MCF 7 and BT 474 breast cancer cell lines have mutated forms of PIK3CA, which can render these cells resistant to Ly294002 mediated cell cycle arrest. Surprisingly, Ly294002 treatment resulted full report in G1 arrest in MCF 7 cells in addition to SK BR 3 cell line. This discrepancy might be explained by the possibility that MCF 7 cell line harbor mutation with no activating function of PI3K. In contrast, no arrest was detected in BT 474, MDA 361 and MDA 436 cells. Taken together, our results confirm the previously observed effects of these inhibitors on cell cycle and sug gest that different breast cancer cell lines have different biological responses to PI3K mTOR pathway inhibitors. especially MDA 436 seems to be resistant to rapamycin and Ly294002 induced cell cycle arrest.

The inhibition of PI3K mTOR pathway with Ly294002 and rapamycin led to similar gene expression alterations in different breast cancer cell lines. Altogether, Inhibitors,Modulators,Libraries 38% Inhibitors,Modulators,Libraries of the differentially expressed genes were altered by both treat ments. Additionally, a number of genes known to be asso ciated with PI3K mTOR pathway were differentially Inhibitors,Modulators,Libraries expressed. For example, the down regulation of eIF4G1 in response to rapamycin and Ly294002 treatments was also shown at protein level. This suggests that PI3K mTOR pathway inhibition leads to the transcriptional deregula tion of a number of critical components of the transla tional machinery. Unlike with PI3K or mTOR inhibition, direct suppression of p70S6K did not seem to down regu late genes involved in eIF 4F initiation complex.

This might be due to the fact that p70S6K is known to regulate the rate of Inhibitors,Modulators,Libraries translation of transcripts encoding elongation factors and ribosomal proteins, but inhibition of p70S6K do not affect on transcriptional activation of these genes. Gene Ontology Categorizer and recently pub lished Connectivity Map were further used to explore the biological processes affected by PI3K mTOR pathway inhibition and drugs with similar mechanism of action. Indeed, GO categories involved in cell killing, mitosis, and G1 phase of the cell cycle were enriched in Ly294002 treated cells, Inhibitors,Modulators,Libraries whereas functional categories like mitosis and translational elongation were among the most enriched classes with lowest p values in rapamycin treated cells.

Also Connectivity Map gave the highest Ponatinib dna scores for Ly294002 and rapamycin in the breast cancer cell lines treated with these inhibitors further validating the gene expression profiles responsive to these PI3K and mTOR inhibitors. Also wortmannin scored high in Con nectivity Map, which is expected, due to its mechanism as a PI3K inhibitor. Other drugs with high statistical signifi cance included rottlerin, a protein kinase inhibitor, and trichostatin A, a known HDAC inhibitor, both of which are known to inhibit proteins interacting with PI3K mTOR pathway.