 Our next speaker is a recognized leader in the fields of nanotechnology and tissue engineering. She uses nanotechnology to examine practical application of engineering solutions to difficult medical problems. More specifically, her research focuses on the synthesis, development, and application of new materials with biological function. As an example, her lab is investigating ways to use a patient's own cells to grow replacement blood vessels. This is done by constructing materials that mimic the extracellular matrix and provide structure for the growth of replacement blood vessels. In other words, bioengineering is scaffolding that the body can use to regenerate itself. Another area of active research in her labs involved the use of gold nanoshells for both diagnostic imaging and treatment of cancerous tissue. I asked Dr. West how she decided on a career in science over dinner one night. She told me that she'd never really considered being a chemist while she was younger. But there were several events that in retrospect seemed like signposts. When she was in sixth grade, for example, her family went on vacation to Washington, D.C. Her dad asked her and her sister what they'd like to see while they were in the area, and she immediately said, I want to go see the National Institutes of Health. I don't know about you, but when I was in sixth grade, I had no idea what the National Institutes of Health was. To her father's credit, he made some phone calls and managed to get a lab tour for her. So I'm not sure if you file that under Kids Say the Darnest Things or Daddy's Little Girl Gets What She Wants. But we should take from that that sometimes those small indulgences pay off big in the end. Dr. West went on to graduate from MIT with a B.S. in chemical engineering and obtained a Ph.D. in biomedical engineering from the University of Texas at Austin. Now she's the Isabelle C. Cameron Professor of Bioengineering at Rice University in Houston, Texas. She's affiliated with the Department of Chemical Engineering and Bioengineering and is the Director of the Institute of Biosciences and Bioengineering at Rice. This institute fosters cross-disciplinary research and education programs in biology, chemistry, and engineering. Dr. West received the National Science Foundation Faculty Early Career Development Award. She also won the prestigious Anuncio Award presented by the Christopher Columbus Fellowship Foundation. This award recognizes individual Americans who are improving the world through ingenuity and innovation. And given the venue of this conference, I think it's appropriate to cite her ingenuity and innovation in teaching the next generation of scientists as well. In April, Dr. West was awarded a Howard Hughes Medical Institute grant to facilitate the development of interdisciplinary and cross-disciplinary courses in teaching this next generation of scientists. So with that, would you please join me in welcoming Dr. Jennifer West? Thank you all and thank you all for staying through the end of the conference. I know it's been a long couple of days, but hopefully this will add a little bit of different flavor to what you've been hearing about. So as mentioned, I'm at Rice University in Houston, which is actually a campus very similar in some ways to this campus. It's a small institution, very strong in math and science, with about the same size student body. And even though we're right in the center of Houston, we have a very beautiful and pastoral setting, much like the one here. I think the main difference is who our next door neighbors are. So literally across the street from our campus, we have the Texas Medical Center here. And that's been key for allowing us to really engage in ways of looking at how we can take technologies developed in the basic science and engineering labs at Rice University and translating them into real changes in how medicine is performed and what types of diagnostic and therapeutic applications we really have as we treat patients. So looking at where we are in developing new ways of treating disease and curing patients, we have tremendous advances in basic research. Some of these you've heard about here at this conference, really the advances as we enter the genomic area that Dr. Bishop talked about, the ability to understand and manipulate what stem cells are doing, as Dr. Pollock talked about. And the issue is how do we really see these through to impacting how patients are treated, what happens to them in the medical setting. And to bridge that gap, we need to have the development of suitable technologies. And some of these are the means and methods to make the different basic research advances such as ways to mass produce DNA materials that can be used as therapeutics. Some of it is looking at the biomaterials that you need to really enable regenerative medicine, new types of devices, the application of computational modeling to understand the systems at the basic research level so that we can facilitate their translation into medicine. And so our lab focuses very specifically as far as technology development on what we can do with materials and material science to help translate things from the basic lab through to medical practice. So I'm going to try and tell you today four short stories that give example of some of this type of work. So I'll first talk about some of the work we've done in treatment of coronary artery disease and then leading from there into talking about regenerative medicine, specifically focused on growing new blood vessels, then talk about our work in cancer therapy that Scott mentioned and finally mention a little bit of our work in ways that we can very rapidly detect infectious disease that may help to deal with some of the issues we heard about yesterday with rapid spread of infectious disease. So looking at cardiovascular disease, cardiovascular disease remains the leading cause of death in the United States. A big proportion of that is due to coronary artery disease. Currently treatment options for coronary artery disease are fairly effective but still very limited. So you have angioplasty and stenting and you have coronary artery bypass grafting, but both of these types of therapies suffer from very high failure rates. So one of the main modes of failure with both angioplasty stenting approaches and with vascular graft approaches is a problem referred to as restenosis. Restenosis is basically the reocclusion of a blood vessel due to the formation of scar tissue within it. So that the lesion now inside the blood vessel isn't the lipid filled atherosclerotic plaque, it's now scar tissue but it's still preventing blood flow through that blood vessel. And unfortunately this leads to failure, for example, in angioplasty in about 35 