Fabrication of Ciprofloxacin-Immobilized Calcium Phosphate Particles for Dental Drug Delivery.

Calcium phosphate (CaP) particles immobilizing antibacterial agents have the potential to be used as dental disinfectants. In this study, we fabricated CaP particles with immobilized ciprofloxacin (CF), a commonly prescribed antibacterial agent, via a coprecipitation process using a supersaturated CaP solution. As the aging time in the coprecipitation process increased from 2 to 24 h, the CaP phase in the resulting particles transformed from amorphous to low-crystalline hydroxyapatite, and their Ca/P elemental ratio, yield, and CF content increased. Despite the higher CF content, the particles aged for 24 h displayed a slower release of CF in a physiological salt solution, most likely owing to their crystallized matrix (less soluble hydroxyapatite), than those aged for 2 h, whose matrix was amorphous CaP. Both particles exhibited antibacterial and antibiofilm activities along with an acid-neutralizing effect against the major oral bacteria, Streptococcus mutans, Porphyromonas gingivalis, and Actinomyces naeslundii, in a dose-dependent manner, although their dose-response relationship was slightly different. The aging time in the coprecipitation process was identified as a governing factor affecting the physicochemical properties of the resulting CF-immobilized CaP particles and their functionality as a dental disinfectant.

In Situ Synthesis of a Tumor-Microenvironment-Responsive Chemotherapy Drug.

Current chemotherapy still suffers from unsatisfactory therapeutic efficacy, multi-drug resistance, and severe adverse effects, thus necessitating the development of techniques to confine chemotherapy drugs in the tumor microenvironment. Herein, we fabricated nanospheres of mesoporous silica (MS) doped with Cu (MS-Cu) and polyethylene glycol (PEG)-coated MS-Cu (PEG-MS-Cu) as exogenous copper supply systems to tumors. The synthesized MS-Cu nanospheres showed diameters of 30-150 nm with Cu/Si molar ratios of 0.041-0.069. Only disulfiram (DSF) and only MS-Cu nanospheres showed little cytotoxicity in vitro, whereas the combination of DSF and MS-Cu nanospheres showed significant cytotoxicity against MOC1 and MOC2 cells at concentrations of 0.2-1 µg/mL. Oral DSF administration in combination with MS-Cu nanospheres intratumoral or PEG-MS-Cu nanospheres intravenous administration showed significant antitumor efficacy against MOC2 cells in vivo. In contrast to traditional drug delivery systems, we herein propose a system for the in situ synthesis of chemotherapy drugs by converting nontoxic substances into antitumor chemotherapy drugs in a specific tumor microenvironment.

X-ray irradiation negatively affects immune responses in the lymphatic network.

Immune checkpoint inhibitor therapy has been attracting attention as a new cancer treatment and is likely to be widely used in combination with radiotherapy. Therefore, examination of the effects of X-ray irradiation on sentinel lymph nodes and lymphatic vessels, which are involved in antigen presentation, is important for therapy. The hindlimbs of mice were irradiated with X-rays (total radiation doses: 2, 10, and 30 Gy), and X-ray computed tomography (CT) imaging was performed using 15-nm or 2-nm gold nanoparticles (AuNPs) as contrast agents on days 7, 14, and 28 after irradiation to evaluate the diameter of the collecting lymph vessels and lymph flow within the irradiated area. X-ray CT imaging data using 15-nm AuNPs on day 28 after irradiation showed that the diameter of the collecting lymph vessels was significantly larger in all irradiated groups compared to the control group (p ≤ 0.01). CT imaging with 2-nm AuNPs showed that lymphatic drainage was significantly reduced in the lymph nodes irradiated with 10 Gy and 30 Gy compared to the lymph nodes irradiated with 2 Gy (p ≤ 0.05). Additionally, immunohistochemical analyses were conducted to evaluate the area density and morphology of high endothelial venules (HEVs) in the lymph nodes, which are important vessels for naive T cells to enter the lymph nodes. The expression level of MECA-79, which specifically localized to HEVs, was significantly decreased in the 10 Gy and 30 Gy irradiation groups compared to the control group (p ≤ 0.05). There was a significant decrease in normal HEV morphology (p ≤ 0.05) and a significant increase in abnormal HEV morphology (p ≤ 0.05) in all irradiated groups. These results also showed that X-ray irradiation induced a time- and radiation dose-dependent increase in the diameter of the collecting lymph vessels, stagnation of intralymphatic lymph flow, and a reduction in the area density of HEVs and their abnormal morphology, demonstrating that X-ray irradiation affected the immune responses. Therefore, these findings suggest that X-ray irradiation to lymph nodes may impair the opportunity for antigen presentation in the lymph nodes, which is the key to cancer immunity, and that for this reason, it is important to carefully plan irradiation of sentinel lymph nodes and develop treatment strategies according to future treatment options.

