Journal of Liposome Research, 2009; 19(4): 310–321

R E S E A R C H A R T I C L E

Role of nanocarrier systems in cancer nanotherapy

M. R. Mozafari1, A. Pardakhty2, S. Azarmi3, J. A. Jazayeri4, A. Nokhodchi5, and A. Omri6

1Department of Biochemistry and Molecular Biology, Monash University, Clayton, Victoria, Australia, 2Department of Pharmaceutics, School of Pharmacy and Pharmaceutical Sciences, Kerman University of Medical Sciences, Kerman, Iran, 3Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, Alberta, Canada, 4Monash University, Faculty of Pharmacy and Pharmaceutical Sciences, Department of Pharmaceutical Biology, Parkville, Victoria, Australia, 5Chemistry and Drug Delivery Group, Medway School of Pharmacy, Universities of Kent and Greenwich, Kent, UK, and 6!e Novel Drug and Vaccine Delivery Systems Facility, Department of Chemistry and Biochemistry, Laurentian University, Sudbury, Ontario, Canada

Address for Correspondence: M. R. Mozafari, Department of Biochemistry and Molecular Biology, Monash University, Wellington Road, Clayton, Victoria 3800, Australia; Fax: +61 3 9905 3166; E-mail: mozafarimr@yahoo.com

(Received 15 January 2009; revised 25 February 2009; accepted 21 March 2009)

Introduction

Despite the magnitude of research in the area of cancer therapy and discovery of many new cytotoxic drugs as potential candidates for the treatment of cancer, this life-threatening disease still causes millions of deaths every year. !e usefulness of most conventional cancer therapeutics is often limited due to insu"cient delivery of therapeutic drug concentrations to the target tumor or due to severe and harmful toxic e#ects on normal tissues. Further, although proteomics and genomics continue to uncover molecular signatures that are unique to cancer, the major challenge remains in targeting and selectively killing cancer cells. It is commonly acknowledged that if a su"cient amount of the antineoplastic agent can be delivered to the target tumor without causing harm to healthy cells, an improved chemotherapy will be achieved. Nevertheless, the accompanying damage to

healthy tissues that occurs once drug doses are elevated poses a severe limitation to this strategy (Uchegbu, 2000). Controlled release systems and particulate car-rier technologies $t well in the chemotherapy of tumors, as simple strategies can be employed to achieve tumor targeting and limit drug toxicity. Various carrier systems have been used until now to achieve high therapeutic concentrations in tumors. !ese include microspheres (Chaurasia et al., 2006; Cheung et al., 2006), nanopar-ticles (Azarmi et al., 2006a, 2006b; Sun et al., 2008), micelles (Mahmud et al., 2007), niosomes (Uchegbu, 2000; Uchegbu and Vyas, 1998), solid lipid nanoparticles (Wong et al., 2007), dendrimers (Kono et al., 2008), and liposomes (Karanth and Murthy, 2007; Shehata et al., 2007). !e possibility of surface engineering of colloi-dal drug-delivery systems has enabled the scientists to prepare polymer-linked vesicles (e.g., PEGylated lipo-somes) or folate-decorated nanoparticles to enhance the

ISSN 0898-2104 print/ISSN 1532-2394 online © 2009 Informa UK LtdDOI: 10.3109/08982100902913204

AbstractCancer continues to be a major cause of morbidity and mortality worldwide. While discovery of new drugs and cancer chemotherapy opened a new era for the treatment of tumors, optimized concentration of drug at the target site is only possible at the expense of severe side e!ects. Nanoscale carrier systems have the potential to limit drug toxicity and achieve tumor localization. When linked with tumor-targeting moieties, such as tumor-speci"c ligands or monoclonal antibodies, the nanocarriers can be used to target cancer-speci"c receptors, tumor antigens, and tumor vasculatures with high a#nity and precision. This article is an overview of advances and prospects in the applications of nanocarrier technology in cancer therapy. Applications of nanoliposomes, dendrimers, and nanoparticles in cancer therapy are explained, along with their preparation methods and targeting strategies.

Keywords: Cancer; dendrimers; nanotherapy; nanocarriers; nanoliposomes; niosomes; nanoparticles

http://www.informahealthcare.com/lpr

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

Role of nanocarrier systems in cancer nanotherapy 311

circulation time of the carrier-drug complex and enable it to reach the target organ. When linked with tumor-targeting moieties, such as tumor-speci$c ligands or monoclonal antibodies, these micro- and nanocarriers can be used to target cancer-speci$c receptors, tumor antigens (biomarkers), and tumor vasculatures with high a"nity and precision.

It has been shown that drug delivery to various sites within the body is directly a#ected by particle size (Hughes, 2005; Kawashima, 2001). While microencap-sulation has been widely studied in the drug-delivery area, researchers are gradually exploiting more nanos-cale technologies to combat cancer and other diseases. Progress in nanoscience and technology has made it pos-sible to manufacture and characterize submicrometric drug carriers on a routine basis. Studies have shown that the encapsulation of drugs and other bioactive agents in nanocarrier systems improves their therapeutic potential by facilitating intracellular delivery and prolonging their retention time inside the cell (Hughes, 2005). Further, nanometer-sized carriers have novel optical, electronic, and structural properties that are not available from either individual molecules or bulk solids (Mozafari, 2006). Accordingly, the focus of the present article will be on the nanocarrier systems and their utilization in the encapsulation of the anticancer drugs. E#orts aimed at diverting antitumor agents away from healthy tissues and toward tumor tissues with some of the most promis-ing nanocarrier systems will be discussed.

Nanocarrier science and technology

Encapsulation of therapeutic agents in biodegradable and biocompatible carriers has been considered a safe way of delivering anticancer drugs. Unprotected drugs, when introduced into the body, exhibit a short in vivo half-life and de$cient antitumor properties. Consequently, the e#ective delivery of anticancer drugs necessitates pro-tection from the hostile immunological and enzymatic environments of the body. Microencapsulation systems and microcarriers have been extensively utilized for the protection and delivery of bioactive agents, including drugs, vaccines, nutrients, and cosmetics, both in the research and development as well as manufacturing of various products. However, targeted controlled release can be provided much more e"ciently by employ-ing nanocarrier technologies (Mozafari, 2006). Novel nanocarrier systems can make it possible to use certain drugs that were previously impractical to prescribe due to toxicities, high cost of manufacture, or because they were impossible to administer. Compared to micron-sized carriers, nanocarriers provide more surface area and have the potential to increase solubility, enhance bioavailability, improve controlled release, and enable

precision targeting of the entrapped drugs to a greater extent (Khosravi-Darani et al., 2007; Mozafari et al., 2006). As a consequence of improved stability and tar-geting, the amount of drug required to exert a speci$c e#ect when encapsulated or incorporated to a nanocar-rier is much less than the amount of drug administered in the free form. A timely, targeted release improves the e#ectiveness of therapeutics, broadens their application range, and ensures optimal dosage, thereby improving cost-e#ectiveness of the product. Reactive or sensitive materials, such as polynucleotides and polypeptides, can be turned into stable ingredients through encapsulation or entrapment by nanocarrier systems. Nanoparticles and other nanocarriers can be used as intravascular infusion preparations, injectable emulsions for both parenteral and enteric administration, as well as vaccine dosage forms for use in subcutaneous or intramuscular injections (Gibaud et al., 1998; Lemoine and Preat, 1998; Song et al., 1997).

Nanocarrier-mediated drug targeting has made it possible to deliver chemotherapy agents directly to tumors, hence reducing systemic side e#ects (Hughes, 2005). In fact, nanocarrier science is the foundation upon which most nanotechnological cancer therapy is based. Nanotechnology is a new scienti$c discipline and the full scope of its contributions to the $eld of human health care is yet to be fully explored. Nevertheless, recent advances suggest that nanotechnology will have a profound impact on disease prevention, diagnosis, and treatment (Cheng et al., 2006; Emerich, 2005; Sahoo and Labhasetwar, 2003; Williams, 2004). Applications of nanotechnology in medicine are very promising and areas such as molecular imaging, disease diagnosis, drug encapsulation, and targeted delivery at speci$c sites in the body are being intensively investigated and some products are undergoing clinical trials (Moghimi et al., 2005; Morrow et al., 2007; Sha#er, 2005; Wilkinson, 2003). In cancer therapy, targeting and localized delivery of drug are the key parameters. In addition, newer gen-erations of molecular therapies, such as gene therapy and siRNA, require intracellular delivery strategies to attain optimized results.

Studies, so far, have indicated that nanocarrier-based formulations can alter a number of pharmacokinetic parameters in a range otherwise di"cult to obtain (Langner and Kozubek, 2006). For instance, it has been shown that, in using nanocarrier-based formulations, the circulation time has been increased up to tens of hours, the severity of side e#ects substantially reduced, and the mechanisms of passive and active targeting discov-ered and exploited (Cammas, 1996; Mehta et al., 1997; Mozafari and Mortazavi, 2005). Extended persistence time is especially valuable in specialized applications, such as boron neutron capture therapy, in which active agents are encapsulated in lipid vesicles (Johnsson et al.,

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

312 M. R. Mozafari et al.

1999), or in medical imaging, in which contrast agents last for extended periods of time and exhibit a"nity toward pathologically altered tissues (Harrington et al., 2000a, 2000b).

