Heat shock protein 90 inhibitor 17‐allylamino‐17‐demethoxygeldanamycin potentiates the radiation response of tumor cells grown as monolayer cultures and spheroids by inducing apoptosis

Abstract

Activation of the PI3K-Akt pathway is known to induce tumor radioresistance. In the current study, we examined the ability of 17AAG, which decreases the levels of Hsp90 client proteins including components of the PI3K-Akt pathway, to sensitize radioresistant human squamous cell carcinoma cells to Xirradiation. Human squamous cell carcinoma cell lines (SQ20B, SCC61 and SCC13) were incubated for 16 h at 37°°°°C in medium containing 17AAG. Radiation sensitivity was determined by clonogenic assays, and protein levels were examined by western blotting. Apoptosis was determined in monolayer cells by AO/EB double staining and in spheroids using the TdT-mediated dUTP nick end labeling assay. 17AAG (0.2 µ µ µ µM) enhanced the radiosensitivity more effectively in radioresistant SQ20B and SCC13 cells than in radiosensitive SCC61 cells. However, in all three cell lines, 17AAG increased radiation-induced apoptosis by reducing the expression of EGFR and ErbB-2 and inhibiting the phosphorylation of Akt. Furthermore, 17AAG (1 µ µ µ µM) sensitized SQ20B spheroids to radiation, and inhibition of Akt activation by 17AAG increased radiation-induced apoptosis in spheroids. The findings suggest that 17AAG effectively sensitizes radioresistant cells to radiation by inhibiting the PI3K-Akt pathway. Targeting the PI3K-Akt pathway with 17AAG could be a useful strategy for radiosensitization of carcinomas. (Cancer Sci 2005; 96: 911-917) R adiation therapy is commonly used for squamous cell carcinoma of the head and neck because of its minimal cosmetic effects and its ability to preserve voice in laryngeal cancer. However, treatment failure correlates with a poorer prognosis and decreased patient survival.The EGFR family contributes to resistance to radiation and chemotherapy in many human tumors, including head and neck cancer.(1-14) ErbB-2, a member of the EGFR family, is also overexpressed in 30% of breast tumors as well as in a significant number of other cancers. (15,16) Blockade of the EGFRmediated signaling pathway enhances the sensitivity of tumor cells to radiation and several chemotherapeutic agents.The PI3K-Akt pathway is a major downstream pathway initiated by activation of the EGFR family members. (17)(18)(19)(20)(21) This pathway plays an important role in cell growth and survival. (22,23) The serine/threonine kinase Akt (also known as protein kinase B) is thought to be a downstream target of PI3K. Akt activation is responsible for desensitizing cells to apoptotic stimuli. This occurs by regulation of the transcriptional activity of both Forkhead family members (24,25) and NF-κB, (26,27) and through phosphorylation and inactivation of the apoptotic machinery including Bcl-2 homolog, Bad (28,29) and caspase-9.(30) Furthermore, Akt is amplified or overexpressed in a wide variety of human tumors.(31-33) PTEN, a tumor suppressor gene, is thought to negatively regulate the PI3K-Akt pathway.(34-36) Mutations causing PTEN to be functionally inactive are frequently detected in many human cancers and enhance Akt activity.(37) Akt acti...

Hsp90 is expressed and represents a therapeutic target in human oesophageal cancer using the inhibitor 17-allylamino-17-demethoxygeldanamycin

“…A further observation was that SER was larger for Kyse450 than for Kyse70, 2.1 and 1.5, respectively. It should be noted that Kyse450 is more radioresistant than Kyse70, and our results are thus consistent with reports that 17-AAG has a better sensitisation effect on radioresistant cell lines (Russell et al, 2003;Machida et al, 2005). Inhibition of the EGFR, which is highly expressed in these cell lines and frequently expressed at high levels in oesophageal tumours, did not influence the radiosensitivity, although we could observe an effective inhibition of EGFR signalling after gefitinib treatment (data not shown).…”

BIIB021, a novel Hsp90 inhibitor, sensitizes head and neck squamous cell carcinoma to radiotherapy

“…In an experiment with 17-AAG in 3 HNSCC cell lines (SQ20B, SCC61 and SCC13), they found that 17-AAG enhanced radiosensitivity more effectively in radioresistant tumor cells than in radiosensitive cells. 31 This effect was observed in both monolayer culture and spheroids and again was achieved by inhibiting the PI3K-Akt pathway. Taken together, these data indicate a general ability of Hsp90 inhibitors to potentiate the effects of radiation in HNSCC cells, given that the tumor cell lines used in each study have diverse genetic backgrounds, including differences in p53 status.…”

Dual induction of PKR with E2F-1 and IFN-α to enhance gene therapy against hepatocellular carcinoma

“…Antibodies and reagents E2F-1 (KH95) and PKR (K-17) antibodies were purchased from SantaCruz Biotechnology (SantaCruz Biotechnology Inc., Heidelberg, Germany), b-actin [20][21][22][23][24][25][26][27][28][29][30][31][32][33] from Sigma (Sigma, Buchs, Switzerland), phospho-PKR (pT451) from Biosource (Biosource Europe S A, Nivelles, Belgium).…”

“…20,17,25,14 To determine their effect on HCC cells, viability assays were performed in the Hep3B cell line. Concentrations were chosen based on previously reported studies.…”

“…Concentrations were chosen based on previously reported studies. [25][26][27][28][29] Increasing concentrations of the PKR modifiers radicicol, heparin and 17-AAG reduced cell viability in a time and dose-dependent manner (Figures 2a-c), whereas IFNa showed no cytotoxic effect (Figure 2d). …”