Int J Curr Pharm Res, Vol 7, Issue 4, 26-33Review Article


EVOLUTION OF LIPOSOMAL CARRIERS INTENDED TO ANTICANCER DRUG DELIVERY: AN OVERVIEW

MARILENE ESTANQUEIRO*, MARIA HELENA AMARAL, JAIME CONCEIÇÃO, JOSÉ MANUEL SOUSA LOBO

Laboratory of Pharmaceutical Technology, Department of Drug Sciences, Faculty of Pharmacy, University of Porto, Portugal
Email: msrestanqueiro@ff.up.pt

Received: 10 Aug 2015, Revised and Accepted: 21 Sep 2015

ABSTRACT

Most current anticancer agents are present lack of specificity, leading to systemic toxicity and adverse effects, and limiting the maximum dose of drug. Liposomes quickly passed from a simple scientific curiosity to “magic bullets” for the delivery of drugs. Liposomal formulations of anticancer drugs have been extensively evaluated, with notorious advances and the market introduction of some of them. In the last years, the research under liposomes has been carried out to increase the circulation time and the specificity to cancer cells. The aim of this work was to make a review about the research carried out about the application of liposomes as carriers for anticancer drugs. Liposomal formulations of anticancer drugs have been extensively evaluated. However, many other liposome based carriers were studied, like immuno liposomes, thermossensitive liposomes, dual functional liposomes and cross linked multifunctional liposomes, intended to increase drug specificity. Additionally, some special types of liposomes, like niosomes, transfersomes and ethosomes were also investigated as cytotoxic drug carriers.

Keywords: Cancer, Drug delivery, Liposomes, Liposomes derivatives, Special liposomes.

INTRODUCTION

Cancer is a term used for a heterogeneous group of malignant diseases in which abnormal cells divide without control and are able to invade other tissues. A mass of cancerous cells is called tumor and an uncontrolled growth of a tumor results in the destruction of health tissue around. The dissemination of tumor cells through blood or lymphatic stream leads to the formation of secondary tumors or metastasis in other organs or tissues. There are more than 100 different types of cancer [1]. Cancer is a multifunctional disease that can arise through the influence of environmental factors, and life style. Some knew risk factors include high-fat diet, smoking and excess alcohol intake, viral infections and immune system perturbations. All cancers begin in cells, which are the body's basic unit of life [1].

According to data of Glubocan project from International Agency for Research on Cancer (IARC), World Health Organization (WHO), there were 14.1 million new cancer cases, 8.2 million cancer deaths and 32.6 million people living with cancer (within 5 y of diagnosis) in 2012, worldwide. A percentage of 57 % of the new cancer cases, 65% of the cancer deaths and 48 % of the 5-year prevalent cancer cases occurred in the less developed regions. Cancer incidence rate is almost 25% higher in men than in women [2].

The main treatments for cancer are surgery and radiation, which are considered local treatments. Local control rates with radiation therapy alone, for patients treated with curative intent is about 50%, with some improvement seen when radiation therapy is combined with chemotherapy [3]. Chemotherapy finds its main use as an adjuvant to surgery and radiotherapy. Although the adverse effects of surgery and radiotherapy, chemotherapy is the third option in cancer management. This can be due to the fact that, although some excellent drugs are available, the efficacy of many existing chemotherapeutic drugs is limited by their inability to reach their therapeutic site of action in sufficient amounts to be efficacious [4]. In most cases only a small fraction of the administered drug reaches the site of action, whilst the rest of the drug is distributed throughout the body. Is this unavoidable distribution into healthy organs and tissues, and the depression of the immune system, that limits the dosage that can be given, and in turn, prevents these drugs from achieving the potential cures that they are clearly capable [5]. It is well recognized that when systemic chemotherapy is used in the treatment of solid tumors, it is almost impossible to achieve the therapeutic levels of drug at the tumor site without damaging healthy organs and tissues [6]. An inability to control the growth of primary (or regional) tumors, however, leads to fatality in a significant number of patients. In approximately one half of patient treated by local therapies, local control can be extremely difficult, the treatment fails, and the tumor grows back [5]. Because of the precarious blood supply and often high interstitial fluid pressure, many cancer chemotherapeutic agents are not-effectively delivered to the tumor region [6]. This situation is heightened by the need for almost 100% cell kill to obtain a cure. The tumor vessel wall represents a significant barrier for many therapeutic agents, and nonspecific delivery can lead to significant systemic toxicities and a low therapeutic index that is often seen with current cancer chemotherapeutics that use injected free drugs [7, 8].

