Phosphatidylinositol 3-Kinase Inhibition Broadly Sensitizes Glioblastoma Cells to Death Receptor– and Drug-Induced Apoptosis

Abstract

The aberrant activity of the phosphatidylinositol 3-kinase (

The pyridinylfuranopyrimidine inhibitor, PI-103, chemosensitizes glioblastoma cells for apoptosis by inhibiting DNA repair

“…Furthermore, we previously showed that inhibition of PI3K sensitizes GBM to chemotherapy-and to death receptor-induced apoptosis (Opel et al, 2008). A similar importance of the PI3K/Akt pathway has also been reported in ovarian, gastric, breast and lung cancer (Jiang and Liu, 2008), as well as in neuroblastoma, for which it was recently shown that aberrant activation of Akt correlates with reduced life expectancy (Opel et al, 2007).…”

“…This might also explain in part the reduced sensitization effect of PI-103 when combined with temozolomide, which is an alkylating agent (Tisdale, 1985) and damages DNA by generating O 6 -alkylguanine, which is repaired primarily independent of DNA-PK (Christmann et al, 2003). Indeed, inhibition of PI3K-mediated signaling also enhances apoptosis sensitivity in GBM cells in an NHEJ-independent manner, as we have previously shown for agents that do not primarily cause DNA damage, that is, death receptor ligands such as TRAIL and anti-CD95 agonistic antibody (Opel et al, 2008). Intriguingly, in line with recent observations regarding the pharmacokinetics of inhibitor-induced cancer cell death (Shah et al, 2008), we also found that a brief, but potent inhibition of PI3K administered concurrently with a DNA-damaging drug is sufficient to commit GBM cells to apoptosis and to suppress clonogenic growth.…”

“…To further verify the clinical relevance of our findings, we also used primary cultured GBM cells generated from surgical specimens (Opel et al, 2008). PI3K signaling was inhibited by PI-103 in these cells at similar concentrations to those used in GBM cell lines (Supplementary Figure 2a).…”

“…Western blot analysis was carried out as previously described (Opel et al, 2008), using the following antibodies: mouse monoclonal Akt (BD Biosciences, Heidelberg, Germany); rabbit polyclonal phospho-Akt (Ser473), rabbit phospho-S6 ribosomal protein (Ser235/236), rabbit anti-S6 ribosomal protein (Cell Signaling, Beverly, MA, USA); mouse monoclonal b-actin (Sigma-Aldrich), followed by anti-mouse or anti-rabbit immunoglobulin G-horseradish peroxidase from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Enhanced chemiluminescence was used for detection (Amersham Bioscience, Freiburg, Germany).…”

“…Cells were transfected with siRNA using TransMessenger transfection reagent (Qiagen, Hilden, Germany) as previously described (Opel et al, 2008), using the following Stealth RNAi constructs from Invitrogen: PIK3CA (PIK3CAHSS10800 6) and PIK3CB (PIK3CBHSS10800 7) for the p110a and b subunits of PI3K; PRKDC (PRKCDHSS10852 7) for the catalytic subunit of DNA-PK; FRAP1 (FRAP1HSS103825 5-7) for mTOR; and Stealth Negative Universal Control Medium RNAi (12935-300) for negative control.…”