Cell lines

1483 head and neck carcinoma cells

Preparation method

The solubility of this compound in DMSO is >23.3mg/mL. General tips for obtaining a higher concentration: Please warm the tube at 37 ℃ for 10 minutes and/or shake it in the ultrasonic bath for a while. Stock solution can be stored below -20℃ for several months.

Reacting condition

1 μM, 1 h

Applications

In 1483 cells pretreated for 1 h with a range of concentrations of PF-543, PF-543 dose-dependently depleted the intracellular level of S1P with EC50 concentration of 8.4 nM and elevated the intracellular level of sphingosine. PF-543 showed no effect on the growth of several other cancer cell lines. PF-543 inhibited S1P formation in human whole blood (IC50=26.7 nM).

Animal models

Mouse hypoxic model of pulmonary hypertension

Dosage form

Intraperitoneal injection, 10 mg/kg, 24 h

Application

In a mouse hypoxic model of pulmonary hypertension, PF-543 showed no effect on vascular remodelling but reduced right ventricular hypertrophy. Administration of 10 mg/kg PF-543 for 24 h to mice decreased SK1 expression in pulmonary vessels.

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.

PF-543 is a novel and cell-permeant inhibitor of Sphingosine kinase (SphK1) with IC50 value of 3.6 nM [1],

SphK1 is a kinase that phosphorylates sphingosine to sphingosine-1-phosphate (SIP). SphK1 is normally a cytosolic protein but can be recruited to the membrane to conduct its activity. It is the major source of production of S1P which promotes cell growth, survival and migration, and also regulate lymphocyte trafficking.

Biochemical study has identified that SphK1 was a sphingosine-competitive inhibitor but not ATP-competitive [1]. In 1483 head and neck carcinoma cell cultures expressing high levels of SphK1 and with an unusually high rate of S1P production, pretreatment of PF-543 for 1 hr decreased the level of endogenous S1P by 10-fold with a proportional increase in the level of sphingosine. It indicated a significant inhibition of SphK1 BY pf-543. However, specific inhibition of SphK1 had no effect on the proliferation and survival of 1483 cell cultures, despite a dramatic change in the cellular S1P/sphingosine rate [1].

The inhibitory activity of PF-543 was also examined ex vivo in human whole blood. Human whole blood with high SphK1 activity was prepared, which was able to quickly convert C17-sphingosine to C17-S1P. PF-543 treatment showed that PF-543 was a potent inhibitor of SphK1, capable of blocking >90% C17-S1P formation [1].

Reference:
[1] Schnute M E et al. , Modulation of cellular S1P levels with a novel, potent and specific inhibitor of sphingosine kinase-1. Biochem J. 2012, 444(1): 79-88.