to 50% of patients within the first year. And so that's going to require repeat procedures or escalation in therapy from angioplasty to bypass grafting and of course that brings with it increased costs as well as morbidity and mortality. So we wanted to look at ways to try and deal with this long-standing problem. And so one of the things we've looked at is making materials that can generate a compound called nitric oxide. And so nitric oxide is very interesting because of its ability to... I'm sorry, I think I'm missing a slide just to see if it's out of order. No, I just somehow deleted it, I'm sorry. At any rate, so nitric oxide is a very interesting compound because as we look at restinosis, there are three sets of key cellular events that are related to the induction of the scar tissue formation. The first is at the site where you have an injured blood vessel, you induce blood clotting or thrombosis and that provides a lot of the initiation signals to cause the scar tissue to form. At the same time, you injure cells called endothelial cells that line the surface of the blood vessel and normally keep cell growth in check and prevent thrombosis. And you've also done some damage to the underlying smooth muscle cells and that leads to a cascade of growth factor release as well. And the end result of that is that the smooth muscle cells that form the wall of the blood vessel proliferate out of control and form scar tissue within the vessel. So nitric oxide has the unique ability to act at all three of those points. It interacts with platelets to prevent their clotting actions. It interacts with the endothelial cells to increase their rate of healing. So it increases their proliferation and migration and then it interacts with the smooth muscle cells to block their proliferation. So it in many ways is a wonder drug for this type of problem and the limitation has been the fact that nitric oxide actually acts in all tissues in the body and so you can't just give someone a pill and assume that it will go in and work in their coronary arteries. It would instead induce side effects in many tissues. So we wanted to look at a way that we could use polymer science and materials to now locally deliver this wonder drug just at the site where the vessel's been injured and try and prevent the restenosis after angioplasty or stenting. So we've made a material where the details of the synthesis aren't really important to show you that we've made different polymer compounds where grafted to it we've put molecules that are reacted with nitric oxide which is NO and these form adducts that will then undergo a hydrolytic reaction in the body to slowly generate nitric oxide. And so if we can control where we put these polymer materials then we can control where that generation of nitric oxide occurs. So this shows that when we make these materials we can have prolonged production of nitric oxide for a period of about 100 days which is a sufficiently long time for healing in the blood vessel to occur. And then when we look at the bioactivity of these we look at how it interacts with platelets and ethereal cells and smooth muscle cells. If we look at platelets first we see that these materials can dramatically reduce platelet adhesion and aggregation as we see on this side here relative to the control where we see many more of these fluorescent platelets adhering and beginning to form large aggregates. And then looking at the effects on endothelial cells and then smooth muscle cells we see that exposure to these materials increases the proliferation of endothelial cells and those are basically the good cells that we want to have come in and heal the surface of the vessel and it decreases the proliferation of the smooth muscle cells that are responsible for generating scar tissue. So they are bio-functional and then we had to look at how we could create very localized coatings at the surface of the blood vessel where we've performed our procedure so that we just generate the nitric oxide right where the tissue needs it. So what we do is we make what are called intraluminal hydrogel coatings that's a very thin coating of this soft polymer material on the inside surface of the blood vessel that adheres and conforms to the vessel wall. And this right here is an electron microscope image of one of these coatings the hydrogel coating is the bright white band and the gray to the side is the tissue that it's adherent to the blood vessel wall this coating is about the thickness of one cell layer so it's very thin it's not going to interfere with blood flow it acts as a mechanical barrier to prevent the platelets in your blood from coming and adhering to the site and forming a clot and it acts to locally deliver the nitric oxide just at the site where you've performed your angioplasty or stent deployment. So when we look at this in animal models these are studies that were done in rats where we did a balloon angioplasty procedure in the carotid artery and then we either had control animals that were untreated just went through a normal angioplasty or ones where we created a very thin layer of the nitric oxide producing hydrogel. So as you see on the left we have a blood vessel that was treated with the nitric oxide hydrogel in the staining here we have very dark staining for a protein called elastin and so we can see these bands called the elastic lamina the red arrows point to a structure called the internal elastic lamina above that structure in a normal vessel you should have one single layer of cells and so across most of the vessel wall that's what we have so it looks fairly normal we have some small areas where we have a very minute amount of scar tissue that formed but by and large this blood vessel is open, patent allowing very effective blood flow whereas in the control case our internal elastic lamina runs here pointed out by these two red arrows we have a very significant amount of scar tissue that has formed when we look at it at lower magnification we can see that more than two thirds of the cross-sectional area that should be available for blood flow has been lost and so looking at some of the quantification of this data we can look either at the average thickness of the scar tissue or we can look at the ratio of the intimal area to medial area both are ways of evaluating restinosis in both cases we see better than 75% reduction in restinosis by just forming these very thin coatings of nitric oxide producing hydrogels so we're very excited about that we think that the ability to deal with this problem of restinosis could have a very dramatic effect it's a very prevalent problem in today's healthcare system related work that we've done is to look at making synthetic vascular graphs that also produce