A12 2120 TARGETING THE SOURCE: THE VALUE OF BIODEGRADABLE NANOPARTICLES FOR CANCER TREATMENT Tyler Alan (tra17@pitt.edu), Justin Blazer (jtb68@pitt.edu) importance of scientists and engineers working to develop new methods of treating this terrible disease. The development of successful new cancer treatments will not only save families the great emotional and economic hardship that cancer can bring upon them, but also save society billions of dollars that are lost each year when people in the prime of their careers are incapacitated by cancer. One of the promising new technologies for cancer treatment is the use of biodegradable nanoparticles. These particles directly target cancer cells and deliver cancer fighting drugs or imaging agents directly to the affected area. These molecules make use of abnormalities in cancer tissue to successfully accumulate and linger in cancer tumors and greatly increase the efficiency of therapeutic agents. The use of biodegradable nanoparticles promises to make both the treatment and imaging of tumors more effective, while minimizing harm to surrounding healthy cells. Biodegradable nanoparticles are tiny particles that consist of a desired agent that is encapsulated on or within a biodegradable polymer, the polymer protects the agent and aids in the delivery and controlled release of the agent. The agent encapsulated on or within the nanoparticle can be either a specific gene, cancer-fighting drug which can fight the tumor cells, or a chemical that aids in the effective imaging of tumors. The protective biodegradable polymer can be broken down by the body into its individual monomer units because they are of a type found in the body naturally due to metabolic processes. The microscopic size of nanoparticles is beneficial because it allows them to be taken up directly into cells through endocytosis (the process by which cells absorb molecules by engulfing them). All of the properties mentioned above will allow biodegradable nanoparticles to improve current cancer treatment. PLGA, Poly(lactic-co-glycolic acid), is a specific type of biodegradable polymer that is Federal Drug Administration (FDA) approved and commonly used in the synthesis of biodegradable nanoparticles. It is a versatile polymer that can be used to encapsulate various agents that aid in cancer treatment. The versatility of PLGA allows it to serve as a perfect example from which various characteristics and properties of biodegradable nanoparticles can be explored. As with all engineering and medical applications, there are ethical codes that must be followed while conducting research to ensure the safety of patients who may eventually receive treatment with biodegradable nanoparticles. Biodegradable nanoparticles show great promise for the treatment of cancer. They will allow tumors to be effectively targeted, treated, and imaged while minimizing the harmful side effects to healthy cells. Engineers must pursue this Abstract - Humanity faces a wide variety of diseases and physical ailments. These conditions can be treated much more effectively if medications are delivered to specific regions of the body. Biodegradable nanoparticles will make this possible. These particles show great promise in creating more effective methods of treating cancer. One of the main problems with current chemotherapy treatment is that it damages the patient’s healthy cells along with the cancer cells. This collateral damage causes many of the adverse side effects that make cancer treatment such a painful process for patients. Biodegradable nanoparticles can be designed to specifically target the cancer cells while minimizing harm to healthy cells making them an attractive option for future cancer treatment. The application of these particles to tumor imaging will be described by outlining the design features that allow them to be attracted by tumors and clearly imaged. The ethical codes that must be followed as this technology is pursued will be noted and discussed. This conference paper will describe in detail the new technology associated with the use of biodegradable nanoparticles for the targeted delivery and controlled release of chemotherapy drugs as well as its application to tumor imaging. The significance of the technology, ethical concerns, and the current research and development being conducted on biodegradable nanoparticles for cancer treatment will be outlined. Key Words – Biodegradable nanoparticles, Cancer, Chemotherapy, PLGA, Ethics, Tumor imaging THE FIGHT AGAINST CANCER For the past 30 years technological breakthroughs have inundated society. They have made their way into every aspect of our lives changing how we operate on a day-to-day basis. The most important breakthroughs have occurred in the medical field and transformed healthcare into an industry that operates with incredible precision and efficiency. Countless lives have been saved or made better by these technological advances. Despite these breakthroughs, the healthcare industry still faces many challenges in the treatment of certain diseases, specifically the treatment of cancer. Currently, a cancer diagnosis remains synonymous with a painful period of treatment that may inevitably lead to one’s death. In America approximately 1500 people die from cancer and over 3400 are diagnosed daily. During 2010 cancer cost society