Speciation analysis of inorganic selenium in wastewater using a highly selective cellulose-based adsorbent via liquid electrode plasma optical emission spectrometry.

Speciation of selenium (Se) is typically carried out using a sophisticated technique such as ICP-MS after preconcentration using an adsorbent; however, the separation and preconcentration of inorganic Se has not been realized in the solutions containing high concentrations of SO42-. A dithiocarbamate-modified cellulose (DMC) was used in this study for the selective extraction and preconcentration of inorganic Se in wastewater, with a portable liquid electrode plasma-optical emission spectrometry (LEP-OES) being employed for quantification. DMC was found to selectively and quantitatively adsorb selenite (SeIV) over a wide range of pH (1.0-8.0); however, less than 3.0% of selenate (SeVI) was adsorbed in a pH range of 3.0-11. Quantitative extraction of SeIV was achieved even in the presence of 3.5 mol L-1 SO42-. The maximum sample volume from which 10 mg of DMC could quantitatively extract SeIV was found to be 500 mL. KOH (0.60 mL, 1.5 mol L-1) was found to quantitatively desorb SeIV retained on the adsorbent and yielded an enrichment factor of 833. The recovery of Se species from synthetic flue-gas desulfurization wastewater containing SeIV and SeVI at concentrations of 5.0 µmol L-1 was 96.2 ± 1.8% and 105.8 ± 1.8%, respectively.

Cancer neovasculature-targeted near-infrared photoimmunotherapy (NIR-PIT) for gastric cancer: different mechanisms of phototoxicity compared to cell membrane-targeted NIR-PIT.

BACKGROUND: Near-infrared photoimmunotherapy (NIR-PIT) constitutes a new class of molecular-targeted theranostics utilizing monoclonal antibody (mAb)-photosensitizer conjugates and NIR light. In this study, we developed a new type of NIR-PIT targeting vascular endothelial growth factor receptor 2 (VEGFR-2) expressed on vascular endothelium in an experimental gastric cancer model and evaluated the feasibility by comparing conventional NIR-PIT targeting cancer cell membrane in vitro and in vivo. METHODS: HER2-positive human gastric cancer cells, NCI-N87, were used for the experiments. Anti-HER2 mAb, trastuzumab and anti-VEGFR-2 mAb, DC101 were conjugated to photosensitizer, IR700. Phototoxicity in response to NIR-PIT were investigated in vitro and in vivo. Microvessel densities, as an indicator of angiogenesis, were counted in harvested xenografts after NIR-PIT to elucidate the mechanism. RESULTS: DC101-IR700 did not induce phototoxic effect in vitro because of the absence of expression of VEGFR-2 in NCI-N87 cancer cells. However, it induced an antitumor effect in NCI-N87 xenograft tumors accompanied with damage in tumor neovasculature as determined by decreasing tumor microvessel density, which represents a different mechanism than that of conventional NIR-PIT targeting antigens expressed on the tumor cell membrane. CONCLUSION: We demonstrated a new approach of NIR-PIT utilizing a target on vascular endothelium, such as VEGFR-2, and this treatment might lead to the development of a new therapeutic strategy for human gastric cancer.

Photoimmunotherapy targeting biliary-pancreatic cancer with humanized anti-TROP2 antibody.

