Keiji Komoriya

Hypouricemic effect of the novel xanthine oxidase inhibitor, TEI-6720, in rodents.

We investigated the xanthine oxidase/xanthine dehydrogenase inhibitory activity and hypouricemic effect of a newly synthesized xanthine oxidase/xanthine dehydrogenase inhibitor, TEI-6720, 2-(3-cyano-4-isobutoxyphenyl)-4-methyl-5-thiazole-carboxylic acid, and compared its effects with those of allopurinol in rodents. TEI-6720 was found to inhibit bovine milk xanthine oxidase, and mouse liver and rat liver xanthine oxidase/xanthine dehydrogenase with IC50 values of 1.4, 1.8 and 2.2 nM, respectively. On bovine milk xanthine oxidase, TEI-6720 exhibited mixed-type inhibition and the Ki value was 0.7 nM. TEI-6720 displayed prolonged urate-lowering activity in normal mice and rats. We evaluated the hypouricemic effect of TEI-6720 on hyperuricemia induced by the uricase inhibitor, potassium oxonate (250 mg/kg s.c., 1 h before the test drugs), and measured the total molarity of both serum allantoin and urate in rats. Oral TEI-6720 and allopurinol had a hypouricemic effect 2 h after their administration to oxonate-pretreated rats with ED50 values of 1.5 and 5.0 mg/kg, respectively. Both compounds also reduced the combined molarity of uric acid and allantoin in rats. The ED50 values of TEI-6720 and allopurinol were 2.1 and 6.9 mg/kg p.o., respectively. These results suggest that TEI-6720 may be useful for the treatment of hyperuricemia.

Effects of ultrasound and 1,25-dihydroxyvitamin D3 on growth factor secretion in co-cultures of osteoblasts and endothelial cells

It has been shown that low-intensity pulsed ultrasound (US) accelerates fracture healing in animal models and in clinical studies. However, the mechanism by which US accelerates fracture healing remains unclear. Systemic factors and several growth factors, such as platelet-derived growth factor (PDGF), are thought to be involved in the process of fracture healing. In the present study, we examined the effects of US and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] on growth factor secretion in a co-culture system of human osteoblastic cells (SaOS-2) and endothelial cells (HUVEC). US was applied to cultured cells for 20 min daily for four consecutive days. US treatment increased the PDGF-AB level in the conditioned media. 1,25-(OH)2D3 (1 x 10(-8) M) also enhanced PDGF-AB secretion. The secretion of PDGF-AB was synergistically increased by the combination of US and 1,25-(OH)2D3. These results suggest that the stimulation of growth factor secretion from cells by US and 1,25-(OH)2D3 treatment may be involved in the acceleration of fracture healing.

Safety and pharmacokinetics of urtoxazumab, a humanized monoclonal antibody, against shiga-like toxin 2 in healthy adults and in pediatric patients infected with shiga-like toxin-producing Escherichia coli.

ABSTRACT Shiga-like toxin-producing Escherichia coli (STEC) infection causes diarrhea, which is often bloody and which can result in potentially life-threatening hemolytic-uremic syndrome (HUS). Urtoxazumab, a humanized monoclonal antibody directed against the Shiga-like toxin 2 (Stx2) produced by STEC, has been developed as a promising agent for the prevention of HUS. Single randomized, intravenous, double-blind, placebo-controlled doses of urtoxazumab were administered to assess its safety and pharmacokinetics in healthy adults (0.1 to 3.0 mg/kg of body weight) and STEC-infected pediatric patients (1.0 and 3.0 mg/kg). No dose-related safety trends were noted, nor were antiurtoxazumab antibodies detected. The disposition of urtoxazumab showed a biexponential decline, regardless of the dose. In healthy adults, the mean terminal elimination half-life was consistent across the dose groups and ranged from 24.6 days (3.0-mg/kg dose group) to 28.9 days (0.3-mg/kg dose group). The mean maximum serum drug concentration (Cmax) ranged from 2.6 μg/ml at 0.1 mg/kg to 71.7 μg/ml at 3.0 mg/kg. The disposition of urtoxazumab following the administration of doses of 1.0 and 3.0 mg/kg in pediatric patients showed mean Cmaxs of 19.6 and 56.1 μg/ml, respectively. Urtoxazumab was well tolerated, appears to be safe at doses of up to 3.0 mg/kg, and is a potential candidate for the prevention of HUS in pediatric patients.