!e anatomical changes and pathophysiological conditions of diseased or in%amed tissues o#er oppor-tunities for the delivery of various nanotechnological products. Drug targeting can be achieved by taking advantage of these speci$c characteristics of abnormal tissues (Vasir and Labhasetwar, 2005; Vasir et al., 2005). An ideal targeting system should have long circulation time, should be present in su"cient concentrations at the target site, and should not lose its activity or thera-peutic e"cacy while in circulation (Sahoo et al., 2007). !e increased vascular permeability, coupled with an impaired lymphatic drainage in tumors, allows an enhanced permeability and retention e#ect of the nano-carriers in the tumors or in%amed tissues (Hashizume et al., 2000; Maeda et al., 2000; Matsumura and Maeda, 1986; McDonald and Baluk, 2002). Hence, this patho-physiological opportunity allows extravasation of the nanosystems and their selective localization in the abnormal tissues, such as tumors (Allen and Cullis, 2004; Hobbs et al., 1998). !e tendency of nanocarriers to speci$cally localize in the reticuloendothelial system also provides an excellent opportunity for passive drug targeting to the macrophages present in the liver and spleen. !is natural mechanism, therefore, can be used for drug targeting for intracellular infections, such as candiasis, leishmaniasis, and listeria. !e macrophages of the infected individual play a role in these diseases; consequently, if the macrophages are destroyed, then the disease will be eliminated (Daemen et al., 1995). !e following sections explain some of the most commonly applied nanocarrier systems and their role in cancer therapy.

Liposomes and nanoliposomes

Although it is known that the $rst man-made liposomes appeared around 40 years ago, there are increasing number of scienti$c evidence that these bilayered structures have been a vital part of living cells—as biomembranes—since the emergence of life some 3.5 billion years ago (Monnard and Deamer, 2001; Mozafari, et al., 2004; Nomura et al., 2001; Pozzi et al., 1996). During the last few decades, liposomes (also known as bilayer lipid vesicles) have moved a long way from being just a model membrane system to become a carrier of choice for numerous practical applications in di#erent areas, including pharmaceutics, cosmetics, and food technol-ogy. Compared with other encapsulation strategies, liposomal carrier systems have distinctive advantages, including the ability to entrap material with di#erent

solubilities, the possibility of being produced by using natural and biodegradable ingredients on industrial scales, and targetability (Mozafari et al., 2006; Mozafari and Mortazavi, 2005; Mozafari et al., 2007; Yurdugul and Mozafari, 2004). Liposomal carriers can protect the drug they carry from free radicals, metal ions, pH, and enzy-matic degradation. !ey can encapsulate or entrap not only lyophilic material, but also lyophobic compounds, separately, or, if required simultaneously, providing a synergistic e#ect (Suntres and Shek, 1995, 1996). !e unique properties of liposomes have triggered numer-ous applications in di#erent scienti$c $elds, from basic studies of membrane structure/function to bioactive agent delivery. Liposomes and nanoliposomes are particularly useful as e"cient drug-delivery systems because of their ability to pass through lipid bilayers and cell membranes, hence improving bioavailability.

Physically, liposomes consist of one or more lipid and/or phospholipid bilayers and can contain other molecules, such as proteins or carbohydrates, in their structure. !e lipidic layer on the liposome con$nes and protects the enclosed drug until the liposome reaches its destination and adheres to the outer membrane of target cancer cells. By this process, drug toxicity to healthy cells is minimized and therapeutic e"cacy can be increased. An onion-shaped liposome composed of a number of concentric bilayers is known as a multila-mellar vesicle (MLV; Figure 1), while one composed of many small nonconcentric vesicles encapsulated within a single lipid bilayer is known as a multivesicular vesicle (MVV). Another type of liposome is known as a unila-mellar vesicle (ULV; Fig. 1) and contains a single lipidic bilayer (Khosravi-Darani et al., 2007). Based on their diameters, unilamellar liposomes are classi$ed as small (SUV, ca. less than 100 nm in diameter) and large unila-mellar vesicles (LUV, ca. larger than 100 nm in diameter). Due to the presence of both lipid and aqueous phases in their structure, liposomes can be utilized in the encapsulation or entrapment of water- and lipid-soluble material in addition to the amphiphilic compounds. !e term nanoliposome has recently been introduced

A B C

Figure 1. (A) A phospholipid vesicle (liposome), which can be uni- or multilamellar with respect to the number of phospholipid bilay-ers. (B) Partially sectioned liposome composed of one phospholipid bilayer (the grey section) enclosing an internal aqueous core (blue) known as a unilamellar vesicle (ULV). (C) Partially sectioned mul-tilamellar vesicle (MLV) comprised of several phospholipid bilayers separated by aqueous compartments.

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

Role of nanocarrier systems in cancer nanotherapy 313

to exclusively refer to nanoscale lipid vesicles (Mozafari and Mortazavi, 2005), since liposome is a general phrase covering many classes of lipid vesicles, with diameters ranging from around 20 nm to several micrometers.

Liposomal drug formulations can be administered to patients via di#erent routes, including oral, nasal, buccal, pulmonary, transdermal, rectal, and ocular. Based on these characteristics, liposome technology is currently a well-developed strategy for encapsulation and delivery of chemotherapy agents and other drugs. Table 1 lists a number of liposomal products already approved for human use. In addition, currently, there are a number of liposomal formulations in clinical trials (Peek et al., 2008).

A preclinical study in mice conducted by Guan and colleagues evaluated the e#ect of liposomes on the type of immune response generated for a cancer-associated antigen (MUC1 mucin peptide) therapeutic cancer vaccine (Guan et al., 1998). !is study revealed that liposome-entrapped MUC1 peptide (BP25) produced a strong, speci$c cytotoxic T-cell response (Guan et al., 1998). Clinical studies have con$rmed that L-BLP25 (also known as Stimuvax®, a lyophilized liposomal for-mulation of BP25 lipopeptide, MPL®, and three lipids) is well tolerated and elicits a cellular immune response in patients with lung cancer (Butts et al., 2005). A phase IIB trial showed that the L-BLP25 vaccine increases survival rates for patients with nonmetastatic NSCLC (non-small-cell lung cancer) tumors (Butts et al., 2005). A phase III clinical trial is currently under way (Peek et al., 2008). Stimuvax is being developed by Merck and Biomira for the treatment of NSCLC that accounts for almost 80% of all lung cancers (Peek et al., 2008).

Methods of liposome and nanoliposome preparation

!e manufacture of both liposomes and nanoliposomes requires input of energy to a dispersion of lipid/phos-pholipid molecules in an aqueous medium (Mozafari and Mortazavi, 2005). !e underlying mechanism for the formation of liposomes and nanoliposomes is basically the hydrophilic-hydrophobic interaction between phos-pholipids and water molecules (Mozafari, 2005). Since liposomes are dynamic entities that tend to aggregate and/or fuse and as a result increase in size, vesicles pre-pared in nanometric size ranges may end up becoming

micrometric particles upon storage. However, nano-liposomes should have su"cient physical stability and should maintain their size and polydispersity index upon storage (Mozafari and Mortazavi, 2005). Most methods of liposome and nanoliposome preparation, which can also be used to prepare other drug-delivery systems, including niosomes and vesicular phospholipid gels, involve solubilization of the ingredients in organic sol-vents, such as chloroform and methanol. !ese solvents not only a#ect the chemical structure of the entrapped drug, but also will remain in the $nal liposome formu-lation and result in toxicity and diminished stability of the vesicles (Cortesi et al., 1999; Deamer and Uster, 1983; Mozafari et al., 2007; Vemuri and Rhodes, 1995). Generally, residual solvents in pharmaceuticals, known as organic volatile impurities (OVIs), have no therapeu-tic bene$ts but can be hazardous to human health and the environment (Dwivedi, 2002). !e other drawback of most liposome preparation methods is employment of sonication, micro%uidisation, homogenization, or other forms of mechanical stress, which can lead to drug denaturation (Kasaai et al., 2003). !ese drawbacks can be addressed by employing robust, scalable prepara-tion techniques, including recently developed methods, such as the heating (Mozafari et al., 2002, 2007) and Mozafari methods (Colas et al., 2007). !ese methods are capable of producing a number of di#erent drug car-riers without subjecting the drug to toxic solvents, high shear forces, low/high pH, or extreme temperatures—a detailed description of which is given elsewhere (Colas et al., 2007; Mozafari, 2005; Mozafari and Mortazavi, 2005; Mozafari et al., 2002, 2007).

