Review Article
Nanotechnology-Based Drug Delivery Systems for Melanoma
Antitumoral Therapy: A Review

Roberta Balansin Rigon, Márcia Helena Oyafuso,
Andressa Terumi Fujimura, Maíra Lima Gonçalez, Alice Haddad do Prado,
Maria Palmira Daflon Gremião, and Marlus Chorilli

School of Pharmaceutical Sciences, Department of Drug and Medicines, São Paulo State University, 14801-902 Araraquara, SP, Brazil

Correspondence should be addressed to Marlus Chorilli; chorilli@fcfar.unesp.br

Received 9 January 2015; Revised 6 April 2015; Accepted 7 April 2015

Academic Editor: Fabio Sonvico

Copyright © 2015 Roberta Balansin Rigon et al.This is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
cited.

Melanoma (MEL) is a less common type of skin cancer, but it is more aggressive with a high mortality rate. The World
Cancer Research Fund International (GLOBOCAN 2012) estimates that there were 230,000 new cases of MEL in the world in
2012. Conventional MEL treatment includes surgery and chemotherapy, but many of the chemotherapeutic agents used present
undesirable properties.Drug delivery systems are an alternative strategy bywhich to carry antineoplastic agents. Encapsulated drugs
are advantageous due to such properties as high stability, better bioavailability, controlled drug release, a long blood circulation time,
selective organ or tissue distribution, a lower total required dose, and minimal toxic side effects. This review of scientific research
supports applying a nanotechnology-based drug delivery system for MEL therapy.

1. Introduction

Malignant melanoma (MEL) are tumors that mainly affect
adult and elderly patients; the highest incidence is at approx-
imately 60 years of age [1]. However, currently, MEL recur-
rence has increased in young adults and can be observed in
children and adolescents [2].

The World Cancer Research Fund International
(GLOBOCAN 2012) estimates that there were 230,000
new cases of MEL in the world in 2012; MEL incidence rates
are much higher in the White population than in the Black
population, and it is uncommon in the Asian population,
likely due to better protection from their skin pigment and
different sun exposure habits; African and Asian societies
consider fair skin beautiful [3]. In addition, rich populations
have a high rate of MEL with a relatively low rate of mortality
from this disease, potentially because MEL is diagnosed in
early stages for this social class [4–6].

Pathogenesis of Melanoma. Melanocytic skin tumors include
a wide variety of benign and malignant skin lesions with
distinct clinical, morphological, and genetic profiles [7–9].

Melanoma describes melanocyte malignance; a
melanocyte is a melanin-producing cell located in the
basal layer of the epidermis [10]. When it functions normally,
the melanocyte provides basic skin pigmentation and
protects against UV radiation damage [11–13].

In summary, the most significant causes of MEL develop-
ment are at personal history of MEL in the family, advanced
age, the presence an atypical nevus, intense exposure to sun-
light, sunburn during childhood [14], and chronic immuno-
suppression [15]; it is especially observed in posttransplant
patients and patients with acquired immunodeficiency syn-
drome (AIDS) or a prior cancer diagnosis [11].

Genetic predisposition plays an important role in MEL
development due to the relative risk of people with a family
history of MEL developing this cancer, which is 2-3 times
greater than in people without such a family history; several
genes (CDKN2A; BRAFV600E; N-Ras codon 61; CKIT;
GNAQ/GNA11; BRCA2; OCA1 and MC1R) related to this
predisposition have been identified [2, 16–20].

UV radiation also has a profound influence on MEL
development. Sunscreens use, which protect the skin against
this radiation, does not prevent MEL development, because

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Volume 2015, Article ID 841817, 22 pages
http://dx.doi.org/10.1155/2015/841817

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2 BioMed Research International

the UV radiation spectrum that causes erythema (UVB) and
that traditional sunscreens protect against differ from the
spectrum that promotes MEL (UVA). Thus, users of sun-
screens are relatively unprotected from UVA radiation [11].
An alternative theory suggests that vitamin D, which inhibits
the signaling pathway involved inMELdevelopment [21] (i.e.,
the MAP kinase pathway that promotes cell proliferation),
is synthesized upon UV radiation, and when radiation is
blocked by sunscreen, vitamin D synthesis stops [22, 23].

The cutaneous MEL is manifested in different regions
of the body through lesions on the head and neck and is
associated with chronic sun exposure and lesions on the
trunk related to the presence of numerous melanocytic nevi
[24].

Almost all MEL lesions are pigmented and flat; malignant
melanocytes growth is restricted to the epidermis (“MEL
in situ”), and the cells are characterized by a relatively
homogeneous brown pigmentation with slightly irregular
edges [25, 26]. Over time, likely many years, these lesions
present with irregular edges and pigmentation. In late stages,
this neoplastic growth is vertical, and the tumor cells infiltrate
through collagen fibers in the reticular dermis [27]. The
subcutaneous tissue is then infiltrated by the tumor, which
forms papules and nodules, and is typically confined to the
lesion area [28]. Partial regression of the lesion is common,
which functions through an immunemediated phenomenon
that promotes malignant melanocyte elimination by cyto-
toxic lymphocytes [29]. However, complete MEL regression
may be associated with the spread of metastasis, which is a
negative, not positive prognostic sign [2, 30].

For melanocyte transformation in MEL, resistance to
apoptosis is necessary [31], and MELs escape from apopto-
sis stimulation through overexpressing apoptosis-inhibiting
genes (e.g., inhibitor of apoptosis proteins (IAPs), especially
survivin) or decreasing apoptosis-inducing gene expression,
which results in apoptosis dysfunction and an increased risk
of metastasis [32]. The serine/threonine kinase Akt/protein
kinase B and transcription factor nuclear factor-𝜅𝛽 (NF-
𝜅𝛽) participate in the cell proliferation control, apoptosis,
and oncogenesis [33], and certain studies suggest that Akt
activation can facilitate MEL progression by increasing cells
survival through NF-𝜅𝛽 regulation with a consequent reduc-
tion in apoptosis [20].

Classification of Melanoma and Diagnoses. MEL is clinically
classified into fourmain groups [34].The first group is lentigo
maligna MEL, which is characterized by an invasive tumor
in the head, neck, or forearms regions [35]. Another group is
superficial-spreadingMEL, which is characterized by a lesion
with irregular edges and pigmentation that grows laterally
and slowly before promoting vertical invasion [36]. The next
group is nodular MEL, which is a more aggressive type that
appears in the body following high levels of sunlight exposure
[37]. The final group is acral lentiginous MEL, which are
pigmented lesions that appear on the palms of the hands, soles
of the feet, and above the nose [38, 39]. Other classifications
include amelanoticMEL, mucosal MEL, and subungualMEL
[2].

For MEL diagnosis, five main characteristics of the
lesion are analyzed: asymmetry, border-color, diameter, and
elevation; MEL diagnoses are more accurate where der-
matoscopy is used [11]. However, for many people, the first
area that metastasizes is the lymph node (sentinel); the next
most common site of metastasis is distant skin. The organs
more frequently affected are the lungs and the liver; the
central nervous system and bones can also be metastasis sites
[40, 41].

The MEL stage can be determined through a complete
clinical examination [42], including sonography [43] of the
superficial lymph nodes and the abdomen, radiography of the
thorax [44], and evaluation of serummarkers, such as lactate
dehydrogenase (LDH) [45], S-100-beta, sialic acid, enolase, 5-
S-cyseinyldopa, 6-hydroxy-5-methyoxy-indole-2-carboxylic
acid, 3,4-dihydroxy-L-phenylalanine (DOPA), L-tyrosine
[46], computer tomography scan [47], magnetic resonance
imaging [48], bone scintigraphy [49], and positron emission
tomography, which are useful for evaluating patients with
metastatic disease [50].

