RESEARCH POSTER PRESENTATION DESIGN © 2012
www.PosterPresentations.com
A large proportion of chemotherapeutic drugs such as paclitaxel (PTX) are
water insoluble and thus require the use of specialized delivery vehicles (e.g.
micelle, liposome, etc.) for parenteral administration. These nano-sized
delivery vehicles are often complex and may be difficult to manufacture,
immunogenic, and/or cause toxic side effects. Recently, exosomes
(extracellular vesicles ~100nm in size, released by almost all cell types and
used for intracellular communication) have begun to be explored for use as
drug delivery vehicles for non-native therapeutics such as nucleic acids and
low MW drugs such as curcumin[1-3] and doxorubicin[4].
Introduction
exoPTX is More Cytotoxic Than Taxol or Paclitaxel In Vitro
• Sonication provided the greatest loading capacity* (28.29 ± 1.64%) as compared to incubation (12.12 ±
3.67%) and electroporation (12.68 ± 3.53%). The loading capacity of the commercial formulations of
paclitaxel, Abraxane and Taxol, are 10% and 1%, respectively. *Loading Capacity calculated by µg exosomal protein
• exoPTX is more cytotoxic than either Taxol or Paclitaxel alone in vitro, when tested against a Lewis Lung
Carcinoma cell line (3LL-M27), as well as MDCK WT and MDCK MDR1 (Pgp+) cell lines.
• Uptake of exosomes by 3LL-M27 cells is greater than uptake of other commonly used nanocarriers
(liposomes or polystyrene (PS) nanoparticles) in vitro .
• Intranasally administered exosomes and their drug payload co-localize with pulmonary metastases in vivo.
• Intranasally administered exoPTX exhibits greater therapeutic efficacy as compared to Taxol in a mouse
model of pulmonary metastases.
• Exosomes do not induce any organ toxicity.
Summary
Exosomes (107 particles/10µL x2) or PBS were intranasally administered to healthy C57/BL mice every other
day for 12d (6x total injections). Paraffin tissue sections of lung tissue were then stained with H&E and
imaged.
exoPTX Exhibits Greater Therapeutic Efficacy than Taxol In Vivo
Drug-Loaded Exosomes Co-localize With Pulmonary Metastases In Vivo
References1.Sun, D., et al., A Novel Nanoparticle Drug Delivery System: The Anti-inflammatory Activity of Curcumin Is
Enhanced When Encapsulated in Exosomes. Mol Ther, 2010. 18(9): p. 1606-1614.
2.Zhuang, X., et al., Treatment of Brain Inflammatory Diseases by Delivering Exosome Encapsulated Anti-
inflammatory Drugs From the Nasal Region to the Brain. Mol Ther, 2011. 19(10): p. 1769-1779.
3.Kalani, A., et al., Curcumin-primed exosomes mitigate endothelial cell dysfunction during
hyperhomocysteinemia. Life Sciences, 2014. 107(1–2): p. 1-7.
4.Tian, Y., et al., A doxorubicin delivery platform using engineered natural membrane vesicle exosomes for
targeted tumor therapy. Biomaterials, 2014. 35(7): p. 2383-2390.
Acknowledgements
1UNC Eshelman School of Pharmacy, Division of Molecular Therapeutics, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7362, USA2Center for Nanotechnology in Drug Delivery, Division of Molecular Pharmaceutics UNC Eshelman School of Pharmacy, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-7362, USA
3Laboratory of Chemical Design of Bionanomaterials, Faculty of Chemistry, M.V. Lomonosov Moscow State University, Moscow 119899, Russia
Myung Soo Kim1, Matthew Haney1, Yuling Zhao1, Richa Gupta1, Onyi Okolie1, Phi Phua1, Shawn Hingtgen1,2, Alexander Kabanov1,2,3, Elena Batrakova1,2
Exosome-Encapsulated Water-Insoluble Small Molecule Chemotherapeutics for the Treatment of Pulmonary Metastases
PBSTaxolexoPTX
3LL-M27 /DAPIBar = 50µm
This work was supported by the United States National Institutes of Health grant 1RO1 NS057748. We would
also like to thank The Center for Nanotechnology in Drug Delivery at UNC for their assistance and for allowing
us to use their instrumentation.
