INTRODUCTION:

Nanotechnology is the new trend in the drug delivery system in the biomedical and pharmacy field. Nanotechnology is derived from the Greek word “nanos” means “dwarf”. This term is originated from the physics, chemistry and biology. It is one of the most important technologies in the future. Nanotechnology developed at several levels like materials, devices and systems. ^[1]

Nanotechnology can simply be defined as the technology at the scale of one-billionth of meters. It is the design, characterization, synthesis and application of materials, structures, devices and systems by controlling shape and size at nanometre scale. ^[2] It is the ability to work at the atomic, molecular and supra molecular levels to create and employ materials, structures, devices and systems with basically new properties. ^[3] Scientifically, nanotechnology is employed to describe materials, devices and systems with structures and components exhibiting new and significantly improved physical, chemical and biological properties as well as the phenomena and processes enabled by the ability to control properties at nanoscale. ^[4] Materials exhibit unique properties at nanoscale of 1 to 100 nanometre (nm). The changes in properties are due to increase in surface area and dominance of quantum effects which is associated with very small sizes and large surface area to volume ratio. ^[5]

There are several examples that illustrate this. Copper which is opaque at macro scale becomes transparent at nanoscale. ^[6-7]while platinum which is inert becomes a catalyst at nanoscale. ^[8-9] Silicon becomes a conductor at nanoscale.^[10-11] the quantum effects at nanoscale determine a material’s magnetic, thermal, optical and electrical properties. It is expected generally, that products at nanoscale will be cheaper due to less quantity of materials utilized.

At present this technology is most advanced which is used scientifically and commercially on large scale due to its size dependent physical and chemical properties. ^[12]Biomedical nanotechnology is one of the fastest-growing fields of research across the world in nanotechnology. However, it is clearly evident in some controversial fields like in cloning and stem cell research. ^[13]

The term nanotechnology was established by Norio Taniguchi in 1974.^[14] Through this technology some scientists gave the new idea that it is some kind of a new production through which high précised small dimension molecules are originated in extreme. Now days the nanotechnology is used in the widened scopes include the devices and systems rather than just materials. The essence of nanotechnology application is preferred by nanoscale and nano dimensions. These materials can be fabricated by using either “bottom up” or “top down” approaches. ^[15]

The main application of nanotechnology is shown in the “nano medicine” field or “nanobiomedicine” and this should be used in monitoring, diagnosis, treatment, controlling the biological systems. ^[16]

History of nanotechnology:

The platform for nanotechnology is believed by many workers in the field of nanotechnology to have been laid by Richard Feynman, a physicist at California Institute of Technology, He presented a technological vision of miniaturization of materials, manipulating and controlling things on a small scale called “Nanotechnology”.

The words ‘nanoscale, nano engineering, nanotechnology and nano-object in the references above have become the modern concept of Feynman’s speech. In 1959 Richard Feyman gives the official idea of the beginning of nanotechnology through his dissertation. The main idea behind this concept is the creation of the things out of tiny pieces rather than smaller. ^[17] The main idea behind the nanotechnology is the structure of an atom.

This word was established by the Norio Taniguchi in 1974. ^[18] A researcher at the University of Tokyo who used it to refer to the ability to engineer materials at nanoscale, Gerd Binnig invented scanning tunnelling microscopy (STM) while Henrich Rohrer invented atomic force microscopy. In 1985, Fullerene C60 was discovered by Kroto’s and Smalley’s research teams. In 1986, Eric Drexler began to promote and popularize nanotechnology through speeches and books “Engines of creation: the coming era of nanotechnology”. In 1991, Scientist discovered carbon nanotubes , the United States government launched the National Nanotechnology Initiative (NNI a Federal visionary research and development programme for nanotechnology based investments through the coordination of 16 various US departments and independent agencies) and these paved way for the progress in research and development in the field of nanotechnology . ^{[19, 20]}

Nanotechnology areas and applications:

Nanotechnology, being an interdisciplinary field, has three main extensively overlapping areas: Nanoelectronics, nanomaterials and nanobiotechnology which find applications in materials, electronics, environment, metrology, energy, security, robotics healthcare, information technology, biomimetics, pharmaceuticals, manufacturing, agriculture, construction, transport, and food processing and storage. ^{[1, 2, 4, 21, 22]}

Nanoelectronics:

Ø Information and computing:

Quantum dots and nanowires in cameras and personal computers.Nanotubes instead of cathode rays in televisions. Semiconductor silicon nanowires containing functioning electronic and optical devices.