Moreover, an immunohistochemical technique can also
be used to diagnosemetastasis because antigens are expressed
on malignant cells’ membrane and cytoplasm surface, which
can be immunohistochemically detected using antibodies
that are specific to these antigens [51]. The antibodies com-
monly used are anti-S100, HMB-45, and MART-1 e NK1/C3.

Conventional Therapeutic Strategies against Melanoma.
Chemoprevention can be used to avoid MEL development;
chemoprevention was originally proposed by Sporn et al.
(1976) [52] and refers to using synthetic or natural agents
to reverse, suppress, or prevent molecular and histological
premalignant lesions that occur with invasive cancer
progression [20]. Reactive oxygen species play a role in MEL
progression because they lead to uncontrolled overregulation
and compartmentalization of melanosomes [53], a diet rich
in antioxidants, particularly carotenoids and vitamins C and
E, which can be used for chemoprevention [54, 55].

The conventional treatment for primary MEL is surgical;
the lesion is removed, and the tissue is analyzed to determine
the MEL stage, which depend on the lesion thickness and
location (epidermis or dermis). The lesion is removed with
a certain safety margin; however, where lesion excision is
inappropriate, such as for MELs in the nasopharyngeal,
sinonasal, and oral regions, radiotherapy is a way to eliminate
the lesion. For patients who present risk of metastasis,
the above indicated laboratory tests are also used, such as
radiography of the thorax [11, 56, 57].

The conventional MEL chemotherapy treatment is per-
formed using dacarbazine, temozolomide (dacarbazine ana-
logue), nitrosoureas (carmustine, lomustine), vinca alkaloids
(vincristine, vinblastine), platinum compounds (cisplatin,
carboplatin), and taxanes (Taxol, docetaxel), but these sin-
gle agents are not an improvement over dacarbazine [32,
58]. Immunotherapy has also been applied for MEL ther-
apy; immunotherapy employs cytokines that stimulate the
patient’s immune system to fight cancer, such as interleukin
(IL), IL-2, IL-5, IL-7, and IL-21, interferon-𝛼 (INF-𝛼), and



BioMed Research International 3

granulocyte macrophage colony–stimulating factor (GM-
CSF) [59].These cytokines have side effects, such as diarrhea,
nausea, constipation, abdominal pain, vomiting, vitiligo,
dermatitis, enterocolitis, hepatitis, toxic epidermal necrolysis,
neuropathy, and endocrinopathy [11].

The benefits of therapywith interferon alfa-2b are directly
related to the MEL stage [60]. However, high interferon alfa-
2b doses have many side effects, such as chronic fatigue,
headaches, weight loss, myelosuppression, and depression
[61].

Vemurafenib and dabrafenib are BRAF inhibitors
approved for use in MEL metastases that express
BRAFV600E and lead to dramatic shrinkage of tumors.
However, they are short-lived and resistance to treatment
eventually emerges in most melanomas. In addition,
treatment with BRAF inhibitors can lead to the induction
of second primary cancers, including squamous cell
carcinomas of the skin and new primary BRAF wild-
type melanomas; other side effects are nausea, diarrhea,
arthralgias, nonspecific skin rashes, fatigue, alopecia, and
photosensitivity [62–64].

Tremelimumab is an antibody against the cytotoxic T
lymphocyte-associated antigen 4 and is well-tolerated, but it
does not offermany benefits over conventional chemotherapy
[65].

Ipilimumab is a humanized antibody against CTLA-4, a
negative regulatory checkpoint protein that is expressed on
T cells surface after activation; the ipilimumab specifically
blocks theCTLA-4 inhibitory signal, resulting in activation of
T cells and tumour infiltrating lymphocytes; this is an indirect
mechanism that enhances the immune response mediated by
T cells. The adverse effects are colitis, dermatitis, hepatitis,
endocrinopathy, and neuritis [63, 66].

Nivolumab and pembrolizumab are anti-PD-1 antibodies;
PD-1, like CTLA-4, is expressed on the surface of activated
T cells and has a function to turn off the T-cell response to
prevent an excessive immune reaction. Anti-PD-1 antibodies
may have higher response and lower toxicity rates than
ipilimumab, as well as improved overall survival compared
to chemotherapy [63, 66, 67].

Studies demonstrated that a combination therapy with
ipilimumab and nivolumad was responsible for more adverse
effects than monotherapy; on the other hand the patient’s
median survival was higher when patients were treated
with combination therapy [66]. Therefore, several studies are
being realized to evaluate the potential survival benefits of
immunotherapy combination [68].

2. Nanotechnology-Based Drug
Delivery Systems

Many active ingredients used in MEL therapy present unde-
sirable properties and, thus, have been discarded [69]. Intro-
ducing a new active ingredient on the market takes several
years of research and involves high costs. The alternative
employed to circumvent these high costs and reintroduce
the active ingredients that were previously discarded is the
development of delivery systems that increase efficiency [70].

Drug delivery systems represent an alternative strategy to
carrier antineoplastic agents. Encapsulated drug could result
in advantages such as high stability, better bioavailability,
controlled drug release, long circulation time in blood,
selective organs or tissue distribution, a reduction of the total
dose required, and minimizing the toxic side effects [71–73].
Nanotechnology-based drug delivery systems arewidely used
to improve the effectiveness of antineoplastic agent; the most
common nanosystems are hydrogel, cyclodextrins, liquid
crystalline phase, and nanoparticulate pharmaceutical drug
delivery systems (NDDSs), as classified by Torchilin (2014),
that include liposomes; polymeric nanoparticles; polymeric
micelles; silica, gold, silver, and other metal nanoparticles;
carbon nanotubes; solid lipid nanoparticles; niosomes; and
dendrimers [74]. This review of scientifically based research
supports the application of nanotechnology-based drug
delivery system for MEL therapy.

2.1. Hydrogels. Polymeric systems can be classified by their
physical forms such as (i) linear polymer chain in solution,
(ii) physically or covalently cross-linked reversible gels, and
(iii) polymer chains grafting or adsorption on the surface
of micro- and nanoparticles [75]. A hydrogel is a network
of polymer chains that are hydrophilic and promote the
drug release through the spaces formed in the network
via dissolution or disintegration of the polymeric matrix.
Swelling in certain non-water-soluble polymer demonstrates
a high water absorption capacity in the reticular structure
(>20%) [76, 77].

An increased interest in hydrogels as a drug delivery
system has been demonstrated as a result of their easy
handling and similar physical properties to animal tissue,
which depend on the polymer employed [78–80].The release
rate depends on the hydrogel properties, initial drug concen-
tration, drug solubility, and drug-polymer interaction [81].

A wide variety of polymeric materials with different
properties have been used to form hydrogels. The required
polymer is selected based on the tissue of interest and the
specific application [80, 82]. For example, poly(vinyl alco-
hol) tetrahydroxyborate (PVA-THB) hydrogels have shown
therapeutic potential for topically treating acute and chronic
wounds due to many benefits, such as controlled release,
bioadhesion, and low toxicity [83, 84]. Moreover, chitosan-
based hydrogels have an additional advantage as a drug
delivery system because the drug can be released under
various environmental stimuli; thus, these hydrogels provide
an anchor for delivering therapeutic payloads to the site of
action [85].

Hydrogels have been employed as a drug delivery system
in MEL therapy because they may act as an intratumoral
chemotherapy depot by promoting accumulation or main-
tenance of high intracellular levels of the chemotherapeutic
agent. In recent years, a hydrogel composed of a cyclodextrin-
containing linear polymer and decorated with PEG as well as
transferrin was approved for commercial use inMEL therapy
[86].

Hydrogels are classified as stimuli-sensitive swelling-
controlled release systems because they can respond to



4 BioMed Research International

various environmental conditions, such as pH, the surround-
ing fluid ionic strength, temperature, an applied electrical
or magnetic field, or glucose level changes. These changes
promote altered network structure, swelling, mechanical
strength and permeability [83]. Thus, hydrogels may be used
to improve drug delivery [87–90]. However, little evidence
supports using hydrogels for topical treatment of MEL.