0
5
10
15
20
25
30
35
Incubation Electroporation Sonication
Load
ing
Cap
acit
y (%
)
Comparison of Drug Loading Methods
Exosomes/DAPI 3LL-M27 /DAPI 3LL-M27 with DOXDOX/DAPI
Bar = 20µm
Drug Cell Line IC50 (ng/mL)
exoPTX 3LL-M27 13.57 ± 1.33
Taxol 3LL-M27 23.16 ± 1.88
Paclitaxel 3LL-M27 126.41 ± 31.31
exoPTX MDCK WT 23.33 ± 3.77
Taxol MDCK WT 69.54 ± 11.50
Paclitaxel MDCK WT 428.77 ± 63.37
exoPTX MDCK MDR1 187.5 ± 38.65
Taxol MDCK MDR1 1708.67 ± 299.93
Paclitaxel MDCK MDR1 >10,000
ResultsSonication Provides the Greatest Drug Loading
0
100
200
300
400
500
600
700
800
0 10 20 30 40 50
# Pa
rtic
les
Mill
ion
s
Time (hrs)
Uptake, 3LL-M27, # Particles vs Time (hrs)
Exosomes PS Nanoparticles
Exosomes are Taken Up More Readily by Lewis Lung Carcinoma Cells than Other Commonly Used Nanocarriers In Vitro
Bar: 20 µm
3hrs:
24hrs:
Exosomes Liposomes PS Nanoparticles
C57/BL mice with pulmonary metastases (3LL-M27) were intranasally administered DiD-labeled doxorubicin-
loaded exosomes. After 18d, sections of lung tissue were stained and imaged by confocal microscopy.
Exosomes Distribute to Lung Tissue In Vivox10 x40
Exosomes were labeled with DiD (red) and intranasally administered to healthy BALB/C mice. 4h later, lung
sections were stained and examined by confocal microscopy.
Future Direction
Our exoPTX formulation exhibits greater cytotoxicity in vitro than Taxol, the commercially available
formulation of PTX. Furthermore, intranasally administered exosomes travel to sites of pulmonary metastases
in vivo and exoPTX exhibits greater therapeutic efficacy than Taxol in vivo. It is worth noting that our exoPTX
formulation appeared to mitigate the drug-resistance phenotype afforded by Pgp expression in MDCK MDR1
cells. The exact mechanism behind this phenomenon has yet to be elucidated and warrants further research.
Thus, exoPTX holds great promise for use in the treatment of pulmonary metastases and may represent a new
platform for the delivery of water-insoluble chemotherapeutics.
Conclusions
A release study of exoPTX will be conducted in parallel with a stability study to determine the release profile
and stability of our exoPTX formulation. The intracellular distribution of PTX when delivered through exoPTX
will also be studied. In addition, the apparent ability of exoPTX to circumvent Pgp-mediated resistance will
be further investigated, and the cytotoxicity of exoPTX against Pgp+ and/or MRP1+ cell lines will be assessed.Exosomes Do Not Induce Any Organ Toxicity
C57/BL mice with pulmonary metastases (red) were intranasally administered exoPTX (107 particles/10µL x2)
or Taxol (equivalent amount of PTX), or PBS every other day for 12d (6x total injections). After 18d, sections
of lung tissue were stained and imaged by confocal microscopy.
Exosomes:
PBS:
Kidney Spleen Liver
AFM Image of exoPTX
HypothesisIncorporation of paclitaxel into exosomes (exoPTX) will increase its solubility and therapeutic efficacy against
pulmonary metastases.
Exosomes & Microvesicles
Macrophage
Exosome Isolation and Drug Loading
Cytotoxicity was assessed using a standard MTT assay.
http://www.ozyme.fr/gammes/sbi/images/exoquick_exosomes_proto_hztl505-351.pnghttp://www.sigmaaldrich.com/content/dam/sigma-aldrich/product1/114/cls3289g.jpg/_jcr_content/renditions/large.jpg
https://immunology.org/image/17-15_Macrophages_1.jpg
Incubate in exosome-
free media for 3 days
Remove cells and cell
debris, take supernatant
Exosome Isolation: ExoQuick-TC™
Drug Loading
HPLC
Nucelosil C18
exoPTX purification
(NAP10 column)
Incubate exoPTX
at 37°C for 30min
http://www.iopan.gda.pl/fizyka/biofizyka/images/stories/biooptyka/hplc%20system%20agilent%201200.jpg
Data Analysis
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