nitric oxide it's a very similar concept but in coronary artery bypass grafting you're generally looking at treating vessels where the disease is too severe to simply insert a balloon catheter and do an angioplasty and so instead you sew in a thin tube like we see here to divert flow from the coronary around the obstructed region of the coronary artery so one of the big issues faced in coronary bypass grafting is that you can't use synthetic materials for this partially because they fail due to restinosis and also to thrombosis or blood clot formation so we've made materials that can be used to make these synthetic graphs that produce nitric oxide in a manner very similar to the hydrogels we use in angioplasty and have shown very dramatic effects where we can now use synthetic materials in small diameter applications and see prevention of blood clots and encouragement of endothelialization of the graft and this work was highlighted by New England Journal of Medicine last year as one of the most significant advances from basic research likely to impact clinical medicine so along those same lines of looking at what we can do about the issues associated with blood vessel replacement there's a huge need for vascular graphs not only for atherosclerosis but also in treatment of aneurysm arteriovenous dialysis, trauma and currently there are synthetic materials that look something like this that are available that are basically plastic tubes that you can sew into place to divert flow where you need it to go and these work wonderfully in applications in very large blood vessels like an abdominal aortic aneurysm but they don't work at all in small diameter applications like coronary artery bypass grafting and as I mentioned that's because of thrombosis the blood clot formation on the surface of these materials and because of scar tissue formation that occurs at the ends where you've sewn these into place so restinosis and so that's been a significant limitation so patients have to use transplanted tissue for coronary artery bypass grafting so typically you could for instance take a saphenous vein and transplant that from the leg to the heart and use that to divert flow and unfortunately there's a very significant population of patients who don't have suitable donor tissue for this procedure so either they've had procedures in the past and they've run out of vessels or they may have peripheral vascular disease and thus their blood vessels in their legs aren't in good shape and they're not considered good candidates for this type of procedure so we felt that being able to recreate a blood vessel structure in the laboratory from a very small sample of a patient's cells would allow us to offer an option to this type of patient and so if we want to make a blood vessel in the lab then we want to basically try and recreate as much of the structure and function of a normal blood vessel as we can so just a quick anatomy lesson this is what a blood vessel looks like it's actually a three-layered structure you have a structure on the inside called the intima this is a lining of endothelial cells that helps prevent blood clot formation in your normal vessels the middle layer here is called the media it's composed of smooth muscle cells and elastin fibers and then surrounding this is a layer of connective tissue called the adventitia and to have a functional blood vessel you at least need to have an intima and a media and in an ideal world you would recreate all three of those structures so if we look at how to form an engineered tissue this was gone over briefly this morning by Professor Pollack the general approach is to take some sort of material that's going to act as the scaffold to determine the size and shape that the tissue should ultimately be and provide mechanical support to the cells to allow them to grow in three dimensions to form a new tissue and then you see this with cells you have lots of options about what type of cell as you heard about this morning we focus mainly just on taking adult tissue cells for example isolating blood vessels out of fat samples and then using those cells to form the new tissue you could also certainly look at a number of stem cell sources and then from that point you have two options in some tissue engineering instances you take that construct and implant it directly into the patient some of the thinking there is that the biological environment is very complex and in general the body functions as a better bioreactor than anything we can construct in the laboratory however if you're forming a tissue that absolutely has to function from the moment you put it in the body and where failure would be a very significant problem then you want to allow new tissue to form outside the body form a tissue that's very highly functional and then implant that back into the person so if I'm making you a coronary artery you definitely want me to take this approach that coronary artery should not be subject to any failure at the point that it goes back into the body so if we look at then what people have been using for these scaffold materials far and away the most common scaffold has been a polyester called polylactic coglycolic acid or PLGA this is a very rigid polymer and people make foams of it like this and then infuse cells through that foam and ideally they grow within that structure and then this polyester slowly degrades over time and that has worked reasonably well in a number of applications but it does have some rather severe limitations for other applications and so if we think back to the anatomy that we just went over we need to be able to control where multiple cell types are able to grow in our construct we need to have a defined intima and a defined media if we have a mixture of endothelial cells and smooth muscle cells rather than this very defined laminated structure the tissue will not work at all so we needed to be able to control what cell types go and what locations we looked at developing alternative materials so the materials that we work with the most are photopolymerized hydrogels where we have a number of different water-soluble polymer precursors here that can have different bioactivity and we can design a material and what type of biofunctions it will have based on what choice of these polymer chains we put into the initial solution so these are all a liquid they're water-soluble and you can mix cells in with this liquid you can then expose it to long wavelength UV light for a few seconds and rapidly convert it from the liquid down to a cross-linked solid and this is a picture of a couple of tubes that we formed this way to use as tissue engineered blood vessels so if we look at what we want to have as far as functionality in these different polymer chains