an estimated $263.8 billion between medical expenses and lost potential earnings due to missed work and early death [1]. These facts emphasize the University of Pittsburgh Swanson School of Engineering 1 February 10, 2012 Tyler Alan Justin Blazer technology due to the potential impact it has for the millions of individuals who suffer from cancer. nanoparticles are created with specific surface modifications that allow them to pass through the body without being tagged as foreign by the immune system and reach the cancer tumors. Polymers are typically added to the surface of the molecules in order to make them hydrophilic and to allow them to avoid being tagged as foreign and attacked [4]. Once the immune system is bypassed, the tiny size of these molecules allows them to travel through organ membranes and be taken up by cancer cells through endocytosis. Molecules of larger size cannot achieve similar maneuverability through various body tissues [3]. This is a major benefit of nanoparticles because they can protect the therapeutic agent all the way until it is delivered to the tumor. Other methods of treatment are not be able to protect the agent, which results in some of the agent not reaching the source, or acting on healthy cells as well as the tumor cells. Once inside of the tumor cells, the biodegradable polymer is broken down and the encapsulated agent is released over a controlled period of time. The rate of release is determined by the composition and molecular properties of the nanoparticle. The ratio of drug to carrier, molecular weight, and the ratio of monomer units of the polymer all affect the rate of release. Variances in composition can allow the drug to be released over a period of several days, or several weeks. In the case of PLGA, a 50:50 ratio of lactic to glycolic acid yields the fastest drug release because at this ratio the polymer is hydrolyzed quickest once at the target. The ability of biodegradable nanoparticles to be designed to release drugs at optimum rates is extremely valuable, and is a defining characteristic of the biodegradable nanoparticles. This trait will allow for drug concentrations to be maintained at the proper level so that treatment is effective and adverse side effects are minimized. BENEFITS OF BIODEGRADABLE NANOPARTICLES FOR D RUG DELIVERY Currently the primary methods of cancer treatment are surgery, chemotherapy, and radiation treatment. All of these which side effects such as pain, fatigue, nausea, anemia, and second-hand cancer later in life [2]. These side effects can be minimized, and in some cases eliminated, through the use of biodegradable nanoparticles due to their ability to treat cancer directly at its source while leaving healthy cells relatively undisturbed. Current advances in research and clinical applications of PLGA-based nanotechnology notes this issue and also states another consequence of chemotherapeutic agents, “Chemotherapeutic agents also damage healthy tissues, leading to systemic toxicity and adverse effects that greatly limit the maximal allowable dose of anticancer drugs and, thus, restricts their therapeutic efficacy [3].” Biodegradable nanoparticles have the potential to greatly increase the effect of both chemotherapeutic and other cancer fighting agents by delivering them directly to tumors. Delivering the agents directly to the tumor will reduce side effects and allow the prescription of larger doses than previously possible. The ability to prescribe larger doses of therapeutic agents could greatly increase the effectiveness with which cancer tumors can be treated. In addition to their ability to allow the delivery of anticancer agents in larger doses, biodegradable nanoparticles can be constructed to release the encapsulated drug at a desired rate. By controlling the rate at which the drugs are released cancer therapy can be improved by maintaining consistent amounts of cancer fighting drugs in tumors to maximize the effect of the therapy. Controlled release can also reduce the frequency at which therapeutic injections need to be delivered. Controlled release will make treatment more pleasant for patients because they will not need to receive as many injections, and will also maintain consistent drug concentrations in the tumors so that they are constantly combatted between a patients’ appointments. Cell specific targeting and accumulation Biodegradable nanoparticles are designed with moieties, which target and cause the molecules to be attracted to receptors on cancer cells. A moiety is a functional group of a molecule. Target moieties take advantage of the fact that tumor targeting “can be enhanced by associating the drugs with molecules that bind to antigens or receptors that are either uniquely expressed or overexpressed on target cells compared with normal tissues [3].” Target moieties allow biodegradable nanoparticles to take advantage of the differences in receptors and antigens in cancer cells to turn the molecules into miniature “homing missiles.” Target moieties direct the biodegradable nanoparticles to the tumors and allow them to accumulate in much higher concentrations in the tumor tissue than in healthy tissue [4]. This accumulation is crucial due to nanoparticles small size and individual ability to carry only small amounts of the desired agent. Another key to biodegradable