Intelligent BiomaterialsIntelligent BiomaterialsProtein Delivery Protein Delivery

Molecular Imprinting and MicropatterningMolecular Imprinting and Micropatterning

Nicholas A. Peppas

Our LaboratoriesOur Laboratories

• 22 Researchers

– 12 Ph.D. students (8 ChEs, 4 BMEs)

– 3 Visiting scientists (Italy)

– 1 Technician

– 6 Undegraduate students• About 3,800 sq.ft. facilities

• Modern equipment including cellular facilities

• Budget of about $ 2M

• Grants from NIH, NSF, industry

The Changing World of Biomaterials, The Changing World of Biomaterials, Drug Delivery and Biomolecular EngineeringDrug Delivery and Biomolecular Engineering

• Formation and fabrication of supramolecular assemblies comprising natural biological elements, structures or membranes.

• Synthesis and preparation of modified biological molecules

• Biomolecules as the basis of nanostructures, molecular adhesives

• Micropatterned and microfabricated arrays

Oral protein deliveryOral protein delivery

Oral Delivery of ProteinsOral Delivery of Proteins

Why?• Increase patient compliance

and comfort over other forms of drug delivery (i.e. injection)

• Mimic physiologic delivery of proteins

• Simple administration• Reduce costs• Potentially improve efficacy

“Oral delivery of peptides and proteins has long been dubbed the ‘Holy Grail’ of drug delivery…”

Challenges of Oral Protein DeliveryChallenges of Oral Protein Delivery

1. Protect the drug– Acidic environment in the stomach– Proteolytic enzymes in the GI tract

2. Improve bioavailability– Increase drug transport across intestinal

epithelium– Localize drug at targeted site of

absorption

3. Maintain biologically active and stable drug

GI Tract is designed to digest proteins and food.

Transport for Oral Drug AbsorptionTransport for Oral Drug AbsorptionTransport Mechanism

a. Transcellular pathway

b. Paracellular pathway

c. Transcytosis and receptor-mediated endocytosis

d. Lymphatic absorption through M cells

e. P-glycoprotein efflux (not shown)

Factors Affecting Transport

1. Molecular mass of drug

2. Drug solubility

In VivoIn Vivo Study with pH-Responsive Study with pH-Responsive Complexation HydrogelsComplexation Hydrogels

• P(MAA-g-EG) microspheres loaded with insulin

• Administered to diabetic rats 40% drop in blood glucose levels

• Prior work done by Tony Lowman

Carrier MediatedCarrier Mediated

• Biodegradable polymers, lectin modified carriers

• Sites of uptake1. M cells (majority of uptake)

2. Transcellular

3. Paracellular

• Poor particle absorption

Goal: Protect drug in the GI tract and be absorbed with drug by epithelial layer.

Florence, A. T. The oral absorption of micro- and nanoparticulates: neither exceptional nor unusual. Pharm Res 1997, 3, 259-266.

MucoadhesionMucoadhesion

Mucosa Mucosa

Decomplexation

Stomach Upper small Intestine

Blood Glucose Response in Healthy and Diabetic Wistar Rats

40

60

80

100

120

140

0 2 4 6 8

Healthy Animal

Diabetic Animal

Serum Glucose (% of Initial Level)

Time t, (h)

Systemic Circulation

Polymeric Carrier

TightJunction

Mucosa

Protein

ProteoliticEnzymes

Caco-2 Cells as GI ModelCaco-2 Cells as GI Model Caco-2 Cells as GI ModelCaco-2 Cells as GI Model

• Advantages

– Spontaneously differentiate– Produce enzymes– Posses tight junctions– Develop microvilii– Transport of inorganic

molecules correlates well with the in vivo absorption

• Disadvantages

– Do not produce mucus– The properties are

determined by the passage number

Nanodevices of Intelligent GelsNanodevices of Intelligent Gelsfor Protein Releasefor Protein Release

GOx

GOx

GlucA

GOx

GOxG

Empty hydrogel absorbs glucoseleading to gluconic acid production

Decrease in pH leads to gel expansionwhich releases insulin

G

G

G

GGGlucA

GlucAGOx

GOx

G

G

G

GGlucA

GlucA

I

I I

I

II

I I

I

I

II

Targetting and Targetting and NanotechnologyNanotechnology

• Targeted delivery for cancer therapy

• Gene delivery

• Long term treatment of chronic diseases

BioMEMS Sensor PlatformBioMEMS Sensor Platform

• Pattern environmentally responsive hydrogels onto silicon microcantilevers to

create a BioMEMS/MEMS sensor device.

