| In vitro: |
| J Biol Chem,1993 Dec 5;268(34):25846-56. | | Inhibition of histamine secretion by wortmannin through the blockade of phosphatidylinositol 3-kinase in RBL-2H3 cells.[Pubmed: 7503989] |
METHODS AND RESULTS:
The surface engagement of high affinity immunoglobulin E receptor (Fc epsilon RI) of rat basophilic leukemia 2H3 (RBL-2H3) cells induced histamine secretion and leukotriene release following activation of the tyrosine kinase Lyn together with phosphatidylinositol 3-kinase (PI3-kinase). Wortmannin inhibited the activity of partially purified PI3-kinase from calf thymus, as well as the PI3-kinase activity in anti-PI3-kinase p85 immunoprecipitates from RBL-2H3 cells, at a concentration as low as 1.0 nM and with IC50 values of 3.0 nM, but did not inhibit PI4-kinase activity. The inhibition of PI3-kinase by Wortmannin was irreversible. Wortmannin inhibited both Fc epsilon RI-mediated histamine secretion and leukotriene release up to 80% with IC50 values of 2.0 and 3.0 nM, respectively. Wortmannin inhibited PI3-kinase activity in intact cells up to 80% with an IC50 value of 2.0 nM, which is almost equal to those for PI3-kinase in vitro and for histamine secretion and leukotriene release. With anti-Wortmannin antibody, we have shown that Wortmannin binds to the 110-kDa protein, but not to PI3-kinase 85-kDa regulatory subunit both in vitro and in whole cells. Furthermore, there was a positive correlation between the potencies of Wortmannin derivatives as inhibitors of PI3-kinase and as inhibitors of histamine secretion. Wortmannin had no effect on the activation of the tyrosine kinase Lyn.
CONCLUSIONS:
These results suggest that PI3-kinase is involved in the signal transduction pathway responsible for histamine secretion following stimulation of Fc epsilon RI and that Wortmannin blocks these responses through direct interaction with the catalytic subunit of this enzyme. | | J Biol Chem,1994 Feb 4;269(5):3568-73. | | Essential role of phosphatidylinositol 3-kinase in insulin-induced glucose transport and antilipolysis in rat adipocytes. Studies with a selective inhibitor wortmannin.[Pubmed: 8106400] |
METHODS AND RESULTS:
Significant activity of phosphatidylinositol 3-kinase (PI 3-kinase) was detected in the membrane fractions, or in the immunoprecipitates prepared with anti-phosphotyrosine antibodies, from rat adipocytes that had been incubated with insulin for 20 min. The PI 3-kinase activity in these preparations as well as in the whole cell lysates of adipocytes not treated with insulin was inhibited by the addition of Wortmannin, a fungal metabolite, to the enzyme assay mixture. The inhibition was dependent on the inhibitor concentration with IC50 being less than 10 nM and perfect inhibition at 100 nM. The effect of insulin to induce membrane PI 3-kinase activity was mostly abolished, but its effects to tyrosine-phosphorylate the beta-subunit of the insulin receptor or other cellular substrate proteins including insulin-receptor-substrate-1 were not at all antagonized, by Wortmannin added to the cell incubation medium. Insulin stimulation of cellular 2-deoxyglucose uptake and inhibition of isoproterenol-induced lipolysis observable in adipocytes under the same conditions were also antagonized by Wortmannin added in the same concentration range as used for the inhibition of insulin-susceptible PI 3-kinase.
CONCLUSIONS:
It is concluded, therefore, that activation of Wortmannin-sensitive PI 3-kinase plays a pivotal role in the intracellular signaling pathways arising from the insulin receptor autophosphorylation and leading to certain metabolic responses. |
|
| In vivo: |
| Cancer Sci,2009 Apr;100(4):770-7. | | Phosphoinositide 3-kinase inhibitor (wortmannin) inhibits pancreatic cancer cell motility and migration induced by hyaluronan in vitro and peritoneal metastasis in vivo.[Pubmed: 19469020] | In order to block peritoneal metastasis of pancreatic cancer cells, we have attempted to block the signal transduction pathway involving hyaluronan (HA), Src, phosphoinositide 3-kinase (PI3K) and Akt.
METHODS AND RESULTS:
We examined the effects of Src, PI3K and Akt inhibitors on pancreatic cancer cell motility, invasion and metastasis. The pancreatic cancer cell line SW1990, known to cause peritoneal metastasis efficiently in nude mice, was used in this study. SW1990 cells were stimulated by HA to induce Akt phosphorylation. Then, the inhibitory effects of PI3K and Src kinase inhibitors were examined. Cell motility and cell migration assays were adopted to assess the cancer cell motility and its migration capability. We also examined the therapeutic efficacies of PI3K inhibitor Wortmannin on peritoneal metastasis of SW1990 cells in the nude mouse model. Stimulation of SW1990 cells by HA markedly induced the Src-PI3K-Akt signaling, thus enhancing cancer cell motility and its migration. Significantly, we found that Wortmannin could exert marked inhibition of the peritoneal metastasis of SW1990 in nude mice in vivo.
CONCLUSIONS:
These findings indicate that the PI3K-Akt signaling pathway plays an essential role in peritoneal metastasis and PI3K inhibitors such as Wortmannin can be novel modalities to prevent peritoneal metastasis of invasive cancers such as pancreatic cancer. |
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