Recent advances in liposomal dry powder formulations: preparation and evaluation

Liposomal drug dry powder formulations have shown many promising features for pulmonary drug administration, such as selective localization of drug within the lung, controlled drug release, reduced local and systemic toxicities, propellant-free nature, patient compliance, high dose carrying capacity, stability and patent protection. Critical review of the recent developments will provide a balanced view on benefits of liposomal encapsulation while developing dry powder formulations and will help researchers to update themselves and focus their research in more relevant areas. In liposomal dry powder formulations (LDPF), drug encapsulated liposomes are homogenized, dispersed into the carrier and converted into dry powder form by using freeze drying, spray drying and spray freeze drying. Alternatively, LDPF can also be formulated by supercritical fluid technologies. On inhalation with a suitable inhalation device, drug encapsulated liposomes get rehydrated in the lung and release the drug over a period of time. The prepared LDPF are evaluated in vitro and in vivo for lung deposition behavior and drug disposition in the lung using a suitable inhaler device. The most commonly used liposomes are composed of lung surfactants and synthetic lipids. Delivery of anticancer agents for lung cancer, corticosteroids for asthma, immunosuppressants for avoiding lung transplantation rejection, antifungal drugs for lung fungal infections, antibiotics for local pulmonary infections and cystic fibrosis and opioid analgesics for pain management using liposome technology are a few examples. Many liposomal formulations have reached the stage of clinical trials for the treatment of pulmonary distress, cystic fibrosis, lung fungal infection and lung cancer. These formulations have given very promising results in both in vitro and in vivo studies. However, modifications to new therapies for respiratory diseases and systemic delivery will provide new challenges in conducting well-designed inhalation toxicology studies to support these products, especially for chronic diseases.     

Liposomal drug delivery systems: an update review

The discovery of liposome or lipid vesicle emerged from self forming enclosed lipid bi-layer upon hydration; liposome drug delivery systems have played a significant role in formulation of potent drug to improve therapeutics. Recently the liposome formulations are targeted to reduce toxicity and increase accumulation at the target site. There are several new methods of liposome preparation based on lipid drug interaction and liposome disposition mechanism including the inhibition of rapid clearance of liposome by controlling particle size, charge and surface hydration. Most clinical applications of liposomal drug delivery are targeting to tissue with or without expression of target recognition molecules on lipid membrane. The liposomes are characterized with respect to physical, chemical and biological parameters. The sizing of liposome is also critical parameter which helps characterize the liposome which is usually performed by sequential extrusion at relatively low pressure through polycarbonate membrane (PCM). This mode of drug delivery lends more safety and efficacy to administration of several classes of drugs like antiviral, antifungal, antimicrobial, vaccines, anti-tubercular drugs and gene therapeutics. Present applications of the liposomes are in the immunology, dermatology, vaccine adjuvant, eye disorders, brain targeting, infective disease and in tumour therapy. The new developments in this field are the specific binding properties of a drug-carrying liposome to a target cell such as a tumor cell and specific molecules in the body (antibodies, proteins, peptides etc.); stealth liposomes which are especially being used as carriers for hydrophilic (water soluble) anticancer drugs like doxorubicin, mitoxantrone; and bisphosphonate-liposome mediated depletion of macrophages. This review would be a help to the researchers working in the area of liposomal drug delivery.     

Recent advances in liposome technologies and their applications for systemic gene delivery

The recent clinical successes experienced by liposomal drug delivery systems stem from the ability to produce well-defined liposomes that can be composed of a wide variety of lipids, have high drug-trapping efficiencies and have a narrow size distribution, averaging less than 100 nm in diameter. Agents that prolong the circulation lifetime of liposomes, enhance the delivery of liposomal drugs to specific target cells, or enhance the ability of liposomes to deliver drugs intracellularly can be incorporated to further increase the therapeutic activity. The physical and chemical requirements for optimum liposome drug delivery systems will likely apply to lipid-based gene delivery systems. As a result, the development of liposomal delivery systems for systemic gene delivery should follow similar strategies.     