Laser beam

θ

Polymer

Silicon

φ > θ

Laser beam

φ

Change in pH, temperature, etc. hydrogel swells

Experimental ProcedureExperimental Procedure

• Surface Modification

• Micropatterning

Organosilane (?- )MPS Surface Treatment

OHOH

OHOH

OHOHOH

OH

O

Si

O

OO

OO

OOOO

O

Si

OO

Si

OO

Si

OO

Si

O

O

Si

O

REACTIVE SITES

Provides inorganic/organic interface

Organosilane ( ?-MPS) Surface Treatment

OHOH

OHOH

OHOHOH

OH

O

Si

O

OO

OO

OOOO

O

Si

OO

Si

OO

Si

OO

Si

O

O

Si

O

REACTIVE SITES

Provides inorganic/organic interface

OHOH

OHOH

OHOHOH

OHOH

OHOH

OHOH

OHOHOH

O

Si

O

OO

OO

OOOO

O

Si

OO

Si

OO

Si

OO

Si

O

O

Si

OO

Si

O

OO

OO

OOOO

O

Si

OO

Si

OO

Si

OO

Si

O

O

Si

O

REACTIVE SITES

Provides inorganic/organic interface

Provides inorganic/organic interface

Silicon substrateSurface treated with organosilane

agent to induce bondingMonomer applied to

treated silicon substrate

Micropatterned polymer bonded on silicon substrate

UV light

Photomask

Masked UV polymerization

Silicon substrateSurface treated with organosilane

agent to induce bonding

Silicon substrateSurface treated with organosilane

agent to induce bondingMonomer applied to

treated silicon substrateMonomer applied to

treated silicon substrate

Micropatterned polymer bonded on silicon substrate

Micropatterned polymer bonded on silicon substrate

UV light

Photomask

Masked UV polymerization

UV light

Photomask

Masked UV polymerization

Micropatterned Hydrogel on Micropatterned Hydrogel on Silicon MicrocantileverSilicon Microcantilever

Top view images obtained utilizing an optical microscope in Nomarski mode showing a silicon microcantilever patterned with an environmentally responsive hydrogel. In A), the focus is on the substrate, while in B), the focus is on the microcantilever tip. Profilometry indicated that the thickness of the patterned hydrogel is approximately 2.2 m.

100 ? m 100 ? m

Silicon cantilever

Polymer

Silicon cantilever Etched well • Volume shrunk as the

polymerization proceeded

• Polymer adhered to silicon surface and could not shrink at the interface, resulting in stress formation in the polymer film

• This stress in the polymer film resulted in bending the microcantilever

A) B)

Confocal Images of MicroarraysConfocal Images of MicroarraysAcrylamide-PEG200DMA with 67% Crosslinking RatioAcrylamide-PEG200DMA with 67% Crosslinking Ratio

3D Projection of micropatterned recatangular array of a biorecognitive networks obtained utilizing a confocal microscope. Profilometry indicated that the thickness of the micropatterns are approximately 13 m.

50 m

Optical and Confocal Images ofOptical and Confocal Images of MicropatternsMicropatternsAcrylamide-PEG200DMA with 67% Crosslinking RatioAcrylamide-PEG200DMA with 67% Crosslinking Ratio

Microcantilever ShapeMicrocantilever Shape

Images of micropatterned biorecognitive networks. In A), an optical image (Nomarski mode) of recognitive network patterned in shape of cantilever is demonstrated. In B) and C), a confocal microscope slice through middle cantilever pattern of a control and recognitive network, respectively, are shown. Profilometry indicated that the thickness of the micropatterns are approximately 13 m.

25 m 25 m

RecognitiveControl

50 m

SiliconPolymerA)

B) C)