The Role And Mechanism Of HER2 In The Radiotherapy Of Gastric Cancer

The present study consisted of three parts.The first part clarified the impact of HER2 in the adjuvant radiotherapy of gastric cancer.The second part investigated the effect and mechanism of pyrotinib on the radiosensitivity of HER2-overexpression gastric cancer cells.The last part explored the anti-tumor effect of the combination of pyrotinib and chemotherapy drugs.Part 1: The effect of HER2 in the adjuvant radiotherapy of gastric cancer.【Objective】To investigate the impact of HER2 in the radiotherapy of gastric cancer.【Method】Patients received gastric cancer surgery in our hospital from January 2013 to June 2015 were enrolled in the present study.The frequency of HER2-overexpression and the association between HER2 and clinicopathological factors were analysed.Patients received gastric cancer surgery and standard adjuvant radiotherapy from January 2013 to December 2017 in our hospital were enrolled in the second part of the study.The effect of HER2 on the survival of these patients were analysed after regular following-up.【Results】HER2 test were received by 99.9% patients after surgery.HER2 positive was presented in 8.313% patients.HER2 expression was significantly associated with lymph node metastasis(P=0.042),but it was not associated with gender,age,tumor size,differentiation,tumor depth or clinical stage.There was no significant difference of the OS between HER2-positive(42.83 months)and HER2-negetive(37.53 months)patients received adjuvant radiotherapy(P=0.7029).【Conclusion】The OS of the gastric cancer patients received adjuvant radiotherapy was not affected by HER2 status.Part 2: The effect and mechanism of pyrotinib on the radiosensitivity of HER2-overexpression gastric cancer cells.【 Objective 】 To explore the impact of pyrotinib on the radiosensitivity of HER2-overexpression gastric cancer cells.【 Method 】 The experiments used NCI-N87 and SKBR3 cells exhibiting HER2 overexpression,and used MKN28 and MCF-7 cell lines with low expression of HER2.The expression of HER2 in these cell lines was determined by western blot and immunofluorescence.NCI-N87 and SKBR3 cells were treated by irradiation or the combination of pyritinib and irradiation.Then the proliferation was tested by CCK-8 assay.Colony formation assay was used to determine the radiosensitivity of cells in vitro.The activation of PI3K/Akt and MEK/MAPK signal pathway and the expression of HER2 in the nuclear were determined by western blot.Apoptosis and distribution of cell cycle were assessed by flow cytometry assay.The DNA damage was detected by western blot and immunofluorescence.The xenograft model in mice was used to explore the effect of pyrotinib on the radiosensitivity in vivo.【Results】HER2 overexpressing cells were selectively sensitive to pyrotinib inhibition.The combination of pyrotinib and irradiation significant inhibited proliferation compared with irradiation alone group(P<0.01).Pyrotinib enhanced the radiosensitivity of NCI-N87 and SKBR3 cells in colony assay(SER=1.375 and 1.326).The combination of pyrotinib and irradiation exhibited a greater antitumor impact than irradiation alone treatment in xenograft tumors.Pyrotinib inhibited irradiation-induced HER2 nuclear transport and DNA damage repair.Meanwhile,pyrotinib increased apoptosis and G2/M arrest induced by irradiation.【Conclusion】Pyrotinib could enhance the radiosensitivity of HER2-overexpressing gastric cancer cells.The mechanism involved in this process was the inhibition of HER2 nuclear transport and DNA damage repair and the increased of apoptosis and G2/M arrest induced by irradiation.This study may provide a new treatment strategy for HER2-overexpressing gastric cancer patients.Part 3: The efficacy of the combination of pyrotinib and cytotoxic drugs.【Objective】To clarify the anti-tumor effect of cytotoxic drugs combined with pyritinib.【 Method 】 The experiments used NCI-N87 and SKBR3 cells exhibiting HER2 overexpression.Chemotherapy drugs,including docetaxel,fluorouracil,cisplatin,and epirubicin,were used in this part.NCI-N87 and SKBR3 cells were exposed to different concentrations of each agent alone or in combination with pyrotinib for 48 h.CCK-8 assay was used to tested the proliferation of the cells.Student’s t test was applied to evaluate the significance of differences between the two groups.Mice bearing xenograft tumors were divided into four groups: control group,cytotoxic drugs alone group,cytotoxic drugs combined with pyrotinib group,cytotoxic drugs combined with pyrotinib and irradiation group.Tumor volume was recorded in each group.【Results】In NCI-N87 cells,the combination of pyrotinib and docetaxel/ cisplatin showed a stronger inhibitory effect than docetaxel/ cisplatin(P<0.01).In SKBR3 cells,the pyrotinib combined with docetaxel/ fluorouracil exerted a stronger inhibitory effect than docetaxel/ fluorouracil(P<0.01).There was no significant difference between pyrotinib combined with epirubicin and epirubicin alone.Furthermore,pyrotinib combined with docetaxel exhibited better anti-tumor effect than docetaxel in animal models(P<0.05).The addition of irradiation to pyrotinib combined with docetaxel further induced tumor regression in gastric cancer models,while not in breast cancer models.【Conclusion】Pyrotinib significantly enhanced the cytotoxicity of docetaxel in HER2 overexpressing tumors,which may provide a new treatment strategy for HER2-overexpressing gastric cancer patients.