Role And Mechanism Of Leptin And Leptin Receptor In Breast Cancer

Objective:Breast cancer is one of the most common malignant tumors in women.The incidence of breast cancer ranks first among female malignant tumors,and it has become a major public health problem in the current society.Epidemiology has found that obesity as a global health problem is one of the important risk factors for breast cancer.Some studies have shown that obesity is an independent prognostic factor for patients with breast cancer who have reduced pathological complete response to neoadjuvant chemotherapy.Leptin as an important adipocyte factor,is closely related to obesity.Overexpression of leptin and its receptor have been detected in breast cancer.The binding of leptin to its receptor can promote the proliferation of cancer cells,inhibit apoptosis,induce angiogenesis and promote the metastasis of breast cancer.However,the relationship between leptin and its receptor and the therapeutic efficacy of breast cancer remains unknown.NF-kappa B is an important nuclear transcription factor.Its signaling pathway plays an important role in the occurrence,development and metastasis of human solid tumors.IKK beta is an important kinase in the classical NF-kappa B signaling pathway,which is involved in regulating downstream gene transcription after activation.Experimental studies have shown that conditional knockout or knockdown of IKK beta in tumor cells can significantly inhibit the growth of solid tumors such as lung cancer,melanoma and glioma in mice,suggesting that IKK beta may be one of the potential targets for clinical treatment of tumors.Moreover,it also has been reported that estrogen receptor alpha(ER alpha)and NF-kappa B can synergistically stimulate tumor proliferation and drug resistance,while through its receptor leptin can regulate and activate ER alpha.However,the relationship of leptin with NF-kappa B signaling pathway in breast cancer has not been further studied.To address the role of leptin and its receptor on the efficacy,prognosis and the pathological complete response rate for neoadjuvant chemotherapy in patients with breast cancer,we examined the expression of leptin and leptin receptor in the PCR,non-PCR and PD groups after neoadjuvant chemotherapy in 325 patients with breast cancer.Meanwhile,in this study,we determined the effects of IKK beta on proliferation,apoptosis and metastasis of breast cancer cells with different expression levels of leptin and its receptor,as well as the underlying mechanism NF-kappa B signaling pathway regulates the expression and function of leptin and its receptor in breast cancer cells,Therefore,our study will give a possible predictive and prognostic reference index for breast cancer chemotherapy,and help to find a new potential therapeutic target.Methods: Detailed information of 325 breast cancer patients were collected in our hospital from January 2014 to December 2017 and preoperative biopsy was well completed.All patients were followed up.The expressions of leptin and leptin receptor in patients' tissue were detected by immunohistochemical method.The expressions of leptin and leptin receptor in the PCR,non-PCR and PD groups were analyzed,and the relationships between leptin and molecular type and prognosis of patients were analyzed.Then IKK? in breast cancer cells was knocked down via virus transfection technology.Then we performed MTT,flow cytometry,cell scratch test and clone formation test and so on to address the effects of IKK beta on proliferation,apoptosis and metastasis of breast cancer cells with different expression levels of leptin and its receptor.Moreover,we detected the downstream genes expressions in IKK beta signal pathway by RT-PCR to find out the relationships between NF-kappa B signal pathway with leptin and its receptor.Results:1.The positive and strong positive expression rates of leptin and leptin receptor in the PCR group were higher than those in the non-PCR group respectively,but there was no statistical significance,suggesting that there were no significant differences in the expressions of leptin and leptin receptor between the PCR group and the non-PCR group.The positive and strong expression rates of leptin and leptin receptor in the PCR group were significantly higher than those in the PD group.There was no statistical significance on the expressions of leptin between the two groups,which implying that patients with high expressions of leptin receptor will have better therapeutic efficacy.2.The positive and strong expression rates of leptin and leptin receptor in HER2 positive PCR group were obviously higher than those in PD group,suggesting that the patients with high expressions of leptin and leptin receptor in HER2 positive patients had better therapeutic efficacy.3.The positive rates of leptin and leptin receptor expressions in HR-positive,HER2-positive and triple-negative patients were significantly different.The expressions of leptin and leptin receptor in triple-negative patients were higher than that in HER2-positive and HR-positive patients.4.The disease progression rates of high expressions group(positive and strong positive expression)and low expressions group(weak positive and negative expression)were 13.25% and 11.32%,respectively,while that of high expressions group and low expressions group were 13.1% and 10.42%,respectively,with no statistical significant difference.It suggests that the prognosis of patients with high expressions of leptin and leptin receptor may be worse.5.The expression levels of leptin and leptin receptor in MCF-7 cells were 1.005 and 1.015,respectively.In MDA-MB-231 cells,the expression levels of leptin and leptin receptor were 1.384 and 1.449,respectively.There were significant differences between the two cell lines.Lentivirus can successfully infect breast cancer cells in vitro and knock down the expression of IKK?.6.Knockdown of IKK? expression reduced the viability of MDA-MB-231 cells,but had little effect on MCF-7.It can effect on cell cycle of both cell lines.After knocking down IKK?,the apoptotic rate of MDA-MB-231 cells increased.There was no significant difference in the apoptotic rate of MCF-7 cells.The number of clones of MCF-7 and MDA-MB-231 cells was significantly decreased.The effect on MDA-MB-231 cells was more significant.After knocking down the expression of IKK beta on MDA-MB-231,scratch mobility decreased significantly,it was statistical significant.7.After knocking down IKK?,the expressions of leptin and leptin receptor,IKBKG,P65 and HIF in MCF-7 cells were significantly decreased compared to control group with statistical significance.However,the expressions of leptin and leptin receptor,IKBKG and P65 was significantly increased in MDA-MB-231 cells.The expression level of IKBKG increased significantly.There was no significant difference in the expression level of HIF.Conclusion:1.Neoadjuvant chemotherapy may be more effective in patients with high leptin and leptin receptor expressions,especially in HER2 positive patients.2.The expressions of leptin and leptin receptor are different in different molecular types of breast cancer,which may have some predictive value for the prognosis.3.The expression levels of leptin and leptin receptor in MDA-MB-231 cells were higher than those in MCF-7 cells,which was consistent with clinical studies.4.After blocking the NF-kappa B pathway,the proliferation,metastasis and apoptosis of MDA-MB-231 cells were inhibited,while the proliferation of MCF-7 cells was inhibited.There was no significant change in apoptosis.The differences between the two cell lines may be related to the different expression levels of leptin and its receptor.5.After knocking down IKK?,the expressions of leptin and leptin receptor in MCF-7 cells decreased,while there was an increasing trend in MDA-MB-231 cells.It may related to the inhibition of leptin expressions in MDA-MB-231 cells by NF-kappa B signaling pathway,the specific reason needs further basic research to confirm.6.After blocking the NF-kappa B pathway,the expressions of genes related to the NF-?B pathway in MDA-MB-231 cells increased,while the expressions of genes related to the NF-?B pathway in MCF-7 cells decreased,which may be related to the different expression levels of leptin and its receptor.7.After knocking down IKK?,the expressions of HIF gene in MCF-7 cells decreased,while that in MDA-MB-231 cell lines were not affected.Whether this difference is related to leptin and its receptor needs further verification.