How Magnetic Composites are Effective Anticancer Therapeutics? A Comprehensive Review of the Literature

Abstract

Chemotherapy is the most prominent route in cancer therapy for prolonging the lifespan of cancer patients. However, its non-target specificity and the resulting off-target cytotoxicities have been reported. Recent in vitro and in vivo studies using magnetic nanocomposites (MNCs) for magnetothermal chemotherapy may potentially improve the therapeutic outcome by increasing the target selectivity. In this review, magnetic hyperthermia therapy and magnetic targeting using drug-loaded MNCs are revisited, focusing on magnetism, the fabrication and structures of magnetic nanoparticles, surface modifications, biocompatible coating, shape, size, and other important physicochemical properties of MNCs, along with the parameters of the hyperthermia therapy and external magnetic field. Due to the limited drug-loading capacity and low biocompatibility, the use of magnetic nanoparticles (MNPs) as drug delivery system has lost traction. In contrast, MNCs show higher biocompatibility, multifunctional physicochemical properties, high drug encapsulation, and multi-stages of controlled release for localized synergistic chemo-thermotherapy. Further, combining various forms of magnetic cores and pH-sensitive coating agents can generate a more robust pH, magneto, and thermo-responsive drug delivery system. Thus, MNCs are ideal candidate as smart and remotely guided drug delivery system due to a) their magneto effects and guide-ability by the external magnetic fields, b) on-demand drug release performance, and c) thermo-chemosensitization under an applied alternating magnetic field where the tumor is selectively incinerated without harming surrounding non-tumor tissues. Given the important effects of synthesis methods, surface modifications, and coating of MNCs on their anticancer properties, we reviewed the most recent studies on magnetic hyperthermia, targeted drug delivery systems in cancer therapy, and magnetothermal chemotherapy to provide insights on the current development of MNC-based anticancer nanocarrier.

Keywords:

• magnetic nanoparticles
• polymer-based magnetic nanocomposites
• hyperthermia
• external magnetic field
• targeted cancer treatments

Introduction

Insights into advanced materials have eminently illuminated several authentic features for medical applications in the past few years to promote a healthy society. Specifically, fascinating and enlightening studies on novel therapies and magnetic nanocomposites (MNCs) are prone to offer solutions for many global health issues including cancer. In a practical example, the treatment of glioblastoma cancer by magnetic hyperthermia therapy (MHT) using magnetic nanoparticles (MNPs) has been approved in Europe.¹ Although MNPs alone have shown some merits in MHT and targeted anticancer drug delivery systems, they have several downsides such as colloidal instability, fast oxidation or sensitivity to air and humidity, self-agglomeration due to dipole–dipole attractions between the bare nanoparticles (NPs), low biocompatibility, undesired biodegradability, low drug encapsulation efficiency, and rapid drug release performance.^2,3 For these reasons, MNPs alone are relatively basic and essential, but not appropriate materials for enhanced pharmaceutical effects in drug delivery systems. To overcome these issues, MNCs with multifunctional counterparts and properties have been widely used in MHT and drug delivery to obtain localized synergistic chemothermotherapy.^4,5

The non-targeted actions of high-dose chemodrugs (such as doxorubicin (DOX) and 5-fluorouracil (5FU)) can cause unwanted side effects.⁶ The Food and Drug Administration (FDA) has already approved five nano-carriers, including inulin, alginate, liposomes, albumin NPs, and polymeric micelles.^7,8 These polymers are pH-sensitive, biocompatible, and biodegradable, which makes them suitable to be incorporated in novel nanotherapies.^9–11 In addition, biocompatible polymers can potentially be naturally eliminated from the body, because of their enzymatic degradation by colonic microbial agents.^12,13 However, polymers with single-modality function (eg, their pH-sensitive structure) are inefficient for multi-stage and targeted cancer treatments; since pH changes in the human body take a long time.^14,15 Incorporating biocompatible polymer in MNCs to fabricate polymer-based magnetic nanocomposites (PMNCs) will result in multifunctional and magnetized polymer structures with the ability to overcome challenges that lie ahead for the multi-stimulus responsive drug nanocarrier systems, MHT, and magnetic targeting.¹⁶ Another factor is determination of the appropriate dosage of administered chemodrug to improve tumor drug uptake and avoid the dose-limiting toxicity. Therefore, development of external-stimuli responsive drug delivery systems by combining chemotherapy, MHT, and magnetic targeting can be used in order to reduce off-target toxicity of the applied drug and subsequently enhance the delivery of sufficient drug dosage to the target tumor with precise accumulation of MNCs onto the heated tumor area. This reflects that MNCs and PMNCs as low-cost and smart nanocarrier systems have gained tremendous attention in remote controllability and magnetothermally assisted drug release. This is due to their excellent structural properties including pH, thermo, and magneto-responsive along with high magnetization, colloidal stability, drug-conjugation, biodegradability, and biocompatibility manner.^4,17,18 In this manner, MNCs have been studied for MHT,^19–23 targeted drug delivery,^24,25 bioseparation,^26,27 biosensing,^28,29 cell labeling,³⁰ targeting and immunoassays,^31–33 targeted cancer therapy,^34,35 and magnetic resonance imaging (MRI).^36–39

In an ideal scenario of chemothermotherapy using MHT, alternating magnetic field (AMF) can generate local heating onto MNCs and also tumor cells for thermal treatment and also a heat-dependent release of drug to sensitize cancer cells.⁷ Improving the heat efficiency in MHT is not only related to externally applied parameters such as AMF and field frequency/amplitude but also attributed to the internal factors, including the type of the cancer cells and concentration, surface modification, morphology, viscosity, magnetic susceptibility/saturation magnetization, the average carrier alignment, size, shape, and nanostructure of MNCs. Another clinically related feature in MHT is the influence of intratumorally produced temperature on the blood flow within the tumor and the surrounding normal cells that a rise in temperature causes blood vessel dilation and subsequent enhancement of the permeability of vascular walls. In reality, the therapeutic benefit of hyperthermia on cancer has multiple causes, including but not limited to enhancement of drug absorption and cytotoxicity, greater sensitivity to radiation, and probably even more important, immunomodulation; generation of heat-shock proteins during hyperthermia can also play roles.

It should be mentioned that MHT alone cannot guide the injected MNCs in different areas of the human body, which is potentially addressed by using an external magnetic field (EMF) to steer and localize MNCs onto the targeted tumor tissue. EMF also reduces the possibility of obstruction in blood vessels and obtains extended retention once the particles are injected into tumor tissues to enhance the accumulation of MNCs for an effective inflammation procedure. Further, EMF can be used to remove MNCs from the tumor tissue. It is highly suggested to remove the administered MNCs from the body after their oxidative damage and cancer-killing effects⁴⁰ since the MNCs might penetrate the blood–brain barrier and generate the iron disbalance in the body causing Alzheimer’s⁴¹ and Parkinson’s disease.⁴² An important challenge is declining the effects of AMF and EMF with distance to appropriately heat and guide the administered MNCs.⁴³ Thus, it is vital to overcome the mentioned issues in order to use MNCs in combination with hyperthermia and targeted cancer therapy as the cornerstone of the nanodrug delivery systems and localized cancer treatments.

Several studies have been devoted to reviewing the use of various MNCs for the development of MHT⁴⁴ and smart drug delivery systems.^45–49 However, there have been no comprehensive reviews on fabrication methods, properties of MNPs as well as drug-loaded MNCs for magnetothermal chemotherapy using MHT and magnetic targeting, in which this study was conducted. As the main objective of this review paper, it is vital to build a bridge between the physicochemical properties of MNCs and a promising strategy of stimulus-triggered drug delivery via incorporating diagnostic and therapeutic agents into a single biocompatible nanoplatform. The purpose of this review article is to minutely discuss the structure of magnetism and different synthesis methods and surface modifications of MNCs for drug delivery systems, MHT, as well as magnetic targeted cancer therapy. This paper mainly focuses on the recent articles with respect to development and interests within the key parameters of synthesis of MNCs for the pH-responsive and magnetothermal-facilitated drug release, heat treatment, and targeted cancer therapy.

Investigation of Magnetic Properties

MNPs are essential counterparts of MNCs. They can be classified into magnetic alloy NPs (eg iron, nickel, and cobalt) and magnetic metal oxide NPs (eg iron oxides, nickel oxide, and lanthanum strontium manganite). Magnetic nano-agents such as ferromagnets (FM) and ferrimagnets (FiM) have potent molecular exchange interactions with the ability to produce a magnetization. Permanent magnets are the well-known FM and FiM, which are not limited to a submicroscopic scale. A material acts as an FM when the exchange interactions between atoms set off magnetic moments in order to support one another, and analogous orientation of magnetic moments would arise below a critical temperature known as Curie temperature (Tc).⁵⁰ A material performs as a paramagnet beyond Tc where its magnetization disappears since thermal energy overcomes the exchange interaction. FiM performs similarly to FM with comparable Tc, quick and high magnetization, and hysteresis, although lower than FM upon EMF. Table 1 shows categories of magnetisms and their magnetic properties.

Table 1 Categories of Magnetism Materials and Their Characteristics

Type	Example, Refs	Net Magnetization	Response to EMF	Response After Removing EMF	Susceptibility
Diamagnetic	Au, Ag, and most of known elements, ^Citation51,Citation52	0	Weak response, aligned contrary to the applied EMF	Do not retention of magnetic moment	Very small and negative (−10-4 to −10-6)
Paramagnetic	Na, Mg, Al, Ga, Li, Ta, Cu,^Citation53	✓	Sectional response, aligned positive to the direction of EMF	No retention of magnetic moment	Small and positive (10–4 to 10–6)
FM	Fe, Co, Ni,^Citation54	✓	Potent response, aligned parallel to EMF	Remains a residual magnetic moment	Very large and positive (104 to 106)
Antiferromagnetic	MnO, CoO, NiO, CuCl2,^Citation55	0	Potent response, aligned parallel to EMF	A residual magnetic moment remains	Small and positive (0.1–1)
FiM	BaFe12O19 Fe3O4, MnFe2O4, Magnetite (Fe3O4) and maghemite (γ-Fe2O3),^Citation56	✓	Potent response, aligned parallel to EMF	A residual magnetic moment exists after the removal of EMF	Large and positive (103 to 105)

The Types of Magnetization Materials

FiM is a magnetic ordering that attributed to the metal ions and electron interactions as the magnetic moments with spontaneous alignment below Tc, causing the formation of magnetization.⁴⁴ The typical difference between FM and FiM (which was not differentiated from FM before 1948) is that the FiM materials consist of different types of atoms in the resulting magnet’s unit cell. Without EMF, the lattice of FiM is a combination of two separate sub-lattices with varied strengths, non-parallel formation, and dissimilar magnetic moments where one direction has a stronger magnetic moment than others to obtain the net magnetization. FiM shows similar net magnetization and hysteresis to FM, nevertheless, with opposite magnetic ordering and they become FM and paramagnets at below and above Tc, respectively. The FiM structure can be indicated in MgFe₂O₄⁵⁷, γ-Fe₂O₃⁵⁸, α-Fe₂O₃⁵⁹, CoFe₂O₄⁶⁰, and MnFe₂O_4.⁶¹

