Study On The Relationship Between The Occurrence Of Triple-negative Breast Cancer And The Expression Control Of Antigen Presenting Machinery (APM) In Uygur And Han Women

Objective:Breast cancer is one of the major malignant tumors in women,and the incidence of breast cancer in the world gradually increased,of which triple negative breast cancer(triple negative breast cancer,TNBC)is a subtype of breast cancer,its main pathological features is negative expressing of progestrone receptor(PR),estrogen receptor(ER)and human epidermal growth factor receptor 2(HER-2).Compared with other subtypes of breast cancer,TNBC is more common in premenopausal women,with mild onset,with higher brain metastases,visceral metastasis rate and local recurrence rate,shorter total survival time and tumor-free survival time.Tumor immune escape plays an important role in the development and progression of breast cancer.T cell-mediated immune surveillance is based on normal function of antigen-processing elements(APM)and human leukocyte antigen class I(HLA-I)function.HLA-I and APM function is normal or not involved in tumor survival and development process.There are a large difference between Xinjiang Uygur and Han women breast cancer prevalence and clinical characteristics,therefore,to carry out TNBC pathogenesis of basic and clinical research,not only help to reveal the tumor subtype research in the field of common problems to provide a scientific basis,but also help to improve the level of prevention and treatment of breast cancer in Xinjiang.In this study,the differences of HLA-I and APM gene expression levels in TNBC tissues of Uygur-Han were investigated by immunohistochemistry and RT-PCR using the resources.Sequenom Mass ARRAY(?) DNA methylation mass spectrometry was used to analyze HLA-I and APM gene methylation levels in TNBC tissues of Uygur women,and to elucidate the apparent genetic modification of TNBC production in Uygur women.Methods:(1)Uygur and Han women breast lesions in patients with paraffin-embedded tissue and fresh tissue were collected.The protein and m RNA expression levels of HLA-A,HLA-B,HLA-C,TAP1,TAP2,calnexin,calreticμlin,Erp57,ERAP1 and Tapasin were identified by immunohistochemistry and semi-quantitative RT-PCR.(2)For TAP1 and ERp57 gene,methylation level of Cp G island in the promoter region of the candidate gene was done by Mass ARRAY methylation detection technique.The differences in methylation levels in the whole and single Cp G sites of TNBC patients,TNBC and non-TNBC patients,breast and breast fibroma were identified.Results:(1)Immunohistochemical analysis showed that the positive expression rates of HLA-A,HLA-B,TAP2,Erp57,ERAP1,calnexin and calreticμlin in malignant breast tissue were significantly lower than those in benign breast tissue(P<0.05),while there was no significant difference in benign and malignant breast tissue for HLA-C,TAP1 and tapasin(P>0.05).There were no significant differences in the expression levels of HLA-A,HLA-B,HLA-C,TAP1,TAP2,calnexin,calreticμlin,tapasin,Erp57 and ERAP1 in Uygur and Han breast cancer patients(P>0.05).There were no significant differences in the expression levels of HLA-A,HLA-B,HLA-C,TAP1,TAP2,calnexin,calreticμlin,Erp57 and ERAP1 in the patients with triple-negative and non-tri-negative breast cancer(P>0.05).86 cases of triple-negative breast cancer patients included tapasin-35 cases(40.70%),+ 26 cases(30.20%),++ 25 cases(29.10%);34 cases of non-triple negative breast cancer patients with tapasin-25 cases(73.50%),+ 7 cases(20.60%),++ 2 cases(5.90%).The positive expression rate of tapasin in patients with triple negative breast cancer was significantly higher than that in non-triple negative breast cancer patients(P<0.05).(2)RT-PCR resμlts showed that there was no significant difference in HLA-A,HLA-B,HLA-C,TAP1,calnexin,calreticμlin and ERAP1 m RNA expression between benign and malignant breast tissues(P>0.05).The expression level of ERp57 and TAP2 m RNA in malignant breast tissue was significantly lower than benign breast tissue(P<0.05).The expression of HLA-A,TAP2,calreticμlin,ERp57 and ERAP1 m RNA in patients with triple-negative and non-tri-negative breast cancer were significantly different(P<0.05).HLA-B,HLA-C,TAP1 and calnexin m RNA expression levels were not statistically significant(P>0.05).(3)The total methylation level showed that there was no significant difference in the total methylation level of TAP1 and ERp57 genes in TNBC tissues of Uygur and Han (P>0.05).The total methylation level of ERp57 gene in TNBC tissues was significantly lower than that in non-TNBC tissues(P<0.01).There was no significant difference in total methylation level of TAP1 genes in TNBC and non-TNBC tissues(P>0.05).The total methylation level of ERp57 gene in breast cancer tissue was significantly lower than that in benign breast tissue(P<0.05),and the total methylation level of TAP1 gene was significantly higher than that of benign breast tissue(P<0.05).(4)There was no significant difference in the methylation level of TAP1 and ERp57 single Cp G in the TNBC tissues of Uygur and Han (P>0.05).ER57-8-Cp G-1,ER57-8-Cp G-2.3.4,ER57-8-Cp G-5,ER57-8-Cp G-6.7,ER57-8-Cp G-8,ER57(P<0.01).ER57-8-Cp G-1,ER57-8-Cp G-2.3.4,ER57-8-Cp G-5,ER57-8-Cp G-8,ER57-8-Cp G-9 in ER57 gene promoter region of breast cancer(P<0.05).The level of methylation of TAP1-14-Cp G-29 in TAP1 gene promoter region was significantly higher than that in benign mammary gland(P<0.05).Conclusion:(1)The expression of APM members in breast cancer cells may be prevented by the correct assembly,loading and presentation of endogenous antigenic peptides after transcription of HLA class I molecules.There was no significant difference in the expression of HLA-I and APM in Uygur and Han breast cancer patients.There was significant difference in the expression of HLA-I and APM in TNBC patients and non-TNBC patients,which was mainly manifested in the low expression of HLA-C,TAP1 and calnexin and the high expression of tapasin.Therefore,the transcriptional expression of HLA-I and APM genes or the lack of protein expression are closely related to the process of breast cancer,which has the potential to screen tumor molecμlar markers.(2)There were no significant differences in the methylation levels of TAP1 and ERp57 genes in TNBC patients of Uygur and Han .(ER57-8-Cp G-1,ER57-8-Cp G-2.3.4,ER57-8-Cp G-5,ER57-8-Cp G-6.7,and ER57-8-Cp G)of the ERp57 gene promoter was closely related to the occurrence of TNBC.(ER57-8-Cp G-1,ER57-8-Cp G-2.3.4,ER57-8-Cp G-5,ER57-8-Cp G-8,ER57-8-Cp G)in the ER57 gene promoter region,and(TAP1-14-Cp G-29)in the TAP1 gene promoter region and one Cp G site are closely related to the occurrence of breast cancer and may be a key site for regμlating the methylation of the corresponding gene promoter region.In conclusion,methylation of the promoter regions of TAP1 and ERp57 genes in APM members plays an important role in the development and progression of TNBC.