Ferritin: A Multifunctional Nanoplatform for Biological Detection, Imaging Diagnosis, and Drug Delivery.

Ferritins are spherical iron storage proteins within cells that are composed of a combination of 24 subunits of two types, heavy-chain ferritin (HFn) and light-chain ferritin (LFn). They autoassemble naturally into a spherical hollow nanocage with an outer diameter of 12 nm and an interior cavity that is 8 nm in diameter. In recent years, with the constantly emerging safety issues and the concerns about unfavorable uniformity and indefinite in vivo behavior of traditional nanomedicines, the characteristics of native ferritin nanocages, such as the unique nanocage structure, excellent safety profile, and definite in vivo behavior, make ferritin-based formulations uniquely attractive for nanomedicine development. To date, a variety of cargo molecules, including therapeutic drugs (e.g., cisplatin, carboplatin, paclitaxel, curcumin, atropine, quercetin, gefitinib, daunomycin, epirubicin, doxorubicin, etc.), imaging agents (e.g., fluorescence dyes, radioisotopes, and MRI contrast agents), nucleic acids (e.g., siRNA and miRNA), and metal nanoparticles (e.g., Fe3O4, CeO2, AuPd, CuS, CoPt, FeCo, Ag, etc.) have been loaded into the interior cavity of ferritin nanocages for a broad range of biomedical applications from in vitro biosensing to targeted delivery of cargo molecules in living systems with the aid of modified targeting ligands either genetically or chemically. We reported that human HFn could selectively deliver a large amount of cargo into tumors in vivo via transferrin receptor 1 (TfR1)-mediated tumor-cell-specific targeting followed by rapid internalization. By the use of the intrinsic tumor-targeting property and unique nanocage structure of human HFn, a broad variety of cargo-loaded HFn formulations have been developed for biological analysis, imaging diagnosis, and medicine development. In view of the intrinsic tumor-targeting property, unique nanocage structure, lack of immunogenicity, and definite in vivo behavior, human HFn holds promise to promote therapeutic drugs, diagnostic imaging agents, and targeting moieties into multifunctional nanomedicines.Since the report of the intrinsic tumor-targeting property of human HFn, we have extensively explored human HFn as an ideal nanocarrier for tumor-targeted delivery of anticancer drugs, MRI contrast agents, inorganic nanoparticles, and radioisotopes. In particular, by the use of genetic tools, we also have genetically engineered human HFn nanocages to recognize a broader range of disease biomarkers. In this Account, we systematically review human ferritins from characterizing their tumor-binding property and understanding their mechanism and kinetics for cargo loading to exploring their biomedical applications. We finally discuss the prospect of ferritin-based formulations to become next-generation nanomedicines. We expect that ferritin formulations with unique physicochemical characteristics and intrinsic tumor-targeting property will attract broad interest in fundamental drug research and offer new opportunities for nanomedicine development.

Thermal Atomization of Platinum Nanoparticles into Single Atoms: An Effective Strategy for Engineering High-Performance Nanozymes.

Although great progress has been made in artificial enzyme engineering, their catalytic performance is far from satisfactory as alternatives of natural enzymes. Here, we report a novel and efficient strategy to access high-performance nanozymes via direct atomization of platinum nanoparticles (Pt NPs) into single atoms by reversing the thermal sintering process. Atomization of Pt NPs into single atoms makes metal catalytic sites fully exposed and results in engineerable structural and electronic properties, thereby leading to dramatically enhanced enzymatic performance. As expected, the as-prepared thermally stable Pt single-atom nanozyme (PtTS-SAzyme) exhibited remarkable peroxidase-like catalytic activity and kinetics, far exceeding the Pt nanoparticle nanozyme. The following density functional theory calculations revealed that the engineered P and S atoms not only promote the atomization process from Pt NPs into PtTS-SAzyme but also endow single-atom Pt catalytic sites with a unique electronic structure owing to the electron donation of P atoms, as well as the electron acceptance of N and S atoms, which simultaneously contribute to the substantial enhancement of the enzyme-like catalytic performance of PtTS-SAzyme. This work demonstrates that thermal atomization of the metal nanoparticle-based nanozymes into single-atom nanozymes is an effective strategy for engineering high-performance nanozymes, which opens up a new way to rationally design and optimize artificial enzymes to mimic natural enzymes.

H-ferritin-nanocaged gadolinium nanoparticles for ultra-sensitive MR molecular imaging.

