35
Ashutosh Tiwari and Anis N. Nordin (eds.) Advanced Biomaterials and Biodevices, (487–522) 2014 © Scrivener Publishing LLC 487 14 Nanoparticles: Scope in Drug Delivery Megha Tanwar, Jaishree Meena and Laxman S. Meena* CSIR-Institute of Genomics and Integrative Biology, Delhi University Campus, Delhi, India Abstract Nanotechnology is the most expeditiously emerging technology in the field of therapeutics as drug delivery systems. It is the manipulation of matter on atomic and molecular scale. It works on devices, structures and materials with at least one dimension sized from 1 to 100 nanometers. Nanoparticle based technology has played a significant role in treatment and prevention of tuberculosis, cancer, etc. Recently a variety of nanocarriers has been evaluated as potential drug delivery systems for various administration routes. Liposomes and microspheres have been developed for sustained delivery of anti-TB drugs. Several anti-cancer drugs includ- ing paclitaxel, doxorubicin, 5-fluorouracil and dexamethasone have been success- fully formulated using nanomaterial. Targeting of drugs to certain physiological sites using nanoparticles has emerged a promising tool in treatment of tuberculosis with improved drug bioavailability and reduction of dosing frequency. Nanotechnology based targeting of drugs may improve the therapeutic success by limiting the adverse drug effects and resulting in more patient’s compliance and attaining high adherence level. Nanotechnology holds merit as a drug carrier because of its high carrier capac- ity, feasibility of incorporation of both hydrophilic and hydrophobic substances, high stability, and also because of feasibility of variable routes of administration like oral application and inhalation. e nanoparticle based drug delivery systems are advantageous over other modes of drug administration as nanoparticles deliver the drug more efficiently and with reduced side effects as well. Most recently, various vesicular systems have been developed such as niosomes which can be explored for achieving maximum effective concentration of the delivered drug. ey can be uti- lized for research on various drugs and may turn out to be most promising mode of drug delivery in treatment of various deadly diseases. Keywords: Nanoparticles, targeted drug delivery, nanomedicine *Corresponding author: [email protected]

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Page 1: Advanced Biomaterials and Biodevices (Tiwari/Advanced) || Nanoparticles: Scope in Drug Delivery

Ashutosh Tiwari and Anis N. Nordin (eds.) Advanced Biomaterials and Biodevices, (487–522)

2014 © Scrivener Publishing LLC

487

14

Nanoparticles: Scope in Drug Delivery

Megha Tanwar, Jaishree Meena and Laxman S. Meena*

CSIR-Institute of Genomics and Integrative Biology,

Delhi University Campus, Delhi, India

AbstractNanotechnology is the most expeditiously emerging technology in the fi eld of

therapeutics as drug delivery systems. It is the manipulation of matter on atomic

and molecular scale. It works on devices, structures and materials with at least one

dimension sized from 1 to 100 nanometers. Nanoparticle based technology has

played a signifi cant role in treatment and prevention of tuberculosis, cancer, etc.

Recently a variety of nanocarriers has been evaluated as potential drug delivery

systems for various administration routes. Liposomes and microspheres have been

developed for sustained delivery of anti-TB drugs. Several anti-cancer drugs includ-

ing paclitaxel, doxorubicin, 5-fl uorouracil and dexamethasone have been success-

fully formulated using nanomaterial. Targeting of drugs to certain physiological sites

using nanoparticles has emerged a promising tool in treatment of tuberculosis with

improved drug bioavailability and reduction of dosing frequency. Nanotechnology

based targeting of drugs may improve the therapeutic success by limiting the adverse

drug eff ects and resulting in more patient’s compliance and attaining high adherence

level. Nanotechnology holds merit as a drug carrier because of its high carrier capac-

ity, feasibility of incorporation of both hydrophilic and hydrophobic substances,

high stability, and also because of feasibility of variable routes of administration like

oral application and inhalation. Th e nanoparticle based drug delivery systems are

advantageous over other modes of drug administration as nanoparticles deliver the

drug more effi ciently and with reduced side eff ects as well. Most recently, various

vesicular systems have been developed such as niosomes which can be explored for

achieving maximum eff ective concentration of the delivered drug. Th ey can be uti-

lized for research on various drugs and may turn out to be most promising mode of

drug delivery in treatment of various deadly diseases.

Keywords: Nanoparticles, targeted drug delivery, nanomedicine

*Corresponding author: [email protected]

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488 Advanced Biomaterials and Biodevices

14.1 Introduction

Th e development of nanoparticles for drug delivery began in the1960s [1]. Recent developments in multifunctional nanoparticles has off ered a great potential for targeted delivery of drugs for treatment of various types of diseases. Nanoparticles are basically solid colloidal particles ranging in size from 1 to 1000nm (μm). Nanoparticles are made from biocompat-ible and biodegradable materials such as polymers, either natural (e.g. gelatin, albumin) or synthetic (e.g. polylactide, polyalkylcyanoacrylates) or solid lipid nanoparticles [2]. Th e drug loaded in nanoparticles is usu-ally released from the matrix by diff usion, swelling, erosion or degrada-tion. Th e reason, why nanoparticles used for medical purposes is large surface to mass ratio which is much larger than other particles. Large surface area provides ability to bind, adsorb and carry other compounds such as drugs, proteins, probes. Th ey consist of macromolecular materials in which active agent (drug or biologically active material) is dissolved, entrapped, encapsulated or to which active agent adsorbed or attached [3]. Unfortunately the conventional therapeutic strategies require unnecessar-ily high systemic administration due to non-specifi c biodistribution and rapid metabolism of free drug molecules before reaching their targeted sites. Th erefore nowadays nanotechnology based targeting improved the therapeutic success by limiting adverse drug eff ects and patient requires less frequent administration regimes, which ultimate results in more

Oral

Routes of

administration

of various

drug delivery

systems

Inhalable

Injectable Implantable

Figure 14.1 Various Routes of Drug Delivery Systems.

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Nanoparticles: Scope in Drug Delivery 489

patients compliance and thus attaining higher adherence levels [4]. Th ere are many advantages of nanoparticles as drug carriers including high stability, high carrier capacity, and feasibility of incorporation of both hydrophilic and hydrophobic substances, feasibility of variable routes of administration including oral administration and inhalation and reduc-tion in dosing frequency [5]. Mainly uptake of nanoparticles occurs by three mechanisms (1) transcytosis (2) intracellular uptake and transport through the epithelial cells lining the intestinal, mucosa, (3) uptake via peyer’s patches [6, 7].

Th ere are various routes of administration of drug delivery systems as described above in Figure 14.1, which are advantageous in one or another way.

1. Oral drug delivery-Most acceptable route for drug delivery, substantial reduction in dosing frequency, economic and ease of preparation [8].

2. Implantable drug delivery-High drug bioavailability and least dosing frequency [8].

3. Injectable drug delivery-It also has high drug bioavailabil-ity and least dosing frequency but painful procedure [8].

4. Inhalable drug delivery-Direct drug delivery to the target site, reduction in dosing frequency and improved bioavail-ability [8].