one of the things we want to have is control over which cell types grow in which location and most importantly to control the intima lining and make sure that we grow the endothelial cells there and nothing else now a complication to this is that endothelial cells grow very very slowly relative to smooth muscle cells or the fibroblasts in the adventitia and so if on day zero I make a structure and I put smooth muscle cells in the inside and I put fibroblasts on the outside and I put endothelial cells in the inside and I put it in the incubator to go form a new blood vessel and I come back and I look at it what I find is I've got fibroblasts growing everywhere I've got a few smooth muscle cells scattered about and I'm hard pressed to even find an endothelial cell so if I put that construct into an animal or into a patient it will instantly form a very solid blood clot and it will not function at all so we want to be able to have an area within our construct where the endothelial cells can adhere, grow and thrive form their lining but the other cell types can't enter so to accomplish that we look at how do cells interact with the world around them the way they do this, the way they adhere and grow in locations is that they have receptors on their surface that recognize specific ligands on the extracellular matrix proteins around them and endothelial cells have different sets of receptors than smooth muscle cells or fibroblasts so if we design a material so that the base material is completely cell non-adhesive doesn't allow any cells to grow on it but then we engineer into that the appropriate ligands that can recognize and bind to the receptors that we're targeting then we can choose what cell types are able to grow in that particular location and so we can make within our constructs an area where we've provided the ligands that allow the endothelial cells to grow and other areas where we put different ligands to allow the smooth muscle cells to grow so when we make the materials we'll have a cross-linked network that makes the material solid but we graft in short peptide sequences so basically just short sequences of certain amino acids that are able to interact very specifically with a targeted receptor on the cell that we want to grow in that area and so this is an example looking at cell adhesion onto these materials where in one case we put the correct cell adhesion peptide, RGDS and we see the cells are adherent to the material they've spread out, they're starting to form tissue here but on a material where we flip two of the amino acids we make it RDGS instead of RGDS the cells can't interact in any way with that material none of the cells are attached they're all round little balls that you can just wash off the surface and how much cell attachment you see is dose dependent just based on how much of the peptide you've provided in your material structure so the next thing we wanted to accomplish in these materials was to have what we term biotargeted degradation so we looked at the polyester materials like PLGA and other materials used in tissue engineering most of them have been designed to degrade via hydrolysis so water can come in and cleave bonds in the polymer backbone and you can do things with designing the polymer structure to determine how quickly it will degrade but you need to design that degradation rate essentially before you start your experiment and so one of the problems if you design it so that it degrades too slowly it acts as a mechanical barrier to tissue formation it's basically solid stuff that's in the way of the cells and they can't push on and make their new tissue and if it degrades too quickly then you have nothing you have a pile of mush in the bottom of your petri dish and nothing that resembles a tissue and one of the key problems we've faced is that if we take cells from 20 different individuals in the audience it's a very hard time predicting which people's cells would grow and form tissue the fastest there's some correlation to age but there are many other factors involved that we don't understand so it's hard to predict ahead of time how you should design your biodegradation but if we look at how this happens in your tissue how during things like embryogenesis and wound healing the cells are able to know when to degrade their scaffold material around them sets of proteins called the extracellular matrix things like collagen they are able to regulate this very carefully and only degrade those materials when they're ready and when they're actively forming new tissue and they do that by secreting proteolytic enzymes at one edge of the cell as they're going through and forming new tissue so we know what enzymes are involved in tissue formation by different cell types and so if we can target now synthetic materials to be substrates for those enzymes and to degrade in response to those we can have materials that are customized to degrade at the right rate no matter whose cells we're trying to make a tissue from and so now if we look in the material the degradable sequences are here within the network that makes the material solid if the enzyme comes in and cleaves one of these sequences it breaks the network apart and the material breaks down and so this is an example looking at the degradation we've done a number of studies to confirm that the materials do degrade and that it's specific just to the enzymes we target we also wanted to be able to actually visualize the process happening and so to do that what we did was we made polymers where in the degradation sequence that's the substrate for the enzyme we put fluorescent tags on each side of that degradation site and these fluorescent compounds that we used when they're very close to each other they auto-clench so one molecule quenched the fluorescence of the other and you'll just have no fluorescence but once they separate once you've come in and cleave the site between them and allowed them to move apart from each other then they turn on their fluorescence and you get a very bright green so in this set of studies here looking at the proteolytic cleavage by cells as they're forming new tissue we have fibroblast cells labeled with red in each of these cases and then we're looking at the degradation products in green we can see it's very highly localized it's something that the cells are just actively doing and we can also visualize where the cell has been and where it's degraded material as it's moved along and continued its process of tissue formation so when we look at then taking these and actually fabricating them into graphs we're using photopolymerizations we're going to be exposing them for a few seconds to long wavelength light to make the tubes and to make the three layers we start off