Molecular imaging, drug delivery, and photodynamic and photothermal therapies using nanomaterials promise to have a major impact on diagnosis and therapy [5,30]. Several products are already available on the market, primarily in the United States and Asia, with an emphasis on the field of oncology, and the results of preclinical and clinical studies using nanostructured carrier systems, nanomaterials as contrast agents, and magnetic nanoparticles have shown improved diagnosis and treatment in terms of drug delivery, image quality, and electromagnetic functions [31–34].

The development of novel nanomaterials for use in medicine has proved viable in clinical and preclinical proof-of-concept experiments. Dendrimers have been applied in diagnostics and therapeutics for the development of a new cardiac test. This test can identify any damage to the heart muscle and significantly decrease the waiting time for blood test results [35,36]. Clinical trials with magnetic and metallic nanoparticles have yielded excellent results in the treatment of cancer via application of hyperthermia, in which the tumor is subjected to an electromagnetic field. For this application, nanoparticles are conjugated to specific antibodies or proteins to mark cancerous cells or tissues [37]. External laser irradiation in these tissues increases the temperature of the tissue to approximately 55°C and destroys tumor cells [38]. The application of nanomedicine is already a reality, and examples of this application are presented later.

Many studies have focused on the encapsulation of new iron oxide nanoparticles with increasingly specific physicochemical characteristics to mark human erythrocytes by internalization of these nanoparticles through the membrane pores of these cells, without affecting cell viability [42]. The effectiveness of magnetic nanoparticles in the identification tumor metastasis is superior to that of other noninvasive methods [43,44]. Magnetic nanoparticles are versatile in the diagnostic imaging of inflammatory processes, including multiple arterioscleroses and rheumatoid arthritis in macrophages [41,45]. Magnetic nanoparticles and fluorophores are combined in preoperative treatments to diagnose glioma based on the response of these nanoparticles to fluorescence and magnetic resonance [46].

A new type of magnetic nanoparticle used in diagnostic imaging is radioluminescent core–shell nanoparticles. Their significant advantage is that a single nanoparticle allows for responses in terms of magnetic resonance, fluorescence, and radioluminescence. That is, they have magnetic and optical properties. Furthermore, magnetic luminescent nanoparticles can be used for controlled drug delivery [47]. Superparamagnetic nanoparticles with a size of 5–10 nm are used as intracellular markers for infectious or viral diseases. These nanoparticles are functionalized with antibodies, peptide ligands, or enzymes to provide greater specificity in the target cells [3,48,49]. Researchers intend to use core–shell magnetic nanoparticles coated with gold as marker probes coupled with drug delivery using an HIV antibody. These nanoparticles would trace and treat the viruses that were not eliminated with the available anti-AIDS cocktails, thereby promoting a revolution in the treatment of AIDS [49]. There are numerous applications of magnetic nanoparticles, either magnetite or core–shell, to mark tumors or lesions in tissues for improved diagnostic imaging.

Current advancements in drug delivery include the development of carrier systems, in which the active ingredient is released only to the target area, for example, cancerous tissues [5]. The challenge today is to develop timed-release formulations in which the active ingredient is released over a period in preset doses [50]. For the drug to reach the target site efficiently, the delivery system should avoid the defense mechanisms of the body so that the system only releases the active ingredient to the target site [54]. The greatest challenge will be to prevent physiological mechanisms from limiting the activity of delivery systems, to block drug dispersion, and to prevent immune defense responses from causing physical or biochemical changes in these systems [55].

Drug delivery systems were initially developed using polymers and later using lipid vesicles. The interest in protecting vitamins from oxidation using polymers in the 1980s marked the beginning of the development of encapsulation systems. Soon afterward, polymers such as poly(lactic-co-glycolic acid) (PLGA) and polylactic acid (PLA) were used in drug delivery systems owing to their biodegradability and biocompatibility [56]. However, their disadvantages were associated with their low solubility and permeability. To address these disadvantages, the development of drug carriers using polypeptide vesicles increased rapidly [51]. However, disadvantages were also associated with the use of peptide vesicles, including low bioavailability due to low stability and high degradation in biological systems, low permeability in biological membranes, and short half-life in the circulatory system. Studies in nanomedicine indicate that several nanoparticles could penetrate epithelial tissue. Therefore, nanoparticles have received more attention than vesicular systems owing to their therapeutic potential and high stability in biological fluids [50,52]. In addition, drug delivery to specific organs began to become a reality. The combination of nanoparticles with antibodies, transferases, lectins, and avidins led to the development of delivery systems with greater specificity (Fig. 3.2).