14.2 Diff erent Forms of Nanoparticles as Drug Delivery

Nanoparticles are used in various forms for drug delivery such as nano-spheres, nanoemulsions, nanocapsules, solid lipid nanoparticles, nanosus-pensions, polymeric nanoparticles, liposomes and micelles, dendrimers and niosomes. Monolithic nanoparticles (nanospheres) are those in which drug is adsorbed, dissolved or dispersed throughout the matrix and nano-capsules are those in which drug is confi ned to an aqueous or oily core sur-rounded by shell like wall [9]. Nanoemulsions referred as miniemulsions or sub-microemulsions by dispersing mainly oil in water. Th ermodynamically stable nanoemulsion (mean size 80.9 nm) of ramipril was developed for oral administration. In vitro studies showed that drug release till 24h from nanoemulsion band was much more important as compared to marketed capsule formation and drug suspension [4]. Th e relative bioavailability of ramipril nanoemulsion to that of conventional capsule was 229.62% and to

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490 Advanced Biomaterials and Biodevices

that of drug suspension was 539.49% which suggested the use of developed ramipril nanoemulsion for paediatric and geriatric patients [10].

Solid lipid nanoparticles are lipid-based submicron colloidal carriers. Th ey were initially designed as pharmaceutical an alternative to liposomes and emulsions. In case of solid lipid nanoparticles, the drug is entrapped in solid lipid matrix to produce lipid nanoparticles of size range 50–100nm by using hot or cold high pressure homogenization technique [4]. Th ey are more stable than liposomes because of their rigid core consisting of hydro-phobic lipids which are solid at room and body temperature, surrounded by a monolayer of phospholipids. Th ey can be stabilized by high level of sur-factants. Th ey are less toxic than polymeric nanoparticles because of ease of biodegradation. Solid lipid nanoparticles have important advantages, such as their physiological compounds and large scale production favoured their feasibility thus avoiding organic solvents in the manufacturing process [11].

Nanosuspensions are poor water soluble drugs dispersed in aqueous phase containing stabilizing agent [4]. Clofazimine, a riminophenazine compound used for treating patients with M. avium infection and because of its poor solubility the drug usage was restricted, now to overcome this problem of solubility. It was formulated as a nanosuspension (385 nm) and was administered to mice intravenously which has resulted in reduction of bacterial loads in the liver, lungs, spleen of mice [12].

Reduce

irritation

Liposomal

drugsFree drug

Stratum

Corneum

Epidermis

Dermis

Blood

supply

Enhance drug

permeation

Prolong

resistance time

Reduce systemic

toxicity

Figure 14.2 Liposomal drug and free drug delivery compared.

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Nanoparticles: Scope in Drug Delivery 491

Liposomes are small spherical vesicles formed of amphiphilic lipids enclosing an aqueous core. Th ey are mainly carrier systems for hydro-philic drugs [13, 14]. Th e fi rst formulation of liposomes was prepared in 1986 by the Christian Dior Laboratory in collaboration with Pasteur Institute. Th e amphiphilic nature of liposomes, the ease of surface modifi -cation and good biocompatibility make them good drug delivery systems. Liposomes are composed of mixture of lipids such as phosphatidylcholine, cholesterol, diacetylphosphate-o-steroylamylopectin, monosialoganglio-side, distearylphoshatidylethanolaminepoly-(ethylene glycol) for targeted delivery of anti-TB drug to the lung [15]. Th e drug encapsulated lipo-somes signifi cantly help in reduction of the bacterial count in spleen and liver as compared to free drug [16]. Several diff erent kinds of liposomes are being widely employed for cancer treatment and for delivering cer-tain vaccines. During cancer treatment they encapsulate drugs, protecting healthy cells from their toxicity and prevent their concentration in tis-sues such as patient’s kidney, liver, pancreas showing details in Figure 14.2 ( previous page).

Th ey also reduce side eff ects related to chemotherapy such as hair loss and nausea [17]. Polymeric nanoparticles have also emerged as a prom-ising tool for targeted and controlled drug delivery. In case of polymeric nanoparticles the drug is encapsulated or entrapped in polymeric core and depending upon method of preparation they are called nanospheres or nanocapsules [18]. Polymeric nanoparticles include those made from chitosan, gelatin, poly-(lactide-co-glycolide), copolymer, polylactic acid, poly-(cyanoacrylate), poly-(methylacrylate). Th ese nanoparticles can be functionalized into other types of nanoparticles to change and improve their biodistribution properties. Th e surface property mainly plays an impor-tant role in drug targeting. When they come in direct contact with cellular membranes their surface properties help in determining the mechanism of internalization and intracellular localization. Biodegradable polymeric nanoparticles such as PGA and PLGA can be formulated to encapsulate several classes of therapeutic drugs but not limited to low molecular weight compounds. Th ey represent as an alternative to liposomes with much improved drug delivery to target sites and with lesser side eff ects. Recently stimuli responsive polymeric nanoparticles have been demonstrated that response to an external or internal stimulus such as pH, redox, magnetic fi eld and light [19]. Pandey et al, developed sustained release RIF, INH and PYZ loaded poly-(lactide-co-glycolide) [PLG] nanoparticles for oral deliv-ery in mice. As a result the drugs were detected in the plasma for up to 4 days for RIF and 9 days for INH and PYZ; whereas in the tissues were detected till 9 to 11 days aft er single oral administration of nanoparticles

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492 Advanced Biomaterials and Biodevices

whereas free drugs were cleared within 12–24h from the plasma. Five oral doses were suffi cient for completely bacterial clearance whereas free drugs took 46 doses to get the same results [20]. Injectable PLGA (poly lactic-co-glycolic acid) nanoparticles were also administered subcutaneously in a murine model. A single subcutaneous dose of PLGA nanoparticles main-tains drug levels in plasma, lungs, and spleen for <1 month and bacterial count remain almost undetectable in these organs [21].

Niosomes are similar to liposomes and are mainly composed of non-ionic surfactant with or without incorporation of lipids. Recently nio-somes were prepared by reverse phase evaporation method and given a charge with a charge-inducing agent, dicetyl phosphate. Drug entrap-ment effi ciency was determined by spectrophotometer. In vitro drug release and cellular uptake studies was also carried out on macrophage J774A. As a result the cellular uptake of drug loaded by macrophage cells was as high as 61.8% a level which is capable for eff ective treatment of tuberculosis [22].

Micelles are sub microscopic aggregates (20–80 nm) of surfactant mol-ecules resulting in liquid colloid [8]. PLA (poly lactic acid) modifi ed chi-tosan oligomer micelles release 35% drug release within 10h followed by more sustained drug release till 5 days suggesting the role of micelles as drug carrier with reduced side eff ects. Alginate nanoparticles are also used as drug delivery for tuberculosis treatment. Zahroor et al used ionotropic gelation method for preparing alginate nanoparticles (235nm) of anti-TB drug. Oral administration of drug to mice resulted in detection of level free drugs in tissues till next day. Whereas in plasma alginate nanoparticles were detected up to 7 days for ETB (ethambutol), 9 days for RIF, 11 days for INH and 15 days for PYZ in tissues [24]. Gold Nanoparticles, Mesoporous Silica Nanoparticles, Quantum Dots are also employed for detection, imag-ing and treatment of various diseases.

Gold nanoparticles preparation involves the chemical reduction of gold salts in aqueous, organic, or mixed solvent system. Under these condi-tions gold surface are extremely reactive as a result aggregation occurs. To reduce aggregation, gold nanoparticles were reduced in the presence of a stabilizer which binds to the surface and remove aggregation via cross link-ing and charge properties. Gold nanoparticles are also used for biological imaging and sensing. A simple gold nanoparticle probe assay was done to detect Mycobacterium tuberculosis and its complex in clinical specimen. An assay using gold nanoparticles derivatized with thiol modifi ed oligo-nucleotides was carried out. Th e gold nanoparticle probes GP-1/GP-2 for IS6110 and GP-3/GP-4 for Rv3618 were designed to hybridize with target DNAs of MTB and MTBC strains. Th en the effi ciency of gold nanoparticle

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Nanoparticles: Scope in Drug Delivery 493

probes assay was evaluated directly, detecting both MTB and MTBC from 600 clinical sputum specimens [25].

