Nanotechnology-Based Approaches for Voriconazole Delivery Applied to Invasive Fungal Infections

Invasive fungal infections increase mortality and morbidity rates worldwide. The treatment of these infections is still limited due to the low bioavailability and toxicity, requiring therapeutic monitoring, especially in the most severe cases. Voriconazole is an azole widely used to treat invasive aspergillosis, other hyaline molds, many dematiaceous molds, Candida spp., including those resistant to fluconazole, and for infections caused by endemic mycoses, in addition to those that occur in the central nervous system. However, despite its broad activity, using voriconazole has limitations related to its non-linear pharmacokinetics, leading to supratherapeutic doses and increased toxicity according to individual polymorphisms during its metabolism. In this sense, nanotechnology-based drug delivery systems have successfully improved the physicochemical and biological aspects of different classes of drugs, including antifungals. In this review, we highlighted recent work that has applied nanotechnology to deliver voriconazole. These systems allowed increased permeation and deposition of voriconazole in target tissues from a controlled and sustained release in different routes of administration such as ocular, pulmonary, oral, topical, and parenteral. Thus, nanotechnology application aiming to delivery voriconazole becomes a more effective and safer therapeutic alternative in the treatment of fungal infections.


Introduction
Fungal infections are a growing threat to global public health. Most of these fungal infections are superficial, but some species can cause life-threatening illnesses. Immunocompromised patients are at higher risk for fungal infections, including organ transplant, oncology, HIV/AIDS, and, more recently, SARS-CoV-2 patients [1][2][3][4][5]. It has also occurred in immunocompetent patients as a secondary infection [6,7]. Systemic fungal infections usually originate either in the lungs, after conidia inhalation (Aspergillus, Cryptococcus), or from endogenous microbiota (Candida spp.) as a result of infected lines or leakage from the gastrointestinal tract, and may spread to many other organs. These pathogens, under certain circumstances, can spread, causing fatal infections responsible for more than one million deaths worldwide each year. Thus, the systemic spread of fungi is a critical step in developing these deadly infections. These infections are associated with high morbidity and mortality rates, especially in some hospital settings [8][9][10][11]. If appropriate therapy is delayed, systemic fungal infections are medical emergencies with high mortality rates.
The distribution of mycoses varies according to several factors, including the region and the epidemiological conditions, and may be classified as superficial [12], cutaneous [13][14][15], subcutaneous [16], and systemic [17]. Among the most common fungal Several species can cause Candida infections, but the most common is Candida albicans. Among the most common clinical manifestations of Candida sp. are invasive candidiasis, oral candidiasis, denture stomatitis, and candidemia neonatal [85][86][87]. Invasive candidiasis is one of the most aggressive forms of this disease, and affects immunocompromised patients, those who have undergone some organ transplant or are undergoing chemotherapy treatment, with a mortality rate above 70% in the latter group [88][89][90]. Candida spp. are common commensal organisms in the skin and gut microbiota, and disruptions in the cutaneous and gastrointestinal barriers (for example, owing to gastrointestinal perforation) promote invasive disease [90].
There are more than 15 species of Candida sp. capable of causing infections in humans; however, the five most common species that cause infections are C. albicans, C. glabrata, C. tropicalis, C. parapsilosis, and C. krusei, where C. albicans is responsible for 40 to 60% of cases worldwide [91][92][93][94]. Another species that has drawn attention is C. auris. Infections and outbreaks caused by this species in hospital settings have recently increased. Difficulty in its identification, multidrug resistance properties, the evolution of virulence factors, high associated mortality rates in patients, and long-term survival on surfaces in the environment make this species particularly problematic in clinical settings [95][96][97]. Associated with the increase in COVID-19 infections, a trend toward bacterial, fungal, and viral superinfection has been observed. An important co-infection agent is C. auris due to its multidrug-resistant nature and easy transmissibility. Patients with comorbidities, immunosuppressive states, and intubated and mechanically ventilated patients are more likely to contract the fungal infection; therefore, being placed in the critical group of human pathogenic fungi by the WHO [21,98,99].
Risk factors for candidemia are using the venous catheter, admission to the intensive care unit, broad-spectrum antibiotics, abdomen surgery, parenteral nutrition, neutropenia, acute renal failure, malignancy, and burns [11].
Candidemia symptoms are related to the risk factors predisposing to this type of infection. However, clinical manifestations such as blood infections related to catheter use, septic shock, and eye involvement are observed [92].
The diagnosis is based on the detection of Candida sp. by the culture method, which in turn has low sensitivity, and other tests are frequently requested, such as the detection of antigen (1,3)-β-D-glucan, tube antibodies C. albicans, and techniques based on molecular biology such as PCR, RT-PCR and MALDI-TOF MS [100][101][102].
The treatment protocols use four different classes of drugs: polyenes, triazoles, echinocandins, and flucytosine [103,104]. The first-choice medicine for treating candidiasis and invasive Candida infections is echinocandins; however, other protocols are used and consider the species causing the infection and the associated clinical conditions [105]. Monotherapy protocols are amphotericin B, in the traditional or liposomal form, anidulafungin, caspofungin, micafungin, fluconazole, and VCZ. However, the combined therapies of amphotericin B and fluconazole or amphotericin B and flucytosine also apply [106,107].