The advantage of using nanoparticles in drug delivery systems is a result of two basic properties. The first is their small size, which allows them to penetrate into cells and small capillaries, causing the accumulation of active ingredient into the delivery systems. The second characteristic is the possibility of using biodegradable materials in the preparation of nanoparticles. The primary benefits of using nanotechnology in drug delivery are the improved delivery of the active ingredient, longer shelf life of the drugs, low cost of new formulations compared with the discovery of new molecules, and particularly the reduction of the doses indicated, which reduces the costs of the drugs.

The research on new biomarkers and drug delivery systems has focused on cardiovascular disease, cancer, and immunological diseases. Nanomaterials such as dendrimers, metal and silica nanoparticles, polymer nanoparticles, quantum dots, and carbon nanotubes (CNTs) have exceptional potential for direct delivery of drugs, including nitric oxide (NO) in patients with endocrine disorders [46]. In this context, the advantage of NO delivery systems using dendrimers includes the drug release control for time and site specificity and improved biocompatibility compared with conventional pharmacological applications. The primary advantage of NO release using dendrimer nanoparticles compared with other release systems is their increased capacity to store NO owing the pore structure of dendrimer molecules [57]. Furthermore, it is possible to change the external characteristics of dendrimer molecules, increasing their solubility or altering a specific function. Another example is the use of CNTs in the preferred delivery of acetylcholine in the brain for the treatment of diseases such as Alzheimer’s, without damaging the mitochondria or nonspecific organelles [58].

The development of drug carriers is highly dependent on the physicochemical properties of the nanoparticles but not on the active ingredient, as was once thought. Therefore, the physicochemical properties of these carriers can be optimized to target a specific tissue or to be nonspecifically absorbed by the target cells. However, to achieve this goal, nanomedicine should be used for the correct development of these carriers by determining the best techniques for encapsulation, the solvents used, the solubility of the active ingredient and solvent, and the types of polymers and stabilizers; in addition, it allows us to control other variables, including nanoparticle size, encapsulation efficiency, temperature, pH, ionic strength, and, most importantly, the residue of the polymer or nanoparticle associated with the drug in the patient’s body. The elimination of the polymer or nanoparticle from the patient’s body is a major cause for concern because of the possible toxicity of these materials [59].

One challenge in developing drug delivery systems is the use of theranostic nanomaterials, which combine diagnostics with therapeutics [60]. An ideal theranostic system will combine diagnosis and treatment in a single step using preventive monitoring before disease symptoms manifest [33,46,60]. This system includes the detection and removal of fat plaques in arteries for the prevention of cardiovascular disease. The development of drug-delivery systems with multiple active ingredients is promising because it allows for the synergistically combined delivery of several drugs to biological targets, for example, an antiangiogenic compound and a DNA intercalating agent, and this approach can increase the therapeutic efficacy.

Standard photodynamic therapy has been applied to cancerous tumors, in which a photosensitive drug is administered to the patient, accumulates near the tumor tissue, and induces cell death after light activation. The photosensitive agents used for therapy since 1990 are porphyrin compounds, and for this reason, the photosensitive agents are classified based on the presence or absence of porphyrin [64]. Although porphyrins have a rapid adsorption and low toxicity, the disadvantages include low selectivity to smaller tumors and high hydrophobicity. Therefore, the development of highly selective photosensitive agents is the primary research topic in this area.

One of the most significant problems in photodynamic therapy is the controlled delivery of the photosensitive agent. Therefore, the development of this type of treatment is directly correlated with advances in systems for controlled drug delivery. Other limitations of photodynamic therapy are the low selectivity to tumors, low solubility, and high aggregation of these agents under physiological conditions [65]. With the advancement of research in drug delivery systems in nanomedicine, new strategies have been developed to increase the solubility of hydrophobic photosensitive agents and selectivity by modifying the surface area of nanoparticles loaded with the agent. The development of highly selective photoactive nanoparticles will ensure that this therapy targets cancer cells and preserves healthy cells [66].

Photosensitive agent formulations using nanoparticles are already available. One of the most promising formulations is the encapsulation of temoporfin as liposome (Foscan) [67], which overcome the limitations of existing commercial agents by causing a more efficient destruction of tumors and reducing damage and toxicity to healthy tissue compared with commercial agents. The encapsulation of porphyrin-based photosensitive agents in dendrimers is also possible. In this case, porphyrins and phthalocyanines are stabilized externally by dendrimers, which prevents aggregation [54]. CNTs are another possibility for delivery of photosensitive agents because they can be modified to increase specificity and adsorption in the body. However, a challenge that needs to be overcome is that CNTs absorb light in the NI region, which may cause the death of healthy cells owing to excessive local heating [68,69].