nanoparticles success is their ability to take advantage of the chaotic and disorganized blood vessel structure inside of tumors. Unlike Tricking the Immune System to Reach the Tumors and Deliver the Payload The body’s immune system is designed to attack particles with similar surface properties to those of biodegradable nanoparticles. Without modification, the surfaces of nanoparticles are negatively charged and hydrophobic. Macrophages, white blood cells within tissues that digest debris and pathogens, see particles of this type as foreign and thus eliminate them through phagocytosis (phagocytosis is the process by which cells engulf and digest particles, in this case particles tagged as foreign). In order to prevent phagocytosis by the macrophages, biodegradable 2 Tyler Alan Justin Blazer normal tissues with organized, logical vascular structure, cancer tumors have illogical, randomly branched, dilated vascular structure. The excess dilation of tumor vessels allows them to be infiltrated readily by nanoparticles because the dilation increases their permeability. Once the nanoparticles have entered the tumor vessels they are cleared much slower than they would be by normal cells due to the chaotic and disorganized vascular structure [3]. This gives the nanoparticles a longer time frame to release their drug load, or increase the ability for the tumor to be effectively imaged. acids that occur naturally in the body due to metabolic processes and the body can easily eliminate them through the Krebs cycle. The Synthesis PGLA based Biodegradable Nanoparticles PGLA based nanoparticles are commonly synthesized using the emulsion/solvent evaporation technique. The emulsion technique of polymerization involves making an aqueous solution of the monomer units of the polymer and adding initiator molecules, which cause monomer units to attach to one another as they collide in the solution. The solvent can then be evaporated leaving behind the nanoparticles. The size of the nanoparticles is influenced by temperature, stirring rate, and presence and concentration of various chemicals during the solvent evaporation process. Typically the molecules range in size from about 10 to 100nm. Though the average size of the particles can be influenced, there are still differences in particle size due to random variation. THE CREATION OF EFFECTIVE PGLA BASED BIODEGRADABLE NANOPARTICLES The production of biodegradable nanoparticles is a complex process consisting of several steps. A large part of the hindrance in their development and release for widespread public use is the difficulty in manufacturing them. There is a vast array of nanoparticles that are currently being researched but the most common biodegradable nanoparticles are based off of a PLGA polymer. Poly(lacticco-glycolic acid) is currently FDA approved for use on humans, and is an incredibly versatile molecule which shows promise in all areas of cancer treatment from diagnostic imaging to drug delivery. FIGURE 1 IMAGE SHOWING THE SIZE OF PLGA BASED NANOPARTICLES [3] Figure 1 shows the variation of size in a batch of PLGA based nanoparticles. The emulsion/solvent evaporation method serves as an effective means of combining drugs with the base polymer to create a useful biodegradable nanoparticle [4]. The two different methods of emulsion that are used to create nanoparticles are single and double emulsion. The single emulsion process is best for encapsulating drugs that are insoluble in water. In the single emulsion process PGLA is first dissolved in a volatile organic solvent with the desired drug agent. This mixture is then added to water and the emulsion process is conducted under the desired temperature and stirring conditions. After the emulsion process the solvent and water are removed either by evaporation or extraction. The final mixture is then centrifuged or filtered to remove the nanoparticles that are desired for use [3]. The double emulsion process is best suited for encapsulating drugs that are hydrophilic. While single emulsion was a water-in-oil process, double emulsion is a water-in-oil-in-water process. In this process an aqueous FIGURE 2 FIGURE SHOWING THE BASIC CHEMICAL STRUCTURE OF THE BIODEGRADABLE PLGA POLYMER [5] PLGA is a biodegradable polymer made up of monomer units of lactic and glycolic acid. Figure 2 shows the basic chemical structure of a small section of a PLGA polymer. There is a hydroxyl group on the left side of the molecule in the diagram and a lone hydrogen on the right side. These are the sites where other monomer units can attach to the PLGA molecule by dehydration synthesis. When dehydration synthesis occurs the hydroxyl group and lone hydrogen are replaced by a peptide bond and a water molecule is given off. This method of bonding between molecules is useful in biodegradable nanoparticles because it can be reversed in the body to break down the polymer. The body adds a water molecule to the bonding site, which breaks the peptide bond and hydrolyzes the polymer into its monomer units once it has reached the target tumor. Both of the monomer units are 3 Tyler Alan Justin Blazer solution of the desired drug is emulsified in an oil based organic solution that contains the desired polymer. This water and oil emulsion is then again emulsified in another water-based solution containing a stabilizing