Photoimmunotherapy (PIT) is a new type of tumor-specific treatment utilizing monoclonal antibody (mAb)-photosensitizer conjugates and near-infrared (NIR) light irradiation. One potential PIT target, the type I transmembrane protein TROP2, is expressed at high levels in many cancers, including pancreatic carcinoma (PC) and cholangiocarcinoma (CC), in which its expression is correlated with poor prognosis and tumor aggressiveness. In this study, we assessed the efficacy of PIT utilizing newly developed humanized anti-TROP2 mAb conjugated to the photosensitizer IR700 (TROP2-IR700) for PC and CC. Immunohistochemistry on PC and CC tissue microarrays confirmed that TROP2 is overexpressed in about half of PC and CC specimens. Using cultured PC and CC cells, TROP2-IR700 localized TROP2-specific and target-specific cell killing was observed after NIR light irradiation. In addition, TROP2-IR700 was localized to mouse xenograft tumors expressing TROP2 after intravenous injection. PC and CC xenograft tumor growth was significantly inhibited by TROP2-targeted PIT relative to controls. The efficacy of TROP2-targeted PIT in vitro and against xenografted tumors in vivo suggests promise as a therapy for human PC and CC, both of which currently have dismal prognoses and limited therapeutic options.

Near-Infrared Photochemoimmunotherapy by Photoactivatable Bifunctional Antibody-Drug Conjugates Targeting Human Epidermal Growth Factor Receptor 2 Positive Cancer.

Near-infrared photoimmunotherapy (NIR-PIT) is a new class of molecular targeted cancer therapy based on antibody-photoabsorber conjugates and NIR light irradiation. Recent studies have shown effective tumor control, including that of human epidermal growth factor receptor 2 (HER2)-positive cancer, by selective molecular targeting with NIR-PIT. However, the depth of NIR light penetration limits its use. Trastuzumab emtansine (T-DM1) is an antibody-drug conjugate consisting of the monoclonal antibody trastuzumab linked to the cytotoxic agent maytansinoid DM1. Here, we developed bifunctional antibody-drug-photoabsorber conjugates, T-DM1-IR700, that can work as both NIR-PIT and chemoimmunotherapy agents. We evaluated the feasibility of T-DM1-IR700-mediated NIR light irradiation by comparing the in vitro and in vivo cytotoxic efficacy of trastuzumab-IR700 (T-IR700)-mediated NIR light irradiation in HER2-expressing cells. T-IR700 and T-DM1-IR700 showed almost identical binding to HER2 in vitro and in vivo. Owing to the presence of internalized DM1 in the target cells, NIR-PIT using T-DM1-IR700 tended to induce greater cytotoxicity than that of NIR-PIT using T-IR700 in vitro. In vivo NIR-PIT using T-DM1-IR700 did not show a superior antitumor effect to NIR-PIT using T-IR700 in subcutaneous small-tumor models, which could receive sufficient NIR light. In contrast, NIR-PIT using T-DM1-IR700 tended to reduce the tumor volume and showed significant prolonged survival compared to NIR-PIT using T-IR700 in large-tumor models that could not receive sufficient NIR light. We successfully developed a T-DM1-IR700 conjugate that has a similar immunoreactivity to the parental antibody with increased cytotoxicity due to DM1 and potential as a new NIR-PIT agent for targeting tumors that are large and inaccessible to sufficient NIR light irradiation to activate the photoabsorber IR700.

Effect of L-Carnitine in Patients With Liver Cirrhosis on Energy Metabolism Using Indirect Calorimetry: A Pilot Study.