Liposomal drug formulations in the treatment of rheumatoid arthritis

Liposomes have been extensively investigated as drug delivery systems in the treatment of rheumatoid arthritis (RA). Low bioavailability, high clearance rates and limited selectivity of several important drugs used for RA treatment require high and frequent dosing to achieve sufficient therapeutic efficacy. However, high doses also increase the risk for systemic side effects. The use of liposomes as drug carriers may increase the therapeutic index of these antirheumatic drugs. Liposomal physicochemical properties can be changed to optimize penetration through biological barriers and retention at the site of administration, and to prevent premature degradation and toxicity to nontarget tissues. Optimal liposomal properties depend on the administration route: large-sized liposomes show good retention upon local injection, small-sized liposomes are better suited to achieve passive targeting. PEGylation reduces the uptake of the liposomes by liver and spleen, and increases the circulation time, resulting in increased localization at the inflamed site due to the enhanced permeability and retention (EPR) effect. Additionally liposomal surfaces can be modified to achieve selective delivery of the encapsulated drug to specific target cells in RA. This review gives an overview of liposomal drug formulations studied in a preclinical setting as well as in clinical practice. It covers the use of liposomes for existing antirheumatic drugs as well as for new possible treatment strategies for RA. Both local administration of liposomal depot formulations and intravenous administration of passively and actively targeted liposomes are reviewed.     

Liposomal drug delivery systems--clinical applications

Liposomes have been widely investigated since 1970 as drug carriers for improving the delivery of therapeutic agents to specific sites in the body. As a result, numerous improvements have been made, thus making this technology potentially useful for the treatment of certain diseases in the clinics. The success of liposomes as drug carriers has been reflected in a number of liposome-based formulations, which are commercially available or are currently undergoing clinical trials. The current pharmaceutical preparations of liposome-based therapeutic systems mainly result from our understanding of lipid-drug interactions and liposome disposition mechanisms. The insight gained from clinical use of liposome drug delivery systems can now be integrated to design liposomes that can be targeted on tissues, cells or intracellular compartments with or without expression of target recognition molecules on liposome membranes. This review is mainly focused on the diseases that have attracted most attention with respect to liposomal drug delivery and have therefore yielded most progress, namely cancer, antibacterial and antifungal disorders. In addition, increased gene transfer efficiencies could be obtained by appropriate selection of the gene transfer vector and mode of delivery.     

Molecular targeting of liposomal nano-particles to lymphatic system

Conventional liposomal drug delivery has been associated with obvious limitations, such as a rapid absorption by the recticulo-endothelial system in the liver and spleen, a short circulation time and a low therapeutic efficacy. Various modifications of liposomal drugs have been developed to prolong the duration of actions of the drugs at target sites, reduce its adverse effects and increase therapeutic index of drugs such as polymeric conjugation and polymeric fixation on the surface of a liposome. The lymphatic system is an important highway to spread the metastasis of most human cancers including breast, colon, and lung, ovarian and prostate. To eradicate those metastatic cancer cells from the lymphatic system, several efforts have been made to develop new and efficient lymphatic targeting drug delivery systems in order to achieve a high initial lymphatic uptake and lymph node localization. Recently, molecule targeting of liposome to lymphatic system may enhance therapeutic efficacy by improving the initial lymphatic uptake and the lymph nodal retention of liposomes such as the ligand-receptor and antibodies binding on the surface of liposome. This article aims to review the emerging liposomal drug, which is targeting the lymphatic system. The significant factors associated with targeting liposomal drugs will also be discussed in more detail in this review.     

Liposomes and niosomes as potential carriers for dermal delivery of minoxidil

The aim of this work was to formulate minoxidil loaded liposome and niosome formulations to improve skin drug delivery. Multilamellar liposomes were prepared using soy phosphatidylcholine at different purity degrees (Phospholipon 90, 90% purity, soy lecithin (SL), 75% purity) and cholesterol (Chol), whereas niosomes were made with two different commercial mixtures of alkylpolyglucoside (APG) surfactants (Oramix NS10, Oramix CG110), Chol and dicetylphosphate. Minoxidil skin penetration and permeation experiments were performed in vitro using vertical diffusion Franz cells and human skin treated with either drug vesicular systems or propylene glycol-water-ethanol solution (control). Penetration of minoxidil in epidermal and dermal layers was greater with liposomes than with niosomal formulations and the control solution. These differences might be attributed to the smaller size and the greater potential targeting to skin and skin appendages of liposomal carriers, which enhanced globally the skin drug delivery. The greatest skin accumulation was always obtained with non-dialysed vesicular formulations. No permeation of minoxidil through the whole skin thickness was detected in the present study irrespective of the existence of hair follicles. Alcohol-free liposomal formulations would constitute a promising approach for the topical delivery of minoxidil in hair loss treatment.     