Fig. 1: Role of key carrier properties in drug delivery (adapted from 11)

Role of carrier characteristics in drug delivery processes

Four key requirements of drug carrier design are essential to the overall function and performance of the carrier system. These requirements are: retain de drug; escape to the immune system, for example, extending the circulation time; target (passive) to the diseased site while avoiding most healthy organs; and release of the drug 9. Drug delivery vehicles introduced in the bloodstream undergo a complex journey prior to arriving at the target site. The carriers need to circulate through the vasculature and interact with the reticuloendothelial system (RES). The RES is the body’s primary mechanism of clearing and corresponds to the phagocytic cells, as macrophages [10]. Additionally, the carrier has to escape to the filtration that takes place in the spleen and in the kidney. If the carrier can overcome these cleaning mechanisms, it is able to adhere at the desired site in the vasculature or permeate through the vasculature into the desired tissue. This is followed by diffusion of the carrier through the interstitial space, attachment to the target cell membrane and endocytosis. The particle parameters that can influence these processes include size, shape, surface chemistry and mechanical flexibility [11]. These particle parameters and the processes that they influence are schematized in fig. 1.

Particle size has a significant impact on the circulation time and for intravenous application the particles need to have a size intended to not clog the smallest capillaries [12]. Size has also impact on splenic and renal clearance of particles. Particles larger than 200 nm are susceptible to elimination through the splenic filtration while particles smaller than 10 nm are cleared by kidneys filtering systems [13]. Particle size influences the extent of cellular uptake by phagocytosis and endocytosis. Particles smaller than 500 nm are usually internalized by endocytosis whereas particles larger than 500 nm are believed to be internalized by phagocytosis [14]. Laverman et al. have studied the effect of liposomes size on the removal rate, using liposomes with identical composition. These authors have observed that liposomes with greater size were removed faster from the blood stream, with a half-life of 0.2 h, comparatively with small vesicles that showed a half-life of 1.5 h [15].

Many studies have demonstrated the accumulation of nanocarriers in the abnormal tumor microenvironment through the enhanced permeability and retention (EPR) effect as an advantage of nanoparticle-based drugs [16-18]. The EPR effect results from the combination of the generally leaky microvasculature and missing or tight lymphatic capillary system [19].

Liposomes

Liposomes or phospholipid-based nanovesicles were first described in the mid-sixties, and have attracted attention because of their potential as drug delivery systems [20]. Following their discovery, liposomes were quickly developed for a range of potential uses and investigations which sought to exploit the capsular and biocompatibility properties of the lipid membrane in drug delivery applications [5,21]. These are small spherical vesicles with diameters varying from nanometers (>25 nm) to few micrometers, essentially constituted by phospholipids and other constituents from cellular membrane, like cholesterol, which when in contact with water can originate double molecular layers, which, in turn form mono or poly-compartmental structures. Fig. 2 illustrates the formation of mono or poly-compartmental vesicles.

Fig. 2: Representation of liposomes formation. When in contact with water, phospholipids form a lipid bilayer that form poly-compartmental (I) or mono-compartmental vesicles (II)

Quickly, liposomes have passed from a simple scientific curiosity to “magic bullets” intended to carry drugs and can be considered one of the most popular nanocarriers for delivering many biologically active substances [22]. Liposomes can encapsulate hydrophilic and/or lipophilic drugs, wherein the first stay in the aqueous compartment and the seconds inserted or adsorbed in the membrane. Because they are biodegradable, biocompatible and no immunogenic, they are extremely versatile in therapeutics, research and analytical applications [23-25]. Liposomal formulations of anticancer drugs have been extensively evaluated for treating cancers [18, 26, 27]. In the last two decades, notorious advances occurred with the commercialization of pharmaceutical liposomal formulations as is the example of liposomal daunorubicin (DAU) (DaunoxomeTM, NeXtar, Inc.) and doxorubicin (DOX) (DoxilTM, Sequus Pharmaceuticals), that showed more prolonged circulation times, greater release, an accumulation in the tumor tissue, increasing the drug efficacy and decreasing the side effects. The most recent examples of antineoplastic drugs encapsulated in liposomes, include MarqiboTM (Inex Pharmaceutical Corporation), a formulation containing vincristine sulfate, and DepoCytTM a formulation for controlled released of cytarabine[28]. Additionally, other liposomal formulations are in clinical trials. Table 1 presents some examples of liposomal platinum drugs under clinical trials [29].