Targeting strategies

Targeted therapy can be achieved e"ciently via lipo-somes and nanoliposomes employing passive or active targeting mechanisms (Mozafari, 2005, 2006). !e use of site-speci$c triggers that can cause drug release speci$cally in the target tissue is one way of improving drug bioavailability at the tumor site. Another method of increasing drug bioavailability is to obtain a higher level of liposome accumulation by active targeting. In addition, the combination of active targeting with active triggering can result in an enhanced, more speci$c drug release at the tumor target site (Andresen et al., 2005).

Table 1. FDA-approved liposome-based drugs on the market.Name of the drug Application ManufacturerAbelcet® (amphotericin B) Invasive fungal infections Enzon Pharmaceuticals; St Mary’s

Pharmaceutical Unit, USAAmBisome (amphotericin B liposome for injection) Antifungal Gilead Sciences; Astellas Pharma Inc., USADaunoXome (daunorubicin citrate liposome injection) Anticancer Diatos, FranceDoxil (doxorubicin HCl liposome injection) Ovarian and breast cancer ALZA corporation; Ortho Biotech. Products, USAMyocet™ (liposome-encapsulated doxorubicin citrate complex)

Anticancer Elan corporation, USA

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

314 M. R. Mozafari et al.

Active targeting is achieved by formulating liposomes sensitive to di#erent stimuli (e.g., pH, temperature, light, ultrasound, etc.), or conjugating them to one or more targeting ligands, such as tissue- or cell-speci$c molecules. A recently developed liposome, employed for active targeting, releases its load at the target tissue upon encountering ultrasound waves. !ese “echogenic liposomes” can be used e#ectively to ablate solid tumors, and a variety of cancers are presently being treated in the clinic using this technology (Frenkel, 2008).

Several researchers have described potential meth-ods for active targeting, such as liposomes coupled to speci$c antibodies (Pan et al., 2008; Sofou and Sgouros, 2008), as well as liposomes coated with targeting pro-teins expressed on cancer cell membranes or endothe-lial cells lining the newly generated blood vessels in the tumor. Examples of such proteins are the folate receptor, induced on the surface of actively growing tumor cells (Lu and Low, 2003; Xiang et al., 2008), and the integrin surface receptor (Hood et al., 2002; Huang et al., 2008), expressed on the endothelial cells in the neovasculature of growing tumors. Other examples include galactoli-pids that have been found to increase transfection sig-ni$cantly by targeting the asialoglycoprotein receptor of human hepatoma HepG2 cells (Zanta et al., 1997). In gene therapy of cancer, targeted delivery to the tumors in various parts of the body can be achieved by using advanced liposomal systems, such as antibody-antigen and ligand-receptor combinations (Dass and Choong, 2006). In an in vivo study, Zhang and colleagues formu-lated liposomes to overcome the blood-brain barrier and target rhesus monkeys’ brains. !ey showed that PEGylated (treated with polyethylene glycol) liposomes, linked to a monoclonal antibody for the human insulin receptor, led to widespread reporter expression in the brains of the tested animals (Zhang et al., 2003).

Studies show that selective delivery of the antican-cer drug, doxorubicin, in PEGylated liposomes for the treatment of patients with breast-carcinoma metastases resulted in a signi$cant improvement in survival rate (Perez et al., 2002). A combination therapy utilizing liposomal doxorubicin and paclitaxel (Schwonzen et al., 2000) or Caelyx (doxorubicin in PEGylated liposomes) and carboplatin (Goncalves et al., 2003) has been used to target breast-carcinoma metastases (Karanth and Murthy, 2007). Caelyx is also in phase II clinical trials for patients with squamous-cell cancer of the head and neck (Harrington et al., 2001) and for ovarian cancer (Johnston and Gore, 2001). Recently, the group of Maruyama developed a novel type of liposome encapsulating the anticancer drug, oxaliplatin, for active targeting of the cancer cells (Suzuki et al. 2008). !is transferrin- modi$ed liposomal formulation is in a phase II clinical trial with Mebiopharm Co. Ltd., Japan, May 5th, 2009. (http://www.mebiopharm.com/english/index.html).

Passive targeting, on the other hand, uses the natural course followed by the drug-carrier complex upon being introduced to the body as the method of site-speci$c delivery of the drug. It is, therefore, based on the physi-cochemical properties of the drug-carrier complex and anatomical conditions of the body. !e clearance kinetics and in vivo biodistribution of carrier systems depend on their physicochemical factors, such as size, charge, and hydrophobicity and can be manipulated to enable pas-sive targeting. Scientists in the $eld of drug delivery have successfully constructed long-circulating liposomes that accumulate in tumor tissues, where the entrapped drugs subsequently leak out of the vesicles by passive di#usion, unless there is an active trigger present. Nanoliposomes of about 100–200 nm in size passively target solid tumors by extravasation into their extracellular space on intra-venous administration (Hashizume et al., 2000; Maeda et al., 2000; Matsumura and Maeda, 1986; McDonald and Baluk, 2002). Extravasation is achieved due to the disorganized tumor vasculature, as shown schematically in Figure 2. Indications, other than cancer, targeted by liposomal formulations include amphotericin B for the treatment of visceral leishmaniasis (Sundar et al., 2003) and long-acting analgesia with liposomal bupivacaine in healthy subjects (Grant et al., 2004).

!e increasing number of liposomal formulations in clinical trials, as well as on the market today, indicates that these formulations have a very promising future.

Dendrimers

Dendrimer is from the Greek word “dendron” (tree) and “meros” (part). Dendrimers consist of a central core molecule in which highly branched, tree-like arms originate in an ordered, symmetric fashion (Dufes et al., 2005; Gardikis et al., 2006). In comparison with

Normal cells

Bloodvessel

Liposome

Tumour cells

Figure 2. Passive tissue targeting is achieved by extravasation of nanocarriers (here, nanoliposomes) through increased permeability of the tumor vasculature and ine!ective lymphatic drainage—(right) liposome extravasation from the disorganized tumor vasculature and (left) liposomes in normal blood vessel.

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

Role of nanocarrier systems in cancer nanotherapy 315

the traditional macromolecular structures, which are polydisperse, dendrimers have monodisperse struc-tures (Svenson and Tomalia, 2005). !e $rst reports of dendrimers were published in mid-1980s (Tomalia et al., 1985) and focused on the synthesis as well as the chemical and physical characteristics of dendrimers. In recent decades, the unique properties of dendrimers were recognized and their applications in drug delivery, gene therapy, magnetic resonance imaging, develop-ment of vaccines, antibacterials, antivirals, and antican-cer agents were studied (Aulenta et al., 2003; Boas and Heegaard, 2004; Patri et al., 2002; Stiriba et al., 2002).

Synthesis of dendrimers

Two major strategies have been introduced for the synthesis of dendrimers: “divergent method” and “convergent growth process.” In the divergent method, the growth of a dendron starts from a core site, which involves assembling of monomeric modules in a radial form (Tomalia, 1996). In contrast to the divergent method, in the convergent growth method, the synthe-sis proceeds from the surface inward to the focal point, leading to the formation of a dendron. Several dendrons are reacted with a multifunctional core to form a den-drimer (Bosman et al., 1999). Figure 3 shows the above-mentioned strategies for the synthesis of dendrimers and modes of drug association to these nanostructures.

Simplifying the synthesis methods of dendrimers was an issue for the pharmaceutical industry to overcome the commercialization obstacles. Two new strategies have

recently been introduced: the “lego” chemistry and the “click” chemistry. In the “lego” chemistry, highly func-tionalized cores and branched monomers are utilized to synthesize phosphorous dendrimers. !ese dendrimers allow easy puri$cation and produce environmentally safe by-products, such as water and nitrogen (Svenson and Tomalia, 2005). In the “click” chemistry, 1,2,3-triazoles are synthesized from azides and alkynes by the catalytic action of Cu(I) to produce dendrimers with dif-ferent surface groups.

In 1984, the $rst complete dendrimer family, poly(amidoamine) (PAMAM) dendrimers, were syn-thesized (Tomalia and Esfand, 1997). !e PAMAM den-drimers were formed based on an ethylenediamine or ammonia core with four and three branching points, respectively (de Brabander-van den Berg et al., 1994; Dufes et al., 2005; Wörner and Mülhaupt, 1993).

Applications of dendrimers in drug delivery

!e unique structure of dendrimers, in addition to improving the solubility and bioavailability of drugs, can also improve the passive targeting of active moie-ties based on enhanced permeation and retention mechanisms (Al-Jamal et al., 2005). First, studies using dendrimers as drug-delivery vehicles were considering their application as unimolecular micelles and “den-dritic boxes” for noncovalent encapsulation of drug molecules. In these studies, the potential of PAMAM dendrimers as carriers of DNA and hydrophobic drugs was investigated (Haensler and Szoka, 1993; Hawker et al., 1993; Jansen et al., 1994, 1995; Newkome et al., 1992). !e advantage of this method for drug deliv-ery, compared to the polymeric micelles, was that the micellar structure was maintained at all concentrations because the hydrophobic segments are covalently con-nected. However, the di"culty in controlling the release of active molecules from dendrimers was a major draw-back (Gillies and Frechet, 2005; Jansen et al., 1995). !e introduction of stabilizing PEG chains on the dendrimer periphery has enabled the researchers to encapsulate anticancer drugs, such as 5-%uorouracil (Bhadra et al., 2003), methotrexate, and doxorubicin (Kojima et al., 2000), and slow the drug-release rates.

