CELL-MEDIATED MAGNETIC HYPERTHERMIA OF PRECLINICAL TUMORS
Sivasai Balivada†, Rajashekhar Rachakatla†, Matthew T. Basel†, Hongwang Wang‡, Gwi Moon Seo†, Tej B. Shrestha†, Marla Pyle†,
Viktor Chikan‡, Stefan H. Bossmann‡, Deryl L. Troyer†
†-Department of Anatomy & Physiology ‡- Department of Chemistry, Kansas State University
ABSTRACT
Using iron nanoparticles (MNPs) to absorb alternating magnetic field (AMF) energy
as a method of generating localized hyperthermia has shown potential as a cancer
treatment. Many attempts have been made to increase the localization of MNPs, for
example by attaching antibodies recognizing tumor-specific epitopes or peptides
binding receptors on tumor cells or neovasculature. Several research groups have
shown reliable results using tumor homing cells as delivery cells for different
therapeutics. Here we hypothesized that tumor homing cells can carry MNPs
specifically to the tumor site, and tumor burden will decrease after AMF exposure.
To test this hypothesis, first we loaded Fe/Fe3O4 bi-magnetic NPs into neural
progenitor cells (NPCs).We observed that NPCs loaded with MNPs travel to
subcutaneous melanoma tumors. After AMF exposure, the targeted delivery of
MNPs by the NPCs resulted in a significant decrease in tumor size. Second,
monocytes/macrophages (Mo/Ma) are known to infiltrate tumor sites, and also have
phagocytic activity which can increase their uptake of MNPs. To test Mo/Mamediated magnetic hyperthermia we transplanted Mo/Ma loaded with MNPs into a
mouse model of pancreatic peritoneal carcinomatosis. We observed that MNPloaded Mo/Ma infiltrated pancreatic tumors and, after AMF treatment, significantly
prolonged the lives of mice bearing disseminated intraperitoneal pancreatic tumors.
Based on these observations we concluded that development of localized
hyperthermia treatment using tumor tropic cells shows promise as a cancer therapy.
MATERIALS & METHODS
• Quantitative iron estimation in MNP: Ferrozine based colorimetric method
• Qualitative iron identification in cells: Prussian blue staining, TEM
• Cell toxicity assays: Trypan blue counting, MTT assay
• Flow analysis of cells: Guava Flow Express Plus flow cytometer
• Cell lines: B16F10 melanoma cells, PanO2 pancreatic adenocarcinoma cells,
C17.2 neural progenitor cells (NPCs), RAW264.7 monocytes/
macrophages (Mo/Ma)
Inductive heater
• Mouse tumor models: subcutaneous melanoma model;
disseminated peritoneal pancreatic cancer model.
•10 KW inductive heater to generate AMF
• Gaussia luciferase-mediated bioluminescence was measured
using an IVIS Lumina II
• Histology: Prussian blue staining and PKH26 fluorescence detection.
HYPOTHESIS
Tumor homing cells can carry MNPs specifically to the tumor site, and tumor
burden will decrease after AMF exposure
MNPs testing on NPCs in vitro:
www.PosterPresentations.com
Histological analysis of tissues from mice treated with MNPs and AMF exposure:
Increase in survival of mice bearing peritoneal pancreatic tumors by AMF
after delivery of MNPs by Mo/Ma:
•Kaplan-Meier test showed that
survival curves were significantly
different for the experimental group
compared with other
groups.(p<0.005)
•MNP/AMF treatment mice
survived substantially longer, with
mice lasting until 33.5 days
•Average increase in survival of
treatment group is 7 days
(A−D) Prussian blue stained tissue sections of melanoma tumor-bearing mice which received
NPCs-MNPs followed by AMF treatment: lung (A), liver (B), and tumor (C). Note the absence of
blue stained NPCs in the tumor sections. (D) NPCs loaded with MNP (blue color cells) in tumor
section of a mouse which received NPCs-MNPs but no AMF treatment. (E,F): green apoptotic
cells in a tumor-bearing mouse with NPCs-MNPs+AMF (E) compared to few apoptotic cells in a
tumor-bearing mouse with saline only treatment (F).