BACKGROUND: L-carnitine supplementation has been suggested to show several favorable effects on patients with liver cirrhosis (LC). However, there have been no reports regarding the effect of L-carnitine on energy metabolism in patients with LC using indirect calorimetry which is a well-established method for assessing the degree of liver malnutrition. We examined the effect of L-carnitine in patients with LC on energy metabolism using indirect calorimetry. METHODS: A total of 13 LC patients who are scheduled to be treated with L-carnitine (1,800 mg/day) were analyzed in this study. None of the patients previously received L-carnitine. An evaluation of the nutritional status was performed at the initiation of L-carnitine therapy and after 4 weeks of L-carnitine therapy. We evaluated the effect of L-carnitine on the nutritional status and energy metabolism by comparing various clinical variables at these two time points. In addition, the changes in the nutritional status of the patients were also evaluated using indirect calorimetry. RESULTS: After 4 weeks of L-carnitine treatment, for all cases, the mean substrate oxidation rates of carbohydrate (%C) increased from 37.6% to 48.2%, the mean substrate oxidation rates of fat (%F) decreased from 40.2% to 31.9% and the mean substrate oxidation rates of protein (%P) decreased from 22.2% to 19.9%. In a subgroup analysis of patients with baseline non-protein respiratory quotient (npRQ) < 0.85, the mean %C increased from 15.3% to 34.2%, the mean %F decreased from 59.9% to 45.1%, and the mean %P decreased from 24.8% to 20.6%. After 4 weeks of L-carnitine treatment, for all cases (n = 13), the mean value of npRQ increased in comparison with the baseline levels, although the difference was not significant (0.868 ± 0.060 vs. 0.838 ± 0.097, P = 0.19). Conversely, in patients with baseline npRQ < 0.85, the npRQ value significantly increased after 4 weeks treatment of L-carnitine compared with the baseline levels (0.827 ± 0.030 vs. 0.760 ± 0.043, P = 0.016). CONCLUSION: L-carnitine supplementation can be useful for improving energy metabolism, especially in patients who have an advanced LC status and lower baseline npRQ values.

Combination photoimmunotherapy with monoclonal antibodies recognizing different epitopes of human epidermal growth factor receptor 2: an assessment of phototherapeutic effect based on fluorescence molecular imaging.

Photoimmunotherapy is a new class of molecular targeted cancer therapy based on a monoclonal antibody (mAb) conjugated to a photosensitizer and irradiation with near-infrared (NIR) light for both imaging and therapy. Here, we sought to determine the feasibility of combining photoimmunotherapy using conjugates of human epidermal growth factor receptor 2 (HER2)-specific mAb-photosensitizer IR700, trastuzumab-IR700 and pertuzumab-IR700. HER2-expressing and non-expressing cells were treated with mAb-IR700 conjugates and irradiated with NIR light. Fluorescence imaging and cytotoxic effects were examined in cultured HER2-expressng cancer cell lines and in a mouse tumor xenograft model. Trastuzumab-IR700 and pertuzumab-IR700 could specifically bind to HER2 without competing, and the combination treatment of both agents yielded stronger HER2-specific IR700 fluorescence signals than with the treatment with either agent singly. A cytotoxicity assay showed that the combination treatment of both trastuzumab-IR700 and pertuzumab-IR700 followed by NIR light irradiation induced stronger cytotoxic effect than with treatment of either agent plus NIR light irradiation. Furthermore, the phototoxic and cytotoxic effects of mAb depended on HER2-specific IR700 signal intensities. Consistent with in vitro studies, in xenograft tumor models also, IR700 fluorescence imaging-guided NIR light irradiation after the combination treatment of trastuzumab-IR700 and pertuzumab-IR700 led to stronger antitumor effects than by treatment with either agent followed by NIR light irradiation. In conclusion, fluorescence molecular imaging can facilitate the assessment of treatment outcomes of molecular targeted photoimmunotherapy, which holds great potential in facilitating better outcomes in cancer patients.

[Two cases of successful sorafenib retreatment with the addition of steroid therapy following sorafenib-induced erythema multiforme in two patients with hepatocellular carcinoma].

Erythema multiforme (EM) is a known side effect of sorafenib therapy in cancer patients; at onset, the causative medication should be permanently discontinued. Here we report two cases of hepatocellular carcinoma (HCC) that developed sorafenib-induced EM. In both cases, retreatment with sorafenib combined with steroid therapy achieved effective tumor control without EM recurrence. The first patient was a 72-year-old woman who showed a dramatic response to sorafenib retreatment, with complete remission after 8 months of therapy. There was no rash recurrence after the steroid dose was gradually tapered and stopped. The second patient was a 69-year-old man who responded to sorafenib and exhibited stable disease, with no recurrence of the rash after the steroid dose was tapered. However, mild hand-foot syndrome persisted throughout sorafenib therapy. Although sorafenib should be discontinued if EM occurs, if there is no suitable alternative treatment, retreatment may be considered with steroid cover in patients with unresectable HCC.