An alternative method for using dendrimers as drug carriers is to utilize their multivalance property and attach drug molecules to their surface by using covalent bonds (Figure 3). Drug release can be controlled by the selection of linkage type between drug and dendrimer periphery. Drug loading can be controlled by chang-ing the generation number of dendrimer (Gillies and Frechet, 2005). For example, the conjugation of cis-platin with PAMAM dendrimers resulted in a complex that exhibits improved solubility, slower release, higher tumor accumulation, and lower toxicity, compared with

Divergent synthesisA

B

Convergent synthesis

+

Encapsulation Complexation

Convergent synthesis

Figure 3. (A) Divergent and convergent strategies for the synthesis of dendrimers. (B) Drug incorporation to dendrimers via encapsulation (entrapment of drugs inside the core of a dendrimer) or complexa-tion (covalent attachment of drug molecules to end groups).

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

316 M. R. Mozafari et al.

free cisplatin (Duncan and Malik, 1996; Malik et al., 1999). Zhou et al. (1999) synthesized 5-%uorouracil conjugated to the periphery of a cyclic tetraamine dendrimer and showed that the conjugates can release 5-%uorouracil when incubated in phosphate-bu#ered saline. In other studies, dendrimer conjugates of Ara-C, a chemotherapeutic drug, were synthesized and the results showed that dendrimer conjugates can increase the blood circulation time of the drug and improve the drug stability, compared with the free drug (Choe et al., 2002; Schiavon et al., 2004).

Another approach was to attach the drug to the peripheral dendron arms by a biodegradable peptide linker. Conjugates of doxorubicin was prepared by the conjugation of poly [N-(2-hydroxypropyl) methacry-lamide] macromonomers to PAMAM dendrimers, fol-lowed by attachment of the drug to the polymer (Wang et al., 2000). !e prepared product showed slower drug release and decreased toxicity, compared to the drug solution. In another research study, Morgan et al. (2006) used a biocompatible polyester dendrimer composed of the natural metabolites, glycerol and succinic acid, for the encapsulation of camptothecin. !ey studied cytotoxicity of the dendrimer-drug complex and free drug in four di#erent human cancer cell lines. Results of their study showed that dendrimer-conjugated drug can increase cellular uptake 16-fold in MCG-7 cells and enhance drug retention within the cells, when compared with free form of the drug (Morgan et al., 2006). In another study performed by Lai et al. (2007), doxorubicin (DOX) was conjugated to PAMAM den-drimers via pH-sensitive and -insensitive linkers and was combined with di#erent photochemical internali-zation (PCI) strategies to evaluate the cytotoxic e#ects. !eir results showed that PCI treatment was e"cient in releasing doxorubicin from the PAMAM-hyd-DOX conjugates (prepared by attaching DOX to the hydrazide drug-binding residues of dendrimer) and also resulted in more nuclear accumulation of the drug and more cell death through synergistic e#ects.

Active targeting of the tumor cells by using folic-acid–decorated dendrimers is another approach in improving the e"cacy of drug delivery. Because the folate receptor is expressed in several human cancers, folic acid is an interesting candidate for the active target-ing of dendrimer-drug conjugates to tumors (Sudimack and Lee, 2000). In order to develop tumor-speci$c target-ing using dendrimers, Kono and coworkers synthesized conjugates of methotrexate and polyaryl ether dendrim-ers with folic acid. !e presence of folic acid on the sur-face of the dendrimer enhances water solubility of the conjugates and improves tumor-speci$c targeting of the dendrimers (Kono et al., 1999). In another study, using folate decoration concept, Quintana and coworkers syn-thesized methotrexate-conjugated PAMAM dendrimers

(Quintana et al., 2002). As expected, the introduction of folic acid into these conjugates increased cellular uptake and localized cytotoxicity of the methotrexate, compared with the free drug in vitro.

Targeted therapeutics using antibodies are an attractive option over conventional cancer chemothera-peutics. Based on this concept, Patri and colleagues syn-thesized J591 anti-PSMA (prostate-speci$c membrane antigen) antibody-dendrimer conjugates containing %uorophores on the dendrimer (Patri et al., 2004). !eir in vitro studies showed that the synthesized dendrim-ers speci$cally bind to cells expressing PSMA. Confocal microscopy experiments showed the binding and inter-nalization of these conjugates. In a recent research, Myc et al. used BH3 peptide conjugated with generation 5 poly(amidoamine) dendrimers (G5-PAMAM) to reverse the antiapoptotic mechanism of cancer cells (Myc et al., 2007). !eir results demonstrated the therapeutic poten-tial of dendrimer-conjugated BH3 peptides as a means of inducing apoptosis by interfering with antiapoptotic proteins within speci$c cancer cells.

Nanoparticles

Nanoparticles are solid colloidal particles with particle sizes smaller than 1,000 nm. However, most of the nano-particles utilized in drug delivery are in the size range of 100–200 nm (Allemann et al., 1993; Moghimi et al., 2006). Nanoparticles can be classi$ed into two main subgroups: nanospheres and nanocapsules (Figure 4). Nanospheres have a matrix-type structure, and drug molecules can

Encapsulated drug

Absorbed drug

Nanocapsules Nanospheres

Figure 4. Schematic presentation of drug-loaded nanoparticles, which are small polymeric colloidal particles with a therapeutic agent either dispersed in the polymer matrix or encapsulated in the polymer or adsorbed to the particle surface.

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

Role of nanocarrier systems in cancer nanotherapy 317

be adsorbed on their surface or entrapped inside their matrix. Nanocapsules have a capsule-like structure and possess the capability of encapsulating the drug mol-ecules inside the capsule or adsorbed to them externally (De Jaeghere et al., 1999). Because these systems have unique characteristics, such as very small particle size, high surface area, and possibility of surface modi$cation, they have been attracting much interest for drug-delivery purposes during recent years (Moghimi et al., 2006).

Synthesis of nanoparticles

Nanoparticles are usually synthesized from either preformed polymeric materials or by polymerization of monomers (Moghimi et al., 2006). Preformed mac-romolecules can be from natural or synthetic origin. Commonly used methods for the preparation of nano-particles include solvent evaporation (Desoguilles et al., 2003; Scholes et al., 1993), solvent di#usion (Niwa et al., 1993), desolvation (Azarmi et al., 2006a), and emulsion polymerization (Vauthier et al., 2003). !e choice of method for the preparation of nanoparticles depends on the drug characteristics, the polymer, the required particle size, and purpose of administration.

In the solvent evaporation method, polymer and drug are dissolved in an organic solvent and the mixture is emulsi$ed to form oil in water emulsion. !is method is used for the encapsulation of hydrophobic drugs with high loading e"ciency. For the encapsulation of hydrophilic drugs, multiple emulsi$cation methods are used (Moghimi et al., 2006). A $nal step in nanoparticle preparation is evaporation of the organic solvent.

In the desolvation method, which is normally used for natural macromolecules, such as gelatin and albumin, desolvation of macromolecule is rendered by the addi-tion of a nonsolvent, such as acetone or alcohol. A $nal cross-linking, using glutaraldehyde or formaldehyde, is used after the formation of nanoparticles (Azarmi et al., 2006a). In the emulsion polymerization method, sponta-neous polymerization of monomers (e.g., cyanoacrylate monomers) happens in a controlled acidic medium. !e particle size of the nanoparticles mostly depends on the pH of the polymerization medium. Stabilizing agents such as dextran (Zanta et al., 1997) or pluronic are usu-ally added into the nanoparticle medium (Vauthier et al., 2003).

Targeted delivery using nanoparticles

Most cancer drugs do not di#erentiate between normal and cancer cells. !ese drugs are administered in high doses to reach the tumor site. !erefore, an optimum drug concentration in the tumor is achieved at the expense of exposing other organs to high drug concentrations, which results in serious side e#ects (Nie et al., 2007).

Nanoparticles o#er a targeted approach, which can be used for improving cancer therapy. However, depending on their surface characteristics, nanoparti-cles will be taken up by the liver, spleen, and other parts of the reticuloendothelial system (RES) (Nie et al., 2007). Hydrophobic nanoparticles will be preferentially taken up by RES organs. It has been shown that hydrophilic particles can remain in the circulation for a longer time and are taken up by liver to a lesser extent (Araujo et al., 1999; Moghimi et al., 2005). Di#erent strategies have been used to make a hydrophilic cloud around the nano-particles and decrease their uptake by RES organs. !ese strategies include coating of nanoparticles with Tween 80 (Gelperina et al., 2002), PEG (Tang et al., 2007), and poloxamers and poloxamines (Moghimi and Hunter, 2000). It is known that the vascular system around the tumor is defective and leaky. !e leaky characteristic of tumor vessels enables the accumulation of nano-particles in a tumor, which is an enhanced permeation and retention mechanism (Chari, 1998; Duncan, 2003; Matsumura and Maeda, 1986).

