Molecular Imaging and Therapy with Perfluorocarbon Nanoparticulates:

Environmental Impact.

Washington University Medical School

Saint Louis, MO

GENERALIZED TARGETING PARADIGMGENERALIZED TARGETING PARADIGM

Targeting system: “Molecular zip codes”(MAb, aptamers, small peptides, peptidomimetics, polysaccharides, etc)

PayloadsImaging agents (Gd3+)DrugsRadionuclides 19FGd3+Lipid coatAnti-fibrinAnti-tissue factorAnti−αvβ3-integrin

Liquid VentilationVitreous

Replacement

Perfluorocarbon Biomedical Applications

Before After

0.5 µ 0.5 µ 0.5 µ

RBC

Fibrin

TargetedContrast

Control Thrombus Targeted Thrombus

1 mm

Clot

Suture1 mm

Clot

Suture

Scanning EMclot labeled with fibrin targeted nanoparticles

Ultrasound contrastenhancement of fibrin clot in vitro using conventional clinical scanner (7.5 MHz)

Fibrin Imaging

Targeted Nanoparticles Support Enormous Paramagnetic Payloads

Targeted Targetedαβ3- Non-α β - Non-

mM)-1 [Particle] 1,800,000 1,950,000

r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000

94,400 92,400Gd3+Ions / Particle 94,400 92,400Gd3+Ions / Particle

19.1 21.1r1 (s*mM)-1 [Gd] 19.1 21.1r1 (s*mM)-1 [Gd]

22.9 24.6r2 (s*mM)-1 [Gd] 22.9 24.6r2 (s*mM)-1 [Gd]

r1 (s*mM)-1 [Particle] 1,800,000 1,950,000r1 (s*mM)-1 [Particle] 1,800,000 1,950,000

r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000

α β - Non-α β - Non-α β - Non-α β - Non-

mM)-1 [Particle] 1,800,000 1,950,000

r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000

94,400 92,400Gd3+Ions / Particle 94,400 92,400Gd3+Ions / Particle

19.1 21.1r1 (s*mM)-1 [Gd] 19.1 21.1r1 (s*mM)-1 [Gd]

22.9 24.6r2 (s*mM)-1 [Gd] 22.9 24.6r2 (s*mM)-1 [Gd]

r1 (s*mM)-1 [Particle] 1,800,000 1,950,000r1 (s*mM)-1 [Particle] 1,800,000 1,950,000

r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000

mM)-1 [Particle] 1,800,000 1,950,000

r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000

94,400 92,400Gd3+Ions / Particle 94,400 92,400Gd3+Ions / Particle

19.1 21.1r1 (s*mM)-1 [Gd] 19.1 21.1r1 (s*mM)-1 [Gd]

22.9 24.6r2 (s*mM)-1 [Gd] 22.9 24.6r2 (s*mM)-1 [Gd]

r1 (s*mM)-1 [Particle] 1,800,000 1,950,000r1 (s*mM)-1 [Particle] 1,800,000 1,950,000

r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000r2 (s*mM)-1 [Particle] 2,160,000 2,270,000

Detection of Angiogenesis in the Vx-2 Model

Masses Seen By MRI Were Not Always Vx-2 Tumors

T2-Weighted MRIAfter

avB3-Nanoparticles

5 mm5 mm

Histology Revealed Rejected Tumor Infiltrated with Inflammatory Cells

No Molecular Imaging of No Molecular Imaging of Angiogenesis for Tumors in Angiogenesis for Tumors in

RejectionRejection

Histology Revealed Rejected Tumor Infiltrated with Inflammatory Cells

Melanoma Angiogenesis: Detection With αvβ3 Integrin-Targeted Paramagnetic NanoparticlesMouse Imaging @ 1.5T