Transition metals of Fe, Ni, and Co and their alloys are the most well-known FM, indicating analogously oriented atomic magnetic moments and magnetic characteristics even after removal of EMF. In FM, the large electron exchange forces may lead to the potent exchange interactions among neighboring moments and the alignment of the two-electron spins with strong magnetizations. FM with small and large coercive fields (Hc) are such soft and hard magnetic materials, respectively. Compared to soft magnetic materials, the magnetization of hard magnetic materials desires more energy and subsequently a wider hysteresis loop. Magnetic responses of FM at below Tc order are aligned in the same direction and provoke the decrease of exchange energy of the material. FM particles maintain their spontaneous magnetization with aligned atoms in one direction without applied EMF. This is due to the lack of tendency in the electron field of FM’ atoms.⁶²

Superparamagnets have indicated a great potential in magnetic nanofluid hyperthermia treatment and targeted cancer therapy.^3,63 Above block temperature (which occurs just before TC)⁶⁴, both small nanosize of FM or FiM particles (20–100 nm) have a tendency of superparamagnetic behavior that the amount of domain walls for each particle decreases to only one wall with a single domain, due to low energy statement and the absence of boundaries. Superparamagnets obtain magnetization quicker than other forms of nanomagnetism, and each atomic magnetic moment preserves its ordered state, causing a non-hysteretic curve with zero Hc and remanent magnetization (Mr).⁶⁵

In the periodic table, chromium is the only element as antiferromagnetism with a weak state of magnetism and positive susceptibility, and become paramagnets above Tc.⁵⁵ Diamagnetism is a common feature of all material resulting from the non-supportive performance of electrons in the presence of EMF. Without EMF exposure, diamagnetic materials exhibit zero magnetization (due to lack of orbital shells and unpaired electrons), whereas with EMF exposure, they show a negative magnetization. In a recently reported study,⁵¹ EMF around the synthesis container could collect the magnetic iron particles, while diamagnetic Bismuth and gold NPs were successfully separated.⁵¹

The Synthesis of Magnetic Nanoparticles

Since 1996, the synthesis of MNPs with various physicochemical properties and morphologies has been widely carried out by several techniques based on chemical, physical, and green routes (Figure 1a).⁶⁶ Ferrous sulfate or iron(II) sulfate is an inorganic salt that can be utilized as precursor in the formation of iron compounds. For many years, the LaMer burst nucleation has been used to explain the procedure of the nucleation and development of NPs.^67,68 In 1950, LaMer and Dinegar declared a model for the mechanisms of nucleation-growth of sulfur particles related to the three phases (Figure 1b).⁶⁸ In the first stage, the metal salts were soluble in the homogeneous media than via increasing salt concentration, the reduction process of metallic NPs called nucleation (Cmin) starts. The continuous increase of the salt concentration leads to a certain critical supersaturation level (Cmax) since the molecules construct nuclei and the media becomes heterogeneous.⁶⁸ In the second stage, the nuclei is resulted from the collisions between ions and molecules of the media in an autonucleation procedure. In particular, the Cmax concentration of the metal salts declines quickly to Cmin because of the development of clusters, and then the amount of nucleation drops instantly to zero. In the third stage, the cluster steadily increases to form a critical size, while the cluster converts into a structure known as “seed”. Lastly, the seed keeps growing to obtain a nanocrystal by enhancing metal atoms as the number of salts in the media declines to a solubility concentration of nanocrystals (Cs) and the NPs fabrication.^68,69 In MHT, MNPs are unique nano agents with magnetic properties, which is a basis of several advanced biomedical demands (Figure 1c).

Figure 1 (a) Schematic of popular methods for synthesis of MNPs, (b) Representation of the principle of nanoparticle nucleation due to LaMer’s mechanism of (sulfur) nucleation.^67,68 (c) MHT using MNCs for different biomedical applications.⁷⁰

Chemical Methods

The chemical co-precipitation method is widely applied in the synthesis of many different MNPs and MNCs, due to its low-cost, simple, and fast chemical route.⁷¹ However, this method may result in low crystallinity and inhomogeneous size distribution. In chemical vapor condensation, solid nanostructures or thin films are deposited on a solid substrate from the reaction of vapor phase metal halide or organometallic precursors at high temperatures to synthesize compounds such as FeSi, CoSi, and MnSi, as well as their alloys (including cubic FeGe nanowires⁷² and Fe_1-xCo_xSi nanowires⁷³). During this process, high-purity gas like argon or helium is used in a heated bubbling device comprising the liquid magnetic precursor. Then, it is vaporized at high temperature for producing a flow of carrier gas to deliver the precursor vapor from the heated tubular furnace to the chamber and finally a collection of the magnetic particles. In an innovative study by Farhanian et al,⁷⁴ photo-initiated chemical vapor deposition was assisted with a jet-assisted fluidized bed configuration for a large-scale synthesis of IONPs. In a different technique, sol–gel reactions of condensation and hydrolysis of magnetic salts were used for the synthesis of MNPs.^75,76 Similar to the sol–gel method but with a different hydrolysis reaction, polyol can be used as a reducing and stabilizer solvent to fabricate less aggregated MNPs.^77,78 Polyol approach is commonly applied for enhancing MNPs solubility.⁷⁷ In an electrochemical technique (such as laser pyrolysis), localized heating of gas mixture by a continuous wave of CO₂ laser controls the chemical reaction.⁷⁹ Table 2 summarizes various fabrication methods and physicochemical properties of MNPs.

Table 2 Various Methods for Synthesis of MNPs

Routes	Methods	Condition	Temp (°C)	Duration (min/hrs/d)	Nano-Size	Capability of Morphology Control	Production Scale	Refs
Chemical	Co-precipitation	Facile in ambient conditions	20–150	Quick (min)	Small	Poor	Large	[80,81]
Chemical	Sol-gel synthesis	Difficult	25–200	Average (hrs)	Small	Good	Normal	[82,83]
Chemical	Electrochemical	Difficult	Room	Slow (hrs - d)	Small	Good	Normal	[84,85]
Green	Biosynthesis	Difficult	Room	Slow (hrs)	Average	Poor	Large	[86,87]
Green	Green-based	Easy	Room	Quick (min - hrs)	Small	Poor	Large	[20,37]
Physical	High-temperature thermal decomposition	Difficult	100–350	Slow (hrs - d)	Very small	Good	Large	[88,89]
Physical	Hydrothermal or solvothermal synthesis	Facile, High pressure	150–220	Slow (hrs - d)	Very small	Great	Large	[90,91]
Physical	Pulsed laser ablation pyrolysis	Difficult	Room	Quick (min - hrs)	Small	Good	Large	[92,93]
Physical	Microwave-assisted	Difficult	Room	Slow (hrs)	Average	Good	Normal	[94,95]
Physical	Microemulsion	Difficult	20–80	Average (hrs)	Small	Good	Small	[96,97]
Physical	Sonolysis or sonochemical	Very facile	20–50	Very quick (min)	Small	Poor	Normal	[98,99]
Physical	Aerosol/vapor	Difficult, controlled conditions	Above 100	Average (min- hrs)	Small	Good	Large	[100,101]

Biosynthesis Methods

Bio or green methods are fascinating to improve the biocompatibility of MNPs due to the usage of natural stabilizers and reducing agents such as microbial enzymes and plant extracts with anticancer compounds.¹⁰² In a study by Kumeria et al,⁸⁶ green fabrication of superparamagnetic iron oxide nanoparticles (SIONPs) were carried out by using bacteria mariprofundus ferrooxydans as a reducing agent. In a different study,⁸⁷ a 15 nm spherical-shaped Fe₃O₄ was prepared by a combination of co-precipitation and green synthesis using the reducing agent of bacterial metabolites. It is also worth mentioning that several studies have used plant-based stabilizer/capping agents in the green synthesis methods of MNPs.^103–106

Physical Methods

The physical method of pulsed wire discharge uses electrical wire explosion in a vacuum, where joule heating vaporizes solid wire to obtain a high-density current over the wire, that subsequently, a shockwave moves out the generated vapor and drops, then it gradually is cooled down at the ambient condition to finally collect the solid MNPs.¹⁰⁷ This technique has various benefits such as a high rate of fabrication, good efficiency of energy transformation, and a reasonable cost of the required devices. As a relatively simple and recent method, microwave-assisted procedure uses the magnetic precursors’ mixture under the external electric field of the microwave for triggering molecule reorientation and homogeneous inner heating reactions to synthesize MNPs. The thermal decomposition technique has been recommended based on two classifications: i) conventional reaction method with primary processes at room temperature and subsequently at high temperature in an open or closed reaction vessel and ii) hot-injection method as precursors were injected into the heated reaction mixture. MNPs with desired physicochemical properties are also fabricated through low-cost hydrothermal methods using organic solvent instead of water in the reaction medium. For instance, Fe₃O₄ NPs were successfully fabricated using a mixture of Fe salts, urea, sodium citrate, and acetate in ethylene glycol solution to obtain a homogeneous dispersion as the reaction mixture was then transferred to an autoclave and heated up to 200 °C for 24-hr.⁸⁸ Another interesting procedure for MNPs fabrication is microemulsion, where the surfactant molecules are presented at the interface border area of oil and water via a monolayer, which can be based on oil/water (oil dispersed in water) or water/oil (water dispersed in oil). In this particular case, the surfactant molecules with hydrophobic and hydrophilic tails and head, respectively, are dissolved within the oil and water phase or inversely.¹⁰⁸ In this method, the most common surfactants are polyvinyl pyrrolidone (PVP), cetyltrimethylammonium bromide, sodium dodecyl sulfate, and bis (2-ethylhexyl) sulfosuccinate. MNPs with a complex design have been fabricated by sonolysis or sonochemical methods using high acoustic waves and cavities or bubbles to generate 5000 °C and 1800 kP pressure in a short reaction time.⁹⁸ In addition to the above-mentioned procedures, MNPs can be fabricated through flow injection (with high reproducibility) and aerosol/vapor technique.

