Abstract 227: Overexpression of EphB1 leads to Stat3 phosphorylation

Abstract

Abstract The Receptor Tyrosine Kinase (RTK) family comprises some important disease drivers. Over-expressed or mutated RTKs in disease may lead to aberrant phosphorylation of intracellular signaling molecules. Therefore it is desirable to monitor simultaneously the activation levels of RTKs and signaling nodes. Here we used a multiplexed antibody array to monitor the phosphorylation of 28 RTKs and 11 intracellular signaling molecules simultaneously. We have observed a strong signal on a spot corresponding to phospho-STAT3 Tyr705 with lysates derived from CHO cells transiently over-expressing EphB1. The strong Phospho-STAT3 signal was not seen in non-transfected cells or in lysates made form cells transfected with several other Eph family members. Western blotting with phospho-specific antibodies showed an increase in STAT3 phosphorylation on Tyr705 in CHO cells upon EphB1 ectopic expression. The amounts of Tyr705 STAT3 phosphorylation positively correlated with the levels of EphB1 protein. Co-precipitation experiments showed that EphB1 and STAT3 interact both in CHO cells and MCF-7 breast cancer cells. Phosphorylated JAK2 was not found in these immuno-precipitates suggesting an involvement of another tyrosine kinase in STAT3 phosphorylation in cells over-expressing EphB1. Consistently with these observations, a potent JAK1 and JAK2 inhibitor INCB018424 inhibited the basal but not the EphB1-induced STAT3 Tyr705 phosphorylation in transfected CHO cells. These results suggest that EphB1 over-expression may bypass the requirement in Jak kinase and lead to STAT3 activation either directly or through another receptor-tethered kinase. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 227. doi:1538-7445.AM2012-227