Ø Sensors:

Nanomaterials used to assess the quality of the soil and water, and determine the state of plants, food and other products

Nanomaterials:

Ø One-dimensional materials:

Thin films and layers used in waterproof fabrics and electronics. Surfaces in fuel cells and as catalysts

Ø Two-dimensional materials:

Inorganic nanotubes such as molybdenum disulphide for catalysis and energy storage. Nanowires such as silicon nanowires for data storage, electronic and optoelectronic devices. Carbon nanotubes for sensors, electric current transmission and antistatic packaging. Nanotubes as containment for hydrogen in hydrogen fuel cells Biopolymers such as DNA molecules

Ø Three-dimensional materials:

Nanoparticles employed in cosmetics, textiles, paints, catalysis and drug delivery. Fullerenes are carbon materials which are employed as lubricants, drug delivery vehicles and in electric circuits. Dendrimers are polymeric molecules used in coatings and inks, for drug delivery and environmental remediation by trapping metals such as copper (II) which is then removed by ultra-filtration.

Nanobiotechnology:

Ø Bionano-sensors:

Combinations of enzymes and silicon chips implanted in humans or animals to monitor health and administer corrective doses of drugs.

Ø Biomimetic structures:

Diagnosis of diseases, molecular imaging and drug delivery

Ø Drug delivery:

New formulations for drug and gene therapies

Ø Tissue engineering:

Reproduction and repair of damaged tissues using nanomaterial based Scaffolds

One of the major impacts of nanotechnology and nanoscience will be in leading development of completely new drugs with more useful behavior and less side effects.

Impact of nanotechnology:

Nanotechnology a wide technological platform for a varying range of potential applications. The basic level of organization of atoms and molecules at which functions for man-made products and living things are defined can be manipulated by nanotechnology. Nanotechnology is interdisciplinary and so it reverses the trend of specialization in specific disciplines. Thus it integrates all disciplines especially biomedicine, engineering and technology. It has broadened and changed manufacturing capabilities, which were more of bulk manufacturing, to include self-assembling and top-down approach. The speed and scope of research and development have been influenced by nanotechnology such that regulators cannot meet up in assessment and environmental impact. Due to the vast areas of applications of nanotechnology, a number of governments such as U.S., Japan,

China and Europe have deemed it fit to invest in nanotechnology. Nanotechnology is currently one of the main propellants for technological, economical change and industrial competitions. ^[23]

ECONOMY OF NANOPHARMACEUTICALS:

According to a report from markets, an industry consulting firm based in Sterling, USA, nanotechnology drug delivery systems will generate over $1.7 billion ($US) in 2009 and over $4.8 billion in 2012. The global drug delivery products and services market is projected to surpass US$67 billion in 2009. Lux Research reported in big Pharma companies are “flat footed” in their initiative about nanotechnology, however, medical devices companies are more aggressive in pursuing the nanotechnological strategies. The nanotechnology shows its impact on design of drug molecule, the benefit of which will be seen in augmented the product life cycle, patent life, along with their therapeutic efficiency. ^[24]

RECENT DEVELOPMENTS IN NANOTECHNOLOGY FIELD

Tissue engineering:

Natural bone surface is quiet rough in nature and 100 nm in surface size but the artificial implanting of the bone leads to the smoothness which causes the rejection through the body. Because of that smooth surface is likely to cause production of a fibrous tissue covering the surface of the implant. This layer reduces the bone-implant contact, which may be resulted in loosening of the implant and further inflammation.