New aerosol drug delivery systems for the treatment of immune-mediated pulmonary diseases

In the lung, unchecked immune responses mediated predominantly by T-lymphocytes and concurrent inflammation can lead to the development of different pathological conditions such as parenchymal disease, interstitial fibrosis, hypersensitivity pneumonitis, bronchiolitis obliterans and bronchiolar asthma. Targeted modulation of uncontrolled T-cell activation and inhibition of cytokine production within different pulmonary compartments is the challenge for the development of novel methods for immunotherapeutic intervention. Utilization of aerosol technology for pulmonary drug delivery represents new potential opportunities for therapeutic application for such immune-mediated pulmonary diseases. For targeted aerosol pulmonary drug delivery, continuous-flow jet nebulizers have several advantages over metered dose or dry powder inhalers since they are the simplest and most effective for aerosol droplet deposition into the peripheral lung tissues. At the present, the major limitations for targeted pulmonary immunosuppression through effective utilization of nebulizer technology has been the conspicuous lack of suitable formulations. The development of liposomal formulations compatible with aerosol delivery with jet nebulizers has expanded the potential for more effective utilization with an array of potent and effective immunosuppressive drugs. For pulmonary therapy, the utilization of liposomes for aerosol delivery has many potential advantages, including universal carrier suitability for most lipophilic drugs, aqueous compatibility, sustained pulmonary release or depot and intracellular delivery. Drug liposomes may also prevent local irritation in the lung, and increase potency with reduced systemic toxicity. Successful utilization of potent immunosuppressive drugs, like cyclosporin, tacrolimus (FK-506), rapamycin, mycophenolate and budesonide, in a variety of immunopathological conditions for other indications demonstrates their potential efficacy for the treatment of many different immune-mediated pulmonary diseases. The route of delivery to the pulmonary tissues can potentially limit adverse effects and markedly affect localized immunosuppressive activity in the lung. Combination of liposomal formulations with topical aerosol delivery to the central and peripheral lung tissues has expanded potential for more effective utilization with these lipophilic immunosuppressive (and antiinflammatory) drugs. Synergistic combinations can also be developed for localized and sustained delivery of therapeutic drug concentrations within the lung to provide multisite immunosuppression. Drug liposome aerosol technology represents one readily available approach for more effective therapeutic intervention in the lung using cyclosporin, FK-506, rapamycin, mycophenolate, budesonide and other lipophilic drugs.     

Nanocarriers for cancer-targeted drug delivery

Nanoparticles as drug delivery system have received much attention in recent years, especially for cancer treatment. In addition to improving the pharmacokinetics of the loaded poorly soluble hydrophobic drugs by solubilizing them in the hydrophobic compartments, nanoparticles allowed cancer specific drug delivery by inherent passive targeting phenomena and adopted active targeting strategies. For this reason, nanoparticles-drug formulations are capable of enhancing the safety, pharmacokinetic profiles and bioavailability of the administered drugs leading to improved therapeutic efficacy compared to conventional therapy. The focus of this review is to provide an overview of various nanoparticle formulations in both research and clinical applications with a focus on various chemotherapeutic drug delivery systems for the treatment of cancer. The use of various nanoparticles, including liposomes, polymeric nanoparticles, dendrimers, magnetic and other inorganic nanoparticles for targeted drug delivery in cancer is detailed.     

Stavudine-loaded mannosylated liposomes: in-vitro anti-HIV-I activity, tissue distribution and pharmacokinetics