Table 1: Liposomal platinum drugs under clinical trials [29]

Formulation Incorporated drug Approx. size Clinical phase Indication
Lipoplatin Cisplatin 110 nm Phase III NSCLC, breast cancer, gastric cancer
SPI-77 Cisplatin 110 nm Phase II Advanced NSCLC, refractory ovarian cancer
Aroplatin(L-NDDP) NDDP 1 µm Phase II Refractory colorectal cancer, malignant pleural mesothelioma
Lipoxal Oxaliplatin 250 nm Phase I Advanced gastrointestinal cancer
MBP-426 Oxaliplatin 100 nm Phase II Gastric, gastroesophageal, esophageal adenocarcinomas

NSCLC: Non-small-cell lung carcinoma; NDDP: cis-bisneodecanoato-trans-R, R-1, 2-diaminocyclohexane platinum, is a cisplatin analog.

Table 2: Susceptible volume to be encapsulated by each liposome category [30]

Diameter (nm) Encapsulated volume (µl/mg lipid)
SUV 20-100 0.5
LUV 100-800 13.7
MLV 500-5000 4.1

By definition, liposomes are spherical vesicles formed by one or various lipid bilayers concentrically arranged, trapping therein one or more aqueous compartments. Their diameter and layers number depends, primarily, from the manufacture process [21]. According their dimensions and compartments, liposomes can be classified in three categories, namely, small unilamellar vesicles (SUV), large unilamellar vesicles (LUV), and multilamellar vesicles (MLV). In table 2 it is indicated the diameter and susceptible volume to be encapsulated by each liposome category [30].

Liposomes are composed of substances with low intrinsic toxicity, and they can be formulated in a large range of sizes and chemical compositions. For clinical use, liposomes are commonly composed of neutral phospholipids and cholesterol and they have average diameters of 50 to 100 nm. The bilayer is impermeable to large molecules (such as proteins and enzymes) and has low permeability to charged molecules, including protons and other cations[20]. Because of the unique structural properties of liposomes, hydrophilic drugs can be entrapped in the aqueous interior of the liposomes and lipid soluble drugs can be incorporated into the hydrophobic core of the phospholipid bilayer. Even drugs of intermediate solubility such as DOX and DAU, can be stably associated with the liposome interior by manipulation of the internal liposomal pH or by the addition of counter-ions to form molecular complexes of the drug within the liposome interior [20].

Although phospholipids are biodegradable and non-toxic amphiphiles, problems arise in practical applications of liposomes, because of the low physical and chemical stability of aqueous suspensions of this type of vesicles [31]. During a prolonged storage, liposomes can suffer various kinds of changes over time, such as physical, chemical and biological, that determine the liposomes half-life [21, 32]. Regarding physical stability, depending on their composition and environment, liposomes can suffer aggregation, fusion, deposition, membrane rupture, or loss of content. Including a small proportion of charged lipids, the aggregation can be controlled by electrostatic repulsion. The permeability and diffusion of the encapsulated drug can be decreased by an addition of cholesterol to the formulation [33]. Chemical instability depends on the liposomes composition, namely from the hydrolysis and oxidation of lipid bilayer constituents [34].

Liposomes derivatives

As previously mentioned, in order to achieve maximum targeting, liposomes should remain in the systemic circulation for a relatively long period of time. However, formulations of liposomes used in the past were rapidly removed from the circulation by the RES [35, 36]. Based on this, several studies have been conducted in order to improve the characteristics of liposomes as carriers of antitumor drugs.

Pegylated liposomes

Many studies have focused on the use of liposomes with polyethylene glycol (PEG) attached in their surface. The presence of PEG reduces serum proteins binding (opsonins) and increases the circulation time to hours or days [37]. PEG also increases vascular permeability to liposomes, facilitating increased accumulation of drug containing liposomes in tumor tissues [38].

Doxil®or Caelyx® is the drug DOX encapsulated in PEG-liposome, approved in USA and Europe, respectively. Notably, the pegylated liposomes encapsulating DOX exhibited an improved safety profile by reducing cardiac toxicity and enhancing penetration and accumulation in solid tumors. Consequently, it has been used in the treatment of a wide range of cancers [39-41].