making the center layer of the construct the medial layer by making up a polymer formulation designed for smooth muscle cells mixing in the smooth muscle cells putting it in this annular tube shaped mold exposing it to light then we can remove that initial tube from that structure then we take another polymer that's designed to support endothelial cells and we use a process called interfacial photopolymerization to create a thin layer of the endothelial cells and their polymer on the inner surface and then we do that a second time on the outer surface with fiber blasts and a polymer design to support them to create the adventitia around the outside and then at that point we have these nice tubes and as I mentioned we need to make sure that those are highly functional tissue at the point we implant them we wouldn't want to implant them at this point where they're mostly polymer with some cells in there we want to implant them when it's all cells and extracellular matrix proteins that they've secreted and they're organized and ready to function so what we do is we take them into a bioreactor that's designed to support their growth over time and vascular cells are very mechanically responsive so the cells in your blood vessel are used to continually experiencing the shear forces generated by the blood flowing past and also cyclic stretch because with each beat of the heart your blood vessels are stretching and it turns out that both of those types of stimuli are very much responsible for causing the blood vessel cells to assemble into appropriate tissue structures so we've built a bioreactor system that allows us to place those graphs in a flow circuit where we can control the pulsatility in the different conditions and provide the cells with the right mechanical environment to encourage them to form the right type of tissue and so when we do this we can control things like pressure profiles pulse rate how much stretch you get across the wall the shear stress at the surface and this movie here shows one of these constructs growing and you can see the walls stretching and they're stretching in a very physiologically relevant amount and rate and so that's giving the cells within this construct now the right cues to start assembling into an appropriate tissue and so then if we monitor these over time we see that by about day 18 we achieve mechanical properties that are similar to saphenous vein so saphenous vein is currently the gold standard for transplantation that is much weaker than a coronary artery but we know from history that those are strengths and mechanical properties sufficient for function in this application and so we can see we can reach that point within a few weeks and then when we implant these we've done one set of studies in planting these in ceramics we implanted six tissue engineered vascular graphs and after a month all four were patent meaning that blood was flowing very smoothly through them they were open and functioning they were intact so there were no problems with rupture or breakdown of the tissues so we're very excited about that and moving forward okay so at this point I want to shift away from the cardiovascular system and talk about some of our work focusing on cancer so cancer remains a leading cause of death but one of the most disturbing facts around cancer mortality is that if you look at the age adjusted changes in mortality cancer is the one disease that's made no improvement over the past 50 years so at the same time when we have this somewhat bleak news there are also very exciting advances in basic research in cancer such as the genomic analyses that we've heard from Dr. Bishop the understanding of biomarkers that differentiate cancer from normal tissue and many of these are on the brink of being exploited for future therapies and so we can start looking at some of these basic research advances and figuring out how to translate them into therapies that might allow us to really start making a change in the mortality rate here so an approach we've taken to try and look at very specifically and very effectively knocking out cancer while minimizing damage to surrounding normal tissues is to use a process that we call photothermal ablation where we design nanoparticles to very strongly absorb light in the infrared and this is light that just passes harmlessly through normal tissue we can add targeting agents to the surface of these nanoparticles to allow them to specifically recognize and bind to tumor cells we can then inject them into the body allow them to circulate through the body and find the tumor site and then we can apply the infrared light from outside the body it has no effect on the normal tissue but when it hits the nanoparticles down to the tumor cells the particles very rapidly heat up through the absorption of light and that local heating causes destruction of the cancerous cells and this is an image that was taken from the June issue of National Geographic they did a very wonderful spread on nanotechnology and I highly recommend looking at this they talk about many different applications of nanotechnology and many different types of technology they did highlight our work in cancer and made this nice cartoon that I would never be able to recreate myself but the idea here is you have these particles now circulating through the body dramatically smaller than the human cells that they're designed to target but when they recognize these regions on the cancer cell that are unique to the cancer cell and not to the normal cell they bind there when you come in with the infrared light as we see here then these particles become activated cause the local heating whereas particles outside that region are causing no problem so there are many types of nanoparticles in medicine the definition of a nanoparticle is a particle under a hundred nanometers in size so nanometer is a billionth of a meter we're talking about technologies that are two to three orders of magnitude smaller than the micro technologies used on most computer chips interestingly many of these materials when we control their dimensions at this size scale give us very unique and special properties often times because we're talking about constructing materials with dimensions smaller than the wavelengths of light they give us very unique optical properties such as the fluorescence we see here in materials called quantum dots or bright colors from different metal colloids and so often times in the popular press when people talk about nanotechnology and medicine they envision something along the lines of this where you have a robot you've injected into the body that can circulate around and turn to cells, repair your DNA talk with its friends send information back to you if you believe