DRUG DELIVERY is a special form of drug delivery system where the pharmacologically active agent or medicament is selectively targeted or delivered only to its site of action or absorption and not to the non-target organs or tissues or cells. The drug may be delivered To the capillary bed of the active sites,To the specific type of cell (or) even an intracellular region. Ex- tumour cells but not to normal cells,To a specific organ (or) tissues by complexing with the carrier that recognizes the targetREASON FOR DRUG TARGETING:In the treatment or prevention or diseases.Pharmaceutical drug instability in conventional dosage form solubility ,biopharmaceutical low absorption, high-membrane bounding, biological instability, pharmacokinetic / pharmacodynamic short halflife, large volume of distribution, low specificity, clinical, low therapeutic index.

To achieve a desired pharmacological response at a selected sites without undesirable interaction at other sites, there by the drug have a specific action with minimum side effects & better therapeutic index. Ex- in cancer chemotherapy and enzyme replacement therapy.

IDEAL CHARACTERISTICS:Targeted drug delivery system should be-biochemically inert (non-toxic), non-immunogenic.both physically and chemically stable in vivo and in vitro.restrict drug distribution to target cells or tissues or organs and should have uniform capillary distribution.controllable and predicate rate of drug release.drug release does not effect the drug action.therapeutic amount of drug release.minimal drug leakage during transit.carriers used must be bio-degradable or readily eliminated from the body without any problem and no carrier induced modulation of diseased state.the preparation of the delivery system should be easy or reasonably simple, reproductive and cost effective.

Targeting of drugs to special cells and tissues of the body without their becoming a part of systemic circulation is a very novel idea. If a drug can be administered in a form such that it reaches the receptor sites in sufficient concentration without disturbing in extraneous tissue cells.Such products are prepared by considering-1.specific properties of target cells.2.nature of markers or transport carriers or vehicles, which convey drug to specific receptors.3.ligands and physically modulated components.

TARGET:Target may be defined as a cell or group of cells in minority, identified to be in the need of treatment. CARRIERS OR MARKERS:Carrier is one of the important entity essentially required for effective transportation of loaded drugs. They are vectors, which sequester, retain drug and transport or deliver it into the vicinity of the target cells. LIGANDS:The ligands confer recognition and specificity upon carrier/vector and lend them to approach the respective target and deliver the drug. Ex-antibodies, polypeptides, endogenous hormones etc.DEFINITIONS

There are various approaches in delivery of a therapeutic substance to the target site as its necessary for designing a controlled drug delivery system.One such approach is using polymeric microspheres as carriers for drugs.Microspheres of biodegradable & non-biodegradable polymers have been investigated for the sustained release depending on the final application.

MICROSPHERES DEFNITION:Microspheres are the free flowing powders containing of encapsulated (drugs) spherical particles of size ideally less than 125m that can be suspended in aqueous vehicle & injected by an 18 (or) 16 number needle. Ex: polyacric acid, polyacetideglycoside.

MAGNETIC MICROSPHERES:-These are the microspheres containing substance inside which can be easily targeted by applying external magnetic field.Magnetic microspheres were developed:a)To minimize renal clearanceb) To increase target site specificityc) To entrap a wide variety of drugs d) In the entrapment of localised tumours in the religions of well defined blood supplye) For controlled release of peptide protein drugs such as LHRH, which have the short half life and otherwise need to be injected once or more daily as conventional parenteral formulation.f)To minimize reticulo endothelial clearance target site specificity.

MICROSPHERES AS DRUG CARRIERS:- The most important characteristic of microsphere is the microphase separation morphology which endows it with a controllable variability in degradation rate and also drug release. As opposed to other physical forms of drug carriers such as rods, cylinders, slabs etc.1.Microspheres based on the biodegradable polymers:- The biodegradable microspheres can be prepared from certain synthesis as well as natural polymers.An important requirement of such polymers is that the degradation products should be non toxic because such products eventually enter the systemic circulation or result in the tissue deposition.

2.Polymer of microspheres:-The preparation of microspheres should satisfy certain criteria. They are: The ability to incorporate reasonably high concentrations of the drug.Stability of the preparation after synthesis with a clinically acceptable shelf-life.Controllable particle-size & dispersion in aqueous vehicles for injection.Release of active agent with good control over a wide time scale.Biocompatibility with a controllable biodegradability &Susceptibility to chemical modification.