Indeed, such liposomal drug formulations do appear to improve accumulation of liposomes at the tumor site. However, slow and incomplete drug release could still lead to low drug bioavailability within tumor tissue, limiting, in turn, therapeutic activity [42-45]. Furthermore, a lack of controlled-release properties of encapsulated drug may lead to toxic side effects, such as palmar-plantar erythrodysesthesia that is thought to result from unwanted drug distribution to skin during prolonged circulation of liposomal DOX [46]. Efforts to design liposomes that are pH sensitive, temperature sensitive, antibody targeted, or fusogenic have all been pursued with various degrees of success [47].

Immuno liposomes

The first attempts to create the immune-conjugated liposomes were performed in the late 70s-early 80s. In vivo studies have revealed that coating liposomes with antibodies leads to enhanced uptake of immuno liposomes by the RES and the immuno targeting efficiency depends on the antibody density on the surface [48, 49]. Given a suitable antibody with high specificity and affinity for the target antigen, the critical factor is the accessibility of target cells to immuno liposomes [37]. The earlier attempts to improve of the circulation longevity of immuno liposomes were performed by simple co-mobilization of an antibody and PEG on the surface of the same liposomal composition and a significant increase of their circulation time was achieved [50-52]. Fig. 3 is an illustration of an immuno liposome, with the antibodies attached to liposome through a PEG chain.

Fig. 3: Immunoliposome with co-mobilization of an antibody and PEG on the surface (adapted from 37)

The majority of long-circulating immuno liposomes are targeted for the delivery of anticancer drugs [53]. Thus, the clinical success of DOX-loaded long circulating pegylated liposomes (Doxyl®/Caelyx®) used in the treatment of metastatic breast cancer, progressive ovarian cancer, multiple myeloma and AIDS-related Kaposi's Sarcoma, stimulated numerous experimental attempts for the improvement of their targeting properties by surface immobilization of different antibodies or their fragments against specific tumor antigens [54, 55].

As an example, the modification of pegylated DOX-liposomes with monoclonal antibodies or their Fab fragments against HER2, a member of the epidermal growth factor receptor (EGFR/ErbB) family frequently over-expressed on various cancer cells, successfully improved the tumor delivery and therapeutic efficiency of liposomal DOX in different HER2-overexpressing mouse xenograft models [56-59].

Another example is CD19, an internalizing receptor over expressed in most types of B-lymphoid malignancies, is a promising targeting antigen. Introduction of anti-CD19 monoclonal antibodies or its Fab' fragments to PEG-liposomes loaded with DOX clearly enhanced targeting and therapeutic efficacy in mice bearing a human CD19+B-lymphoma [60, 61].

Pegylated immuno liposomes modified with anti-HER2 monoclonal antibody fragments are internalized much better by cancer cells compared to non-modified liposomes, which allows higher drug dose delivery inside cancer cells for, i.e., more efficient cancer cell killing [62].

The therapeutic effects are dependent on the type of the encapsulated drug and the rate of drug release from the immuno liposomes in the targeted areas [61, 63]. The cytotoxic efficiency of immuno liposomes is also dependent on the surface density of the membrane antigen against which liposomes were targeted. It was estimated that about 4×104 antigen sites per single cell are required to exert the immuno liposomal targeting effect [64,65]. Another essential factor that determines the degree of immuno liposomal targeting is the extent of heterogeneous expression of antigens in the targeting area. It was suggested that a co-mobilization of antibodies against different antigens on a single immuno liposome will provide better and more uniform targeting of all cells within the tumor. Additionally, some cells can be killed by the “bystander effect”, i.e. the drug released by the immuno liposome attached to a cancer cell expressing a specific antigen can act over the neighboring cancer cells devoid of a similar receptor [63, 66].

Thermo sensitive liposomes

In order to produce a more controlled, rapid and complete release of drug from a lipid carrier, local intervention techniques make use of temperature change (hyperthermia), light (photodynamic therapy) or mechanical disruption (e. g. by ultrasound), to initiate the breakdown or change the phase of the membrane capsule composed of specifically engineered lipids 5,[67-70]. In the opinion of Koning et al., hyperthermia can be of great importance to achieve success in both suggested strategies [71]. In 1978, Yatvin et al. suggested the use of temperature sensitive liposomes to control and produce a burst release that can be an essential step to provide efficacious levels of drug in the tumor [72]. Hyperthermia can be applied to augment liposomal drug delivery by increasing tumor blood flow and micro vascular permeability to liposomes [8, 73].