Certain studies suggest using topical hydrogels; topi-
cal ibuprofen-releasing hydrogels promote lower metastatic
spread of primary MEL through significantly lower tumor
necrosis factor (TNF)-𝛼 levels, which is the major proinflam-
matory cytokine that induces MEL cell migrations [91].

Injectable hydrogels have been widely explored for cancer
therapy [85, 92]. Interleukin-2 was given as pulse in cancer
immunotherapy because it is a potent immunomodulator
that can induce antitumor activity [93]. Recombinant human
interleukin-2 (rhIL-2) loaded, in situ gelling, and physically
cross-linked dextran hydrogels slowly release rhIL-2 and
maintain the rhIL-2 protein in an intact form that is both bio-
logically and therapeutically active, which greatly enhances
the clinical applicability of rhIL-2 immunotherapy [94].

Subcutaneous injection of a doxorubicin-loaded hydro-
gel composed of sugar beet pectin (SBP) associated with
biodegradable gelatin (SBP/gelatin) successfully suppressed
mouse MEL B16F1 cell tumor growth in nude mice [95].

The human MEL cell line Me665/2/21 derived from a
cutaneous metastasis was treated for 48 h with a cisplatin-
loaded hydrogel and it showed similar and, in certain cases,
higher cytotoxic activity towards the MEL cell line compared
with free cisplatin at the same concentration [79].

A novel system for incorporating paclitaxel has been
investigated to lower toxicity and improve efficacy [96].
Tumor activity upon using a paclitaxel (PTX) loaded hydro-
gel composed of a pH- and temperature-sensitive block
copolymer, the poly(𝜀-caprolactone-co-lactide)–poly(ethyl-
ene glycol)–poly(𝜀-caprolactone-co-lactide) (PCLA–PEG–
PCLA) block copolymer, was analyzed in vivo using B16F10
MEL cells. After 2 weeks of subcutaneously injecting the
B16F10 MEL cells into male mice, the tumors were allowed
to grow, and the results demonstrate that saline-treated mice
produced a tumor volume of approximately 17 cm3, while the
PTX-treated mice tumors were smaller than 7 cm3, which
demonstrates that a PTX-loaded block copolymer hydrogel
can effectively suppress tumor development [97].

2.2. Liposomes andMicelles. Recently, research has described
the importance of lipids in drug carrier systems such as
liposomes [98]. According to Fahy and coworkers (2005)
[99], lipids are hydrophobic molecules that are soluble in
organic solvents. However, certain lipids have amphiphilic
characteristics due to both hydrophilic and hydrophobic
segments [100]. These compounds can form carrier systems,
such as micelles and liposomes, because they can self-
assemble in the presence of water [101]. Micelles are formed
when amphiphilic components concentration exceeds a cer-
tain threshold concentration. The micelles’ size and shape
depend on pH, temperature, constituent geometry, and inter-
molecular interactions [98, 99, 102, 103].

Hydrophobic drug
Hydrophilic head

Hydrophobic tail

Hydrophobic core 

Figure 1: Micelle with hydrophobic compounds.

Hydrophilic core 

Hydrophilic drug
Hydrophilic head

Hydrophobic tail

Figure 2: Micelle with hydrophilic compounds.

The liposomes are microscopic spherical nanostructured
with awell-defined shape and size, which varies from 10 nm to
severalmicrometers, depending on the technique used to cre-
ate them [104].These vesicles are formed by an external phase
with double phospholipid membranes and an internal phase
formed by an aqueous medium [105]. These components
provide an amphiphilic character due to the organized double
phospholipid layer that surrounds the aqueous compartment.
Thus, they can encapsulate both hydrophobic andhydrophilic
compounds [106, 107]. Figures 1, 2, and 3 schematically show
the micelles and liposomes structures.

Liposomes have high versatility because they can bemod-
ified based on pharmacological and pharmaceutical needs.
Thus, the size, surface, lamellarity, lipid composition, volume,



BioMed Research International 5

Hydrophobic bilayer

Hydrophilic core 

Hydrophilic head

Hydrophobic tail

Hydrophobic drug
Hydrophilic drug

Figure 3: Liposome encapsulated hydrophobic and hydrophilic
compounds.

and inner aqueous medium composition can be modified in
these vesicles [108].

Liposomes can be formed with natural lipids, such as
sphingomyelins, as well as lecithins and synthetic lipids, such
as dimyristoyl, distearoyl, dipalmitoyl, and dioleoyl [109].
Currently, there are several methods to obtain liposomes.
Under certain conditions, they can undergo spontaneous
rearrangement and be derived from preformed micelles by
changing the solution or applying external energy, such as by
extrusion through filter membranes, sonication, or agitation
[98, 110, 111].

The extrusion technique forces a lipid suspension to pass
through a polycarbonate membrane with a well-defined pore
size [112]. This method can produce vesicles with a diameter
near the membrane pore size used to prepare the liposomes.
Over time, studies have shown several advantages from
this technique; for example, the average size of the vesicles
formed is reproducible due to the physical process involved
in liposome formation, residual organic solvent removal at
the end of the technique is unnecessary, and a large variety
of lipids can be used to prepare the vesicles [111, 113, 114].

For the sonication technique, liposomes are prepared
using a sonicator to mix the lipid suspension. The pressure
exerted by the sonicator stirring causes a decrease in the
larger vesicles sizes. Thus, the stirring time is decisive for
liposome size formed. The main advantage of this technique
is less time in liposome preparation [111, 115].

Liposomes have attracted the attention of the scientific
community due to their high versatility. Liposomes have
greater therapeutic efficacy than conventional pharmaceu-
tical system because they promote slow drug release at the
target site [107, 116, 117]. Furthermore, liposomes are less
toxic, nonimmunogenic, and biocompatible with organic
tissues.They can decrease systemic toxicity and improve drug
efficacy, especially for antibiotics, antifungals, and anticancer
drugs [107, 118, 119]. Thus, using liposomes as a delivery

system for chemotherapeutic agents offers great prospects for
cancer treatment [108].

Wolf et al. (2000) [120] incorporated the DNA repair
enzyme, T4 endonuclease V, into liposomes composed of
phosphatidylcholine, phosphatidylethanolamine, oleic acid,
and cholesteryl hemisuccinate (2 : 2 : 1 : 5 molar ratio) and
applied it to human patients with a previous history of skin
cancer after ultraviolet exposure.These liposomeswere devel-
oped by encapsulating a purified recombinant T4 endonucle-
ase V. The researchers observed that the enzyme was present
in skin cells, which, in the skin, tends to improve DNA
repair. Moreover, they reported that the T4 endonuclease
V liposome prevented ultraviolet-induced upregulation of
tumor necrosis factor-alpha and interleukin-10 mRNAs as
well as interleukin-10 protein.

Another study using T4 endonuclease V in liposomes
was conducted by Yarosh et al. (2001) [121]. They observed
30 patients with xeroderma pigmentosum in a double-blind
study. The patients were randomly assigned to use either the
T4N5 liposome or a placebo liposome lotion, daily for 1 year.
To produce the T4 endonucleaseV liposome lotion, they used
1mg/L T4 endonuclease V encapsulated in liposomes in a
1% hydrogel lotion. The placebo lotion was prepared with
the same liposomes in a 1% hydrogel solution but without
the enzyme T4 endonuclease V. Patients with xeroderma
pigmentosum have a genetic error in a DNA repair enzyme.
Thus, the incidence of skin cancer in these patients is more
frequent than in other people. The researchers noted a
decrease in the xeroderma pigmentosum incidence rate and
skin cancer in the groups treated with the T4 endonuclease
V liposome. Furthermore, no significant adverse effects were
reported.