CONCEPT OF TARGETING OF MICROSPHERES: As we know that the capillaries of the human body are in the microns, so one can easily target the capillaries of lungs, blood, liver etc. by the use of microspheres.

CONCEPT OF MAGNETIC TARGETING OF MICROSPHERES: Ideally, magnetic microspheres are injected into an artery that supplies a given site. As the microspheres would be selectively & magnetically localized at the capillary level, they would have free flow access through the large arteries

IMPORTANT CHARACTERISTICS OF MICROSPHERES & MAGNETIC MICROSPHERES:

Particle size of a drug carrier can affect the degree of drug entrapment.Increase in size of the albumin microspheres due to the hydration can alter its body distribution.Use of sub-micron size microspheres minimises the incidence of pulmonary embolism that often encounters with particles greater than 7m or particles which aggregate upon their vivo administration.The retention of magnetic microspheres at the target site is dependent on the magnetic content of the carrier & the magnitude of the applied magnetic field.Although high magnetic content allows the use of smaller magnetic fields, it reduces the effective space available within the carrier for the drug entrapment.In targeting using MM, the magnetic field content of the carrier & the magnitude of applied magnetic field are important.

Incorporation of drugs & magnetic needs are to be delicately balanced. Depending on the type of drugs & the desired target site, the optimum magnetic content would vary between 20% & 50% magnetic by dry weight of the drug carrier complex retention. The microspheres migrated from endothelial cells into the intestinal compartment & formed a depot for sustained drug release over an extended period of time.

For example: Magnetite.

MAGNETITE: Its also called as ferric ferrous oxide, tri iron tetra oxide, & black iron oxide. Magnetic iron oxide chemical formula FeO.Fe2O3 having a molecular weight of 231.55 with a chemical composition of Fe=72.36%, O=27.64%. The ferromagnetic materials when incorporated into microspheres makes them magnetically responsive, so that they can be concentrated to the desired site by applying some external magnetic field. Iron although showing Ferro magnetic substance, due to its local tissue irritation & other toxic manifestations cannot be included into microspheres.

Drug content depends on the size. Drug content which in turn is governed by solubility characters of drug & their method of preparation.Size of microspheres if size less than target, the drug delivery will be more, but due to static attraction aggregation may take place following thrombo-embolism.Rate of hydration of polymer effects the distribution of magnetic microspheres in the body.The magnetic content & the magnitude of magnetic field govern the retention of microspheres at the target site.In microspheres with high magnetic content, the external magnetic field strength required is less.

Two main techniques of preparation are:

phase separation emulsion polymerisation (PSEP)continuous solvent evaporation(SCE)

COMPARISION BETWEEN PSEP & CSE

PSEPCSEA narrow range of the chemotherapeutic agents can be incorporated into the microspheres by PSEP because this process requires aqueous solubility of the agent to be incorporated into the microsphere is essential prerequisite for this technique.Wide range of the chemotherapeutic agent could be covered by CSE techniques, due to availability of organic solvents with varying polarity.Disadvantage:CSE gives microspheres of size >50 which leads to problems of thromboembolism.

The release of drugs from biodegradable microspheres can be classified broadly into four different categories:Degradation controlled monolithic system,Diffusion controlled monolithic system,Diffusion controlled reservoir system,Erodible polyagent system.

In degradation controlled monolithic microsphere systems, the drug is dissolved in the matrix and is distributed uniformly throughout. The drug is slowly attached to the matrix and is released only on degradation of the matrix. The diffusion of the drug is slow compared with the degradation of matrix. When the degradation is done by the homogeneous bulk mechanism, drug release is slow initially and increases rapidly and then rapid bulk degradation starts. If the degradation is by heterogeneous mechanism, degradation confined to the surface. Release from a sphere is governed by the equation, where Mt is the amount of the agent released at the time t, M is the amount at time t & t is the time for total erosion. e.g. progesterone release from polymer films containing 10 weight% steroid. Mt/M = 1 - [(1- t/t)]3

Here the active agent is released by diffusion prior to or concurrent with the degradation of the polymer matrix. Degeneration of the polymer matrix affects the rate of release and has to be taken into account. Rate of release also depends on whether the polymer degrades by homogenous or heterogeneous mechanism.

