Nanotechnology for therapeutics and diagnostics

Project 1: Photothermal and Photochemical Response to Laser Therapy with Nanoparticles
Graduate Student Researchers: Jon Whitney and Saugata Sarkar (graduated)

Laser therapy based on photothermal (heat generation) and photochemical (reactive oxygen species production (ROS)) can provide minimally-invasive and potentially more effective alternatives to conventional resection, chemotherapy, and radiation, reducing complication rates and increasing patient recovery and quality of life. Although laser therapies have enormous promise, their effectiveness is limited due to nonspecific excitation or heating of target tissue, which can lead to healthy tissue injury. When laser therapies are utilized as cancer treatments their therapeutic efficacy can be compromised due to induction of molecular chaperones known as heat shock proteins (HSP) in regions of the tumor where non-lethal temperature elevation occurs. Thermal induction of HSP expression by tumor cells can increase tumor recurrence by enhancing tumor cell viability and diminish effectiveness of chemotherapy and radiation treatments generally employed in conjunction with thermal therapy.

Nanoparticles utilized in combination with laser therapy can improve treatment effectiveness by enhancing photothermal and photochemical mechanisms and increasing treatment selectivity when nanoparticles are targeted to tumor cells. When incorporated in cancer therapy, the selective photothermal/photochemical capability afforded by nanoparticles increases tumor cell death and diminishes HSP expression in the tumor through targeted heat generation and production of reactive oxygen species. We are measuring the response of breast, brain, prostate, and kidney cancer cells and tumors to a variety of nanoparticles such as carbon nanotubes (CNTs) and novel embodiments of carbon nanotubes including single walled carbon nanohorns and carbon nanotube peapods. Figure 1 shows the varying types of nanoparticles we are investigating. CNTs are graphene sheets of carbon atoms rolled into a tube and capped by fullerene hemispheres. The two major types of CNTs are single walled carbon nanotubes (SWNTs) which have one seamless tube and multi-walled nanotubes (MWNTs) which possess two or more concentric tubes. The diameter and length of SWNTs are 1.5-3.0 nm and 20-1000 nm respectively whereas the corresponding dimensions are 5.0-100 nm and 1-50 µm for MWNTs. Another unique embodiment of SWNTs are single-walled carbon nanohorns (SWNHs) which are composed of an aggregate of SWNTs with overall diameters of 50-100 nm.

We are also exploring the capability of carbon CNT/SWNH peapods for dual imaging and therapy (Figure 2). These structures are comprised of a CNT or SWNH (pod) encapsulated with endohedral metallofullerenes (EMFs) (a.k.a. buckyballs), The EMFs are spherical fullerenes containing gadolinium. This embodiment leverages CNTs/SWNHs as photoabsorbers and buckyballs containing gadolinium as imaging agents providing a nanoparticle capable of photothermal treatment and diagnostic imaging. Our lab is investigating the optimal nanoparticle properties and laser parameters to achieve selective and effective tumor treatment using both in vitro and in vivo experiments and computational modeling. We are measuring the impact inclusion of these nanoparticles has on optical and thermal tissue properties. We employ infrared imaging to determine the spatial temperature distribution in vitro and magnetic resonance thermometry to determine the temperature distribution in vivo in response to nanoparticle enhanced therapies. We have recently developed a novel cell viability algorithm to determine the temporal and spatial tumor response to therapies involving nanoparticles and laser therapy. Figure 3 shows a representative fluorescence image with live cells fluorescing green due to calcein staining (A), brightfield image showing dead cells stained wit trypan blue (B), and corresponding quantified viability distribution showing live (red) and dead (blue) cells (C). There is close correspondence between measured and quantified cell viability distributions. There is a high degree of tumor death in the center ring which is encompassed by the laser beam, however in the dish periphery where lower temperature elevations occurs, cell viability remains high. Radial viability measurements were computed and the radial viability percentages in 1000 pixel increments and are shown in Figure 3D.

