Ferritin nanocages for early theranostics of tumors via inflammation-enhanced active targeting.

Engineered nanocarriers have been widely developed for tumor theranostics. However, the delivery of imaging probes or therapeutic drugs to the tumor pre-formation site for early and accurate detection and therapy remains a major challenge. Here, by using tailor-functionalized human H-ferritin (HFn), we developed a triple-modality nanoprobe IRdye800-M-HFn and achieved the early imaging of tumor cells before the formation of solid tumor tissues. Then, we developed an HFn-doxorubicin (Dox) drug delivery system by loading Dox into the HFn protein cage and achieved early-stage tumor therapy. The intravenous injection of HFn nanoprobes enabled the imaging of tumor cells as early as two days after tumor implantation, and the triple-modality imaging techniques, namely, near-infrared fluorescence molecular imaging (NIR-FMI), magnetic resonance imaging (MRI), and photoacoustic imaging (PAI), ensured the accuracy of detection. Further exploration indicated that HFn could specifically penetrate into pre-solid tumor sites by tumor-associated inflammation-mediated blood vessel leakage, followed by effective accumulation in tumor cells by the specific targeting property of HFn to transferrin receptor 1. Thus, the HFn-Dox drug delivery system delivered Dox into the tumor pre-formation site and effectively killed tumor cells at early stage. IRDye800-M-HFn nanoprobes and HFn-Dox provide promising strategies for early-stage tumor diagnosis and constructive implications for early-stage tumor treatment.

Nanocages displaying SIRP gamma clusters combined with prophagocytic stimulus of phagocytes potentiate anti-tumor immunity.

Antigen-presenting cells (APCs), including macrophages and dendritic cells (DCs), play a crucial role in bridging innate and adaptive immunity; thereby, innate immune checkpoint blockade-based therapy is an attractive approach for the induction of sustainable tumor-specific immunity. The interaction between the cluster of differentiation 47 (CD47) on tumor and signal-regulatory protein alpha (SIRPα) on phagocytic cells inhibits the phagocytic function of APCs, acting as a "don't eat me" signal. Accordingly, CD47 blockade is known to increase tumor cell phagocytosis, eliciting tumor-specific CD8+ T-cell immunity. Here, we introduced a nature-derived nanocage to deliver SIRPγ for blocking of antiphagocytic signaling through binding to CD47 and combined it with prophagocytic stimuli using a metabolic reprogramming reagent for APCs (CpG-oligodeoxynucleotides). Upon delivering the clustered SIRPγ variant, the nanocage showed enhanced CD47 binding profiles on tumor cells, thereby promoting active engulfment by phagocytes. Moreover, combination with CpG potentiated the prophagocytic ability, leading to the establishment of antitumorigenic surroundings. This combination treatment could competently inhibit tumor growth by invigorating APCs and CD8+ T-cells in TMEs in B16F10 orthotopic tumor models, known to be resistant to CD47-targeting therapeutics. Collectively, enhanced delivery of an innate immune checkpoint antagonist with metabolic modulation stimuli of immune cells could be a promising strategy for arousing immune responses against cancer.

Ferritin nanocage: A promising and designable multi-module platform for constructing dynamic nanoassembly-based drug nanocarrier.

Ferritin has been widely recognized as an ideal drug delivery vehicle owing to its unique cage-like structure. Coupled with intrinsic targeting ability and excellent biosafety, ferritin-based drug delivery system, recently coined as ferritin drug carrier (FDC), has sparked great interest among researchers and shown promising application potential in the biomedical field. However, the flexibility and accuracy of traditional FDCs are limited when facing with complex disease microenvironments. To meet the fast-growing requirements for precision medicine, ferritin can serve as a designable multi-module platform to fabricate smarter FDC, which we introduce here as dynamic nanoassembly-based ferritin drug carrier (DNFDC). Compared to conventional FDC, DNFDCs directly integrate required functions into their nanostructure, which can achieve dynamic transformation upon stimuli to specifically activate and exert therapeutic functions at targeted sites. In this review, we summarize the superior characteristics of ferritin that contribute to the on-demand design of DNFDC and outline the current advances in DNFDC. Moreover, the potential research directions and challenges are also discussed here. Hopefully, this review may inspire the future development of DNFDC.

Nephron loss detected by MRI following neonatal acute kidney injury in rabbits.

BACKGROUND: Acute kidney injury affects nearly 30% of preterm neonates in the intensive care unit. We aimed to determine whether nephrotoxin-induced AKI disrupted renal development assessed by imaging (CFE-MRI). METHODS: Neonatal New Zealand rabbits received indomethacin and gentamicin (AKI) or saline (control) for four days followed by cationic ferritin (CF) at six weeks. Ex vivo images were acquired using a gradient echo pulse sequence on 7 T MRI. Glomerular number (Nglom) and apparent glomerular volume (aVglom) were determined. CF toxicity was assessed at two and 28 days in healthy rabbits. RESULTS: Nglom was lower in the AKI group as compared to controls (74,034 vs 198,722, p < 0.01). aVglom was not different (AKI: 7.3 × 10-4 vs control: 6.2 × 10-4 mm3, p = 0.69). AKI kidneys had a band of glomeruli distributed radially in the cortex that were undetectable by MRI. Following CF injection, there was no difference in body or organ weights except for the liver, and transient changes in serum iron, platelets and white blood cell count. CONCLUSIONS: Brief nephrotoxin exposure during nephrogenesis results in fewer glomeruli and glomerular maldevelopment in a unique pattern detectable by MRI. Whole kidney evaluation by CFE-MRI may provide an important tool to understand the development of CKD following AKI.

Ferritin drug carrier (FDC) for tumor targeting therapy.

Ferritin is an iron storage protein that plays a key role in iron homeostasis and anti-oxidation of cells. Due to its unique architecture of 24 self-assembling subunits and hollow cavity capable of encapsulating drugs, and an outer surface that can be modified genetically and chemically for additional functionality, ferritin has recently emerged as a promising drug delivery vehicle. Recent research demonstrated that unmodified human heavy chain ferritin binds to its receptor, transferrin receptor 1 (TfR1), in different types of tumor tissues, including lung and breast cancer, thus highlighting the potential use of ferritin for tumor-targeting applications. In this review, we consider the many favorable characteristics of ferritin drug carriers (FDCs) for tumor drug delivery. In particular, compared with antibody-drug conjugates (ADCs), ferritin exhibits superiority in a range of attributes, including drug loading ability, thermostability, and ease of production. Thus, the emergence of FDCs may be the next step in targeted cancer therapy.

Ferritin nanovehicle for targeted delivery of cytochrome C to cancer cells.

In this work, we have exploited the unique properties of a chimeric archaeal-human ferritin to encapsulate, deliver and release cytochrome c and induce apoptosis in a myeloid leukemia cell line. The chimeric protein combines the versatility in 24-meric assembly and cargo incorporation capability of Archaeglobus fulgidus ferritin with specific binding of human H ferritin to CD71, the "heavy duty" carrier responsible for transferrin-iron uptake. Delivery of ferritin-encapsulated cytochrome C to the Acute Promyelocytic Leukemia (APL) NB4 cell line, highly resistant to transfection by conventional methods, was successfully achieved in vitro. The effective liberation of cytochrome C within the cytosolic environment, demonstrated by double fluorescent labelling, induced apoptosis in the cancer cells.

Inlaying Radiosensitizer onto the Polypeptide Shell of Drug-Loaded Ferritin for Imaging and Combinational Chemo-Radiotherapy.

Rationale: Ferritin with unique hollow cavity is an emerging protein-based nanoplatform for anticancer-drug delivery, but the in vivo chemotherapeutic effectiveness is still unsatisfactory with such a monotherapy modality, which is urgently in need of improvement. Methods: Here a novel ferritin nanotheranostic with anticancer-drug doxorubicin encapsulated into its hollow interior and nanoradiosensitizer bismuth sulfide nanocrystals inlayed onto its polypeptide shell was synthesized for combinational therapeutic benefits. The formation mechanism of bismuth sulfide nanocrystals based on ferritin has been analyzed. The in vitro and in vivo treatment effects were carried out on HeLa cancer cells and tumor-bearing mice, respectively. The biocompatibility and excretion of the ferritin nanotheranostic have also been evaluated to guarantee their biosafety. Results: The polypeptide shell of ferritin provides nucleation sites for the bismuth sulfide nanocrystals through coordination interaction, and simultaneously inhibits the further growth of bismuth sulfide nanocrystals, rendering the bismuth sulfide nanocrystals like rivets inlaying onto the polypeptide firmly, which can not only strengthen the architectural stability of ferritin to prevent drug burst leakage during systemic circulation, but also act as excellent computed tomography contrast agents and nanoradiosensitizers for in vivo imaging-guided cancer combinational treatments. Conclusions: The design concept of inlaying bismuth sulfide nanocrystals onto the polypeptide shell of doxorubicin-encapsulated ferritin significantly inhibits the tumor growth and simultaneously further broadens the application of ferritin in nanomedicine.