Active targeting is normally achieved by conjugating a targeting moiety to the nanoparticle. !e targeting moiety is selective to tumor site and can increase the accumulation of nanoparticles in the tumor (Nie et al., 2007). Antibody-oriented (Patri et al., 2004) and folate-decorated nanoparticles (Park et al., 2005) are examples of active targeting. Overexpression of antigens in human cancer cells provides a means for active targeting and allows those cancer antigens to be targeted by using nanoparticles conjugated with speci$c antibodies. !is mechanism results in the accumulation of the drug-loaded nanoparticles at the tumor site (Blagosklonny, 2003; Tsuruo, 2003). Folate receptors are also overex-pressed in some human cancer cells, which enabled researchers to develop folate-decorated nanoparticles for targeted cancer therapy (Leamon and Low, 2001; Leamon and Reddy, 2004).

Delivery of chemotherapy drugs

In this section, some examples are given for the role and usefulness of nanocarrier systems in the delivery of anti-cancer agents.

Paclitaxel

Paclitaxel is a widely used chemotherapeutic agent in the treatment of ovarian cancer as well as breast, colon, and lung cancers (Brannon-Peppas and Blanchette, 2004). Paclitaxel has poor aqueous solubility and is soluble in organic solvents. !e available formulations of paclitaxel contain Cremophor EL (polyethoxylated castor oil), which is toxic and shows hypersensitivity,

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

318 M. R. Mozafari et al.

nephrotoxicity, and neurotoxicity reactions (Singla et al., 2002). Biodegradable nanocarriers could be an ideal solution to such problems and could achieve controlled, targeted drug delivery with better e"cacy and fewer side e#ects. Nanoparticles containing paclitaxel have been formulated by using PLGA (polylactide-co-glycolide) with very high loading e"ciencies. !ese formulations have shown more cytotoxic e#ect when incubated with lung cancer cells in vitro, compared with the free drug solution (Wang et al., 1996). A study involving the HT-29 cancer cell line showed that after 24 hours of incubation, the cell mortality caused by a nanoparticle formulation of paclitaxel was more than 13 times higher than the mortality caused by the free drug under similar condi-tions (Wei et al., 2007). An albumin-paclitaxel nanopar-ticle (Abraxane) has been recently approved by the U.S. Food and Drug Administration (FDA) for the treatment of metastatic breast cancer (Gradishar, 2006).

Doxorubicin

Doxorubicin is another potent, widely used chemother-apeutic agent. Several studies were performed on the formulation and cytotoxicity of doxorubicin nanoparti-cles in vitro and in vivo. Doxorubicin-loaded polyalkyl cyanoacrylate nanoparticles have shown higher cytotoxic e#ect against lung cancer cells, compared with doxoru-bicin solutions in vitro (Azarmi et al., 2006b). In another study, it was shown that mice injected intravenously with doxorubicin-loaded chitosan nanoparticles showed a signi$cant decrease in tumor size, whereas treatment with doxorubicin solution did not decrease tumor size (Mitra et al., 2001). Another group studied the in vivo e#ectiveness of doxorubicin-loaded PLGA nanoparticles and showed that a single injection of doxorubicin-PLGA nanoparticles can suppress tumor growth for up to 12 days postadministration (Yoo and Park, 2000).

Camptothecin

Camptothecin is the prototype compound of a relatively new class of anticancer drugs known as topoisomerase inhibitors (e.g., irinotecan, topotecan). !ese therapeu-tic agents stabilize the binding of topoisomerase I to DNA and cause fragmentation of DNA in the G2-phase of the cell cycle, which result in cell death. !e anticancer activity of topoisomerase inhibitors is based on the func-tionality of the lactone ring of the drug molecule (Wong et al., 2007). Camptothecin-based drugs, due to their poor solubility and labile lactone ring, pose challenges for drug delivery. A possible solution to the application of camptothecin is its encapsulation in nanocarrier systems, such as dendrimers or nanoparticles. In an in vitro study, Morgan and colleagues showed that encap-sulation of camptothecin drugs in dendrimers increased

their solubility, cellular uptake, and cellular retention, which, consequently, improved the anticancer activity of the drug (Morgan et al., 2006). Further, it was shown that nanoparticle formulations of irinotecan possess better e"cacy, compared with the free form of the drug (unen-capsulated drug) (Williams et al., 2003). In an in vivo study, mice were injected twice-weekly for 2 weeks or daily for 10 days, with a camptothecin-loaded nanopar-ticle formulation. A longer tumor regression and survival time was observed for mice injected with the nanopar-ticle formulation, compared to Camptosar® (irinotecan hydrochloride injection) (Williams et al., 2003). !ese studies clearly demonstrate the advantages of employ-ing nanocarrier technologies in cancer therapy.

Conclusions

Compared to other therapeutic agents, anticancer drugs and, particularly, the cytotoxic compounds are more reactive, unstable, and toxic. Nanotechnology-based car-rier systems could be an ideal solution to such problems and could achieve controlled and targeted drug delivery with better e"cacy and fewer side e#ects. Nanocarriers provide a new means of anticancer drug application, exploiting various routes of drug delivery, ranging from oral to transdermal. Current nanocarrier technologies have been shown to be superior to conventional drug solutions in almost every aspect, such as drug e"cacy, pharmacokinetics, and drug biodistribution. Given the remarkable advantages of the nanocarrier-based systems, it is not surprising that this new class of drug carriers is quickly being adopted for the delivery of vari-ous anticancer compounds. !e coming years promise a revolutionary change in the delivery of anticancer drugs for millions of su#erers worldwide.

Acknowledgments

Declaration of interest: !e authors report no $nancial con%icts of interest. !e authors alone are responsible for the content and writing of this paper.

References

Al-Jamal KT, Ramaswamy C, Florence AT. (2005). Supramolecular structures from dendrons and dendrimers. Adv Drug Deliv Rev. 57:2238–2270.

Allemann E, Gurny R, Doelker E. (1993). Drug-loaded nanoparticles—preparation methods and drug targeting issues. Eur J Pharm Biopharm. 39:173–191.

Allen TM, Cullis PR. (2004). Drug delivery systems: entering the mainstream. Science. 303:1818–1822.

Andresen TL, Jensen SS, Jorgensen K. (2005). Advanced strategies in liposomal cancer therapy: problems and prospects of active and tumor speci"c drug release. Prog Lipid Res. 44:68–97.

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

Role of nanocarrier systems in cancer nanotherapy 319

Araujo L, Lobenberg R, Kreuter J. (1999). In#uence of the surfactant concentration on the body distribution of nanoparticles. J Drug Targ. 6:373–385.

Aulenta F, Hayes W, Rannard S. (2003). Dendrimers: a new class of nanoscopic containers and delivery devices. Eur Polym J. 39:1741–1771.

Azarmi S, Huang Y, Chen H, McQuarrie S, Abrams D, Roa W, et al. (2006a). Optimization of a two-step desolvation method for preparing gelatin nanoparticles and cell uptake studies in 143B osteosarcoma cancer cells. J Pharm Pharm Sci. 9:124–132.

Azarmi S, Tao X, Chen H, Wang Z, Finlay WH, Lobenberg R, et al. (2006b). Formulation and cytotoxicity of doxorubicin nano-particles carried by dry powder aerosol particles. Int J Pharm. 319:155–161.

Bhadra D, Bhadra S, Jain S, Jain NK. (2003). A PEGylated dendritic nan-oparticulate carrier of #uorouracil. Int J Pharm. 257:111–124.

Blagosklonny MV. (2003). Targeting cancer cells by exploiting their resistance. Trends Mol Med. 9:307–312.

Boas U, Heegaard PM. (2004). Dendrimers in drug research. Chem Soc Rev. 33:43–63.

Bosman AW, Janssen HM, Meijer EW. (1999). About dendrimers: structure, physical properties, and applications. Chem Rev. 99:1665–1688.

Brannon-Peppas L, Blanchette JO. (2004). Nanoparticle and targeted systems for cancer therapy. Adv Drug Deliv Rev. 56:1649–1659.

Butts C, Murray N, Maksymiuk A, Goss G, Marshall E, Soulieres D, et al. (2005). Randomized phase IIB trial of BLP25 liposome vaccine in stage IIIB and IV non-small-cell lung cancer. J Clin Oncol. 23:6674–6681.

Cammas SK. (1996). NATO ASI Series, Series E. In: Webber SE (Ed.), Solvents and Self-Organization of Polymers (pp 83–113). Amsterdam, $e Netherlands: Kluwer Academic.

Chari RV. (1998). Targeted delivery of chemotherapeutics: tumor-activated prodrug therapy. Adv Drug Deliv Rev. 31:89–104.