Cryptococcosis
Cryptococcosis is a systemic fungal infection caused by Cryptococcus spp., the main species are Cryptococcus neoformans, and Cryptococcus gattii can affect both immunocompetent and immunosuppressed patients in the pulmonary, extrapulmonary, and disseminated form through the Central Nervous System (CNS), causing meningoencephalitis and rarer cases of cutaneous or transplant-associated mycosis [108]. Cryptococcal meningitis has become an infection of global importance reaching 1 million new infections per year; despite both species having many characteristics in common, there are some differences regarding geographic distribution, environmental niches, host predilection, and clinical manifestations that should be emphasized [109,110].
Cryptococcosis was recently highlighted by the world health organization as the first highest-priority microorganism, surpassing even C. auris, more recently described and of worldwide concern due to its resistance. Inserting new therapies for mycoses, especially those referred to here, is essential in this context, according to the number of deaths from HIV associated cryptococcol meningitis, an estimated 181,000 cases worldwide represent 15% of all AIDS-related deaths [1,21].
The pathogenic species of Cryptococcus sp. are divided into five different serotypes subdivided into different molecular types described in Table 1. Cryptococcus spp. virulence is associated with the polysaccharide capsule, which increases pathogenicity, modulates the immune response, and protects against oxidative stress [114][115][116]. The presence of melanin in the cell wall protects against antifungal drug attacks at elevated temperatures and contributes to the modulation of the immune response [117,118]. Furthermore, the secretion of enzymes, such as urease, has played a significant role in fungal metabolic pathways [119,120].
The transmission occurs from inhaling desiccated airborne yeast cells, or possibly sexually produced basidiospores, into the lungs from the feces of pigeons in the environment or different tree species [121]. Risk factors for Cryptococcus spp. infection are HIV, rheumatic diseases, solid organ transplantation, corticosteroid or immunosuppressive therapies, chronic decompensated liver disease, diabetes mellitus, sarcoidosis, kidney problems, use of monoclonal antibodies, lymphoid diseases, and overexpression syndrome IgE and IgM [1,110,122].
Patients with pulmonary infection caused by Cryptococcus spp. are usually asymptomatic and account for a third of the number of cases, and the diagnosis is made by the presence of nodules on radiographic examination [113]. Some patients also have symptoms like pneumonia and can progress quickly to acute respiratory distress syndrome even without CSN involvement [110]. According to authors [113], the most common symptoms are fever and dry cough, but the patient may have dyspnea, chest pain or discomfort, malaise, or no symptoms.
There are some options to diagnose Cryptococcus sp, isolation of the fungus in biological fluids using culture and identification, histopathology, serological detection of the cryptococcal capsular polysaccharide antigen (CrAg), and molecular methods [122].
Treatment of patients with cryptococcosis considers risk factors or other pathologies that affect the patient concurrently being categorized by the host's immune status, organ involvement, and respiratory cryptococcosis with or without CNS problems [1]. The treatment is based on using antifungal agents or drainage of cerebrospinal fluid and surgical resection [123].
The first-choice treatment of cryptococcosis is using Amphotericin B, despite the reported hepatotoxic and nephrotoxic effects [124]. Following the use of fluconazole, mainly because it has a lower cost, some studies indicate a reduction in susceptibility to this class of antifungals [125]. In addition, Cryptococcus sp. has intrinsic resistance to echinocandins [126]. In this sense, the treatment of cryptococcosis is a challenge, and the search for new alternatives has grown in recent years [127], such as the combination of antifungal drugs [128], the use of nanotechnology, and the search for new molecular targets [129][130][131].  [132,133]. It has low solubility in water (0.098 mg/mL), log P 1.82, and pKa 2.01 and 12.7 [134]. VCZ acts by inhibiting cytochrome P450 (CYP 450)-dependent 14α-lanosterol demethylation, an important step in cell membrane ergosterol synthesis by fungi [135,136]. It is fungicidal against most molds, except Mucorales. It is active against all species of Aspergillus, including A. terreus, which is generally resistant to amphotericin B, hyaline fungi, including Fusarium spp. and members of the Scedosporium apiospermum complex, except S. prolificans [137][138][139]. VCZ is fungistatic, like all azoles, against Candida spp. Species that are inherently resistant to fluconazole, such as C. krusei, are susceptible to VCZ and some C. glabrata that are resistant to fluconazole are susceptible to VCZ, but many strains develop resistance to fluconazole also become resistant to VCZ [140,141]. Candida albicans was the predominant species, causing up to two-thirds of all cases of invasive candidiasis. However, a change to Candida spp. non-albicans, such as C. glabrata and C. krusei, with reduced susceptibility to commonly used antifungals, have been recently observed. On the other hand, Candida auris, an emerging pathogen, is highly tolerant to azoles and often resistant to several drugs [142,143]. VCZ shows good in vitro activity against other yeasts, including Cryptococcus neoformans, Trichosporon asahii, and Saccharomyces cerevisiae [144,145]. Finally, VCZ has activity against Blastomyces dermatitidis, Coccidioides spp., Histoplasma capsulatum and Paracoccidioides brasiliensis, but it is not active against Sporothrix schenckii [146][147][148].
Although it is still widely used in the treatment of invasive aspergillosis, the consequences of the non-linear pharmacokinetics of VCZ are one of its main limitations [53,149]. Since dosages have wide interpersonal variation, they can reach subtherapeutic or toxicity levels [150,151]. Antifungals of the same class, such as posaconazole and isavuconazole, have been used as an alternative in risk groups for treatment with VCZ [152,153].

