the effects of RAD001 compared to rapamycin on Akt phosphorylation in a band of lung HCV protease inhibitor cancer cell lines following a prolonged treatment. Both RAD001 and rapamycin at 10 nM increased p Akt levels while inhibiting p70S6K phosphorylation in every of the cell lines after a 24 h treatment. We also handled H157 and A549 lung cancer cells with 1 nM RAD001 or rapamycin for an extended time frame from 24 to 96 h and then harvested the cells for analysis of Akt phosphorylation. As shown in Fig. 1B, p Akt levels remained elevated at most of the tested times on the extended period of time, even if reduced p p70S6K levels returned at 96 h. This result plainly demonstrates mTOR inhibitors induce a sustained Akt activation in the tested cell lines. We observed that g p70S6K degrees restored at 96 h post treatment with RAD001, but not with rapamycin. Since we treated cells only one time, it’s likely that rapamycin could have a lengthier half-life in cell culture than RAD001, resulting in better efficiency than RAD001 in inhibiting mTOR signaling. More over, we examined the results of prolonged treatment with rapamycin or RAD001 on Akt phosphorylation in two cell lines, where Akt phosphorylation was diminished by prolonged treatment with Urogenital pelvic malignancy rapamycin, in an even more detailed way. Past studies used 100 nM rapamycin or 1000 nM CCI 779, which reduced g Akt levels following a 24 h treatment. In our study, we’re able to continue this result after both 24 and 48 h solutions with 100 nM rapamycin in PC 3 cells. But, once the attention of rapamycin was paid off to 1 nM, we consistently observed a rise in Akt phosphorylation at both 24 h and 48 h solutions. CX-4945 solubility Similar results were also obtained from cells treated with RAD001. In though at 10 nM or 100 nM p Akt levels were decreased by them U937 cells, prolonged therapy with either 1 nM rapamycin or RAD001 plainly enhanced the levels of p Akt. Similar results with RAD001 were also noticed in Jurkat cells. We noted that both rapamycin and RAD001 at 1 nM effectively inhibited mTORC1 signaling evidenced by reduction of p S6 or p p70S6K levels. Thus, the results of prolonged therapy with mTOR inhibitors on Akt phosphorylation are clearly dose dependent in these cell lines. We also noted that both rapamycin and RAD001 at 1-100 nM improved Akt phosphorylation at Thr308 in a dose dependent manner in PC 3 cells, indicating that mTOR inhibitors also stimulate PDK1 kinase. We mentioned that our information here on Akt phosphorylation at Thr308 by rapamycin or RAD001 in PC 3 cells are different from previous report that rapamycin at 100 nM somewhat diminished Akt phosphorylation at Thr308 after having a 24 h treatment. The cause of this inconsistency isn’t clear, but might be as a result of various ways the cells were treated by us and other investigators.