RGD-Modified Apoferritin Nanoparticles for Efficient Drug Delivery to Tumors

Ferritin (FRT) is a major iron storage protein found in humans and most living organisms. Each ferritin is composed of 24 subunits, which self-assemble to form a cage-like nanostructure. FRT nanocages can be genetically modified to present a peptide sequence on the surface. Recently, we demonstrated that Cys-Asp-Cys-Arg-Gly-Asp-Cys-Phe-Cys (RGD4C)-modified ferritin can efficiently home to tumors through RGD–integrin α_vβ₃ interaction. Though promising, studies on evaluating surface modified ferritin nanocages as drug delivery vehicles have seldom been reported. Herein, we showed that after being precomplexed with Cu(II), doxorubicin can be loaded onto RGD modified apoferritin nanocages with high efficiency (up to 73.49 wt %). When studied on U87MG subcutaneous tumor models, these doxorubicin-loaded ferritin nanocages showed a longer circulation half-life, higher tumor uptake, better tumor growth inhibition, and less cardiotoxicity than free doxorubicin. Such a technology might be extended to load a broad range of therapeutics and holds great potential in clinical translation

Protein-Adsorbed Magnetic-Nanoparticle-Mediated Assay for Rapid Detection of Bacterial Antibiotic Resistance

Antibiotic susceptibility tests have been used for years as a crucial diagnostic tool against antibiotic-resistant bacteria. However, due to a lack of biomarkers specific to resistant types, these approaches are often time-consuming, inaccurate, and inflexible in drug selections. Here, we present a novel susceptibility test method named protein-adsorbed nanoparticle-mediated matrix-assisted laser desorption–ionization mass spectrometry, or PANMS. Briefly, we adsorb five different proteins (β-casein, α-lactalbumin, human serum albumin, fibrinogen, and avidin) onto the surface of Fe₃O₄. Upon interaction with bacteria surface, proteins were displaced from the nanoparticle surface, the amounts of which were quantified by matrix-assisted laser desorption ionization mass spectrometry. We find that the protein displacement profile was different distinctive among different bacteria strains and, in particular, between wild-type and drug-resistant strains. More excitingly, we observe bacteria resistant to drugs of the same mechanisms share similar displacement profiles on a linear discriminant analysis (LDA) map. This suggests the possibility of using PANMS to identify the type of mechanism behind antibiotic resistance, which was confirmed in a blind test. Given that PANMS is free of drug incubation and the whole procedure takes less than 50 min, it holds great potential as a high-throughput, low-cost, and accurate drug susceptibility test in the clinic

Noncovalent Ruthenium(II) Complexes–Single-Walled Carbon Nanotube Composites for Bimodal Photothermal and Photodynamic Therapy with Near-Infrared Irradiation

To enhance the efficacy and optimize the treatment of cancers, the integration of multimodal treatment strategies leading to synergistic effects is a promising approach. The coassembly of multifunctional agents for systematic therapies has received considerable interest in cancer treatment. Herein, Ru­(II) complex-functionalized single-walled carbon nanotubes (Ru@SWCNTs) are developed as nanotemplates for bimodal photothermal and two-photon photodynamic therapy (PTT-TPPDT). SWCNTs have the ability to load a great amount of Ru­(II) complexes (<b>Ru1</b> or <b>Ru2</b>) via noncovalent π–π interactions. The loaded Ru­(II) complexes are efficiently released by the photothermal effect of irradiation from an 808 nm diode laser (0.25 W/cm²). The released Ru­(II) complexes produce singlet oxygen species (¹O₂) upon two-photon laser irradiation (808 nm, 0.25 W/cm²) and can be used as a two-photon photodynamic therapy (TPPDT) agent. Based on the combination of photothermal therapy and two-photon photodynamic therapy, Ru@SWCNTs have greater anticancer efficacies than either PDT using Ru­(II) complexes or PTT using SWCNTs in two-dimensional (2D) cancer cell and three-dimensional (3D) multicellular tumor spheroid (MCTS) models. Furthermore, <i>in vivo</i> tumor ablation is achieved with excellent treatment efficacy under a diode laser (808 nm) irradiation at the power density of 0.25 W/cm² for 5 min. This study examines an efficacious bimodal PTT and TPPDT nanoplat form for the development of cancer therapeutics