agent. This makes double emulsion a water-in-oil in water process. Beyond this step double emulsion is identical to the single emulsion process in the solvent removal and particle retrieval steps [6]. quantum dots can be safely used without poisoning the human body. Production methods have been developed that prepare-folate decorated nanoparticles of biodegradable polymers, such as PLGA, for quantum dot formulation. This recent research has lead to improvements in targeted and sustained imaging. These developments will allow tumors to be imaged more effectively because the quantum dots will accumulate in the tumors due to their attractive properties and allow them to clearly show up in images. This will lead to an increase in the number of cancer diagnoses in the early stages. BIODEGRADABLE NANOPARTICLES APPLICATION TO TUMOR IMAGING The first step in the fight against cancer is tumor detection. Without this step, cancer cannot be treated at all and will likely go unnoticed until the patient begins to feel physical symptoms. Once the patient feels these symptoms, the cancer may be to far along in its development to be effectively treated. The most important aspect of detection is early detection. The earlier that cancer is detected the greater a patient’s chances are for surviving the disease. If cancer is diagnosed at an early stage, by regular precautionary screenings, the 5-year survival rate for cancer could increase to 95% [7]. Considering that the current 5-year survival rate is only 68%, this fact demonstrates the impact that discovering cancer tumors in their early stages has on a patient’s chances of survival. When the tumor is found after it has metastasized, current treatment methods involve infusions of toxic chemicals, and blasts of radiation. These treatments are detrimental to the physical health of the individual and cause cancer treatment to be incredibly strenuous. When cancer is found in the late stages, the prognosis is typically just as miserable as the treatment experience [8]. Early detection is extremely important for treatment to be less arduous and more successful. It greatly increases the probability that patients will avoid reaching the fourth stage of the cancer cycle. Figure 3 gives a graphical representation of how large of an impact early detection of cancer has on patients; survival rates. If cancer is found in stage one the 5-year survival rate is above 50% for most types of cancers and is nearly 90% for other types. After this stage the survival rate drops continually as the stages increase. Biodegradable nanoparticles can be used in order to detect cancer cells in the earlier stages of cancer. One way that biodegradable nanoparticles are able to improve the imaging of cancer cells involves the use of quantum dots. Quantum dots are widely studied luminescence probes that have many advantages in the modern day field of medical imaging. They have beneficial optical and chemical properties that include tunable emissions, broad excitation spectra, high quantum yield of fluorescence, strong brightness, photostability, and high resistance to photobleaching. Recent research has yielded fantastic results that that has ridden quantum dots of the toxic features that previously plagued their reputation. The newly developed FIGURE 3 THIS CHART DISPLAYS THE IMPORTANCE OF EARLY DETECTION [7] Developing Biodegradable Nanoparticles with Quantum Dots With the knowledge that quantum dots have the potential to detect cancer more effectively at early stages when combined with biodegradable nanoparticles, researchers have begun to develop numerous methods of creating such particles. One method begins with the synthesis of TPGSCOOH, TPGS being (D-α-tocopheryl polyethylene glycol 1000 succinate), and folate-NH2. This is used to help in the formulation of quantum dot loaded nanoparticles with folate decoration and free quantum dots. The newly formulated particles undergo a series of technical steps, including sonication, (sonification is a process that uses sound waves to evenly disperse nanoparticles in solution) and eventually become mercaptoaceric acid coated quantum dots. The aforementioned process created particles that after testing confirmed the original objective of increasing specificity and sensitivity of quantum dots image by labeling various cancer cells with folate receptors on the surface. The key to the success of these particles is the folate receptors on the surface of the cancer cells which the biodegradable nanoparticles are designed to be attracted to. This allows these particles to accumulate around the tumor cells so they can be effectively imaged. 