Synergistic tumor growth inhibition by combined treatment with green tea epigallocatechin-3-gallate (EGCG) and an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), erlotinib in squamous cell carcinoma of the head and neck (SCCHN)
Authors: Xin Zhang, John E. Lee, Mourad Tighiouart, Hyung J. Shin, Fadlo R. Khuri, Zhuo (georgia) Chen, Dong M. Shin
Journal: Cancer Research (2006)

Proc Amer Assoc Cancer Res, Volume 47, 2006

4892 

EGCG is the main polyphenol extracted from green tea. It has shown anti-tumor or chemopreventive activity in several tumors including SCCHN by regulating EGFR-mediated pathways. We hypothesize that a combination of EGCG and an EGFR-TKI (erlotinib) may synergistically inhibit tumor growth of SCCHN. Five head and neck cancer cell lines (Tu212, Tu177, 886LN, SQCCY1, and 38) were used for this study. Cell growth inhibition assay showed that treatment with two drugs as single agent for 72 hrs inhibited growth of all five SCCHN cell lines in a dose dependent manner. Combination index (CI) analysis demonstrated that the combined treatment of EGCG with erlotinib for 72 hrs achieved synergistic effects (CI < 1) of cell growth inhibition on four of five cell lines except an additive effect on 886LN. We selected two representative cell lines, Tu212 and Tu177 to further investigate the underlying mechanisms using fixed concentration of EGCG (30 μM) and erlotinib (0.5 μM). Cell cycle analysis showed a significant G1 arrest after the combined treatment of the two cell lines for 24 and 48 hrs compared to either the single treatment or the control group. Annexin-V binding assay demonstrated more apoptotic cells in the combined treatment group compared to either the single agents or the control group. Immunoblotting analyses showed that the combined treatment significantly reduced the phosphorylation of EGFR and AKT. The combined treatment also induced activation of caspase-8, caspase-9, caspase-3 and PARP compared to either the single agents or the control group. In our in vivo study, twenty five nude mice were randomly divided into four groups. Each group was orally gavaged with the vehicle control (1% tween 80), EGCG (125 mg/kg), erlotinib (50 mg/kg) or a combination of EGCG and erlotinib at the same doses for 7 days followed by a subcutaneous injection with 2 × 106 of Tu212 cells. The animals were continuously administered the agents 5 days a week. The tumor size in each mouse was measured three times per week until all mice were sacrificed once the tumor size of any mouse reached 2cm in diameter. Our results showed that the combination treatment (443 ±120 mm3) significantly inhibited the tumor growth compared to the control (2743 ± 910 mm3), EGCG alone (1198 ± 406 mm3) or erlotinib alone (1176 ± 392 mm3), (p = 0.005). The mean time (days) to reach a tumor volume of 400 mm3 was significantly delayed in the combined treatment (34.2 ± 2.4) as compared with the control (21.2 ± 2.8), EGCG (23.7 ± 2.1), or erlotinib (23.7 ± 1.3), (p = 0.005). Taken together, both our in vitro and in vivo results demonstrate significant anti-tumor activity for the combined treatment with EGCG and erlotinib. This combination provides a novel and promising chemopreventive regimen for SCCHN. (Supported by NCI UO1 101244)