Nanotechnology in BioMedical Sciences

EarlJ.Bergey,BruceA.Holm,TapasDe,YudhisthiraSahoo,David
Rodman, ChristopherFriend,IndrajitRoy,andParasN.Prasad

InstituteforLasers,PhotonicsandBiophotonicsand
SchoolofMedicineandBiomedicalSciences
TheStateUniversityofNewYorkatBuffalo
Buffalo,NewYork14260

Abstract

Nanotechnology is rapidly expanding into the biomedical field. At the Institute for Lasers,
Photonics and Biophotonics, research in nanotechnology has focused on the development of
multifunctional nanosized particles we have termed as nanoclinics. They consist of a nanosized
bubblefabricatedtocarrytherapeuticordiagnosticcomponentstotargetedcells.Theoutersurface
of these particles can be functionalized to incorporate biologically active targeting agent. This
technology provides a platform for the development of new imaging, diagnostic and therapeutic
modalities. For bioimaging and diagnostics, nanoclinics are fabricated containing rareearth ions,
whichexhibittwophoton,antistokesluminescencebyfrequencyupconvertinginfraredtovisible
light.AttheInstitute,wehavesuccessfullypreparedthephosphorscontainingnanoclinicshavinga
size ~25nm with a silica shell around it which helps in aqueous dispersability and also allows
functionalization for covalent attachment of bioprobes for targeting. The coupling of specific
peptides,proteinsornucleicacidsequencestothesilicashellwillallowfortheselectivedetection
of biological entities that will have applications in microscopy, flow cytometry, ELISA and
DNA/RNA hybridization systems. This adaptable nanoparticle platform was used to develop
nanoclinics,whichselectivelytargetspecificcellortissuetypes.Thesetargetednanoclinicscan
carrytherapeuticdrugs,DNAforgenetransfertherapy,orprovideexternallyactivatedtherapyasin
photodynamictherapyormagneticfieldactivation.Thecoreofourprototypicnanoclinicconsists
of either Fe2O3 or Fe3O4 to which a fluorophore is coupled. These nanoparticles are encased in
eithersilicaorbyafattyacidbilayer,respectively.Thefinalstepistoattachthetargetingligand
LHRH(luteinizing hormone-releasing hormone) tocompletethenanoclinic.Thisdesignallows
for detection of specific cells or tissues using standard MRI equipment and can be used
therapeutically with the same instrumentation (stronger DC magnetic field). These nanoclinics
cause magnetocytolysis of LH-RH receptor-positive cancers, in vitro, in a DC magnetic field..
These therapeutic nanoclinic systems could provide the clinician with a new tool in the fight against
human diseases.

NanoClinics
Composed of Silica or CaPO4 shell encapsulating , Rare-earth
elements, iron oxide, DNA or Drugs
Less than 50 nm in size (tunable)
Produced by a reverse micelle process
Surface can be modified (targeting)
Rare Earth elements
Efficient upconversion (blue, red and green)
Stable (little photobleaching)
Diagnostic Applications in Bioimaging and Sensing in
Microscopy, Flow Cytometry, ELISA, Fluorescent Imaging
systems, MRI contrast agent
Therapeutic Applications in Gene therapy, Drug Delivery
Systems, Photodynamic Therapy, and Direct Cancer Therapy

First Generation of Nanoclinics

Cancer diagnosis and targeted therapy

Luteinizing Hormone-Releasing Hormone (LH-RH)

Targeted Nanoclinic
LHRH

Silica

Iron Oxide

Nanoclinic

Two photon dye

LHRH

LHRH

Uptake of LH-RH-Nanoclinics in MCF-7 Cells

transmission

Fluorescence
TPLSM image of
breast carcinoma cells.
The arrow shows
nanoparticles
accumulating on and
inside the cell.

Particles saturation with time

3min

9min

15min

21min

27min

LH-RH +

4
3

LH-RH
2

4
3

7 2 h rs .

2 h rs .

U C I-1 0 7

M C F -7

U C I-1 0 7

N u m b e r o f C e lls L y s e d (x 1)0

LH-RH +

M C F -7

N u m b e r o f C e lls L y s e d (x 1)0

Specificity of Magnetocytolysis by Nanoclinics

Nanoclinics incubated with adherent cell for 24 hours. Cells washed, released by
trypsinization and subjected to DC magnetic field. Magnetocytolysis determined by
counting remaining cells.
MCF-7: Human breast carcinoma - LHRH receptor positive
UCI-107: Human uterine carcinoma - LHRH receptor negative

Effect of Up and Down Regulation of the LH-RH Receptors

on Magnetocytolytic Activity of LH-RH-Nanoclinics
C LH-RH +

LH-RH -

Num ber of Lysed Cells (x 10 )

12
10
8
6
4
2
0

EGF (epidermal growth factor) up-regulation

RC160 (synthetic somatostatin) down-regulation

Surface Topography of MCF-7 Cells Incubated with

LHRH-Nanoclinics - AFM

Effect of Cellular Interaction

Magnetic Properties of Nanoclinics
1.2

b
1

M(au)

0.8

Particles with cells

Particles in media

0.6
Cell Bound particles

Free particles

0.4

B
a

0.2

cell
0
0

500

1000

B
b

cell
1500

H(Gauss)

2000

2500

Upconversion Emission by
Rare-earth Nanophosphors

Tm/Yb:Y2O3

Tm - thulium
Er - erbium
Yb - ytterbium
Y2O3 - yttrium oxide

Er:Y2O3

Er/Yb:Y2O3

Upconverting Nanotechnology for

Bio-Imaging

KB Cells

Emission from
Nanophosphor

KB Cells and
Emission from
Nanophosphor

TEM of DNA-Doped CaPO4 Nanoparticles

In Vitro Transfection Efficiency of

DNA-Doped CaPO4 Nanoparticles

Cell line = Jurkat

DNA pSVgal (-galactosidase)
Positive Control Superfect

See acknowledgements

In Vivo Transfection with DNA-Doped CaPO4 Nanoparticles and

I.P. Injection

I.M. Injection

Swiss albino mice 15 gr.

Galactose-Surface Modified DNA-Doped CaPO4 Nanoparticles

I.P. Injection

Swiss albino mice 15 gr

I.M. Injection

See acknowledgements
See acknowledgements

Conclusions
Nanoclinics provide the following advantages:
Multifunctional nanoparticulate platform
Surface modifiable for coupling of targeting agents
Can carry different payloads (Therapeutic or Imaging)
Can exert therapy using external activation
Up-conversion bioimaging with reduced background

Safer in vivo gene transfer system (compared to viral)

Acknowledgements
The gene transfer studies were performed by Dr. I Roy
under the direction of :
Dr. A.N. Maitra
Department of Chemistry
University of Delhi
Delhi, India
as partial requirements for the completion of his Ph.D.
Thesis.