Ferritin Nanocages To Encapsulate and Deliver Photosensitizers for Efficient Photodynamic Therapy against Cancer

Photodynamic therapy is an emerging treatment modality that is under intensive preclinical and clinical investigations for many types of disease including cancer. Despite the promise, there is a lack of a reliable drug delivery vehicle that can transport photosensitizers (PSs) to tumors in a site-specific manner. Previous efforts have been focused on polymer- or liposome-based nanocarriers, which are usually associated with a suboptimal PS loading rate and a large particle size. We report herein that a RGD4C-modified ferritin (RFRT), a protein-based nanoparticle, can serve as a safe and efficient PS vehicle. Zinc hexadeca­fluoro­phthalo­cyanine (ZnF₁₆Pc), a potent PS with a high ¹O₂ quantum yield but poor water solubility, can be encapsulated into RFRTs with a loading rate as high as ∼60 wt % (<i>i.e.</i>, 1.5 mg of ZnF₁₆Pc can be loaded on 1 mg of RFRTs), which far exceeds those reported previously. Despite the high loading, the ZnF₁₆Pc-loaded RFRTs (P-RFRTs) show an overall particle size of 18.6 ± 2.6 nm, which is significantly smaller than other PS–nanocarrier conjugates. When tested on U87MG subcutaneous tumor models, P-RFRTs showed a high tumor accumulation rate (tumor-to-normal tissue ratio of 26.82 ± 4.07 at 24 h), a good tumor inhibition rate (83.64% on day 12), as well as minimal toxicity to the skin and other major organs. This technology can be extended to deliver other metal-containing PSs and holds great clinical translation potential

Cumulative ascites volume, tumor weight and CA125 levels at the end of experiments after single MAb C595 and DTX treatment in OVCAR-3 animal model.

<p>Following euthanasia, the peritoneal cavity of each mouse was washed with 2 mL of normal saline, and after aspiration, the volume of ascites present was recorded. A. Cumulative volumes of ascites collected from each animal from initiation of therapy [MAb C595 (H and L), MAb IgG₃, DTX (H and L) are shown. The obvious difference was seen between DTX(H)-treated group and others-treated group (<i>P</i><0.05). B. Tumor weights (mg) at the end of experiments after single MAb C595, MAb IgG₃, DTX or vehicle treatments with different doses. The tumor weigh was obviously lower in MAb-treated and DTX-treated groups compared to MAbIgG₃-treated and HPMC-treated control groups (<i>P</i><0.05). C. Effect of single MAb C595, MAb IgG₃, DTX or vehicle on suppressing the increase in the tumor marker CA125 (CA125 Ku/L) in the ascites fluid (peritoneal wash) at the end of experiments. The level of CA125 was obviously lower in MAb C595 (H)-treated and DTX-treated groups compared to MAbIgG₃-treated and HPMC-treated control groups (<i>P</i><0.05). H: high dose; L: low dose. Representative graphs are shown.</p

The intensity of immunohistochemical staining of MUC1, CD31, Ki-67, TUNEL, Caspase-3 (Active) and PARP-1 (Cleaved p85) in tumor xenografts from combination test, combination control, MAb C595 control and vehicle control.

<p>All sections were prepared at the end of experiments.</p><p>Abbreviations: −, negative; +, weak; ++, moderate; +++, strong.</p><p>*indicates that obvious difference was found between combination test and controls.</p><p><b>A</b>: active; <b>C</b>: Cleaved p85.</p

Tumor weight and survival curve in combination test (control).

<p>A. Tumor weight changes at the end of experiments after combination test, combination control or vehicle treatments with different doses (<i>P</i><0.05). B. The tumor volume in combination test-treated mice was significantly lower than in vehicle control-treated mice (<i>P</i><0.01). C. For all animals, the intended duration of treatment was 4 weeks. Mice (10 per group) were euthanized if due to ill health, they were expected to become moribund within a short time. Survival was calculated as the number of days lapsed between initiation of treatment and euthanasia, and % mice surviving was the number of animals remaining in each group (×10) at the end of each week following initiation of treatment. The survival rate of animals in the combination test group was much better than that in the combination control group or vehicle group (<i>P</i><0.05). Representative images and graph are shown.</p

Representative images of Ki-67, TUNEL, Caspase-3 (Active) and PARP-1 (Cleaved 85) at the end of experiments after combination and control treatments.