Pierre et al. (2001) [122] proposed a topical delivery sys-
tem for 5-aminolevulinic acid (5-ALA) based on liposomes
with a similar composition to the stratum corneum to treat
skin cancer. They prepared these liposomes using a reverse
phase evaporation technique and the following components:
ceramide, cholesterol, palmitic acid, cholesteryl sulfate, and
𝛼-tocopherol. 5-ALA is used in photodynamic, which had
been shown effective in topical treatment for a variety of
skin diseases. 5-ALA liposomal delivery system targeted and
delivered 5 ALA to skin layers (viable epidermis and dermis)
compared with the aqueous solutions typically applied in a 5-
ALA-PDT clinical procedure.Thus, liposomes were a suitable
delivery system for 5 ALA.

Chen et al. (2012) [123] developed a transdermal drug
delivery system for curcumin-loaded liposomes and investi-
gated in vitro skin permeation and the antineoplastic effect
in vivo. Soybean phospholipids, hydrogenated soybean phos-
pholipids, and egg yolk phospholipids were used to obtain
the liposomes. Curcumin-loaded liposomes composed of
soybean phospholipids promoted greater in vitro drug per-
meation. Moreover, the liposomes were effective against
MEL and the liposomes composed of soybean phospholipids
showed a higher capacity to inhibit MEL cells growth.

Nobayashi et al. (2002) [124] evaluated the efficiency of
cationic multilamellar liposome-mediated gene transfer in
murine MEL cell lines and an experimental gene therapy for
subcutaneous MEL. They used B16F10, which is a murine



6 BioMed Research International

MEL cell line and cationic liposomes composed of N-(𝛼-
trimethylammonioacetyl)-didodecyl-D-glutamate chloride
(TMAG), dilauroyl phosphatidylcholine (DLPC), and
dioleoyl phosphatidylethanolamine (DOPE) at molar ratio
1 : 2 : 2. They observed that repeated exposure to liposomes
increased the transduction efficiency in murine MEL cells
and experimental subcutaneous MEL tissue; thus, the
therapy was effective for the intended purpose.

Liu and colleagues (2013) [125] developed liposomes
with quercetin to improve its delivery into human skin
and evaluate the potential anti-UVB effect. The liposomes
were composed of soybean phosphatidylcholine, cholesterol,
tween 80, and span 20. The researchers prepared liposomes
with high entrapment efficiencies and a prolonged drug
release. The group yielded good results; the quercetin lipo-
somes enhanced cell viability compared with a quercetin
solution in UVB-irradiated HaCaT cells, the reactive oxygen
species levels decreased, the edema and inflammation were
alleviated and 3.8-fold more quercetin liposomes permeated
the skin compared with quercetin solution.

2.3. Cyclodextrins. Cyclodextrins (CDs) are a family of nat-
ural cyclic oligosaccharides with 𝛼-(1-4) linked glucopyra-
nose subunits bonds [126–128]. They are produced from
starch via enzymatic conversion using cyclodextrin glycosyl
transferases (CGTases) [129, 130]. CDs have received more
attention as a pharmaceutical excipient, because they can
form drug complexes [130–132]. Furthermore, CDs are bio-
compatible and can be used to reduce in vitro and in vivo
toxicity and the delivery profile can be modulated with great
flexibility by changing the guest components [133].
𝛽-cyclodextrin (𝛽CD) is a CD that comprises seven 𝛼-

(1,4)-linked 𝛼-d-glucopyranose units and is used extensively
due to its ready availability and because its cavity size is
suitable for a varied drug range [131]. Many hydrophilic,
hydrophobic, and ionic CD derivatives have been developed
to increase CDs versatility and decrease undesirable drug
properties [134, 135].

Recent studies have demonstrated that CDs are efficient
drug delivery systems for targeting cancer cells [136–138].
A complex formed between CD and a gemini surfactant
(CDgemini) was used to carry curcumin analogue, and the
cytotoxic effect of this system in MEL cells was analyzed.
The results indicate that the drug-loaded CDgemini showed
higher caspase 3/7 activity levels compared with drugs
dissolved in DMSO, which enhance their ability to trigger
apoptosis. Further, the researchers demonstrated that this
treatment was more specific for MEL cells than for healthy
keratinocytes [139].

In general, the pH surrounding tumor tissues tends to be
more acidic (i.e.,∼ 5.5 to 6.5) than normal tissue (i.e., 7.4) [140,
141]. Thus, pH-triggered drug release systems are promising
for intracellular delivery of anticancer drugs [142]. Certain
substances exhibit pH-sensitive host-guest interactions with
cyclodextrin and may be used as pH-triggered drug release
systems [143]. He and coworkers (2013) [144] synthesized a
pH-responsive material through acetonating 𝛼-CD for PTX
delivery. Results from in vitro drug release studies show a
more rapid release profile in pH 5.0 buffer comparison with

physiological conditions (pH 7.4). Moreover, this system can
be effectively internalized by tumor cells; it demonstrates
a superior cytotoxic activity and a longer incubation time
results in higher efficiency. In addition, treatment with PTX
loaded pH-sensitive 𝛼-CD inhibited tumor growth even at
the lower PTX dose (1.1mg/kg) [144].

Polypseudorotaxanes are inclusion complexes formed
between cyclodextrins and linear macromolecules such as
polymers [145]. Doxorubicin (DOX) loaded polypseudoro-
taxanes were developed by Chang and colleagues (2013)
[146] and in vitro antitumor studies including cellular
uptake and inhibition efficiency were analyzed for B16 MEL
cells. The results indicate that doxorubicin-loaded polypseu-
dorotaxanes inhibited MEL cells proliferation. The loaded
doxorubicin showed slower endocytosis than doxorubicin
hydrochloride, perhaps due to the larger system size. The
cellular uptake of loaded doxorubicin was greater upon
increasing the incubation time. Polypseudorotaxanes may be
a promising carrier for DOX as antitumor MEL therapy.

4-Hydroxynonenal (4-HNE) is the end product of lipid
peroxidation, which has been broadly used to inducer oxida-
tive stress, and it produces a cytotoxic effect in cancer
cells [147, 148]. The 4-HNE inclusion complex with the
derivative 𝛽CD (PACM-𝛽CD) was developed by Pizzimenti
and coworkers (2013) to enhance 4-HNE stability [149]. The
results demonstrate that the inclusion complex HNE/PACM-
𝛽CD was stable and significantly reduced more viable cells
among the several cell lines tested, including human MEL
A375 cells, than untreated control cells and cells treated with
10 𝜇M free HNE.

Disrupting the lipid rafts’ integrity, which are plasma
membrane microdomains rich in cholesterol, may mod-
ify tumorigenic processes by altering the functionality of
CD44, which is a cell surface receptor involved in cell
migration and tumor metastasis [150, 151]. Murai and col-
leagues (2011) [152] showed that cholesterol reduction might
be effective for preventing and treating malignant tumors
progressions. Methyl-𝛽-cyclodextrin (M𝛽CD) forms soluble
inclusion complexes with cholesterol and depletes cholesterol
in plasma membranes [153]. A study conducted by Onodera
et al. (2013) [154] investigated the potential of M𝛽CD to
cause apoptotic cell-death in a highly pigmented human
MEL cell line. The results demonstrate that M𝛽CD induced
apoptosis through cholesterol depletion in lipid rafts, which
activated caspase-3/7 and promoted cancer cell apoptosis.
Thus, M𝛽CD provides a potential strategy for treating MEL
via lipid rafts modulation.

Mazzaglia and coworkers (2013) [155] developed an
amphiphilic cyclodextrin (ACD) system for incorporating
porphyrin derivatives to improve their water solubility and
their selectivity towards MEL cells. The complexes formed
showed higher cytotoxic activity in MEL cells than the free
porphyrin derivative in water; thus, apoptotic cell death was
observed at lower concentrations, and both cell proliferation
and changes in cellular morphology were inhibited.

