425

The candidate tumor suppressor PTEN (phosphatase and tensin homolog)/MMAClITEPl encodes a protein tyrosine phosphatase known to play a critical role in regulating signaling pathways involved in cell growth and apoptosis. Functional loss of PTEN has been implicated in the onset and progression of cancer including glioblastoma and endometrial, lung, breast, and prostate cancer. PTEN acts as a negative regulator of the phosphoinositide-3-kinase (PI3K)/ Akt signaling pathway, suppressing activation of proliferative and anti-apoptotic processes. Inactivating mutations in the PTEN gene or deletion of one or both PTEN alleles occur with high frequency and can result in constitutive activation of Akt signaling pathways potentiating inhibition of apoptosis, hyperplasia, and tumorigenesis. The insulin-like growth factor receptor (IGF-IR) is known to play a critical role in normal development and tissue homeostasis, but is also recognized as an important signaling molecule implicated in the development and maintenance of cancer. As a result, targeted therapies inhibiting this pathway are being investigated in clinical testing. Since the PI3K/Akt pathway is one of the major mechanisms for transducing the IGF-IR signal, it is possible that deregulation of this pathway by loss of PTEN may effect reduced sensitivity of tumor cells to therapy mediated by IMC-A12, a fully human (IgG1λ.) monoclonal antibody (mAb) antagonist of IGF-IR. We have observed that certain PTEN negative cancer cell lines which exhibit constitutive phosphorylation of Akt in serum starved cells do not respond to IGF-IR therapy. We were therefore interested in determining if abrogation of PTEN in an IGF-IR responsive tumor line would confer reduced sensitivity to targeted anti-IGF-IR mAb therapy. In this study, we utilized the MCF7 human breast cancer line, which is highly responsive to anti-IGF-IR mAb treatment. To study the effect of PTEN loss, we generated PTEN knockdown cell lines by siRNA. Western analysis demonstrated a greater than 80% reduction in total PTEN levels in a representative MCF7 siRNA transfected cell line. In proliferation and soft-agar clonal assays responding to ligand or to serum, anti-IGF-IR mAb IMC-A12 inhibited the growth of both mock transfected and PTEN knockdown cells. Western blot analysis demonstrated that IMC-A12 inhibited AKT phosphorylation regardless of PTEN status indicating that Akt activation in MCF7 cells occurs in response to IGF-IR signaling and thus is sensitive to targeted anti-IGF-IR mAb therapy. To the contrary, if MCF7 cells were transfected with a constitutively active Akt (myristylated-Akt), IMC-A12 did not inhibit cell proliferation or growth in soft agar. These results demonstrate that sensitivity to IGF-IR therapy is indicated in tumor cells in which Akt activation is mediated by IGF signaling, irrespective of PTEN status.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA