ultrasonic+microbubble+destruction+for+therapeutic+arteriogenesis.">Targeted delivery of nanoparticles bearing fibroblast growth factor-2 by ultrasonic microbubble destruction for therapeutic arteriogenesis.</a> Small. 10, 1769-1777 (2008).</li><li>Chomas, J. E., Pollard, R., Wisner, E., Ferrara, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Subharmonic+Phase-Inversion+for+Tumor+Perfusion+Estimation.">Subharmonic Phase-Inversion for Tumor Perfusion Estimation.</a> IEEE. 2, 1713-1716 (2001).</li><li>Davda, J., Labhasetwar, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+of+nanoparticle+uptake+by+endothelial+cells.">Characterization of nanoparticle uptake by endothelial cells.</a> Int J Pharm. 233, 51-51 (2002).</li><li>Hynynen, K., McDannolod, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Noninvasive+MR+imaging-guided+focal+opening+of+the+blood-brain+barrier+in+rabbits.">Noninvasive MR imaging-guided focal opening of the blood-brain barrier in rabbits.</a> Radiology. 220, 640-646 (2001).</li><li>McDannold, N., Vykhodtseva, N., Hynynen, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microbubble+contrast+agent+with+focused+ultrasound+to+create+brain+lesions+at+low+power+levesl:+MR+imaging+and+histological+study+in+rabbits.">Microbubble contrast agent with focused ultrasound to create brain lesions at low power levesl: MR imaging and histological study in rabbits.</a> Radiology. 241, 95-106 .</li><li>McDannold, N., Vykhodtseva, N., Hynynen, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Targeted+disruption+of+the+blood-brain+barrier+with+focused+ultrasound:+Association+with+cavitation+activity.">Targeted disruption of the blood-brain barrier with focused ultrasound: Association with cavitation activity.</a> Phys Med Biol. 51, 793-807 .</li><li>McDannold, N., Clement, G. T., Black, P., Jolesz, F., Hynynen, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transcranial+magnetic+resonance+imaging-+guided+focused+ultrasound+surgery+of+brain+tumors:+initial+findings+in+3+patients.">Transcranial magnetic resonance imaging- guided focused ultrasound surgery of brain tumors: initial findings in 3 patients.</a> Neurosurgery. 66, 323-332 (2009).</li><li>Meairs, S., Alonso, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ultrasound+microbubblesand+the+blood+brain+barrier.">Ultrasound microbubblesand the blood brain barrier.</a> Progr Biophys Mol Biol. 93, 354-362 (2007).</li><li>. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Optison, products insert. , (2009).</li><li>Sadlowskie, A., Chromas, J., Pollard, R., Bloch, S., Griffey, S., Wisner, W., Ferrara, K. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mean+Flow+Rate+and+Intergrated+Perfusion+Estimates+Obtained+with+Contrast-Assisted+Ultrasound.">Mean Flow Rate and Intergrated Perfusion Estimates Obtained with Contrast-Assisted Ultrasound.</a> IEEE Ultrasonics Symposium. , 1977-1980 (2002).</li><li>Song, J., Chappell, J. C., Qi, M., VanGieson, E. J., Kaul, S., Price, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Influence+of+injection+site,+microvascular+pressure+and+ultrasound+variables+on+microbubble-mediated+delivery+of+microspheres+to+muscle.">Influence of injection site, microvascular pressure and ultrasound variables on microbubble-mediated delivery of microspheres to muscle.</a> J Am Coll. Cardiol. 39, 726-731 (2002).</li><li>Sheikov, N., McDannold, N., Vykhodtseva, N., Jolesz, F., Hynynen, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cellular+mechanisms+of+blood-brain+barrier+opeing+induced+by+ultrasound+in+the+presences+of+microbubbles.">Cellular mechanisms of blood-brain barrier opeing induced by ultrasound in the presences of microbubbles.</a> Ultrasound Med. Biol. 30, 979-989 (2004).</li><li>Tanter, J., Fink, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Focusing+and+steering+through+absorbing+and+aberrating+layers:+Application+to+ultrasonic+propagation+through+the+skull.">Focusing and steering through absorbing and aberrating layers: Application to ultrasonic propagation through the skull.</a> J Acoust Soc Am. 103, 2403-2410 (1998).</li><li>Yeh, C. K., Kruse, D. E., Lim, M. C., Redline, D. E., Ferrara, K. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+New+High+Frequency+Destruction/Reperfusion+System.">A New High Frequency Destruction/Reperfusion System.</a> IEEE. 1, 433-436 (2003).</li></ol>

No conflicts of interest declared.

