The mTOR-targeting drug temsirolimus enhances the growth-inhibiting effects of the cetuximab-bevacizumab-irradiation combination on head and neck cancer xenografts.

We previously reported on head and neck tumor xenografts that the tumor regression induced by a triple combination of irradiation (RT), anti-EGFR and anti-angiogenic therapies was followed, after treatment arrest, by tumor re-growth characterized by activation of the AKT signaling pathway. Since mTOR is the main AKT-related messenger, the aim of this study was to add the mTOR inhibitor temsirolimus to a tri-therapy with RT plus anti-EGFR and anti-angiogenic drugs in order to improve anti-tumor effects. The human head and neck cancer cell line CAL33 (over-expressing EGFR and secreting VEGF-A) was xenografted in nude mice. Treatment (20 mice per treatment group) was administered for 2 weeks and consisted of either vehicle (control), temsirolimus (5mg/kg i.p. five times a week), tri-therapy with RT (6 Gy three times a week) combined with cetuximab (0.5mg/kg i.p. five times a week) and bevacizumab (5mg/kg i.p. five times a week) or the temsirolimus-tri-therapy association. The time to reach a tumor volume of 2000 mm(3) was significantly different between the four treatment groups (Log Rank p<0.0001), with a median of 29.5, 44.5, 67.0 and 70.0 days for control, temsirolimus, tri-therapy and combination groups, respectively. The combination of temsirolimus plus tri-therapy produced the longest growth-inhibiting effects (tri-therapy versus combination, p=0.01). No significant interaction was observed between temsirolimus and the tri-therapy, suggesting that temsirolimus, on the one hand, and RT-cetuximab-bevacizumab, on the other, exert additive effects on tumor growth inhibition. These decreases observed on tumor growth were corroborated by the parallel decreases observed on tumor proliferation (Ki67) and on anti-apoptotic markers (Bcl2). These results suggest that temsirolimus exhibits synergistic antiproliferative effects when administered in combination with irradiation, anti-EGFR and anti-angiogenic therapies in head and neck cancer patients.

Bozec A ，Etienne-Grimaldi MC ，Fischel JL ，Sudaka A ，Toussan N ，Formento P ，Milano G ... -  《-》

Combination of mTOR and EGFR targeting in an orthotopic xenograft model of head and neck cancer.

Recent preclinical and clinical studies on head and neck squamous cell carcinoma (HNSCC) revealed synergistic effects when combining anti-EGFR agents with conventional chemotherapeutic drugs. Activation of the PI3-kinase/AKT/mTOR signaling pathway has been identified as an important mechanism implicated in tumor progression and resistance to EGFR inhibitors. The aim of this study was to investigate the effects of combining the mTOR inhibitor temsirolimus (Tem) with the anti-EGFR agent cetuximab (Cet) and conventional chemotherapeutic drugs (cisplatin and fluorouracil (C/F)) on an orthotopic model of HNSCC. Preclinical in vivo study. We evaluated the anti-tumor efficacy (measured tumor volume) of Tem, Cet, and C/F, administered alone or in combination. Investigations were performed using a human HNSCC cell line, CAL33, injected into the mouth floor of nude mice. As compared with the control, the combination of Tem and Cet led to the highest tumor inhibition and induced almost complete tumor growth arrest (P = 0.001). Tem significantly enhanced the impact of the Cet-C/F combination on tumor growth (P < 0.001). The highest inhibitory effects of treatments on cell proliferation (Ki67 labeling), MAPK (pP42/44 labeling), and PI3K/AKT/mTOR (pS6R labeling) signaling pathways were found with the Tem-Cet association. In this orthotopic HNSCC model, the combination of Tem with Cet produced synergistic effects on tumor growth. These results were corroborated by a strong inhibition of both MAPK and PI3K-mTOR signaling pathways. N/A.

Bozec A ，Ebran N ，Radosevic-Robin N ，Sudaka A ，Monteverde M ，Toussan N ，Etienne-Grimaldi MC ，Nigro CL ，Merlano M ，Penault-Llorca F ，Milano G ... -  《-》

Sustained antitumor activity by co-targeting mTOR and the microtubule with temsirolimus/vinblastine combination in hepatocellular carcinoma.