Cells of the mononuclear phagocyte system (MPS) are important hosts for human immunodeficiency virus (HIV). Lectin receptors, which act as molecular targets for sugar molecules, are found on the surface of these cells of the MPS. Stavudine-loaded mannosylated liposomal formulations were developed for targeting to HIV-infected cells. The mannose-binding protein concanavalin A was employed as model system for the determination of in-vitro ligand-binding capacity. Antiretroviral activity was determined using MT-2 cell line. Haematological changes, tissue distribution and pharmacokinetic studies of free, liposomal and mannosylated liposomal drug were performed following a bolus intravenous injection in Sprague-Dawley rats. The entrapment efficiency of mannosylated liposomes was found to be 47.2 +/- 1.57%. Protein-carbohydrate interaction has been utilized for the effective delivery of mannosylated formulations. Cellular drug uptake was maximal when mannosylated liposomes were used. MT2 cells treated continuously with uncoated liposomal formulation had p24 levels 8-12 times lower than the level of free drug solution. Further, the mannosylated liposomes have shown p24 levels that were 14-20 and 1.4-2.3 times lower than the level of free drug and uncoated liposomal formulation treatment, respectively. Similar results were observed when infected MT2 cells were treated overnight. Stavudine, either given plain or incorporated in liposomes, led to development of anaemia and leucocytopenia while mannosylated liposomes overcame these drawbacks. These systems maintained a significant level of stavudine in the liver, spleen and lungs up to 12 h and had greater systemic clearance as compared with free drug or the uncoated liposomal formulation. Mannosylated liposomes have shown potential for the site-specific and ligand-directed delivery systems with desired therapeutics and better pharmacological activity.     

An update on liposomes in drug delivery: a patent review (2014-2018)

ABSTRACT

Introduction: Pharmacotherapy is limited by the inefficient drug targeting of non-healthy cells/tissues. In this pharmacological landscape, liposomes are contributing to the impulse given by Nanotechnology to optimize drug therapy.

Areas covered: The analysis of the state-of-the-art in liposomal formulations for drug delivery purposes have underlined that lately published patents (since 2014) are exploring alternative compositions and ways to optimize the stability and drug loading content/release profile. These improvements are complemented by improved long-circulating structures and further liposome functionalizations, which have definitively opened the road for the (co-)delivery of therapeutics to the site of action. Liposomes are also contributing to new drug delivery approaches involving the generation of extracellular vesicles by targeted cells, while opening new ways to combine disease diagnosis and therapy (theranosis).

Expert opinion: Patent publications on liposomal formulations have expanded new ways in drug delivery. New lipid compositions and strategies to optimize stability and drug vehiculization capabilities have settle solid pillars in liposome fabrication. Despite, their architecture has been satisfactorily adapted for combining passive and active drug targeting concepts, new inputs of liposomes into the disease arena should answer for: a simple/scalable/cost-effective formulation; a safe/stable/controllable formulation meeting quality control regulations; and, a confirmed therapeutic efficiency in clinical investigations.     

Pulmonary pharmacokinetics of cyclosporin A liposomes

The development of liposomal formulations for aerosol delivery with jet nebulizers has expanded the possibilities for effective utilization of aerosol based therapies in the treatment of pulmonary diseases. The property of sustained release or depot effect of liposomes has been studied using different tracer molecules to monitor absorption and clearance of liposomes from the lung. With liposomal drug formulations, few studies have simultaneously monitored phamacokinetics of both the phospholipid carrier and the therapeutic agent. We have developed a cyclosporin A (CsA)-dilauroylphosphatidylcholine (DLPC) liposomal formulation for aerosol delivery to the lung. Recent studies of CsA-liposomes have reported that CsA displays a unique property of rapid bilayer membrane exchange with dissociation between CsA and its liposome carrier in vivo following intravenous delivery. The purpose of this study was to determine the pharmacokinetics of both CsA (determined by HPLC) and liposomal carrier (labeled with ^99mtechnetium (^99mTc)) to study potential dissociation after delivery to normal mouse lungs. Furthermore, the effects of pulmonary inflammation on the clearance of CsA-DLPC liposomes were compared with ^99mTc labeled human serum albumin (HSA). Results indicate that ^99mTc-DLPC liposome carrier is retained up to 16.9 times longer than the CsA half-life in normal lung and 7.5 times longer in inflamed lungs. Similar values were obtained for ^99mTc-labeled albumin (14.8 times for normal CsA half life (6.8 times in inflamed lungs)). These pharmacokinetic results help to delineate the most effective therapeutic regimens for pulmonary CsA-liposome aerosol delivery.     

Targeted thermosensitive liposomes: an attractive novel approach for increased drug delivery to solid tumors

Introduction: Currently available chemotherapy is hampered by a lack in tumor specificity and resulting toxicity. Small and long-circulating liposomes can preferentially deliver chemotherapeutic drugs to tumors upon extravasation from tumor vasculature. Although clinically used liposomal formulations demonstrated significant reduction in toxicity, enhancement of therapeutic activity has not fully met expectations.