Chaurasia M, Chourasia MK, Jain NK, Jain A, Soni V, Gupta Y, et al. (2006). Cross-linked guar gum microspheres: a viable approach for improved delivery of anticancer drugs for the treatment of colorectal cancer. AAPS PharmSciTech. 7:74.

Cheng MM, Cuda G, Bunimovich YL, Gaspari M, Heath JR, Hill HD, et al. (2006). Nanotechnologies for biomolecular detection and medical diagnostics. Curr Opin Chem Biol. 10:11–19.

Cheung RY, Rauth AM, Ronaldson PT, Bendayan R, Wu XY. (2006). In vitro toxicity to breast cancer cells of microsphere-delivered mitomycin C and its combination with doxorubicin. Eur J Pharm Biopharm. 62:321–331.

Choe YH, Conover CD, Wu D, Royzen M, Gervacio Y, Borowski V, et al. (2002). Anticancer drug delivery systems: multiloaded N4-acyl poly(ethylene glycol) prodrugs of ara-C. II. E%cacy in ascites and solid tumors. J Contr Rel. 79:55–70.

Colas JC, Shi W, Rao VS, Omri A, Mozafari MR, Singh H. (2007). Microscopical investigations of nisin-loaded nanoliposomes prepared by Mozafari method and their bacterial targeting. Micron. 38:841–847.

Cortesi R, Esposito E, Gambarin S, Telloli P, Menegatti E, Nastruzzi C. (1999). Preparation of liposomes by reverse-phase evaporation using alternative organic solvents. J Microencapsul. 16:251–256.

Daemen T, Hofstede G, Ten Kate MT, Bakker-Woudenberg IA, Scherphof GL. (1995). Liposomal doxorubicin-induced toxicity: depletion and impairment of phagocytic activity of liver macro-phages. Int J Cancer. 61:716–721.

Dass CR, Choong PF. (2006). Selective gene delivery for cancer ther-apy using cationic liposomes: in vivo proof of applicability. J Contr Rel. 113:155–163.

De Brabander-van den Berg EMM, Nijenhuis A, Mure M, Keulen J, Reintjens R, Vandenbooren F, et al. (1994). Large-scale pro-duction of polypropylenimine dendrimers. Macromolecular Symposia, International Symposium on New Macromolecular Architectures and Supramolecular Polymers. 77:51–62.

De Jaeghere F, Doelker E, Gurny R. (1999). Nanospheres. In: Mathiowitz E. (Ed.), Encyclopedia of Controlled Drug Delivery (pp 641–656). New York: John Wiley & Sons.

Deamer DW, Uster PS. (1983). Liposome preparation methods. In: Ostro MJ (Ed.), Liposomes (pp 27–51). New York: Marcel Dekker.

Desoguilles S, Vauthier C, Bazile D, Vacus J, Grossiord JL, Veillard M, et al. (2003). $e design of nanoparticles obtained by solvent evaporation: a comprehensive study. Langmuir. 19:9504–9510.

Dufes C, Uchegbu IF, Schtzlein AG. (2005). Dendrimers in gene deliv-ery. Adv Drug Deliv Rev. 57:2177–2202.

Duncan R. (2003). $e dawning era of polymer therapeutics. Nat Rev Drug Discov. 2:347–360.

Duncan R, Malik N. (1996). Dendrimers: biocompatibility and potential for delivery of anticancer agents. Proceedings of the International Symposium on Controlled Release of Bioactive Materials. 23:105–106.

Dwivedi AM. (2002). Residual solvent analysis in pharmaceuticals. Pharm Technol Eur. 14:26–28.

Emerich DF. (2005). Nanomedicine—prospective therapeutic and diagnostic applications. Expert Opin Biol !er. 5:1–5.

Frenkel V. (2008). Ultrasound mediated delivery of drugs and genes to solid tumors. Adv Drug Deliv Rev. 60:1193–1208.

Gardikis K, Hatziantoniou S, Viras K, Wagner M, Demetzos C. (2006). Interaction of dendrimers with model lipid membranes assessed by DSC and RAMAN spectroscopy. In: Mozafari MR (Ed.), Nanocarrier Technologies: Frontiers of Nanotherapy (pp 207–220). $e Netherlands: Springer.

Gelperina SE, Khalansky AS, Skidan IN, Smirnova ZS, Bobruskin AI, Severin SE, et al. (2002). Toxicological studies of doxorubicin bound to polysorbate 80-coated poly(butyl cyanoacrylate) nanoparticles in healthy rats and rats with intracranial glioblas-toma. Toxicol Lett. 126:131–141.

Gibaud S, Rousseau C, Weingarten C, Favier R, Douay L, Andreux JP, et al. (1998). Polyalkylcyanoacrylate nanoparticles as carriers for granulocyte-colony stimulating factor (G-CSF). J Contr Rel. 52:131–139.

Gillies ER, Frechet JM. (2005). Dendrimers and dendritic polymers in drug delivery. Drug Discov Today. 10:35–43.

Goncalves A, Braud AC, Viret F, Genre D, Gravis G, Tarpin C, et al. (2003). Phase I study of pegylated liposomal doxorubicin (Caelyx) in combination with carboplatin in patients with advanced solid tumors. Anticancer Res. 23:3543–3548.

Gradishar WJ. (2006). Albumin-bound paclitaxel: a next-generation taxane. Expert Opin Pharmacother. 7:1041–1053.

Grant GJ, Barenholz Y, Bolotin EM, Bansinath M, Turndorf H, Piskoun B, et al. (2004). A novel liposomal bupivacaine formu-lation to produce ultralong-acting analgesia. Anesthesiology. 101:133–137.

Guan HH, Budzynski W, Koganty RR, Krantz MJ, Reddish MA, Rogers JA, et al. (1998). Liposomal formulations of synthetic MUC1 peptides: e!ects of encapsulation versus surface display of peptides on immune responses. Bioconjug Chem. 9:451–458.

Haensler J, Szoka FC, Jr. (1993). Polyamidoamine cascade polymers mediate e%cient transfection of cells in culture. Bioconjug Chem. 4:372–379.

Harrington KJ, Lewanski C, Northcote AD, Whittaker J, Peters AM, Vile RG, et al. (2001). Phase II study of pegylated liposomal doxorubicin (Caelyx™) in patients with inoperable, locally advanced squamous cell cancer of the head and neck. Eur J Cancer. 37:2015–2022.

Harrington KJ, Rowlinson-Busza G, Syrigos KN, Abra RM, Uster PS, Peters AM, et al. (2000a). In#uence of tumour size on uptake of (111)ln-DTPA-labelled pegylated liposomes in a human tumour xenograft model. Br J Cancer. 83:684–688.

Harrington KJ, Rowlinson-Busza G, Syrigos KN, Uster PS, Abra RM, Stewart JS. (2000b). Biodistribution and pharmacokinetics of 111In-DTPA-labelled pegylated liposomes in a human tumour xenograft model: implications for novel targeting strategies. Br J Cancer 83:232–238.

Hashizume H, Baluk P, Morikawa S, McLean JW, $urston G, Roberge S, et al. (2000). Openings between defective endothelial cells explain tumor vessel leakiness. Am J Pathol. 156:1363–1380.

Hawker CJ, Wooley KL, Frechet JMJ. (1993). Unimolecular micelles and globular amphiphiles: dendritic macromolecules as novel recyclable solubilization agents. J Chem Soc. 1:1287–1297.

Hobbs SK, Monsky WL, Yuan F, Roberts WG, Gri%th L, Torchilin VP, et al. (1998). Regulation of transport pathways in tumor vessels:

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

320 M. R. Mozafari et al.

role of tumor type and microenvironment. Proc Natl Acad Sci U S A. 95:4607–4612.

Hood JD, Bednarski M, Frausto R, Guccione S, Reisfeld RA, Xiang R, et al. (2002). Tumor regression by targeted gene delivery to the neovasculature. Science. 296:2404–2407.

Huang G, Zhou Z, Srinivasan R, Penn MS, Kottke-Marchant K, Marchant RE, et al. (2008). A%nity manipulation of surface-conjugated RGD peptide to modulate binding of liposomes to activated platelets. Biomaterials. 29:1676–1685.

Hughes GA. (2005). Nanostructure-mediated drug delivery. Nanomedicine. 1:22–30.

Jansen JF, de Brabander-van den Berg EM, Meijer EW. (1994). Encapsulation of guest molecules into a dendritic box. Science. 266:1226–1229.

Jansen JFGA, Meijer EW, De Brabander-van den Berg EMM. (1995). $e dendritic box: shape-selective liberation of encapsulated guests. J Am Chem Soc. 117:4417–4418.

Johnsson M, Bergstrand N, Edwards K. (1999). Optimization of drug loading procedures and characterization of liposomal formula-tions of two novel agents intended for boron neutron capture therapy (BNCT). J Liposome Res. 9:53–79.

Johnston SR, Gore ME. (2001). Caelyx: phase II studies in ovarian cancer. Eur J Cancer. 37(Suppl 9):S8–S14.