Mistletoe extract is often used in complementary cancer
therapy [156]; it has been shown to stimulate cytokine
production, modify intracellular protein synthesis, induce
cell necrosis, and inhibit tumor colonization [157]. Strüh



BioMed Research International 7

et al. (2013) [158] solubilized mistletoe triterpenoids with
cyclodextrins and observed lower tumor growth, tumor
necrosis, apoptotic cells, and prolonged survival in mice.
These results indicate that solubilized mistletoe triterpenoids
enhanced the antitumor effect of mistletoe extract.

Betulin (BET) is found in Betula sp. and has been
used to treat skin diseases due to its therapeutic prop-
erties, including antitumor activity [159, 160]. Complexes
formed between BET and a novel CD derivative, octakis-[6-
deoxy-6-(2-sulfanyl ethanesulfonate)]-𝛾-CD (GCDG) were
developed, and in vitro and in vivo experimental animal
model experiments were conducted to verify antineoplasic
activity in system.The results showed that BET complexation
with CD improved BET solubility, which was an important
property for enhancing BET antitumor activity. Moreover,
BET promoted a lower MEL size, which was attributed to its
antiangiogenic effect [160].

Interleukin-2 (IL-2) promotes immune recognition of
MEL, while sparing normal cells [161]. However, secretion
of certain immunosuppressive factors, such as TGF-𝛽 (trans-
forming growth factor-𝛽), can decrease ability of the immune
system to identify the tumor as being composed of foreign
cells [162]. A system composed of methacrylate-f-CD to
solubilize the TGF-𝛽 inhibitor and liposomes loaded with a
biodegradable crosslinking polymer and IL-2 cytokine was
developed to sustain cytokines release to the tumor microen-
vironment and induces antitumor immune responses in a
B16/B6mouse.The results show that the TGF-𝛽 inhibitor and
IL-2 reduced tumor growth. Furthermore, the natural killer
cells’ activity increased [163].

Cancer photodynamic therapy (PDT) combines a pho-
tosensitizer or photosensitizing drug with a specific type of
light source to treat cancers [164]. A nontoxic carrier was
prepared using 2-hydroxypropyl-cyclodextrins (hpCDs) and
metallocomplex meso-tetrakis(4-sulfonatophenyl)porphyrin
(ZnTPPS4) as the photosensitizer. The results demonstrate
that low irradiation doses do not promote a substantial
damaging effect on MEL cells, whereas higher irradiation
doses induce cell death. Cell apoptosis or tissue necrosis
depends on the radiation intensity. ZnTPPS4 complexation
was an efficient sensitizer in human MEL cells [165].

2.4. Liquid Crystalline Phases. Pharmaceutical companies
have shown an interest in developing nanostructured sys-
tems, such as liquid crystals, which have advantages that are
mainly related to controlled drug release, and protect the
active ingredients from thermal degradation or photobleach-
ing [166, 167].

Liquid crystalline systems can compartmentalize drugs
in the inner phase droplets, which have different physico-
chemical properties than the dispersing medium, and induce
changes in the biological properties of the incorporated
substances [168, 169].

Lehmann described an intermediate state in the thermal
transformation from solid to liquid, which became known as
liquid crystals (CLs) [170–172].

Liquid crystals are classified as lyotropic and ther-
motropic. When these systems are formed through adding
solvents, they are lyotropic; thermotropic formation is

Figure 4: Polarized light microscopy of the lamellar phase
(anisotropic system).

Figure 5: Polarized light microscopy of the hexagonal phase
(anisotropic system).

temperature-dependent. As the surfactant concentration
changes occur, different liquid-crystalline forms can be gen-
erated, such as lamellar, hexagonal (hexasomes), and cubic
(cubosomes) forms.The lamellar phase is formed by parallel,
planar layers of surfactant bilayers separated by a solvent
layer, which form a one-dimensional network. Beginning
in the hexagonal phase, aggregates are formed through an
arrangement of long cylinders that form two-dimensional
structures. In the cubic phase systems, the molecules are
arranged in a three-dimensional system that consists of two
corresponding water channel networks surrounded by lipid
bilayers or surfactant [169].

Polarized lightmicroscopy is an important tool to identify
and classify liquid crystalline materials. Photomicrographs
are used to demonstrate the observed textures, typically using
polarized light [173]. Under polarized light plane, the sample
is anisotropic if it can divert the plane of incident light
that is isotropic and does not deflect light. The lamellar
and hexagonal mesophases are anisotropic, while the cubic
mesophase is isotropic [169, 174].

Figures 4 and 5 showmicroscopy systems in lamellar and
hexagonal phases, respectively.

Liquid crystals have increasingly been used as delivery
systems; Bitan-Cherbakovsky and colleagues (2013) [175]
evaluated the release of gallic acid in cancer treatments.
Liquid crystalline systems were studied as a dermal delivery
system with ascorbyl palmitate to prevent skin aging [176].



8 BioMed Research International

Cubosomes present potential utility as a drug delivery
system in skin cancer therapy, such as for MEL, due to
their bioadhesion properties and enhancer penetration [177].
Bei and coworkers (2010) formulated dacarbazine-loaded
cubosomes composed of glycerol monooleate RYLO MG
19 (GMO), poloxamer 407 (F127), phosphate buffer saline
(PBS), and DTIC (5-(3, 3-dimethyl-1-triazeno) imidazole-
4-carboxamide) and characterized their physicochemical
properties. Currently, dacarbazine is a first-line chemother-
apy agent against MEL. Due to the material’s bioadhesion
properties, it presents a potential for use in MEL therapy
[178].

5-FC phytanyl (5-FCPhy) is an amphiphile prodrug,
carried in a lyotropic liquid crystalline system, and its in vivo
efficacy as a chemotherapy agent against breast cancer has
been investigated. The results show that the 5-FCPhy-loaded
lyotropic liquid crystalline system reduced tumor size in a
dose-dependentmanner; the smallest average tumor volumes
were observed with highest 5-FCPhy doses.Thus, a 5-FCPhy-
loaded lyotropic liquid crystalline system may be used as
a controlled drug delivery system for chemotherapeutic
treatments such as for MEL [179].

von Eckardstein et al. (2005) developed an intracavitary
carrier system composed of cubosomes that encapsuled
carboplatin and paclitaxel; the release kinetics, the antitumor
activity against glioma, and the prolonged survival were
analyzed. The results show a significantly smaller tumor in
animals treated with paclitaxel/carboplastin compared with
the control group although survival did not differ among the
groups studied. Both the drugs carried in the crystalline cubic
phases presented cytotoxic activity in tumor cells, which
indicates that they play an important role in cancer therapy
[180]. The same researchers clinically observed 12 patients
with a recurrent glioblastoma multiforme, who received
an intracavitary application of paclitaxel and carboplatin
cubosomes in different doses. The results indicate that this
system is feasible and safe [181].

Many studies show the advantages of liquid crystals as
a drug delivery system. However, most studies conducted
using liquid crystals as a chemotherapy drug delivery sys-
tems remain at an early development stage. Several studies
have been executed to characterize certain systems without
efficacy trials [178, 182–186]. However, more studies are
necessary to better understand the role of liquid crystals as
a drug delivery system in MEL therapy.

2.5. Nanoparticles. The Food and Drug Administration
(FDA) defines a nanoparticle as any material with a dimen-
sional range of approximately 1 to 100 nm or end products
with a dimension up to 1𝜇m that exhibit properties or biolog-
ical phenomena (chemical, physical, and biological effects)
[187–192]. Nanotechnology-based drug delivery systems have
gained scientific notoriety due to variety of applications and
many benefits; these systems may include polymeric and
lipid-based nanoparticles.

In 1996, Müller and Lucks introduced the term solid
lipid nanoparticle (SLN) to patent a manufacturing process
using high pressure homogenization [193]. SLNs are the
first generation of lipid nanoparticles (LN), which can be

constructed by only using solid lipids (i.e., lipids that are solid
at room temperature) [194].