Pharmacokinetics
VCZ is a class II drug with low solubility and high permeability [154][155][156]. Low water solubility makes it difficult to administer by different routes of administration [157]. VCZ is commercially available in tablets, oral suspension, and intravenous solutions [158][159][160]. An essential point of the VCZ pharmacokinetics is that they are non-linear, presenting wide inter-individual variability according to cytochrome P450 polymorphisms [161].

Absorption
VCZ is rapidly absorbed, and its plasma levels depend on body weight, age, route of administration, presence of polymorphisms, and inflammation, among other factors [162][163][164][165]. After oral administration, the maximum plasma concentration occurs within 1 to 2 h, reaching about 96% [166]. However, a wide variation of absorption (35% to 83%) appears in clinical studies associated with the high or low activity of the CYP2C19 enzyme [167][168][169].

Distribution
The estimated volume of distribution is between 2 to 4.6 L/kg, suggesting intravascular and extravascular distribution [41]. Binding to plasma proteins has not been defined yet, despite efforts to measure it through in vitro and in vivo studies. The binding proteins reported are albumin and glycogen-α-1-acid [170,171].
Factors such as pre-existing diseases and the presence of CYP450 polymorphisms are responsible for the non-linear metabolism of the VCZ, resulting in dose-dependent auto-inhibition and saturation of metabolism [161,[174][175][176]. In addition, polymorphisms can lead to unwanted interactions with other drugs [177].