It was demonstrated by creating nano-sized features on the surface of the hip or knee prosthesis could reduce the chances of rejection as well as to stimulate the production of osteoblasts. ^[25] The osteoblasts are the cells responsible for the growth of the bone matrix and are found on the advancing surface of the developing bone. The effect was demonstrated with polymeric, ceramic and, more recently, metal materials. More than 90% of the human bone cells from suspension adhered to the nanostructured metal surface, but only 50% in the control sample. Titanium is a well-known bone repairing material widely used in orthopaedics and dentistry. It has a high fracture resistance, ductility and weight to strength ratio. Unfortunately, it suffers from the lack of bioactivity, as it does not support cell adhesion and growth well. These coatings were suffered from thickness non-uniformity, poor adhesion and low mechanical strength. In addition, a stable porous structure is required to support the nutrients transport through the cell growth. It was shown that using a biomimetic approach a slow growth of nano structured apatite film from the simulated body fluid resulted in the formation of a strongly adherent, uniform nano porous layer. The layer was found to be built of 60 nm crystallites, and possess a stable nanoporous structure and bioactivity. A real bone is a nanocomposite material, composed of hydroxyl apatite crystallites in the organic matrix, mainly composed of collagen. The actual nanoscale mechanism leading to this useful combination of properties is still debated. An artificial hybrid material was prepared from 15–18 nm ceramic nanoparticles and poly (methyl methacrylate) copolymer. ^[26]

Cancer therapy:

Cancer therapy is based on the destruction of the cancer cells by laser generated atomic oxygen, which is cytotoxic. A greater quantity of a special dye was used to generate the atomic oxygen was taken by the cancer cells when compared with a healthy tissue. Hence, only the cancer cells are destroyed then exposed to a laser radiation. Unfortunately, the remaining dye molecules migrated to the skin and the eyes and make the patient very sensitive to the daylight exposure. This effect can last for up to six weeks. To avoid this side effect, the hydrophobic version of the dye molecule was enclosed inside a porous nanoparticle. The dye stayed trapped inside the Ormosil nanoparticle and did not spread to the other parts of the body. At the same time, its oxygen generating ability has not been affected and the pore size of about 1 nm freely allowed for the oxygen to diffuse out. ^[27]

Improving public health:

Through nanotechnology and biological combinations will provide biosensors, biomaterials and new breeds of biochips for treating life-threatening conditions, including cancer and heart disease. Such bioengineered devices, in the form of body implants, will deliver smart drugs or carry new cells to repair damaged tissue.

Protein detection:

Proteins are the important part of the cell machinery and structure, and understanding their functionalities is extremely important for further progress in human well being. Gold nanoparticles are widely used in immune histo-chemistry for identifying protein-protein interaction. However, the multiple simultaneous detection capabilities of this technique are fairly limited. Surface-enhanced Raman scattering spectroscopy was a well-established technique for detection and identification of single dye molecules. By combining both these methods in a single nano particle probe can drastically improves the multiplexing capabilities of protein probes. The nanoparticles are coated with hydrophilic oligonucleotides containing a Raman dye at one end and terminally capped with a small molecule recognition element (e.g. biotin). Moreover, this molecule was catalytically active and will be coated with silver in the solution of Ag (I) and hydroquinone. After the probe attachment to the small molecule or antigen detection, the substrate was exposed to silver and hydroquinone solution. A silver-plating is happening close to the Raman dye, which allows for dye signature detection with a standard Raman microscope. Apart from being recognized small molecules this probe can be modified to contain antibodies on the surface to recognize the proteins. When tested in the protein array format against both small molecules and proteins, the probe has shown no cross-reactivity. ^[28]

Current application areas:

Nanotechnology was already having an impact on products as diverse as novel foods, medical devices, chemical coatings, personal health testing kits, sensors for security systems, water purification units for manned space craft, displays for hand-held computer games, and high-resolution cinema screens. ^[29]

METHOD OF SYNTHESIS:

There are basically two approaches for the synthesis of nanostructures, irrespective of the field or discipline: ‘Bottom-Up’ approach and ‘Top-Down’ approach.

‘Bottom-Up’ approach:

The building of nanostructures is achieved by growing or assembling of atoms or molecules which are the building blocks. The buildings blocks may be manipulated through controlled chemical reactions to self assemble and make nanostructures such as nanotubes and quantum dots. ^[30] Atoms or molecules may also be physically manipulated to form nanostructures using minute probes. Self-assembling of atoms or molecules can be achieved by templating and non-templating. ^[31] Templating involves the interaction of bio macromolecules under the influence of a specific sequence, pattern, structure, external force or spatial constraint. For instance, non-ionic surfactants and block co-polymers are used as templates in the formation of nanostructures. Two dimensional hexagonal nanostructures formed by cylindrical amphiphilic polymeric micelles were used as templates to fabricate nanotubes of cadmium sulphide, a semiconductor.^[32] Non-templating is the formation of nanostructures from atoms or molecules with external influence. Self assembly lithography which is cost-effective and efficient is one technique that can be used to produce nanostructures below 100nm. Bottom-Up’ is considered to be an ideal approach for nanotechnology. Bottom-Up unfolds almost unlimited possibilities in the design and construction of artificial molecular devices capable of performing specific functions upon stimulation with external energy input. ^[33] However, one of the challenges of the bottom-up approach is the random movement exhibited by atoms which has to be overcome. ^[34]