Studies performed by Needham and collaborators goal to explore the use of hyperthermia, as a way to initiate a temperature-dependent change in the physical structure of a liposome membrane capsule that then might lead to an enhanced permeability of the encapsulating membrane and a rapid triggered release of the drug at clinically attainable temperatures 5. One of the studies performed by the cited authors consisted in developing a new thermo sensitive drug delivery system containing DOX that has been optimized for both, mild hyperthermic temperatures (39º to 40ºC) that are readily achievable in the clinic and rapid release times of drug (ten seconds). The liposomes were prepared by the lipid film hydration and extrusion method and DOX encapsulation into the liposomes was carried out using the pH gradient-driven loading protocol. This new thermo sensitive liposomal formulation (lysolecithin containing thermo sensitive liposome DOX) was composed by 1,2-dipalmitoyl-sn-glycero-3-phosphocholine, 1-palmitoyl-2-hydroxy-sn-glycero-3-phosphocholine, 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-polyethylene glycol 2000. This one differs from the traditional thermo sensitive liposomes regarding the composition. The authors of this study have observed that the release of encapsulated DOX from the new thermo sensitive liposomes was extremely fast (some seconds) upon heating the liposomes to 42ºC, compared with the other liposome formulations, including traditional thermo sensitive liposomes. Studies carried out in a human squamous cell carcinoma tumor xenograft model showed that the tumor growth, during 60 d after treatment, is lower when heating at 42ºC and this is more evident, using thermo sensitive liposomes than free DOX, non-thermo sensitive and traditional thermo sensitive liposomes [8, 67].

Fig. 4: Schematic mode of action of “smart” stimulus-sensitive long circulating immuno liposomes (adapted from 54)

Dual functional liposomes

The attachment of certain stimulus-sensitive moieties to the nanocarriers provides a novel strategy for the assembly of “smart” multifunctional nanocarriers, which function in response to intrinsic or externally applied stimuli in a coordinated manner to maximize the antitumor efficacy and minimize the drug side effects [74, 75]. These multi functionalities can be used to detach the long-circulating polymer (PEG) chains and release the nanocarrier contents at pathological areas, in the presence of certain intrinsic stimuli such as decreased pH, hyperthermia, altered enzyme levels or redox conditions characteristics of these zones [76].

In the case of pH-responsive systems, the pH-cleavable bond stabilize the nanocarrier in normal tissues and blood, but they disintegrate and release the drug load in areas with lowered pH, including neoplasic, ischemic and inflamed tissue, or endosomes or cell cytoplasm [54]. The long-circulating targeting and stimulus-responsive functions of nanocarriers can also be combined with certain cell penetrating proteins and peptides (CPPs), such as trans-activating transcriptional activator (TATp) for improved intracellular drug delivery [77, 78]. These CPPs effectively translocate across the plasma membrane directly into cell cytoplasm, avoiding the endocytic pathway and so prevent lysosomal drug degradation and lead to increased cellular uptake [79, 80]. Fig. 4 represents a schematic mode of action of “smart” stimulus-sensitive long circulating immuno liposomes. Briefly, the mode of action of “smart” stimulus-sensitive long circulating immuno liposome include the shielding long-chain PEG with or without targeting monoclonal antibodies attached to the liposomal surface via low pH-degradable bonds. After the accumulation in the tumor due to passive-accumulation and/or active targeting, pH dependent de-shielding of the temporarily hidden cell-penetrating function allows for carrier penetration into tumor cells [54].

Jiang et al., developed dual functional liposomes with pH-responsive CPP and active targeting hyaluronic acid (HA) for tumor target drug delivery [81]. CPPs, facilitating the cellular uptake of various cargos without causing any cellular injury, have been widely investigated in the fields of gene and drug delivery for cancer therapy [82-84]. However, CPPs with effective tumor targeting are still lacking and remain highly desirable, which present more accumulations in tumor cells but less in normal cells. In the light of this, the pH gradient between the tumor milieu and physiological environment draws more attention to designing pH-responsive CPPs for tumor-targeted drug delivery, which can be used to conjugate drugs or modify nanocarriers [81]. Unfortunately, recent studies have suggested that CPPs on the surface of liposomes and micelles are susceptible to enzymatic cleavage by enzymes present in human plasma [85]. Additionally, for intravenous injection, positively charged nanoparticles, including cationic CPP-modified liposomes, cause severe toxicity, instability and a rapid clearance from the blood compartment, thereby limiting their applications in vivo [86-88]. To address this dilemma, surface pegylation of CPP-modified nanoparticles (CPP-NPs) is regarded as a gold standard for improved safety; bioavailability and blood persistence resulted from the reduced interactions between CPP-NPs and opsonins by the hydrophilic shell of PEG [89-92].

