Interestingly, STAT3Tyr705 phosphorylation was not inhibited by treatment with AG1478 except for, a partial inhibition maybe that was observed in BxPC3 cells treated for 96 h with higher concentrations of AG1478. STAT3Tyr705 phosphorylation is considered to be a down stream target of EGFR signaling in some cell types. However, other studies showed that inhibiting EGFR signaling did not affect STAT3Tyr705 phosphorylation. Skin biopsies of patients treated with the EGFR inhibitor Gefitinib showed a decreased EGFR activation that was associated with an increase in STAT3Tyr705 phosphorylation. In a majority of the HNSCC cells lines tested, inhibition of EGFR signaling by AG1478 did not affect the overall STAT3Tyr705 phosphorylation Inhibitors,Modulators,Libraries levels, while EGFR, ERKs and STAT3Ser727 phosphorylation was inhibited.
In agreement with these latter studies, the data presented here indicates that constitutive STAT3Tyr705 phosphory lation does not require EGFR signaling in the four human PDAC cell lines that were examined. As anticipated, treat ment with Inhibitors,Modulators,Libraries AG1478 of the four PDAC cell lines used in this study did show inhibition of phosphorylation of EGFR, AKT and ERKs. Thus the growth sup pressive effect of AG1478 may be attributable to a reduc tion of the phosphorylation of AKT or ERKs, which are also known to play a role in tumor progression. However, even after effective inhibition of EGFR signaling, the pres ence of constitutive STAT3Tyr705 phosphorylation may decrease the response to chemotherapy Inhibitors,Modulators,Libraries by inducing pro survival pathways.
Similar to this observation, treatment of cells with gemcitabine Inhibitors,Modulators,Libraries either alone or in com bination with AG1478 did not affect the constitutive STAT3Tyr705 phosphorylation. The presence of constitutive phosphorylation of STAT3Tyr705 following treatment with AG1478 or gemcitabine prompted us to investigate whether inhibiting STAT3 would increase the sensitivity of PDAC cells to chemotherapy. Interestingly, PDAC cells with knockdown of STAT3 demonstrated a similar exponential growth rate as Inhibitors,Modulators,Libraries the control cells in vitro. However, PDAC cells with STAT3 knocked down showed a decreased colony forming ability when plated at low density suggesting a reduced onco genic phenotype. Cells where STAT3 was knocked down showed a significant increase of growth inhibitory response to gemcitabine. STAT3 knockdowns of PANC 1 and UK Pan 1 cells showed significant growth inhibition from 0.
5 ngml dose of gemcitabine as compared to 4 and 6 ngml of gem citabine required to cause significant growth inhibition of their respective control cells. BxPC3 and MIA PaCa 2 cells showed a greater Diabete resistance to gemcitabine compared to PANC 1 and UK Pan 1. Knockdown of STAT3 in the gemcitabine resistant PDAC cell lines resulted in a significant increase of growth sup pression.