Abstract
SB202190, a widely used inhibitor of p38 MAPKα and β, was recently described to induce autophagic vacuoles and cell death in colon and ovarian cancer cells lines and, therefore, this effect was supposed to be specific for transformed cells and to open therapeutic options. Here, we demonstrate that SB202190 and the structurally related inhibitor SB203580 induce pro-autophagic gene expression and vacuole formation in various cancer and non-cancer cell lines of human, rat, mouse and hamster origin. This effect seems to induce defective autophagy leading to the accumulation of acidic vacuoles, p62 protein and lipid conjugated LC3. Using further p38 inhibitors we show that p38 MAPK inhibition is not sufficient for the autophagic response. In line with these results, expression of a SB202190-resistant mutant of p38α, which significantly increases activity of the p38 pathway under inhibitory conditions, does not block SB202190-dependent vacuole formation, indicating that lack of p38α activity is not necessary for this effect. Obviously, the induction of autophagic vacuole formation by SB203580 and SB202190 is due to off-target effects of these inhibitors on post-translational protein modifications, such as phosphorylation of the MAPKs ERK1/2 and JNK1/2, ribosomal protein S6, and PKB/Akt. Interestingly, the PI3K-inhibitor wortmannin induces transient vacuole formation indicating that the PI3K-PKB/Akt-mTOR pathway is essential for preventing autophagy and that cross-inhibition of this pathway by SB202190 could be the reason for the early part of the effect observed.
Highlights
Small molecule protein kinase inhibitors are largely being developed for the treatment of a variety of human diseases [1,2]. p38 MAPK has been identified as a potential target of such small molecules for the treatment of cancer and inflammation [3]
Induction of autophagy is characterized by the conversion of microtubule-associated protein 1 light chain 3 (LC3) to a lipid conjugated form, LC3-II, and its recruitment to autophagic vacuoles forming a distinct punctuate staining pattern [20]
The cell type-specific induction of defective autophagic vacuoles by SB202190 and SB203580 seen in colon cancer HT29 cells, which we extended to other transformed and non-transformed cells types including primary cells, is a p38 MAPK-independent effect of these compounds
Summary
Small molecule protein kinase inhibitors are largely being developed for the treatment of a variety of human diseases [1,2]. p38 MAPK has been identified as a potential target of such small molecules for the treatment of cancer and inflammation [3]. Small molecule protein kinase inhibitors are largely being developed for the treatment of a variety of human diseases [1,2]. P38 MAPK has been identified as a potential target of such small molecules for the treatment of cancer and inflammation [3]. The gate keeper threonine (T) 106 in the ATP-binding groove of p38a was shown to be the major determinant for the specificity of this class of compounds. The SB compounds were thought to inhibit the a and b isoforms of p38 MAPK leading to suppression of inflammatory gene expression, later studies identified several further protein kinase targets of these compounds including GAK, GSK3b, RICK (RIP2), Casein kinase I, Type-II TGF receptor, LCK, CRAF (Raf-1), BRAF and PDK1 [5,6,7]. At higher concentrations SB compounds were shown to have inhibitory effects on several non-protein kinase targets, such as hepatic cytochrome P450 enzymes [8], cyclooxygenases and thromboxane synthase [9]
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