Nanomedicine

1. Nanomedicine Jonathan P. Rothstein

2. Introduction • What is nanomedicine? • It is nanotechnology used for the treatment, diagnosis, monitoring and control of biological systems • It includes the delivery and targeting of pharmaceutical, therapeutic, and diagnostic agents using nanoparticles to cancer and other cells • It includes nanomaterial for bone, cartilage, vascular, bladder and neural applications • What isn’t nanomedicine? • Flesh eating/repairing nanorobots

3. Introduction – Goals of Nanomedicine • End goal of nanomedicine is improved diagnostics, treatment and prevention of disease • For a great review see http://www.wtec.org/nano2/Nanotechnology_Research_Directions_to_2020/

4. Introduction – Goals of Nanomedicine • Ultimate goal is to integrate detection, diagnostics, treatment and prevention of disease into a personalized single platform

5. Introduction • Nanotechnology holds key to a number of recent and future breakthroughs in medicine

6. BioMEMS for Screening and Diagnostics • Goal is to develop handheld diagnostic devices for personalized medical testing and treatment Biomedical Analysis and Communication System Disposable Diagnostic BioChip

7. BioMEMS – Micro and Nanofluidics

8. Nanoparticles for Pathogen Detection Fluorophore Release Nanoparticle Probe Targeted RNA • Gold nanoparticles can be functionalized with thiolated oligonucleatides. • Bound to the oligonucleatides are fluorophores which are quenched by their proximity to the nanoparticle. • When the targeted RNA (H2N2, HIV or a cancer) bindes to the oligonucleatide, the fluorophore is released and becomes fluorescence. • The fluorescence can be detected in a BioMEMS device. • Challenge is developing oligonucleatides with high selectivity for the target RNA.

9. Nanoparticles for Targeted Detection of Cancer Healthy Cells Breast Cancer Cells • As an example, nanoparticle probes were developed by Chad Mirkin at Northwestern Univ. that target the survivin RNA sequence known to exist in a certain breast cancers. • Experiments are done ex-vivo. • On the left, cancer cells fluoresce. • On the right, healthy cells show minimal fluorescence.

10. Nanoparticles for In-vivo Detection of Pathogens • Fluorescence is not a viable option in-vivo, but magnetic tagging works very well. • Harmless virus can used as a building block to produce contrast agents that can be used in Magnetic Resonance Imaging (MRI). • Here, magnetic metal ions are bonded to the virus as are molecules that bind to cancer cells. • A full body MRI scan detects these contrast agents and even very small tumors throughout the body

11. Nanoparticle Encapsulation for Drug Delivery • Nanoparticle shells can be formed around spherical droplets • A.D. Dinsmore, et al., Science 298, 1006 (2002), Y. Lin, et al., Science 299, 226 (2003) • Shells are porous at lengthscales much smaller than size of nanoparticle. A: Scanning electron microscope of a dried 10-μm-diameter colloidosome composed of 0.9- μm-diameter polystyrene spheres.

12. Why Particles Adsorb to Interfaces [Pickering (1907); Pieranski PRL 45, 569 (1980)] I. Particle (P) away from interface: Interfacial Area = A P surface tension (Oil) Energy = AgO/W + 4pR2gP/O (Water) II. Particle sitting astride the interface (half-in, half-out): Energy = (A-pR2)gO/W + 2pR2gP/O+ 2pR2gP/W • If |gP/O –gP/W| < gO/W/2, then adsorption reduces surface energy.

13. oil-nanoparticle suspension, w/ droplets water droplet: mm to mm nm Nanoparticles At Interfaces • Nanoparticles can be functionalized, cross linked or sintered to make shell permanent, strengthen shell or change shell permeability.

14. Nano-Encapsulation for Drug Delivery • By making the holes between nanoparticles approximately the same size as the drug you want to administer you can get a constant release rate – avoids spikes in dosage. • Can also allow encapsulation of hydrophobic drugs which are difficult to get into you mostly water body. Standard Diffusion Based Drug Delivery Nano-Encapsulated Drug Delivery Drug Concentration in Patient Time

15. Targeted Delivery to Tumors • Goal is to inject treatment far from tumor and have large accumulation in tumor and minimal accumulation in normal cells/organs.

16. Cancer Treatments • Tumor penetration is a key issue for successful chemotherapy

17. Nanoparticle use in Cancer Treatments • Because of their small size, nanoparticles can pass through interstitial spaces between necrotic and quiescent cells. • Tumor cells typically have larger interstitial spaces than healthy cells • Particles collect in center bringing therapeutics to kill the tumor from inside out.