Investigating the Toxicity of Magnetic Nanomaterials

Toxicity issues are vital aspects of the drug delivery systems and cancer therapy. The toxicity of MNPs can be relevant to various parameters, including dose-dependency, duration, concentration, and physicochemical characteristics (such as surface modification, dimension, and shape) of MNPs.¹⁰⁹ García et al investigated the in vitro toxicity of copper, titanium dioxide, CuZnFe₂O₄, and IONPs at concentrations ranging between 20 and 100 μg/mL and found no toxicity from the tested IONPs.¹¹⁰ Despite this, the surface of bare MNPs may exert toxicity associated with their large surface area and the generated reactive oxygen species (ROS), due to the inadequate reduction of oxygen. ROS can be produced via the leaching of metal ions, or release of oxidants via enzymatic degradation of MNPs.¹¹¹ The inflammatory response of cells with an increased amount of ROS triggers oxidative stress and subsequently attacks the cellular proteins, lipids, and DNA, causing carcinogenesis, necrosis, apoptosis, and finally cell death.¹¹² This reflects that the amount of administered MNPs must be precisely controlled. After the administration of MNPs, the ratio of ROS generation can be analyzed via dichlorofluorescein diacetate as a fluorescent marker. It is worth mentioning that ROS may also eliminate damaging cancer, for instant.¹¹³ In a report,¹¹⁴ 25 µg/mL spherical Fe₃O₄ MNPs (72.6±0.6 nm) showed damages against erythrocytes for in vitro study to analyze the eryptosis indices. For the in vivo evaluation using female CD^® IGS Rats at 8 weeks of age, 12 mg/kg Fe₃O₄ NPs caused apoptosis of circulating erythrocytes. In short, Fe₃O₄ NPs caused pathological changes in cell membranes, and oxidative stress caused cancer cell elimination in vitro and also in vivo.¹¹⁴ In order to decrease the toxicity against normal cells, MNPs have been incorporated with various counterparts and coating agents to synthesize MNCs and PMNCs with increased multifunctional properties, biocompatibility, and free magnetic ions onto the target organ causing oxidative stress and toxicity against target cancer cells. In a study, bare MNPs and n-octyltriethoxysilane-coated MNPs were fabricated for anticancer analysis using ReNcell VM (human neural stem cells) and PC12 (rat pheochromocytoma) at different concentrations from of 0–64 µL for 24 h.¹¹⁵ The results showed the higher safety and biocompatibility from coated MNPs than uncoted MNPs. In a different study,¹¹⁶ bare nanomagnetite, starch-coated nanomagnetite, and dextran-coated nanomagnetite were utilized in toxicity assays on rat pheochromocytoma PC12 tumors at a concentration range of 0.01–0.5 mg/mL for 1, 2, 3, 24, 48 and 72-hr. In the XTT cytotoxicity assay, bare nanomagnetite was on the outer surface and unable to enter the cells without any killing effects up to 0.1 mg/mL; however, 49% cell death was indicated at 0.25 mg/mL after 72-hr. Starch-coated nanomagnetite had insignificant cell death during 24-hr, whereas it caused 30% cell death at 0.1 mg/mL after 72-hr. In addition, at 0.25 mg/mL, it indicated almost 100% killing effects against PC12 tumors after 72-hr. The anticancer effects of dextran-coated nanomagnetite were ineffective at concentrations up to 0.1 mg/mL; however, it was significantly enhanced at 0.25 mg/mL after 72-hr and also 24-hr. Overall, this study aimed to indicate the positive influence of coating for MNPs and also the effects of incubation time and concentration of MNPs in the elimination of PC12 cells. It is worth to mention that the pharmacokinetics and degradation pathway of counterparts of MNCs such as MNPs cores and polymer coating agents could be the potential limitations for the clinical usage of innovative MNCs.¹¹⁷

The specific dosage of the magnetic complex to eliminate 50% of the tested animals in a required duration can be measured by LD-50.¹¹⁸ In an interesting study, LD-50 of dextran-coated IONPs (2000–6000 mgFe/kg) was almost ten times higher than that of uncoated IONPs (300–600 mgFe/kg body).¹¹⁹ Similarly, IONPs coated with carboxy-dextran revealed the LD-50 value of 35 mmol Fe/kg,¹²⁰ showing the positive effects of coating MNPs in the body’s innate biological response related to an appropriate dose of the administered MNPs.¹¹⁸ In vivo study indicated that almost 100% dextran-coated IONPs were cleared from the blood of rats 84 days post-injection.¹²¹ It should be mentioned that there is still insufficient evidences to understand the degradation and extended circulation time of magnetic materials in the body, since it requires long-duration monitoring for in vivo toxicity (months or even years), and also ethical issues of immolating numerous animals for LD-50 analysis. In addition, FDA does not consider the customary LD-50 analysis anymore. As an alternative procedure, in vitro analysis was suggested; however, it caused low number of metabolic actions of cell lines with less required regulatory parameters for various types of cells. In the future toxicity studies, it is therefore vital to evaluate further toxicity evaluations of MNCs in the human body.

Application of Polymer in Stabilizing Magnetic Nanocomposites

Polymer-based nanocomposites with pH-responsive structures could be promising nanodrug systems for future cancer treatment.^122,123 However, changing pH in the human body is a long time process with incapability to be adjusted externally. This issue can be tackled via using polymers as solid supports or coating agents for MNPs as fillers to fabricate multifunctional polymer-based magnetic nanocarriers as remotely controlled drug delivery systems.^124,125 Biopolymer is the most abundant polymer on earth with low cost, desired biocompatibility, biodegradability, and physicochemical properties.¹²⁶ It should be considered that some synthetic and also natural polymers have issues of instability, low biocompatibility, undesired biodegradability, and high-cost chemical modifications.¹² As natural excipients, various polysaccharides such as chitosan, alginate, dextran, nanocellulose, pectin, hyaluronic acid, starch, pullulan, guar gum, glycogen, beta mannan and inulin can be used in advanced anticancer nano-drug systems.¹² Table 3 shows some polysaccharides utilized in the fabrication of biocomposites. Only two polysaccharides of chitosan and curdlan are cationic, while other listed polysaccharides are anionic or neutral. Polysaccharides can be obtained from different plants, vegetables, fruits, grains, glycogen, and red alga (such as Glaucocystis, and Chaetomorpha melagonicum), liver and muscles of animals such as ascidians, and some bacteria.^127,128 In polysaccharides, glycosidic linkages bound many monosaccharides with long chains. In drug delivery applications, natural cellulose and chitosan are the first and second most popular polysaccharides, respectively. The polysaccharide-based products possess desirable properties, including the capability to bind with various drugs, great swelling behavior, pH gradient behavior, and high biodegradability for innovative antitumor drug delivery systems.¹²⁹ Each property has its particular pros and cons for developing a topical nanodrug formulation with improved stability and therapeutic effects.¹³⁰ The vitality of drug-loaded polysaccharides and polymers is a consequence of their potential to deliver a desirable amount of drugs to cancer cells without significant effects against normal cells.^130,131 For example, curcumin was loaded onto crystalline nanocellulose and caused almost three times higher colorectal cancer cell death than that of curcumin alone.¹³² The side-effects of chemotherapy have been alleviated by loading a sufficient dosage of chemo-drugs onto various polymer-based carrier systems, for instance, poly(2-vinyl pyridine)-b-poly(ethylene oxide) nanomicelles,¹³³ chitosan microspheres,^134,135 carboxymethyl cellulose,¹³⁶ and crystalline nanocellulose to name a few.¹³⁷ Thus, with the immense demand for using biocompatible nanocarriers in anticancer drug delivery systems, natural polysaccharides should be explored for advanced cancer treatments.

Table 3 Various Polysaccharides Utilized in Composites

Polysaccharide	Source	Charge	Main Functional Groups	Ref.
Cellulose	Botanical and bacteria	Negative	OH	[138]
Chitosan	Shells of crustaceans such as shrimp, lobster, crayfish and oyster	Positive	OH, COO−, NH2	[9]
Starch	Microbial product in wine, glucose, plants	Neutral	OH	[139]
Alginate	The cell wall of brown marine algae	Negative	OH, COO^−	[140]
Mannan	Plant	Neutral	OH	[141]
Hyaluronic acid	Connective, epithelial, and neural tissues	Negative	OH COOH	[142]
Heparin	Animal tissues	Negative	OH OSO3H	[143]
Pullulan	Fungus Aureobasidium pullulans	Neutral	OH	[144]
Levan	Archaea, fungi, bacteria, and a limited number of plant specie	Neutral	OH	[145]
Elsinan	Fungi, bacteria	Neutral	OH	[146]
Gellan	Bacterium Sphingomonas elodea	Negative	OH	[147]
Curdlan	Fungi, bacteria	Positive	OH, COO^−	[148]
Succinoglycan	Agrobacterium sps, Rhizobium sps, Rhizobium meliloti, Alcaligenes faecalis	Negative	OH	[149]
Zooglan	Zoogloea ramigera	Negative	OH	[150]
Xanthan	Xanthomonas campestris	Negative	OH	[151]
Dextran	Leuconostoc mesenteroides, Lactobacillus spp and Streptococcus mutans	Neutral	OH	[152]

In MNCs, various coating materials and surface functionalization with peptides, antibodies, small molecules, and aptamers could improve the colloidal stability, functionalities, and other advantages of MNPs for biological regimes and magnetothermal chemotherapy to obtain combination therapy of MHT and magnetic targeting, switchable synthetic cell surfaces, and magnetothermal chemotherapy. The most well-known coating for MNPs are organic materials (like surfactants, polymers, and biological molecules), and inorganic materials (such as metals and metal oxides) (Figure 2). The popular biocompatible polymers used in MNCs are dextran, PVP, starch, albumin, polyethylene glycol (PEG), chitosan, ethylcellulose, polyethylenimine (PEI), polymethyl methacrylate (PMMA), polyacrylic acid (PAA), polylactic acid (PLA), polyvinyl alcohol (PVA), citric acid, glucuronic acid, gelatin lipids, liposomes, dendrimers, polyacrylamide, and bisphosphonates. Various studies have reported the fabrication of different MNCs (with various size ranges) by using chitosan-coated MnFe₂O₄ (18 nm),¹⁵³ dextran-coated Fe₃O₄ (21 nm),¹⁵⁴ cellulose matrix (87.12 nm)/Fe₃O₄ fillers (11.01 nm),⁸⁰ PEG-coated NiFe₂O₄ (16 nm),¹⁵⁵ phosphate-coated Fe₃O₄ (14 nm),¹⁵⁶ tetraethyl orthosilicate coated MnFe₂O₄ (14 nm),¹⁵⁷ Zn_0.9Fe_0.1Fe₂O₄ (11 nm),¹⁵⁸ stevioside-coated Fe₃O₄ (3 nm),¹⁵⁹ citric acid-coated Mn_xFe_3-xO₄ (34 nm),¹⁶⁰ aminosilane-coated Fe₃O₄ (100 nm),¹⁶¹ and oleic acid-coated Fe₃O₄ (45 nm).¹⁶²

Figure 2 Inorganic and organic materials and their properties regarding their use as functionalizing and coating agents for the synthesis of MNCs.¹⁶³

MNCs have various shapes and structures, including core-shell, shell-core, matrix dispersed, shell-core-shell, yolk, dumbbell, and Janus (Figure 3).^164,165 In core-shell, either organic or inorganic materials coat the magnetic core.^166,167 However, if the magnetic core is not centered, it is called “yolk” form.¹⁶⁸ When the magnetic core is placed between two functional materials, it is termed shell-core-shell NPs. In a matrix dispersed system, a matrix acts as the dispersant agent for MNPs to avoid aggregation over large clusters and possibly maintain superparamagnetism. Janus NPs are associated with two sides of a magnetic core and different functional materials.^169,170

Figure 3 Different coating structures for synthesis of MNCs and common shapes of the magnetic core.¹⁷¹