Impacto de la anticoagulación y antiagregación plaquetaria en la anemia y los eventos hemorrágicos y ateroscleróticos de pacientes con enfermedad renal crónica en estadios 3 y 4 / Impact of anticoagulation and platelet antiaggregation on anaemia and haemorragic events in patients with chronic kidney disease stages 3 and 4

Introducción y objetivo: Existe controversia sobre el riesgo/beneficio de anticoagular/antiagregar a pacientes con enfermedad renal crónica (ERC). Analizamos el impacto de la anticoagulación/antiagregación en pacientes con ERC sobre el riesgo hemorrágico, cardiovascular y la mortalidad. Pacientes y métodos: Se estudió a 232 pacientes (81 controles, 91 anticoagulados y 60 antiagregados) con ERC en estadios 3 y 4, que fueron seguidos durante un tiempo medio de 33,7 ± 14,8 meses. Se recogieron eventos hemorrágicos, cardiovasculares y mortalidad. Resultados: La hemoglobina sérica y los niveles de ferritina fueron significativamente mayores en pacientes controles (hemoglobina 13,7 ± 1,6; 13,3 ± 1,8 y 12,7 ± 1,9g/dl; p = 0,004; ferritina 170 ± 145; 140 ± 138; 105 ± 99μg/l; p = 0,023). Durante el seguimiento hubo 36 eventos hemorrágicos: 4 en pacientes control, 23 en anticoagulados y 9 en antiagregados (log rank 12,5; p = 0,002). En un modelo de Cox ajustado para edad, función renal y niveles de hemoglobina, la anticoagulación aumentó el riesgo de sangrado 4veces (HR 4,180; 1,955-8,937; p = 0,001) y la antiagregación en casi 3veces (HR 2,780; 1,257-6,149; p = 0,012). Se registraron 64 eventos cardiovasculares, 21 de los cuales fueron clasificados como eventos ateroscleróticos: 10 en el grupo de antiagregación, 8 en el grupo control y 3 en el grupo de anticoagulación (log rank: 8,351; p = 0,015). El tratamiento anticoagulante demostró un efecto protector frente al riesgo de padecer eventos ateroscleróticos (HR 0,136; 0,033-0,551; p = 0,005), mientras que el tratamiento antiagregante no lo modificó (HR 1,566; 0,569-4,308; ns). Conclusiones: La anticoagulación y la antiagregación aumentan el riesgo hemorrágico en pacientes con ERC y empeoran la anemia. La anticoagulación disminuye el riesgo de eventos cardiovasculares ateroescleróticos en más de un 85% y la antiagregación no lo modifica

Background and objective: There is controversy concerning the risk/benefit of anticoagulation/antiaggregation in chronic kidney disease (CKD) patients. We analysed the impact of anticoagulation/antiaggregation on anaemia and haemorrhagic events in CKD patients. Patients and methods: A total of 232 CKD patients stages 3 and 4 were followed during a mean follow-up time of 36.7 ± 11.6 months: 81 patients did not receive any anticoagulation or antiaggregation treatment, 91 received anticoagulation treatment and 60 patients received platelet antiaggregation. Haemorrhagic and cardiovascular events were recorded. Results: Haemoglobin and ferritine levels were significantly higher in patients who did not receive anticoagulation or antiaggregation (Hb 13.7 ± 1.6, 13.3 ± 1.8 and 12.7±1.9g/dl, p=0.004; ferritine 170 ± 145, 140 ± 138, 105 ± 99μg/l, p=0.023). During follow up, 36 haemorrhagic events were registered: 4in the control group, 23 in the anticoagulation group and 9in the antiaggregation group (log rank 12.5; p=0.002). In a Cox model adjusted by age, renal function and haemoglobin levels, the anticoagulation increased the risk of bleeding by 4times (HR 4.180, 1.955-8.937); p=0,001) and antiaggregation by almost 3times (HR 2.780, 1.257-6.149, p=0.012). A total of 64 cardiovascular events were registered, 21 of which were classified as atherosclerotic events: 10 in the antiaggregation group, 8in the control group and 3in the anticoagulation group (log rank: 8.351; p=0.015). Anticoagulation treatment showed a reduction in the risk of atherosclerotic events (HR 0.136, 0.033-0.551, p=0.005) while platelet antiaggregation did not modified this risk (HR 1,566, 0.569-4.308). Conclusions: Anticoagulation and antiaggregation increase haemorrhagic risk in patients with CKD and worsen anaemia. Anticoagulation reduces atherosclerotic events by more than 85% while platelet antiaggregation does not modify this risk

Anemia in Heart Failure: Still Relevant?

One-third of all patients with heart failure have anemia, and its presence is associated with more symptoms, increased rates of hospitalization, and increased mortality. The etiology of anemia is multifactorial, complex, and varies between patients. The most important factors leading to anemia in heart failure are inadequate erythropoietin production resulting from renal failure, intrinsic bone marrow defects, medication use, and nutritional deficiencies such as iron deficiency. Erythropoiesis-stimulating agents (ESAs) have been proven to successfully correct hemoglobin levels, albeit without significant improvement in clinical outcome. On the contrary, the use of ESAs has led to increased rates of thromboembolic events and ischemic stroke. This use of ESAs for the treatment of anemia in heart failure, therefore, cannot be recommended. In addition, these results question whether anemia is a therapeutic target or merely a marker of disease severity. Other therapies are being studied and include agents targeting the erythropoietin receptor, hepcidin pathway, or iron availability. This review focuses on the pathophysiology of anemia in heart failure, explains why investigated therapies might not have led to the desired results, and discusses promising future therapies.

Measuring rat kidney glomerular number and size in vivo with MRI.

number is highly variable in humans and is thought to play an important role in renal health. Chronic kidney disease (CKD) is the result of too few nephrons to maintain homeostasis. Currently, nephron number can only be determined invasively or as a terminal assessment. Due to a lack of tools to measure and track nephron number in the living, the early stages of CKD often go unrecognized, preventing early intervention that might halt the progression of CKD. In this work, we present a technique to directly measure glomerular number ( Nglom) and volume in vivo in the rat kidney ( n = 8) using MRI enhanced with the novel contrast agent cationized ferritin (CFE-MRI). Adult male rats were administered intravenous cationized ferritin (CF) and imaged in vivo with MRI. Glomerular number was measured and each glomerulus was spatially mapped in 3D in the image. Mean apparent glomerular volume (a Vglom) and intrarenal distribution of the individual glomerular volume (IGV), were also measured. These metrics were compared between images of the same kidneys scanned in vivo and ex vivo with CFE-MRI. In vivo Nglom and a Vglom correlated to ex vivo metrics within the same kidneys and were within 10% of Nglom and a Vglom previously validated by stereologic methods. This is the first report of direct in vivo measurements of Nglom and a Vglom, introducing an opportunity to investigate mechanisms of renal disease progression and therapeutic response over time.

Dietary nonheme iron is equally bioavailable from ferritin or ferrous sulfate in thalassemia intermedia.

Transfusion-independent patients with thalassemia intermedia (TI) develop fatal iron overload from excessive iron absorption triggered by ineffective erythropoiesis. More information about iron pharmacokinetics and nonheme, dietary iron absorption in such patients is needed to optimize management. To obtain more information, different forms of supplemental nonheme iron sources (ferritin and ferrous sulfate) were compared in 4 TI (hemoglobin <9 g/dL) and 6 control (hemoglobin 12-16 g/dL) patients. Serial serum iron concentrations were measured during the 24 hours following consumption of 1 mg/kg of elemental iron as ferritin or ferrous sulfate. Serum iron concentrations were also measured for one TI patient and one control patient 2 hours after the ingestion of 2 mg/kg of dietary iron in ferritin or ferrous sulfate. Maximum serum iron concentrations were observed 4 hours after the consumption of either dietary iron source. However, the serum iron values were unchanged for either dietary iron source, even at the higher doses of consumed iron. Thus, the bioavailability of dietary iron, either as ferritin or ferrous sulfate, was equivalent in both groups of patients. The pilot data support ferritin as an alternative dietary iron supplement to ferrous sulfate. ABBREVIATIONS: CRP C-reactive protein; Hb hemoglobin; IDA iron-deficient anemia; ICP inductively coupled plasma; IE ineffective erythropoiesis; SCD sickle cell disease; sTf transferrin saturation; TI thalassemia intermedia; TIBC total iron binding capacity; TM thalassemia major; Tf transferrin.

