| In spite of being preventable,the morbidity and mortality of cervical cancer have remained been high,particularly,in some so-called "third world" countries with underdeveloped economy.The current treatment for cervical cancer has begun to take shape.Surgery,radiotherapy and chemotherapy will be implemented based on the patient's situation.Adjuvant therapy helps to enhance the anti-tumor effect of chemotherapy.For advanced and recurrent cervical cancer patients,the main treatment is still chemotherapy.Paclitaxel is one of the most effective chemotherapy drugs for cervical cancer.Paclitaxel is widely recognized as a microtubule stabilizer.It was also reported that Paclitaxel can induce DNA damages,however,the mechanism remains unclear.In the long time chemotherapy,acquired chemoresistance to Paclitaxel obviously limits the efficacy in cervical cancer treatment.One explanation on resistance to Paclitaxel is the high efficiency of DNA damage repair system in tumor cells,making DNA-damaging chemotherapeutic drugs less effective.So we infer that combination of Paclitaxel with the inhibitor targeting DNA repair pathway may greatly enhance the anti-tumor effect of Paclitaxel.FEN1,an important protein in DNA repair system,participating in LP-BER as well as double-strand break repair,plays a major role in RNA primer removal during Okazaki fragment maturation of the lagging strand during DNA replication.In view of the dual role of FEN1 in DNA repair and DNA replication,we targeted FEN1 protein and synthesized a novel FEN1 inhibitor SC13.Our study showed that the combination of Paclitaxel and SC 13 had a synergistic effect in cervical cancer treatment.We first examined the FEN1 expression in cervical cancer and found that the both of mRNA and protein of FEN1 in cervical cancer was up-regulated.Next,two kinds of cervical cancer cell lines:Hela and Siha were treated by SC 13.Data showed that Hela was more sensitive to SC 13 compared to Siha.Cell growth inhibition assay and colony formation assay were performed when SC 13 treatment was combined with Paclitaxel in Hela.We found that the combination therapy significantly inhibited Hela cell growth.Isobologram method was used to analyze the synergistic effect in the combination therapy.To investigate the synergistic mechanism of the combination therapy,we first studied the cell cycle distribution in Hela after Paclitaxel and SC 13 treatment.Results showed that the cell cycle in the combination group was blocked in the S phase and the G1 phase.Next,cell apoptosis was examined as the Hela cells were found to be obviously killed during the combined treatment.Results showed that the percentage of apoptotic cells increased significantly in the combination group.To further explore the mechanism in molecular level,we examined DNA damage in Hela after Paclitaxel and SC 13 combination treatment.Detection of DNA damage markers showed that ?H2AX and 53BP1 were significantly increased in the combination group.The accumulation of DNA double-strand breaks may cause chromosomal fragmentation,so we next examined the number of abnormal chromosomes.More anomalous chromosomes were observed after combination therapy compared to other treatments.We injected Hela cells into the nude mice subcutaneously to construct a nude mice xenograft model.Results showed slight effects on tumor growth inhibition after the use of SC 13 alone or Paclitaxel alone.Significant inhibitory effect was observed after treatment with the combination of two compounds.In addition,immunohistochemistry assay was performed to show that more apoptosis were observed and more DNA damages were accumulated in the combination group compared to the mono-treatment group or the untreated group which consisted with the results of cell experiments.Above all,SC 13 as an inhibitor of FEN1 can be used as a potent sensitizer for the Paclitaxel treatment either in vitro or in vivo.Our study contributes to the idea of combining the bio-targeted therapy with the chemotherapy and makes an effort to optimize the treatment in cervical cancer. |
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