In a study by Wulandari et al,¹⁷² Fe₃O₄ NPs were fabricated by the ex-situ co-precipitation method and cross-linked with chitosan using tripolyphosphate/sulfate. The results from scanning electron microscopy (SEM) and X-ray diffraction (XRD) spectrum revealed that the increase of chitosan coating agent enhanced and declined the SEM size and crystallite size of the NPs, respectively. In different investigations, the in-situ fabrication of MNPs was reported via dispersing the iron ions in the polymer particles for subsequent MNP nucleation as conjugated strongly with functional groups of the polymer to synthesize PMNCs.^50,173,174 Recently, Malhotra et al¹⁷⁵ synthesized bare Fe₃O₄ NPs through co-precipitation method followed by the use of hydrothermal method at 200 °C and carbonized organic starch (0.19 g) to simultaneously functionalize and also coat the bare Fe₃O₄ (0.5 g) abbreviated as Fe₃O₄@C (Figure 4a–g). Compared to the bare MNPs, the transmission electron microscope (TEM) image of Fe₃O₄@C exhibited better dispersion and bigger sizes ranging from 16 to 52 nm with around 2 nm thickness of the carbon coating. Raman spectrum of both Fe₃O₄ and Fe₃O₄@C displayed peaks at 660–680 nm attributed to the presence of MNPs, whereas Fe₃O₄@C showed additional peaks between 1100 and 1800 cm⁻¹ related to the formation of diamond-like carbon (Figure 4C). Fe₃O₄ and Fe₃O₄@C exhibited analogous XRD results (Figure 4D) with an average crystallite size of 18 nm and 24 nm, respectively, since the hydrothermal process did not damage the crystalline structure of Fe₃O₄@C. From Figure 4E, Fourier-transform infrared spectroscopy (FTIR) spectrum of the samples indicated the magnetite formation with the peaks at 572 cm⁻¹ and 700 cm⁻¹. Thermogravimetric analysis (TGA) for Fe₃O₄ and Fe₃O₄@C displayed a weight loss of ~2.6% and ~0.4% at 40 °C−115 °C, respectively. Interestingly, the saturation magnetization (M_s) of Fe₃O₄@C (135 emu/g) was higher than Fe₃O₄ (94 emu/g) (Figure 4G); thus, the coated Fe₃O₄ NPs revealed better multifunctionality and physicochemical properties than the uncoated Fe₃O₄ NPs. Furthermore, from the results of cytotoxicity assays using model organisms of adult zebrafish, Fe₃O₄@C was safer and less toxic than uncoated Fe₃O₄. The above examples indicate the positive effects of polymer coating for MNPs to fabricate PMNCs with high colloidal stability, desired size ranges, improved functional groups, and biocompatibility.

Figure 4 TEM morphologies and size distribution of (A) Fe₃O₄ NPs and (B) PMNCs of Fe₃O₄@C. (C) Raman spectrum, (D) XRD, (E) FTIR spectrum (F), TGA curves, and (G) M_s of Fe₃O₄ and Fe₃O₄@C.¹³⁶

Studying the Methods Used in the Hyperthermia and Cancer Treatment Based on Magnetic Nanocomposites

In advanced drug delivery systems, MNCs can show controlled and precise release of drug under adjustable conditions, including electric and magnetic fields, ultrasound waves, heat, light, enzymes, and pH changes.⁴⁵ MHT (known as magnetic thermal ablation or magnetic thermotherapy) has been a distinctive technique in cancer treatment for nearly past five decades with the vitality related to using a noncontact, safe, and low-cost external heating source of AMF (typically 50–1000 kHz). In this treatment method, the temperature is commonly ranged from 42 °C to 48 °C onto accumulated MNPs at the target malignant tissue to effectively kill cancer cells with negligible or no damage against healthy cells.⁴⁴ MHT can use various drug-loaded MNCs to i) closely control the heating yield by adjusting the applied AMF, ii) maintain the efficiency of the cancer treatments without detrimental side effects, and iii) consumption of less drug-administered to overcome the treatment shaping problems in the conventional chemotherapy alone.^17,176 Moreover, AMF is potentially capable of achieving “on/off” drug release from MNCs in target tissue with treatment scale-up capability compared to chemotherapy and surgery, which are confined to poor cancer treatment index and harmful side effects. The AMF strengths regulate hyperthermia temperature in the cell medium through the heating capacity of a magnetic nano heater measured by specific absorption rate (SAR) or the specific loss power (SLP).^37,177 Figure 5a shows the schematic magnetic hyperthermia procedure and thermal dissipation procedure, including Néel and Brownian relaxation processes from MNCs under AMF. The increased temperature positively increases physical rotation of magnetic particles, which subsequently can improve M_s, magnetic anisotropic constant (K), and a Brownian heating loss mechanism.^44,178,179 In this manner, the SAR value of MNPs is related to M_s (Figure 5bi), whereas the highest SAR value is at a K (Figure 5bii) and specific NPs size (Figure 5biii) for obtaining enhanced biological effects. Furthermore, the SAR value is attributed to morphology, composition, and concentration of MNPs, as well as the amount of the generated heat by a unitary ratio of magnetic particles per time under the exposure of AMF.^19,180 As schematically shown in Figure 5a, during the hyperthermia process, magnetic nanofluids are placed in the middle of a helical coil with several turned-loops connected to a water-cooled system. Also, currents of AMF introduce different frequencies and field strengths in order to increase the temperature for paramount effects of MHT. For the precise measurement of SAR value, the dT/dt graph is obtained from the initial slope of ‎the temperature and time curves using the linear fitting method classified into the initial slope, full-curve fit method, and corrected slope. The initial slope method relies on the initial temperature changes without measuring the complete heating curve as a non-linear temperature rises to underestimate SAR values by 25%. The full-curve fit technique is able to estimate 10% accuracy for the SAR value. The third and most accurate technique is the corrected slope. More details on several proposed methods to calculate SAR from heating data are explained well in different studies.^181,182 The SAR values of several MNPs and MNCs are listed in Table 4. In a recent study, the applied AMF increased the temperature of MNPs (13 nm) up to 42 °C after 128 sec with the SAR value of 350 W/g.¹⁸³ This study concluded that the fabricated MNPs with a TEM size of around 10 nm is appropriate for the hyperthermia test, while it did not consider that enhanced temperature of MNPs within the secure hyperthermia range (42–48 °C) is appropriate for MHT. It should be noted that different studies stated that the secure hyperthermia range can be in the range from ~42 °C to ~48 °C.^{20,37,44,184,185} The generated temperature from AMF ranging from 42 to 48 °C causes the cells to favorably undergo apoptosis, however above this range, they mostly undergo necrosis,^186,187 in which the cell elimination by apoptosis is preferable over necrosis to prevent unwanted inflammation and metastasis.¹⁸⁷ Furthermore, above 50 °C and 200 °C may eventuate in coagulation and undesired burn at the affected areas.

Figure 5 (a) Schematic presentation of thermal dissipation process (Néel and Brownian relaxation processes) from MNCs under AMF.^188,189 (b) The SAR value of MNPs related to (i) M_s, (ii) K, and (iii) particle size (r) with SLP, respectively.^188,189 (c) Illustration of different methods for hyperthermia treatment.¹⁹⁰

Table 4 Overview of the Recent Studies on SAR Values of MNPs, MNCs, and PMNCs with Respect to Size, Morphology, and Factors Used in Magnetic Hyperthermia Analysis