Here the active agent is encapsulated by a rare controlling membrane through which the agent diffuses and the membrane erodes only after its delivery is completed. In this case the drug release is unaffected by the degradation of the matrix. Polymers that remain as such till the complete release of drug and then degrade by homogenous mechanism, so that the device is removed from the body is better for this type of delivery.

In this case the active agent is chemically attached to the matrix and the rate of biodegradation of the matrix is slow compared to the rate of hydrolysis of drug polymer bond. The rate of diffusion of active agent from the matrix to the surrounding is rapid, the rate limiting step is the rate of cleavage of bond attaching the drug to the polymer matrix

Targeting is achieved by exploiting the natural distribution pattern of a drug carrier called passive targeting or by changing the natural distribution pattern of the carrier by some means there by directing the drug to the specific organ or tissue; this is called as active targeting.

PASSIVE TARGETING: Particles administered into the body intravenously will distribute itself in different organs depending on the size of the particles.Particles

ACTIVE TARGETING: Active targeting includes coating of the microspheres with hydrophilic coating agents which suppress the opsonisation. When colloidal particles are administered into the blood stream, they may be coated with the proteins such as albumin, globulin etc., depending on the nature of the material surface charge & hydrophobicity of the particles. This is called as opsonisation. TARGETING USING MAGNETIC MICROSPHERES:Another approach in this area is by using magnetic microspheres. In this method the magnetic loaded microspheres. In this method the magnetic loaded microspheres is infused into an artery supplying a given target site.For example:- the anti-cancer effect of magnetic albumin microspheres containing Adriamycin in a rat model is found that the particles can be guided to the target site by magnetic means & a sustained release will be observed.

INTRACELLULAR TARGETING:Certain cytotoxic drugs are active intracellular, but are normally discarded due to their poor intracellular influx. The poor efficacy of many therapeutic substances for intracellular bacterial & parasitic therapy is known. The intracellular delivery of the drugs by suitable means can obviate these problems.For e.g. the biologically active streptomycin was released from albumin microspheres inside the phagocytic cells after ingestion and intracellular degradation of microspheres.

Advantages:-Incorporation of magnetically responses materials into microspheres makes them susceptible to applied magnetic field, so that they are concentrated to the target site by the application of magnetic field externally to that site. Due to this rapid clearance of these microspheres by RES is prevented. Difference occurs maximally in capillary network so efficient delivery of drug to diseased tissue is achieved.Microspheres can transit into extravascular space creating an extravascular depot of drug for sustained released of drug with in the target areas.Increase of tumour targeting microspheres can be internalizing by tumour cells due to its which increased phagocytic activity as compared to normal cell. So the problem of drug resistance due to inability of drug to be transported across the cell membrane can be prevented.

Disadvantages:One of the major limitations of this system is, the drug cannot be targeted to deep seated organism in the body. This approach is confined to the targeting of drugs to superficial tissues like skin, superficial tumours or to the joints.Thrombosis at the site of catharization.The unknown toxicity of magnetic beads.The possible unwanted localisation of the product in the liver and at the regions of RES and the dangerous effects of self-flocculation of the magnetic particles causing vascular obstruction to vital organs in the body.

Nanoparticles are defined as particulate dispersions or solid particles with a size in the range of 10-1000nm. The drug is dissolved, entrapped, encapsulated or attached to a nanoparticle matrix. The major goals in designing nanoparticles as a delivery system are to control particle size, surface properties and release of pharmacologically active agents in order to achieve the site-specific action of the drug at the therapeutically optimal rate and dose regimen.