To improve the selectivity and specificity of the photosensitive agent in cancer cells, a combination of nanostructures with monoclonal antibodies and antibodies against transferrin receptors that are expressed on the surface of many solid tumors can be used [70,71]. Nanostructures can also be combined with other ligands, including vitamins, glycoproteins, peptides, aptamers, DNA, and growth factors. Another approach is the combination of different treatment modalities. For example, a single type of nanoparticle can be used in photodynamic therapy and radiotherapy. In this case, luminescent nanoparticles combined with photosensitive agents can be exposed to radiation such as X-rays, which causes the nanoparticles to emit luminescence, which in turn activates the photosensitive agents [72]; this is a major advantage for in vivo applications because the nanoparticles are luminescent and therefore no external light source is required.

Nanoparticles used in upconversion usually consist of nanostructured systems doped with lanthanides, that is, rare-earth trivalent ions such as Yb³⁺, Tm³⁺, Er³⁺, and Ho³⁺, dispersed in nanostructures in a suitable dielectric medium [75,76]. These lanthanide dopants act as optically active centers and emit light when excited with the appropriate wavelength. These optical properties can be adjusted by selecting the lanthanide dopants to produce systems with different emission wavelengths, ranging from visible to NI or ultraviolet. Furthermore, the selection of the nanostructured host system, that is, the nanomaterials that will be doped with lanthanides, is essential for the production of highly efficient systems. Characteristics such as transparency in the spectral region of interest and chemical stability are critical in the selection of this system [73].

Nanoparticles of sodium yttrium fluoride doped with ytterbium and erbium (NaYF₄: Yb,Er) are efficient in converting infrared radiation into visible radiation and have shown promise in the detection of biological interactions [77]. The modification of dopants of NaYF₄ nanoparticles to Tm³⁺ and Yb³⁺ allows for the upconversion from NI to visible radiation, resulting in high penetration of these particles into biological tissues and acquisition of images with high optical contrast, which is supported by in vitro and in vivo experiments [78]. Several other host nanostructures have been explored in the development of these systems, including ZrO₂ [79] and LaF₃ [80].

Furthermore, the possibility to combine different materials into a single structure has made it possible to obtain systems with multiple and synergistic properties. For example, multifunctional nanoparticles can be manufactured by adsorption of Fe₃O₄ nanoparticles on the surface of upconverting nanostructures followed by the formation of a thin layer of gold on the surface [81]. Therefore, these materials have optical and magnetic properties and can be exploited as contrast agents in magnetic resonance imaging. Alternatively, the introduction of Gd³⁺ to these structures allows it to be used as a T1 contrast agent in magnetic resonance imaging [82]. The presence of gold nanoparticles on the surface of upconverting nanoparticles enables the modulation of these systems, increasing their versatility [83].

Therefore, these nanoparticles have many advantages for tumor imaging applications, including stable emission, high resistance to photodegradation, the ability to be detected in biological tissues using light in the NI region, and the possibility of biofunctionalization of their surface. Despite their great potential, rapid heating can overheat the tissue around the tumor, and lanthanides can increase the toxicity of these materials; therefore, new strategies for synthesis and functionalization are required to minimize these adverse effects [34,73,74].

Gold nanoparticles can be manufactured using the precipitation method, in which a reducing agent is added to a gold salt solution in the presence of a stabilizer. In the most common route of synthesis, nanoparticles are synthesized in an aqueous medium in the presence of citric acid or sodium citrate, which acts as a reducing agent and stabilizer because of its negative charge [85]. Other stabilizing molecules have been explored, including polymers. This functionalization allows us to obtain organic and inorganic hybrid nanomaterials, whose parameters, including size, shape, and surface functional groups, can be adjusted by the appropriate choice of the surface polymer [86].

Sulfur establishes a strong bond with atomic gold [87] and has been extensively explored for the preparation and stabilization of gold nanoparticles. In one method of synthesis, gold nanoparticles stabilized with sodium 3-mercaptopropionate molecules are obtained by the simultaneous addition of a citrate salt and sodium 3-mercaptopropionate to an aqueous solution of chloroauric acid (HAuCl₄) under boiling [88]. The particle size can be controlled by modifying the ratio between the concentration of stabilizer and the concentration of gold ions in solution [88].

Using only iron (II) salt, the reaction occurs by different mechanisms and the diameter of the obtained nanoparticles varies between 30 and 50 nm:

This method is quite simple, but it has limitations on the control of the shape and size of the obtained particles. Methods that use organic solvents and high temperatures have been the most efficient to date. In one such method, nanoparticles (MFe₂O₄) are obtained from metal acetylacetonate (M = Fe, Co, Mn) in 1,2-hexadecanodiol in the presence of oleic acid and olamine at high temperatures [90]. After synthesis, these particles can be washed by magnetic separation and functionalized with polymeric molecules for use in an aqueous medium.

