5-(1,3-Benzothiazol-6-yl)-4-(4-methyl-1,3-thiazol-2-yl)-1H-imidazole derivatives as potent and selective transforming growth factor-ß type I receptor inhibitors.

A series of 5-(1,3-benzothiazol-6-yl)-4-(4-methyl-1,3-thiazol-2-yl)-1H-imidazole derivatives was synthesized as transforming growth factor-ß (TGF-ß) type I receptor (also known as activin-like kinase 5 or ALK5) inhibitors. These compounds were evaluated for their ALK5 inhibitory activity in an enzyme assay and for their TGF-ß-induced Smad2/3 phosphorylation inhibitory activity in a cell-based assay. As a representative compound, 16i was a potent and selective ALK5 inhibitor, exhibiting a good enzyme inhibitory activity (IC(50) = 5.5 nM) as well as inhibitory activity against TGF-ß-induced Smad2/3 phosphorylation at a cellular level (IC(50) = 36 nM). Furthermore, the topical application of 3% 16i lotion significantly inhibited Smad2 phosphorylation in Mouse skin (90% inhibition compared with vehicle-treated animals).

Blocking S1P interaction with S1P1 receptor by a novel competitive S1P1-selective antagonist inhibits angiogenesis.

Sphingosine 1-phosphate receptor type 1 (S1P(1)) was shown to be essential for vascular maturation during embryonic development and it has been demonstrated that substantial crosstalk exists between S1P(1) and other pro-angiogenic growth factors, such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor. We developed a novel S1P(1)-selective antagonist, TASP0277308, which is structurally unrelated to S1P as well as previously described S1P(1) antagonists. TASP0277308 inhibited S1P- as well as VEGF-induced cellular responses, including migration and proliferation of human umbilical vein endothelial cells. Furthermore, TASP0277308 effectively blocked a VEGF-induced tube formation in vitro and significantly suppressed tumor cell-induced angiogenesis in vivo. These findings revealed that S1P(1) is a critical component of VEGF-related angiogenic responses and also provide evidence for the efficacy of TASP0277308 for anti-cancer therapies.

Design, synthesis, and evaluation of novel 4-thiazolylimidazoles as inhibitors of transforming growth factor-ß type I receptor kinase.

A novel series of 4-thiazolylimidazoles was synthesized as transforming growth factor-ß (TGF-ß) type I receptor (also known as activin receptor-like kinase 5 or ALK5) inhibitors. These compounds were evaluated for their ALK5 inhibitory activity in an enzyme assay and their TGF-ß-induced Smad2/3 phosphorylation inhibitory activity in a cell-based assay. N-{[5-(1,3-benzothiazol-6-yl)-4-(4-methyl-1,3-thiazol-2-yl)-1H-imidazol-2-yl]methyl}butanamide 20, a potent and selective ALK5 inhibitor, exhibited good enzyme inhibitory activity (IC(50)=8.2nM) as well as inhibitory activity against TGF-ß-induced Smad2/3 phosphorylation at a cellular level (IC(50)=32nM).

Lymphopenia induced by a novel selective S1P(1) antagonist structurally unrelated to S1P.

Sphingosine 1-phosphate (S1P) regulates lymphocyte trafficking via type-1 S1P receptor (S1P(1)) and participates in many pathological conditions. We developed a novel type S1P(1)-selective antagonist, TASP0251078, which is structurally unrelated to S1P. This competitive antagonist inhibited binding of S1P to S1P(1) resulting in reduced signaling downstream of S1P(1), including GTPγS-binding and cAMP formation. TASP0251078 also inhibited S1P-induced cellular responses such as chemotaxis and receptor-internalization. Furthermore, when administered in vivo, TASP0251078 induced lymphopenia in blood, which is different from previously reported effects of other S1P(1)-antagonists. In a mouse contact hypersensitivity model, TASP0251078 effectively suppressed ear swelling, leukocyte infiltration, and hyperplasia. These findings provide the chemical evidence that S1P(1) antagonism is responsible for lymphocyte sequestration from the blood, and suggest that the effect of S1P(1) agonists on lymphocyte sequestration results from their functional antagonism.

