pki-587 and Squamous-Cell-Carcinoma-of-Head-and-Neck

Radiation remains a mainstay for the treatment of nonmetastatic head and neck squamous cell carcinoma (HNSCC), a malignancy characterized by a high rate of PI3K/mTOR signaling axis activation. We investigated the ATP-competitive dual PI3K/mTOR inhibitor, PF-05212384, as a radiosensitizer in preclinical HNSCC models.. Extent of radiation enhancement of two HNSCC cell lines (UMSCC1-wtP53 and UMSCC46-mtP53) and normal human fibroblast (1522) was assessed by in vitro clonogenic assay with appropriate target inhibition verified by immunoblotting. Radiation-induced DNA damage repair was evaluated by γH2AX Western blots with the mechanism of DNA double-strand break repair abrogation investigated by cell cycle analysis, immunoblotting, and RT-PCR. PF-05212384 efficacy in vivo was assessed by UMSCC1 xenograft tumor regrowth delay, xenograft lysate immunoblotting, and tissue section immunohistochemistry.. PF-05212384 effectively inhibited PI3K and mTOR, resulting in significant radiosensitization of exponentially growing and plateau-phase cells with 24-hour treatment following irradiation, and variable radiation enhancement with 24-hour treatment before irradiation. Tumor cells radiosensitized to a greater extent than normal human fibroblasts. Postirradiation PF-05212384 treatment delays γH2AX foci resolution. PF-05212384 24-hour exposure resulted in an evident G1-S phase block in p53-competent cells. Fractionated radiation plus i.v. PF-05212384 synergistically delayed nude mice bearing UMSCC1 xenograft regrowth, with potential drug efficacy biomarkers identified, including pS6, pAkt, p4EBP1, and Ki67.. Taken together, our results of significant radiosensitization both in vitro and in vivo validate the PI3K/mTOR axis as a radiation modification target and PF-05212384 as a potential clinical radiation modifier of nonmetastatic HNSCC.

Topics: Animals; Carcinoma, Squamous Cell; Cell Cycle; Cell Line, Tumor; Disease Models, Animal; DNA Breaks, Double-Stranded; DNA Repair; Dose-Response Relationship, Radiation; Gene Knockdown Techniques; Head and Neck Neoplasms; Humans; Mice; Morpholines; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Protein Kinase Inhibitors; Radiation Tolerance; Radiation-Sensitizing Agents; Signal Transduction; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Triazines; Tumor Burden; Xenograft Model Antitumor Assays

Head and neck squamous cell carcinomas exhibit variable sensitivity to inhibitors of the PI3K/mTOR pathway, an important target of genomic alterations in this cancer type. The mitogen-activated protein kinase kinase (MEK)/ERK/activator protein 1 (AP-1) and nuclear factor-κB (NF-κB) pathways are also frequently co-activated, but their roles in resistance mechanisms to PI3K/mTOR inhibitors and as therapeutic targets in head and neck squamous cell carcinoma (HNSCC) are not well defined.. We determined the IC50s of dual PI3K/mTOR inhibitor PF-05212384 (PF-384) by XTT assays in 14 HNSCC lines with PI3K/Akt/mTOR cascade alterations. In two resistant models, we further characterized the molecular, cellular, and in vivo attributes and effects of combining PF-384 with MEK inhibitor PD-0325901 (PD-901).. PF-384 IC50s varied between 0.75 and 133 nmol/L in 14 HNSCC lines with overexpression or mutations of PIK3CA, and sensitivity correlated with increased phospho-AKT(T308/S473). In resistant UMSCC-1 and -46 models, PF-384 increased G0-/G1-phase accumulation but weakly induced sub-G0 cell death. PF-384 inhibited direct targets of PI3K/mTOR, but incompletely attenuated co-activated ERK and UMSCC-1 xenograft growth in vivo. PD-901 strongly inhibited MEK/ERK targets, and the combination of PF-384 and PD-901 inhibited downstream NF-κB and AP-1 transactivation, and IL8 and VEGF production in vitro. PD-901 potently inhibited tumor growth alone and with PF384, enhanced antiproliferative, apoptotic, and anti-angiogenesis activity in vivo.. PI3K/mTOR inhibitor PF-384 exhibits variable activity in a panel of HNSCC cell lines with differing PIK3CA expression and mutation status. MEK inhibitor PD-901 overcomes resistance and enhances antitumor effects observed with PF-384 in vivo.

Topics: Animals; Antineoplastic Agents; Benzamides; Carcinoma, Squamous Cell; Cell Line, Tumor; Cell Proliferation; Cell Survival; Cell Transformation, Neoplastic; Cytokines; Diphenylamine; Disease Models, Animal; Drug Resistance, Neoplasm; Drug Synergism; Gene Expression; Genes, Reporter; Head and Neck Neoplasms; Humans; Inflammation Mediators; Mitogen-Activated Protein Kinase Kinases; Morpholines; NF-kappa B; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Protein Kinase Inhibitors; Signal Transduction; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Transcription Factor AP-1; Transcriptional Activation; Triazines; Xenograft Model Antitumor Assays

Cetuximab is the only targeted agent approved for the treatment of head and neck squamous cell carcinomas (HNSCC), but low response rates and disease progression are frequently reported. As the phosphoinositide 3-kinase (PI3K) and the mammalian target of rapamycin (mTOR) pathways have an important role in the pathogenesis of HNSCC, we investigated their involvement in cetuximab resistance.. Different human squamous cancer cell lines sensitive or resistant to cetuximab were tested for the dual PI3K/mTOR inhibitor PF-05212384 (PKI-587), alone and in combination, both in vitro and in vivo.. Treatment with PKI-587 enhances sensitivity to cetuximab in vitro, even in the condition of epidermal growth factor receptor (EGFR) resistance. The combination of the two drugs inhibits cells survival, impairs the activation of signalling pathways and induces apoptosis. Interestingly, although significant inhibition of proliferation is observed in all cell lines treated with PKI-587 in combination with cetuximab, activation of apoptosis is evident in sensitive but not in resistant cell lines, in which autophagy is pre-eminent. In nude mice xenografted with resistant Kyse30 cells, the combined treatment significantly reduces tumour growth and prolongs mice survival.. Phosphoinositide 3-kinase/mammalian target of rapamycin inhibition has an important role in the rescue of cetuximab resistance. Different mechanisms of cell death are induced by combined treatment depending on basal anti-EGFR responsiveness.

Topics: Animals; Antibodies, Monoclonal, Humanized; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Autophagy; Carcinoma, Squamous Cell; Caspase 3; Cell Line, Tumor; Cell Proliferation; Cetuximab; Drug Resistance, Neoplasm; ErbB Receptors; Head and Neck Neoplasms; Humans; Mice; Mice, Inbred BALB C; Mice, Nude; Morpholines; Phosphoinositide-3 Kinase Inhibitors; Squamous Cell Carcinoma of Head and Neck; TOR Serine-Threonine Kinases; Triazines; Xenograft Model Antitumor Assays