Top-Down’ approach:

Bulk materials are reduced by some processes to form nanostructures. ‘Top down’ is achieved by breaking, cutting or etching techniques.^[31] which is achieved by bulk or film machining, surface machining and mold machining employing lithography ^[30] Bulk machining employs photolithography which applies the etching process while mold machining employs soft lithography. Other techniques are electron beam lithography, x-ray lithography and micro-electro-mechanical systems lithography. ^[35] However, photo lithography and related techniques have limitations of fabricating nanostructures ofsub-100nm and so cannot be applied for nanostructures below 100 nm.

Benefits of nanotechnology:

Based on its broad sectors of applications, nanotechnology has numerous benefits globally both in developed and developing countries: Creation of new products and improvement on existing products. Availability of stronger, tougher and lighter materials for construction and engineering. Cleaner drinking water due to the creation of filters that can entrap organisms and toxins. Cleaner environment through remediation to remove pollutants from the environment Improved healthcare by fabrication of devices and drug delivery systems for better monitoring, diagnosis and treatment of chronic diseases. Improvement on transport systems Cheaper and clean energy

Risks of nanotechnology:

Despite the great potentials of nanotechnology, its safety in humans, animal and plants, and effects on the environment are of concern. Also, military application is of concern as chemical weapons fabricated from nanoparticles will be more deadly than present chemical weapons.

This is due to the fact that the smaller a particle, the greater its impact either positively or negatively. Some nanoparticles show increased toxicity due to their increased surface area.Studies have shown carbon nano tubes to be cytotoxic and to induce granulomas in lungs of laboratory animals. Also, metals and metallic oxide nanoparticles such as copper, cobalt, titanium oxide and silicon oxide have inflammatory and toxic effects on cells. ^[36] However, studies and debates are going on about the benefits and risks of nanotechnology. ^[37-38] Optimistically, the benefits of nanotechnology are enormous and so studies which include the health, environmental, ethical and safety issues should indicate how to maximize the benefits and reduce the risks. Macro and micro technologies had their risks, yet the benefits were accepted.

NANOTECHNOLOGY IN DRUG DELIVERY:

Some of the challenges of most drug delivery systems include poor bioavailability, in vivo stability, solubility, intestinal absorption, sustained and targeted delivery to site of action, therapeutic effectiveness, side effects, and plasma fluctuations of drugs which either fall below the minimum effective concentrations or exceed the safe therapeutic concentrations. However, nanotechnology in drug delivery is an approach designed to overcome these challenges due to the development and fabrication of nanostructures at submicron scale and nanoscale which are mainly polymeric and have multiple advantages. Generally, nanostructures have the ability to protect drugs encapsulated within them from hydrolytic and enzymatic degradation in the gastrointestinal tract; target the delivery of a wide range of drugs to various areas of the body for sustained release and thus are able to deliver drugs, proteins and genes through the peroral route of administration. ^{[39 40]} This technique delivers drugs that are highly water insoluble; can bypass the liver, thereby preventing the first pass metabolism of the incorporate drug. ^{[41, 42]} They increase oral bioavailability of drugs due to their specialized uptake mechanisms such as absorptive endocytosis and are able to remain in the blood circulation for a longer time, releasing the incorporated drug in a sustained and continuous manner leading to less plasma fluctuations thereby minimizing side-effects caused by drugs. ^[41] Due to the size of nanostructures, they are able to penetrate into tissues and are taken up by cells, allowing efficient delivery of drugs to sites of action. The uptake of nanostructures was found to be 15-250 times greater than that of microparticles in the 1- 10μm range. ^[43] Through the manipulation of the characteristics of polymers, release of drug from nanostructures can be controlled to achieve the desired therapeutic concentration for the desired duration. For targeted delivery, nanostructures can be conjugated with targeting moieties such that the linkage between the polymer and the active substance can be manipulated to control the site and duration at which the drug is released. The linkage may be achieved by incorporation of amino acids, lipids, peptides or small chains as spacer molecules. ^[44] Drug targeting is crucial in chemotherapy, where a drug delivery system can target only the malignant tumour while shielding the healthy cells from uniform distribution of chemotherapeutics in the body and their harmful effects. The use of nanostructures such as polymeric nanoparticles is a non-invasive approach of penetrating the blood brain barrier for management of neurodegenerative disorders, cerebrovascular and inflammatory diseases.^[45,46]Research and development of new drugs are capital- and time- intensive which requires that pharmaceutical companies, in addition, search for other means of meeting up with market demands. New drug delivery methods enable pharmaceutical companies reformulate existing drugs in the market. Nanotechnology is strategic in developing drug delivery systems which can expand drug markets. Nanotechnology can be applied to reformulate existing drugs thereby extending products’ lives, enhance their performance, improve their acceptability by increasing effectiveness, as well as increase safety and patient adherence, and ultimately reduce health care costs. ^{[42, 47]} It provides drug delivery carriers, as well as treatment and management of chronic diseases which include cancer, HIV/AIDS and diabetes.