Karanth H, Murthy RS. (2007). pH-sensitive liposomes—principle and application in cancer therapy. J Pharm Pharmacol. 59:469–483.

Kasaai MR, Charlet G, Paquin P, Arul J. (2003). Fragmentation of chitosan by micro#uidization process. Innov Food Sci Emerg Technol. 4:403–413.

Kawashima Y. (2001). Panoparticulate systems for improved drug delivery. Adv Drug Deliv Rev. 47:1–2.

Khosravi-Darani K, Pardakhty A, Honarpisheh H, Rao VS, Mozafari MR. (2007). $e role of high-resolution imaging in the evaluation of nanosystems for bioactive encapsulation and tar-geted nanotherapy. Micron. 38:804–818.

Kojima C, Kono K, Maruyama K, Takagishi T. (2000). Synthesis of polyamidoamine dendrimers having poly(ethylene glycol) grafts and their ability to encapsulate anticancer drugs. Bioconjug Chem. 11:910–917.

Kono K, Kojima C, Hayashi N, Nishisaka E, Kiura K, Watarai S, et al. (2008). Preparation and cytotoxic activity of poly(ethylene glycol)-modi"ed poly(amidoamine) dendrimers bearing adri-amycin. Biomaterials. 29:1664–1675.

Kono K, Liu M, Frechet JM. (1999). Design of dendritic macromol-ecules containing folate or methotrexate residues. Bioconjug Chem. 10:1115–1121.

Lai PS, Lou PJ, Peng CL, Pai CL, Yen WN, Huang MY, et al. (2007). Doxorubicin delivery by polyamidoamine dendrimer conju-gation and photochemical internalization for cancer therapy. J Contr Rel. 122:39–46.

Langner, M., Kozubek, A. (2006). Pharmacokinetic modula-tion with particulate drug formulations. In: Mozafari MR (Ed.), Nanocarrier Technologies Frontiers of Nanotherapy (pp 113–138). $e Netherlands: Springer.

Leamon CP, Low PS. (2001). Folate-mediated targeting: from diag-nostics to drug and gene delivery. Drug Discov Today. 6:44–51.

Leamon CP, Reddy JA. (2004). Folate-targeted chemotherapy. Adv Drug Deliv Rev. 56:1127–1141.

Lemoine D, Preat V. (1998). Polymeric nanoparticles as delivery sys-tem for in#uenza virus glycoproteins. J Contr Rel. 54:15–27.

Lu Y, Low PS. (2003). Immunotherapy of folate receptor-expressing tumors: review of recent advances and future prospects. J Contr Rel. 91:17–29.

Maeda H, Wu J, Sawa T, Matsumura Y, Hori K. (2000). Tumor vascular permeability and the EPR e!ect in macromolecular therapeu-tics: a review. J Contr Rel. 65:271–284.

Mahmud A, Xiong XB, Aliabadi HM, Lavasanifar A. (2007). Polymeric micelles for drug targeting. J Drug Target. 15:553–584.

Malik N, Evagorou EG, Duncan R. (1999). Dendrimer-platinate: a novel approach to cancer chemotherapy. Anticancer Drugs. 10:767–776.

Matsumura Y, Maeda H. (1986). A new concept for macromolecular therapeutics in cancer chemotherapy: mechanism of tumori-tropic accumulation of proteins and the antitumor agent smancs. Cancer Res. 46:6387–6392.

McDonald DM, Baluk P. (2002). Signi"cance of blood vessel leakiness in cancer. Cancer Res. 62:5381–5385.

Mehta J, Kelsey S, Chu P, Powles R, Hazel D, Riley U, et al. (1997). Amphotericin B lipid complex (ABLC) for the treatment of con-"rmed or presumed fungal infections in immunocompromised patients with hematologic malignancies. Bone Marrow Transpl. 20:39–43.

Mitra S, Gaur U, Ghosh PC, Maitra AN. (2001). Tumour targeted delivery of encapsulated dextran-doxorubicin conjugate using chitosan nanoparticles as carrier. J Contr Rel. 74:317–323.

Moghimi SM, Hunter AC. (2000). Poloxamers and poloxamines in nanoparticle engineering and experimental medicine. Trends Biotechnol. 18:412–420.

Moghimi SM, Hunter AC, Murray JC. (2005). Nanomedicine: current status and future prospects. FASEB J. 19:311–330.

Moghimi SM, Vega E, Garcia ML, Al-Hanbali OAR, Rutt KJ. (2006). Polymeric nanoparticles as drug carriers and controlled release implant devices. In: Torchilin VP (Ed.), Nanoparticulates as Drug Carriers (pp 29–42). London: Imperial College Press.

Monnard PA, Deamer DW. (2001). Nutrient uptake by protocells: a liposome model system. Orig Life Evol Biosph. 31:147–155.

Morgan MT, Nakanishi Y, Kroll DJ, Griset AP, Carnahan MA, Wathier M, et al. (2006). Dendrimer-encapsulated camptothecins: increased solubility, cellular uptake, and cellular retention a!ords enhanced anticancer activity in vitro. Cancer Res. 66:11913–11921.

Morrow KJ, Jr, Bawa R, Wei C. (2007). Recent advances in basic and clinical nanomedicine. Med Clin North Am. 91:805–843.

Mozafari MR, Reed CJ, Rostron C. (2004). Formation of the initial cell membranes under primordial Earth conditions. Cell Mol Biol Lett. 9 (S2), 97–99.

Mozafari MR. (2005). Liposomes: an overview of manufacturing techniques. Cell Mol Biol Lett. 10:711–719.

Mozafari MR. (2006). Bioactive entrapment and targeting using nano-carrier technologies: an introduction. Nanocarrier Technologies Frontiers of Nanotherapy. $e Netherlands: Springer.

Mozafari MR, Flanagan J, Matia-Merino L, Omri A, Suntres ZE, Singh H. (2006). Recent trends in the lipid-based nanoencap-sulation of antioxidants and their role in foods. J Sci Food Agric. 86:2038–2045.

Mozafari MR, Mortazavi SM. (2005). Nanoliposomes: from Fundamentals to Recent Developments. Oxford, UK: Tra!ord Publishing Ltd.

Mozafari MR, Reed CJ, Rostron C. (2007). Cytotoxicity evaluation of anionic nanoliposomes and nanolipoplexes prepared by the heating method without employing volatile solvents and deter-gents. Pharmazie. 62:205–209.

Mozafari MR, Reed CJ, Rostron C, Kocum C, Piskin E. (2002). Construction of stable anionic liposome-plasmid particles using the heating method: a preliminary investigation. Cell Mol Biol Lett. 7:923–927.

Myc A, Patri AK, Baker JR, Jr. (2007). Dendrimer-based BH3 conju-gate that targets human carcinoma cells. Biomacromolecules. 8:2986–2989.

Newkome GR, Nayak A, Behera RK, Moore"eld CN, Baker GR. (1992). Cascade polymers: synthesis and characterization of four- directional spherical dendritic macromolecules based on ada-mantane. J Organic Chem. 57:358–362.

Nie S, Xing Y, Kim GJ, Simons JW. (2007). Nanotechnology applica-tions in cancer. Annu Rev Biomed Eng. 9:257–288.

Niwa T, Takeuchi H, Hino T, Kunou N, Kawashima Y. (1993). Preparations of biodegradable nanospheres of water-soluble and insoluble drugs with D,L-lactide/glycolide copolymer by a novel spontaneous emulsi"cation solvent di!usion method, and the drug release behavior. J Contr Rel. 25:89–98.

Nomura SM, Yoshikawa Y, Yoshikawa K, Dannenmuller O, Chasserot-Golaz S, Ourisson G, et al. (2001). Towards proto-cells: “primi-tive” lipid vesicles encapsulating giant DNA and its histone complex. Chembiochem. 2:457–459.

Pan H, Han L, Chen W, Yao M, Lu W. (2008). Targeting to tumor necrotic regions with biotinylated antibody and streptavidin modi"ed liposomes. J Contr Rel. 125:228–235.

Park EK, Lee SB, Lee YM. (2005). Preparation and characterization of methoxy poly(ethylene glycol)/poly(epsilon-caprolactone)

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.

Role of nanocarrier systems in cancer nanotherapy 321

amphiphilic block copolymeric nanospheres for tumor-speci"c folate-mediated targeting of anticancer drugs. Biomaterials. 26:1053–1061.

Patri AK, Majoros IJ, Baker JR. (2002). Dendritic polymer macro-molecular carriers for drug delivery. Curr Opin Chem Biol. 6:466–471.

Patri AK, Myc A, Beals J, $omas TP, Bander NH, Baker JR, Jr. (2004). Synthesis and in vitro testing of J591 antibody-dendrimer con-jugates for targeted prostate cancer therapy. Bioconjug Chem. 15:1174–1181.

Peek LJ, Middaugh CR, Berkland C. (2008). Nanotechnology in vac-cine delivery. Adv Drug Deliv Rev. 60: 915–928.