Michael Crichton gang up with each other and take over the world so this is definitely science fiction the very cutting edge of technology today is the ability to simply control a dimension at the nanometer scale the ability to make robots where we can communicate send instructions, get information back is still a long way off and I don't think that we have to worry about these types of materials ganging up and taking over the world any more than we should worry about empty coat cans doing the same so the particular nanomaterials that we've been working with are called nanoshells these were invented at Rice about 10 years ago by Naomi Hallis and these are materials that are basically the structure of a malted milk ball but much much smaller so they're a core and a shell where the inside part of the malted milk ball is just anything that doesn't conduct electricity and for the studies I'll talk about today we've used glass for that material and then around that we create a very thin coating that now instead of being chocolate is a very thin coating of a metal and for biomedical applications we make that be gold because it has good biocompatibility and we can easily bind our targeting agents to it now the thing that's special about these materials is that by changing the design of the nanoparticle the size and composition of each of the two layers we have an optically tunable particle where as shown here we can shift where the absorption peaks for the material occur just by changing the structure so for these examples here we have four batches of nanoshells that all have the same size core nanoparticle but we grew different thicknesses of shells ranging from a 20 nanometer thick shell down to a 5 nanometer thick shell and then we shift the optical properties around and we can extend that over a much larger range by also changing the size of the core and by changing compositions and just to give you some idea of what those peaks really then translate into these are batches of nanoshells where again as we move from left to right we have progressively thinner shells and so we go across the visible and then out into the infrared regions of the spectrum and one of the things that was very readily apparent to us with this was that we had the ability to make materials with their optical properties in an area called the near infrared and that's an area of the spectrum between about 650 and 900 nanometers where you have very deep penetration of light through tissue and this is because you're above the regions where you have absorption of light by chromophores such as hemoglobin or melanin but you're below where water starts to absorb light so in this region here you have very deep penetration of light through tissue, depths over 15 centimeters and so that would create a lot of interest for being able to either image non-invasively or perform therapies non-invasively since you can send the light in from outside and the limitation has been the ability to have materials that give you strong optical properties in this region without giving you toxicity but now with these nanoparticles we can design them so that their optical absorption is very strong in this region and that they have the appropriate properties for whatever application we're trying to develop so to make these we first make the core nanoparticle and then we grow the gold shell around the outside so this is a TEM image of the silica cores we form we react to them with a reagent that puts positively charged amine groups on the outer surface and we use those to absorb very small, negatively charged gold colloid onto the surface of the silica particle and we use those small dots of gold as nucleation sites so that we can reduce more gold and basically grow our shell and that process is shown in this series of TEM images where we start off with the gold colloid absorbed to the nano shell surface and we progressively are growing those they're getting larger, coalescing and finally forming a complete shell and fortunately the optical properties of these materials are very well predicted by something called Mie scattering theory and that allows us to computationally predict what size and composition we need for our final nano shell material to achieve whatever optical properties we need for an application and then we can set up our chemical reactions and stoichiometries to just make that happen so as we look at trying to see whether or not these will be effective in cancer therapy we've done animal studies where we grow human tumors within mice, we're able to inject the nano shells intravenously, allow them to circulate through the body find and bind to the tumor sites and then after that's happened we apply light to the animal from outside and then monitor the efficacy of the therapy so first this is an MRI image taken in collaboration with John Hazelet and the Anderson Cancer Center this mouse has two tumors one about here and one here and after injecting nano shells as we apply the near infrared light to the animal up in this direction if the pointer follows we were able to do MR thermal imaging to allow us to monitor the temperature profiles in the tissue hopefully there we go so as we see the color changing we see regions where we've achieved a change in temperature of at least 15 degrees C which is sufficient to cause irreversible damage to the tumors we see that the normal tissue in between the two sites doesn't heat up and isn't harmed by the process so when we take these animals then and monitor the efficacy of the therapy we first look at changes in tumor size and we start off on the day we're doing the treatment with three groups in green we have a group that receives a nano shell injection in treatment with the laser we have then a group that has the laser treatment but was injected just with saline solution no nano shells and then in blue a completely untreated control group and at the start of the study they all have basically the same size tumors but by day 10 what we saw was complete regression of all tumors in nano shell treated mice there was no detectable tumor tissue left in any of those animals whereas the two control groups continued to grow very rapidly and then when we look at survival of these animals what we found were that the two control groups all of the animals perished due to excessive tumor growth within three weeks of treatment whereas the nano shell treated mice continued to live tumor free for a year following the treatment date we saw no regrowth of any tumor tissue in any of these animals so we're also working with Dr. Rebecca Dresik at RICE looking at applying these materials as contrast agents for optical imaging there's also been a lot of interest in near infrared light for imaging it's been used for instance