Evaluation of DNA vaccines encoding M. Tb gene Bfrb and Mpt32 in mice model.

Tuberculosis (TB) is a life threatening infectious disease which is prevalent throughout the world. Mycobacterium bovis based Bacille Calmette-Gue´rin (BCG) is the only vaccine available against TB however, this (single) vaccine is not enough to eradicate it. Furthermore, numbers of researches from different parts of the World have shown its efficacy as variable. Hence other (better) vaccines like DNA vaccines are needed in addition to BCG in order to achieve desired goal of TB eradication. The current study was aimed to develop subunit based DNA vaccines against TB and to check their efficacy. Two constructs Bfrb-pND14 and Mpt32-pND14 were made and used as DNA vaccines. Endotoxin free DNA preparations were made and used in immunization studies. Twenty Balb/c female mice of age eight weeks were used in trial. Two experimental groups each comprising eight animals were used to inoculate Mpt32-pND14 and Bfrb-pND14 vaccines respectively. A group of four animals was used as negative control. Animals were bled through tail periodically and finally through cardiac puncture before euthanization. Antibodies were confirmed through dot blot and Agar Gel Immuno Diffusion test (AGID). All the animals immunized with both vaccines were found positive as tested through dot blot and AGID. The results of this study have indicated that both the M. tb genes have produced strong immune response in mice model through pND14 vector and proved themselves as good subunit based DNA vaccines.

Ferritin nanocage with intrinsically disordered proteins and affibody: A platform for tumor targeting with extended pharmacokinetics.

Ferritin nanocages are of particular interest as a novel platform for drug and vaccine delivery, diagnosis, biomineralization scaffold and more, due to their perfect and complex symmetry, ideal physical properties, high biocompatibility, low toxicity profiles as well as easy manipulation by genetic or chemical strategies. However, a short half-life is still a hurdle for the translation of ferritin-based nanomedicines into the clinic. Here, we developed a series of rationally designed long circulating ferritin nanocages (LCFNs) with 'Intrinsically Disordered Proteins (IDP)' as a stealth layer for extending the half-life of ferritin nanocages. Through predictions with 3D modelling, the LCFNs were designed, generated and their pharmacokinetic parameters including half-life, clearance rate, mean residence time, and more, were evaluated by qualitative and quantitative analysis. LCFNs have a tenfold increased half-life and overall improved pharmacokinetic parameters compared to wild-type ferritin nanocages (wtFN), corresponding to the low binding against bone marrow-derived macrophages (BMDMs) and endothelial cells. Subsequently, a tumor targeting moiety, epidermal growth factor receptor (EGFR)-targeting affibody peptide, was fused to LCFNs for evaluating their potential as a theragnostic platform. The tumor targeting-LCFNs successfully accumulated to the tumor tissue, by efficient targeting via active and passive properties, and also the shielding effect of IDP in vivo. This strategy can be applied to other protein-based nanocages for further progressing their use in the field of nanomedicine.

Drug delivery systems improving chemical and physical properties of anticancer drugs currently investigated for treatment of solid tumors.

Cancer is the second leading cause of death worldwide. Conventional cancer treatment like chemotherapy do not fulfil the expectations of both patients and physicians and there is a pressing need for a new kind of therapies that will increase drug delivery to the tumor mass. Standard chemotherapy does not show either specific tumor-targeting, or selective mode of action for cancer cells. Moreover, tumor microenvironments additionally disturb drug perfusion and diffusion. Currently approved anticancer drugs have many limitations and therefore special delivery systems improving their chemical and physical properties are beneficial. In the present review paper we discuss various drug delivery systems for solid tumors that are actually at various stages of pre-clinical tests or approved for therapy.