Liposomes and solid lipid nanoparticles because of their poor chemical stability and degradation by the serum resulted in decrease in the drug delivery to the target cells and increasing the potential systemic toxicity [26]. Due to some of the disadvantages related to the liposomes and solid lipid nanoparticles, there was a discovery of mesoporous silica nanopar-ticles. Mobil fi rst discovered the mesoporous silica based nanomaterials MCM-41 in 1992. Th ey are 100nm sized silica nanoparticles with 2nm sized pores. Th ese pores run parallel through mesoporous sized nanopar-ticles and form hexagonal pattern [27]. Th ese biocompatible solid mesopo-rous silica nanoparticles framework of hexagonal pattern provides it more intrinsic stability as compared to the liposomes, copolymers or polymeric nanoparticles drug delivery platforms. Some of the in vivo experiments resulted in more enhanced blood stability of mesoporous silica nanopar-ticles as compared to liposomes and polymeric nanoparticles and have favourable biodegradability, biocompatibility and excretion properties [28, 29, 30].

14.3 Tuberculosis Targeting Nanoparticles

Tuberculosis is a ubiquitous and highly contagious chronic granulomatous bacterial infection. It is one of the leading causes of mortality and mor-bidity worldwide. In the year 1993, World Health Organization (WHO) declared tuberculosis as a global emergency. Tuberculosis is caused by M. tuberculosis, which infects about one third of the World’s population and causes approximately 9 million new cases of active tuberculosis and 1.7 million deaths annually [31]. Most of the cases occurred in South East Asia (55%) and the African regions (30%). Th e fi ve countries with largest num-bers of the cases include India, China, South Africa, Nigeria and Indonesia. Out of the 9 million new cases of TB, 15% were HIV positive, 78% of these HIV positive cases were in the Africa and 13% of the cases were in South East Asia regions [8]. It serves as second most common cause of death aft er HIV/AIDS. Treatment of active TB is becoming more and more complex due to the emergence of MDR-XDR strains. Th e identifi cation of multidrug-resistant (MDR) strains, makes mycobacteria resistant to at least Rifampicin (RIF) and Isoniazid (INH) (two fi rst line anti-TB drugs) and extensively drug resistant (XDR) strains defi ned as MDR strains with additional resistance to the fl uoroquinolone. MDR-TB has become one of the major obstacles to eff ective global TB control. To solve these problems

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494 Advanced Biomaterials and Biodevices

of drug resistance WHO (World’s Health Organization) implemented DOTS (Directly Observed Treatment, short course), program which was not successful in solving the patients non-compliance [32]. As Figure 14.3 below discloses, most patients fail to adhere to multidrug therapy for lon-ger periods of time.

Tubercle bacilli are slender, acid fast, non-motile gram-positive bacilli. Th ese bacteria remain viable in the air for longer period of time as a result of which they get inhaled by the lungs and engulfed by alveolar macro-phages (white blood cells) where they start replicating within 2–3 weeks [33]. If these bacteria is not completely destroyed, they remain dormant for several days and may reactivate years later. Th e cell wall has high lipid content which allows the bacteria to survive within macrophages. Because of the impermeable nature of the cell wall, drugs are only partially eff ective and organisms develop resistance [4]. Th e bacilli can spread from the site of infection in the lung through the lymphatic or blood to the other parts of the body. Extra pulmonary TB of the pleura, bone, genitourinary sys-tem, skin or peritoneum, meninges occurs in approximately 15% of the TB patients [8]. Risk of development of active disease varies according to time according to infection, age, host immunity, however life time risk of death for a newly infected child has been estimated 10% [34–36]. Available TB treatment involves daily administration of four oral antibiotics for a period

Free

Mycobacterium tuberculosisreleased from granuloma center

Formation of caseum

(cells are in dormant phase)

Formation of fibrous cuff made up of

extracellular matrix around

foamy macrophages

Differentiation of infected

macrophages into

foamy macrophages

Infected macrophages entered

into blood vessel

Life Cycle of

Mycobacterium tuberculosis

Mycobacterium tuberculosisfrom air get engulfed by

alveolar macrophages

Figure 14.3 Lifecycle of Mycobacterium tuberculosis.

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Nanoparticles: Scope in Drug Delivery 495

of six months or more. Th e daily administration of antibiotics has side eff ects like nephrotoxicity and ototoxicity and longer duration of treatment has resulted in low patience adherence [37]. To improve the treatment and reduction in death cases due to tuberculosis, various chemotherapeutic strategies against TB include :- (1)-Derivatization of the existing Anti-tubercular drugs (ATDs) into more potent compounds. (2)-Screening of the compounds which are active against replicating as well as latent bacilli. (3)-Identifi cation of novel drug targets and designing of appropriate inhib-itors. (4)-Targeting of host-pathogen related processes which are essential for the survival in the human diseases [38].

Th e currently used vaccine for the tuberculosis treatment is Bacillus-Calmette-Guerin (BCG) which provides extremely limited protection. Recently, several Mycobacterium tuberculosis antigens have been identi-fi ed for potential use as vaccine antigens, including three protein Antigen 85 complex (Ag85A, Ag85B and Ag85C), the surface exposed lipoproteins PstS (PstS-1, PstS-2 and PstS-3) and early secretary antigenic target pro-tein ESAT-6 [39–41]. Cationic liposomes entrapped antigenic tuberculosis ESAT-6 protein, complexed with TLR agonists was evaluated as a prophylac-tic vaccine system by Zaks research group [42]. Th ese control measures for TB such as BCG vaccination and chemoprophylaxis has proven to be unsat-isfactory, so there come anti-TB drugs the only option for TB treatment. Th e goal of treatment with drugs is to cure without relapse, to prevent death, to stop transmission, and to prevent the emergence of drug resistance [43]. As suggested by WHO, treatment of TB and drug resistant cases required multi-drug therapy, comprising (a): an initial intensive phase of rifampicin (RIF), isoniazid (INH), pyrazinamide (PYZ), ethambutol (ETB) for 2 months; and (b): a continuation phase of RIF and INH for further 4 months, either daily or 3 times per week has to be administered [44]. (Figure 14.4)

14.3.1 Action of anti-TB drugs

Rifampicin (RIF)-Inhibition of bacterial RNA synthesis by binding to the β subunit of bacterial DNA dependent-RNA polymerase, as a result it inhibits RNA synthesis. It is one of the most eff ective anti-tuberculosis agents and is bactericidal for extra and intracellular bacteria [45, 46].

Isoniazid (INH) - It is most active drug for treatment of TB caused by susceptible strains. It is a prodrug activated by katG, which exerts its lethal eff ects by inhibiting the synthesis of mycolic acids, an essential component of mycobacterial cell walls through formation of covalent complex with an acyl carrier protein (AcpM) and KasA, a beta-ketoacyl carrier protein synthetase [45, 47].

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496 Advanced Biomaterials and Biodevices

Pyrazinamide (PYZ) - It gets converted into pyrazanoic acid, which lowers the pH of the surroundings of M. tuberculosis and thus organism unable to grow. It is also antimetabolite of nictoniamide and interferes with the synthesis of NAD, thus inhibiting synthesis of short chain fatty acid precursors [45, 47].

Ethambutol (ETH) - It inhibits mycobacterial arabinosyl transferases involved in the polymerization of D-arabinofuranose to arbinoglycan, an essential cell wall component of mycobacterium [45, 47].

INH, ETB along with streptomycin helps in eradicating most of the rapidly replicating bacilli in fi rst 2 weeks of treatment. RIF and PYZ are two drugs which play an important role in sterilisation of lesions by eradi-cating organisms, and are crucial for 6 months treatment regimens. RIF is responsible for killing non-replicating organisms and high sterilising eff ect of PYZ act on semi-dormant bacilli and not aff ected by any other TB agents, in sites hostile to the penetration and action of the other drugs [48, 49].

Despite of having availability of highly eff ective treatments of TB, cure rates remain low. Patients have to consume large amount of drugs, the major cause of patient’s non-compliance. Th ese short course regimens results in decrease of the therapeutic potential of patient resulted in escala-tion in the mortality rate and increased risk of developing acquired drug

Mycolic acid

Arabinogalactan

Short chain fatty acid

precursors

RNA polymerase

(beta subunit)

Systematic diagram for the site of action of principle anti-TB drugs

Cyt

op

lasm

Ce

ll w

all

an

d c

yto

pla

smic

me

mb

ran

eINH

ETB

PYZ

RIF

Figure 14.4 Site of action of principal anti-tuberculosis drugs against M. tuberculosis H37

Rv.

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Nanoparticles: Scope in Drug Delivery 497

resistance [50, 51, 52]. Resistance of M. tuberculosis to anti-TB agents is a worldwide problem in both immunocompetent and HIV-infected popula-tions [53, 54]. Th ough many new antibiotics have come into existence but treatment of such intracellular pathogens still remains a problem as the infection remain localized within phagocytic cells and most of the antibi-otic are highly active in vitro, so they do not actively pass through cellular membrane and therefore it’s diffi cult to achieve the relatively high concen-tration of the drugs within the infected cells [55, 56].

To solve this problem of intracellular chemotherapy there is a need to design such a carrier system for antibiotics that could effi ciently endocy-tosed by phagocytic cells and once inside the cells should prolong release of antibiotics so that the number of doses frequency and drug toxicity can be reduced as in Figure 14.5 (above).

All these problems associated with the chemotherapy led to the inves-tigation of drug carriers for treating intracellular pathogens such as anti-biotics loaded into liposomes, microspheres, polymeric nanoparticles, and nanoplexes [57, 58]. Present eff orts are being in progress in improving treatment of diseases by shortening time period of treatment or using new carrier based drug delivery strategies in addition to alternative adminis-tration routes, which have important role in improving anti-tubercular chemotherapy effi cacy, thus enhancing patient’s compliance, and reduc-ing dosing frequency. Nanotechnology can be defi ned as formation of the

Encapsulation

Nanoparticle

Nanoparticle get

endocytosed by the

cell

Degradation of nanoparticle to

release drug

Uptake of nanoparticle through intracellular

membrane; localisation and internalisation

Nanoparticle with drugDrug

Figure 14.5 Systemic uptake of nanoparticle through intracellular membrane,

localization and internalisation.

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498 Advanced Biomaterials and Biodevices

submicron colloidal particles, which has become a great advancement in the drug delivery [9]. Modifi cation of new drugs as a nanoparticle-based delivery system is feasible, cost-eff ective, and readily available alternative to chemotherapy. In turn, nanoparticle carrier based drug delivery system hold a signifi cant importance in the reduction of drug resistance TB cases. Th is system of drug delivery through nanotechnology enhances the eff ec-tiveness of approved drugs and extends their applicability by overcoming technological limitations, such as low availability, resistance, cellular and anatomical barriers, among others [59].Diff erent nanoparticulate strategies are being developed to specifi -cally deliver chemotherapeutic compounds to target disease sites. Th ese nanoparticles increase drugs therapeutic index by localizing its pharmaco-logical activity on its target site or organ of action. Th e particle size between 50 to 200 nm is desired for maximum drug localization upon administra-tion by inhalation [60]. In case of lungs, particles deposition takes place by inertial impaction, sedimentation and diff usion [61, 62]. Large particles (>5μm) get deposited by impaction in the extra-thoracic cavities, particles (1–5μm) deposited deeper in the lungs by inertial impaction and sedimen-tation while very small particles (<1μm) are taken up by diff usion and the most of the particles which remain suspended are exhaled out [63].

Recently, the microencapsulation of pharmaceutical substances in bio-degradable polymers is an emerging technology. Many natural and syn-thetic carriers are used in drug delivery systems. Natural carriers mainly include lipids (liposomes and solid lipid nanoparticles), alginic acid, gela-tin, dextrins etc whereas synthetic carriers include poly-(DL-lactide-co glycolide) (PLG), polylactic acid (PLA), polymethyl acrylates, polyanhy-drides, carbomer etc. Carriers not only help in designing diff erent delivery system but also provide fl exibility for selecting the route of drug delivery system [64]. PLG (poly-DL-lactide-co-glycolic acid), copolymer of lactic acid and glycolic acid is completely biodegradable, biocompatible and has role in medical procedures as well as used for encapsulating antibiotics, antigens, peptides in order to develop sustained-release delivery system [65]. Various nanoparticulate systems include polymeric nanoparticles, lipid nanoparticles, nanosuspensions, nanoemulsions etc can be used to treat several types of parasitic infections [66–70]. Dutt and Khuller have entrapped anti-TB drugs such as INH and RIF in PLG polymers. When these nanoparticles were injected subcutaneously as a single dose in mice, the microparticles with a diameter range of 11.75μm to 71.95μm resulted in sustained release of drugs over 6–7 weeks [71]. Sheogkar explored nanoparticulate systems for the anti-TB therapy and briefl y described three drugs under clinical trials [72]. Sosnik et al. reviewed the

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Nanoparticles: Scope in Drug Delivery 499

development of nano-based drug delivery systems for encapsulation and release of antitiberculosis drugs [73]. Pandey et al. reported the formula-tion of three anti-TB drugs i.e. RIF, INH, and PYZ encapsulated in PLG nanoparticles. In M. tuberculosis infected mice, aft er oral administration of drug loaded nanoparticles at every 10th day, resulted in no detection of the tubercle bacilli in the tissues aft er 5 oral doses of treatment. Th ese oral nano based drug delivery of anti-TB drugs has resulted in reduction of dosing frequency and better management of tuberculosis [20]. Prabakaran developed an osmotically regulated capsular multi-drug oral delivery sys-tem made of asymmetric membrane coating and dense semipermeable membrane coating-capsular systems for the controlled administration of RIF and INH for the treatment of TB [74]. Various anti-TB drugs have been formulated in dry microparticles for pulmonary delivery of drugs. Th ese microparticles provided promising strategy in targeting those TB sites, which can be directly administered to the lungs with reduced sys-temic side eff ects.

According to the experiment three formulations of PLG (each encap-sulating rifampicin) were developed i.e. non-porous (based on their drug release behaviour), hardened (based on the use of polyvinyl alcohol as a hardening agent) and porous. Out of three, the hardened PLG microcap-sules which showed 12–14% encapsulation of rifampicin and sustained drug release for 42 days in all the organs can be used as controlled drug delivery [75].

Nanoparticle with drug released

in blood capillary

Blood capillary

Drug release on degradation

Endolytic

vesicle

Lysozyme

Release of nanoparticle encapsulated

drug into the infected macrophage

Figure 14.6 Release of nanoparticles encapsulated drug into infected alveolar

macrophages of human.

