A (-)-MK 801 Maleate A 205804 All Your Co-Workers Is Talking About

We identified that treatment with NSC114792 decreased (-)-MK 801 Maleate the tyrosine phosphorylation of each JAK3 and STAT5 within a dose dependent manner. Furthermore, we identified that BKO84 cells handled with NSC114792 have significantly decreased viability within a time and dose dependent manner.

To more evaluate the specificity of NSC114792 for JAK3 inhibition, we employed the rat pre T lymphoma cell line Nb2 plus the murine myeloid progenitor cell line 32D stably expressing IL 2Rb, each of which are previously employed to study cytokine dependent activation of JAK proteins. We first examined the effects of NSC114792 on phospho JAK2 and phospho (-)-MK 801 Maleate JAK3 induced by PRL and IL 2 treatment, respectively, in Nb2 cells. Cells were incubated in the presence of NSC114792 for 16 hours and then stimulated by PRL or IL 2 for 10 minutes. While phospho JAK2 and phospho JAK3 were barely detectable in cells without stimulation, their levels were increased in response to PRL and IL 2 stimulation, respectively. As expected, NSC114792 could not inhibit PRL induced JAK2/ STAT5 phosphorylation at the concentrations up to 20 umol/L.

In 32D/IL 2Rb cells in the NSCLC absence of cytokine stimulation, phospho JAK2 and phospho JAK3 were barely detectable. However, consistent with the previous report, JAK2 and JAK3 become tyrosine phosphorylated in response to treatment with IL 3 and IL 2, respectively. Consistent with the results from Nb2 cells, NSC114792 did not affect IL 3 induced JAK2/STAT5 phosphorylation, whereas it did block IL 2 induced JAK3/ STAT5 phosphorylation. Once again, the pan JAK inhibitor AG490 non selectively inhibited JAK2 and JAK3 phosphorylation induced by IL 3 and IL 2, respectively. These findings strongly suggest that NSC114792 has selectivity for JAK3 over JAK2. We further assessed if NSC114792 can specifically inhibit JAK3, but not other JAKs, using various cancer cell lines where constitutively active JAK kinases are expressed.

By contrast, the compound did not alter phospho JAK1 and JAK2 levels in HDLM 2, MDA MB 468, and DU145 cells. In addition, NSC114792 did not inhibit IFN a induced TYK2 phosphorylation in U266 cells at the concentrations up to 20 umol/L. As expected, AG490 profoundly reduced the phosphorylation levels of all JAKs tested in those (-)-MK 801 Maleate cells. Our results thus far indicate that NSC114792 selectively inhibits JAK3. To assess the functional outcome of this inhibition, we monitored the phosphorylation of a JAK3 target. We chose STAT3, which is phosphorylated by JAKs on Y705, as its persistent activation is the most common STAT form found in human cancers. We found that NSC114792 inhibits phospho STAT3 levels in a dose dependent manner in L540 cells, which have elevated phospho JAK3 levels.

In contrast, at the concentrations up to 20 umol/L, NSC114792 did not inhibit the phosphorylation of STAT3 in cells that lack persistently active JAK3.