# Use of Ferritin-Based Metal-Encapsulated Nanocarriers as Anticancer Agents
## 1 Introduction  
Nanocarrier (NC) systems are suitable for several applications, including cell-specific or tissue-specific drug delivery, biomedical imaging and therapy [1; 2]. Many types of materials are used for the synthesis of NC: among these, special attention is given to protein cage structures, such as ferritins (Fts), because they are self-assembling, highly symmetrical proteins, with a strictly controlled size. Fts are easy to be expressed and purified at high levels and at low costs in _Escherichia coli_, and are exceptionally stable over wide ranges of temperature and pH. Fts are small enough to penetrate capillary spaces and large enough to avoid renal clearance. Notably, Fts are physiological materials, very soluble in aqueous solutions and in the blood, with low toxicity, susceptible to chemical modifications and being modified by molecular biology techniques [3; 4; 5; 6; 7; 8; 9; 10]. Materials such as Fe\({}_{3}\)O\({}_{4}\), Co\({}_{3}\)O\({}_{4}\), Mn\({}_{3}\)O\({}_{4}\), Pt, CoPt, Pd, CdS, CdSe, ZnSe, CaCO\({}_{3}\), SrCO\({}_{3}\), Au, Ag and BaCO\({}_{3}\) have been produced and characterized in different ferritin templates [11]. Fts are the main intracellular iron storage proteins in both prokaryotes and eukaryotes, and are large (480 kDa) complexes of 24 subunits, capable of containing as many as 4500 atoms of iron ions (Fe\({}^{3+}\)) within a hydrous ferric oxide core [12] (Figure 1). In mammals, two distinct classes of ferritin subunits exist, heavy (H) and light (L) chains, with molecular weights of about 21 kDa and 19 kDa, respectively, which share about 54% sequence identity. The H- and L-subunits have different functions,in that the L-subunit enhances the stability of the iron core, while the H-subunit is endowed with a ferroidase activity that is necessary for the rapid uptake of ferrous iron [13]. The H-subunit is very important for the organism because the absence of the H-ferritin (HFt) is embryonically lethal [14; 15]. In contrast, bacterial ferritins contain a single subunit type that resembles the H chains [16; 17]. HFt can deliver iron to many tissues, including the brain [18]. The brain imposes challenges to iron acquisition because of the highly developed tight junctions that bind neighbouring endothelial cells that make up the brain microvasculature. These junctions prevent the flux of molecules into the brain. The resulting blood-brain barrier (BBB) is a highly effective mechanism for protecting the brain from potentially harmful substances that circulate in the blood. A consequence of such a blockade, however, is that specific transport mechanisms must be designed for the trophic substances required for normal brain function. Transferrin and Fts are the main sources of iron for brain cells. Transferrin is considered the primary mechanism for cellular iron delivery to the brain, and a transferrin-mediated transport system has been identified in the BBB, based on transferrin receptors (TfRs, or CD71) [19; 20]. CD71 was identified in endothelial cells in culture and in rat brain microvasculature [21]. HFt also enters the cells via CD71: upon interaction with CD71, membrane invagination takes place, with the formation of Ft-containing early endosomes [22]. HFt is, therefore, an important means for transporting iron across the BBB: in fact, iron can be delivered to the brain also in hypotransferrinemic mice [23]. CD71 expression is highly dependent on cellular iron requirement, and is increased by 1-2 orders of magnitude in malignant, metastatic and/or drug resistant tumors (for a review, see [24]). This, together with the possibility to load Ft with high amount of chemotherapeutics, and the exceptional physico-chemical properties mentioned above, makes H-subunit Ft-based nanocarriers (HFt-NCs) ideal vehicles for drug delivery to cancers [4; 6]. Our group has recently synthesized Ft-based nanoparticles (NPs) designed to carry large amounts of drugs [25]. These molecules are able to bind metals and have been used against leishmaniasis [11; 26].