Excretion
Only 2% VCZ is excreted unchanged in the urine, with an elimination half-life of approximately six hours [178,179].

Toxicity and Drug Monitoring Therapeutics
Therapeutic monitoring has been suggested in clinical practice to reduce adverse effects and increase the therapeutic efficacy and safety of VCZ [180,181]. Its unpredictable nonlinear pharmacokinetics, with wide variation in serum levels between individuals, makes therapeutic regimens difficult due to the polymorphisms associated with its primary metabolism, drug interactions, and oral bioavailability [135,182].
An investigation conducted by Zonios with 95 patients at a medical center, about 7 to 18% of patients treated with VCZ experienced adverse effects, including hallucinations, visual disturbances, photosensitivity, and hepatotoxicity [183]. Among the adverse effects in patients with prolonged therapy, hepatotoxicity and neurotoxicity were frequent [42,184,185]. In another study carried out by Epaulard, phototoxicity was present in 8% of patients treated in 61 case reports [186].
Therapeutic monitoring of VCZ levels has been investigated due to the effects of its non-linear pharmacokinetics, especially in children [187], the elderly [37], patients in Intensive Care Units (ICU), and for hematological and inflammatory diseases [188,189]. The central monitoring measures have been the evaluation of CYP2C19 polymorphisms [190] and plasma drug concentration [189,191], in addition to the evaluation of liver function and other measures [192,193]. Monitoring VCZ plasma concentrations is carried out with well-standardized Liquid Chromatography coupled with Mass Spectrometry and High-Performance Liquid Chromatography methods [194,195].
It has been observed an increase in clinical efficacy, reduction in adverse effects [196][197][198], and reduction in intra and inter-individual variability [199] with dose monitoring, being a safe alternative for using VCZ in the treatment of invasive fungal infections [200].