CBET 0955072 (M.N. Rylander) 5/10/10-5/9/15 
National Science Foundation 
CAREER: Holey Scaffold Sensing System for Characterization of the Spatiotemporal Tumor Response to Nanoparticle-Mediated Photothermal and Photochemical Therapy

1R21CA156078-01 (C.G. Rylander) 8/10/2010-8/09/2012 
Fiberoptic Microneedle Device for Nanoparticle-Enhanced Photothermal Therapy of Aggressive, Infiltrative Bladder Cancer

CBET 1034026 (J. Freeman) 8/10/10-8/9/13 
National Science Foundation 
A Novel Treatment for Connective Tissue in Ehlers-Danlos Patients and Strained and Sprained Ligaments Investigating Carbon Nanostructure Enhanced Prolotherapy

ZRG1 BCMB-S (50) (S. Torti) 4/01/2008 – 3/31/2013 NIH/NCI 
Nanotubes in Tumor Imaging and Therapy

CBET 0933571 (C.G. Rylander) 8/16/2009 – 8/15/2012 
National Science Foundation 
Fiberoptic Microneedle Device for Combined Light and Nanomedicine Delivery: Mimicking Nature’s Design of a Mosquito

CBET 0731108 (M.N. Rylander) 8/1/2007 – 7/31/2011
National Science Foundation 
Characterization and Model Development for the Cellular Response to Nanotube-Mediated Laser Therapy

1R21CA135230-01 (M.N. Rylander) 7/1/2008 – 6/31/2011 NIH/NCI 
Trimetallic Nitride Templated Endohedral Metallofullerenes as Dual Imaging and Multimodal Therapeutic Agents for Laser Cancer Therapy

1 R41 EB008907-01 (Naha) NIH/ ZRG1 SSMI-Q (10)7/1/2008 – 6/31/2010 
A Novel Nanomaterials Approach for Cancer Imaging and Therapeutic Treatment

Related Papers
Photothermal or Hyperthermia Therapy alone or in combination with Nanoparticles (most recent to earliest)

