nutlin-3a and Esophageal-Neoplasms

Esophageal squamous cell carcinoma (ESCC) is highly resistant to chemoradiation therapy. We aimed to examine whether Nutlin-3, a molecule that suppresses murine double min 2 (MDM2)-mediated p53 and Retinoblastoma (RB) protein degradation leading to downregulation of DNA methyltransferases (DNMTs), can be a novel therapeutic agent for ESCC. We used wild-type and chemoradiation-resistant ESCC cell lines in this study. The expression of DNMTs, p53 and RB, and methylation level of tumor suppressor genes (TSG) were analyzed upon Nutlin-3 treatment. The antitumor efficacy of Nutlin-3 was investigated in ESCC cell lines and xenograft tumor model. TSG protein expression was checked in the excised tumor tissue. Nutlin-3 induced upregulation of p53 and RB and downregulation of DNMTs proteins in the chemoradiation-resistant and aggressive ESCC cells. The methylation level of TSGs was decreased by Nutlin-3. Nutlin-3 inhibits clonogenic growth of ESCC cells and exerts a synergistic cytotoxic-effect when combined with chemotherapeutic agent cisplatin. Moreover, xenograft tumor growth in SCID mice was suppressed by Nutlin-3. The protein expression level of DNMTs was downregulated, and that of TSGs was upregulated by Nutlin-3 treatment in the excised tumor tissue. In conclusion, Nutlin-3 is a potential therapeutic agent that can potentiate the treatment efficacy of chemoradiation-resistant ESCC.

Topics: Animals; Apoptosis; Cell Line, Tumor; DNA; Enzyme Inhibitors; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Humans; Methyltransferases; Mice; Mice, SCID; Proto-Oncogene Proteins c-mdm2; Retinoblastoma Protein; Tumor Suppressor Protein p53

Murine double minute 2 (MDM2) is well known to inhibit p53 function and its over-expression is associated with poor prognosis in several human malignancies. Nutlin-3, a small-molecule inhibitor of MDM2, exerts antitumor effects on various solid tumors harboring wild-type p53. We aimed to clarify its effects on esophageal cancer.. We first examined the potential antitumor effects of nutlin-3 according to MDM2 status using esophageal carcinoma cell lines (KYSE 170/180). We then immunolocalized MDM2 immunoreactivity in 62 surgical cases of esophageal squamous cell carcinoma undergoing neoadjuvant chemotherapy followed by esophagectomy and correlated the findings with clinicopathological variables.. MDM2 mRNA expression in KYSE 170 was significantly higher than that in KYSE 180 cells. No significant changes were detected in both cell lines when nutlin-3 was added. However, cell proliferation was significantly decreased in KYSE 170 cells treated with nutlin-3 and cisplatin compared to cisplatin alone but not in KYSE 180. MDM2 immunoreactivity was also significantly associated with poor sensitivity to neoadjuvant chemotherapy in the cases examined.. The combination of nutlin-3 with chemotherapeutic agents may become a novel therapeutic strategy in esophageal cancer over-expressing MDM2.

Topics: Cisplatin; Esophageal Neoplasms; Esophageal Squamous Cell Carcinoma; Humans; Imidazoles; Piperazines; Proto-Oncogene Proteins c-mdm2; Tumor Suppressor Protein p53

Murine double minute 2 (MDM2) negatively regulates the activity of the p53 protein and plays a vital role in cell cycle arrest, apoptosis, and senescence mediated by p53. Nutlin-3, an antagonist of MDM2, is frequently used in anti-cancer studies. In many human tumors, nutlin-3 stabilizes p53 status and enhances p53 expression in cells with wild-type p53. However, the effect of nutlin-3 combined with radiotherapy on esophageal squamous cancer (ESCC) has not been reported. In this study, we examined whether nutlin-3 increases the radiosensitivity of ESCC in vitro and in vivo.We chose two cell lines, ECA-109 (wild-type p53) and TE-13 (p53 mutated), for the following experiments. Cell proliferation and clonogenic survival experiments showed that nutlin-3 inhibits the cell growth and colony formation of ECA-109 cells in a dose-dependent manner. Flow cytometry analysis showed that the apoptosis rate of ECA-109 cells co-treated with nutlin-3 and irradiation(IR) was significantly increased compared with cells treated with irradiation or nutlin-3 alone. Western blotting detected the expression of apoptosis-associated proteins in ECA-109 cells in response to nutlin-3 and irradiation. These effects were not evident in TE-13 cells. Xenograft mouse models indicated that nutlin-3 suppresses tumor growth and promotes radiosensitivity in the ESCC cell line ECA-109 in vivo. We have demonstrated that co-treatment of nutlin-3 with irradiation can significantly inhibit the growth and improve the radiosensitivity of ESCC cells with wild-type p53. The study suggests that nutlin-3 may be a potent therapeutic agent in conjunction with radiotherapy in ESCC.

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Carcinoma, Squamous Cell; Cell Proliferation; Esophageal Neoplasms; Gamma Rays; Humans; Imidazoles; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Mutation; Piperazines; Proto-Oncogene Proteins c-mdm2; Radiation Tolerance; Tumor Cells, Cultured; Tumor Suppressor Protein p53; Xenograft Model Antitumor Assays