Magnetic Composites	Type of Analysis, Shape, Size (nm)	Field (kA/m)	Frequency (kHz)	SAR (W/g)	Ref.
Fe3O4/poly(styrene-co-maleic anhydride)	TEM, nanoclusters, 33	23.8	302	253	[191]
Fe3O4/PVP	TEM, spherical, 45	32.5	400	1100	[192]
Fe3O4/graphene oxide	TEM, spherical, 45	32.5	400	5160	[192]
CoFe2O4	TEM, spherical, ~ 10	30.1	265	91.84	[193]
Ag-Fe3O4	TEM, spherical, ~ 10	13.9	274	43	[194]
NiFe2O4	TEM, spherical, 4.4	23.7	170	~ 11	[195]
Fe3O4/PEG	TEM, cubes, 19	29	520	2452	[196]
CoFe2O4	TEM, spherical, 5.4	95.6	329	~76	[197]
Co0.03Mn0.28Fe2.7O4/SiO2	TEM, spherical, ~22	33	380	~3417	[198]
Fe3O4/citric acid	TEM, spherical, ~10	35.88	316	49	[199]
Fe3O4/PEG	TEM, spherical, ~10	35.88	316	56	[199]
Fe3O4/ethylene diamine	TEM, spherical, ~10	35.88	316	77	[199]
Fe3O4/cetyl-trimethyl ammonium bromide	TEM, Spherical, ~10	35.88	316	47	[199]
Fe3O4/citric acid/PEG	TEM, spherical, ~10	35.88	316	60	[199]
Zn0.5Ca0.5Fe2O4	TEM, spherical, 12–14	10.2	354	~14.8	[200]
Zn0.33Fe2.67O4	TEM, spherical, ~8	35.7	360	~178	[201]
Gd-doped Mg-Zn ferrites	TEM, spherical, 50–200	5.0	600	~15–27.5	[202]
Fe3O4/garcinia mangostana fruit peel extract	TEM, spherical, ~13	75	318	98	[20]
		100	313	130
		125	312	179
Fe3O4/Punica Granatum fruit peel extract	TEM, spherical, ~11	75	318	196	[203]
		100	313	212
		125	312	232
Fe3O4	Nr	0.450	400	46.8	[204]
Fe3O4@agar	TEM, spherical, 9.2	0.450	400	23.6	[204]
Chitosan/trimethylammonium/ SPION	TEM, spherical, 13.7	27.57	360	168	[205]
Chitosan/curcumin/alginate/SPION	Nr	27.57	360	280	[205]
Peptide nucleic acid oligomers/SPION	TEM, spherical, ~15	17	183	65	[206]
CoFe2O4/meso-2,3-dimercaptosuccinic acid/DOX	TEM, spherical, 13	30	307	94.58	[207]
Chitosan/Fe3O4/5FU	SEM, spherical, 125	35	180	Nr	[208]
Fe3−δO4@silane and a thermoresponsive copolymer shell composed of 2-(2-methoxy)ethyl methacrylate and oligo(ethylene glycol)methacrylate moieties (1.14 mL)	TEM, spherical, 10	23.8	536.5	25.2	[209]
Fe3-δO4@silane and a thermoresponsive copolymer shell composed of 2-(2-methoxy)ethyl methacrylate and oligo(ethylene glycol)methacrylate moieties	Dynamic light scattering (DLS), Not reported (Nr), 55–80	23.8	536.5	61.7	[209]
Fe3-δO4@silane	DLS, Nr, 20–30	23.8	536.5	100	[209]
Graphene quantum dots-Fe3O4/SiO2	TEM, Nr, 100	14.3	409	44	[210]
DOX loaded Fe3O4/MamC/proteinAR-3 mAbs (IgG1)	TEM, Nr, 36	12.5	273	53	[211]
			205	41
			163	22
			143	19
Magnetite	TEM, spherical, 30	12.5	273	68	[211]
			205	47
			163	28
			143	24
Alginate-chitosan-Fe3O4	Nr	40	265	720	[212]
CuFe2O4	TEM, spherical, 17.3	28.5	126	192	[213]
		25.7	126	185.2
		22.2	126	177.8
		18.6	126	171.5
CuFe2O4	TEM, spherical, ~19.9	19	120	44.9	[91]
		16	120	38.0
		13	120	24.8
CuFe2O4	TEM, spherical, ~25	13.5	331	6.48	[95]
Mn0.5Zn0.5Fe2O4	TEM, spherical, ~14	6.37	178	28.38	[214]
MgFe2O4	TEM, spherical, ~9	9.2	198	~19	[215]
Solvent optimized redox tuned iron oxide	TEM, spherical, ~25	50	100	~700	[216]
Thermosensitive polymer poly(2-(dimethylamino)ethyl methacrylate) coated Fe3O4	SEM, irregular–quasi-spherical, 98	16.15	205	150	[217]
Fe3O4/hexadecylamine/oleic acid/1-octanol	TEM, rods, ~56 (length) × 10 (width)	63.81	310	862	[218]
Graphene oxide-Fe3O4-PEG	TEM, spherical, ~20	12.57	293	557.38	[219]
PLGA/Fe3O4/DOX/PVA	TEM, spherical, 172	2	205	36.27	[220]
PLGA/Fe3O4	Nr	2	205	35.73	[220]
Cetyltrimethylammonium bromide@Fe3O4	TEM, cubic, ~ 80	63.66	315	1036	[221]
Fe3O4	TEM, Pseudo spherical, 12.97	15.9	252	63.4	[222]
Fe3O4@citric acid	DLS, 46.9	15.9	252	65.8	[222]
Fe3O4@(3-aminopropyl)triethoxysilane	DLS, 68.1	15.9	252	67.2	[222]
Fe3O4@dextran	DLS, 76.6	15.9	252	55.6	[222]
SPIONs coated with PEG4.9kD-PLA6.0kD	TEM, Cubic, 18	59.6	346	558	[223]
IONPs	TEM, Cubic, 30–35	63.3	310	800	[224]
Chitosan coated Fe3O4	TEM, spherical, 37	14	335	595	[84]
Fe3O4	TEM, cubic, 37	23.9	571	213	[98]
mSiO2@Fe3O4	TEM, spherical, ~100	22	120	13.84	[89]
Poly(Nisopropyl acrylamide-co-acrylic acid)@mSiO2@Fe3O4	TEM, spherical, ~100	22	120	8.22	[89]
Fluorescent IONPs/PLGA conjugated with human epidermal growth factor receptor 2	DLS, Nr, 524	16.27	440	183	[225]
IONPs - (poly(maleic anhydride-alt-1-octadecene)- (tetramethylrhodamine 5(6)-carboxamide cadaverine - N-(3 dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride - 4-aminophenyl β-D glucopyranoside	TEM, spherical, 11.3	20	829	104	[226]
PEG coated SPIONs	TEM, spherical versus cubic, 33	9.35	300	1010	[227]
PEG coated SPIONs	TEM, spherical versus cubic, 40	20.7	300	1026	[227]
Chitosan coated Fe3O4	TEM, spherical, 30–40	20	335	711	[228]
Graphene oxide-Fe3O4	TEM, spherical, 10–21	16.72	325	543	[85]
Silica coated lanthanum strontium manganese oxide	TEM, irregular, 35	13	335	295	[229]
Chitosan-g-N-isopropylacrylamide coated Fe3O4-silica	TEM, spherical, 30–50	3	100	9.75	[230]
Fe3O4	TEM, spherical, 5–10	3	100	15.80	[230]
Trimesic acid coated SPIONs dispersed in triethylene glycol	TEM, spherical, 9	10.9	751.5	276.3	[231]
		13.8	262.2	81.2
Tetraethyl orthosilicate coated La0.7Sr0.3MnO3	TEM, wormhole-like mesopores, 45	14	350	255	[232]
Citric acid coated HoxFe3xO4	TEM, spherical, 10–15	23.87	488	337.3	[233]
Pluronic F127 coated Mn0.6Ga0.4Fe2O4	Nr	10.18	354	160.9	[234]
Cetyltrimethylammonium bromide and polycaprolactone coated Fe3O4	TEM, spherical, 21	15.75	312	255.12	[235]
Aminosilane coated Fe3O4	DLS, Nr, 100	23.87	557	320	[161]
Dextran coated Fe3O4	TEM, irregular, 21	11.93	150	52.3	[154]
Polyphenol coated Fe3O4-γFe2O3	TEM, spherical, 10–14	23.87	570	Nr	[236]
Fe3O4	TEM, spherical, 27	Nr	13,560	725	[237]
Citric acid coated MnxFe3-xO4	TEM, spherical, 34	13.36	405	Nr	[160]
Stevioside coated Fe3O4	TEM, spherical, 3	13.36	405	80	[159]
Oleic acid coated Fe3O4	TEM, Nr, 45	26.67	265	80	[162]
Chitosan coated MnFe2O4	TEM, cubic, 18	60	307	270	[153]
PEG NiFe2O4	TEM, rod, 16	3.89–5.49	260	17–22	[155]
Phosphate coated Fe3O4	TEM, spherical, 14	0.819	126	11.1	[156]
Tetraethyl orthosilicate coated MnFe2O4	TEM, spherical, 14	5.49	260	47.8–84.65	[157]
Zn0.9Fe0.1Fe2O4	TEM, spherical, 11	2.73	700	36	[158]

The Methods of Hyperthermia Therapy

Hyperthermia treatment with noninvasive properties is among procedures that have been exposed to intensive studies for treatments of various tumors with diverse biological regimes and sizes at different parts of the body.^238,239 Even without using a magnetic nano heater, hyperthermia techniques have been used in development of desired heat induction for eliminating tumor tissues with negligible damage to the surrounding normal cells.^185,240 The hyperthermia systems with the capability to regulate the topical heat induction and effectiveness have conferred to be classified into the following methods: local, localized deep, whole-body, part-body, perfusion, and interstitial hyperthermia (Figure 5c).

Investigation of Local and Localized Deep Hyperthermia

External local hyperthermia using heating devices is a coordinated therapy to target small areas of superficial tumors and selectively raise the therapeutic temperature (39–45 °C) of tumor cells under the skin. This method was used for the treatment of laryngeal cancer, neck cancer, and dendritic cell.²⁴¹ For example, local hyperthermia decreased 25% of the tumor size located at tongue squamous cell carcinoma; however, it showed the ineptitude of heat dose control and localized treatment for tumors located at supraclavicular, head, and neck areas.²⁴¹ Endocavitary and interstitial hyperthermia are subtypes of local hyperthermia to eliminate the superficial tumors via implanting the applicator (with diameter below 5 cm) onto the tumor and provide heat source of radio wave, microwave, or ultrasound using antennas, which can be challenging and uncomfortable for the patients.^239,242 Antennas or applicators release microwaves or radio waves, to increase the temperature on the surface of superficial tumors via a contacting medium.²⁴² For endocavity hyperthermia, the applicator is placed within the hollow organs including the rectum, urethra, cervix, and vagina, whereas, for interstitial hyperthermia, the applicator is placed within the interstitial area for the treatment of recurrent head and neck, breast, and prostate cancer.²³⁹ Rather than the whole-body treatment, localized deep hyperthermia increases the controlled heat around 42 °C to trigger the desired thermal damage at targeted deep-seated tumor area for the elimination of tumor cells such as peritoneal.²³⁹ In principle, it is normally applied for treatments of diseases in deeper tissues by microwaves radiofrequency, and ultrasound methods.²⁴³

Study of Whole-Body, Part-Body, and Perfusion Hyperthermia

Whole-body hyperthermia has been used as treatment of many different diseases for several decades.²⁴⁴ It was traditionally used for fever therapy by covering the patient with plastic sheets, hot liquid, a hot water jacket suit, and a heated bed to increase the blood flow and the whole body temperature up to 40 °C for increasing the permeability of cancer cell membrane with higher heat sensitivity than normal cells for subsequent treatment of the metastatic tumors.²³⁹ The most popular procedures of whole-body hyperthermia are flexible infrared chambers by heating the patient’s room and wrapping them with a warm blanket. The maximum heat of 42 °C for around 1-hr can be appropriate,²⁴⁵ albeit some systemic issues were reported for heart, blood vessels, and main body organs along with major side effects such as diarrhea, nausea, and vomiting.²³⁹ Interestingly, a combination of chemotherapy and whole-body hyperthermia treated several kinds of cancer, for instance, ovarian cancer was treated by using whole-body hyperthermia (at 41.8 °C) in combination with carboplatin chemotherapy.²⁴⁶ In the 1960s, new whole-body hyperthermia via using infrared irradiation started.²⁴⁴ Based on the report by Jia et al²⁴⁷ in 2010, commercially available medical whole-body hyperthermia devices are ET Space (Energy Technology, Shenzhen, China) and Oncotherm WBH2000 (Oncotherm, Budaörs, Hungary) and also currently available devices are IRATHERM1000 (von Ardenne Institute of Applied Medical Research, Dresden, Germany) and heckel-HT3000 (Hydrosun Medizintechnik, Müllheim, Germany). More details for hyperthermia therapy can be found from recent publications on whole body hyperthermia²⁴⁴, thermochemotherapy²⁴⁸, reirradiation and hyperthermia plus chemotherapy²⁴⁹, hyperthermia in cancer care²⁵⁰, and microwave hyperthermia²⁵¹.

Intraperitoneal perfusion is another type of hyperthermia therapy to treat a metastatic tumor in the abdominal area via using hot liquid to raise the temperature to around 45 °C²³⁹ for treatments of various types of cancer, including colorectal and ovarian carcinoma.²⁵² The hyperthermic intraperitoneal chemotherapy was effective in ovarian cancer treatment²⁵³ and resulted in a faster wound healing after abdominal surgeries than that of using chemotherapy alone.²⁵² Various in vitro and in vivo studies have indicated the therapeutic and anticancer effects of water bath heat treatment (as a kind of whole-body hyperthermia treatments) for example at 43.5 °C and 45 °C (30 min),²⁵⁴ 47 °C and 37 °C (1 min heat shock),²⁵⁵ 37, 39, and 37 °C (24-hr).²⁵⁶ As a novel option for cancer patients, combining hyperthermia and chemotherapy at considerably reduced-drug doses can indicate an exceptional efficacy, for example, therapy of breast cancer²⁴⁹ and ovarian cancer²⁵⁷. Further, clinical experiences have indicated a combination treatment of radiotherapy with chemotherapy as a desired therapy for superficial tumors, including skin carcinomas, Merkel cell carcinoma, and malignant melanoma.²⁴⁴ In addition to the mentioned treatments, combined therapy methods such as hyperthermia, chemotherapy, and immunotherapy can be used in the prospective studies.

