Pharmaceutical nanotechnology and nanomedicines
Yvonne Perrie
Chapter contents
Applications of pharmaceutical nanotechnology
Rationale for polymer conjugation
Polymer-drug conjugates case studies
Antibodies and antibody-drug conjugates
Dendrimer systems – case studies
Polymeric micelles – case studies
Nanosized drug particles and drug nanocrystals
Liposomes and bilayer vesicles
Clinical application of liposomes
Formulation design considerations for liposomes
Key points
Introduction
In general terms, pharmaceutical nanotechnology is a term applied to the design, characterization and production of pharmaceutical materials, structures and products that have one or more dimension between approximately 1 and 100 nm. However this classification remains open to debate, and a degree of ambiguity remains over what is considered nanotechnology, particularly regarding the size range considered. Currently an internationally accepted definition of nanotechnology is lacking and frequently particles in larger size ranges are considered as nanotechnology, e.g. the United States Food and Drug Administration (FDA) often considers 1000 nm as an appropraite upper limit regarding the screening of materials for consideration as nanotechnology. However, with the use of nanotechnology growing, both the European Medicines Agency and the FDA continue to refine their regulatory guidelines concerning nanotechnology in recognition of the key properties that nanomedicines can offer, i.e. their small size and high surface area to volume ratio. Yet it is generally agreed that using size alone as the defining factor for nanomedicines may be misleading. It is also useful to consider if a product exhibits different physical, chemical or biological properties that are attributed to its dimensions, even if these dimensions fall outside the nanoscale range. It is important to be aware that the functional effects, of the product, such as bioavailability, toxicity and/or potency may be influenced by the product’s dimensions.
By applying this general definition, pharmaceutical nanotechnology can encompass many systems from macromolecules, such as antibodies (e.g. Herceptin®) and polymer-protein conjugates (e.g. PegIntron®) to nanoscale particles (e.g. Emend®), through to colloidal and particulate constructs in the nano size range, such as liposomal formulations (e.g. Ambisome®) and nanoparticle systems (e.g. Abraxane®). Examples of the range of such pharmaceutical nanotechnologies, sometimes referred to nanomedicines, are shown in Figure 45.1.
Applications of pharmaceutical nanotechnology
The application of nanotechnology encompasses the formulation and development of nanomedicines to improve drug potency and efficacy and the use of nanomaterials in tissue engineering and implants to fabricate structures to support tissue regeneration within the body. Nanotechnology can also include the development of devices in the nano-range such as implantable sensory systems (nanodiagnostics) for improved diagnostic measurements. This chapter will focus on the use of nanotechnology in drug formulation, which can offer particular advantages including:
There are already several products authorized for clinical use that can be classified as nanomedicines (Table 45.1). Indeed some of these products have been approved for several years, having been originally approved for registration prior to the recognized classification of nanotechnology products. This includes some of the liposome-based products which have been licensed for clinical use since the mid-1990s. For example, the liposome formulation of doxorubicin (licensed in the US as Doxil® and marketed within Europe as Caelyx®) was the first liposomal product approved by the FDA. This product is a suspension of polyethylene glycol-coated-liposomes entrapping doxorubicin. Due to the ability of the liposome formulation to enhance targeting of doxorubicin to tumour sites, it was first developed for the treatment of AIDS-related Kaposi’s sarcoma, and is now licensed for other anti-tumour indications including metastatic breast cancer, advanced ovarian cancer and relapsed/refractory multiple myeloma. Table 45.1 shows that nanotechnology systems can offer a variety of attributes and each of these types of system is discussed in further detail in this chapter.
Polymer-drug conjugates
To improve the drug solubility and/or the delivery of drugs, drug molecules may be conjugated to polymers producing polymer-drug conjugates. These polymer-drug conjugates are considered as new chemical entities in their own right and, as their overall size is generally below 100 nm, these systems can be classified within the general area of nanotechnology. To build polymeric-drug conjugates there is a large range of synthetic polymers that can be produced having appropriate quality and stability attributes, and they can be custom-made to have distinct characteristics, including specified molecular weight, size, charge, etc. As these polymers are synthetic they are generally less immunogenic than naturally derived macromolecules. For the production of polymer-drug conjugates for parenteral administration, water soluble polymers are used.
A polymer-drug conjugate can be described as being built of three basic components:
A water soluble polymer backbone.