Perez AT, Domenech GH, Frankel C, Vogel CL. (2002). Pegylated liposomal doxorubicin (Doxil) for metastatic breast cancer: the Cancer Research Network, Inc., experience. Cancer Invest. 20(Suppl 2):22–29.

Pozzi G, Birault V, Werner B, Dannenmuller O, Nakatani Y, Ourisson G, et al. (1996). Single-chain polyprenyl phosphates form primitive membranes. Angewandte Chemie. 35:177–180.

Quintana A, Raczka E, Piehler L, Lee I, Myc A, Majoros I, et al. (2002). Design and function of a dendrimer-based therapeutic nanode-vice targeted to tumor cells through the folate receptor. Pharm Res. 19:1310–1316.

Sahoo SK, Labhasetwar V. (2003). Nanotech approaches to drug delivery and imaging. Drug Discov Today. 8:1112–1120.

Sahoo SK, Parveen S, Panda JJ. (2007). $e present and future of nan-otechnology in human health care. Nanomedicine. 3:20–31.

Schiavon O, Pasut G, Moro S, Orsolini P, Guiotto A, Veronese FM. (2004). PEG-Ara-C conjugates for controlled release. Eur J Med Chem. 39:123–133.

Scholes PD, Coombes AGA, Illum L, Daviz SS, Vert M, Davies MC. (1993). $e preparation of sub-200-nm poly(lactide-co- glycolide) microspheres for site-speci"c drug delivery. J Contr Rel. 25:145–153.

Schwonzen M, Kurbacher CM, Mallmann P. (2000). Liposomal doxo-rubicin and weekly paclitaxel in the treatment of metastatic breast cancer. Anticancer Drugs. 11:681–685.

Sha!er C. (2005). Nanomedicine transforms drug delivery. Drug Discov Today. 10:1581–1582.

Shehata M, Mukherjee A, Sharma R, Chan S. (2007). Liposomal doxo-rubicin in breast cancer. Women’s Health. 3:557–569.

Singla AK, Garg A, Aggarwal D. (2002). Paclitaxel and its formula-tions. Int J Pharm. 235:179–192.

Sofou S, Sgouros G. (2008). Antibody-targeted liposomes in cancer therapy and imaging. Expert Opin Drug Deliv. 5:189–204.

Song CX, Labhasetwar V, Murphy H, Qu X, Humphrey WR, Shebuski RJ, et al. (1997). Formulation and characterization of biodegradable nanoparticles for intravascular local drug deliv-ery. J Contr Rel. 43:197–212.

Stiriba SE, Frey H, Haag R. (2002). Dendritic polymers in biomedical applications: from potential to clinical use in diagnostics and therapy. Angew Chem Int Ed Engl. 41:1329–1334.

Sudimack J, Lee RJ. (2000). Targeted drug delivery via the folate recep-tor. Adv Drug Deliv Rev. 41:147–162.

Sun B, Ranganathan B, Feng SS. (2008). Multifunctional poly(D,L-lactide-co-glycolide)/montmorillonite (PLGA/MMT) nanopar-ticles decorated by trastuzumab for targeted chemotherapy of breast cancer. Biomaterials. 29:475–486.

Sundar S, Jha TK, $akur CP, Mishra M, Singh VP, Bu!els R. (2003). Single-dose liposomal amphotericin B in the treatment of vis-ceral leishmaniasis in India: a multicenter study. Clin Infect Dis. 37:800–804.

Suntres ZE, Shek PN. (1995). Liposome-associated antioxidants for pulmonary applications. In: Shek PN (Ed.), Biomedical Applications of Liposomes (pp 179–198). Singapore: Harwood Academic.

Suntres ZE, Shek PN. (1996). Alleviation of paraquat-induced lung injury by pretreatment with bifunctional liposomes contain-ing alpha-tocopherol and glutathione. Biochem Pharmacol. 52:1515–1520.

Suzuki R, Takizawa T, Kuwata Y, Mutoh M, Ishiguro N, Utoguchi N, et al. (2008). E!ective anti-tumor activity of oxaliplatin encapsulated in transferrin-PEG-liposome. Int J Pharm. 346(1–2):143–150.

Svenson S, Tomalia DA. (2005). Dendrimers in biomedical applications—re#ections on the "eld. Adv Drug Deliv Rev. 57:2106–2129.

Tang N, Du G, Wang N, Liu C, Hang H, Liang W. (2007). Improving penetration in tumors with nanoassemblies of phospholipids and doxorubicin. J Natl Cancer Inst. 99:1004–1015.

Tomalia DA. (1996). Starburst dendrimers—nanoscopic supermol-ecules according to dendritic rules and principles. Macromolec Symp. 101:243–255.

Tomalia DA, Baker H, Dewald J, Hall M, Kallos G, Martin S, et al. (1985). A new class of polymers: starburst-dendritic macromol-ecules. Polymer J. 17:117–132.

Tomalia DA, Esfand R. (1997). Dendrons, dendrimers, and dendri-grafts. Chem Industry. 11:416–420.

Tsuruo T. (2003). Molecular cancer therapeutics: recent progress and targets in drug resistance. Intern Med. 42:237–243.

Uchegbu, I.F. (2000). Niosomes and other synthetic surfactant vesi-cles with antitumour drugs. In: Uchegbu IF (Ed.), Synthetic Surfactant Vesicles (pp 115–133). $e Netherlands: Harwood Academic.

Uchegbu IF, Vyas SP. (1998). Non-ionic surfactant based vesicles (niosomes) in drug delivery. Int J Pharm. 172:33–70.

Vasir JK, Labhasetwar V. (2005). Targeted drug delivery in cancer therapy. Technol Cancer Res Treat. 4:363–374.

Vasir JK, Reddy MK, Labhasetwar V. (2005). Nanosystems in drug tar-geting: opportunities and challenges. Curr Nanosci. 1:45–62.

Vauthier C, Dubernet C, Fattal E, Pinto-Alphandary H, Couvreur P. (2003). Poly(alkylcyanoacrylates) as biodegradable materials for biomedical applications. Adv Drug Deliv Rev. 55:519–548.

Vemuri S, Rhodes CT. (1995). Preparation and characterization of liposomes as therapeutic delivery systems: a review. Pharm Acta Helv. 70:95–111.

Wang D, Kopeckova JP, Minko T, Nanayakkara V, Kopecek J. (2000). Synthesis of star-like N-(2-hydroxypropyl)methacrylamide copol-ymers: potential drug carriers. Biomacromolecules. 1:313–319.

Wang YM, Sato H, Adachi I, Horikoshi I. (1996). Preparation and characterization of poly(lactic-co-glycolic acid) microspheres for targeted delivery of a novel anticancer agent, taxol. Chem Pharm Bull 44:1935–1940.

Wei C, Wei W, Morris M, Kondo E, Gorbounov M, Tomalia DA. (2007). Nanomedicine and drug delivery. Med Clin North Am. 91:863–870.

Wilkinson JM. (2003). Nanotechnology applications in medicine. Med Device Technol. 14:29–31.

Williams D. (2004). Nanotechnology: a new look. Med Device Technol. 15:9–10.

Williams J, Lansdown R, Sweitzer R, Romanowski M, LaBell R, Ramaswami R, et al. (2003). Nanoparticle drug delivery system for intravenous delivery of topoisomerase inhibitors. J Contr Rel. 91:167–172.

Wong HL, Bendayan R, Rauth AM, Li Y, Wu XY. (2007). Chemotherapy with anticancer drugs encapsulated in solid lipid nanoparticles. Adv Drug Deliv Rev. 59:491–504.

Wörner C, Mülhaupt R. (1993). Polynitrile- and polyamine-functional poly(trimethylene imine) dendrimers. Angewandte Chemie Int Ed Engl. 32:1306–1308.

Xiang G, Wu J, Lu Y, Liu Z, Lee RJ. (2008). Synthesis and evaluation of a novel ligand for folate-mediated targeting liposomes. Int J Pharm 356:29–36.

Yoo HS, Park TG. (2000). In vitro and in vivo anti-tumor activities of nanoparticles based on doxorubicin-PLGA conjugates. Polym Prep. 41:992–993.

Yurdugul S, Mozafari MR. (2004). Recent advances in micro- and nanoencapsulation of food ingredients. Cell Mol Biol Lett. 9 (S2):64–65.

Zanta MA, Boussif O, Adib A, Behr JP. (1997). In vitro gene delivery to hepatocytes with galactosylated polyethylenimine. Bioconjug Chem. 8:839–844.

Zhang Y, Schlachetzki F, Pardridge WM. (2003). Global non-viral gene transfer to the primate brain following intravenous administra-tion. Mol $er. 7:11–18.

Zhou RX, Du B, Lu ZR. (1999). In vitro release of 5-#uorouracil with cyclic core dendritic polymer. J Contr Rel. 57:249–257.

Journ

al o

f L

iposo

me

Res

earc

h D

ow

nlo

aded

fro

m i

nfo

rmah

ealt

hca

re.c

om

by T

he

Lib

rary

F

or

per

sonal

use

only

.