S. Sarkar, Abhijit A. Gurjarpadhye, C. Rylander, M. N. Rylander, 2011, “Optical Properties of Breast Tumor Phantoms Containing Carbon Nanotubes and Nanohorns,” Journal of Biomedical Optics, Accepted and in press
S. Sarkar, K. Zimmermann, W. Leng, P. Vikesland, J. Zhang, H. Dorn, T. Diller, C. Rylander, and M. N. Rylander, 2011, “Measurement of the Thermal Conductivity of Carbon Nanotube-Tissue Phantom Composites with the Hot Wire Probe Method,” Ann Biomed Eng. [Epub ahead of print].
J. Whitney, S. Sarkar, J. Zhang, T. Do, T. Young, M. K. Manson, T.A. Campbell, A. A. Puretzky, C. M. Rouleau, K. L. More, D. B. Geohegan, C. G. Rylander, H. C. Dorn, M. N. Rylander, 2011, “Carbon Nanohorns as Photothermal Agents for Cancer Therapy,” Lasers in Surgery and Medicine, 43(1):43-51.
J. W. Fisher, C. F. Buchanan, C. Szot, S. Sarkar, C. Rylander, and M. N. Rylander, 2010, “Photothermal Response of Human and Murine Cancer Cells to Multiwalled Carbon Nanotubes and Laser Irradiation,” Cancer Research, 70 (23): 1-10.
M. N. Rylander, Y. Feng, K. Zimmermann, and K. R. Diller, 2010, “Measurement and Mathematical Modeling of Thermally Induced Injury and Heat Shock Protein Expression Kinetics in Normal and Cancerous Prostate Cells,“ International Journal of Hyperthermia Special Issue on Prostate Cancer Therapy, 26(8): 748-764.
J. Zhang, J. Ge, M. Shultz, E. Chung, G. Singh, C. Shu, P. Fatouros, S. Henderson, F. Corwin, D. Geohegan, A. Puretzky, C. Rouleau, K. More, C. Rylander, M. N. Rylander, H. Gibson, H. Dorn, 2010, “In Vitro and In Vivo Studies of Single-Walled Carbon Nanohorns with Encapsulated Metallofullerenes and Exohedrally Functionalized Quantum Dots,” Nano Letters, 10(8): 2843-2848.
S. Sarkar, C. G. Rylander, and M. N. Rylander, 2010, Photothermal Response of Tissue Phantoms Containing Multi-walled Carbon Nanotubes: Modified Optical and thermal properties,” Journal of Biomechanical Engineering, 132: 044505-1-044505-5.
M. Kosoglu, R. Hood, Y. Chen, Y. Xu, M. N. Rylander, and C. Rylander, 2010, “Fiberoptic Microneedles for Transdermal Light Delivery: Ex Vivo Porcine Skin Experiments”, Journal of Biomechanical Engineering, 132(9): 091014-1-091014-7.
C. Shu, J. Ge, J. Zhang, J. Hyun Sim, B. G. Burke, K. A. Williams, M. N. Rylander, T. Campbell, D. Geohegan, A. R. Esker, H. W. Gibson, and H. C. Dorn, 2010,"A Facile High-Speed Vibration Milling Method to Water-Disperse Single-Walled Carbon Nanohorns,” Chemistry of Materials, 22, 347–351.
A. Burke, X. Ding, Ravi Singh, R. A. Kraft, M. N. Rylander, C. Szot, C. Buchanan, J. Whitney, J. Fisher, N. Levi-Polyachenko, H. C. Hatcher, R. D’Agostino Jr., N. Kock, P. M. Ajayan, D. L. Carroll, F. M. Torti, S. V. Torti, 2009, “Long-term survival following a single treatment of kidney tumors with multiwalled carbon nanotubes and near-infrared radiation,” PNAS, 4, pp. 12897-12902.
Computational Models for Cancer Laser Therapy alone or with Nanoparticles (most recent to earliest)

Y. Feng, D. Fuentes, A. Hawkins, J. Bass and M. N. Rylander, 2009, "Optimization and Real-Time Control for Laser Treatment of Heterogeneous Soft Tissues," Computer Methods in Applied Mechanics and Engineering, 198, Issues 21-26, 1742-1750.
Y. Feng, D. Fuentes, A. Hawkins, J. Bass, M. N. Rylander, A. Elliott, A. Shetty, R. J. Stafford, and J. T. Oden, 2009, "Nanoshell-Mediated Laser Surgery Simulation for Prostate Cancer Treatment," Special issue on Computational Biomedical Engineering for Journal of Engineering with Computers, 25, 3-13.
Y. Feng, J. T. Oden, and M. N. Rylander, 2008, " A Two-State Cell Damage Model Under Hyperthermic Conditions: Theory and In Vitro Experiments," Journal of Biomechanical Engineering, 130: 0410161-10.
J. Fisher and M. N. Rylander, 2008, “Effective Cancer Laser Therapy Design Through the Integration of Nanotechnology and Computational Treatment Planning Models,” Proc. SPIE, 6869, 68690D1-11.
J. T. Oden, K. R. Diller, C. Bajaj, J. C. Browne, J. Hazle, I. Babuška, J. Bass, L. Biduat, L. Demkowicz, A. Elliott, Y. Feng, D. Fuentes, S. Prudhomme, M. N. Rylander, R. J. Stafford, Y. Zhang, 2007, “Dynamic Data-driven Finite Element Models for Laser Treatment of Cancer,” Numerical Methods For Partial Differential Equations, 23, 904-922.
M. N. Rylander, Y. Feng, J. Bass, and K. R. Diller, 2007, “HSP Expression and Damage Optimization Algorithm for Prostate Cancer Therapy Design,” Lasers in Surgery and Medicine, 39, 731-746.
J. T. Oden, K. R. Diller, C. Bajaj, J. C. Browne, J. Hazle, I. Babuska, J. Bass, L. Demkowicz, Y. Feng, D. Fuentes, S. Prudhomme, M. N. Rylander, R. J. Stafford, and Y. Zhang, 2006, “Development of a Computational Paradigm for Laser Treatment of Cancer,” Lecture Notes in Computer Science, 3993: 530-537.
Y. Feng, M. N. Rylander, J. Bass, J.T. Oden, and K. R. Diller, 2005, “Optimal Design of Laser Surgery for Cancer Treatment Through Nanoparticle-Mediated Hyperthermia Therapy,” NSTI-Nanotech. 1, 39-42.
M. N. Rylander, Y. Feng, Y. Zhang, J. Bass, R. J. Stafford, J. Hazle, and K. R. Diller, 2005, “Optimizing HSP Expression in Prostate Cancer Laser Therapy Through Predictive Computational Models,’ Journal of Biomedical Optics, 11, 0411131-16.
Project 2: Measurement of Nanoparticle Transport at the Cellular and Tissue Level
Graduate Student Researcher: Kristen Zimmermann