Subsequent modifications to SLNs have been described,
which are nanostructured lipid carriers (NLC) and are the
second generation of LN [194]. Both SLN and NLC are con-
structed from lipid solid. However, they can be distinguished
by their internal structures. The internal SLN structures only
have solid lipids and NLCs are constructed using a blend
of solid and liquid lipids, which produces imperfections in
the crystal lattice [195, 196], as shown in Figure 6. These
imperfections have also been observed for SLNs because
SLNs that contain multiple solid lipid components with
distinct structural featuresmay improve the drug entrapment
efficiency [195, 197, 198].

In addition to LN, polymeric nanoparticles (PN) may be
constructed from organic polymers or inorganic materials,
such as silica [199]. Polymers or lipids form solid NP nuclei,
which promotes more stable systems, sustained drug release,
and a uniform particle size distribution [200].

PN can be referred to as nanocapsules or nanospheres
depending on their composition [166], as shown in Figure 7.
The presence of oil promotes a vesicular structure in
nanocapsules that forms reservoir-based drug delivery sys-
tems [201, 202], while nanospheres form matrix organized
polymeric chains in the absence of oil [203, 204].

Drugs are entrapped in PN throughmixing the drug and
polymer solution. Drug compounds are physically entrapped
in the nanoparticle through polymer self-assembly [200].
PNs using different encapsulation mechanisms, such as
dissolving it, disperse it or chemically adsorbs it in the
constituents of the polymer matrix [166, 205, 206].

Both types of nanoparticles (lipid and polymeric
nanoparticles) can be used as a drug delivery system with
antitumor properties in MEL therapy.

Identifying tumor microenvironment properties is criti-
cally important for accumulating the most nanoparticles at
the site of action, which decreased drug toxicity and adverse
effects. Pathological systems’ metabolism, cell morphology,
and microenvironment have peculiar characteristics [207].
Through knowing these characteristics, specific biomarkers
(antibodies, aptamers, peptides, and small molecules) can be
identified, andmolecules can be attached to the nanoparticles
surface for successful targeted drug delivery to the site of
action [208, 209].

Nonspecific interactions may appear in addition to spe-
cific biomarkers such as van der Waals bonds and elec-
trostatic and steric affinities that can be used to predict
the propensity for nanoparticle adhesion and uptake [210].
Thus, the particular nanoparticles structural components
(lipids, surfactants, and polymers: Table 1) may improve drug
targeting to the tumor tissue [198, 211], prevent opsonization
and a consequent decrease in nanoparticles degradation
by the immune system [212–214], improve the interactions
between the surface nanoparticles and tumor cell membrane
[215–217], alter the normal function of organelles, and induce
apoptosis, which would increase tissue-specific cytotoxicity
in cancer cells [197, 218, 219].



BioMed Research International 9

Drug dissolved or dispersed
Solid lipid

(a)

Drug dissolved or dispersed

Blend of liquid and solid lipid or
blend of solid lipids

(b)

Figure 6: The image shows an SLN-organized lipid matrix composed of only solid lipids (a) and imperfections in the crystal lattice (b) on
NLC or SLN that are composed of multiple solid lipid components with distinct structural features that are distorted upon forming a perfect
crystal.

Drug dispersed
Polymeric structure

(a)

Drug entrapment

Inner core

Polymeric structure

(b)

Figure 7: Polymeric nanoparticles schematics: nanospheres (a) and nanocapsules (b).

2.5.1. Benefits of Using Nanoparticles for MEL-Targeted Drug
Delivery. Recent studies have shown improved SLN uptake
and accumulation in tumor tissue [220], due to physiological
tumor tissue characteristics, such as abnormalities and a
dysfunctional tumor vasculature, which allow SLN in the
range 30–100 nm to easily permeate the tumor. Moreover,
higher SLN concentrations are maintained in the tumor for
long periods of time due to low venous return and lymphatic
drainage [221–223].

Xu and coworkers (2009) development a docetaxel-
loaded SLN composed by egg phosphatidylcholine,
dioleoylphosphatidylethanolamine, trimyristin, and
lactobionic acid that showed 2.4-fold greater accumulation
in tumors compared with the nonencapsulated drug 6
hours after intravenous administration. Galactosylation
of the nanoparticle surfaces enhanced the cellular uptake
of docetaxel and promoted passive targeting of the drug
into the tumor cell, which reduced systemic toxicity [224].



10 BioMed Research International

Table 1: Particular nanocarrier structural components for improving drug targeting to the tumor tissue.

Components for successful
targeted drug delivery in
antitumor

Benefits in anticancer therapy References

Active targeting

Cholesterol

Cancer cells take up 100-fold more low density lipoprotein (LDL) than normal
tissue due to upregulated LDL receptors in cancer cells for membrane synthesis
during cell division associated with malignant transformation processes. Thus, LDL
has been proposed as a drug carrier for anticancer agents.

[208, 239–245]

Polyunsaturated fatty acids
(𝛼-linolenic acid; linoleic acid;
arachidonic acid;
eicosapentaenoic acid; and
docosahexaenoic acid).

They can be attached to the tumor cell membrane more easily, which results in
disruption and fluidity of the cell membranes. Tumor progression is reduced by
modulating p53, p16, and p27 expression and cell cycle regulation, as well as by
inducing cell death by apoptosis and necrosis.

[246–248]

Hyaluronic acid

Hyaluronic acid is an extracellular matrix compound that specifically binds CD44,
which is an extracellular membrane protein that regulates various cellular
responses. CD44 is overexpressed in cancer cells, while normal cells underexpress
this protein. Thus, CD44 is a good candidate biomarker for cancer cells.

[249–251]

Folic acid

Folate is important for producing and maintaining new cells because it can
participate in nucleotide synthesis. Folates receptors are highly overexpressed in
cancer cells. In addition, only the malignant cells, not normal cells, transport
folate-conjugates; thus, the folate-drug conjugation can improve tumor-targeted
drug delivery.

[248, 252–254]

Passive targeting
Polysaccharides; polyacrylamide;
polyvinyl alcohol;
polyvinylpyrrolidone; PEG;
PEG-containing copolymers
(poloxamers; poloxamines;
polysorbates; and PEG
copolymer).

They prevent the opsonin binding to the nanoparticle surfaces and, consequently,
recognition as well as phagocytosis of the nanoparticles by the mononuclear
phagocytic system, which enhances the blood circulation time.

[212, 255–258]

Cationic surfactants
The positive charge of a cationic surfactant interacts through electrostatics with the
negatively charged phospholipids that are preferentially expressed on the cancer cell
surface.

[252, 259–261]

Higher cationic nanoparticle uptake in HeLa cells compared
with anionic nanoparticles was observed [225], which
demonstrates that nanoparticle uptake is influenced by the
nanoparticle surface molecules [226, 227].

Guo et al. (2010) investigated the antitumor effects of
resveratrol (RES) bovine serum albumin nanoparticles. The
results showed that the concentration of RES was greatly
increased in the target tissue when the RES-loaded nanopar-
ticle was injected. High levels of RES were observed in
bloodstream for long periods of time after the RES sus-
pension was administered (nonencapsulated RES), which
illustrated incomplete RES distribution.Moreover, the results
show that RES-loaded nanoparticles promoted greater tumor
growth inhibition [228]. Teskač and Kristl (2010) showed
that where NLC (Compritol 888ATO and Phospolipon 80H
as the oil phase and Lutrol as the steric stabilizer) was
used to incorporate RES, it crossed the cell membrane,
was delivered throughout the cytosol, and was located
in the perinuclear region without inducing cytotoxicity.
They found that RES solubility, stability, and intracellular
release were also enhanced in the RES-loaded nanoparticle
[229].