To decrease the toxicity and occurrence of adverse rxnsBetter drug utilizationControlling the rate and site of drug releaseProvide a more predictable drug delivery systemProviding greater convenience and/or better patient complianceThe therapeutic effectiveness of the drug i.e. overall pharmacological response per unit dose is enhancedThey are reproducibleThey can be freeze dried, so obtain in a dried powder formNon toxic and biodegradableRelatively cheaper and stable

Due to their smaller size the nanoparticles disperse readily in water to form a clear colloidal dispersion which is suitable for injection through fine gauze needle.The preparation is sterile and apyrogenic.No gross change occurs when the gelatin nanoparticles were autoclaved at 121oC for 156 mints.The experimental data, the nanoparticles were found to be non antigenic.

Nanoparticles can be prepared from a variety of materials such as proteins, polysaccharides and synthetic polymers. The selection of matrix materials is dependent on many factors including:-Size of nanoparticles required; Inherent properties of the drug, e.g., aqueous solubility and stability;Surface characteristics such as charge and permeability;Degree of biodegradability, biocompatibility and toxicity;Drug release profile desired; andAntigenicity of the final product. Between the two phases leading to the formation.

Nanoparticles have been prepared most frequently by three methods:

(1) Dispersion of preformed polymers;(2) Polymerization of monomers; and (3) Ionic gelation or coacervation of hydrophilic polymers.

However, other methods such as supercritical fluid technology and particle replication in non-wetting templates have also been used in production of nanoparticles.

Dispersion of preformed polymers is a common technique used to prepare biodegradable nanoparticles from poly (lactic acid) (PLA).This technique can be used in various ways as described below.Solvent evaporation method: In this method, the Polymer is dissolved in an organic solvent such as dichloromethane, chloroform or ethyl acetate.The mixture is then emulsified in an aqueous solution containing a surfactant or emulsifying agent to form oil in water (o/w) emulsion. After the formation of stable emulsion, the organic solvent is evaporated either by reducing the pressure or by continuous stirring. In order to produce small particle size, often a high-speed homogenization or ultra sonication may be employed.

Spontaneous emulsification or solvent diffusion method: In this method, the water miscible solvent along with a small amount of the water immiscible organic solvent is used as an oil phase. Due to the spontaneous diffusion of solvents an interfacial turbulence is created between the two phases leading to the formation of small particles. As the concentration of water miscible solvent increases, a decrease in the size of particle can be achieved.

In this method, monomers are polymerized to form nanoparticles in an aqueous solution.Drug is incorporated either by being dissolved in the polymerization medium or by adsorption onto the nanoparticles after polymerization completed. The nanoparticle suspension is then purified to remove various stabilizers and surfactants employed for polymerization by ultracentrifugation and re-suspending the particles in an isotonic surfactant-free medium. Nanocapsules formation and their particle size depend on the concentration of the surfactants and stabilizers used.

In this method the preparation of nanoparticles using biodegradable hydrophilic polymers such as chitosan, gelatin and sodium alginate is done. The method involves a mixture of two aqueous phases, of which one is the polymer chitosan and the other is a polyanionsodium tripolyphosphate.In this method, positively charged amino group of chitosan interacts with negative charged tripolyphosphate to form Coacervates with a size in the range of nanometer. Coacervates are formed as a result of electrostatic interaction between two aqueous phases, whereas, ionic gelation involves the material undergoing transition from liquid to gel due to ionic interaction conditions at room temperature.

Drug loadingIdeally, a successful Nano particulate system should have a high drug-loading capacity thereby reduce the quantity of matrix materials for administration. Drug loading can be done by two methods:Incorporating at the time of nanoparticles production (incorporation method)Absorbing the drug after formation of nanoparticles by incubating the carrier with a concentrated drug solution. Macromolecule or protein shows greatest loading efficiency when it is loaded at or near its isoelectric point when it has minimum solubility and maximum adsorption.

Drug releaseTo develop a successful Nano particulate system, both drug release and polymer biodegradation are important consideration factors. In general, drug release rate depends on: (1) Solubility of drug(2) Desorption of the surface bound/ adsorbed drug(3) Drug diffusion through the nanoparticle matrix(4) Nanoparticle matrix erosion/degradation; and (5) Combination of erosion/diffusion process.

Paul Ehlrichs magic bullet concept has become a reality with the development of hybridoma tech. if possible to produce virtually unlimited quantity of homogenous antibodies having defined specifically i.e. monoclonal antibodies (MoAbs), which have found use insensitive immune diagnostic tests. The therapeutic use of MoAbs and their conjugates still in its infancy.

