| Objective:Ovarian cancer is often diagnosed at an advanced stage and has already metastasized widely.Invasion and metastasis of ovarian cancer is abundant in fat tissue.A variety of cytokines in tumor microenvironment can regulate cell metabolism and influence the invasion and metastasis of tumor cells.How to excavate new diagnostic markers to predict the occurrence and development of ovarian cancer in advance and how to develop a treatment plan for metastatic targets are important issues to be solved in clinical diagnosis and treatment of ovarian cancer.Recent studies have shown that growth differentiation factor 15(GDF15),as an exocrine protein,is expressed at elevated levels in various malignant tissues and at low levels in normal tissues.Furthermore,the high expression of GDF15 is significantly associated with tumor invasion and metastasis and poor prognosis,while the function of GDF15 in ovarian cancer remains unclear.This study will investigate the function and potential mechanism to phenotype of GDF15 in ovarian cancer.Methods:This study was carried out in the following parts:1.The evaluation of GDF 15 levels on various aspects in ovarian cancer was detected:First of all,this study included 114 cases advanced ovarian serous adenocarcinoma patients as clinical samples.The serum level,mRNA and protein expression levels of GDF 15,were measered by enzyme-linked immunosorbent experiment,quantitative reverse transcription polymerase chain reaction and protein immunoblot respectively.2.Analyze the association between GDF 15 and ovarian cancer prognosis:prognosis data database was established through follow-up.The effects of serum GDF 15 and protein levels on the prognosis of ovarian cancer patients were evaluated.Kaplan-Meier survival analysis and Cox regression model were performed for survival analysis of prognosis to determine the effect of GDF 15 on the prognosis of ovarian cancer.3.Functional study on the promotion of GDF 15 in ovarian cancer metastasis:human recombinant GDF 15 protein(rhGDF15)and GDF 15 neutralizing antibody(GDF 15Ab)were used to treat ovarian cancer cell lines TOV112D and OVCAR3,respectively,and Transwell assay was used to evaluate the the changes in their migration and invasion ability.In addition,the migration and invasion ability of GDF15 overexpressing stable cell lines and underexpressing transient cell lines were evaluated by the same method,and the expression levels of downstream epithelial and stromal characteristic proteins were detected.4.The signaling pathway downstream of GDF15:GDF15 plasmid was transfected into ovarian cancer TOV112D cell line,and GDF15 transient cell line with high expression of GDF15 was constructed to detect the association between GDF 15 and Akt phosphorylation.The PI3K/Akt pathway was blocked by Akt inhibitor to observe whether the ability of GDF 15 to promote ovarian cancer invasion and migration was affected.5.Explore the mechanism of GDF 15 promoting ovarian cancer metastasis:after exogenous addition of rhGDF15 and overexpression of GDF 15,the ATP level and oxygen consumption rate of cell lines were detected at different time points,and on this basis,the effect of GDF 15 on lipid oxidation and decomposition level of ovarian cancer cells was detected.The effects of GDF 15 on lipid content and lipid-synthesisrelated protein expression in ovarian cancer cells were detected.To explore the relevant pathways downstream of GDF 15 and the influencing mechanism,the PI3K/Akt pathway was blocked to observe the effect of GDF 15 on the level of lipid anabolism.The key factors of lipid of lipid biosynthesis were blocked to investigate the association between lipid biosynthesis and the promotion of GDF 15 in metastasis of ovarian cancer.Results:1.Based on the results of collected ovarian cancer samples,it was suggested that the expression of serum GDF 15 was increased in ovarian cancer tissues compared with normal controls.IHC results of ovarian cancer clinical samples showed that GDF 15 could be expressed in ovarian cancer epithelial cells and tumor stromal cells.Compared with normal controls,GDF 15 protein expression was significantly increased in ovarian cancer patients.At both protein level and mRNA level,we found that the expression of GDF 15 was significantly higher in the metastatic ovarian cancer than in the primary ovarian cancer,suggesting that GDF 15 may promote the metastasis of ovarian cancer.2.Survival analysis based on the clinical sample database of this study showed that compared with patients with low GDF15 secretion level,ovarian cancer patients with high GDF15 expression in serum had significant differences,and those with high GDF15 expression had poor prognosis.3.In in vitro studies,the proliferation ability of ovarian cancer cells was significantly enhanced after treatment with rhGDF15 or overexpression of GDF15,as well as the migration and invasion ability,while drug sensitivity was significantly decreased.Treatment with GDF15Ab or knockdown of GDF15 led to decrease of the proliferation ability,migration and invasion