NOVEL APPLICATIONS OF NANOTECHNOLOGY IN MEDICINE:

Current techniques for diagnosis and treatment of various diseases, especially cancer have major limitations such as poor sensitivity or specificity and drug toxicities. Newer and improved methods of cancer detection based on nanoparticles are being developed. They are used as contrast agents, fluorescent materials, molecular research tools and drugs with targeting antibodies. Paramagnetic nanoparticles, quantum dots, nanoshells and nanosomes are few of the nanoparticles used for diagnostic purposes.

Liposomes:

Liposomes discovered in mid 1960s were the original models of nanoscaled drug delivery devices. They are spherical nanoparticles made of lipid bilayer membranes with an aqueous interior but can be unilamellar with a single lamella of membrane or multilamellar with multiple membranes. They can be used as effective drug delivery systems. Cancer chemotherapeutic drugs and other toxic drugs like amphotericin and hamycin, when used as liposomal drugs produce much better efficacy and safety as compared to conventional preparations.

Targeting of liposomal drugs:

Liposome can be targeted to specific organ or tissue by passive as well as active methods. As the liposomal drug acts minimally on other tissues, the safety profile is better than non-liposomal drug. The vascularity in tumour tissue is poorly organized and significant leak occurs from blood vessel in the tumour tissue. The liposomal drugs get accumulated in the tumour tissue passively and produce enhanced effects. Active targeting of the drug can be achieved by using immunoliposomes and ligand directed liposomes.

Immunoliposomes are liposomes conjugated with an antibody directed towards the tumour antigen. The antibody can be conjugated to the surface of a stealth liposome, the polyoxyethylene coating of a stealth liposome or on the surface of a non stealth liposome. These immunoliposomes when injected into the body, reaches the target tissue and gets accumulated in its site of action. This reduces unwanted effects and also increases the drug delivery to the target tissue, thus enhancing its safety and efficacy. ^[48] Antibody directed enzyme prodrug therapy (ADEPT) consists of liposomes conjugated with an enzyme to activate a prodrug and an antibody directed to a tumour antigen (enzyme linked immunoliposomes). These are administered prior to administration of a prodrug. The antibody directs the enzyme to the target tissue where it activates the prodrug selectively and converts it to its active form. This way, action of the drug is avoided in other normal tissues, thus minimizing the toxicity of drug. ^[49-50] such studies are being tried with epirubicin and doxorubicin ^{[51, 52]} Ligand bearing liposomes are conjugated with specific ligands which are directed towards target structures. In ovarian cancer, overexpression of folate receptors by the tumour tissue occurs. The liposomal drug can be conjugated with folate so as to direct the molecule to the tumour. ^[53]

Nanopores:

Nanopores designed in 1997 by Desai and Ferrari ^[54^] consists of wafers with high density of pores (20 nm in diameter). The pores allow entry of oxygen, glucose and other products like insulin to pass through. However, it does not allow immunoglobulin and cells to pass through them. Nanopores can be used as devices to protect transplanted tissues from the host immune system, at the same time, utilizing the benefit of transplantation. β cells of pancreas can be enclosed within the nanopore device and implanted in the recipient’s body. Nanopores can also be employed in DNA sequencing. Branton’s team at Harvard University ^[55]has been working on modified nanopores that have the ability to differentiate DNA strands based on differences in base pair sequences.