Changes in plasma vascular endothelial growth factor at 8 weeks after sorafenib administration as predictors of survival for advanced hepatocellular carcinoma.

BACKGROUND: A new predictive biomarker for determining prognosis in patients with hepatocellular carcinoma (HCC) who receive sorafenib is required, because achieving a reduction in tumor size with sorafenib is rare, even in patients who have a favorable prognosis. Vascular endothelial growth factor (VEGF) receptor is a sorafenib target. In the current study, the authors examined changes in plasma VEGF concentrations during sorafenib treatment and determined the clinical significance of VEGF as a prognostic indicator in patients with HCC. METHODS: Plasma VEGF concentrations were serially measured in 63 patients with advanced HCC before and during sorafenib treatment. A plasma VEGF concentration that decreased >5% from the pretreatment level at 8 weeks was defined as a "VEGF decrease." An objective tumor response was determined using modified Response Evaluation Criteria in Solid Tumors 1 month after the initiation of therapy and every 3 months thereafter. RESULTS: Patients who had a VEGF decrease at week 8 (n=14) had a longer median survival than those who did not have a VEGF decrease (n=49; 30.9 months vs 14.4 months; P=.038). All patients who had a VEGF decrease survived for >6 months, and the patients who had both a VEGF decrease and an α-fetoprotein response (n=6) survived during the observation period (median, 19.7 months; range, 6.5-31.0 months). In univariate analyses, a VEGF decrease, radiologic findings classified as progressive disease, and major vascular invasion were associated significantly with 1-year survival; and, in multivariate analysis, a VEGF decrease was identified as an independent factor associated significantly with survival. CONCLUSIONS: A plasma VEGF concentration decrease at 8 weeks after starting sorafenib treatment may predict favorable overall survival in patients with advanced HCC.

Extracts of Larix Leptolepis effectively augments the generation of tumor antigen-specific cytotoxic T lymphocytes via activation of dendritic cells in TLR-2 and TLR-4-dependent manner.

Type-1 immunity plays a crucial role in host defense against various tumors and infectious diseases. Here, we first demonstrated that extract of Larix Leptolepis (ELL), one of the most popular timbers at Hokkaido area in Japan, strongly activated Type-1 immunity. ELL induced production of Type-1 cytokines such as IL-12 and TNF-α from bone marrow-derived dendritic cells (BMDCs) in TLR2- and TLR4-dependent manner and remarkably up-regulated the expression of MHC and co-stimulatory molecules. In addition, antigen-specific CTLs were significantly augmented by the combined administration of ELL, antigen and BMDCs. Finally, we revealed that combination therapy using ELL, antigen and BMDCs significantly inhibited the growth of established tumor in mouse model. Thus, these findings suggested that ELL would be a novel adjuvant for inducing an activation of Type-1-dependent immunity including activation of BMDCs and induction of tumor-specific CTLs, which is applicable to the therapy of cancer and infectious diseases.

Toll-like receptor-dependent IL-12 production by dendritic cells is required for activation of natural killer cell-mediated Type-1 immunity induced by Chrysanthemum coronarium L.