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Liposomes are widely used drug carriers for macrophage-specifi c antibacterial drug delivery (see Figure 14.6 above). In one of the experi-ment, streptomycin loaded liposomes were intravenously injected into the infected mice led to the decrease of the mycobacterium count in the spleen but not in the lungs but prolonged mouse survival and reduced drug toxicity was observed as compared to the free drugs [76]. According to the Klemens et al, gentamicin loaded liposomes were evaluated for the antibacterial activity in M. avium infected mouse model. It signifi -cantly reduced the bacterial count in spleen as well as liver compared to free drug [77]. Deol and Khuller developed stealth liposomes for the targeted delivery of anti-TB drugs to the lungs. Liposomes composed of phosphatidylcholine, cholesterol, dicetylphosphate-o-steroylamylopectin and monosialogangliosides / distearylphosphatidylethanolaminepoly (ethylene glycol) 2000. Aft er intravenous administration in healthy and tuberculosis infected mice, increase in accumulation from 5.1% for con-ventional liposomes to 31% for Poly ethylene glycolated liposomal sys-tems aft er 30 min was observed. Drug uptake levels in the lungs increased to approximately 40% for the poly ethylene glycolated nanocarriers when administered to pretreated infected animals aft er 30 min 3[78]. Ahmed et al. developed various nanoemulsions of RIF (47 and 115nm) using GRAS listed excipients (US-FDA). As a result the entrapment effi -ciency was 99% with excellent stability over 3 months with slight increase in particle size and initial burst drug release of 40–70% aft er 2h [79]. Anisimova et al. encapsulated RIF, INH and streptomycin within poly-(n- butylcyanoacrylate) (PBCA) and poly-(isobutylcyanoacrylate) (PIBCA) nanoparticles and their accumulation in the human blood monocytes was tested in vitro [80]. Econazole and moxifl oxacin loaded PLG nanopar-ticles were prepared by the multiple emulsion and solvent evaporation technique. As a result the drug levels in lungs, liver and spleen lasted till 6 days as compared to the pure drugs which were cleared within 12–24 h. In M. tuberculosis infected mice, only 8 doses of polymeric nanopar-ticles were suffi cient to suppress bacterial clearance as compared to the pure drug which requires 56 doses daily of moxifl oxacin and 112 doses of econazole twice a day. Further to improve treatment there was addition of third drug for tubercular chemotherapy [81].

Encapsulation of RIF, INH, and PYZ in alginate microspheres and oral administration to guinea pigs, maintain the drug concentration in plasma for 4–5 days and in the organs for 7–9 days. Weekly treatment of alginate microspheres resulted in the complete bacterial clearance in the organs of infected guinea pigs aft er 8 oral doses daily as compared to the admin-istration of free drugs [82]. Alginate-chitosan coated microcapsules were

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developed as oral sustained delivery carriers for the anti-TB drugs. Th ese microparticles were developed using ionotropic/external gelation method. Microparticles were formulated into three diff erent forms containing rifampicin, isoniazide and pyrazinamide in the ratio of 1:2:2 (drug: sodium alginate: chitosan). Th ese prepared microcapsules were then evaluated by SEM analysis, size analysis, spherecity, drug content, encapsulation effi -ciency, swelling studies and mucoadhesion which was then compared to the pure drug. In vitro release studies were carried out and the amount of drug released was analysed and was compared with the free drug [83].

Sung tested PA-824 (an alternative anti-TB candidate) which resulted in sustained release of the drug and maintained its level in the lungs for 32h [84]. PLG nanoparticles encapsulating anti-TB drugs such as PYZ, RIF, INH and ETB remained in the circulation for 72h. As a result, PLG encap-sulated INH was found to be higher than its MIC value (0.1mg/ml) [85]. A single subcutaneous dose of PLG encapsulated nanoparticles maintained drug level in the plasma, lungs, and spleen concentrations for more than 1 month and led to undetectable bacterial counts in the diff erent organs [86]. Stearic acid encapsulating RIF, INH and PZA drugs nanoparticles aft er a single oral administration, the therapeutic concentration was maintained in the plasma for 8 days and in the organs for 10 days whereas free drugs get cleared within 1–2 days [87].

Th e nebulization of the drug loaded PLG nanoparticles with anti-TB drugs RIF, INH, and PZA was detected in plasma aft er 6h and therapeutic concentrations were detected until day 6 for RIF and day 8 for both INH, PZA. Nebulization of the nanoparticles to the M. tuberculosis-infected guinea pigs at every 10th day, no detection of the tubercle bacilli in the lungs was observed aft er only 5 doses of treatment, whereas 46 daily doses of orally administered drug required obtaining an equivalent therapeutic benefi t [88].

In tracheal administration of Ofl oxacin-loaded hyaluronan particles resulted in 50% lower serum bioavailability with respect to the intrave-nous or oral ofl oxacin. Th is observation led to the conclusion that inhaled nanoparticles reduce systemic side eff ects, but it also suggested that extra pulmonary TB cannot be treated only by the inhaled therapies [89]. Saraogi et al prepared mannosylated gelatin nanoparticles for the selective delivery of INH to the alveolar macrophages and concluded that these nanopar-ticles can be a potential carrier for the safer and effi cient management of TB through targeted drug delivery [90].

Ohasi et al designed RIF loaded biodegradable PLGA nanoparticles which were incorporated into the mannitol microspheres in single step by means of a four-fl uid nozzle spray drier. As a result due to mannitol, the in

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vivo uptake of the drug by alveolar macrophages in rat lungs was improved as compared to the RIF containing PLGA [91].

Chitosan has also been used for the development of eff ective drug deliv-ery systems because of its unique physiochemical and biological proper-ties. Th e primary hydroxyl and amine groups located on the backbone control its physical properties. Th e small size of chitosan nanoparticles favours intravenous administration of drugs to target sites [92]. Th ey are widely used as drug delivery systems for low molecular drugs, peptides and genes [93, 94].

Machida et al evaluated the potential of lactosaminated N-succinyl-chitosan (Lac-Suc) synthesized by the reductive amination between the N-succinyl-chitosan and lactose in the presence of sodium cyanoborohy-dride, as a liver specifi c drug carrier [95]. Recently, Liu et al prepared the polyion complex micelles (PIC micelles) based on methoxypoly-(ethylene glycol) (PEG)-graft -chitosan and lactose-conjugated PEG-graft -chitosan for targeted delivery of diammonium glycyrrhizinate (DG) to liver [96].

Chitosan nanoparticles with PEG have gain attention because of its potential in the therapeutic applications [97]. PEGylation mainly increases physical stability and prolongs their circulation time in blood. Recently, the eff ect of PEG conjugation on PTX loaded N-octyl-sulfate chitosan nanoparticles were investigated by Qui et al. Th ey found that these con-jugated particles were less phagocytised as compared to the unconjugated nanoparticles by the reticuloendothelial system. Th ey have also been investigated as carriers for the delivery of diff erent types of small molecules drugs such as paciltaxel, camptothecin, methotrexate and all trans-retionic acid (ATRA) [98–102].

Dendrimers are well defi ned, regularly hyper branched and 3D archi-tecture having relatively low molecular weight, polydispersity and high adjustable functionality. Th e unique structure is responsible for the encap-sulation and delivery of anti-TB agents. Kumar et al developed manno-sylated polypropylimine dendrimeric nanocarriers for the delivery of RIF to macrophages. Th e RIF loaded dendrimers in alveolar macrophages in lungs of rat showed an increase in the intracellular concentration of the antibiotic [103].