The drug release profile was modulated using drug-
loaded LN. A recent study demonstrates that the camp-
tothecin release rate can be modified by changing lipid
nanoparticle inner phases. The SLN composed of precirol as
the solid lipid showed the most sustained release (45% of
the total drug was released after 30 hours) while the NLC
composed of precirol as the solid lipid and squalene as the
liquid lipid showed more rapid release (65% of the total drug
was released after 30 hours). Drug mobility decreased when
solid or crystalline substances were incorporated into the
nanoparticles, which decreased the drug levels released as a
function of time. Greater inhibition of MEL cell proliferation
was observed when the cells were treated with nanoparticles,
which may be because MEL cells exhibit excellent uptake
endocytosis [230].

Docetaxel-loaded NLC (DTX-NLC) composed of stearic
acid, glyceryl monostearate, soya lecithin, and oleic acid
showed as sustained-release drug profile (77% of the total
drug after 24 hours), while Duopafei (docetaxel injec-
tion provided by Qilu Pharmaceutical Co., Ltd. in China)
showed 100% release after 24 hours. In addition, DTX-NLC
showed greater cytotoxicity against MEL cells compared with



BioMed Research International 11

Duopafei through enhanced apoptosis. Moreover, DTX-NLC
showed low cytotoxicity for healthy cells because the drug is
only released after endocytosis by a target cell [231].

Camptothecin was encapsulated into NLC, which was
composed of cetyl palmitate, coconut oil, and Myverol
associated with a quantum dot (metallic compounds at
the core of the semiconductor NLC) as oil phase and
Pluronic 68 solution as water phase. Camptothecin-loaded
NLC presented superior cytotoxicity against MEL cells and
the greater cell uptake compared with other carriers. Cellular
endocytosis was essential for cell viability and the quantum
dots showed a minimal capacity to influence proliferation. In
addition, camptothecin accumulation in the MEL increased
by approximately 6.4-fold following administration of the
camptothecin-loaded nanoparticle. In vivo real-time mon-
itoring showed that a camptothecin-loaded nanoparticle
associated with a quantum dot was strongly localized at
tumors with a persistent signal for 24 hours.The drug-loaded
NLC was directed using quantum dot to efficiently transmit
sustained tumor bioimaging, in addition to promoting drug
release. This system offers the potential for diagnosing or
monitoring evolution of the tumor through bioimaging and
for drug delivery through nanocarrier [232].

Multiple synthetic and natural biodegradable polymers
may be used in antitumor drug delivery systems, such as
polyesters (e.g., polylactic acid, PLA), polyamino acids (e.g.,
polyaspartic acid), and polyoxypropylenes (e.g., poloxamers)
[233].

Interleukin-2 was delivered by a polymeric nanoparticle
composed of a lowmolecular weight polyethylenimine linked
by 𝛽-cyclodextrin and conjugated with folate; this molecule
was analyzed as a potential MEL antitumor therapy. High
levels of tumor suppression were observed after peritumoral
injection of the polymeric nanoparticle with interleukin-2,
which prolonged survival inmice; thus, it is a promising gene-
based therapeutic strategy for MEL. The antitumor effect
can be attributed mainly to activation, proliferation, and
infiltration of effector T cells and NK cells into the tumor;
the therapeutic efficiency was dose-dependent and presented
low cytotoxicity [234].

Polymeric nanoparticle using polylactic-co-glycolic acid
(PLGA) as a polymer to incorporate coumarin increased the
cellular uptake rate 2-fold versus nonencapsulated coumarin.
In addition, molecular signals for mRNA expression were
used to demonstrate that the coumarin-loaded nanoparticle
downregulated cyclin-D1, proliferating cell nuclear antigen
(PCNA), survivin, and Stat-3, and it upregulated p53 and
caspase-3, promoting enhanced apoptosis ofMEL tumor cells
compared with nonencapsulated coumarin [235].

As a drug delivery system for apigenin, PLGA-PN pro-
motes faster mobility and site-specific activity in MEL in
addition to efficiently preserving apigenin photodegradation.
The results also showed increased free radical accumulation
and antioxidant enzymes depletion inside tumor cells, which
exacerbated DNA damage and results in apoptosis through
mitochondrial dysfunction [236].

A polymer-based delivery vehicle for cisplatin composed
of chitosan and carboxymethylcellulose showed enhanced
cytotoxicity (approximately 10-fold greater) in MEL tumor

cells comparedwith nonencapsulated cisplatin. Further, rapid
intracellular drug release was observed upon endocytosis of
this system by a tumor cell, and only high-density NPs and
positively charged-surfaces were capable of releasing cisplatin
into MEL. Moreover, it decreased drug loss during blood
circulation [237].

Superparamagnetic iron oxide nanoparticles consist of a
carboxydextran shell and show increased uptake in human
mesenchymal stem cells; the nanoparticle uptake efficiency
was related to a higher density of carboxyl groups on the
nanoparticle surface [238].

3. Advantages and Disadvantages of
Nanocarrier Systems

This paper describes umpteen benefits to use nanocarriers
system to vehiculate drugs used in melanoma therapy. But,
to choose the better system type, it is also necessary to
analyze the disadvantage of each system. Table 2 describes
main advantages and disadvantages of each system.

4. Nanocarriers Application in Animal Model
or Clinical Studies

Particular nanocarrier structural components were previ-
ously described for improving drug targeting to the tumor
tissue (Table 1). Now, some animal models studies or clinical
efficacy will be portrayed.

A study realized by Shi and coworkers (2014) demon-
strated that microRNA-34a and paclitaxel-loaded functional
cationic solid lipid nanoparticles presented a synergistic
anticancer efficacy. In vivo test was conducted and this system
was much more potent inhibitor of B16F10-bearing tumor
growth and can eliminate melanoma metastasized to the
lungs cells compared to single drug-loaded SLN [273].

C57BL/6 mice were inoculated subcutaneously with B16-
F10 melanoma cells (1 × 106 in 100 𝜇L/animal) to verify the
effect of free curcumin (CUR) andCUR-loaded nanocapsules
on melanoma tumor growth. Results showed that treatments
significantly inhibited the tumor growth, 59.6% (1128.4mm3
tumor volume) after treatment with free curcumin, 61.4%
(1078.2mm3 tumor volume) after treatment with Cur-
loaded lipid nanocapsule suspensions, and 71.3% (801.4mm3
tumor volume) after treatment with Cur-loaded polymeric
nanocapsule suspensions, when compared to the control
group treated with cell culture medium only (2791.0mm3
tumor volume). Cisplatin was used as positive control and
decreased 72.4% (770.0mm3) of tumor volume [274].

Interleukin-2-loaded polymeric nanoparticle inhibited
the tumor growth and can lengthen survival in mice B16F1-
bearingmelanoma.The antitumor effect was dose-dependent
and the system demonstrated low toxicity, representing a new
strategy in drug delivery system for melanoma gene therapy
[234].

Cai et al., 2012, carried out a study to verify the influence
of tumor-targeting nanocarrier in long-circulation effects
promoted by PEGylated liposome. Results demonstrated that
paclitaxel-loaded targeted PEGylated liposomes (TL-PTX)



12 BioMed Research International

Table 2: Main advantages and disadvantages of each system.