ability of ovarian cancer cells and increase of drug sensitivity.4.After the overexpression of GDF15,epithelial characteristic protein decreased and stromal characteristic protein increased,indicating that GDF15 promoted epithelial mesenchymal transformation of ovarian cancer cells.5.GDF15 promoted the phosphorylation of Akt and activated the PI3K/Akt signaling pathway,while blocking of PI3K/Akt pathway by Akt inhibitor could effectively inhibit the effect of GDF15 promoting the migration and invasion of ovarian cancer.6.After 24 hours of stimulation of TOV112D ovarian cancer cells by rhGDF15,the overall ATP level of the cells was significantly up-regulated at different time points,and the oxygen consumption rate of ovarian cancer cells was also significantly increased in the treatment of rhGDF15.7.After sequencing and verification,we found that mRNA levels of lipid metabolism-related genes such as FABP4,CPT1 and CD36 were significantly different in ovarian cancer cell line TOV112DD-oeGDF15 with overexpression of GDF15.8.The lipid content of TOV112DoeGDF15 was significantly higher than that of TOV112D-vector.Based on this,the level of total triglyceride,free fatty acids and phospholipids were significantly increased,and the expression of some key enzymes in lipid synthesis was also significantly increased.9.Treatment with Akt inhibitor can significantly reduce the lipid content in the cytoplasm of TOV112D-oeGDF15,and treatment with Akt agonist can significantly increase the lipid content in the cytoplasm of TOV112D-vector.10.Inhibitor of key enzymes in lipid synthesis can significantly inhibit the migration and invasion effect of GDF15 on ovarian cancer.Conclusion:In conclusion,the expression of GDF15 is increased in ovarian cancer,and the high expression of GDF15 in the ovarian cancer is the risk factor of the poor prognosis of ovarian cancer.GDF15 is associated with malignant phenotypes of ovarian cancer.GDF15 induces the activation of PI3K/Akt pathway,up-regulates energy metabolism,promotes lipid anabolism,and thus enhances the migration and invasion ability of ovarian cancer.Objective To analyze the prognosis and appropriate treatment modalities of the patients with recurrence of early cervical squamous cancer primarily treated with radical hysterectomyMethods This retrospective study included patients with International Federation of Gynecology and Obstetrics(FIGO)stage Ⅰ B and Ⅱ A recurrent cervical squamous cancer who underwent radical hysterectomy primarily from January 2007 to July 2015.Kaplan-Meier method and Cox regression analysis were performed to analyze possible prognostic factors of overall survival and progression-free survival,which contained age,postoperative therapy,the site of recurrence,therapy-free interval(TFI)and treatment modality.We selected the patients who were treated with palliative chemotherapy after recurrence as a subgroup.The responses of palliative chemotherapy were evaluated and contrasted in their effect on different factors,including TFI,the site of recurrence and chemotherapy regime.Results Of the 2071 patients,116 relapsed and were included in our study with the average age of(45.6±7.2)years old.3-year progression-free survival rate and 3-year overall survival rate after recurrence were 42.2%and 30.2%,respectively.Univariate analysis implied that postoperative radiotherapy,recurrence site,TFI and treatment modality were associated with progression-free survival(P<0.05),while postoperative radiotherapy,TFI and treatment modality with overall survival(P<0.05).Multivariate analysis showed that TFI and treatment modality were independent prognostic factors for progression-free survival(P<0.05),while postoperative radiotherapy at initial treatment,therapy-free interval and treatment modality were independent prognostic variables for overall survival(P<0.05).In the analysis of treatment modality,3-year progression-free survival rate and 3-year overall survival rate of 47(40.5%)patients who were treated with definitive local therapy were significantly higher than that of 68(59.5%)patients who were treated with palliative chemotherapy(P<0.01).In the subgroup analysis of palliative chemotherapy,15 patients achieved complete response(CR,21.7%)and 16 displayed partial response(PR,23.2%).The overall response rate(ORR)was 44.9%.TFI(P<0.01)and chemotherapy regime(P<0.05)were significant factors associated with ORR.The ORR of TFI>12 months was significantly higher than that of TFI<12 months.Besides,the ORR of paclitaxel plus platinum chemotherapy was prominently higher than that of other regimes,while there is no significant difference in the ORR of paclitaxel plus cisplatin and other platinum(P=0.408).Conclusions For recurrent cervical squamous carcinoma treated with radical hysterectomy,use of definitive local therapy for suitable patients is advised to achieve better prognosis.In terms of palliative chemotherapy,longer TFI may mean better ORR and the combination of paclitaxel plus platinum is preferred. |
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