Th e therapeutic drugs with polymeric nanoparticles and solid lipid nanoparticles, results in more sustained drug release and the ability to tar-get specifi c cells and organs. Delivery of the drugs to the lungs has to face many challenges such as formulation instability due to particle-particle interactions and poor delivery effi ciency due to the exhalation of low iner-tia nanoparticles. Concerning all these problems led to the invention of novel methods of formulating nanoparticles into the form of micron scale

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dry powders. Th rough these nanoparticles the lungs can be targeted for the drug delivery to specifi c lung cells such as alveolar macrophages for the treatment of tuberculosis [104].

Signifi cant advances in medical aerosol development began in the 1950s with a focus on the delivering of the asthma drugs directly to the lungs, the target organ, thereby resulting in reduction in the amount of systemic drug and their adverse eff ects [105]. Since mid-1800s nebulizers have been used for delivering solutions of the drugs to target site, by generation of aerosol droplets from liquid with the help methods such as ultrasonic or air jet technology.

Recently, nanoscale aerosol vaccines have also been developed which perfuse more throughout the respiratory tract and also increases the amount of drug reaching to the target alveoli. Garcia ontreas et al. have recently synthesized a particle system both micrometer and nanometer dimensions for aerosolized delivery of the attenuated tuberculosis vaccine, BCG. Th e aerosol delivery of BCG encapsulated nanomicroparticles in guinea pigs increased their resistance to tuberculosis infection, and gen-eration of better immune protection than a standard parenteral BCG for-mulation [106]. Th e nanoparticulate delivery of aerosolized IFN-gamma through the pulmonary route has been more effi cient new adjunct treat-ment for tuberculosis [107].

Para-aminosalicyclic acid (PAS) is a tuberculostatic agent recently being formulated into large porous particles for direct delivery into the lungs via inhalation. Th ese particles possess some optimized physical properties for deposition through respiratory tract; the drug was loaded with 95% by weight over 4 weeks at elevated temperatures. PAS concentrations were measured in the plasma, lung lining fl uid and homogenized whole lung tissue. As a result the PAS get cleared within 3h from the lung lining fl uid and plasma. Th e above experiment led to the conclusion that the inhala-tion delivery of PAS help in the reduction of total dose delivered [108]. Dry powder inhalation systems have also come into existence with a potential of storing the drug in a dry state, which confers long term stability and sterility. Th e fi rst type of dry powder inhalation system utilized the patients breathing system for dispersing and delivering of the milled micron sized particles. Th e large geometric sized particles improved the dispersion properties and higher lung deposition of the delivered dose (upto 59%) [109]. Vyas formulated aerosolised liposomes with the incorporation of RIF through a cast-fi lm method. Th e liposomes coated with the alveolar macrophage-specifi c ligands resulted in the more accumulation in alveolar macrophages, thus maintaining high concentrations of RIF in the lungs, even aft er 24h [110].

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Recently, encapsulation, characterization and in vitro release of anti-TB drug (RIF) using chitosan-polyethylene glycol nanoparticles were devel-oped. Chitosan polyethylene glycol 600 (PEG) nanoparticles were pre-pared by ionic gelation technology. Th e preparation of these nanoparticles is based on the interaction between positively charged chitosan solution and negatively charged TPP solution. When PEG binds with chitosan-rifampicin it changes the character and the surface of the nanoparticles and slightly increases its particle size, as well as encapsulation of drugs also increased. PEG bind with CS-RIF has resulted in more prolonged retention as compared with non-coated CS-RIF. Various parameters and method-ologies such as loading capacity, encapsulation effi ciency, SEM (Scanning electron microscopy), FITR (Fourier transmission infra-red microscopy) and in vitro release of drugs have been utilised for characterization of nanoparticles [111].

Recently, there was targeted intracellular delivery of anti-TB to Mycobacterium infected macrophages through functionalized mesopo-rous silica nanoparticles. Mesoporous silica nanoparticle (MSNP) drug delivery system was coated with polyethyleneimine (PEI) to release rifam-picin. Th ese mesoporous silica nanoparticles get internalized by human macrophages and delivered to the lysosomes and to the acidifi ed endo-somes as a result intracellular release of high concentrations of antituber-culosis drugs occurred. Coated MSNP have shown much greater loading capacity than uncoated MSNP. Th e amount of RIF loaded on the PEI coated nanoparticles was determined aft er elution by spectrophotometer at the wavelength 475nm against RIF standards [112]. Microspheres are spheri-cal, free fl owing particles ranging in average particle size 1–50 microns. Currently, the potential of microspheres as carriers for target drug delivery systems has been exploring. Microspheres are prepared from the diff erent methods such as protein gelation technique, sonication technique, solvent evaporation technique, spray and freeze technique, polymerization tech-nique and solvent extraction method [113].

Oral nanoparticles are also being used for the delivery of the antitu-berculosis drugs. In an experiment, orally administered poly-lactide-co-glycolide (PLG), a synthetic polymer nanoparticle encapsulating antituberculosis drugs such as RIF, INH and PZA was developed for cere-bral drug delivery in murine model. Th ese nanoparticles were prepared by multiple emulsion and solvent evaporation technique and then they were administered orally to mice for their biodistribution, pharmokinetic and chemotherapeutic studies. As a result of this experiment, a single oral dose to mice maintains sustained drug levels for 5–8 days in the plasma and for 9 days in the brain. As well as in M. tuberculosis H

37Rv infected mice, fi ve

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oral doses of these nanoparticles formulation was administered every 10th day which in turn resulted in absence of tubercle bacilli in the meninges, on the basis of CFU and histopathology [114].

Niosomes can also be used as an alternative to liposomes for specifi c tar-get drug delivery. Th ey are basically thermodynamically stable liposomes like vesicles produced by the hydration of cholesterol and charge induc-ing components such as charged phospholipids (e.g. Dicetylphosphate and stearyl amine) and non ionic surfactants (e.g. monoalkyl or dialkyl polyoxyethylene ether) [115]. Micron sized RIF loaded niosomes contain-ing Span-85 as surfactants were prepared by the Jain and Vyas [116]. As a result upto 44% of the drug was localized in the lungs by adjusting the size of the carrier. Th e same group of the scientist further extended their studies to investigate the biodistribution of niosomes of smaller sizes with the diff erent sorbitan esters (Span 20, 40, 60, 80 and 85) and cholesterol in 50:50 mol fraction ratios [117]. In vitro studies showed 80% maximal and 52% minimal levels for Span-20 and Span-85 based systems. All these studies led to the conclusion that more lipophilic the surfactant, slower was the drug release in the aqueous medium.

Nanoparticles of size 250 nm were used for delivery of the anti-TB drugs such as isoniazid, rifampicin and streptomycin. Th e accumulation of these drugs was checked in human monocytes as well as thieir antimi-crobial activity against M. tuberculosis residing in the human-monocyte derived macrophages. Th e result was that the intracellular concentration of the free INH was equal to or slightly higher than that of the extracellular fl uid [118].

