| Background:The incidence of breast cancer has remained high among females with malignant tumors in recent years,and breast cancer is becoming a threat to women’s health worldwide.Previous studies have shown that tumor cells interact with the tumor microenvironment(TME)as a whole to promote tumor occurrence and development.The prognosis of breast cancer is affected not only by the biological characteristics of tumor cells but also by the status of the TME.The TME is mainly composed of tumor cells,multiple mesenchymal cells,and the extracellular matrix.All these components participate in inducing tumor proliferation,stimulating angiogenesis,inhibiting cell apoptosis,and suppressing the immune system,ultimately hindering the antitumor response and promoting the metastasis of breast cancer.Previous treatments for breast cancer,including traditional radiotherapy,chemotherapy,and targeted therapy,mostly target and kill tumor cells through corresponding drugs.However,these treatments have limitations because of drug resistance.To survive,tumor cells activate a variety of drug resistance mechanisms to evade drugs.Once drug resistance occurs,the efficacy of the administered drugs greatly reduces.Recent progress in tumor-infiltrating lymphocytes and immune checkpoint inhibitors has promoted immunotherapy against advanced triple-negative breast cancer(TNBC),During the past few years,researchers have studied the regulatory effects of the TME on breast cancer and its potential therapeutic value,aiming to regulate the“soil environment" of tumor cells through TME transformation.Moreover,treatments targeting the TME will provide new treatment choices for patients with breast cancerObjective:To study the importance of the TME in breast cancer,we analyzed related factors and prognostic impacts based on retrospective research.The purpose was to explore dynamic changes in relevant indicators in the TME of breast cancer through immunohistochemical methods and cell co-culture models.By revealing the underlying mechanisms of prognostic effects,this study may provide new ideas for breast cancer-related treatments.Part One:Prognostic analysis of systemic immune inflammation indicators in patients with breast cancerMethods:The preoperative systemic immune-inflammationindex(SII)was obtained from 380 patients with breast cancer who were continuously treated in our hospital and met the selection criteria.We compared the clinicopathological characteristics of these patients based on the SⅡ levels.Survival analysis was performed based on the rate of breast cancer-related death.Based on the follow-up results,we performed a subgroup analysis among 36 patients with TNBC.Results:Based on the receiver operating characteristic curve,we divided all patients into high and low SII subgroups(222 and 158 patients,respectively).The SⅡ level was related to the maximum diameter and the tumor-node-metastasis(TNM)stage of the tumor.We found that patients with a tumor diameter measuring>5 cm(P=0.013)and a tumor in stage Ⅲ(P=0.004)usually had high SII levels.There was no significant difference in the SⅡ level in terms of age group,surgical method,histological grade,and molecular subtype.Univariate and multivariate analyses showed that the TNM stage and preoperative SⅡ level may be independent risk factors of breast cancer-specific survival,and they also have prognostic significance in the TNBC subgroup.Part Two:Changes in related indicators in the microenvironment and analysis of triple-negative breast cancer prognosisMethods:We enrolled patients from our hospital who underwent surgery because of primary TNBC and were reoperated because of breast cancer recurrence and/or metastasis during follow-up.Of the 62 patients who met the criteria,the molecular classification in 43 patients remained as TNBC.We used post-recurrence or post-metastasis survival as the outcome event.All patients were analyzed based on clinical pathological related factors and peripheral blood SII levels before reoperation.Immunohistochemical staining was later performed on primary and relapsed/metastatic lesions.The Ki-67 index,p53 status,and expression of CD8,programed cell death-ligand 1(PD-L1),T cell immunoglobulin and mucin domain-3(TIM-3),and fibroblast activation protein(FAP)were compared and analyzed.Results:The average interval between the two operations was 29.05 ± 25.79 months.In 69.4%patients,the molecular classification remained as TNBC,and the pathological features were characterized by a high Ki-67 index and a positive p53 status.According to the molecular classification of relapsed/metastatic cases,distant metastasis was the reason for>50%reoperations in the non-TNBC group,while local recurrence was the reason for reoperation s in the TNBC group(P=0.029).There were no statistical differences in age,menstrual status,surgical method,pathological stage,histological characteristic,peripheral blood SⅡ level before reoperation,adjuvant therapy,and rescue therapy between the two groups.Univariate and multivariate analyses showed that the maximum diameter of the initial tumor and SⅡ levels before reoperation may be independent risk factors of post-metastasis survival in this group of patients.A comparative analysis between immunohistochemical indicators of primary and relapsed/metastatic tumors was performed in 43 patients.The study showed that relapsed/metastatic tumors were mainly characterized by increased positive staining for p53,FAP,PD-L1,and TIM-3 and decreased positive staining for CD8+ tumor-infiltrating lymphocytes.Part Three:Changes in the morphology and immune status of breast cancer cells induced by co-culture with mesenchymal stem cellsMethods:To simulate the effect of mesenchymal stem cells(MSCs)on tumor cells in the TME of breast cancer,the breast cancer cell lines MCF-7 and MDA-MB-231 were co-cultured with human adipose-derived MSCs(hAD-MSCs),which were functionally stable after isolation,culture,and identification.We observed the morphological changes in breast cancer cells on days 0,1,3,7,and 10 during co-culture and collected the RNA and protein from the two types of cells.We performed quantitative real-time PCR(qRT-PCR)and western blotting to evaluate gene and protein expression,respectively,and compared changes in the immune status of PD-L1 during co-culture.Results:When co-cultured with hAD-MSCs,MCF-7 breast cancer cells were round,whereas MDA-MB-231 cells were fusiform.The co-culture models induced morphological changes;the epithelioid-like state of breast cancer cells was changed to a mesenchymal-like state.Further,the expression of PD-L1 was gradually upregulated during the co-culture process,suggesting that co-culture with hAD-MSCs may cause changes in the immune status of breast cancer cells.Conclusions:The prognosis of breast cancer is not only related to the biological characteristics of tumor cells but also closely linked to their TME.The SⅡ index is an indicator based on peripheral blood lymphocyte,neutrophil,and platelet counts,which may more comprehensively reflect the inflammation and immune status of patients with breast cancer.Hence,the SⅡ index possesses important predictive value for the disease-specific survival in patients with breast cancer.Hormone receptor and human epidermal growth factor receptor 2 status may change during the process of relapse or metastasis.Further,because of malignant transformation and enhanced invasiveness,they may have depleted CD8+T cells,an increased expression of negative costimulatory molecules,and an increased rate of fibroblast activation protein.Rebiopsy in patients with relapsed/metastatic breast cancer under clinically permitted conditions is helpful for subsequent treatment strategies.In the co-culture model,hAD-MSCs induced morphological changes in breast cancer cells,leading to a gradual upregulation of PD-L1,which may be related to the formation of a suppressive immune TME.In conclusion,the co-culture model based on hAD-MSCs can provide promising new insights for studying the interaction between breast cancer and the TME and the associated underlying mechanisms involved. |
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