4 Tyler Alan Justin Blazer What This Means For The Fight Against Cancer Potential Uses In Ultrasound Imaging It is clear that biodegradable nanoparticles are capable of being produced so that they can not only detect cancer cells, but also so that they are capable of enhancing the medical images of the cancer cells they target. Both of these properties of biodegradable nanoparticles can drastically change the way in which cancer is treated. Biodegradable nanoparticles will increase the frequency of cancer detection in its earliest stage. Due to biodegradable nanoparticles’ ability to be designed with specific surface modifications, they can be made to specifically target the cells in cancer tumors. When large concentrations of the nanoparticles carrying imaging agents reach the tumors that they are made to detect it will be easier for doctors to notice the tumors in medical images due to the high concentrations of imaging agents that they will deliver to the tumor sites. This enhanced imaging will increase the rate of early detection, and therefore also increase the chances of survival for millions of future cancer patients. The clear results of research on the use of biodegradable nanoparticles to improve cancer imaging further shows the great impact that these particles can have on cancer treatment as a whole. Early detection is extremely important in the fight against cancer. The further development of biodegradable nanoparticles can vastly increase the rate at which cancer is discovered in its early stages. The faster that cancer can be treated, the greater a patient’s chances of survival are. Biodegradable nanoparticles give cancer patients the upper hand in the fight against the disease. One of the most well-known and publicized forms of cancer today is breast cancer. It affects millions of women every year. For this reason breast cancer research receives incredible financial support from groups like the Susan G. Komen for the Cure foundation. When conducting precautionary scans for Breast Cancer it is very common for doctors to use ultrasound imaging. In recent studies it has been shown that ultrasound imaging along with mammography still prove to be insufficient in detecting breast cancer in its earliest stages [9]. Early detection is a crucial element in the fight against cancer. If ultrasound imaging can be improved to increase the ability for doctors to detect breast cancer earlier in its development and with greater success, the probability of surviving breast cancer can be significantly increased. Biodegradable nanoparticles are the solution to increasing early detection with ultrasound imaging. Research has shown them to be capable of enhancing the images produced by ultrasound. They accomplish this by targeting tumors, and delivering chemicals that are easily detected by ultrasound scanners to them. Figure 4 displays the extent to which nanoparticles are capable of enhancing an ultrasound image so that target of the imaging can be seen more clearly. This will reduce the chance of a cancer tumor being over looked in an image. The particle on the left of Figure 4 could easily be overlooked, as it is not very bright and could easily blend in with other structures within the body. ETHICAL CONCERNS W ITH THIS RESEARCH Biodegradable nanoparticles are a promising technology for the future and have the potential to completely transform the healthcare industry with regards to cancer treatment. Researchers from all parts of the world have begun research to show the capabilities of biodegradable nanoparticles. To date great advances have been made to bring this technology closer to the hospital room, but the technology is still incomplete. There isn’t yet a significant amount of research stating that biodegradable nanoparticles are ready for use in human subjects. This is obviously the most important step, and a step that brings about the ethical concerns associated with biodegradable nanoparticles. The American Institute of Chemical Engineers (AIChE) provides a list of ethical codes that should be followed by their members and chemical engineers in general. One of these goals states that members should “hold paramount the safety, health and welfare of the public and protect the environment in the performance of their professional duties [11].” Research and development of biodegradable nanoparticles is directly linked to this stated code. Chemical engineers should pursue nanoparticle technology because the technology has the potential to improve the health of the public at large, as is shown in the imaging section. Cancer is not going to go away on its own, FIGURE 4 COMPARING IMAGING WITHOUT NANOPARTICLES TO IMAGING WITH NANOPARTICLES [10] Both of the samples that are shown contain the same number and concentration of cells, and were deposited on the same piece of gel. Figure 4 displays the obvious difference between the untreated and nanoparticle-treated samples after ultrasound viewing. These nanoparticles were produced with polylactic acid, and then had Anti-Her2 monoclonal antibody Herceptin covalently linked to the polylactic acid nanoparticle using a carbodiimide technique. This altered the surface of the particle accounts for the difference in imaging results. This is concrete evidence that polymeric nanoparticle agents are capable of targeting specific cells, and marking them so that they display more clearly in ultrasound images. 