14.4 Cancer & Tumor Targeting Nanoparticles

Nanoparticles are very well suited materials for targeted tumor delivery because of their ability to circulate in the bloodstream for relatively longer period of time as well as their ability to accumulate in the tumor spaces. Some of the in vivo and in vitro experiments have shown nanoparticles to be fruitful for the tumor treatment. Th ere is a growing evidence which sug-gests that many nanoparticles accumulated at the tumor site is indepen-dent of the presence or absence of the targeting ligand. In many studies, nanodelivery systems with target ligands have shown better performance than non-targeted system (Figure 14.7, below). Th ese entire conclusions led to the improvement in the performance of the nanoparticles asso-ciation with target cell membranes and target cell internalization. EPR (enhanced permeability and retention) eff ect is one of the dominating

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mechanisms for localization of tumour site [119]. Active tumour targeting involved the use of molecules which specifi cally interact with the physi-ological target whereas the passive targeting involves the use of natural properties and processes in the tissues for localization of the delivery agent at a desired target site. Passive tumour targeting is mainly by EPR eff ect. Particles within the range of 100–200 nm have been shown to accumulate in tumour by EPR eff ect.

Wang et al studied the biodistribution of targeted nanoparticles com-posed of heparin-folate-paclitaxel conjugates loaded with paclitaxel and compared it with the non-targeted nanoparticles of heparin-paclitaxel loaded with paclitaxel, as a result, this targeted system loaded with the drug reduces the tumour volume over nanoparticle and paclitaxel con-trol in the KB-3–1 human nasopharyngeal carcinoma-xenograft bearing mouse model [120]. Another most common targeting ligand such as trans-ferrin (Tf) has been conjugated to a variety of targeted delivery systems for targeting over expressive Tf receptors which are common in many can-cerous cells. Th ey basically improve the drug delivery [121]. Liposomes possessing an anti-HER2 monoclonal antibody (Mab) composed of phos-phatidylcholine and polyethylene glycol (PEG) modifi ed distearoyl phos-phatidylethanolamine with an average particle diameter of 100 nm has resulted in improve antitumor effi cacy of doxorubicin over the various control formulations [122]. Kirpotin et al. further evaluated the biodistri-bution and uptake of these targeted liposomes containing the anti-HER2 Mab and compared it with that of non -targeted liposomes [123]. In 1975,

Nanocarrier

Blood capillary

Through

enhanced

permeation

effect

Ligand

ReceptorFree drugs

Cancer cell

Increase in intracellular

drug concentration

Nanoparticles targeting Cancer cells

Figure 14.7 Nanoparticles target cancer cells through ligand receptor binding.

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Ringsdorf fi rst proposed the concept of polymer-drug conjugates for deliv-ery of the hydrophobic smaller drugs to their target site of action [124]. Th is polymer-drug conjugates composed of a water-soluble polymer and is chemically conjugated to the drug via a biodegradable spacer. Th is spacer basically cleaved at the target site by hydrolysis or enzymatic degradation. Th ey can be accumulated at the tumour site by the EPR eff ects, followed by release of the drug through the cleavage of spacer [125].

In recent years, chitosan anticancer drug conjugates have also been investigated. Doxorubicin conjugated glycol chitosan (DOX-GC) with cis-aconityl spacer was synthesized by the chemical attachment of N-cis-aconityl DOX to GC using carbodiimide. DOX-GC conjugates contain-ing 2–5wt% DOX form self-assembled nanoparticles in aqueous condition but if DOX is present in higher concentration i.e. above 5.5wt% in the nanoparticles, it will get precipitated due to increase in hydrophobicity. Th e loading contents of DOX in the nanoparticles increased upto 38.9wt%. Th e release rate of DOX from the nanoparticles is dependent on the pH of the media because the cis- aconityl spacer is cleavable at low pH [126]. When these DOX-GC conjugated nanoparticles were administered into the mice, they get preferentially accumulated in the tumour tissue thus describing EPR eff ect. A variety of hydrophobic drugs can be loaded into the chitosan nanoparticles; the loading effi ciency depends on the physio-chemical characteristics and preparation methods used.

For the cancer therapy, a hydrophilic 5-fl uorouracil was loaded into the chitosan nanoparticles (250–300 nm in diameter) using water- in-oil emulsion method, followed by the chemical cross linking of the chitosan in the presence of glutaraldehydes [127]. Some of the drug loaded solid nanoparticles could release the drugs at the rate which is in turn dependent on the type of hydrophobic moiety, degree of substitution and the physio-chemical properties of the drugs. Th ese chitosan based nanoparticles have been reported to be selectively accumulated at the tumour site, primarily owing to the EPR eff ect. As a result these drug loaded nanoparticles have shown better therapeutic effi cacy than the free drug in vivo.

Magnetic targeting is also an attractive physical targeting technique, generating a substantial attention for the drug delivery applications. Th e therapeutic agents to be delivered are either immobilized on the surface or they are encapsulated into the magnetic micro or nanoparticle carriers. Upon intravenous administration, they get concentrated at the tumour site using an external high-gradient magnetic fi eld [128]. Aft er the accumula-tion of the magnetic carrier at the target tumour site in vivo, drugs are released from the magnetic carrier and then eff ectively taken up by the tumour cells. Th e effi ciency of the carrier accumulation mainly depends

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on various parameters such as intensity of the magnetic fi eld, rate of blood fl ow and the surface characteristics of carriers.

Gallo et al developed magnetic chitosan microspheres containing oxantrazole (MCM-OX), an anti cancer drug, for the treatment of brain tumour [129]. He monitored the level of OX in the brain aft er administer-ing intra-arterial injections of MCM-OX to male Fischer 344 rats under the magnetic fi eld of 6000G for 30 min. As a result, a 100 fold increase in OX concentrations in the brain aft er administration of MCM-OX was observed as compared to the OX in solution. In a similar study, Chen et al prepared chitosan-bound magnetic nanoparticles loaded with epirubi-cin, an anthracycline drug used for cancer chemotherapy. Th e magnetic nanoparticles were stable at pH 3–7 and approximately 80% of the drug was released aft er 150–300 min in a biological buff er. In vitro experiment showed the effi ciency of anticancer property of drug loaded nanoparticles as compared with that of free drugs [130].

Magnetic fi eld enhances the cellular effi ciency uptake of the mesopo-rous silica nanoparticles. Th e internalization of M-MSNs (magnetic mes-oporous nanoparticles) by A549 cancer cells could be enhanced by the magnetic fi eld. Th e endocytosis studies indicate that the M-MSNs inter-nalization by A549 cells is mainly energy dependent pathway, namely clathrin-induced endocytosis. With the help of magnetic fi eld, anticancer drug loaded M-MSNs induced cancer cell growth inhibition. Delivery of hydrophilic and hydrophobic drugs through magnetic mesoporus silica nanoparticles (MMSNs) inhibits cancer cells growth [131].

Recently, a report on the development of drug delivery system for pho-tosensitive delivery of an anticancer drug campothtecin along with cyto-toxic cadmium sulphide from a magnetic drug nanocarrier was developed. During this experiment, core–shell nanoparticles consisting of magnetic iron-oxide-cores and mesoporous silica shells were synthesized with a high surface area (859 m2 g−1) and hexagonal packing of mesopores (2.6 nm) in diameter. Th e mesopores were loaded with an anticancer drug camp-tothecin and the entrances of the mesopores were blocked with 2-nitro-5-mercaptobenzyl alcohol functionalized CdS nanoparticles through a photo cleavable carbamate linkage. Camptothecin release from the mag-netic delivery system was measured by the fl uorescence spectroscopy upon irradiation by the UV light. As a result, treatment of cancer cells with these drugs lead to the decrease in viability of the cells because of the activity of capping of CdS nanoparticles. Th e capping of Cds nanoparticles and loaded camptothecin exert an anticancer activity [132].