This can include synthetic polymers such as poly(ethyleneglycol) (PEG), poly(ethyleneimine) (PEI), poly(vinylpyrrolidone) (PVP) and polyvinylalcohol (PVA), poly(glutamic acid) (PGA), and hydroxypropylmethacrylate (HPMA) copolymers. Alternatively natural polymers such as dextran, chitosans, hyaluronic acid and proteins can be used. Of the polymers, PEG is the most widely used; it is approved by the FDA for human use and offers properties including low immunogenicity, antigenicity and toxicity. PEG chains also offer high hydration and flexibility which is useful in improving solubility and drug delivery. Another important property of PEG is their low polydispersity (in terms of molecular weight). The ease with which PEG can be modified and conjugated to drugs and proteins also offers an advantage. However, conjugation of PEG to proteins may in some instances reduce their biological activity so the conjugation site of the PEG on the protein is an important consideration. Within clinically approved products PEG molecular weights of 5000 to 40 000 Da are used.
A linker group.
Whilst a drug can be directly covalently bonded to a polymer, it is more common to attach the drug via a linker or spacer group, to help avoid the therapeutic action of the drug being blocked by the polymer. The linker can also be designed to be cleaved under certain conditions, such as changes in pH, enzymatic degradation or hydrolysis. This property can be used to promote the triggered release of the drug from the polymer conjugate under suitable conditions, thereby enhancing drug targeting. Examples of linker groups that can be used include amine, carbamate and ester groups, with an amide linker being the most common option.
Drug.
Commonly drugs delivered using these conjugates are those used in anti-cancer chemotherapy, such as doxorubicin and paclitaxel. This is because polymer conjugates can improve delivery and reduce unwanted side effects for these drugs which have narrow therapeutic windows. A second group of drugs that benefits from formulating as polymer conjugates is proteins e.g. L-asparaginase or interferons. Generally, proteins suffer from short half-lives and low stability after administration into the body. By conjugating proteins to polymers it is possible to increase their half-life by protecting the proteins from enzyme degradation and reducing clearance rates.
In addition to the three components of the system, targeting groups can be added to the polymer-conjugate with the aim of enhancing specificity and cellular uptake. However, there are no licensed products currently on the market which adopt this active targeting method. Examples of polymer-drug conjugates on the market are given in Table 45.2. As can be seen, PEG is used in several of these formulations as the polymer backbone and the majority of the systems are used to deliver protein therapeutics.
Table 45.2
| Name | Polymer-Drug | Indication |
| Adagen® | PEG-adenosine deaminase | SCID syndrome |
| Cimzia® | PEG-anti TNF Fab’ fragment | Crohn’s disease and rheumatoid arthritis |
| Oncaspar® | PEG-asparaginase | Acute lymphoblastic leukaemia |
| PEG-Intron® | PEG-Interferon 2b | Hepatitis C |
| Pegasys® | PEG-Interferon 2a | Hepatitis B and Hepatitis C |
| Neulastra® | PEG-Granulocyte colony-stimulating factor | Prevention of neutropenia associated with cancer chemotherapy |
| Macugen® | PEG-anti-VEGF aptamer | Age-related macular degeneration |
| Somavert® | PEG-growth hormone receptor antagonist | Acromegaly |
| Zinostatin Stimalamer® | Syren-maleic anhydride copolymer-Neocarzinostatin | Hepatocellular carcinoma |
Rationale for polymer conjugation
As noted above, the conjugation of drugs to polymers can improve the therapeutic action of the drug by improving solubility, protecting the drug from enzyme degradation, enhancing plasma circulation times and/or enhancing drug targeting. This is achieved through various actions.
Improving solubility
The conjugation of low-solubility drugs (e.g. paclitaxel, camptothecin or palatinate derivatives) to water soluble polymers can enhance the solubility of the overall system. For example OPAXIO®, a polymer-conjugate currently under development, comprising paclitaxel conjugated to poly-(L)-glutamic acid. The paclitaxel-conjugate has enhanced solubility compared to paclitaxel, therefore the conjugate can be administered without further solubilising agents.
Enhancing bioavailability and plasma half-life
The increased hydrodynamic volume of the polymer-drug conjugate compared to the free drug can reduce excretion rates via the kidneys. The renal clearance of compounds from the circulation is dictated by their molecular weight, with clearance rates decreasing with increasing molecular weight up to a threshold of around 45 kDa. Above 45 kDa, renal excretion cannot occur and larger polymers are more susceptible to clearance by the mononuclear phagocytic system (MPS). So, for example, the conjugation of molecules such as paclitaxel (~850 Da), and proteins such as interferon (~20 kDa) to water-soluble polymers increases their overall molecular weight enhancing drug circulation times and reducing kidney clearance rates.
Protecting against degradation after administration
The polymeric chains in the polymer-drug conjugate can also prevent the approach of antibodies and proteolytic enzymes to the drug. Water-soluble polymers become strongly hydrated and these hydrated polymer strands can promote steric hindrance, and block enzymes and antibodies reaching the drug. This protects the drug from degradation and enhances their plasma half-life and bioavailability. This is of particular advantage to protein-based therapeutic agents that are rapidly degraded by enzymes. However, it has been reported that antibodies against PEG can be generated in vivo and these can remove and neutralize PEG-conjugate products.