Fullerenes:

Fullerenes, a carbon allotrope, also called as “buckyballs” were discovered in 1985. ^[56] The buckminster fullerene is the most common form of fullerene measuring about 7 Å in diameter with 60 carbon atoms arranged in a shape known as truncated icosahedrons. It resembles a soccer ball with 20 hexagons and 12 pentagons and is highly symmetrical. ^[57]

Types of fullerenes:

Alkali doped fullerenes are structures with alkali metal atoms in between fullerenes contributing valence electrons to neighboring fullerenes ^[58]. Endohedral fullerenes have another atom enclosed inside the bucky ball. Exohedral fullerenes also called as fullerene derivatives are synthesized by chemical reaction between the fullerene and other chemical groups. These are also called as functionalized fullerenes. Such fullerenes can be used as photosensitizers in photodynamic therapy for malignancies. Hetero fullerenes are fullerene compounds where one or more carbon atoms are replaced by other atoms like nitrogen or boron. Fullerenes are being investigated for drug transport of antiviral drugs, antibiotics and anticancer agents. ^{[59, 60]} Fullerenes can also be used as free radical scavengers due to presence of high number of conjugated double bonds in the core structure. These are found to have a protective activity against mitochondrial injury induced by free radicals. ^[61]

Nanotubes:

Carbon nanotubes discovered in 1991 ^[62] are tubular structures like a sheet of graphite rolled into a cylinder capped at one or both ends by a buckyball. Nanotubes can be single walled carbon nanotube (SWCNT) or multiwalled carbon nanotube (MWCNT) in concentric fashion. Single walled nanotube has an internal diameter of 1-2 nm and multiwalled nanotube has a diameter of 2-25 nm with 0.36 nm distance between layers of MWCNT. These vary in their length ranging from 1 μm to a few micrometers. ^[63] These are characterized by greater strength and stability hence can be used as stable drug carriers. Amphotericin B nanotubes has shown increased drug delivery to the interior of cells compared to amphotericin B administration without nanotubes. ^[64]

The efficacy of amphotericin B nanotubes was greater as an antifungal agent compared to amphotericin B alone and it was effective on strains of fungi which are usually resistant to amphotericin B alone. DNA can be attached to the tips of nanotubes or can be incorporated within the tubes. Gene silencing studies with small interfering RNA (siRNA) have been done as a modality of cancer therapy where tumour cells will be selectively modulated. Functionalized single walled carbon nanotubes can be used with siRNA to silence targeted gene expression. ^[65] It was observed that carbon nanotubes, except acetylated ones, when bonded with a peptide produce a higher immunological response compared to free peptides. This property can be used in vaccine production to enhance the efficacy of vaccines. Further, it was also found that compounds bound to nanotubes increase the efficacy of diagnostic methods like ELISA.

Quantum dots:

Quantum dots are nanocrystals measuring around 2-10 nm which can be made to fluorescence when stimulated by light. Their structure consists of an inorganic core, the size of which determines the colour emitted an inorganic shell and an aqueous organic coating to which biomolecules are conjugated. The bio molecule conjugation of the quantum dots can be modulated to target various biomarkers. ^[66] Quantum dots can be used for biomedical purposes as a diagnostic as well as therapeutic tool. These can be tagged with biomolecules and used as highly sensitive probes. The fluorescence produced by quantum dots is much brighter than those produced by conventional dyes when used with NIR fluorescence system. ^[67]

Nanoshells:

Nano shells were developed by West and Halas ^[68] at Rice University as a new modality of targeted therapy. Nanoshells consist of nanoparticles with a core of silica and a coating of thin metallic shell. These can be targeted to desired tissue by using immunological methods. This technology is being evaluated for cancer therapy. Hirsh et al ^[^69] used nano shells which are tuned to absorb infra red rays when exposed from a source outside the body to demonstrate the thermal property of nanoshells. The nano shells when exposed to NIR region of the electromagnetic spectrum get heated and cause destruction of the tissue. Gold nanoshells can be coupled to antibodies and the size can be modulated so that it responds to NIR wavelength, which has the ability to penetrate whole blood specimens.