In vitro Studies on Magnetic Nanocomposites for Hyperthermia and Cancer Therapy

MNCs are gaining attention in smart and remotely guided drug delivery systems owing to their magneto effects and guide-ability by EMF, heating capability under AMF, and visualization properties for monitoring their pharmacokinetics. With implantable and/or external EMF, drug-loaded MNCs can operate the targeted actions and time-controlled drug release inside the body for guided treatment of cancer cells. Therefore, there are plenty of room for novel multifunctional drug-loaded MNCs in tumor targeting that EMF and MHT can potentially improve targeting actions and anticancer effects.^258,259 Anticancer drugs are basically conjugated onto MNCs via chemical and/or physical binding methods. Physical binding is facile and achieved through the hydrophilic/hydrophobic, host-guest self-assembly, electrostatic, and affinity interactions.²⁶⁰ Chemical binding is obtained from the chemical interaction between the drug molecules and surface of MNCs with various functional groups such as carboxyl, amino, and thiol. Table 5 summarizes the studies devoted to various MNCs for in vitro targeted drug delivery and MHT.

Table 5 Overview of Some Recent in vitro Studies on MNCs for MHT and Targeted Drug Delivery Systems

Magnetic Nanoagent	Type of Analysis, Size (nm)	Loaded Drug	Drug Release	Origin, Type, Cell Lines	Results	Ref.
Monoclonal antibody conjugated Fe3O4	TEM, 36	DOX	~60% release at pH 5.0 medium with AMF	Gastric, cancer, GTL-16	Chemotherapy and combined therapy caused 20% and 78% cancer cell death, respectively	[261]
Silica-Fe3O4	TEM, ~ 30	Maytansinoid	Nr	Macrophage, cancer, IC21	Chemotherapy, MHT, and combined therapy caused 20%, 15%, and 78% cancer cell death, respectively	[262]
Oleosome-ZnFe2O4	TEM, ~80	Carmustine	~4% and ~23% release at pH 7.2 medium without and with AMF, respectively	Breast, cancer, SK-BR-3	Chemotherapy, MHT, and combined therapy caused 60%, 40%, and 80% cancer cell death, respectively	[263]
Fe3O4@agar	TEM, 9.2	DOX	85% release at pH 7.0 medium	Colon, cancer, HT-29	Combined therapy caused ~85% cancer cell death	[204]
MnFe2O4/ω-hydroxyacid-co-poly(d,l-lactic acid)	DLS, 160	Paclitaxel	~98% release within 18 days at 7.4 medium	Colon, cancer, Caco-2 and hASCs	The elimination of cancer cell was dose-dependent and significantly increased after treatment with increasing sample concentration (1000 µg/mL)	[264]
Malic acid/Fe3O4	TEM, 9.2	DOX	~79% release at pH 6.5 medium. ~22% release at pH 7.4 medium	Breast, cancer, MCF-7	65% cancer cell death without AMF	[265]
Magnetic nano/micro‑particles based on clinoptilolite‑type of natural zeolite	SEM, 75	Nr	Nr	Nr	The temperature in the MCZ injected chicken wing is increased from 27.5 °C to 32.1 °C in 5 min and to 39.7 °C in 22 min with AMF	[266]
Mn0.5Zn0.5Fe2O4@PEG nanofluid	TEM, 6.5	Nr	Nr	Retinal, cancer, ganglion	Controlling the recovery time during MHT affects the cell death rate	[267]
CuFe2O4	TEM, 17.3	Nr	Nr	Colon, cancer, HT-29	53% cancer cell death without AMF	[213]
Graphene oxide/ CoFe2O4	TEM, 5	Nr	Nr	Breast, cancer, MCF-7	42% and 70% cancer cell death without and with AMF, respectively	[268]
PVA-Mg-Co-Fe2O4	DLS, <120	5FU	Nr	Breast, cancer, MCF-7 and cervical, cancer, HeLa	>65% cancer cell death without AMF	[269]
Chitosan/Fe3O4	SEM, 125	5FU	45% release at pH 7.4 medium	Glioblastoma, cancer, A-172	25% cancer cell death with AMF and no significant elimination of normal cells fibroblasts	[207]
Fe3−δO4@silane and a thermoresponsive copolymer shell composed of 2-(2-methoxy)ethyl methacrylate and oligo(ethylene glycol)methacrylate moieties (1.14 mL)	TEM, 10	DOX	100% release after 52-hr at 42 °C and pH 7.4 medium. 62% release after 52-hr at 37 °C and pH 7.4 medium	Ovary, cancer, SKOV-3	~70% cancer cell death at 41 °C	[209]
Fe3−δO4@silane and a thermoresponsive copolymer shell composed of 2-(2-methoxy)ethyl methacrylate and oligo(ethylene glycol)methacrylate moieties (1 mL)	DLS, 55–80	DOX	100% release after 52-hr at 42 °C and pH 7.4 medium. 70% release after 52-hr at 37 °C and pH 7.4 medium	Nr	Nr	[209]
Graphene quantum dots/Fe3O4/SiO2	Nr	DOX	30% and 45% release at pH 5.0 medium with 37 °C and 50 °C, respectively	Breast, cancer, 4T1	18%, 80%, and 92% cancer cell death without, with AMF one, and two times, respectively. Chemotherapy, MHT, and combined therapy caused 40%, 65%, and 80% cancer cell death, respectively	[210]
Graphene quantum dots/Fe3O4/SiO2	TEM, 100	Nr	Nr	Breast, cancer, 4T1	44% and 52% cancer cell death without and with AMF, respectively.	[210]
Fe3O4/MamC/proteinAR-3 mAbs (IgG1)	TEM, 36	DOX	44% and 25% release at pH 7.4 medium with and without AMF. 6% and 2% at pH 5.0 medium with and without AMF	Colorectal, cancer, HT-29	7% and 18% cancer cell death without and with AMF, respectively	[211]
La0.7Sr0.3MnO3/PEG	TEM, 15–20	DOX	46% and 14% release at pH 5 and 7.4 medium without AMF, respectively	Breast, cancer, MCF-7	29% and 90% cancer cell death without and with AMF, respectively	[270]
Human serum protein/La0.7Sr0.3MnO3/PEG	Nr	DOX	39% and 12% release at pH 5 and 7.4 medium without AMF, respectively	Breast, cancer, MCF-7	16% and 79% cancer cell death without and with AMF, respectively	[270]
Alginate-chitosan-Fe3O4	TEM, ~14	DOX	22.5% and 0.2% release in tumor site with and without AMF, respectively	Breast, MCF-7	Chemotherapy, MHT, and combined therapy caused 85%, 63%, and 95% cancer cell death, respectively	[212]
ZIF-90 on polydopamine-coated Fe3O4	SEM, 200	DOX	17.3%, 70.8%, and 88.7% release at pH 7.4, 6.0, and 4.5 medium after 24 h, respectively	Cervical, cancer, HeLa	Chemotherapy, MHT, and combined therapy caused 32%, 50%, and 72% cancer cell death, respectively	[271]
Ferumoxytol-medical chitosan	Nr	DOX	~100% drug release at 43 °C and pH 5.2 medium	Colon, cancer, HT-29	Chemotherapy, MHT, and combined therapy caused 32%, 50%, and 72% cancer cell death, respectively	[272]
Lipid based Fe3O4	TEM, ~50	Temozolomide	61.3% and 100% at pH 7.4 and 4.5 medium after 7 days with AMF	Glioma, cancer, Uppsala 87	Chemotherapy, MHT, and combined therapy caused 10%, 44%, and 50% cancer cell death, respectively.	[273]
IONPs stabilized with trimethoxysilylpropyl-ethylenediamine triacetic acid as a carrier of DOX	TEM, 4.76	DOX	40% and 65% release within 2-hr at pH 7 and 5 medium, respectively	Mouse brain-derived microvessel endothelial, bEnd.3, Madin–Darby canine kidney transfected with multi-drug resistant protein 1 (MDCK-MDR1), and human U251 GBM cells	The combination of magnetic enhanced convective diffusion and the cadherin binding peptide for transiently opening the blood–brain barrier tight junctions are expected to enhance the efficacy of cell death (over 90%) within 48-hr of treatment using the DOX-EDT-IONPs	[274]
PEG coated NiFe2O4	TEM, 55	DOX	Release rate of 1.33% per min.	Nr	Nr	[275]
IONPs/PLGA fiber	TEM, 600 nm fiber and 20 nm IONPs	Bortezomib	18% and 70% release at 7.4 and 5.5 medium.	Fibroblast, cancer, NIH3T3 and mouse mammary carcinoma, 4T1	Many of the cells exhibited a shrunken morphology with membrane blebbing on their surface	[276]
Lectrospun chitosan/cobalt ferrite/titanium oxide nanofibers	SEM, 110	DOX	~85% and ~52% release at 5.3 and 7.4 medium with AMF. ~63% and ~40% release at 5.3 and 7.4 medium without AMF	Melanoma, cancer, B16F10	78% cancer cell death with AMF	[277]
Graphene Oxide-Fe3O4-PEG	TEM, ~20	DOX	~58% and ~19% release at 5 and 7.4 medium without AMF	Murine, cancer, colorectal, CT26	80% cancer cell death with AMF	[219]
PLGA/Fe3O4	TEM, 172.1	DOX	Nr	Colon, cancer, CT26	~80% cancer cell death with AMF	[220]
Mg0.13-Fe2O3	TEM, 7	Nr	Nr	Glioblastoma, cancer, U87MG	75% and 100% cancer cells death at a temperature of 48.4 °C and 63.5 °C with AMF, respectively	[278]
Ether triad modified core-shell magnetic mesoporous silica	TEM, 50–60	DOX	55.3%, 73.5%, and 85% release at pH 6.5, 5.0, and 4.0 medium, respectively, after 24-hr without AMF. 3.6%, 4.5% and 6.2% release at 25 °C, 37 °C, and 45 °C in pH 7.4 medium without AMF. 80%, 88%, and ~ 95% release at 25 °C, 37 °C, and 45 °C, respectively, at pH 7.4 medium with AMF.	Breast, cancer, MDA-MB-231	~5% and 77% cancer cell death without and with AMF, respectively	[279]
Fe3O4@PLA-grafted P (HEMA-co-MAA-co-NIPAAm-coTMSPM)	SEM, 96	DOX, Methotrexate	40% MTX and 75% DOX release at 41 °C in pH 4 medium after 6 days	Nr	Nr	[280]
Poly(Nisopropyl acrylamide-co-acrylic acid)-co-AAc)@mSiO2@Fe3O4	TEM, ~100	5FU	7.8% and 47% release after 20-hr with AMF at 37 °C and 45 °C, respectively	Nr	Nr	[89]
IONPs - (poly(maleic anhydride-alt-1-octadecene)- (tetramethylrhodamine 5(6)-carboxamide cadaverine - N-(3 dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride - 4-aminophenyl β-D glucopyranoside	TEM, ~11	Nr	Nr	3D pancreatic, cancer tumor, MIA PaCa-2	72% cancer cell death with AMF	[226]
Chitosan coated Fe3O4	TEM, 30–40	Nr	Nr	Lung, cancer, A549	50–70% cancer cell death with AMF	[228]
Graphene oxide - Fe3O4	TEM, 10–21	Nr	Nr	Lung, cancer, A549	80.5% cancer cell death with AMF	[85]
Silica coated lanthanum strontium manganese oxide	TEM, 35	Nr	Nr	Lung, cancer, A549	~75% cancer cell death without AMF	[229]
Chitosan-g-N-isopropylacrylamide coated magnetic-silica	TEM, 30–50	DOX	83.30% release after 5-hr with AMF at 45 °C and pH 4.0	Neck, cancer, Cervical	90.83% cancer cell death with AMF	[230]
Trimesic acid coated SPIONs dispersed in triethylene glycol	TEM, 9	Nr	Nr	Breast, cancer, MCF-7	90% cancer cell death with AMF	[231]
Tetraethyl orthosilicate coated La0.7Sr0.3MnO3	TEM, 45	Nr	Nr	Lung, cancer, A549	80% cancer cell death with AMF	[232]
Cetyltrimethylammonium bromide and polycaprolactone coated Fe3O4	TEM, 21	Nr	Nr	Liver, cancer, HepG2	~60% cancer cell death with AMF	[235]
CoFe2O4	TEM, ~10	Nr	Nr	Mouse fibroblast, normal, L929	15% normal cell death without AMF, showing high biocompatibility	[193]
Oleic acid/PEG coated CoFe2O4	TEM, ~10	Nr	Nr	Mouse fibroblast, normal, L929	0% normal cell death with AMF, showing high biocompatibility	[193]
Co0.03Mn0.28Fe2.7O4/SiO2	TEM, ~22	Nr	Nr	Osteosarcoma, normal, MG-63 and MEF	Less than 3% normal cell death without AMF, showing high biocompatibility	[198]
Contractible hydroxypropyl methyl cellulose/Fe3O4 hydrogel	TEM, 200	DOX	57.6% and 32.3% release with and without AMF in 24-hr at pH 7.4 medium. 78.8% and 41.7% with and without AMF at pH 5.5 medium	4T1 breast cancer	No tumor recurrences occurred in the chemo-thermal therapy group after 21 days of MHT	[207]
Fe3O4 NPs-silica-poly(Nisopropyl acrylamide-co-acrylic acid)	Nr	5FU	7.8% and 47% release after 20-hr at 37 °C and 45 °C, respectively	Nr	Nr	[89]
IONPs coated with graphite-like shell and labeled with Alexa 647 fluorescent marker	Nr	Nr	Nr	Nr	With an applied field of 38 kA/m at 980 kHz, tumors could be heated to 60 °C in 2 minutes, durably ablating them with millimeter (mm) precision, leaving surrounding tissue intact	[281]