Areas covered: Low drug bioavailability from liposomal formulations and limited tumor accumulation remain major challenges to further improve therapeutic activity of liposomal chemotherapy. The aim of this review is to highlight strategies addressing these challenges. A first strategy uses hyperthermia and thermosensitive liposomes to improve tumor accumulation and trigger liposomal drug bioavailability. Image-guidance can aid online monitoring of heat and drug delivery and further personalize the treatment. A second strategy involves tumor-specific targeting to enhance drug delivery specificity and drug internalization. In addition, we review the potential of combinations of the two in one targeted thermosensitive-triggered drug delivery system.

Expert opinion: Heat-triggered drug delivery using thermosensitive liposomes as well as the use of tumor vasculature or tumor cell-targeted liposomes are both promising strategies to improve liposomal chemotherapy. Preclinical evidence has been encouraging and both strategies are currently undergoing clinical evaluation. A combination of both strategies rendering targeted thermosensitive liposomes (TTSL) may appear as a new and attractive approach promoting tumor drug delivery.     

Liposomes and liposome-like nanoparticles: From anti-fungal infection to the COVID-19 pandemic treatment

The liposome is the first nanomedicine transformed into the market and applied to human patients. Since then, such phospholipid bilayer vesicles have undergone technological advancements in delivering small molecular-weight compounds and biological drugs.Numerous investigations about liposome uses were conducted in different treatment fields,including anti-tumor, anti-fungal, anti-bacterial, and clinical analgesia, owing to liposome’s ability to reduce drug cytotoxicity and improve the therapeut...     

Liposomes as drug carrier systems. Preparation, classification and therapeutic advantages of liposomes]

Bangham et al. (1965) created first the concept of the liposome as a microparticulate lipoidal vesicle separated from its aqueous environment by one or more lipid bilayers. Later Gregoriadis and Ryman (1972) suggested to use liposomes as drug carrier systems. Nowadays liposomes are under extensive investigation for improving the delivery of therapeutic agents, enzymes, vaccines and genetic materials. Liposomes offer an excellent opportunity to selective targeting of drugs which is expected to optimize the pharmacokinetical parameters, the pharmacological effect and to reduce the toxicity of the encapsulated drugs. To understand the system it is important to know the basic properties of these lipoidal vesicles. Our aim was to focus on the lipid composition and the method of liposome preparation what determine the liposomal membrane fluidity, permeability, vesicle size, charge density, steric hindrance and stability of the liposomes as principle factors those influence the fate of liposomes, their interactions with the blood components and other tissues after systemic administration or local use.     

Drug delivery systems. 1. site-specific drug delivery using liposomes as carriers

Drug delivery systems, offering controlled delivery of biologically active agents, are rapidly gaining importance in pharmaceutical research and development. To achieve controlled drug delivery, i.e., the administration of drugs so that optimal amount reaches the target site to cure or control the disease state, increasingly sophisticated systems containing different carriers have been developed. Macromolecules represent one of the carriers involved, and they have taken on a significantly prominent role in various modes of administration of therapeutic agents. Among macromolecules, for example, synthetic copolymers, polysaccharides, liposomes, polyanions and antibodies, as drug carriers, liposomes have proved most effective for diseases affecting the reticuloendothelial system and blood cells in particular. Liposomes, which are vesicles consisting of one or more concentrically ordered assemblies of phospholipids bilayers, range in size from a nanometer to several micrometers. Phospholipids such as egg phosphatidylcholine, phosphatidylserine, synthetic dipalmitoyl-DL-alpha-phosphatidylcholine or phosphatidylinositol, have been used in conjunction with cholesterol and positively or negatively charged amphiphiles such as stearylamine or phosphatidic acid. Alteration of surface charge has been shown to enhance drug incorporation and also influence drug release. Because of the multifold characteristics as drug carriers, liposomes have been investigated extensively as carriers of anticancer agents for the past several years. Liposomal entrapments include a variety of pharmacologically active compounds such as antimalarial, antiviral, anti-inflammatory and anti-fungal agents as well as antibiotics, prostaglandins, steroids and bronchodilators to name a few. The liposomal entrapment has been shown to have considerable effect on the pharmacokinetics and tissue distribution of administered drugs. Despite the potential value of liposomes as unique carriers, the major obstacles are the first order targeting of a systemically given liposomes, physical stability and manufacture of the liposomal products and these problems still remain to be overcome. Drug delivery systems evolving in the 1980s have become increasingly dependent on fundamental cell-biology and receptor-mediated endocytotic mechanisms. Drug delivery systems during the 1990s may take advantage of the specificity of receptor-mediated uptake mechanisms as well as polymer chemistry and cell-biology in order to introduce more precise and efficient target-specific delivery systems that are based especially on the liposome technology.     