Reducing aggregation, immunogenicity and antigenicity
The hydrophilic coating offered by the polymers to the conjugate compound is the key to this property. The hydrated polymer chains can mask the hydrophobic regions in the protein, improve solubility and provide a steric shield that can help prevent protein-protein association, and reduce aggregation. For example, the native proteins in Neulastra® and PEG-Intron® have a high tendency to aggregate, however, PEG conjugation (referred to as PEGylation or pegylation) of these proteins can reduce aggregation and subsequently reduce associated immunogenic and antigenic problems. As already noted, the presence of the hydrated polymer in the conjugate can reduce antibody interactions, also reducing immunogenicity. PEGylation of proteins can also help stabilize proteins during lyophilization so helping to produce products with acceptable storage conditions.
Promoting targeting to specific organs, tissue or cells
By conjugation of drugs or proteins to water-soluble polymers, not only can their half-lives be improved, but the specific accumulation of the drug or protein can also be promoted in certain tissues. This can be achieved through the use of targeting groups or the phenomenon known as the enhanced permeability and retention (EPR) effect. This EPR effect can be described as passive targeting, whereby the distribution of the conjugate is dictated by local physiological conditions at the target site. Normally after a drug enters the systemic circulation, the drug is required to cross the endothelial lining of the vasculature before it can reach the target site. In most parts of the body, the endothelial lining is continuous with the endothelial cells situated on a basal membrane, and tight junctions between adjacent cells. This makes transport across this barrier difficult. However, the structure of the blood capillary wall varies in different organs and tissues, with three general types of endothelial cells being recognized:
Continuous.
Continuous endothelial cells are the most common. These cells have tight junctions and a continuous basement membrane. Continuous endothelial lining is found in areas, such as capillaries in the brain, lung and muscles.
Fenestrated.
This type of endothelial cells have gaps of between 20 and 80 nm between them, and this can allow the passage of small molecules out of the systemic circulation. Fenestrated endothelial linings are found in the capillaries in the kidneys and gastrointestinal tract.
Sinusoidal.
Here there are gaps between the endothelial cells of up to 150 nm. The basement membrane is either discontinuous, as in the capillaries in the spleen, or absent altogether, as in the case of the capillaries in the liver.
Additionally, the integrity of the endothelial barrier can be disturbed by inflammatory processes or by tumour growth. This can result in defective hypervasculature, leading to endothelial fenestrations as large as 200 to 300 nm being present in the endothelial lining. In addition, due to rapid tumour growth, deficient lymphatic drainage is also an issue. This modified permeability of the endothelium at such sites allow nanomedicines, including polymer-conjugates, to escape from the central circulation into the tumour site, where they are retained due to the poor tumour lymphatic drainage. This phenomenon is the EPR effect described above. Thus, conjugation of a drug or protein to an appropriate soluble polymer will result in a construct with a large hydrodynamic volume which will have reduced kidney excretion and therefore an enhanced systemic circulation time. Due to the EPR effect the polymer conjugates can escape from the systemic circulation into tumour sites where they will accumulate, enhancing drug action at the tumour site and reducing unwanted side effects elsewhere in the body.
The use of targeting groups to dictate the distribution of drugs and drug carriers can also be considered to promote targeting to specific site. This is commonly referred to as active targeting. Here, the designer of the drug-delivery system is relying on the interactions between a targeting group, which can be covalently attached to the polymer, and a corresponding receptor to facilitate the targeting of the system to a specific site. Examples of targeting groups include the use of antibodies due to their ability to specifically recognize and bind specific antigens (see below) and folate to target folate receptors which are overexpressed in tumour cells. Similarly, lectins are over-expressed on the surface of many tumour cells and can be targeted via the use of glycoproteins.
Polymer-drug conjugates case studies
OPAXIO® – a small-molecule conjugate.
In this polymer-drug conjugate, paclitaxel is conjugated to poly-L-glutamic acid (PGA) via an ester linker. Conjugating paclitaxel to the water-soluble PGA overcomes the poor aqueous solubility of paclitaxel, and the congugate can be infused into the body without the addition of solvents. This conjugate has a high drug content (~37% w/w) and is stable in the circulation and, while it remains bound to the polymer, paclitaxel is inactive. Due to its construct, the conjugate can passively target tumour sites via the EPR effect. The drug is then released intracellularly via degradation of PGA by lysosomal proteases, and the ester linker is degraded by esterases or acid hydrolysis. OPAXIO® is currently undergoing clinical trials as a potential treatment for non-small cell lung cancer and ovarian cancer.