Critical Steps
Cannulation of mouse tail vein:
Intravenous injection into the mouse tail vein can be a challenging procedure. However, a tail vein catheter can greatly improve the likelihood of a successful injection. To make the catheter, repeatedly bend back and forth a 25 gauge needle until it breaks from the hub. Insert the blunt end into the end of PE 20 tubing and seal the connection with silicon glue. To prepare the catheter for cannulation, attach a syringe ...

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		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>ApoptTag kit</td>
<td>&nbsp;</td>
<td>Intergen Co.</td>
<td>S7110</td>
<td>&nbsp;</td>
<tr>
<td>un-capped 85:15 poly(lactic-co-glycolic acid) (PLAGA)</td>
<td>&nbsp;</td>
<td>Lakeshore Biomaterials</td>
<td>Custom</td>
<td>&nbsp;</td>
<tr>
<td>Vivo Tag 680</td>
<td>&nbsp;</td>
<td>VisEn Medical</td>
<td>10120</td>
<td>Used to Tag BSA</td>
</tr>
<tr>
<td>Poly(vinyl alcohol)</td>
<td>&nbsp;</td>
<td>Sigma-Aldrich</td>
<td>363136</td>
<td>&nbsp;</td>
<tr>
<td>MicroTip Sonicator</td>
<td>&nbsp;</td>
<td>Misonix</td>
<td>S-4000</td>
<td>&nbsp;</td>
<tr>
<td>Sequoia</td>
<td>&nbsp;</td>
<td>Simons Medical</td>
<td>P.O.A</td>
<td>Equipped with CPS </td>
</tr>
<tr>
<td>FreeZone 2.5</td>
<td>&nbsp;</td>
<td>Labconco</td>
<td>7670020</td>
<td>Equipped with Nitrogen Trap</td>
</tr>
<tr>
<td>Methylene chloride (CH2Cl2)</td>
<td>&nbsp;</td>
<td>Fisher Scientific</td>
<td>D37-500</td>
<td>&nbsp;</td>
<tr>
<td>FMT 250</td>
<td>&nbsp;</td>
<td>VisEn Medical</td>
<td>P.O.A</td>
<td>&nbsp;</td>
<tr>
<td>F-12K Nutrient Mixture</td>
<td>&nbsp;</td>
<td>Gibco</td>
<td>21127-022</td>
<td>&nbsp;</td>
<tr>
<td>polyethyleneglycol-40 stearate </td>
<td>&nbsp;</td>
<td>Sigma Chemical </td>
<td>9004-99-3</td>
<td>&nbsp;</td>
<tr>
<td>distearoyl phosphatidylcholine</td>
<td>&nbsp;</td>
<td>Avanti Polar Lipids</td>
<td>770365</td>
<td>&nbsp;</td>
<tr>
<td>Multisizer Coulter Counter</td>
<td>&nbsp;</td>
<td>Beckman Coulter </td>
<td>P.O.A</td>
<td>&nbsp;</td>
<tr>
<td>Waveform Generator </td>
<td>&nbsp;</td>
<td>Tektronix, Inc.</td>
<td>AFG-310</td>
<td>&nbsp;</td>
<tr>
<td>water-based ultrasound gel </td>
<td>&nbsp;</td>
<td>Parker Laboratories</td>
<td>Aquasonic 100</td>
<td>&nbsp;</td>
<tr>
<td>Infusion pump </td>
<td>&nbsp;</td>
<td>Harvard Apparatus</td>
<td>Harvard Apparatus PHD 2000</td>
<td>&nbsp;</td>
<tr>
<td>1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) </td>
<td>&nbsp;</td>
<td>Pierce Biotechnology</td>
<td>25952-53-8</td>
<td>&nbsp;</td>
<tr>
<td>N-hydroxysulfosuccinimide (Sulfo-NHS)</td>
<td>&nbsp;</td>
<td>Pierce Biotechnology</td>
<td>106627-54-7</td>
<td>&nbsp;</td>
<tr>
<td>Succinic anhydride</td>
<td>&nbsp;</td>
<td>Sigma Aldrich</td>
<td>603902</td>
<td>&nbsp;</td>
<tr>
<td>Power Amplifier </td>
<td>&nbsp;</td>
<td>Electronic Navigation Industries</td>
<td>ENI 3100LA</td>
<td>&nbsp;</td>
<tr>
<td>Needle Thermocouple Probe </td>
<td>&nbsp;</td>
<td>Omega</td>
<td>HYP1-30-1/2-T-G-60-SMPW-M</td>
<td>&nbsp;</td>
<tr>
<td>BioGel (P100, medium)</td>
<td>&nbsp;</td>
<td>Bio-Rad</td>
<td>150-4170</td>
<td>&nbsp;</td>
<tr>
<td>.75&#x2019;&#x2019; diameter 1 MHz unfocused transducer</td>
<td>&nbsp;</td>
<td>Panametrics</td>
<td>A314S </td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