Liposomes in ultrasonic drug and gene delivery

Liposome-based drug and gene delivery systems have potential for significant roles in a variety of therapeutic applications. Recently, liposomes have been used to entrap gas and drugs for ultrasound-controlled drug release and ultrasound-enhanced drug delivery. Echogenic liposomes have been produced by different preparation methods, including lyophilization, pressurization, and biotin-avidin binding. Presently, significant in vivo applications of liposomal ultrasound-based drug and gene delivery are being made in cardiac disease, stroke and tumor therapy. Translation of these vehicles into the clinic will require a better understanding of improved physical properties to avoid rapid clearance, as well as of possible side effects, including those of the ultrasound. The aim of this review is to provide orientation for new researchers in the area of ultrasound-enhanced liposome drug and gene delivery.     

Liposomal encapsulated anti-cancer drugs

Among several drug delivery systems, liposomal encapsulated anti-cancer agents represent an advanced and versatile technology. Several formulations of liposomal anthracyclines are approved, e.g. for the treatment of metastatic breast cancer (pegylated and non-pegylated liposomal doxorubicin) or AIDS-related Kaposi's sarcoma (pegylated liposomal doxorubicin and liposomal daunorubicin). Meanwhile, virtually all anti-cancer drugs have been encapsulated in liposomes using different technologies. This review will summarize preclinical and clinical data of approved and exemplary emerging liposomal anti-cancer agents.     

Rheological characterization and permeation behavior of poloxamer 407-based systems containing 5-aminolevulinic acid for potential application in photodynamic therapy

Although anti-retroviral therapy is the most efficient disease management strategy for HIV-AIDS, its applications are limited by several factors including the low bioavailability and first pass metabolism of the drugs. Nanocarriers such as liposomes have been developed to circumvent some of these problems. We report here preparation of novel liposome formulations for efficient delivery of anti-retroviral drugs to mammalian cells in culture. The liposomes were prepared and surface was modified using poly (ethylene glycol). Encapsulation efficiency of the anti-retroviral drug saquinavir was found to be approximately 33% and also exhibited sustained release of the drug. Although PEGylated liposomes were more stable in protein-supplemented media, had better colloidal stability and exhibited lesser sonochemical stability due to lower cavitation threshold. The cell viability studies using Jurkat T-cells revealed that the PEGylated liposomes loaded with saquinavir were less cytotoxic as compared to the non-PEGylated liposomes or free drug confirming the potential of the liposomes as a sustained drug-release system. The drug delivery potential of the liposomes loaded with Alexa flour 647 was evaluated using Jurkat T-cells and flow cytometry showing uptake upto 74%. Collectively, our data demonstrate efficient targeting of mammalian cells using novel liposome formulations with insignificant levels of cytotoxicity.     

Organic nanocarriers for cancer drug delivery

A major focus in translational cancer research is the study of nanocarriers as novel delivery systems for chemotherapeutics. Organic vesicular nanocarriers, such as liposomes and micelles, have the advantage of low toxicity and the versatility to carry diverse drugs and conjugate to targeting agents. This offers the potential for combining treatment and diagnosis (theranostics). Successful incorporation into these nanoformulations has been demonstrated for classical chemotherapeutic drugs that are mostly hydrophobic, small interfering RNA, biological therapeutics and specific nanoparticles, such as superparamagnetic nanoparticles. Liposomes and micelles appear to take advantage of the enhanced permeability and retention (EPR) effect in solid tumours to increase accumulation at the target site (passive targeting). This translates to the clinic, where liposomal drug formulations are reported to exhibit higher efficacy and less side effects. Multidrug formulations and combinations with other treatments, for example, radiation or radiofrequency ablation, to trigger drug release from the nanocarrier at the target site, are mostly at the pre-clinical stage. More complex formulations that incorporate treatment agents together with targeting (active targeting) and imaging molecules have also been investigated in in vivo models with encouraging results.