Angiogenesis around 2 mm melanoma tumor

2 mm tumor

syringe

Time 0 min

Time 120 min

Time course

1

1

1

1

1

1

J

J

J

J

J

J

H

H

HH

H

H0

20

40

60

80

100

120

140

160

180

0 20 40 60 80 100 120

1 Targeted

J Control

H Competition

Time course of MRI Contrast Enhancement P

erce

nt

En

han

cem

ent

Time (min) Post Injection

Site-Targeted Drug Delivery

D

Target Cell

D

D

0.5 µ

2. Particles bind to cell surface

1. Load with drug 3. Drug effect

Targeting Element

Drug Loaded Outer

Lipid Monolayer

Exchange of

Lipids and Drug

Components

Target Cell Membrane

Epitopes

Novel Mechanisms Of Drug Delivery: “Contact Facilitated Drug Delivery*”

Rhodamine-labeled NP

GFP-linked endocytotic markers rab5 and rab7 in transfected

C32 melanoma cells

*Lanza GM, Wickline SA et al: Targeted antiproliferative drug delivery to vascular smooth muscle cells with magnetic resonance imaging nanoparticle contrast agent: implications for rational therapy of restenosis. Circulation 2002; 106:2842-2847

Antiproliferative Effects of Free versus Tissue Factor - Targeted Paclitaxel Nanoparticles on

Vascular Smooth Muscle Cells

Control Taxol 0.2 mol % Taxol 2.0 mol %0

5

10

15

20

25

30C

EL

L C

OU

NT

X 1

0,0

00

NANOPARTICLE FORMULATION

NON -TARGETED NANOPARTICLES

TARGETED NANOPARTICLES

* *

Control3/3

αβ3DXR-NP(4/4)

Rejection of Mouse Melanoma 7d following αβ3-Targeted, DXR, Paramagnetic Nanoparticles

Proton and Fluorine Imaging

Proton Imaging Fluorine Imaging

Clot

y = 0.2665x - 2.1857R2 = 0.9462

-10010203040

0 20 40 60 80 100Emulsion percentage

y = 0.0493x - 0.641R2 = 0.9178

-2

0

2

4

6

0 25 50 75 100Estimated % of Emulsion Volume

15:5 Crown-Ether

PFOB

Dosimetry of Targeted Nanoparticle Levels

Alliance Pharmaceutical

Oxygent Production

Typical Pharmaceutical Emulsification Process

President Bush: reduce the greenhouse gas intensity of the U.S. economy by 18% over the next ten years.

Perfluorocarbons have 10,000 times the global warming impact of carbon dioxide

Major PFC applications:

• electronic insulators

• aluminum smelting

• refrigerants

• subsurface barrier testing

EPA suggested four options for its formal definition of "leak rate." :

• "annualizing" method.

• "rolling-average" method

• the higher calculated leak rate.

• consistent method

Non-VOC with no significant participation in ozone formation in ambient air.

MedicalGregory Lanza, MD, PhDSamuel Wickline, M.D.

TechnicalMary Watkins, R.T. (MR)Todd Williams, B.S., R.T. (MR)Rosa Lin, M.S.Brian Hennery, B.S.Ralph FuhrhopHuiying Zhang, M.D.Stacy Allen, B.S.Liz Lacy, B.S.Peggy Brown, RDCSMichael Scott, BSAnne Schmeider, M.S.Kathy Crowder, MS

MR & US SciencePatrick Winter, Ph.D.Michael Hughes, Ph.DJon Marsh, Ph.DFrank Hockett, M.S.E.E.Michal Lijowski PhDGrace Hu, M.S.Anagana Pan Ph.D

WU Personnel

Key Collaborators

Philips Medical Systems

David Rollo MD

Horace Hines PhD

Ling Shao PhD

Carmen van Vilsteren PhD

Christopher Hall PhD

Shelton Caruthers PhD

The Dow Chemical Co.

Jaime Simon PhD

Garry Kiefer PhD

Phillip Athey PhD

Gyongyi Gulyas PhD

Keith Frank PhD

Consultants

Sam Achilefu PhD

Wynn Volkert PhD

Richard Holmes PhD

Patrick Gaffney PhD

Bristol-Myers Squibb Medical Imaging

D. Scott Edwards

Stephen Haber

Thomas Harris