Nanotechnology-Based Voriconazole Delivery Systems
The interest in studying nanostructured systems has been maintained over the years due to their numerous advantages for developing of science and society. In pharmaceutical and biomedical fields, the application of nanoparticles has modified the way drug transporting in the body [201]. Hydrophilic or hydrophobic molecules, with highly variable molecular weight, labile (proteins, nucleic acids, vaccines) or not, can be carried by nanostructures and safely transported to exert their pharmacological activities [202,203]. The modification of several drug parameters has already been reported after their nanoencapsulation, such as the improvement in solubility [204], controlled release, improvement in pharmacokinetics [205], protection against degradation [206,207], and targeting to specific tissues [208]. Furthermore, these systems can be used as diagnostic [209] and therapeutic tools [210].
Nanoparticles are mainly classified according to their composition, and depending on the method used to obtain them, different supramolecular structures are obtained. A variety of materials has been used to obtain organic nanoparticles, such as polymers (natural or synthetic), lipids, surfactants, or proteins (animal or vegetable) [211]. These materials require biocompatibility, biodegradability, and specific mechanical and/or thermal properties. Based on these parameters, structures such as nanocapsules, nanospheres, liposomes, solid lipid nanoparticles, nanostructured lipid carriers, nano and microemulsions, cyclodextrins, and others can be developed [212,213]. The different supramolecular arrangements give these nanostructured systems differences in size, shape, drug loading capacity, drug release profile, biological half-life, interaction with cells and biodistribution [214]. The material's chemical composition confers different electrical charge characteristics to the surface of the particles, which interferes with its physical and biological stability and interaction with cells and tissue internalization. The surface of nanoparticles can be chemically modified by interaction and cellular targeting [215].
Considering all limitations regarding the use of VCZ, nanotechnology-based delivery systems represent an excellent approach. Furthermore, the loading of VCZ in nanoparticles, in addition to improving the physicochemical and biological aspects of the drug in the conventional routes of administration (oral and parenteral), could propose alternative routes (topical, intranasal, pulmonary, ocular, vaginal, and others), as can be represented in Figure 1. Table 2 lists some studies of nanoparticle formulations containing VCZ. We briefly present the composition of formulations, preparation methods, route of administration, and the in vitro and in vivo results.
microemulsions, cyclodextrins, and others can be developed [212,213]. The different supramolecular arrangements give these nanostructured systems differences in size, shape, drug loading capacity, drug release profile, biological half-life, interaction with cells and biodistribution [214]. The material's chemical composition confers different electrical charge characteristics to the surface of the particles, which interferes with its physical and biological stability and interaction with cells and tissue internalization. The surface of nanoparticles can be chemically modified by interaction and cellular targeting [215].
Considering all limitations regarding the use of VCZ, nanotechnology-based delivery systems represent an excellent approach. Furthermore, the loading of VCZ in nanoparticles, in addition to improving the physicochemical and biological aspects of the drug in the conventional routes of administration (oral and parenteral), could propose alternative routes (topical, intranasal, pulmonary, ocular, vaginal, and others), as can be represented in Figure 1. Table 2 lists some studies of nanoparticle formulations containing VCZ. We briefly present the composition of formulations, preparation methods, route of administration, and the in vitro and in vivo results.     There was no difference in MIC for VCZ-NLC and free VCZ. However, at low concentrations, the inhibition rate of planktonic cells of C. albicans was higher for VCZ-NLC; there was also a reduction in the biofilm cell density. There is an increase in the efficiency of the VCZ.
[235] The addition of mucoadhesive polymers increases mucoadhesion and sustained drug release of VCZ.
VCZ uptake was higher when administered in the VCZ HP-β-CD formulation, whose Cmax was 7.13 µg/g at two h post-dose, an increase 3.4× greater than HP-β-CD or VCZ in dispersion. [236] Mannitol Accumulation of VCZ in the liver and kidneys was lower in the liposomal form.
Candida sp. was more susceptible than strains of Aspergillus sp., but there was no difference between VCZ liposome and VFEND ® .
There was a difference in the pharmacokinetic parameters for liposome and VFEND ® . Liposome reduced the deputation by half; the AUC 0-24 increased 2.5× and reduced the volume of distribution of the VCZ. The percentage of VCZ release was higher for NLC (40%) than for liposome (15%) after six hours. The gel formulation showed significant accumulation only with liposomes. Liposome deposition in the follicle produces the greatest amount in the stratum corneum. While NLC has a faster and deeper release. The MIC50 to Trichophyton rubrum result was similar for both formulations. [240] Tween 80, ethanol and oleic acid. Microemulsion Topic 10 nm In the pork skin permeation test, the accumulation of VCZ in the stratum corneum and the rest of the skin was higher for the microemulsion concerning the commercial formulation. The antifungal activity was better for the microemulsion containing VCZ compared to the one without VCZ in Candida sp.
[241] In PBS medium, the drug release was limited by the dissolution rate of the drug particles, while in SLF medium, the release occurred by the diffusion/erosion mechanism.
There was variation in the concentration of VCZ in the pulmonary lobes of the animals during treatment with intravenous injection, a situation that did not occur with pulmonary administration.
In addition, there was greater retention of VCZ in lung tissue from the nanoparticles. [248]

