Kinase assays

The oxidation of L-[14C]arginine was monitored by the conversion of L-[14C]arginine to L-[14C]citrulline. The enzyme was assayed at 37℃ in 20 mM Hepes, pH 7.4, with 2.5 mM dithiothreitol, 125 μM NADPH, 10 μM tetrahydrobiopterin, 10 μM FAD, and 0.5-20 μM L-[14C]arginine (10,000 cpm). When eNOS or nNOS were assayed, calmodulin (10 μg/ml) and CaCl2 (2.5 mM) were included. When time courses were measured, bovine serum albumin (1 mg/ml) was included.

Animal models

Rat model of endotoxin-induced microvascular injury

Dosage form

0.1-10 mg/kg, subcutaneous

Application

1400W was a potent inhibitor against iNOS in a rat model of endotoxin-induced vascular injury. When administered 3 h after endotoxin, 1400W completely suppressed leakage into the ileum. 1400W has an effect of endotoxin -induced vascular injury in the ileum.

Other notes

Please test the solubility of all compounds indoor, and the actual solubility may slightly differ with the theoretical value. This is caused by an experimental system error and it is normal.

1400W dihydrochloride is a potent and selective inhibitor of inducible nitric oxide synthase with Kd value of 7 nM [1].

Inducible nitric oxide synthase (iNOS) is an enzyme catalyzing the production of nitric oxide (NO) and is involved in immune response. iNOS produces NO as an immune defense mechanism.

1400W dihydrochloride is an extremely slowly reversible and selective iNOS inhibitor [1]. 1400W inhibited iNOS with Kd value of 7 nM and rapidly reversibly inhibited human neuronal NOS (nNOS) and endothelial NOS (eNOS) with Ki values of 2 and 50 μM, respectively. L-arginine was a competitive inhibitor of 1400W with Ks value of 3.0 μM [1].

In an endotoxin-induced vascular injury rat model, 1400W exhibited 50-fold more potent selectivity against iNOS than eNOS [1]. In focal cerebral ischaemia rats, 1400W (20 mg/kg) inhibited neurological dysfunction and weight loss and significantly decreased ischaemic lesion volume by 31%. Also, 1400W reduced iNOS activity by 36% in the infarct [2]. In the EMT6 murine mammary adenocarcinoma, 1400W (10 or 12 mg/kg/h) significantly reduced tumor weight and inhibited iNOS activity [3].

References:
[1].  Garvey EP, Oplinger JA, Furfine ES, et al. 1400W is a slow, tight binding, and highly selective inhibitor of inducible nitric-oxide synthase in vitro and in vivo. J Biol Chem, 1997, 272(8): 4959-4963.
[2].  Thomsen LL, Scott JM, Topley P, et al. Selective inhibition of inducible nitric oxide synthase inhibits tumor growth in vivo: studies with 1400W, a novel inhibitor. Cancer Res, 1997, 57(15): 3300-3304.
[3].  Parmentier S, Bohme GA, Lerouet D, et al. Selective inhibition of inducible nitric oxide synthase prevents ischaemic brain injury. Br J Pharmacol, 1999, 127(2): 546-552.