<p>A. Very low Ki-67 expression is seen in the combination test; reduced Ki-67 expression in the combination control and MAb C595 control while high Ki-67 expression is seen in the vehicle control (<i>P</i><0.05). B. Obvious TUNEL-positive cells are shown in combination test; some TUNEL-positive cells are shown in the combination control and MAb C595 control while no TUNEL-positive cells are seen in the vehicle control (<i>P</i><0.05). C. High caspase-3 (active) expression is shown in combination test; low caspase-3 (active) expression is shown in combination control and MAb C595 control while negative caspase-3 (active) expression is shown in vehicle control (<i>P</i><0.05). D. High PARP-1 (cleaved p85) expression is shown in combination test; low PARP-1 (cleaved p85) expression is shown in combination control and MAb C595 control while negative PARP-1 (cleaved p85) expression is shown in vehicle control (<i>P</i><0.05). The brown color indicates nuclear staining in Ki-67, caspase-3 (active) and PARP-1 (cleaved p85), while blue indicates nuclear staining with hematoxylin. In the TUNEL assay, the brown color indicates nuclear chromatin condensation and fragmentation with methylgreen nuclear staining. Magnification ×20 in B; Magnification ×40 in A, C, D. Representative images are shown.</p

Cumulative ascites volume and CA125 levels at the end of experiments after combination test and combination control treatments.

<p>A. At the end of experiments, no obvious signs of ascites formation are seen in combination test (5 mg/kg MAb C595+10 mg/kg DTX)-treated mice (a); signs of ascites formation was found in combination control (5 mg/kg MAb C595+10 mg/kg DTX)-treated mice (b); obvious signs of ascites formation was found in vehicle (1/2 saline+1/2 HPMC)-treated mice (c). B. The cumulative ascites volumes from combination test, combination control or vehicle-treated mice are shown (<i>P</i><0.05). C. Effect of combination test, combination control or vehicle on suppressing the increase in tumor marker level (CA 125 Ku/L) in the ascites fluid (peritoneal wash) at the end of experiments (<i>P</i><0.05). Representative image and graphs are shown.</p

Dose-tolerance studies for escalating single-dose administration of single or combination treatments for the first 3 weeks in nude mice without tumors.

<p>Average percentage weight changes compared with day 0 (i.e., day of single or combination administration). A. Dose-tolerance relationship in mice by single MAb C595. ▪: MAb C595 (1 mg/kg); ▴: MAb C595 (5 mg/kg); ▾: MAb C595 (10 mg/kg); ♦: MAb C595 (15 mg/kg); •: MAb C595 (20 mg/kg). The significant difference was found between 20 mg/kg of MAb C595-treated group and other MAb C595-treated groups (5–15 mg/kg) (<i>P</i><0.05). B. Dose-tolerance relationship in mice by single DTX. ▪: DTX (3 mg/kg); ▴: DTX (5 mg/kg); ▾: DTX (7 mg/kg); ♦: DTX (10 mg/kg); •: DTX (15 mg/kg). The significant difference was found between 15 mg/kg of DTX-treated group and other DTX-treated groups (5–15 mg/kg) (<i>P</i><0.05). C. Dose-tolerance relationship in mice by combination test [MAb C595 (5 mg/kg)+DTX (3–10 mg/kg)]. ▪: MAb C595+DTX (3 mg/kg); ▴: MAb C595+DTX (5 mg/kg); ▾: MAb C595+DTX (7 mg/kg); ♦: MAb C595+DTX (10 mg/kg). D. Dose-tolerance relationship in mice by combination control [MAb IgG₃ control (5 mg/kg)+DTX (3–10 mg/kg)]. ▪: MAb IgG₃+DTX (3 mg/kg); ▴: MAb IgG₃+DTX (5 mg/kg); ▾: MAb IgG₃+DTX (7 mg/kg); ♦: MAb IgG₃+DTX (10 mg/kg). Points, mean (n = 5 in each group); bar, SD.</p