Nanobubbles:

Cancer therapeutic drugs can be incorporated into nanoscaled bubble like structures called as nanobubbles. These nanobubbles remain stable at room temperature and when heated to physiological temperature within the body coalesce to form microbubbles. These have the advantages of targeting the tumour tissue and delivering the drug selectively under the influence of ultrasound exposure. This results in increased intracellular uptake of the drug by the tumour cells. It also provides an additional advantage of enabling visualisation of the tumour by means of ultrasound methods. ^{[70, 71]}

Paramagnetic nanoparticles:

Paramagnetic nanoparticles are being tried for both diagnostic and therapeutic purposes. Diagnostically, paramagnetic iron oxide nanoparticles are used as contrast agents in magnetic resonance imaging. These have a greater magnetic susceptibility than conventional contrast agents. Targeting of these nanoparticles enables identification of specific organs and tissues. ^[72] The use of iron oxide in MRI imaging faces limitations like specificity and internalization by macrophages. ^[73] Paramagnetic nanoparticles conjugated with antibodies to HER-2/neuwhich are expressed on breast cancer cells have been used with MRI to detect breast cancer cells in vitro. ^[^74] Study done by Leuschneret al ^[^75] has demonstrated the in vivo detection of breast cancer cells using paramagnetic nanoparticles conjugated with luteinizing hormone releasing hormone as breast cancer cells express LHRH receptors.

Nanosomes:

Raoul Kopelman’s group at the University of Michigan, USA, has been working on nanosomes also called as PEBBLEs (Probes Encapsulated by Biologically Localized Embedding) which integrate various aspects of medical applications such as targeting, diagnosis and therapy. These nanosomes are being developed for treatment of various tumours, in particular CNS tumours. Silica coated iron oxide nanoparticles coated with polyethylene glycol. ^[76] and affixed with targeting antibody and contrast elements like gadolinium are used to access specific areas of brain involved with tumour. Targeting aids in binding the nanoparticle specifically to the tumour cells and the contrast elements helps in better detection with magnetic resonance imaging. Subsequent treatment with laser can destroy the cells loaded with these nanoparticles by the heat generated by iron oxide particles by absorbing the infra red light.

Dendrimers:

Dendrimers are nanomolecules with regular branching structures. The number of branching determines the size of the dendrimer which can be controlled. The branches arise from the core in shape of a spherical structure by means of polymerization. This results in formation of cavities within the dendrimer molecule which can be used for drug transport. The ends of the dendrimer molecule can be attached with other molecules for transport. These molecules give the dendrimers various functional applications. ^[77]

Dendrimers can be used for gene therapy where these can replace conventional viral vectors. Dendrimers have been tested in mammalian cell types and in animal models. Dendrimer based drugs are being tried for antiretroviral therapy ^[78] and it is in stages of clinical trial after getting clearance from US-FDA on July 2003. This molecule was found to successfully prevent simian HIV infection. ^[55] Those dendrimers with a hydrophilic surface escape renal clearance and have a greater circulation time. ^[79]

Nanotechnology in gene therapy:

Gene therapy is a newer modality of approach for treatment of many genetic disorders including diabetes mellitus. ^[80]cystic fibrosis ^[81] and alpha 1 antitrypsin deficiency. ^[82] Viral vectors used for gene transfer have the limitations of safety concerns and stimulation of immune system with production of antibodies against the viral vectors. Further, naked DNA cannot cross the negatively charged cell membrane as these are also negatively charged ^[83] Hence, there is a need for other modes of transfer of genetic material such as nanoparticle based gene therapy.

Liposomes measuring less than 100 nm can be used for delivery of genetic material into cells. Liposomes incorporated with polyethylene glycol and galactose target liver cells effectively due to their rapid uptake by liver Kupffer cells. Thus gene therapy may be tried with such liposomal nanoparticles for various liver disorders such as Wilson’s disease and hereditary hemochromatosis. ^[84]

Respirocytes:

Respirocytes are hypothetical artificial red blood cells are nanodevices which can function as red blood cells but with greater efficacy. These have higher capacity to deliver oxygen to tissues, supplying 236 times more oxygen per unit volume than natural red blood cells. These devices have sensors on the surface which can detect changes in the environment and the onboard nanocomputer will regulate the intake and output of the oxygen and carbon dioxide molecules. Respirocytes, considered as a device by FDA are regulated under the provisions of the Medical Device Amendments of 1976, the Safe Medical Devices Act of 1990, and the Medical Device Amendments of 1992. ^[85]