Crosslinkedmultilamellar liposomes

A strategy to improve liposome-based anticancer drugs should involve the development of a stable liposomal formulation with improved drug release from the carrier in a controlled and sustained manner, thereby enhancing bioavailability. Based on this idea, Jooet al., have developed a new liposomal formulation involving the creation of a robust multilamellar structure of the liposome by covalently crosslinking inter-lipid bilayers. The main goal of these authors was to generate a liposomal formulation with improved bioavailability of liposomal drugs and enhanced vesicle stability. The multilamellar vesicles were formed through covalently crosslinking functionalized head groups of adjacent lipid bilayers. As a nano carrier platform for chemotherapy drug delivery applications, this study demonstrates that these crosslinked multilamellar liposomes (CMLs) can lower systemic toxicity and enhance therapeutic efficacy. Effectively, this study demonstrated that the enhanced delivery of CML-DOX to tumor cell in vitro and in vivo improved anticancer activity and led to better tumor reduction and inhibition of tumor progression, when compared with the antitumor activity of non-CML with the same lipid composition [93].

Special types of liposomes

Niosomes

Niosomes are similar, in terms of structure and physical properties, to liposomes [94]. Niosomes are non-ionic surfactant vesicles made up from single chain surfactant molecules often in combination with cholesterol. They alleviate the disadvantages associated with liposomes, like chemical instability and variable purity of phospholipids [95]. Additionally, the research interest in niosomal formulations is recently widening because surfactants are easily derivative and give a higher versatility to the vesicular structure and moreover they have lower costs than phospholipids [31]. They have longer shelf life, stability and ability to deliver drug at target site in a controlled or sustained manner which enhances bioavailability [94, 96, 97]. Nonionic surfactants used due to their ability to enhance solubility are used to increase bioavailability of poor water soluble drugs. Nonionic surfactants increase both permeability and fluidity of biological membranes and drugs show enhanced bioavailability by the transdermal route via niosomes. Furthermore, nonionic surfactants are preferred due to less irritation power which decreases in order of cationic>anionic>amphoteric>nonionic [98]. They are inhibitors of p-glycoprotein, hence increasing bioavailability of some anticancer drugs, HIV drugs and other class of drugs [99-101].

Kong et al., have developed a novel drug nanocarrier HA niosome, which combine transdermal delivery and tumor targeting. These authors have concluded that incorporating HA significantly promoted the endocytosed amount of nanocarrier by tumor cell. HA-niosome is not only efficient and secure for transdermal permeation, but it also offers a useful and promising carrier for tumor therapy through percutaneous administration [102]. Tavanoet al., have developed magneto-niosomes, in which both the magnetic material and antitumoral drug have been incorporated into the niosome aqueous compartment. Vesicles have been prepared by Tween® 60 and Pluronic® L64 surfactants and DOX was used as a model drug. Magneto-niosome formulations were stable for long periods and exhibited a controlled drug release. It has been concluded that DOX loading and release behavior of magneto-niosomes could probably promote them as effective functional materials for magnetically controlled cancer therapy [103]. Paolino and collaborators have made innovative bola-surfactant niosomes as topical delivery systems of 5-FU (5-Fluorouracil) for the treatment of skin cancer. 5-FU-loaded bola-niosomes showed an improvement of the cytotoxic effect with respect to the free drug. Bola-niosomes also provided an increase of the drug penetration of 8-and 4-folds with respect to a drug aqueous solution and to a mixture of empty bola-niosomes with a drug aqueous solution [104].