Nanocarrier Advantages Disadvantages References

Hydrogels

Cells and fragile drugs, like peptides, proteins,
DNA, and oligonucleotides, could be protected
by aqueous environment
Good transport of nutrients to cells and
products from cells
Cell adhesion ligands easily modified them
Can be injected as a liquid at body temperature;
Usually biocompatible

Can be difficult to manufacture
Usually mechanically weak
Difficulty in encapsulating the drug
Problems connecting with the cells
Difficult to sterilize

[262]

Liposomes

They can be formed by natural or synthetic
lipids
Biodegradable
Nontoxic
Thermosensitive
Hydrophilic and lipophilic molecules can be
incorporated

High-energy sonication frequently causes
oxidation and degradation of
phospholipid
Low-energy sonication requires long
periods of sonication and can also be
destructive to phospholipid
High-pressure homogenization can
confer decreased stability
Application of volatile organic solvents

[263–266]

Micelles
Ease to prepare
Good stability
Many administration routes available

Risk of disintegration after administration [267]

Cyclodextrins
Potential solubilizing and stabilizing agents
Higher order complexes are possible
Targeting water-insoluble drugs to the oral
route

Some cyclodextrins have been shown to
be irritants
Safety concerns limited their use for
parenteral administration

[268, 269]

Liquid crystals
They are easy to prepare
Thermodynamically stable
Composed of simple chemicals
In situ phase transformations

Difficult to prepare and administer due to
high viscosity
Toxicity related to high surfactant
concentration

[267, 270, 271]

Nanoparticles

They can be prepared by different methods
Hydrophilic and lipophilic molecules can be
incorporated
They can change the surface
Increased drug stability
Possibility of controlled drug release and drug
targeting

Toxicological assessment is uncompleted
Low drug-loading capacities
It is difficult to develop an analytical
method for drug delivery
Difficult to scale up the production
Stability problems during storage

[195, 272]

lengthen the half-life of paclitaxel 2.01-fold of conventional
liposome and 3.40-fold of free paclitaxel in plasma. Higher
accumulation of TL-PTX in tumor tissue, liver, and spleen
was observed compared to conventional liposome and free
paclitaxel [275].

Doxil, the first FDA-approved nanodrug [276], is cor-
roborated to development of nanomedicine for melanoma
therapy. After that, a lot of clinical trials have been done to
verify the efficacy of nanodrugs to improve the survival and
quality life in patients with melanoma, especially with poor
prognosis [277].

In a clinical phase II study, Ugurel et al. (2004) ver-
ified that patients treated with liposomal doxorubicin as
monotherapy present survival benefit.Outpatients setting (30
patients) were included on this study. Liposomal doxorubicin
is used at 50mg/m2 i.v. on days 1, 22, 43 and 64, subsequently
at 40mg/m2 at day 85 before first staging and in 4-week
intervals thereafter. The results demonstrated that 7 patients
stay alive more than 300 days and 5 patients more than 400
days [278].

Patients with cancer stage IV melanoma participated in
an open-label, phase II study conducted byHwu and cowork-
ers (2006). Patients received a combination of 75mg/m2 per
day of temozolomide, during 6 weeks, there was a 2-week
break between cycles, and they were continuously subcuta-
neously administrated PEGylated IFN-2b at 0.5 g/kg/week.
Results showed that patient’s median survival was 12 months
and they were followed for 16 months and brain metas-
tases were developed in any patients. Researchers concluded
that a combination therapy promotes antitumor activity in
metastatic melanoma [279].

5. New Approaches and Challenges

The wide range of compositions, morphologies, and particle
sizes exhibited by drug delivery systems makes it difficult to
understand their cellular uptakemechanisms.Thus, elucidat-
ing fundamental cellular processes that cells used to import
and export select extracellular molecules may contribute to
understanding the cellular internalization mechanisms of
the systems and aid in selecting the appropriate system to



BioMed Research International 13

transport active compounds [280]. Endocytosis of particles
into cells depends not only on particle size, but also on surface
coating and cell type [226, 227, 238].

Advances in nanotechnology based drug delivery systems
have improved our understanding of the biological effects
of nanotechnology-based systems, which will undoubtedly
lead to important, clinically relevant improvements in drug
delivery. New challenges in developing nanotechnology-
based drug delivery systems for MEL antitumoral therapy
include the feasibility of upscaling processes to quickly bring
the innovative therapeutic techniques to the market and the
potential for multifunctional systems that will fulfill several
biological and therapeutic requirements, such as the system
needing to be able to target tumor cells or tumor environment
after systemic delivery. Further challenges include researches
on efficiency of targeted anticancer therapies and imaging
agents as well as international standards regarding their
toxicology and biocompatibility.

So, the possibility of nanocarriers can promote the tar-
geted cancer therapy and potentially early detection of cancer
lesions; noninvasive imaging that permits determination of
molecular signatures induces the concept of personalized
medicine [281]. But not only that, the personalized medicine
based on adjusted therapy to individual differences that can
be detected by genetic test, guiding the choose of drug
and their dosage. So, combining clinical and molecular
biomarkers in nanomedicine contribute to improvement of
the disease management [282].

6. Author’s Opinion

Drug delivery systems represent an alternative strategy to
carrier antineoplastic agents. Many advantages of drug deliv-
ery system have been described in recent studies such as
better drug stability, better bioavailability, controlled drug
release, long circulation time in blood, selective organs or
tissue distribution, a reduction of the total dose required, and
minimizing the toxic side effects.

Certain drug delivery characteristic can distinguish
their application such as hydrogel that are stimuli-sensitive
swelling-controlled release system. Liposome can encapsu-
late both hydrophobic and hydrophilic compounds. CD can
form drug complexes and are biocompatible. LC protects
the active ingredients from thermal degradation or pho-
tobleaching. SLN and NLC are maintained in the tumor
for long period of the time due to low venous return and
lymphatic drainage. PN forms reservoir-based drug delivery
systems in nanocapsules and matrix organized polymeric
chains in nanospheres. The wide range of compositions,
morphologies, and particle sizes exhibited by drug delivery
systems makes it variable mechanism for successful targeted
delivery, while making it difficult to understand their cellular
uptake mechanisms.

Another important aspect is identifying pathological
systems’ metabolism, tumor cell morphology, and microen-
vironment properties for accumulating the most drug deliv-
ery system at the site of action, at which decreased drug
toxicity and adverse effects and biomarkers (antibodies,
aptamers, peptides, and small molecules) can be identified,

and molecules can be attached to the systems surface for
successful targeted drug delivery to the site of action.

Thus, elucidating fundamental cellular processes that
cells used to import and export select extracellular molecules
may contribute to understanding the cellular internalization
mechanisms of the systems and aid in selecting the appro-
priate system to transport active compounds. Endocytosis of
particles into cells depends not only on particle size, but also
on surface coating and cell type.

Several studies on cancer have been conducted world-
wide, but peculiarities of tumor cells that distinguish them
from normal cells are not completely elucidated, which made
the delineation of targeted drug delivery for cancer therapy
difficult.

Another problem is that chemotherapy drug delivery
systems remain at an early development stage. Several studies
have been executed to physicochemically characterize certain
systems. However, the influence of systems to improve drug
biological properties is understudied.

Tumor microenvironment plays an important role in
tumorigenesis and may also influence the success rate of
melanoma therapy. The drug delivery systems need to cross
anatomical and physiological barriers of tumor microenvi-
ronment. However, many mysteries emphasize the complex-
ity of the task.

In the recent decade, one of the most studied fields is
nanotechnology-based drug delivery and various targeting
mechanisms were discovered such as cancer-specific lig-
and for receptor-mediated active targeting (i.e., folate and
hyaluronic acid); microenvironment-responsive molecules
that respond to changes in pH, temperature, light, chemicals,
and electromagnetic fields; PEGylation-induced passive tar-
geting; electrostatics interaction and molecules that prevent
the opsonization.

Drug delivery system for melanoma therapy may target
the several pathways involved in melanoma development
such as three-tiered Ras/Raf/MEKmitogen-activated protein
kinase (MAPK); PI(3)K; NF-kappaB; p16INK4a/RB and ARF
signalling pathways.

Although breakthrough in melanoma antitumor therapy
research has been observed, more studies are necessary to
better understand the role of drug delivery system in MEL
therapy.

Conflict of Interests

The authors declare that there is no conflict of interests
regarding the publication of this paper.

Acknowledgments

This work was supported by Coordenação de
Aperfeiçoamento de Pessoal de Nı́vel Superior (CAPES),
Fundação de Amparo à Pesquisa do Estado de São
Paulo (FAPESP), Conselho Nacional de Desenvolvimento
Cient́ıfico e Tecnológico (CNPq), and Programa de Apoio
ao Desenvolvimento Cient́ıfico da Faculdade de Ciências
Farmacêuticas (PADC-FCF-UNESP).



14 BioMed Research International

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