5 Tyler Alan Justin Blazer and if chemical engineers want to truly hold paramount the health and safety of the public then they must pursue nanoparticle technology. This said, it is also essential that this technology is not rushed into use. If biodegradable nanoparticles are used in humans prematurely and fail, the consequences could be great. For example, patients could be severely harmed if large doses of chemotherapeutic drugs are administered via encapsulation in nanoparticles and the nanoparticles fail. This situation may occur if a doctor prescribes larger doses of drugs than previously used for treatment due to biodegradable nanoparticles presumed ability to deliver the drugs to tumors while leaving healthy cells unharmed. If the nanoparticles fail and the drug is released into healthy cells the patient could suffer great physical harm. Engineers must ensure that this product is completely ready before it is released to the public due to the terrible outcome that could result if the product fails. Another equally important code of ethics that chemical engineers must follow states that they must ‘perform professional services only in areas of competence [11].” The meaning of this code applies to the pursuit of nanoparticle technology in a different way than the previous code. As was mentioned earlier, some of the original materials used in the creation of biodegradable nanoparticles are toxic before they undergo the development and synthesis processes. Engineers must be competent when developing these nanoparticles for use on human subjects because an error in the development process can result in long-term side effects, or even death in human subjects. Additionally, the biodegradable particles that are developed must actually be biodegradable and parts must be able to be easily eliminated from the body. This code is extremely important for engineers to follow because when engineers make mistakes, the consequences are often severe and involve injury or death. Engineering applications, such as the development of biodegradable nanoparticles, that are applied directly within the human body, must be performed by engineers that know exactly what they are doing so that people are not harmed. It is of the utmost importance that engineers continuously pursue this technology. The potential to save the lives of millions of current and future cancer patients makes it unethical to put off the research and development of biodegradable nanoparticles. Only the most competent researchers and engineers should focus on this technology. Incompetence can lead to faulty particles that could ruin millions of lives instead of saving them. The AIChE created a code of ethics with a greater understanding of how the engineering world operates and how society is affected by the work of engineers. It is the ethical responsibility of engineers to heed these codes and continue to develop new technologies that will solve the world’s many problems. Applications such as the development of biodegradable nanoparticles that can directly save lives and reduce the immense hardship the cancer places on society are among the most noble. If engineers follow all applicable ethical codes when pursuing this technology, they can have a profound impact on the treatment of cancer. HOPE FOR THE FUTURE Thanks to many technological advances in the treatment of cancer, it is no longer viewed as a death sentence as it once was. Current research concerning biodegradable nanoparticles has great potential to increase the survival rate of cancer even further in the future. Biodegradable nanoparticles application to cancer treatment could drastically increase the survival rate of those diagnosed with cancer and also make the treatment process more bearable for cancer patients. Biodegradable nanoparticles’ ability to navigate around the immune system and deliver drugs directly to tumors can allow them to be an incredibly effective means of treating cancer in the future. No other current method of drug delivery is able to direct the drug to the proper location in the body and protect it until it reaches the target like biodegradable nanoparticles can. Their production is an intricate process that relies on complex chemistry. Hopefully this production process will soon be mastered so these nanoparticles can be mass-produced and used to better the process of cancer treatment. In addition to their drug delivery capabilities, their ability to locate specific cancer cells is just as important in the initial diagnosis of cancer. One of the best factors for beating cancer is early detection. Biodegradable nanoparticles can be equipped with materials that increase the resolution of medical scanning devices. This enhanced imaging through the use of biodegradable nanoparticles will result in an increase in early cancer detection, which will in turn increase the survival rate among patients. Biodegradable nanoparticles should be continuously researched and engineered for years to come due to their cancer fighting abilities. Based on the incredible extent of their current capabilities, the future research on biodegradable nanoparticles and their use for the treatment of cancer will have a profound impact on the detection and treatment of cancer. Biodegradable nanoparticle technology brings humanity one-step closer to the long-term goal of a totally effective cure for cancer. REFERENCES [1] (2011). “Cancer Facts and Figures 2011.” American Cancer Society. [Online]. 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Available: http://www.smartplanet.com/blog/smart-takes/biodegradable-nanoparticlesaim-to-give-antibiotics-a-turbo-charge/15266 ACKNOWLEDGEMENTS 7 Tyler Alan Justin Blazer We would like to thank our writing instructor Barbara Edelman for her help with focusing our topic and editing our conference paper. We would also like to thank our conference co-chair Katie Brown for her suggestions and advice that helped us to refine our paper. 8