contrast ultrasound targeted treatment of gliomas in mice via drug-bearing nanoparticle delivery and microvascular ablation

We are developing minimally-invasive contrast agent microbubble based therapeutic approaches in which the permeabilization and/or ablation of the microvasculature are controlled by varying ultrasound pulsing parameters.  Specifically, we are testing whether such approaches may be used to treat malignant brain tumors through drug delivery and microvascular ablation.  Preliminary studies have been performed to determine whether targeted drug-bearing nanoparticle delivery can be facilitated by the ultrasound mediated destruction of "composite" delivery agents comprised of 100nm poly(lactide-co-glycolide) (PLAGA) nanoparticles that are adhered to albumin shelled microbubbles. We denote these agents as microbubble-nanoparticle composite agents (MNCAs). When targeted to subcutaneous C6 gliomas with ultrasound, we observed an immediate 4.6-fold increase in nanoparticle delivery in MNCA treated tumors over tumors treated with microbubbles co-administered with nanoparticles and a 8.5 fold increase over non-treated tumors. Furthermore, in many cancer applications, we believe it may be desirable to perform targeted drug delivery in conjunction with ablation of the tumor microcirculation, which will lead to tumor hypoxia and apoptosis. To this end, we have tested the efficacy of non-theramal cavitation-induced microvascular ablation, showing that this approach elicits tumor perfusion reduction, apoptosis, significant growth inhibition, and necrosis. Taken together, these results indicate that our ultrasound-targeted approach has the potential to increase therapeutic efficiency by creating tumor necrosis through microvascular ablation and/or simultaneously enhancing the drug payload in gliomas.

Watch this Scientific Journal Video about Contrast Ultrasound Targeted Treatment of Gliomas in Mice via Drug-Bearing Nanoparticle Delivery and Microvascular Ablation at JoVE.com

Contrast Ultrasound Targeted Treatment of Gliomas in Mice via Drug-Bearing Nanoparticle Delivery and Microvascular Ablation

Insonation of microbubbles is a promising strategy for tumor ablation at reduced time-averaged acoustic powers, as well as for the targeted delivery of therapeutics. The purpose of the present study is to develop low duty cycle ultrasound pulsing strategies and nanocarriers to maximize non-thermal microvascular ablation and payload delivery to subcutaneous C6 gliomas.

We are developing minimally-invasive contrast agent microbubble based therapeutic approaches in which the permeabilization and/or ablation of the ...

contrast-ultrasound-targeted-treatment-gliomas-mice-via-drug-bearing

Research

JoVE Journal

Bioengineering

15.8K vistas.  University of Virginia. Insonación de microburbujas es una estrategia prometedora para la ablación del tumor en un promedio de reducción de tiempo de potencias acústicas, así como para la distribución selectiva de la terapéutica. El propósito del presente estudio es desarrollar estrategias de ultrasonido de baja deber ciclo de pulsación y nanotransportadores para maximizar no de ablación térmica microvascular y la entrega a la carga por vía subcutánea C6 gliomas.