both in breast and brain imaging the issue has been you're looking for very subtle differences in the optical properties between the diseased tissue and the normal tissue and so it's hard to discriminate between the two the way you normally deal with this in imaging technologies is to use substances called contrast agents that will accumulate at specific sites and cause the signal there to go up the issue in your infrared imaging again has been that there have been very few non-toxic compounds that you could use for this so now the hope is that nano shells can do that and so for therapy as we've talked about so far we designed the nano shells so that almost all of the light is absorbed so that it's converted to heat and can kill cells for imaging we actually want the opposite scenario where instead of being absorbed most of the light is scattered so it will come back out of the tissue and you can use it to reconstruct an image of what was happening inside the body so we can design nano shells to do either thing by adjusting their structure so this is an example looking at breast carcinoma cells that bear a marker called HER2 looking at nano shells targeted to recognize that marker we see the cells light up very vividly when we treat them with these nano shells and we have high enough resolution based on the scattered light to make out individual cancer cells and without the application of nano shells we see a completely black screen with nothing discernible from the information so we're very excited we think that can really give a lot of information about what molecular markers are present in a tumor and really provide more advanced diagnostic information we were also struck by the possibility of integrating imaging and therapy as a single approach in cancer so looking at using nano shells now designed to be 50% absorbing 50% scattering to hopefully accomplish both tasks sending back enough light to generate an image absorbing enough light to induce heating and cause destruction of the cancer so here we're looking at breast carcinoma with anti HER2 again and wanted to look at whether or not we could accomplish both objectives so in cell culture first we have three conditions where the first two are controls and the third column is our appropriately targeted nano shells we see that we can nicely light up the cells based on scattered light we can see what's happening this is at a lower magnification and then we come back and we expose them to higher intensity light and we see that within the laser beam we're able to kill almost all of the tumor cells due to the heating caused by the near infrared light so with this single material we're able to have a multifunctional platform that can accomplish both tasks and as we look at moving this into animal studies we do similar studies with tumors grown in mice inject the nano shells, allow them to find the tumors but now before doing the treatment we add a step where we have a fiber optic probe for a technique called optical coherence tomography we can place in contact with the skin we just shave to the hair and put a little jelly on the skin to allow the probe to slide we generate our image first and then we come back and we do our treatment and so when we do this these are first looking at the images if you look at an animal where you've done a saline injection and then you image by optical coherence tomography which is clinically used you see a statistically significant change in the optical properties and contrast between normal tissue in blue and cancerous tissue in black so it is statistically significant sometimes in science is the benchmark that you're shooting for but it's actually if you look at the images relatively difficult to discern the differences between the normal here in panel A and the tumor tissue in panel C but if we look at an animal that received nano shells we see a much more dramatic difference between the normal in blue and the tumor tissue in black and even just glancing at the images there's a striking difference between panels B and D that would be easy to pick up and discern and so we feel this is providing much better tumor contrast and then going on and looking at the effectiveness in therapy if we monitor changes in size we see a dramatic reduction in tumor size by day 12 so this isn't complete regression in all of the animals in this case it was in some of the animals still a very good result here and then when we look at long term survival we see a very this top line here is the nanoshell treatment we see a very profound increase in long term survival of the animals we're currently looking at what we can do to shift this study was done with 50% absorption 50% scattering but if we adjust that 60, 40, 70, 30 how far can we go and still get very wonderful images with single cell resolution and very good discrimination of tumor versus normal but get 100% efficacy if it's possible so we're excited and moving forward with that we feel one of the key places where this could have a big impact is in treatment of cervical cancer where there's a lot of problems with following up patients after an abnormal pap smear who live in socio-economically disadvantaged conditions or in third world countries we think that this could be an option to work around some of those difficulties so the last technology I want to mention quickly is one that we've been working on to try and rapidly detect compounds in the blood so immuno assays things such as techniques called ELISAS are used to detect proteins, bacteria and viruses and biological samples most of them require many hours to several days to perform and most of them require optically transparent samples such as these so for instance if you're drawing blood you have to spin out all of the red blood cells before you proceed with your test but if you could have an assay that instead of taking hours to days gave you results from blood samples within minutes you would have the ability to screen large populations quickly to provide results within an emergency setting it could really alter some of the way medicine is currently practiced and so an approach we've taken to this is to develop something called a near infrared ramen biosensor so just briefly ramen scattering has to do with just whether or not the light that comes back after a scattering event is at the same wavelength or if it's been slightly altered so when a scattering event happens the vast majority of the light scatters elastically comes back at the same wavelength and a tiny fraction will have a characteristic shift depending on what molecules were involved in the scattering events and that shift is referred to as the ramen signature and so you could use that to tell you about what molecules were present and in what