Recently, nanoenabled drug delivery systems are gaining application in the pharmaceutical industry, since nanoparticle-based drugs may have improved solubility, pharmacokinetics, and biodistribution compared to small molecule drugs. Nanoparticle-carried drugs may also be easier to administer, have fewer side effects, and offer a market advantage to their developer. With the use of targeting, nanoparticle-based drugs can be efficacious at lower doses, which means lower costs and fewer deleterious side effects. Nanoparticle-based targeting has the potential to improve a whole host of conventional pharmaceuticals, and many nanoparticle formulations of existing drugs are already commercially available. 1
Many more will become available to patients in the near future: in 2006 alone, almost 250 nanotechnology-based products entered U.S. pharmaceutical pipelines.
However, the application of nanoparticles as intravenous drug delivery platforms may depend on avoiding rapid elimination from the systemic circulation by cells of the immune system. When nanoparticles enter the bloodstream, they immediately encounter a complex environment of plasma proteins and immune cells. Nanoparticle uptake by the immune cells may occur both in the blood stream by monocytes, platelets, leukocytes, and dendritic cells (DC) and in tissues by resident phagocytes (e.g., Kupffer cells in liver, DC in lymph nodes, macrophages and B cells in spleen). Nanoparticle uptake by immune cells may occur through various pathways2
and can be facilitated by the adsorption of opsonins (plasma proteins) to the particle’s surface. This uptake may cause a nanoparticle to be routed away from the site of its intended application and thus greatly reduce the number of nanoparticles available to reach the target site, effectively reducing the efficacy of the drug. As such, understanding nanoparticle hematocompatibility is an important step during the initial characterization of nanomaterials. There are three main categories considered relevant to nanoparticle biodistribution to the immune cells and organs; they are hemolysis, thrombogenicity and complement activation.
The term “hemolysis” is commonly used to describe damage to red blood cells (erythrocytes) leading to the leakage of the iron-containing protein hemoglobin into the bloodstream and potentially life-threatening conditions such as anemia. A nanoparticle which induces hemolysis may adsorb some quantity of the released hemoglobin and/or adhere to cell debris which in turn increases its likelihood of elimination by macrophages via scavenger receptor- and phosphatidylserine-mediated phagocytosis.3–5
Erythrocytes occupy a larger volume fraction of the blood than do mononuclear phagocytic cells, so an intravenously injected nanoparticle is likely to interact with red blood cells prior to encounters with other immune cells. This position early in the cascade of blood contact interaction, as well as the intensity of the adverse physiological outcome (severe hemolysis may lead to life threatening conditions such as anemia), makes an examination of hemolytic activity an important aspect of preclinical characterization of nanoparticles.
Thrombogenicity is the propensity of a material to induce blood clotting and partial or complete occlusion of a blood vessel by a thrombus (a mixture of red blood cells, aggregated platelets, fibrin and other cellular elements). Some early studies have indicated a relationship between nanoparticle thrombogenicity and phagocytosis by macrophages.6
Furthermore, nanoparticle-based drugs are often engineered to have longer systemic circulation times than small molecule drugs. These extended circulation times in the bloodstream increase the duration of the nanoparticle’s contact with blood components including the coagulation system, potentially amplifying activation of the coagulation cascade and blood clotting.
Lastly, the complement system is a group of proteins linked to each other in a biochemical cascade which removes pathogens from the body and may also contribute to altered nanoparticle biodistribtion in the form of rapid clearance from the systemic circulation via complement receptor-mediated phagocytosis. In addition, complement activation by systemically administered drugs is responsible for hypersensitivity (allergic) reactions and anaphylaxis, a life-threatening condition. As such, nanoparticles intended for systemic administration should be tested for tendency to activate the complement system.
The above effects confer increased importance to the preclinical testing of nanoparticle interaction with the immune system. Such preclinical testing is challenged by several factors. First, there is no harmonized procedure or regulatory guidance document currently available, though both national and international efforts are being made to rectify this situation.7–12
Second, even though a few standards for selecting biological test methods for medicinal materials and devices exist,13
material biocompatibility testing itself must rapidly evolve as basic research data continually suggests new and improved methods. Third, nanoparticles are a broad class of materials intended for a broad spectrum of clinical applications (e.g., as medical devices, components of vaccine formulations, or drug carriers for therapeutic intervention of malignant, inflammatory, viral, neurodegenerative and other types of disorders). The potentially different physicochemical properties of nanoparticles pose novel methodological and regulatory challenges.
A comprehensive overview of nanoparticle interaction with, and effects on, the immune system is provided elsewhere.2
Here, we will summarize some of the available research on testing nanomaterial hematocompatibility and their interaction with macrophages. Both of these types of testing are particularly important for nanoparticle-based drugs.