MNPs testing on RAW 264.7 Mo/Ma in vitro:
A. TEM of Fe/Fe3O4/aminosilaxane/stealth nanoparticles; B. Bright-field image of NPCs loaded
with MNPs showing positive Prussian blue staining (magnification 20×); C. TEM image of NPC
loaded with MNPs (magnification 30 000×); D. In vitro cell viability of NPCs cultured in medium
containing MNPs; E. Loading efficiency of iron MNPs; F. AMF-induced temperature changes in
vitro.* p < 0.05, ** p < 0.1 when compared with the control
SUMMARY
•At the concentrations used here, MNPs cause minimal toxicity on NPCs
and Mo/Ma; loading of MNPs into cells is concentration dependent.
•MNP-loaded NPCs and Mo/Ma can increase the temperature of tumors
after AMF exposure.
•NPCs can migrate to melanoma tumors after intravenous transplantation.
•Mo/Ma can migrate to pancreatic tumors after intraperitoneal
transplantation.
•AMF exposure resulted in significant regression of melanoma tumor
size for a short period of time.
•AMF exposure resulted in a significant increase in survival of mice
bearing pancreatic tumors after delivery of MNPs by Mo/Ma (average
increase: 7 days).
REFERENCES
NPCs migration toward mouse melanoma and attenuation of melanoma
growth by AMF after delivery of MNPs by NPCs:
A. Homing of
intravenously
injected NPCs
expressing Gaussia
luciferase to
metastatic
melanoma. B.
Tumor volumes in
mice injected with
B16-F10 melanoma
cells (*p < 0.05)
when compared with
saline control.
RESEARCH POSTER PRESENTATION DESIGN © 2011
RESULTS
RESULTS
BACKGROUND
•problem in cancer therapy: metastatic tumors that are smaller than clinically
detectable level; inoperable tumors
•Therapies that can specifically target to tumors can solve the problem.
•Using iron nanoparticles (MNPs) to absorb alternating magnetic field (AMF)
energy as a method of generating localized hyperthermia has been shown to be a
potential cancer treatment.
•MNPs were injected directly into the tumor for magnetic hyperthermia treatment in
most of the studies reported in the literature.
•Several research groups tried to target MNPs to tumors by attaching antibodies
recognizing tumor-specific epitopes or peptides binding receptors on tumor cells or
neovasculature.
•Immune cells and stem cells can migrate towards tumors; using these cells as
delivery vehicles for MNPs may help to achieve tumor-targeted magnetic
hyperthermia.
RESULTS
A. Toxicity of nanoparticles B. Loading of nanoparticles C. Percent of cells loaded D. Bright-field
image of cells loaded with MNPs showing positive for Prussian blue staining
Homing studies of PKH26-loaded RAW 264.7 Mo/Ma on disseminated
peritoneal pancreatic cancer:
Mo/Ma only infiltrate Pan02
tumors. PKH26-labeled
mo/ma were injected
intraperitoneally into mice
bearing intraperitoneal Pan02
tumors. Mice were
euthanized 3 days after
Mo/Ma injection, and organs
were harvested and imaged.
Scale bars are 100 μm
•Attenuation of Mouse Melanoma by A/C Magnetic Field after Delivery of Bi-Magnetic
Nanoparticles by Neural Progenitor Cells
Raja Shekar Rachakatla*, Sivasai Balivada*, Gwi-Moon Seo, Carl B. Myers, Hongwang Wang, Thilani N.
Samarakoon, Raj Dani, Marla Pyle, Franklin O. Kroh, Brandon Walker, Xiaoxuan Leaym, Olga B. Koper,
Viktor Chikan, Stefan H. Bossmann, Masaaki Tamura, and Deryl L. Troyer ACS Nano 2010 4 (12), 70937104, * co-first authors
•Cell-delivered magnetic nanoparticles caused hyperthermia-mediated increased survival in a
murine pancreatic cancer model
Matthew T. Basel, Sivasai Balivada, Hongwang Wang, Tej B. Shrestha, Gwi Moon Seo, Marla Pyle,
Gayani Abayaweera, Raj Dani, Olga B. Koper, Masaaki Tamura, Viktor Chikan, Stefan H. Bossmann, and
Deryl L. Troyer Int J Nanomedicine. 2012; 7: 297–306.I
ACKNOWLEDGEMENTS
This work was supported by NIH 1R21CA135599, NIH-COBRE grant P20 RR0117686, NSF IIP 0930673,
HHSN261200800059C from the Small Business Innovation Research Development Center (SBIR) of the National
Cancer Institute, the Terry C. Johnson Center for Basic Cancer Research at Kansas State University, Kansas State
University Targeted Excellence, Kansas State Legislative Appropriation, and Kansas Agricultural Experiment Station
Kansas Bioscience Authority.