Carbon nanomaterials have been used for a variety of biomedical applications, from biosensing to drug delivery. Over the past decade, groups in Japan and China have begun investigating a new type of carbon nanomaterial, called single-walled carbon nanohorns (SWNHs). This material is unique in structure, yet exhibits the excellent mechanical, chemical, and thermal properties characteristic of carbon nanotubes (CNTs). The unique structure of SWNHs provides advantages over CNTs, such as larger surface areas both externally and internally to allow for greater surface modification or drug loading. Additionally, SWNHs can be synthesized without the use of a metal catalyst, reducing potential toxicity. Due to such advantages, SWNHs have been studied previously as drug delivery systems for chemotherapeutic agents and have also been investigated for nanoparticle-enhanced laser therapies for cancer malignancies; however, studies have been limited by lack imaging techniques. Therefore, our group is developing a highly fluorescent SWNH that can be used to study the transport of these particles in vitro and in vivo at both the cellular and tissue levels. The fluorescently tagged SWNHs will serve to image these particles optically, study their transport mechanisms and distribution both in vitro and in vivo, and to detect tumor margins in vivo once targeted with a receptor-specific ligand.

Kristen is funded by a National Science Foundation Graduate Research Fellowship. 
CBET 0955072 (M.N. Rylander)5/10/10-5/9/15 
National Science Foundation 
CAREER: Holey Scaffold Sensing System for Characterization of the Spatiotemporal Tumor Response to Nanoparticle-Mediated Photothermal and Photochemical Therapy

Project 3: Nanoparticles and Electroporation

Graduate Student Researchers: Chris Arena and Jose Umanzor-Alvarez

This research is focused on utilizing carbonaceous nanomaterials to enhance the ablation of tumors through irreversible electroporation (IRE). IRE employs high-voltage, ultra-short electric pulses to destabilize cell membranes. Nanoparticles afford us the ability to localize the pulsed electric field based therapy to targeted cancer cells.

Additionally, we are evaluating the heat shock response to IRE. While IRE is inherently non-thermal, it is hypothesized that the stress caused by the permeabilization of membranes could lead to the induction of heat shock proteins (HSP), which are known to play a role in tumor recurrence.

Relevant Papers:
C. B Arena, J. L Caldwell, M. N. Rylander, R. V. Davalos, 2009, “Theoretical study for the treatment of pancreatic cancer using electric pulses.” Conf Proc IEEE Eng Med Biol Soc., 2009:5997-6000.

C. B Arena, J. L Caldwell, M. N. Rylander, R. V. Davalos, 2010, “Theoretical Considerations of Tissue Electroporation with High Frequency Bipolar Pulses,” IEEE Trans Biomed Eng., Published online. Funding Information: CBET 0933335 (R. Davalos) 8/1/2009 – 7/31/2012 National Science Foundation Combinatorial Brain Cancer Therapy through Irreversible Electroporation and Carbon Nanotubes