Amelioration of collagen-induced arthritis by a novel S1P1 antagonist with immunomodulatory activities.

Sphingosine 1-phosphate (S1P) regulates lymphocyte trafficking through the type 1 sphingosine 1-phosphate receptor (S1P(1)) and participates in many pathological conditions, including autoimmune diseases. We developed a novel S1P(1)-selective antagonist, TASP0277308, which is structurally unrelated to S1P. This antagonist competitively inhibited S1P-induced cellular responses, such as chemotaxis and receptor internalization. Furthermore, differing from previously reported S1P(1) antagonists, TASP0277308 demonstrated in vivo activities to induce lymphopenia, a block in T cell egress from the thymus, displacement of marginal zone B cells, and upregulation of CD69 expression on both T and B cells, all of which recapitulate phenotypes of S1P(1)-deficient lymphocytes. In a mouse collagen-induced arthritis model, TASP0277308 significantly suppressed the development of arthritis, even after the onset of disease. These findings provide the first chemical evidence to our knowledge that S1P(1) antagonism is responsible for immunosuppression in the treatment of autoimmune diseases and also resolve the discrepancies between genetic and chemical studies on the functions of S1P(1) in lymphocytes.

Effects of TS-022, a newly developed prostanoid DP1 receptor agonist, on experimental pruritus, cutaneous barrier disruptions and atopic dermatitis in mice.

TS-022, {4-[(1R, 2S, 3R, 5R)-5-Chloro-2-((S)-3-cyclohexyl-3-hydroxyprop-1-ynyl)-3-hydroxycyclopentyl] butylthio} acetic acid monohydrate, inhibits ADP-induced platelet aggregation, an effect significantly antagonized, as in the case of prostaglandin D(2) by the prostanoid DP(1) receptor antagonist (BW A868C). TS-022 is a prostanoid DP(1) receptor agonist, originally developed as a novel anti-pruritic drug for patients with atopic dermatitis. We examined the effects of TS-022 on experimental pruritus, cutaneous barrier disruption, and atopic dermatitis and in in vitro immune function tests. Topically applied TS-022 significantly suppressed scratching in skin-lesioned NC/Nga mice from a concentration of 2.5 nM, and this scratch-suppressive activity was significantly antagonized by BW A868C. Tacrolimus (FK-506) and dexamethasone, used as reference drugs for atopic dermatitis, also exhibited suppressive effects against scratching, but only at concentrations of 125 and 25,000 microM. TS-022 applied topically, once a day for 2 days, significantly accelerated repair of the cutaneous barrier disruption caused by mechanical scratching, from concentrations of 2.5 nM. This acceleration of repair of the disrupted cutaneous barrier by this drug was also significantly antagonized by BW A868C. FK-506 and dexamethasone showed no beneficial effects on the repair of the disrupted cutaneous barrier. Repeated topical application of 2.5 microM of TS-022 and 12.5 microM of FK-506 once a day for 6 weeks significantly improved the skin inflammation scores in the NC/Nga mice. In regard to the effects of TS-022 in vitro, the inhibitory activity of TS-022 against concanavalin A-induced cytokine production by splenocytes was marginal as compared with that of FK-506 or dexamethasone. These results suggest that the beneficial therapeutic effects of TS-022 in NC/Nga mice with atopic dermatitis are mediated by its suppressive effect on scratching and its effect of accelerating repair of the disrupted cutaneous barrier, both effects being attributable to its prostanoid DP(1) receptor agonistic activity.

[Use of complementary and alternative medicine and health problems].