The mammalian target of rapamycin (mTOR) and the microtubules are prominent druggable targets for hepatocellular carcinoma (HCC). PI3K/Akt/mTOR activation is associated with resistance to microtubule inhibitors. Here, we hypothesized that co-targeting of mTOR (by mTOR inhibitor temsirolimus) and the microtubule (by microtubule-destabilizing agent vinblastine) would be more efficacious than single targeting in HCC models. In vitro studies showed that effective inhibition of mTOR signaling with temsirolimus alone was able to suppress HCC cell growth in a dose-dependent manner. Among five cell lines tested, Huh7 was the most temsirolimus-sensitive (IC(50)=1.27±0.06μM), while Hep3B was the most temsirolimus-resistant (IC(50)=52.95±17.14μM). We found that co-targeting of mTOR (by temsirolimus) and the microtubule (by vinblastine, at low nM) resulted in marked growth inhibition in Huh7 cells and synergistic growth inhibition in Hep3B cells (achieving maximal growth inhibition of 80-90%), demonstrating potent antitumor activity of this novel combination. In vivo studies showed that temsirolimus treatment alone for 1 week was able to inhibit the growth of Huh7 xenografts. Strikingly, the temsirolimus/vinblastine combination induced a significant and sustained antitumor activity (up to 27 days post-treatment), with effective reduction of tumor vessel density in both Huh7 and Hep3B xenograft models. Mechanistic investigation revealed that this marked antitumor effect was accompanied by specific and concerted down-regulation of several key anti-apoptotic/survival proteins (survivin, Bcl-2, and Mcl-1), which was not observed in single agent treatments. Our findings demonstrated that the potent anti-cancer activity of this co-targeting strategy was indeed mediated in parts by inhibition of these key survival/anti-apoptotic proteins.

Zhou Q ，Lui VW ，Lau CP ，Cheng SH ，Ng MH ，Cai Y ，Chan SL ，Yeo W ... -  《-》

Schedule-dependent interaction between temsirolimus and cetuximab in head and neck cancer: a preclinical study.

Lattanzio L ，Milano G ，Monteverde M ，Tonissi F ，Vivenza D ，Merlano M ，Lo Nigro C ... -  《-》

Antitumor activity of cetuximab associated with the taxotere-cisplatin-fluorouracil (TPF) combination on an orthotopic head and neck cancer model.

Cetuximab (Cet), an anti-epidermal growth factor receptor (EGFR) monoclonal antibody, is a standard of care in advanced head and neck (HN) cancer when combined with irradiation or cisplatin. The TPF association combining docetaxel (T), cisplatin (P) and fluorouracil (F) has become a chemotherapy of reference in the management of HN tumors. The aim of the study was to evaluate the anti-tumor activity of the association of Cet with TPF. In addition, the potential benefit from adding bevacizumab (Bev), an anti-VEGF monoclonal antibody, was examined. Investigations were performed on CAL33 human head and neck cancer cell line (high content of EGFR) injected as an orthotopic xenograft into the mouth floor of nude mice. The anti-tumor efficacy of Cet (2.5 mg/kg), Bev (20 mg/kg) and of TPF (at the MTD: T 20 mg/kg, P 6 mg/kg, F 17 mg/kg), administered alone or in combination on day 3 and day 10 after tumor cell injection, was assessed on day 12 (animal sacrifice for ethical reasons, 10 animals per treatment group). Tumor volume, tumor histology, tumor cell proliferation (Ki67, immunochemistry) and apoptosis (Bcl2, immunochemistry) were examined. As compared to controls, TPF-Cet and TPF-Cet-Bev had a significant impact on tumor volume and histological response. The highest efficacy was observed with the TPF-Cet combination. The effects of the TPF-Cet-Bev association were not significantly different from those observed with TPF-Cet. Data were corroborated by a diminution in Ki67 labeling and Bcl2 expression. The association of Cet with TPF can be considered a beneficial clinical option in advanced HN cancer patients.

Bozec A ，Sudaka A ，Etienne-Grimaldi MC ，Brunstein MC ，Fischel JL ，Milano G ... -  《-》