An interesting MNCs was fabricated via using pH/thermo-sensitive polymer of poly(2-(dimethylamino)ethyl methacrylate) to coat MNPs for dual response in controlled drug delivery systems.²¹⁷ In a recent study,²³³ round-shaped SPIONs (~15 nm) (Figure 6a) was homogeneously coated with citric acid (2 nm thickness) abbreviated as SPION@CA (Figure 6b) for delivering DOX (SPION@CA-DOX) (Figure 6c) and epirubicin (SPION@CA-epirubicin) (Figure 6d). As presented in Figures 6c and d, drug loading did not damage the structure and morphology of the MNCs. In a study by Wang et al,²⁰⁴ DOX-Fe₃O₄@agar PMNCs (Figure 6e–k) with an average size of 9.2±3.2 nm (Figure 6e) and Ms of 41.9 emu/g (Figure 6f) showed a magnetocaloric temperature of 43 °C under AMF only after 5 min (Figure 6h), whereas it indicated a maximum drug release rate of 85±3% after 56 min at pH 7 to simulate the intestinal environment (Figure 6g). Figure 6i indicates the schematic illustration of DOX-Fe₃O₄@agar for eliminating HT-29 cancer cells under AMF that the red fluorescence was found in the image of HT-29 cell treated with DOX-Fe₃O₄@agar for 1-hr (Figure 6j). Then, the release of DOX caused 44% and 86.6% cancer cell death without and with applied AMF, respectively (Figure 6k). The SAR value of DOX-Fe₃O₄@agar increased considerably with increasing AMF strengths owing to the heating loss mechanisms, similar to different reports on Fe₃O₄^19,20 and polycaprolactone-coated Fe₃O₄,²³⁵ γ-Fe₂O₃,²⁸⁵ and CoMn IONPs.²⁸⁶ In another study,²⁸⁷ Parekh et al used a surfactant of the small-chain fatty acid-coated IONPs to trigger temperature sensitivity, and subsequently 75% of HeLa cancer cell death under 24-hr exposure to AMF. In a different study,⁸⁰ a novel and low-cost PMNCs were fabricated using Fe₃O₄ nanofillers supported on rice straw cellulose fibers in 5FU drug delivery systems. It showed the pH and thermo-sensitive drug release and increased selectivity and anticancer activities towards colorectal cancer cell lines in 2D monolayer and 3D tumor spheroid models under EMF and heat induction of 44.2 °C.⁸⁰ In conclusion, the use of polymers as coating or solid supports in MNCs is one of the key points shown by various studies on magnetic drug composites for possible clinical settings of MHT and magnetic targeting in future.

Figure 6 TEM images of (a) SPIONs, (b) SPION@CA, (c) SPION@CA-Dox, and (d) SPION@CA-epirubicin.²³³ (e) TEM image and the particle size distribution analysis of DOX-Fe₃O₄@agar, (f) Magnetic strength of Fe₃O₄ and DOX-Fe₃O₄@agar, (g) Drug release profiles of DOX-Fe₃O₄@agar at pH 7, (h) The SAR values and temperature achieved by Fe₃O₄, DOX-Fe₃O₄@agar, and DOX-Fe₃O₄@agar in HT-29 cells under applied AMF (applied field = 400 A, frequency = 250 kHz), (i) Schematic of effects of DOX-Fe₃O₄@agar for eliminating HT-29 cancer cells under hyperthermia, (j) fluorescence microscopy images of HT-29 cells after 1-hr of incubation with DOX-Fe₃O₄@agar, and (k) Cell survival of HT-29 cells treated by DOX-Fe₃O₄@agar with and without MHT.

Notes: The significance levels denote as * (p < 0.05), ** (p < 0.01), and *** (p < 0.001) indicate the degree of statistical significance when compared to the control group. (i, j, and k) Reprinted from Wang Y-J, Lin P-Y, Hsieh S-L, et al. Utilizing edible agar as a carrier for dual functional doxorubicin-Fe3O4 nanotherapy drugs. Materials. 2021;14(8):1824. Creative Commons.²⁰⁴

In vivo Studies on Magnetic Nanocomposites for Hyperthermia and Cancer Therapy

The therapeutic nature of MNCs with the arsenal of magnetothermal drug delivery systems has manifested several advantages over the conventional cancer therapy methods due to their low damage to normal cells and ability to deliver sufficient drug dosage to the tumor. Various studies have recommended the use of MHT and magnetic targeting as an efficient anti-tumoral therapy for in vivo studies using animal models. This can be done through intravenous injection of MNCs (with or without loaded-drug) in tumor site under EMF to guide MNCs onto the targeted tissue and then inducing AMF to selectively heat the tumor and control the drug release for effective elimination of cancer cells without harmful damages for normal cells. Table 6 summarizes the in vivo studies on different MNCs for MHT and targeted drug delivery systems.

Table 6 The in vivo Studies on MNCs for MHT and Targeted Drug Delivery

Magnetic Nanoagent	Type of Analysis, Size (nm)	Loaded Drug	Drug Release	Cell Lines	NP Administration Route, Animal Model	Results	Ref.
PLGA/Fe3O4	TEM, 172	DOX	Nr	CT26 colon cancer in 5 mice	Intratumoral injection, Mouse	The tumor size decreased 4 times with AMF	[220]
Commercially available “biocompatible” type of MNPs	TEM, 15	Nr	Nr	Nr	Injected intravenously via a tail vein, Mouse	By direct intratumoral injection of MNPs, tumor size decreased 60% in 14 days (amount of Fe ∼17 mg). In addition, 5 out of 5 mice were tumor free at 120 days. They had an initial tumor size of 1.5 cm (amount of Fe ∼6 mg + injections for retreatment).	[282]
Commercially available “biocompatible” type of MNPs	TEM, 11	Nr	Nr	OVCAR-3 ovarian tumor cells	Injected intravenously via a tail vein, Mouse	7 out of 9 (78%) tumor free after 160 days	[282]
Fluorescent IONPs/PLGA conjugated with human epidermal growth factor receptor 2	DLS, 524	Gemcitabine	23% release in 5 minutes at 440 kHz frequency (16.27 kA/m) and sustained drug release for 11 days	MIAPaCa-2 in SCID mice	Intravenous injection, Mouse	The tumor regression study at the end of 30 days showed significant tumor regression (86±3%) with MHT	[225]
Mg0.13-Fe2O3	TEM, 7	Nr	Nr	Hep3B xenografted animal models	Injected into the tumor directly, Mouse	It was clearly observed that the tumor was completely killed after 2 days with AMF.	[278]
Fe3O4/Poly(ethylene glycol) methyl ether	SEM, 73	Nr	Nr	Nude mice xenografted with breast cancer MCF-7	Injected into the tumor directly, Mouse	Following treatment, the tumor volumes were monitored for up to 40 days. For the control mice, tumor size increased 25 fold by the fortieth day. With AMF (1.2 × 109 A m^−1 s^−1), from the sixth day onwards, tumors were completely eliminated without recurrence within the experimental period	[283]
Biomimetic magnetic NPs mediated by magnetosome proteins	TEM, 36	DOX	100, 300, and 500 µg of BMNPs were exposed at (130 kHz and 18 kA m^−1) for 20 min	4T1 and MCF-7 in Balb/c female mice of about six weeks old	Tail-vein injection, Mouse	Around 40% of the tumor size decreased by MHT and chemotherapy, which was higher than chemotherapy alone.	[284]
Graphene oxide-cobalt ferrite	TEM, 5	Nr	Nr	MCF7	Injected in situ to the multiple points of the tumor, Mouse	BALB/c mice and the results indicated the smaller size of the tumor with the concentrations of 0.001 and 0.002 gr/mL and the respective magnetic frequencies of 400 and 250 kHz for 10 min after 27 days	[268]
IONPs-(poly(maleic anhydride-alt-1-octadecene)- (tetramethylrhodamine 5(6)-carboxamide cadaverine-N-(3 dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride-4-aminophenyl β-D glucopyranoside	TEM, ~11 DLS, ~85	Nr	Nr	Pancreatic tumor cell line (MIA PaCa-2)	Intratumoral injection, Rat tail	∼47% and ∼14% antitumor effects in heterotopic xenograft mouse model with and without AMF, respectively.	[226]

Thermo-chemosensitization means increasing the temperature for the enhancement of drug cytotoxicity to obtain advanced cancer treatment. Oltolina et al²⁸⁴ successfully encapsulated anticancer drug DOX onto biomimetic MNPs (BMNPs) via the succor of magnetosome proteins for both in vitro and in vivo MHT of breast cancer model (Figure 7a and b). From the TEM images at various time points, the interaction of BMNPs with 4T1 cell membrane internalized under applied magnetic field. In addition, DOX-BMNPs with AMF-induced hyperthermia decreased the size and weight of the tumor, proving that hyperthermia successfully enhanced targeted drug release to obtain potent antitumor actions.