Lipid Nanoparticles
Lipid nanocarriers are potential drug delivery systems because they allow a controlled and specific target release [250,251]. The main lipid nanocarriers are the liposomes, solid lipid nanoparticles (SLNs), and nanostructured lipid carriers (NLCs) [252,253].
Liposomes are spherical vesicles formed by one or more phospholipid bilayers, surrounded by an aqueous compartment. Hydrophilic drugs can be loaded in the aqueous core while hydrophobic drugs are trapped into the lipid bilayer [254].
SLNs consist of carriers composed of a solid lipid core surrounded by a surfactant layer. The lipid is presented as a solid at room temperature and is smelt at higher temperatures. Hydrophobic drugs are better loaded in SLNs than hydrophilic ones. This first generation of SLNs presented limitations such as low physical stability, limited encapsulation efficiency, and expulsion of the drug during storage. It occurs due to the formation of a perfectly crystallized lipid matrix after solidification of the lipid. The second generation of SLNs are the NLCs, the core of which is constituted by a solid lipid and a liquid lipid (oil), forming a disordered lipid arrangement, avoiding the drawbacks of SLNs [255].
Among the solid lipids commonly used for SLNs are those with a low melting point and solidity at body temperature, in addition to surfactants and co-surfactants [256]. The development of SLN has improved the encapsulation efficiency and stability of drugs using a mixture of liquid, solid lipids, and surfactants [257,258].
SLNs plays a significant role in improving pharmacokinetic characteristics and reducing the adverse effects of antifungals [259]. Amphotericin B was the first antifungal to be commercialized in colloidal dispersion systems, lipid complexes, or liposomes to improve drug solubility and reduce its adverse effects, [45,260].
The ability to improve drug solubility through lipid formulations has also been explored in VCZ delivery systems, to topical [160,232] and ocular administration [157]. In the study carried out by Andrade, an NLS was developed with 75% encapsulation efficiency that allowed safe release into ocular tissue in ex vivo tests [232].
In a study by Liu et al. (2023), a liposome containing voriconazole showed a greater capacity for binding to the chitin of the fungal cell wall of C. albicans. Furthermore, the in vivo study improved the delivery efficacy of voriconazole [261].
In vivo study of vaginal infections by candida albicans showed a significant reduction in infection after 48 and 72 h compared to free voriconazole, indicating sustained release of the drug [220].
In general, lipid nanoparticles containing voriconazole have been developed mainly for ocular delivery and this is justified by the compatibility and permeation capacity of these systems in this tissue, favoring a safe delivery of drugs. Studies with SLNs have shown increased permeation, mainly ocular, and allowed controlled and sustained release, increasing the therapeutic efficacy of these systems for VCZ delivery [46,243,244].

Polymeric Nanoparticles
Among the administration systems and targeted delivery of drugs and molecules, polymeric nanoparticles play a prominent role in this scenario. The main feature is the ease of interacting with pharmacokinetic parameters in administering these systems, such as absorption, bioavailability, and excretion [262,263].
The methods for obtaining nanostructured systems with polymeric nanoparticles are based on two main techniques, dispersion and polymerization [273]. However, these systems still present some challenges as drug delivery systems: the non-scalable procurement methods, the safety and cost of development, and limitations as a system, such as overcoming some biological and stability barriers [274].
In this sense, several systems have been studied in different pathophysiological conditions to overcome these challenges. Among the main diseases that have already been approved and have effective results is cancer, in systems that release doxorubicin and paclitaxel, for example [275][276][277]. Studies also use neurodegenerative [278] and cardiovascular diseases [279,280].
Despite the challenges, polymeric nanoparticles are promising for their use and development advantages [281]. They provide the encapsulated molecule with protection against degradation in some routes of administration, such as oral administration [282]. They enable the controlled release of the target-specific drug and surface modification with the incorporation of ligands [283,284].
Some nanoparticles have also been explored for VCZ delivery. Chitosan-coated PLGA nanoparticles were developed by Paul (2018) using the solvent evaporation emulsification method to improve the bioavailability and release of VCZ through the pulmonary route. The study obtained nanoparticles with 68.57% encapsulation efficiency and a sustained and slow release in vitro and in vivo, which allowed the retention of VCZ in the lung and plasma [230].
In another study by Rençber and Karavana (2018), chitosan nanoparticles were developed to carry VCZ by the emulsification method for topical administration. The nanoparticles had almost 99% encapsulation efficiency. The release was characterized by diffusion, and the method was considered promising for topical administration. The authors justify the result by the association of chitosan, which has mucoadhesive properties [229]. Das (2015) was able to increase the bioavailability of VCZ via the pulmonary route in PLGA nanoparticles developed by solvent evaporation emulsification, with an average diameter of approximately 300 nm. The system allowed increased pulmonary deposition and prolonged release, thus improving the pharmacokinetic characteristics of VCZ [248]. Another study with PLGA was developed by Sinha, Mukherjee and Pattnaik (2013), and the system allowed more significant pulmonary deposition and prolonged release of VCZ [285].
The characteristic of allowing the incorporation of different molecules into the polymeric nanoparticle allows for an increase in the interaction of the nanostructured system with the target tissue [229,283,284]. This has been explored in the literature with chitosan, which is known to have a mucoadhesive property and allows the increase in the delivery of VCZ in the target tissue [218,221,230,247].

