Differential proteomic analysis reveals that EGCG inhibits HDGF and activates apoptosis to increase the sensitivity of non-small cells lung cancer to chemotherapy.

PURPOSE: To search for regulated proteins in response to green tea (-)-epigallocatechin-3-gallate (EGCG) in A549 lung cancer cells. EXPERIMENTAL DESIGN: 2DE and ESI/multistage MS (ESI-MS/MS) were performed to identify modulated proteins in A549 cells treated with EGCG. Cell migration was evaluated by transwell assays. RNA interference was used to silence the hepatoma-derived growth factor (HDGF). Caspase-3, caspase-9, and HDGF were immunodetected by Western blot assays. Flow cytometry was used for detection of mitochondrial membrane potential and apoptosis. RESULTS: We found that HDGF expression was threefold suppressed by EGCG treatment. Downregulation of HDGF by EGCG was confirmed using anti-HDGF antibodies in three lung cancer cell lines. EGCG treatment and HDGF abrogation by RNA interference resulted in a decreased migration of A549 cells. In addition, EGCG induced a marked synergistic effect with cisplatin in cell death. Consistently, an enhanced cytotoxicity in HDGF-silenced cells was also found. Cell death was associated to increased apoptosis, disruption of the mitochondrial membrane potential, and activation of caspase-3 and caspase-9. CONCLUSION AND CLINICAL RELEVANCE: Our data suggest for the first time that abrogation of HDGF by EGCG enhances cisplatin-induced apoptosis and sensitize A549 cells to chemotherapy. Therefore, we propose that decreasing the HDGF levels by using EGCG may represent a novel strategy in lung cancer therapy.

RAD50 targeting impairs DNA damage response and sensitizes human breast cancer cells to cisplatin therapy.

In tumor cells the effectiveness of anti-neoplastic agents that cause cell death by induction of DNA damage is influenced by DNA repair activity. RAD50 protein plays key roles in DNA double strand breaks repair (DSBs), which is crucial to safeguard genome integrity and sustain tumor suppression. However, its role as a potential therapeutic target has not been addressed in breast cancer. Our aim in the present study was to analyze the expression of RAD50 protein in breast tumors, and evaluate the effects of RAD50-targeted inhibition on the cytotoxicity exerted by cisplatin and anthracycline and taxane-based therapies in breast cancer cells. Immunohistochemistry assays on tissue microarrays indicate that the strong staining intensity of RAD50 was reduced in 14% of breast carcinomas in comparison with normal tissues. Remarkably, RAD50 silencing by RNA interference significantly enhanced the cytotoxicity of cisplatin. Combinations of cisplatin with doxorubicin and paclitaxel drugs induced synergistic effects in early cell death of RAD50-deficient MCF-7, SKBR3, and T47D breast cancer cells. Furthermore, we found an increase in the number of DSBs, and delayed phosphorylation of histone H2AX after cisplatin treatment in RAD50-silenced cells. These cellular events were associated to a dramatical increase in the frequency of chromosomal aberrations and a decrease of cell number in metaphase. In conclusion, our data showed that RAD50 abrogation impairs DNA damage response and sensitizes breast cancer cells to cisplatin-combined therapies. We propose that the development and use of inhibitors to manipulate RAD50 levels might represent a promising strategy to sensitize breast cancer cells to DNA damaging agents.