Transfersomes

Deformable liposomes (Transfersomes®) are the first generation of elastic vesicles introduced by Cevcet al. and were reported to penetrate intact skin carrying therapeutic concentrations of drugs, but only when applied under non-occluded conditions [105]. These systems consist of phospholipids and an edge activator. An edge activator is often a single chain surfactant that destabilizes lipid bilayers of the vesicles and increases deformability of the bilayers [106]. Sodium cholate, Span® 80, Tween® 80 and dipotassium glycyrrhizinate were employed as edge activators [107, 108]. Ultra-deformable liposomes have shown potential as carriers for topical drug delivery systems because they can penetrate the intact skin, improving drug delivery of various drugs, with efficiency comparable with subcutaneous administration [107-110].

Maghraby et al., have evaluated the potential use of deformable and standard liposomes as skin drug delivery systems of 5-FU. These authors have observed that ultra-deformable formulation was superior to standard liposomes in the skin delivery of 5-FU and concluded that ultra-deformable vesicles are promising agents for skin delivery of drugs [111].

Recent work has shown that bleomycin can be encapsulated in ultra-deformable liposomes and it has been suggested that this preparation may be useful for topical chemotherapy of non-melanoma skin cancer [112]. Hiruta and collaborators have incorporated beta-sitosterol 3-β-d-glucoside (Sit-G) as an absorption enhancer into ultra-deformable liposomes containing bleomycin to attenuate drug toxicity in human keratinocytes. The presence of Sit-G increased drug entrapment and improved in vitro stability. Furthermore, treatment with preparations incorporating Sit-G resulted in elevated epidermal and dermal concentrations of bleomycin. Ultra-deformable formulation containing Sit-G maintained flexibility for penetration through the skin, increased entrapment efficiency of bleomycin and stability in vitro, and significantly increased the distribution of bleomycin in epidermis and dermis compared with the formulations without Sit-G [113].

Ethosomes

Modern approaches for drug delivery via skin have resulted in the design of modified liposomes. Basically these approaches use the non-toxic and biodegradable, characteristics of phospholipids and for this reason they are able to prolong the half-life of a drug to attain a sustained-release effect. On the other hand, previous studies demonstrated that phospholipids can exhibit their enhancing effect on the skin in the presence of organic solvents such as ethanol, as in the case of ethosomes [114]. Ethosomesare non-invasive delivery carriers that enable drugs to reach the deep skin layers and/or the systemic circulation. These are soft, malleable vesicles tailored for enhanced delivery of active substances. They are composed mainly of phospholipids, (phosphatidylcholine, phosphatidylserine, phosphatidic acid), high concentration of ethanol and water [115]. Ethanol is known as an efficient permeation enhancer [116]. The high concentration of ethanol makes the ethosomes unique, as ethanol is known for its disturbance of skin lipid bilayer organization; therefore, when integrated into a vesicle membrane, it gives the vesicle the ability to penetrate the stratum corneum. Also, because of their high ethanol concentration, the lipid membrane is packed less tightly than conventional vesicles but has equivalent stability, allowing a more malleable structure and improving drug distribution ability in stratum corneum lipids [117].

Paolinhoet al., have developed paclitaxel-loaded ethosomes for potential topical treatment of squamous cell carcinoma, a malignant transformation of actinic keratosis. The paclitaxel-loaded ethosomes were proposed as topical drug delivery systems for treatment of this pathology due to their suitable physicochemical characteristics and enhanced skin permeation ability for deep dermal delivery. The obtained results showed that the proposed formulation enhanced the drug permeation through the skin and also increase its anti-proliferative activity compared to the free drug [118].

Conclusion

The research for novel drug delivery systems that could provide a controlled release, increase efficacy or reduce side effects of antitumor drugs is an important field that is at the forefront of the pharmaceutical technology. Particularly, the nanocarriers have been extensively studied in the last years, demonstrating compliance with the referred objectives. The main advantages of nanocarriers can be a real asset to carry cytotoxic drugs, because, due to lack of specificity, these drugs exhibit various harmful effects typical from chemotherapy.

Liposomes represent one of the most popular nano carriers for the delivery of anticancer drugs. From its discovery to date, liposomes have been intensively investigated in the context of its application as anticancer drug carriers. Liposomes are proven to be efficient drug delivery systems. In a general manner, this research has been carried out with the intention to increase the specificity to cancer cells. Additionally, the special types of liposomes, that is, niosomes, transfersomes and ethosomes, were also investigated as carriers to cytotoxic drugs, showing very promising results.

CONFLICT OF INTERESTS

Declared None

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