Type-1 immunity has an essential role for our host defenses against cancer and outer pathogens such as bacteria and virus. We demonstrated here that the edible plant extract of Chrysanthemum coronarium L. (C. coronarium) remarkably activates Type-1 immunity in a Toll-like receptor (TLR)2-, TLR4-, and TLR9-dependent manner. In the present experiments, the extract of C. coronarium significantly induces interferon (IFN)-γ production by mouse spleen cells. In addition, the IFN-γ production by spleen cells was completely blocked by the addition of anti-Interleukin (IL)-12 monoclonal antibodies. We confirmed that NK1.1(+) natural killer (NK) cells, NKT cells, and CD11c(+) dendritic cells (DC) were immediately activated after the stimulation with the extract of C. coronarium and the IFN-γ production was abolished in NK1.1(+) cell-depleted spleen cells. The stimulation with the extract of C. coronarium caused DC maturation involving with up-regulations of surface expression levels of MHC class I, MHC class II, CD40, and CD86 as well as induction of IL-12 production. The IFN-γ production induced by the extract was significantly reduced in the spleen cells depleted CD11c(+) cells. Furthermore, the IFN-γ production after the stimulation was strongly reduced in TLR4- and partially in TLR2- and TLR9-deficient spleen cells. Thus, we demonstrated the cellular mechanism for the activation of Type-1 immunity via NK cells, NKT cells, and DC by the extract of C. coronarium. These findings strongly suggest that C. coronarium would be a promising immuno-improving adjuvant, which might be useful for prevention of infectious, cancer, and allergic diseases through the activation of Type-1 immunity.

The extract of Japanese soybean, Kurosengoku activates the production of IL-12 and IFN-γ by DC or NK1.1(+) cells in a TLR4- and TLR2-dependent manner.

During the search for immuno-improving foods, we found that a variety of the Japanese soybean, Glycine max cv. Kurosengoku (Kurosengoku), which activated Type-1 immunity in a Toll-like receptor (TLR)4- and TLR2-dependent manner. Namely, the extract of Kurosengoku first caused production of IL-12 from DC and sequentially induced IFN-γ production by NK1.1(+) NK cells and NKT cells. The IFN-γ production was significantly blocked by neutralizing mAb against IL-12 or TLR4- and TLR2-deficient condition, indicating that TLR4- and TLR2-dependent activation of DC to produce IL-12 was essential for the production of IFN-γ from spleen cells by Kurosengoku. Moreover, the extract of Kurosengoku also enhanced production of IFN-γ from human PBMC by co-stimulation with anti-CD3 mAb in a TLR2- and TLR4-dependent manner. Thus, our findings strongly suggest that Kurosengoku might a novel immuno-improving food, which would be a useful tool for preventing the tip of immune balance in developed countries.

IFN-gamma-induced SOCS-1 regulates STAT6-dependent eotaxin production triggered by IL-4 and TNF-alpha.

The production of eotaxin, which is a critical mediator for airway inflammation, is inhibited by IFN-gamma. Here, we investigated the precise mechanisms underlying IFN-gamma-dependent inhibition of eotaxin production using mouse embryonic fibroblasts (MEF). MEF produced high levels of eotaxin in STAT6-dependent manner when they were cultured with both IL-4 and TNF-alpha. However, the eotaxin production by MEF was strongly inhibited by addition of IFN-gamma. Western-blotting analysis demonstrated that IFN-gamma downmodulated STAT6 phosphorylation induced by IL-4 and TNF-alpha. Moreover, IFN-gamma did not exhibit its inhibitory effect on both STAT6-phosphorylation and eotaxin production in MEF obtained from deficient mice in STAT1, a key molecule of IFN-gamma signaling. We also demonstrated that SOCS-1, a potent inhibitory molecule of IL-4 signaling, was induced by IFN-gamma in STAT1-dependent manner. This indicated that SOCS-1 might be involved in IFN-gamma-mediated STAT1-dependent inhibition of eotaxin production. In SOCS-1(-/-) MEF, IFN-gamma inhibited neither STAT6 phosphorylation nor eotaxin production induced by IL-4 and TNF-alpha. Conversely, retroviral transduction of SOCS-1 into MEF inhibited STAT6 phosphorylation and eotaxin production induced by IL-4 and TNF-alpha, in the absence of IFN-gamma. Thus, we demonstrated that IFN-gamma-induced inhibition of STAT6 phosphorylation and eotaxin production were mediated by SOCS-1 induced in STAT1-dependent manner.