Protein cage nanostructure as drug delivery system: magnifying glass on apoferritin.

INTRODUCTION: New frontiers in nanomedicine are moving towards the research of new biomaterials. Apoferritin (APO), is a uniform regular self-assemblies nano-sized protein with excellent biocompatibility and a unique structure that affords it the ability to stabilize small active molecules in its inner core. Areas covered: APO can be loaded by applying a passive process (mainly used for ions and metals) or by a unique formulative approach based on disassemby/reassembly process. In this article, we aim to organize the experimental evidence provided by a number of studies on the loading, release and targeting. Attention is initially focused on the most investigated antineoplastic drug and contrast agents up to the most recent application in gene therapy. Expert opinion: Various preclinical studies have demonstrated that APO improved the potency and selectivity of some chemotherapeutics. However, in order to translate the use of APO into therapy, some issues must be solved, especially regarding the reproducibility of the loading protocol used, the optimization of nanocarrier characterization, detailed understanding of the final structure of loaded APO, and the real mechanism and timing of drug release.

Iron Metabolism in African American Women in the Second and Third Trimesters of High-Risk Pregnancies.

OBJECTIVE: To examine iron metabolism during the second and third trimesters in African American women with high-risk pregnancies. DESIGN: Longitudinal pilot study. SETTING: Large, university-based, urban Midwestern U.S. medical center. PARTICIPANTS: Convenience sample of 32 African American women with high-risk pregnancies seeking care at an urban maternal-fetal medicine clinic. METHODS: Nonfasting venous blood was collected in the second and third trimesters to assess iron status, hepcidin, and systemic inflammation. Anthropometric and survey data were obtained via self-report. Descriptive statistics were calculated from these data, and changes in the clinical parameters between the second and third trimesters were evaluated via paired t tests. Associations among demographic, reproductive, anthropometric, inflammatory, and iron-related parameters were also assessed in each trimester. RESULTS: The mean age of participants was 28.3 (± 6.8) years, and mean prepregnancy body mass index was 31.9 (± 10.7) kg/m2. In the longitudinal analysis, significant (p < .05) declines in serum iron, ferritin, transferrin saturation, and C-reactive protein were observed between the second and third trimesters. There was no statistically significant change in hepcidin between trimesters. When using a ferritin level cut-point of less than 15 ng/ml and soluble transferrin receptor level of greater than 28.1 nmol/L, 48% of the participants (14 of 29) were classified with iron deficiency in the third trimester. CONCLUSION: In this pilot study, iron deficiency was prevalent among a small cohort of African American women with high-risk pregnancies. Hepcidin concentrations were greater than previously reported in healthy, pregnant, primarily White women, which suggests decreased iron bioavailability in this high-risk group.

Biodegradable Magnetic Silica@Iron Oxide Nanovectors with Ultra-Large Mesopores for High Protein Loading, Magnetothermal Release, and Delivery.

The delivery of large cargos of diameter above 15nm for biomedical applications has proved challenging since it requires biocompatible, stably-loaded, and biodegradable nanomaterials. In this study, we describe the design of biodegradable silica-iron oxide hybrid nanovectors with large mesopores for large protein delivery in cancer cells. The mesopores of the nanomaterials spanned from 20 to 60nm in diameter and post-functionalization allowed the electrostatic immobilization of large proteins (e.g. mTFP-Ferritin, ~534kDa). Half of the content of the nanovectors was based with iron oxide nanophases which allowed the rapid biodegradation of the carrier in fetal bovine serum and a magnetic responsiveness. The nanovectors released large protein cargos in aqueous solution under acidic pH or magnetic stimuli. The delivery of large proteins was then autonomously achieved in cancer cells via the silica-iron oxide nanovectors, which is thus a promising for biomedical applications.

The Effectiveness of Ferritin as a Contrast Agent for Cell Tracking MRI in Mouse Cancer Models.