Antibodies are of two types:

1. They are produced without apparent antigenic stimulate Natural antibodies = antigen stimulant.

2. Those arising only on exposure to certain antigens.Acquired antibodies = exposure and certain antigens.

Types:IgG, IgA, IgM, IgE, IgO The specificity of particular antibody is determined by the amino acid sequence.Structure of antibodyThe antibody is y shaped molecule which contains two light chains and two heavy chains joined by the disulphide bond. Each of the heavy chain contains a carbohydrate residue. A bottom trunk portion of the antibody molecule is known as constant region since its amino acid sequence is similarThe upper arm, the antigen binding agent (Sab) is known as the variable region since the sequence is determined by the antigen response for its formation. The variable region in turn as several hyper variable regions is also known as complementary determining region (CDR) which shows greater variability than rest of the variable region. The base of the molecule, the Sc fragment retains the Antigenicity and is responsible for the recognition of the molecule by other components.

Monoclonal antibodiesAn antibody is called MONOCLONAL when each immunoglobulin is produced by a single clone of cells and hence is identical to every other molecule in the preparation in terms of heavy as well as chain structure.Polyclonal antibodiesAfter an Ag is injected into an animal by a region designated to induce an optimal immune response, serum can be collected and immunoglobulin fraction isolated. These antisera are enriched with antibody specific for the original Ag. Since large number of lymphocytes are involved in the production of the antisera, anti-bodies produced by this classical method are called POLYCLONAL

Targeting with Ab depends on the presence of new Ag from the tumor cells Ab the ability to obtain specific Ab against them in normal cell the antigen are absent.The antigen associated with tumor cells are called as the TUMOR MAKER. Antibodies produced as the results of the specific markers monoclonally can be conjugated with drug molecule which in turn can be targeted to the specific cells or tumor tissues.

Targeting antibodies with drugs involve the following steps:Identification of the antigen produced by the tumor cells. Production of antibody monoclonally against the identified antigen.Formation (or) producing drug antibody conjugate or complexes.These complexes concentrate at the tumor site and deliver the drug.Drug antibody conjugate advantage:There are several advantages when drugs are delivered as antibody conjugates. The conjugates can specifically reach the target cells without causing any damage to the normal tissue. The drug antibody conjugate could be expected to be the ideal agents for drug targeting in chemotherapy.

Steps:-

Mouse is immunized with antigen of interest (tumor specific antigen).Beta lymphocytes from the immunized animals spleen or lymph nodes are harvested into a single cell suspension.These cells are fused with myeloma (tumor cells) of the same species.Myeloma cells are immortal and have cytoplasmic machinery to survive longer. They contain an enzyme called as hypoxanthine guanine phosphoribosil tranferase (HGPRT), which is responsible for their survival in the presence of folic acid antagonist Aminopterin.Fusion is usually accomplished in PEG medium, which triggers fusion.The suspension of cells is distributed into the bore of microliter plate in a medium (tissue culture medium) containing hypoxanthine Aminopterin thymidine (HAT) medium where only the fused cells (hybridoma)survive due to the presence of (HGPRT).The hybrids are cloned by limited dilution of one cell per well (bore in the culture medium in which cell multiplication occurs).The cell supernatant is purified by column or affinity chromatography to get pure antibody.Alternatively the antibody producing cells can be injected into the peritoneal cavity of the mice for the production of ascites fluids, which can be purified to get antibodies.

Some aspects to be considered while preparing monoclonal antibodiesIn designing drug antibody conjugate, consideration must be given to the studies on the SAR of drugs as well as the methods of the conjugation.Most of the antineoplastic agents have complex chemical structures and their cell killing properties are sensitive to any chemical modification. For e.g. In this case of methotrexate, which is folic acid antagonist, the pteridine moiety with its free amino groups in position 4 should be intact in order to retain its activity. Drug carrying capacity of antibody:The no. of drug molecules carried per antibody molecules is usually too low to be highly effective. Increasing the no. of drug molecules conjugated to antibody will eventually destroy its antigen binding capacity.

References:

Controlled and novel drug delivery by N.K. JAINwww.google.com