Transferrin conjugated paclitaxel biodegradable nanoparticle were also tested in the murine model for treatment of prostate cancer. It was

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hypothesized that nanoparticle conjugated to transferrin ligand enhances the therapeutic effi ciency of the drug. Nanoparticles of 220 nm diameter loaded with 5.4% w/w paclitaxel drug under in vitro condition exhibited sustained release of 60% encapsulated drug in 60 days. Th e anti-prolifera-tive activity of NPs was then determined in human prostate cancer cell line (PC3) and their eff ect on tumour inhibition was observed in the murine model of prostate cancer. Animals which received a single-dose of intra-tumoral injection of transferrin conjugated nanoparticles resulted in com-plete tumor regression and greater survival rate [133].

Lymphatic drainage plays an important role in uptake of particulates in respiratory system and also being associated with the spreading of lung cancer through metastasis development. Recently, solid lipid nanoparticles (SLN) have used as carriers of anti-tumoral drugs.

Nanoparticles of about 200 nm diameter, radiolabelled with 99m Tc using the lipophilic chelator D,L-hexamethylpropyleneamine oxime (HMPAO) were developed for the pulmonary uptake. Th e biodistribution studies were also carried out following aerosolisation and administration of a 99mTc-HMPAO-SLN suspension to a group of adult male Wistar rats. As a result, 60 min dynamic image followed by the static image were col-lected at 30 min intervals for up to 4h post inhalation which has shown the signifi cant uptake of the radiolabelled SLN into the lymphatic system aft er inhalation and thus controlling spreading of lung cancer [134].

Recently, nanoparticle-aptamer bioconjugate, a new approach for treat-ment of prostate cancer was developed. In this nucleic acid ligands (aptam-ers) are well suited for therapeutic targeting of the encapsulated drugs and controlled release of polymer particles in cells or tissues. A bioconjugate was synthesized, mainly composed of controlled release polymer nanopar-ticles and aptamers. Its effi cacy was examined for targeted delivery to pros-tate  cancer  cells. Nanoparticles composed of poly-(lactic acid) blocked polyethylene glycol (PEG) copolymer with a terminal carboxylic acid func-tional group (PLA-PEG-COOH) and encapsulated rhodamine-labeled dextran (as a model  drug) within PLA-PEG-COOH was synthesized. Nanoparticle-aptamer bioconjugates with RNA aptamers which bind to the prostate specifi c membrane antigen overexpressed on prostate acinar epi-thelial cells. As a result, these bioconjugates can be effi ciently targeted and taken up by the prostate LNCaP epithelial cells. Th is was the fi rst report on targeted drug delivery through nanoparticle aptamer bioconjugates [135].

Recently, Heparin is used as a carrier for cancer targeting and imaging. Heparin-anticancer drug conjugates shows higher anticancer activity than free drug. Th e conjugated heparin (heparin-deoxycholate sodium) retained its ability to bind with angiogenic factors, thus resulting in a

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signifi cant decrease in endothelial tubular formation. Secondly, targeting ligands conjugated with that of heparin derivatives have been used for the receptor mediated delivery of anticancer drug. Heparin-folic acid-retinoic acid (HFR) bioconjugates are being used for treatment of cancer cells [136].

Recently, a pH responsive drug carrier is developed, based on chondrio-tin sulfate functionalized mesostructured silica nanoparticles (NMChS-MSNs) i.e., the amidation between NMChS macromer and amino group functionalized MSNs. Th ese nanoparticles were spherical in shape with a mean diameter of about 74nm. A well known anti-cancer drug, Doxorubicin hydrochloride (DOX) was loaded into the channels of NMChS-MSNs through the electrostatic interactions between drug and matrix. As a result, drug release rate was pH dependent and it increases with the decrease in pH. In vitro cytotoxicity test also proves that these nanoparticles are highly biocompatible and can be used as a drug carrier. Th e above experiment proves that these chondriotin sulfate mesostructured functionalized silica nanoparticles are good platforms for pH dependent controlled drug deliv-ery systems for cancer therapy [137].

Pancreatic cancer is a highly lethal disease with a 5-year survival rate less than 5% due to the lack of an early diagnosis method and eff ective therapy. Recently, a multifunctional nanoimmuno liposome with high loading of ultra small super paramagnetic iron oxides (USPIOs) and doxo rubicin (DOX) was prepared by transient binding and reverse-phase evaporation method and then conjugated with anti-mesothelin mono-clonal antibody by post-insertion method to target anti-mesothelin-overexpressed pancreatic cancer cells. In vivo studies have shown that, comparing with FD (free DOX) and PLDU, M-PLDU possessed higher inhibitory eff ect on tumour growth and the tissue distribution assay fur-ther proved that M-PLDUs get selectively accumulated in the tumour xenograft [138].

A novel magnetic nanoparticle for drug carrier has been recently dis-covered for the enhanced cancer chemotherapy. Magnetic nanoparti-cles loaded with antitumor drugs in presence of external magnetic fi eld resulted in improvement in cancer treatment. In this experiment, DOX-PGMNPs nanoparticles were synthesized and cytotoxicity was assessed in vitro. Along with this, intravenous administration of DOX-PGMNPs to H22 hepatoma cell tumour bearing mice, biodistribution of DOX was also measured in diff erent tissues [139].

One of the nanoparticle poly-(D,L-lactide-co-glycolide)/montmo-rillonite were decorated by Trastuzumab, an epidermal growth fac-tor receptor-2(HER-2) antibody and was being used for targeted breast

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cancer chemotherapy with paclitaxel as a model anticancer drug. Th ese nanoparticles were prepared by solvent extraction/evaporation method. Internalization of the coumarin-6-loaded PLGA/MMT NPs with or with-out the antibody decoration by both of Caco-2 colon adeno carcinoma cells and SK-BR-3 breast cancer cells were visualized by confocal laser scanning microscopy and quantitatively analyzed, which shows that the antibody decoration achieved signifi cantly higher cellular uptake of the NPs to treat breast cancer [140].

Development of functionalized super paramagnetic iron oxide nanopar-ticles were being reported to interact with human cancer cells. Th e capacity of interaction of the cells with these nanoparticles as well as cytotoxicity was evaluated in human melanoma cells. Out of the four formulations of nanoparticles, only the polyvinyl alcohol super magnetic iron oxide nanoparticles interact with cells and cytotoxicity was negative in human melanoma cells [141].

14.5 Conclusion

Nanotechnology is globally emerging technology for targeted drug deliv-ery. Due to some disadvantages related to the unconventional strategies, such as larger amount of drug consumption by patients, more time in recovering lead to the more use of diff erent kinds of nanoparticles as drug carriers. Th ey basically act by crossing the biological barriers and target-ing the site. Nanoparticles such as solid lipid nanoparticles, polymeric nanoparticles, liposomes, mesoporous silica nanoparticles are being used for treatment of various types of diseases. Several carriers based drug deliv-ery system incorporating anti-TB drugs have been developed for target site action. Nanoparticles encapsulating anticancer drugs such as doxorubicin have been also developed for treatment of various types of cancers such as prostate cancer, breast cancer, pancreatic cancer and lung cancer. All these developments in drug delivery reports have resulted in signifi cant merits such as improved drug bioavailability, reducing dosing frequency, versatil-ity of routes of administration and long term stability which become the basis of better management of diseases. Recently, aerosol vaccines are being in progress for drug delivery. Besides having so many advantages, some of the toxicological issues related to understanding the fate of nanocarri-ers, polymeric constituents in the body as well as elimination of residual should be deleted. harmful residual organic solvents have to be resolved. Future research on vectorized delivery system of drug has been focussed for large amount of drug delivery and for better results.

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