Rationale: Magnetic resonance imaging (MRI) is a powerful diagnostic technology by providing high-resolution imaging. Although MRI is sufficiently valued in its resolving morphology, it has poor sensitivity for tracking biomarkers. Therefore, contrast agents are often used to improve MRI diagnostic sensitivity. However, the clinically used Gd chelates are limited in improving MRI sensitivity owing to their low relaxivity. The objective of this study is to develop a novel contrast agent to achieve a highly sensitive tracking of biomarkers in vivo. Methods: A Gd-based nanoprobe composed of a gadolinium nanoparticle encapsulated within a human H-ferritin nanocage (Gd-HFn) has been developed. The specificity and sensitivity of Gd-HFn were evaluated in vivo in tumor-bearing mice and apolipoprotein E-deficient mice (Apoe-/-) by MRI. Results: The Gd-HFn probe shows extremely high relaxivity values (r1 = 549 s-1mM-1, r2 = 1555 s-1mM-1 under a 1.5-T magnetic field; and r1 = 428 s-1mM-1 and r2 = 1286 s-1mM-1 under a 3.0-T magnetic field), which is 175-fold higher than that of the clinically standard Dotarem (Gd-DOTA, r1 =3.13 s-1mM-1) under a 1.5-T magnetic field, and 150-fold higher under a 3.0-T magnetic field. Owing to the substantially enhanced relaxivity values, Gd-HFn achieved a highly sensitive tracking for the tumor targeting receptor of TfR1 and enabled the in vivo MRI visualization of tumors approaching the angiogenic switch. Conclusions: The developed Gd-HFn contrast agent makes MRI a more powerful tool by simultaneously providing functional and morphological imaging information, which paves the way for a new perspective in molecular imaging.

Cargo loading within ferritin nanocages in preparation for tumor-targeted delivery.

Ferritins are spherical iron storage proteins within cells, composed of 24 subunits of two types, heavy-chain ferritin (HFn) and light-chain ferritin. Ferritins auto-assemble naturally into hollow nanocages with an outer diameter of 12 nm and an interior cavity 8 nm in diameter. Since the intrinsic tumor-targeting property of human HFn was first reported in 2012, HFn has been extensively explored for tumor-targeted delivery of anticancer drugs and diagnostic molecules, including radioisotopes and fluorophores, as well as inorganic nanoparticles (NPs) and chemotherapeutic drugs. This protocol provides four detailed procedures describing how to load four types of cargoes within HFn nanocages that are capable of accurately controlling cargo loading: synthesis of inorganic metal nanoparticles within the cavity of a wild-type human HFn nanocage (Procedure 1, requires ~5 h); loading of doxorubicin into the cavity of a wild-type human HFn nanocage (Procedure 2, requires ~3 d); loading Gd3+ into the cavity of a genetically engineered human HFn nanocage (Procedure 3, requires ~20 h); and loading 64Cu2+ radioisotope into the cavity of a genetically engineered human HFn nanocage (Procedure 4, requires ~3 h). Subsequent use of these HFn-based formulations is advantageous as they have intrinsic tumor-targeting capability and lack immunogenicity. Human HFn generated as described in this protocol can therefore be used to deliver therapeutic drugs and diagnostic signals as multifunctional nanomedicines.

Biochemistry of mammalian ferritins in the regulation of cellular iron homeostasis and oxidative responses.

Ferritin, an iron-storage protein, regulates cellular iron metabolism and oxidative stress. The ferritin structure is characterized as a spherical cage, inside which large amounts of iron are deposited in a safe, compact and bioavailable form. All ferritins readily catalyze Fe(II) oxidation by peroxides at the ferroxidase center to prevent free Fe(II) from participating in oxygen free radical formation via Fenton chemistry. Thus, ferritin is generally recognized as a cytoprotective stratagem against intracellular oxidative damage The expression of cytosolic ferritins is usually regulated by iron status and oxidative stress at both the transcriptional and post-transcriptional levels. The mechanism of ferritin-mediated iron recycling is far from clarified, though nuclear receptor co-activator 4 (NCOA4) was recently identified as a cargo receptor for ferritin-based lysosomal degradation. Cytosolic ferritins are heteropolymers assembled by H- and L-chains in different proportions. The mitochondrial ferritins are homopolymers and distributed in restricted tissues. They play protective roles in mitochondria where heme- and Fe/S-enzymes are synthesized and high levels of ROS are produced. Genetic ferritin disorders are mainly related to the L-chain mutations, which generally cause severe movement diseases. This review is focused on the biochemistry and function of mammalian intracellular ferritin as the major iron-storage and anti-oxidation protein.

Nanozymes: A New Disease Imaging Strategy.

Nanozymes are nanomaterials with intrinsic enzyme-like properties. They can specifically catalyze substrates of natural enzymes under physiological condition with similar catalytic mechanism and kinetics. Compared to natural enzymes, nanozymes exhibit the unique advantages including high catalytic activity, low cost, high stability, easy mass production, and tunable activity. In addition, as a new type of artificial enzymes, nanozymes not only have the enzyme-like catalytic activity, but also exhibit the unique physicochemical properties of nanomaterials, such as photothermal properties, superparamagnetism, and fluorescence, etc. By combining the unique physicochemical properties and enzyme-like catalytic activities, nanozymes have been widely developed for in vitro detection and in vivo disease monitoring and treatment. Here we mainly summarized the applications of nanozymes for disease imaging and detection to explore their potential application in disease diagnosis and precision medicine.