OBJECTIVE: Remarkable growth in use of alternative and complementally medicine (CAM) has recently been noted from consume to trends, detail surveys are limited. In this study, to clarify the actual state of use of CAM and associated problem, we performed a cross sectional study in a town using a self-administrated questionnaire. METHODS: The questionnaire including demographic variables, subjective health status and health practices was addressed by people in Oguni town in Kumamoto. Use of kampo, supplements/healthy food, chiropractic, massage, yoga/meditation, acupuncture, kiko/thai-chi, aromatherapy/herbal medicine and hot springs was assessed in the questionnaire in terms of frequency, prescription or advice from physicians, purpose, and satisfaction. RESULTS: The response rate was 83.33%. Use increased with aging and female employed CAM more frequently than male subjects. Most frequently consumed were supplement/health foods in both females (47.0%) and males (35.3%). The most prescribed was Kampo in both sexes (24.8% and 11.4%) About 70% of the subjects had visited chiropractics therapies. CONCLUSION: From 57.0% of subjects had used at least one CAM in the past six months, a high value compared with results from other countries. The rates were particularly large in female and elderly subjects. It is thus possible that the impact of CAM on health promotion policy is not inconsequential.

20-hydroxyeicosatetraenoic acid (20-HETE): structural determinants for renal vasoconstriction.

The effects of natural and synthetic eicosanoids on the diameter of rat interlobular arteries studied in vitro were compared to that of the potent, endogenous vasoconstrictor 20-HETE. Vasoconstrictor activity was optimum for chain lengths of 20-22 carbons with at least one olefin or epoxide between located between C(13)-C(15) and an oxygen substituent at C(20)-C(22). The presence of delta (Zou et al. Am. J. Physiol. 1996, 270, R228; Gebremedhin, D. et al. Am. J. Physiol. 1998, 507, 771)-, delta (Carroll et al. Am. J. Physiol. 1996, 271, R863; Vazquez et al. Life Sci. 1995, 56, 1455)-, or delta (Imig et al. Hypertension 2000, 35, 307; Lopez et al. Amer. J. Physiol. 2001, 281, F420)-olefins had no influence on the vasoconstrictor response whereas the introduction of a C(7)-thiomethylene enhanced potency. A sulfonamide or alcohol, but not a lactone, could replace the C(1)-carboxylate. These data were used to construct a putative binding domain map of the 20-HETE receptor consisting of: (i) a comparatively open, hydrophilic binding site accommodating the C(1)-functionality; (ii) a hydrophobic trough spanning the olefins; (iii) a shallow pocket containing a critical pi-pi binding site in the vicinity of the pi (Ito et al. Am. J. Physiol. 1998, 274, F395; Quigley, R.; Baum, M.; Reddy, K. M.; Griener, J. C.; Falck, J. R. Am. J. Physiol. 2000, 278, F949)-olefin; and (iv) an oxyphilic binding site proximate to the omega-terminus.

Eicosanoid regulation of vascular endothelial growth factor expression and angiogenesis in microvessel endothelial cells.

12(R)-Hydroxy-5,8,14-eicosatrienoic acid (HETrE) is a potent inflammatory and angiogenic eicosanoid in ocular and dermal tissues. Previous studies suggested that 12(R)-HETrE activates microvessel endothelial cells via a high affinity binding site; however, the cellular mechanisms underlying 12(R)-HETrE angiogenic activity are unexplored. Because the synthesis of 12(R)-HETrE is induced in response to hypoxic injury, we examined its interactions with vascular endothelial growth factor (VEGF) in rabbit limbal microvessel endothelial cells. Addition of 12(R)-HETrE (0.1 nm) to the cells increased VEGF mRNA levels with maximum 5-fold increase at 45 min. The increase in VEGF mRNA was followed by an increase in immunoreactive VEGF protein. 12(R)-HETrE (0.1 nm) rapidly activated the extracellular signal-regulated kinases (ERKs) ERK1 and ERK2. Moreover, preincubation of cells with PD98059, a selective inhibitor of MEK-1, inhibited 12(R)-HETrE-induced VEGF mRNA. Addition of VEGF antibody to cells grown in Matrigel-coated culture plates inhibited 12(R)-HETrE-induced capillary tube-like formation, suggesting that VEGF mediates, at least in part, the angiogenic response to 12(R)-HETrE. The results indicate that in microvessel endothelial cells, 12(R)-HETrE induces VEGF expression via activation of ERK1/2 and that VEGF mediates, at least in part, the angiogenic activity of 12(R)-HETrE. Given the fact that both VEGF and 12(R)-HETrE are produced in the cornea after hypoxic injury, their interaction may be an important determinant in the development of neovascularized tissues.