PURPOSE: We aimed to investigate the effectiveness of ferritin as a contrast agent and a potential reporter gene for tracking tumor cells or macrophages in mouse cancer models. MATERIALS AND METHODS: Adenoviral human ferritin heavy chain (Ad-hFTH) was administrated to orthotopic glioma models and subcutaneous colon cancer mouse models using U87MG and HCT116 cells, respectively. Brain MR images were acquired before and daily for up to 6 days after the intracranial injection of Ad-hFTH. In the HCT116 tumor model, MR examinations were performed before and at 6, 24, and 48 h after intratumoral injection of Ad-hFTH, as well as before and every two days after intravenous injection of ferritin-labeled macrophages. The contrast effect of ferritin in vitro was measured by MR imaging of cell pellets. MRI examinations using a 7T MR scanner comprised a T1-weighted (T1w) spin-echo sequence, T2-weighted (T2w) relaxation enhancement sequence, and T2*-weighted (T2*w) fast low angle shot sequence. RESULTS: Cell pellet imaging of Ad-hFTH in vitro showed a strong negatively enhanced contrast in T2w and T2*w images, presenting with darker signal intensity in high concentrations of Fe. T2w images of glioma and subcutaneous HCT116 tumor models showed a dark signal intensity around or within the Ad-hFTH tumor, which was distinct with time and apparent in T2*w images. After injection of ferritin-labeled macrophages, negative contrast enhancement was identified within the tumor. CONCLUSION: Ferritin could be a good candidate as an endogenous MR contrast agent and a potential reporter gene that is capable of maintaining cell labeling stability and cellular safety.

The Effectiveness of Ferritin as a Contrast Agent for Cell Tracking MRI in Mouse Cancer Models

PURPOSE: We aimed to investigate the effectiveness of ferritin as a contrast agent and a potential reporter gene for tracking tumor cells or macrophages in mouse cancer models. MATERIALS AND METHODS: Adenoviral human ferritin heavy chain (Ad-hFTH) was administrated to orthotopic glioma models and subcutaneous colon cancer mouse models using U87MG and HCT116 cells, respectively. Brain MR images were acquired before and daily for up to 6 days after the intracranial injection of Ad-hFTH. In the HCT116 tumor model, MR examinations were performed before and at 6, 24, and 48 h after intratumoral injection of Ad-hFTH, as well as before and every two days after intravenous injection of ferritin-labeled macrophages. The contrast effect of ferritin in vitro was measured by MR imaging of cell pellets. MRI examinations using a 7T MR scanner comprised a T1-weighted (T1w) spin-echo sequence, T2-weighted (T2w) relaxation enhancement sequence, and T2*-weighted (T2*w) fast low angle shot sequence. RESULTS: Cell pellet imaging of Ad-hFTH in vitro showed a strong negatively enhanced contrast in T2w and T2*w images, presenting with darker signal intensity in high concentrations of Fe. T2w images of glioma and subcutaneous HCT116 tumor models showed a dark signal intensity around or within the Ad-hFTH tumor, which was distinct with time and apparent in T2*w images. After injection of ferritin-labeled macrophages, negative contrast enhancement was identified within the tumor. CONCLUSION: Ferritin could be a good candidate as an endogenous MR contrast agent and a potential reporter gene that is capable of maintaining cell labeling stability and cellular safety.

Engineered Human Ferritin Nanoparticles for Direct Delivery of Tumor Antigens to Lymph Node and Cancer Immunotherapy.

Efficient delivery of tumor-specific antigens (TSAs) to lymph nodes (LNs) is essential to eliciting robust immune response for cancer immunotherapy but still remains unsolved. Herein, we evaluated the direct LN-targeting performance of four different protein nanoparticles with different size, shape, and origin [Escherichia coli DNA binding protein (DPS), Thermoplasma acidophilum proteasome (PTS), hepatitis B virus capsid (HBVC), and human ferritin heavy chain (hFTN)] in live mice, using an optical fluorescence imaging system. Based on the imaging results, hFTN that shows rapid LN targeting and prolonged retention in LNs was chosen as a carrier of the model TSA [red fluorescence protein (RFP)], and the flexible surface architecture of hFTN was engineered to densely present RFPs on the hFTN surface through genetic modification of subunit protein of hFTN. The RFP-modified hFTN rapidly targeted LNs, sufficiently exposed RFPs to LN immune cells during prolonged period of retention in LNs, induced strong RFP-specific cytotoxic CD8+ T cell response, and notably inhibited RFP-expressing melanoma tumor growth in live mice. This suggests that the strategy using protein nanoparticles as both TSA-carrying scaffold and anti-cancer vaccine holds promise for clinically effective immunotherapy of cancer.