18. Making Gold Nanoparticles • AuCl4- salts are reduced using NaBH4 in the presence of thiol capping ligands • The core size of the particles formed can be varied from <1 nm to ~ 8 nm • The surface functionality can be controlled through the choice of thiols • Diffusion speed can be controlled by length of thiols

19. Nanoparticles as Sensors and Therapeutics • Glutathione (GSH) provides a selective and tunable release mechanism • Once inside cells, fluorophores and drugs selectively dissociate

20. Nanoparticle Success • Both cationic and anionic particles penetrate and accumulate in tumors. • However, only cationic particles diffuse fully throughout the tumor. • Work of Neil Forbes and Vince Rotello at UMASS

21. Nanoparticle Targeting and Accumulation • To maximize their effectiveness, the microenvironment of the tumor must be quantified and vectors developed to specifically target the tumor. • These treatment approaches have shown great promise in mice. Necrotic Quiescent Proliferating Therapeutic

22. First Successful Nanomedicine - Abraxane

23. Alternatives to Nanoparticles - Surfactants • Surfactants are composed of a hydrophilic head and a long hydrophobic tail • When dissolved in water above the critical micellar concentration (CMC) surfactants can self-assemble into large aggregate • Spherical micelles are around 10nm in size • Hydrophobic drugs can be encapsulated and in their core and delivered throughout the body or to a specific target.

24. Nanotechnology in Tissue Engineering – Cartilage Replacement • Samuel Stupp at Northwestern has shown that nanotechnology can be use to regenerate severed spinal cords. • Two polypeptides amphiphiles are used that when mixed in an aqueous solution self assemble into a nanotube • As seen on right, these nanotubes display peptide growth factors. • In mice, these systems have been shown to promote axonal outgrowth and bridging of injured areas (bottom right).

25. Nanotechnology in Tissue Engineering – Cartilage Replacement • Because cartilage doesn’t have vasculature and cannot repair itself, accepted treatments have been mostly mechanical in their approach. • Joint lubricants: • Simple and effective at short-term pain relief but do not address cause of the problem or repair any damage. • Debridement/lavage/microfracture: • Small lesions are repaired by shaving or shaping contour of cartilage. • Microfracture penetrates subchondral plate (bone) and actually causes growth of fibrocartilage – a lesser form, not desirable. • Total joint replacement: • Addresses problem and generally allows full repair, but • Very invasive procedure, native tissue removed • Prostheses do not last a lifetime in active patients. • Nanotechnology approach • Regrow patient’s own cartillage in-vivo to repair damage www.hughston.com/hha/

26. ACT Methods • A popular tissue engineering approach has been to introduce new cells, via autologous chondrocyte transplantation/implantation (ACT/ACI). • Some of the earliest work by Benya and Shaffer (1982) showed it was possible to isolate and culture chondrocytes. • More interesting result was that when cultured in vitro, the cells differentiated and changed their phenotype to produce a lesser quality collagen. • Need better tissue scaffolds – nanotech. Important to tissue engineering:Cells will differentiate purely based on mechanical stimulus. Genzyme ACT method: FDA approved 1997 biomed.brown.edu

27. Hydrogels – Self Assembly • Hydrogels have applications in drug delivery and tissue engineering • Regenerating cartilage and other tissue requires scaffolds with similar modulus and other mechanical properties → Need to develop stiffer, tunable hydrogels • We investigated Polylactide-Polyethylene Oxide-Polylactide triblock copolymers. • Systems are biocompatible with a hydrophobic ends (PLA) and a hydrophilic center (PEO) which self-assembles in water and can form a gel under the right conditions Reinforced Through Addition of Nanoparticles Gelation CMC Micelle Triblock Copolymer Gel

28. Rheology of Hydrogels • The hydrogels formed are very stiff with elastic modulus on the order of 1-10 kPa. • Within range of moduli of several human tissues including cartilage. • Gels formed from polymers with higher degree of polymerization maintain a high storage modulus even at physiological temperatures (370C). • In-vivo applications feasible. • Rheological response of these polymers can be easily tuned by varying the crystallinity or block length of PLA or through addition of nanoparticles. R-Lactide Amorphous Core L-Lactide Crystalline Core Khaled et al. Biomaterials (2003)

29. Photocrosslinking Hydrogels for Cartilage Replacement • An alternate approach is to make the hydrogel from polymers that can be crosslinked after injection. • From Jennifer Elisseeff’s lab at Johns Hopkins University. • Photo-polymerizing the hydrogel increases its modulus, allowing the appropriate phenotype of cartilage to be expressed and protecting damaged area from wear.

30. Keeping Things Clean – Antimicrobial Surfaces • Silver is an excellent anti-microbial agent • Silver nanoparticles are now being added to fibers of clothing and bandages as well as being incorporated into surfaces in hospitals to reduce the rate of bacterial infections • When co-extruded with a polymer like PLLA, the silver is released slowly over time and has been shown to effectively kill bacteria