Protein Nanocarriers
Protein nanoparticles are also described as polymeric nanoparticles, as they are classified as natural biopolymers. The main proteins used in developing these systems are proteins of animal origin, such as albumin, gelatin, collagen, and fibroin, and proteins of vegetable origin, such as zein and gliadin [286]. The procurement of nanoparticles occurs through different techniques, such as emulsification, desolvation, complexation by coacervation, electrospray deposition, and nanoprecipitation [287,288]. The protein nanoparticles can be classified, according to composition, into several types: solid spherical nanoparticles, plate-shaped nanoparticles, and nanogels [289]. Its applications involve different areas such as drug delivery, nutrient and metabolite delivery, gene delivery, tissue engineering, photodynamic therapy, growth factor delivery, vaccines, and cosmetics [288,290].
Human serum albumin nanoparticles containing VCZ were successfully developed by nab TM technology. The aim of the investigation was to optimize critical process parameters such as the homogenization pressure, the number of homogenization cycles, and the organic solvent. After determining the critical parameters, the authors obtained nanoparticles in the range of 35 to 85 nm, whose size was adequate for the proposed parenteral administration, with an encapsulation efficiency of around 69%. After 6 h, the system had already released 85% of the encapsulated VCZ, which made this system promising for further studies [231].
In addition, nanoparticles for delivering voriconazole are promising, although their application varies according to the need for the route of administration. It is possible to see that lipid formulations seem to improve delivery by the ocular and topical route and protein and polymeric formulations were explored for oral and pulmonary delivery [217,222,234,237,240]. The physicochemical characteristics of nanoparticles are important for the treatment of fungal infections, as they allow a better interaction between the fungus and the nanostructure.
In general, using nanotechnology for voriconazole encapsulation and release is a strategy to increase therapeutic activity since studies have reported increased tissue permeation with reduced toxicity, mainly due to the observed sustained and controlled release [211,216]. In addition, it is possible to observe an improvement in the solubility and bioavailability of VCZ [230,237].
Despite this, there is still a need for more comprehensive studies regarding the cytotoxicity and antifungal potential of these nanoparticles, mainly in in vivo studies.

Cubosomes
Cubosomes are reversed bicontinuous cubic phase liquid crystalline nanoparticles capable of transporting and releasing drugs at the specific target site and have a high rate of encapsulation efficiency. These characteristics are due to the high surface area between the hydrophilic and hydrophobic regions present in its structure. They are composed of a lipid and surfactant/stabilizer, which extends in three dimensions and two nanochannels interwoven, but not connected [291][292][293].
Recently, cubosomes have been studied to increase and control the bioavailability of VCZ after ocular application. VCZ suspensions showed a low ocular residence time, which leads to an increase in the frequency of applications, while VCZ-containing cubosomes demonstrated a prolonged release profile compared to the suspension. Cubosomes were also coated with chitosan, increasing transcorneal permeation and residence time at the site [224].

Cyclodextrins
Cyclodextrins (CD) are cyclic oligosaccharides composed of six or more glucose units formed from an enzymatic reaction on starch. The most important naturally occurring forms are the a-, b-, and g-CD, whose structural shape resembles a cone, where the inner cavity has hydrophobic characteristics and the outer part is hydrophilic. Among the advantages of using DC in nanotechnology is its ability to improve drug solubility and organoleptic properties, stability, and safety [294,295].
CDs were used to improve VCZ solubility, dissolution rate, and chemical stability through complexation in an aqueous solution, followed by spray-dryer drying [294]. Another study carried out the complexation of CD and VCZ to produce a thermosensitive gel based on Poloxamer 407 and Poloxamer 188 for vaginal application. VCZ formulation complexed as CD showed greater uptake of the drug by the vaginal tissue compared to the formulation that used dispersed VCZ [236].
One of the most recent studies involves CD to improve the solubility of VCZ and the production of hydrogels for ophthalmic application since there is no formulation for this purpose. The authors compared the residence time and ophthalmic safety of b-and g-CD complexes compared to the control (VFEND). The results demonstrated good corneal permeability of VCZ, longer residence time, and ophthalmic safety [296]. Therefore, the use of DC to improve the performance of VCZ has shown promise.