Hypouricemic effect of allopurinol and the novel xanthine oxidase inhibitor TEI-6720 in chimpanzees

The hypouricemic effect of a newly synthesized xanthine oxidase/xanthine dehydrogenase inhibitor, TEI-6720, 2-(3-cyano-4-isobutoxyphenyl)-4-methyl-5-thiazolecarboxylic acid, was investigated and compared with that of allopurinol in male chimpanzees (n = 3). When allopurinol (10 mg/kg) was administered orally once a day for three consecutive days, it cumulatively reduced serum urate levels by 29.7, 50.1 and 60.2%, 24, 48 and 72 h, respectively, after the initial dose. This effect was dose dependent at doses of 3 and 10 mg/kg. At 3 mg/kg, the mean serum urate levels were 3.1, 2.4, 2.5 and 2.3 mg/dl before and 24, 48 and 72 h, respectively, after the initial dose. Animals treated with 10 mg/kg of allopurinol showed serum urate levels of 3.3, 2.3, 1.6 and 1.3 mg/dl, respectively. The urate-lowering effect of TEI-6720 was then compared with that of allopurinol at a daily dose of 5 mg/kg (n = 3). Both compounds caused striking reductions in serum and urinary uric acid levels accompanied by an increase in urinary xanthine levels. These effects of TEI-6720 were more potent than those of allopurinol. TEI-6720 reduced serum urate levels by 55.9, 69.6 and 73.6%, 24, 48 and 72 h, respectively, after the first dose, whereas the corresponding values after allopurinol were 28.1, 41.6 and 45.1%. These results suggest that the hypouricemic effect of TEI-6720 may be more potent than that of allopurinol in the treatment of hyperuricemia and gout, and that TEI-6720 may become an effective alternative drug.

HUMAN NEUTROPHILS EXPRESS MESSENGER RNA OF VITAMIN D RECEPTOR AND RESPOND TO 1α,25-DIHYDROXYVITAMIN D3

ABSTRACT 1α,25-Dihydroxyvitamin D3 (1,25(OH)2D3) has been shown to modulate the production of various cytokines or the expression of certain differentiation markers in human T cells or monocytes. Its effects on neutrophils, however, are poorly understood. In this paper, we show several lines of evidence indicating that neutrophils express functional vitamin D receptors (VDR). Sort-purified neutrophils from human peripheral blood expressed VDR mRNA at a level comparable to that of monocytes. As reported to occur in monocytes, protein expression of CD14 on the cell surface of neutrophils was augmented when the cells were incubated with 1,25(OH)2D3. To investigate the physiological roles for VDR in neutrophils, we investigated possible modulating effects of 1,25(OH)2D3 on the expression of several genes in lipopolysaccharide-stimulated neutrophils by using differential display analysis. Of the genes we identified, trappin-2/elafin/SKALP, which was originally reported to be an inhibitor of elastase, was induced in neutrophils by lipopolysaccharide, but was suppressed significantly in the presence of 1,25(OH)2D3. Under the same conditions, interleukin-1β expression was also inhibited. These findings suggest that 1,25(OH)2D3 has a potential to affect the inflammatory process by modulating the expression of neutrophil genes.