Development of X-ray contrast agents using single nanometer-sized gold nanoparticles and lactoferrin complex and their application in vascular imaging.

The technology to accurately image the morphology of tumor vessels with X-ray contrast agents is important to clarify mechanisms underlying tumor progression and evaluate the efficacy of chemotherapy. However, in clinical practice, iodine-based contrast agents present problems such as short blood retention owing to a high clearance ability and insufficient X-ray absorption capacity when compared with other high atomic number elements. To resolve these issues, gold nanoparticles (AuNPs), with a high atomic number, have attracted a great deal of attention as contrast agents for angiography, and have been employed in small animal models. Herein, we developed novel contrast agents using AuNPs and captured changes in tumor vessel morphology with time using X-ray computed tomography (CT). First, glutathione-supported single nanometer-sized AuNPs (sAu/GSH) (diameter, 2.2 nm) were fabricated using tetrakis(hydroxymethyl)phosphonium chloride as a reducing agent. The sAu/GSH particles were intravenously injected into mice, remained in vessels for a few minutes, and were then excreted by the kidneys after 24 h, similar to the commercial contrast agent iopamidol. Next, the Au/GSH and lactoferrin (sAu/GSH-LF) (long axis size, 17.3 nm) complex was produced by adding lactoferrin to the sAu/GSH solution under the influence of a condensing agent. On intravenously administering sAu/GSH-LF to mice, the blood retention time was 1-3 h, which was considerably longer than that observed with iopamidol and sAu/GSH. Moreover, we succeeded in imaging morphological changes in identical tumor vessels for several days using X-ray CT with sAu/GSH-LF.

The anti-angiogenic agent lenvatinib induces tumor vessel normalization and enhances radiosensitivity in hepatocellular tumors.

The evaluation of angiogenesis inhibitors requires the analysis of the precise structure and function of tumor vessels. The anti-angiogenic agents lenvatinib and sorafenib are multi-target tyrosine kinase inhibitors that have been approved for the treatment of hepatocellular carcinoma (HCC). However, the different effects on tumor vasculature between lenvatinib and sorafenib are not well understood. In this study, we analyzed the effects of both drugs on vascular structure and function, including vascular normalization, and investigated whether the normalization had a positive effect on a combination therapy with the drugs and radiation using micro X-ray computed tomography with gold nanoparticles as a contrast agent, as well as immunohistochemical analysis and interstitial fluid pressure (IFP) measurement. In mice subcutaneously transplanted with mouse HCC cells, treatment with lenvatinib or sorafenib for 14 days inhibited tumor growth and reduced the tumor vessel volume density. However, analysis of integrated data on vessel density, rates of pericyte-covering and perfused vessels, tumor hypoxia, and IFP measured 4 days after drug treatment showed that treatment with 3 mg/kg of lenvatinib significantly reduced the microvessel density and normalized tumor vessels compared to treatment with 50 mg/kg of sorafenib. These results showed that lenvatinib induced vascular normalization and improved the intratumoral microenvironment in HCC tumors earlier and more effectively than sorafenib. Moreover, such changes increased the radiosensitivity of tumors and enhanced the effect of lenvatinib and radiation combination therapy, suggesting that this combination therapy is a powerful potential application against HCC.

Fabrication of silica-coated gold nanorods and investigation of their property of photothermal conversion.

This study described the preparation of silica-coated Au nanorods (AuNR/SiO2) in a colloidal solution, assessed their property of photothermal conversion, and investigated their ability to kill cancer cells using photothermal conversion. Au-seed nanoparticles were produced by reducing hydrogen tetrachloroaurate (HAuCl4) with sodium borohydride (NaBH4) in aqueous n-hexadecyltrimethylammonium bromide (CTAB) solution. AuNRs were then fabricated by reducing HAuCl4 and silver nitrate (AgNO3) with l-ascorbic acid in the aqueous CTAB solution in the presence of Au-seed nanoparticles. The as-prepared AuNRs were washed by a process composed mainly of centrifugation to remove the CTAB. The washed AuNRs were coated with silica by mixing the AuNR colloidal solution, an aqueous solution of (3-aminopropyl)trimethoxysilane, and tetraethylorthosilicate/ethanol solution with a water/ethanol solution. We found that the addition of AuNR/SiO2 in water, in mice, and in a culture medium with cancer cells, followed by irradiation with a laser, cause an increase in temperature, demonstrating that AuNR/SiO2 have the ability of photothermal conversion. In addition, the cancer cells in the culture medium were found to be killed due to the increase in temperature caused by the photothermal conversion.