Clinical Trials
Currently, on the Clinical Trials website, there are 189 records of studies with VCZ, of which only two registered studies involve the evaluation of nanocarrier formulations. The PHASE 2 study under registration NCT04110860 conducted by the Minia University of Egypt evaluated the application of a nanoemulsion-based gel containing VCZ in patients with tinea versicolor once or twice a day, compared to a placebo (nanoemulsion-based gel without VCZ). The study included 30 volunteer patients aged between 10 and 60 years, both sexes, with tinea versicolor, excluding pregnant women, nursing mothers, and immunocompromised patients. Patients' clinical improvement, satisfaction, and duration of treatment were evaluated. Adverse effects were recorded, photographs of the lesions were taken before the start of treatment, during and at the end, and the clinical improvement criteria used by the physicians used a quartile rating scale. No results from this study have been published on the Clinical Trials website [297].
The PHASE 1 study under registration NCT01657201 conducted by Samyang Biopharmaceuticals Corporation of South Korea evaluated intravenous administration of VCZ-loaded-PNP 200 mg versus Vfend 200 mg in 59 healthy male patients aged 20 to 45 years to assess pharmacokinetic parameters and safety. The study was randomized, crossover, and an open intervention model; the pharmacokinetic parameters evaluated during the 24 h of the study were AUClast, Cmax, AUCinf, Tmax, T1/2, and CL [298]. Through this brief research, we can conclude that studies involving the application of nanostructured systems containing VCZ are still limited and that the studies carried out are still in the initial stages of research.

Future Prospects
It was observed that advances in the production of nanostructures containing voriconazole with a focus on topical, ocular, and pulmonary administration are significant in relation to other drug application routes. Despite the diversity of developed systems, there are still limitations regarding efficacy and safety studies. Most of the studies compiled in this work employ in vitro tests to demonstrate its performance, while in vivo tests are rarely explored. The reduced applicability of in vivo tests and the lack of use of alternative models, both used for toxicity and pharmacokinetic assessments, indicate the need for efforts in this area. Another point is the development of new drugs using nanotechnology, where clinical studies are rare, which indicates that there is still much to be done for research to proceed and benefit the population. In this sense, the application of nanotechnology is encouraged for better performance of fungal drugs, which allow a higher efficacy and safety for the treatment of different fungal infections that explore the interaction of nanoparticles with yeast and filamentous fungi, improving the prospective application of these nanoparticles.

Conclusions
The therapeutic management of opportunistic fungal infections such as aspergillosis, candidiasis, and cryptococcosis is still a challenge, which is associated with pharmacokinetic and toxicity limitations of currently available antifungals. Among them, VCZ has a wide sub-and supratherapeutic variation due to its metabolism and CYP450 polymorphisms, which reduce its therapeutic efficacy and amplify its adverse effects.
Nanocarrier systems loaded-VCZ offer numerous advantages. Among the results demonstrated, we can highlight the improvement in solubility and dissolution rate of VCZ in an aqueous medium. The release control presented different release profiles, most of them allowing a prolonged or sustained release in the presence of an initial burst effect. In addition, pharmacokinetic parameters improve the increase in mucosal adhesion and safety for topical applications. The physicochemical characteristics change according to the nanostructured system, and these data are verified according to widely disseminated techniques for characterization. Nanotechnology applied to the use of VCZ has the potential to improve parameters related to drug targeting, release, and therapeutic effects for overcoming the biological and physicochemical hurdles.