Sphingosine analogue drug FTY720 targets I2PP2A/SET and mediates lung tumour suppression via activation of PP2A-RIPK1-dependent necroptosis.
Bottom Line: Mechanistically, targeting I2PP2A/SET by FTY720 mediated PP2A/RIPK1-dependent programmed necrosis (necroptosis), but not by apoptosis.The RIPK1 inhibitor necrostatin and knockdown or genetic loss of RIPK1 prevented growth inhibition by FTY720.Expression of WT- or death-domain-deleted (DDD)-RIPK1, but not the kinase-domain-deleted (KDD)-RIPK1, restored FTY720-mediated necroptosis in RIPK1(-/-) MEFs.
Affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC, USA.Show MeSH
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Mentions: Targeting I2PP2A/SET exogenously with a sphingolipid analogue drug that mimics ceramide and/or sphingosine could potentially bind I2PP2A/SET and reactivate PP2A tumour suppressor signalling (Mukhopadhyay et al, 2009). Since exogenous (Mukhopadhyay et al, 2009) and endogenous sphingosine (Fig 3A) bind wt-I2PP2A/SET, but not K209D-I2PP2A/SET, in A549 cells (Fig 3A), we examined whether sphingosine analogue FTY720, which was shown to inhibit tumour growth (Liu et al, 2010; Neviani et al, 2007; Pchejetski et al, 2010; Wallington-Beddoe et al, 2011), binds/targets I2PP2A/SET. We performed molecular docking studies based on I2PP2A/SET–ceramide modelling (Fig 1A and B), which predicted that one of the primary hydroxyl groups of FTY720 might bind to the K209 residue of I2PP2A/SET (Fig 3B and Supporting Information Fig S7). Biotin-labelled FTY720 (B-FTY720) was incubated with purified I2PP2A/SET, drug-bound proteins were then separated through an avidin column, and B-FTY720-bound I2PP2A/SET was detected by Western blotting using anti-I2PP2A/SET antibody. Remarkably, FTY720 (5–10 µM) also bound directly to purified I2PP2A/SET in vitro or endogenously expressed I2PP2A/SET in A549 cells (Fig 3C).
Affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, SC, USA.