Another prominent candidate for application in medicine is nanoparticles of Fe₃O₄@Au. The advantage of this system compared with iron oxide nanoparticles is the possibility of developing systems that combine optical and magnetic properties in a single process. Furthermore, the coating of iron oxide nanoparticles protects the Fe₃O₄ core against oxidation without drastically reducing their magnetic properties [92].

Several strategies have been developed to form such structures. One example is the reduction of gold ions in the presence of Fe₃O₄ nanoparticles, 1,2-hexadecanodiol, oleic acid, and oleylamine in phenyl ether under magnetic stirring, argon atmosphere, and a temperature of 190°C [93].

Another synthesis strategy is the initial coating of iron oxide nanoparticles with silica using tetraethylorthosilicate (TEOS) and ammonium hydroxide in an alcohol solution. These particles are then functionalized with 3-aminopropyltriethoxysilane (APTES). This system is added to a suspension of gold colloids in the presence of tetrakis(hydroxymethyl)phosphonium chloride (THPC), which interacts with amino groups from the APTES. The formation of a gold nanoparticle-THPC suspension involves the reduction of chloroauric acid (HAuCl₄) with THPC, leading to the formation of small particles (approximately 2 nm). Finally, the nanoshell is prepared via reduction of HAuCl₄ with formaldehyde in a solution containing potassium carbonate (K₂CO₃), and the magnetic nanoparticles are coated with silica prepared beforehand [94,95].

For example, gold nanoparticles conjugated to oligonucleotides are of great interest because the complementarity of DNA base pairs increases the specificity of these complexes and allows their use in biosensors for the detection of specific DNA sequences associated with abnormalities or diseases [101], intracellular gene regulation [102], and supramolecular organization of nanostructures [103]. Gold nanoparticles can be functionalized with oligonucleotides by modifying the oligonucleotides with thiol groups, which form highly stable bonds with gold [104].

Metallic and magnetic nanoparticles are biocompatible because they are made of inert material. However, nanoparticles made of inert material can produce morphological changes, loss of function, inflammation, and even irreversible cell damage. One study on the toxicity of gold nanoparticles in human sperm indicated changes in motility, morphology, and sperm fragmentation [128]. However, the morphology of primary endothelial cells from rat brains (rBMEC) did not change after 24 h of exposure to gold nanoparticles [129].

PLGA polymeric nanoparticles were not toxic to the A549 cell line, even at high concentrations [130]. Silica nanoparticles administered to A549 cells in a time- and dose-dependent manner reduced cell viability and increased the levels of oxidative stress, indicating a cytotoxic effect [131]. Silver nanoparticles at concentrations higher than 50 μg mL⁻¹ applied to fibroblast cells (NIH3T3) caused apoptosis associated with ROS production [132]. The administration of silver nanoparticles at concentrations close to 2 μg mL⁻¹ reduced the viability of HEK cells and induced the production of cytokines responsible for immune responses [133]. In contrast, silver nanoparticles at a concentration of 6.3 μg mL⁻¹ showed no evidence of causing cell death or damage to A431 human carcinoma cells [134].

In summary, toxicological studies of nanoparticles in in vitro systems are still inconsistent regarding the adverse and toxic effects of nanoparticles, indicating that more research is needed to address this issue [135]. In addition, further systematic in vitro studies are necessary to regulate the medical use of nanoparticles.

In rats, the accumulation and potential adverse effects of CNTs in several organs, including the liver, lung, and spleen, are reported [110]. Various studies have evaluated the toxicity of carbon-based nanomaterials, including multiwall CNTs, carbon nanofibers, and carbon nanoparticles, on the basis of the radius and surface chemistry of these materials. The nanotoxicity was tested in vivo using a tumor cell line culture. The results indicated that these materials are toxic in a dose-dependent manner. Furthermore, cytotoxicity was enhanced when the surface of these nanomaterials was functionalized after acid treatment [138]. Another study inoculated CNTs into primary skins and subjected this material to conjunctival irritation and sensitivity tests. The negative metagenesis results suggested that SWCNTs are not carcinogenic. In addition, the lethal dose for hamsters was 2000 mg kg⁻¹ body weight, a dose higher than will ever be used in nanomedicine [139].

Changes in gene expression were observed in pregnant mice exposed to titanium oxide (TiO₂) nanoparticles. Exposure of mouse fetuses to TiO₂ nanoparticles during the prenatal period affected the expression of genes related to the development and function of the central nervous system [140]. Considering the potential toxicity of nanoparticles and nanomaterials in general and the lack of legislation governing such effects, new analytical methods and additional systematic studies are necessary to assess the nanotoxicity of these materials. It is expected that, as the field of nanomedicine progresses, new concepts in nanotoxicity will guide the creation of laws to regulate the use of nanomaterials.