Microbivores:

Microbivores ^[86] are hypothetical structures which function as white blood cells in the blood stream designed to trap circulating microbes. They are expected to have greater efficacy than cellular blood cells. The microbivores surface is arranged with processes which can extend in length and secure the microbe which gets in contact with it. The Application of the microbivores in human circulation could theoretically clear the blood stream in septicaemia at a much greater rate than the natural defence mechanism with antibiotics. ^{[55, 86]}

Regulatory challenges with nano medicines:

Regulatory issues play a major role in the development of nano formulation drugs. These include, the type of nano drug produced and the various regulatory requirements that the manufacturers must follow during the manufacturing of nano drugs. A nano formulation of a drug which is based on a previously approved drug in micro formulation can undergo a shorter approval pathway by means of abbreviated new drug application if bioequivalence can be demonstrated to its micro formulation drug. However, if bioequivalence cannot be demonstrated, it would necessitate approval of all the stages of new drug application. Further, when a nano drug is designed as a new chemical entity, the evaluation procedure becomes more stringent. ^[87] Nano drug manufacturers must comply with FDA’s Current Good Manufacturing Practices (CGMP) and Quality System Regulations (QSR) ^[88] Non compliance with these regulations would warrant enforcement actions by the FDA. Further, the training must be conducted at adequate frequency and there must be adequate number of qualified personnel for the assigned duties. ^[89] A nano drug manufacturer must invest considerable amount of financial resources to have such qualified personnel in the working unit. Maintenance of equipment for manufacture of nano drugs and control of contamination are also regulatory requirements for manufacturers. The drug products are purified by the use of filters and CGMP demands that the filters do not release fibres. ^[90] However, when liquid filtration is used, nano drug manufacturers will not be able to comply with CGMP, since the smallest filtration level available is approximately 15 nm and nano drugs could be at the range of 5 to 6 nm long. ^[87]The FDA centres namely, the Center for Drug Evaluation and Research (CDER), the Center for Devices and Radiological Health (CDRH) and the Center for Biologics Evaluation and Research (CBER) regulate drugs, devices, and biologics respectively and are responsible for regulating nano medical products. FDA classifies medicinal products as drug, device or biologics according to their primary mode of action to assign a centre for their primary jurisdiction during the evaluation process. In case of a nano drug it is difficult to classify it as a drug, device or biologics since it tends to have a combination of the above. Hence, the assignment of the Centre becomes difficult. Further, the drug has to pass through all the Centres of FDA owing to its complexity. This results in greater time period for approval of the drug. The staff of FDA must also be sufficiently educated and trained in nanotechnology in the field of medicine to evaluate nano drug products^{. [91]}

Potential hazards of nanoparticles:

Nanoparticles, as a result of their extreme microscopic dimension, which gives unique advantage, have potential hazards similar to particulate matter ^[92]. These particles have the potential to cause varied pathologies of respiratory, cardiovascular and gastrointestinal system1. Intratracheal instillation of carbon nanotube particles in mice has shown that carbon nanotubes have the potential to cause varied lung pathologies like epitheloid granuloma, interstitial inflammation, peribronchial inflammation and necrosis of lung. The toxicity produced by carbon nanotube was found to be greater than that produced by carbon black and quartz. ^[93] Nanoparticles can enter the central nervous system either directly through axons of olfactory pathway or through systemic circulation that C60 fullerene can cause oxidative stress and depletion of GSH in brain in fishes by entering through the olfactory bulb. ^[94] Involvement of olfactory bulb in humans is possible in case of inhalational exposure. Studies done on monkeys and rats have shown accumulation of carbon and manganese nanoparticles in the olfactory bulb through the olfactory pathway. ^{[95, 96]} This shows that nanoparticle mediated delivery can in future provide a means of alternate route, circumventing the blood brain barrier. However, this can also result in the inflammatory reactions in the brain which needs to be evaluated.. It was also observed that fullerenes do not have the property of inducing platelet aggregation. Thus, for designing nanoparticle based drug delivery systems, fullerenes may be a safer approach as compared to nanotubes.^[97]

REFERENCES:

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Received on 14.03.2013 Modified on 30.03.2013

Research J. Pharm. and Tech. 6(5): May 2013; Page 486-495