ABSTRACT
Ectopic expression of mutant forms of phosphatase and tensin homologue deleted on chromosome 10 (PTEN) lacking lipid (G129E) or lipid and protein (C124S) phosphatase activity decreased sensitivity of MCF-7 breast cancer cells, which have wild-type PTEN, to doxorubicin and increased sensitivity to the mammalian target of rapamycin (mTOR) inhibitor rapamycin. Cells transfected with a mutant PTEN gene lacking both lipid and protein phosphatase activities were more resistant to doxorubicin than cells transfected with the PTEN mutant lacking lipid phosphatase activity indicating that the protein phosphatase activity of PTEN was also important in controlling the sensitivity to doxorubicin, while no difference was observed between the lipid (G129E) and lipid and protein (C124S) phosphatase PTEN mutants in terms of sensitivity to rapamycin. A synergistic inhibitory interaction was observed when doxorubicin was combined with rapamycin in the phosphatase-deficient PTEN-transfected cells. Interference with the lipid phosphatase activity of PTEN was sufficient to activate Akt/mTOR/p70S6K signaling. These studies indicate that disruption of the normal activity of the PTEN phosphatase can have dramatic effects on the therapeutic sensitivity of breast cancer cells. Mutations in the key residues which control PTEN lipid and protein phosphatase may act as dominant-negative mutants to suppress endogenous PTEN and alter the sensitivity of breast cancer patients to chemo- and targeted therapies.
Subject(s)
Breast Neoplasms/enzymology , Drug Resistance, Neoplasm , Mutation, Missense , PTEN Phosphohydrolase/metabolism , Protein Kinases/metabolism , Signal Transduction , Amino Acid Substitution , Antibiotics, Antineoplastic/agonists , Antibiotics, Antineoplastic/pharmacology , Antibiotics, Antineoplastic/therapeutic use , Breast Neoplasms/drug therapy , Breast Neoplasms/genetics , Cell Line, Tumor , Doxorubicin/agonists , Doxorubicin/pharmacology , Doxorubicin/therapeutic use , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Drug Synergism , Female , Gene Expression , Humans , PTEN Phosphohydrolase/antagonists & inhibitors , PTEN Phosphohydrolase/genetics , Protein Kinases/genetics , Proto-Oncogene Proteins c-akt/genetics , Proto-Oncogene Proteins c-akt/metabolism , Ribosomal Protein S6 Kinases, 70-kDa/genetics , Ribosomal Protein S6 Kinases, 70-kDa/metabolism , Signal Transduction/drug effects , Signal Transduction/genetics , Sirolimus/agonists , Sirolimus/pharmacology , Sirolimus/therapeutic use , TOR Serine-Threonine Kinases , TransfectionABSTRACT
The Insulin-like growth factor-1 receptor (IGF-1R) is overexpressed in a variety of tumors including breast, prostate and myeloma. Thus, IGF-1R and its downstream signaling effectors are good candidates for molecular-based targeted antitumor therapies. Indeed, protein inhibitors of IGF-1R signaling and IGF-1R blocking antibodies are undergoing clinical trials. Herein, the molecular basis for antibody-mediated IGF-1R signal inhibition has been investigated in a hematopoietic cell line model, FDC-P1, that has been rendered interleukin-3 independent in a ligand-dependent manner through retroviral-mediated expression of IGF-1R (FD/IGF-1R). Furthermore, the ability of an anti-IGF-1R antibody to synergize with signal-transduction pathway inhibitors and induce apoptosis was determined. The alphaIGF-1R antibody, A12, was capable of arresting IGF-1 or insulin-induced FD/IGF-1R cell proliferation in the G1 phase of the cell cycle and resulted in apoptotic induction. A12 effectiveness could be potentiated through combination treatment with small molecule inhibitors of the Ras/Raf/MEK/ERK or PI3K/Akt/mTOR pathways. These results validate the use of the FD/IGF-1R cells to evaluate the effectiveness and mechanisms of targeted IGF-1R therapeutic strategies.