quantity the problem is that this ramen signal is very small and in a complex biological environment it's usually difficult to impossible to detect it however there's a phenomena referred to as surface enhanced ramen scattering which is still actually a very poorly understood phenomena but at the same point it is being now exploited where it's known that if you have molecules very close to the surface of a metal material you can have enormous enhancements million to billion fold enhancements in the amount of ramen scattering so now instead of being a tiny signal it becomes a very large signal that's easy to see but now to get the SIRS effect to really happen well you have to have the wavelength of light that you put into your system match what's called the plasma resonance wavelength of your material that's basically just where the peak optical properties of that material occur and so for most metals they're the visible or UV regions of the spectrum which aren't going to be useful if you're trying to detect compounds in blood or tissue but now because we can make metal nanoshells that have their plasma resonance in the near infrared we can move this to a set of wavelengths where the biological fluids and blood and tissue are essentially transparent and so to develop an assay make the nanoshells so that we have their plasma resonance right in the near infrared these examples at 1064 nanometers these are highly scattering we place an antibody on the surface so this is a protein associated with your immune system that recognizes and binds to one specific agent it may be a specific part of a virus or bacteria and then we put that into our sample if the appropriate compound is present then that's able to bind to that antibody we mix in a second antibody that will also bind there and now has a very unique tag on it that gives us a Raman signal that's very different from those associated with proteins because one of the issues is that if we look at the biomolecules involved here they're all made out of the same set of amino acids so it's fairly difficult to detect the difference in signal between the antibody and the bacteria or virus but now we put on this unique tag that gives us a signal somewhere else and we come in we put in our laser light at 1064 nanometers and then we detect the light that gets scattered and we look at the light that specifically has been shifted in wavelength so we put the antibodies on the surface using basically polymer linkers we also have to come in and add another polymer to the surface a compound called polyethylene glycol that prevents any non-specific interactions with the surface of the nanoshell so that we're only looking at the very specific antibody antigen interactions and once we do that we look at how much antibody we get there and how much is active we get very high coverage with antibodies and we retain greater than 60% activity of the antibodies and we can look at that using a dual enzyme label where we put one enzyme on the antibody a different enzyme on the analyte we can add reagents that change color with one or the other and look at their relative activities and so that gives us some of these numbers but we can also look at direct visualization so this is a nanoshell here where we've conjugated antibodies to the surface that we tagged with small gold colloid dots so that we can see them under the electron microscope and see that they're covering the surface very nicely so when we look at the SIRS signal the Raman signal from binding events on the surface of the nanoshell we start off detecting the presence of a specific compound and looking just at how changes in the concentration of that compound related to the Raman intensity at the peak we were monitoring so we see a very linear correlation over most of the region and once we've basically covered all of the sites on the nanoshells we hit a plateau region if we need to be able to detect higher levels we add more nanoshells but generally the issue is with making sure that you have low enough detection thresholds and we've been at sub-nanogram per mill detection limits which are very much competitive with other amino assay technologies the difference here now is this is done in minutes without any preparation steps so to confirm that we can translate that into biologically relevant fluids we looked at a series of fluids one in phosphate buffered saline so basically just the analyte we're trying to detect in a clear completely plain solution then one in bovine serum so this is looking at all the components of blood except for the cells looking at whether or not any of those proteins and lipids would interfere with the signal and then the last one looking in whole blood there's nothing different and so in each of these cases the peak we're using to quantify is at 1140 right here and as we move from saline to serum we see absolutely no change when we move to whole blood we see a slight attenuation in the signal due to the scattering of light from the surface of cells but we still have a very quantifiable signal and the attenuation is very minimal and we can still detect sub nanogram per mil quantities of analyte and this detection from putting the needle into the arm withdrawing the blood to having an answer about whether or not the bacteria or virus you're concerned about was present is under five minutes and so if you were trying to do screening in an emergency room this would be very easy also we have our senior engineering students do design projects two years ago their challenge was to make portable battery operated hand held systems that would allow you to do these measurements and to make them for under $200 they had to fit in a backpack so that they could end within a certain cubby in the back of an ambulance and all eight teams made functioning devices that were able to do these measurements and for under $200 so I think that so I think part of this message is not all technology will drive up the cost of health care so to conclude I think that the coming decade is going to offer many tremendous advances in medicine we've seen just huge changes in the basic research environment the elucidation of the human genome to really understand its meaning the understanding of stem cell biology and we're seeing now the translation of these advances into medical care and my message to the students there's still much work to be done I hope you're excited about this willing to roll up your sleeves and join us in the effort and with that I'd like to thank a number of people involved in the work especially my collaborators Naomi Hallis Rebecca Drezek and Karen Hershey all students and postdoctoral fellows who have contributed to the work you've seen and our research funding I'll be happy to take questions