Allopurinol induces renal toxicity by impairing pyrimidine metabolism in mice

We investigated the relationship between the toxic effect of allopurinol and pyrimidine metabolism in mice. Allopurinol-induced increases in plasma transaminase levels in dinitrofluorobenzene (DNFB)-sensitized mice were not affected by uridine. In contrast, plasma creatinine and BUN tended to decrease 18 hr after the last injection of uridine. Both plasma and urinary orotidine (OD) were detected in DNFB-sensitized mice after administration of a single dose of allopurinol. In contrast, TEI-6720, a newly synthesized xanthine oxidase/xanthine dehydrogenase inhibitor, caused neither pyrimidine metabolism abnormality nor renal impairment in DNFB-sensitized mice. Also, normal mice administered high doses of allopurinol showed abnormal pyrimidine metabolism together with renal toxicity which could be ameliorated by uridine, indicating that allopurinol essentially causes pyrimidine metabolism abnormality leading to renal impairment. In DNFB-sensitized mice, allopurinol increased urinary OD excretion to an extent similar to that in normal mice administered the same dose of allopurinol. However, renal impairment by allopurinol was more striking in DNFB-sensitized mice than in normal mice. Histopathological observations showed that allopurinol induced calculus formation in the collecting tubules and papillary duct. Calculus formation was increased by DNFB and decreased by uridine. These observations indicate that the enhancement of the renal toxicity of allopurinol by DNFB-sensitization may be due to some biological interactions between DNFB and allopurinol. In humans, it is possible that there are some biological interactions which serve to enhance the toxicity of allopurinol, resulting in the development of allopurinol hypersensitivity syndrome (AHS). In contrast, TEI-6720, had no effect on pyrimidine metabolism and showed no toxic effect.

Effect of interleukin-10 on Anti-CD40- and interleukin-4-induced immunoglobulin E production by human lymphocytes

We studied the effect of IL-10 on the in vitro synthesis of IgE and IgG by human peripheral blood mononuclear cells (PBMC) following stimulation with anti-CD40 monoclonal antibody and IL-4. Anti-CD40- and IL-4-stimulated PBMC showed an increase in IL-10 synthesis together with increases in IgE and IgG production. Addition of anti-IL-10 antibody to this system suppressed IgE as well as IgG production without affecting the proliferation of PBMC. Addition of IL-10 enhanced IgE and IgG production if PBMC were activated with anti-CD40 and IL-4. PBMC costimulated with anti-CD40, IL-4 and IL-10 showed a remarkable increase in IL-6 production, but had no effect on IFN-gamma production. Addition of IL-10 to purified human tonsillar B cells stimulated with anti-CD40 and IL-4 enhanced B cell proliferation and IgG production, but not IgE production. These results suggest that IL-10 accelerates IgE production by anti-CD40- and IL-4-stimulated PBMC by enhancing IL-6 production through activation of T lymphocytes.

Pharmacological profiles of high-concentration (20 μg/g) tacalcitol ointment: Effects on cutaneous inflammation, epidermal proliferation, and differentiation in mice

This study focused on the effects of tacalcitol (1,24(R)(OH)2D3, TV‐02) ointment (20 μg/g) on cutaneous inflammation, epidermal proliferation, and differentiation and compared them with tacalcitol ointment (2 μg/g) and other anti‐psoriatic ointments using hairless mice. Tacalcitol ointment (0, 2 and 20 μg/g) significantly inhibited 12‐O‐tetradecanoylphorbol 13‐acetate (TPA)‐induced cutaneous inflammation, histopathologically. The effect of tacalcitol ointment (20 μg/g) on cutaneous inflammation was much stronger than that of tacalcitol ointment (0, 2 μg/g), and as effective as calcipotriol ointment (50 μg/g) or betamethasone valerate ointment (1.2 mg/g). Tacalcitol ointment (20 μg/g) also significantly inhibited TPA‐induced myeloperoxidase (MPO) activity, as effectively as calcipotriol ointment (50 μg/g) or betamethasone valerate ointment (1.2 mg/g). The effect of tacalcitol ointment on epidermal proliferation [ornithine decarboxylase (ODC) activity] and differentiation [transglutaminase (TGase) activity] was dose‐dependent from 0 μg/g to 20 μg/g. The effect of tacalcitol ointments on epidermal proliferation was significant at the doses of 2 μg/g and 20 μg/g, and that on epidermal differentiation was significant at the doses of 0.2 μg/g or more. The effect of tacalcitol ointment (20 μg/g) on epidermal differentiation was significantly stronger than tacalcitol ointment (2 μg/g). In this study, tacalcitol ointment (20 μg/g) was found to have a marked effect on cutaneous inflammation and improved effect on epidermal differentiation, although tacalcitol ointment (2 μg/g) also had significant effects on epidermal proliferation and differentiation. These findings support the clinical effectiveness of tacalcitol ointment (20 μg/g) against psoriasis.