Figure 7 (a) Interaction of BMNPs with 4T1 cells in the presence/absence of a continuous gradient magnetic field analyzed at TEM, micrographs of the cells incubated with the 100 µg/mL of BMNPs for different periods of time. The micrographs are representative of alternate serial cuts of the cell pellets of each sample. Scale bar, 5 µm. (b) In vivo antitumor activity of BMNPs under the influence of AMF, Images were captured using a thermic camera of a representative mouse without (left) and with (right) injected BMNPs during AMF treatment. Note the different colors within the circle on the backside of the mouse.²⁸⁴

In a study by Garanina et al,²⁸¹ hydrochloric acid treatment was used to coat the iron core NPs followed by additional layers of onion-like carbon coating (Fe@C core-shell NPs) to save the iron core from oxidation. With the presence of a constant magnetic field (150 mT), the magnetic characteristics of superparamagnetic Fe@C core-shell NPs and their endocytosis process through living tumor cells were analyzed. According to the results of correlative light and electron microscopy, the microstructure of Fe@C core-shell NPs was maintained even after internalization by the living cells. Alignment of internalized Fe@C core-shell NPs and their magneto controllability were achieved by exposure to the magnetic field and long-term observation using a confocal microscope revealed that the biocompatibility of the magnetic sample was attributed to carbon onion-like shells. Therefore, Fe@C core-shell NPs were safe without toxicity towards the cell physiology and did not trigger apoptosis.

Biodegradation of Magnetic Nanocomposites in the Body

The safety of MNCs in medical application is associated with i) short-term stability such as their blood circulation time and tissue or organ biodistribution and ii) long-term effects, including their toxicity, degradation, and clearance from the body.²⁸⁸ In the blood circulation, stability of MNCs can vary from minutes to several days and is characterized by their surface coating and the composition of the protein corona.^289,290 The excess MNCs mostly accumulated in the liver and spleen, and are removed from the bloodstream.^291,292 In regard to the long-term effects, factors such as the rate of metal ion release, iron scavenges from the iron metabolic protein compound, formation of ferritin via gradual transition of the particles, and the degradation effects for potentially de novo particle fabrication play the important roles.²⁹³ It was reported that MNCs with diverse magnetic core structure and counterparts have negative impact on the long-term pharmacokinetics.²⁸⁸

It should be noted that the human body’s iron content is presented in transferrin (0.1%), ferritin (15–30%), and myoglobin (4%), and within hemoglobin protein (65%).²⁹⁴ The intracellular degradation of IONPs has a comparable mechanism with those of ferritin. After the degradation of IONPs, the main iron-protein complexes of ferritin (hydrodynamic size ~13 nm) and transferrin support the storage and transport of the iron ions in the body, respectively.²⁹⁵ The lysosomal proteases cause the dissolving of the protein shell around ferritin and subsequently, the internal iron ions are degraded in the acidic condition of the lysosomes. This leads to the presence of extra iron in the body, which is regulated by the inherent clearance systems of the body. It was stated that the biodegradation of MNCs can be more attributed to the initial intracellular amount of MNPs compared to the coating itself.²⁹⁶ The coating and surface modification of MNCs advantageously decrease the water penetration to increase the degradation time in the macrophages. The size of MNCs may influence the surface chemistry, charge, and the bioavailability of the particles in the human body.^297–299 It was stated that the various drug nanocarriers such as MNCs with sizes between 10 and 100 nm trigger longer half lives in the blood and can also go through capillaries.³⁰⁰ Circulation of the plasma transferrin triggers the transfer of iron to different areas of the body. The liver and spleen are responsible for the clearance of NPs from the bloodstream.³⁰¹ Renal clearance might remove the particles with sizes below 5–7 nm,³⁰² however, particles with sizes above 5 microns may result in capillary blockage.³⁰³ It should be noted that the excess MNPs get accumulated in tissues including lung and adipose tissue, which may cause unwanted toxicity.³⁰⁴ It is still required to analyze the toxicity, biodistribution, biodegradation, and clearance of different new MNCs via short and long time evaluation for advanced anticancer drug delivery nanosystems.

Future Perspectives in the Use of Magnetic Nanocomposites

MNCs as multifunctional carrier systems can be externally guided by EMF and produce heat by AMF for the multi-stage and on-demand drug release procedures.^4,80 Figure 8a schematically shows the magneto, pH, and thermoresponsive drug delivery systems using drug-loaded MNCs. Figure 8b indicates the schematic of prospective use of drug-loaded MNCs for hyperthermia and magnetic targeted cancer therapy. Several studies have reported that the long-term accumulation of nanocarriers can lead to unwanted toxicity, health risks, and harmful side-effects such as oxidative stress, fibrosis, inflammation, genotoxicity, and subsequently cell death.³⁰⁵ Although MNPs may potentially penetrate the blood–brain barrier and generate the iron imbalance causing Alzheimer’s and Parkinson’s disease^41,42, they can be effectively removed through their high magnetization and guideability following the drug release in the body.^306,307 This can decrease the long-term NPs accumulation and unwanted damage to the human body.⁴⁰ To achieve this, there are several important considerations. First, a direct method can be applied using needles inserted into the target tumor for the administration of drug-loaded MNCs. Second, EMF can magnetize the tip of the needle to effectively direct and deliver the drug-loaded MNCs to the target tumor. Third, it is necessary to understand the performance and mechanisms of the drug release in the distributed tumor tissues and organs. Fourth, the use of magnetic targeting systems can clear MNCs from the body to minimise the side effects in the targeted tissue and body.³⁰⁸ In targeted therapy, particles penetrate tissues from capillaries, such that strong magnetic fields are needed to maintain MNCs in large arteries since the blood flow decreases the accumulation of magnetic carriers at the target tissue.⁴³ In particular, the velocity of blood flow in the capillaries (±0.5 cm/s) is 50–100 times lower than the linear velocity of blood in large arteries. A different matter related to magnetic carriers is obtaining the desired depth penetration of the magnetic field towards the target area. Since the magnetic field decreases with distance, it is challenging to transfer MNCs near the target areas within the body with depth more than 2 cm from the skin.⁴³ Importantly, the higher interstitial fluid pressure in tumors (10–30 mmHg) than that in the healthy tissue (1–3 mmHg)³⁰⁹ prevents convective flow from the vasculature into the interstitial tissue as can be limited even if the extracellular matrix of tumors has a pore size around hundreds of nanometers.^310,311 Therefore, MNCs penetration into the tumor mainly reflects the level of MNCs diffusion,³¹² and delivering the NPs onto the tumor is mostly limited to perivascular areas. Noticeably, without convection for heat transfer using the bulk motion of a fluid, the larger NPs move slowly over the extracellular matrix, causing ineffective tumor therapy. For instance, 100 nm micelles indicated a lack of tissue penetration and subsequently lower therapeutic effect than 30 nm micelles.³¹³ Thus, the efficiency of tumor therapy is strongly linked to increased NPs penetration into tumors, which can be obtained via using smaller NPs³¹⁴. It is worth to mention that bigger particles are more strongly attracted by EMF. Thus, in magnetically guided drug delivery systems, the size of MNCs should satisfy both tissue penetration efficiency and attraction to the magnet. Due to the transparency of body tissues to EMF, magnetophoresis is considered as a bioorthogonal probe for improving NPs accumulation and penetration into the tumor. It is reported that the magnetic effect of an applied single EMF only from one side would be quickly reduced with distance since the single EMF is effective for magnetophoresis with only less than a 5 mm distance from the tumor.³¹⁵ Deeper body tissues can be targeted by catheter-related magnetophoresis methods using steering coils to guide MNCs to reach the tumor vasculature. The underlying limitation of this strategy is its invasive procedure as the catheter must be physically steered to reach the tumor area.³¹⁶ This challenge has been tackled by using two oppositely polarized EMFs to enhance the accumulation and subsequence penetration of MNCs into the targeted solid tumors^316,317. The two oppositely EMFs polarized form a zero point at the middle of the configuration for delivering MNCs onto the solid tumors, even at the deep tissues. EMF can improve the accumulation of magnetoliposomes for an effective inflammation process. In addition, it can decrease the amount of iron located in the plasma, liver, and spleen of mice.

Figure 8 (a) Schematic of pH, magneto, and thermo-responsive drug delivery systems using drug-loaded MNCs. (b) The schematic of prospective use of drug-loaded MNCs for hyperthermia and magnetic targeted cancer therapy.^220,318

Due to the practical problems and difficult process, there have been very limited reports on the application of magnetic drug delivery systems in humans. An example of a human trial after successful animal experiments is a study on 14 patients with colon adenocarcinoma or hypernephroma.³²⁰ These patients were given epirubicin-loaded ferrofluids with a size below 100 nm via intravenous injection and guided through EMF (0.8 T). There was around 0.5 cm distance between EMF and the surface of the tumor. It was shown that 120 min of EMF exposure was sufficient to localize MNPs to obtain magnetic resonance spectroscopy (MRS) and histology. In another study, MHT in combination with both radiation and chemotherapy was accomplished for 22 patients affected by recurrent tumor entities,³²¹ of which all patients responded and tolerated the applied heat treatment well.

It was mentioned earlier that the carriers with single functionality could respond to only one external or internal drug release procedure. As can be understood from Figure 8, anticancer drug-loaded pH-sensitive MNCs under AMF and EMF can perform a desired multifunctional therapy of which the individual hyperthermia, magnetic targeting, and chemotherapy are incapable.

MNCs have magnetothermal responses to be utilized in multimodal drug delivery systems for targeted cancer therapy in an eco-friendly manner. Magnetic targeting combined with hyperthermia therapy can tackle most, if not all, of the issues related to single-modality cancer treatments. In multi-stage cancer studies, promising results were reported for the combination therapy using MNCs, hyperthermia, and tumor pH targeting. In addition to the high drug loading capacity and therapeutic efficacy of MNCs, the external magnetic field and alternating magnetic field can be used for remotely switchable on-demand drug release and precise steering of the magnetothermal effects for local ablation of cancer cells. This comprehensive review discusses and highlights the use of MNCs in hyperthermia and targeted cancer therapy. Future studies are anticipated to scale up and generate affordable, safe, target-specific and potent MNCs for anticancer drug delivery.

Disclosure

The authors report no conflicts of interest in this work.

Acknowledgments

This work was supported by Takasago Thermal Engineering Co. Ltd. Grant (R.K130000.7343.4B422) from the research management center (RMC) of Universiti Teknologi Malaysia (UTM) and Malaysia-Japan International Institute of Technology (MJIIT).