SB-431542 Is a Potent and Specific Inhibitor of Transforming Growth Factor-β Superfamily Type I Activin Receptor-Like Kinase (ALK) Receptors ALK4, ALK5, and ALK7

Abstract

Small molecule inhibitors have proven extremely useful for investigating signal transduction pathways and have the potential for development into therapeutics for inhibiting signal transduction pathways whose activities contribute to human diseases. Transforming growth factor ␤ (TGF-␤) is a member of a large family of pleiotropic cytokines that are involved in many biological processes, including growth control, differentiation, migration, cell survival, adhesion, and specification of developmental fate, in both normal and diseased states. TGF-␤ superfamily members signal through a receptor complex comprising a type II and type I receptor, both serine/threonine kinases. Here, we characterize a small molecule inhibitor (SB-431542) that was identified as an inhibitor of activin receptor-like kinase (ALK)5 (the TGF-␤ type I receptor). We demonstrate that it inhibits ALK5 and also the activin type I receptor ALK4 and the nodal type I receptor ALK7, which are very highly related to ALK5 in their kinase domains. It has no effect on the other, more divergent ALK family members that recognize bone morphogenetic proteins (BMPs). Consistent with this, we demonstrate that SB-431542 is a selective inhibitor of endogenous activin and TGF-␤ signaling but has no effect on BMP signaling. To demonstrate the specificity of SB-431542, we tested its effect on several other signal transduction pathways whose activities depend on the concerted activation of multiple kinases. SB-431542 has no effect on components of the ERK, JNK, or p38 MAP kinase pathways or on components of the signaling pathways activated in response to serum.

TGF‐β signaling directly regulates transcription and functional expression of the electrogenic sodium bicarbonate cotransporter 1, NBCe1 (SLC4A4), via Smad4 in mouse astrocytes

“…Cultures were treated for 20 min either with 100 µM 4AP alone, or in the presence of SB431542 (10 µM) an Alk4, Alk5, and Alk7 inhibitor (Inman, 2002), and therefore inhibitor of TGF‐β receptor II‐, or with 2 ng/mL exogenous TGF‐β. Initial experiments have ensured that SB431542 has the same effect on cortical astrocytes as α‐TGF‐β.…”

“…This effect is specific for 4AP, since inhibition of TGF‐β signaling in control astrocytes had no effect on NBCe1 (Figure 2b–d). We have used SB431542 to block TGF‐β signaling (Inman, 2002). Because this inhibitor inhibits Alk4, 5, and 7, we have performed initial experiments with α‐TGF‐β 1,2,3 and ensured that α‐TGF‐β 1,2,3 and SB31542 have the same effects on 4AP‐dependent NBCe1 regulation, suggesting that the results of this work indeed derive from inhibition of TGF‐β signaling.…”

TGF-β-mediated activation of RhoA signalling is required for efficient V12HaRas and V600EBRAF transformation

“…Utilizing the recently characterized ALK5 kinase inhibitor SB-431542 (Inman et al, 2002) we show that TGF-b-mediated RhoA activation is kinase dependent. We also show that ALK5 regulates basal RhoA activity in serum-starved rodent fibroblasts in a Smad-independent manner.…”

Protein phosphatase 5 modulates SMAD3 function in the transforming growth factor‐β pathway

“…As the tail-phosphorylated SMADs are observed in the nucleus, the phosphatases that catalyse their dephosphorylation are most likely nuclear. Further observations, including that receptor kinase inhibition results in pronounced cytoplasmic distribution of dephosphorylated SMAD2/3, support the existence of R-SMAD phosphatases in the nucleus (12)(13)(14). Consistent with this notion, PPM1A (also known as PP2C) was reported to be a SMAD2/3-tail phosphatase in the nucleus (11).…”