IMMUNOPHAFMACOLOGICAL PROFILE OF TEI-3096: A NEW IMMUNOMODULATOR

TEI-3096 [6-p-chlorobenzyl-5H-2,3,6,7-terahydro-5,7-dioxothiazolo (3,2-a) pyrimidine], a novel thiazolopyrimidine compound has been shown to suppress adjuvant arthritis in rats without any effect on conventional inflammation. We examined the immuno-pharmacological profile of TEI-3096 in murine lymphocytes. The blastformation induced by Con A or LPS was inhibited by addition of 50-500 microM concentrations of TEI-3096. This compound suppressed elevated plaque-forming cell (PFC) response against T cell-dependent antigen. However, TEI-3096 had no effect either on normal PFC response or on antibody formation against T cell independent antigen. Although orally administered TEI-3096 had no obvious effect on anti-SRBC PFC response in normal mice, it normalized the colchicine-induced enhancement of antibody formation against TNP-HGG. TEI-3096 also enhanced the delayed type hypersensitivity in mice and rats. These results suggests that TEI-3096 restores the abnormal immune response. Therefore, it may be useful for the treatment of auto-immune disease such as rheumatoid arthritis.

Beneficial effects of a novel anti-hypoxemic agent, TEI-7322, on bleomycin-induced experimental hypoxemia in rats

Almitrine bismesylate is known to be an anti-hypoxemic agent that acts via the enhancement of hypoxic pulmonary vasoconstriction. However, screening for this class of compounds has been minimal, owing, in part, to a lack of convenient hypoxemic models in small animals. The present study was designed to establish a convenient model of hypoxemia induced by bleomycin and to evaluate anti-hypoxemic agents including a newly synthesized compound. TEI-7322, 2-allylamino-4-tert-butyl-amino-7-methyl-7H-pyrrolo[2,3- d]pyrimidine hydrochloride by using this model. Bleomycin was intratracheally instilled into rats. After 3 weeks, the arterial blood gas pressures were monitored in the animals in the conscious state. Then, prednisolone, doxapram, almitrine or TEI-7322 was administered to the bleomycin-treated rats to monitor changes in arterial blood gas pressures. Bleomycin-treated rats showed a decrease in the arterial blood O2 pressure (PaO2). The blood CO2 pressure (PaCO2) increased, along with an increase in the alveolar-arterial oxygen difference (AaDO2). These blood gas pressures in bleomycin-treated rats were not affected by treatment with prednisolone. Doxapram decreased the PaCO2 but did not change the PaO2. However, administration of almitrine or TEI-7322 significantly improved the PaO2 of bleomycin-treated rats with a decrease in the PaCO2. In conclusion, (1) bleomycin-induced lung injury causes hypoxemia in rats, probably resulting from ventilation-perfusion inequality; thus this model may be useful for evaluating anti-hypoxemic agents; and (2) TEI-7322, as well as almitrine, showed anti-hypoxemic effects in this model with different properties from those of doxapram, possibly due to improvement of ventilation-perfusion inequality, indicating that TEI-7322 may be a potent candidate for the treatment of hypoxemia.