Research On The Correlation Between Peripheral Blood Of Breast Cancer And Tumor Immune Microenvironment Based On Gene Expression Profiling

Breast cancer is a major cause of morbidity and the leading global cause of cancer motality among women. Clinical management relies on known prognostic factors, such as hormone receptor(HR) and HER2 status, for predicting responsiveness to therapies. However, even when morphological characteristics and ER phenotypes are similar, patients have varying prognosis or chemotherapy response. Gradually increasing resistance against conventional therapies requires improving strategies to diagnose and treat breast cancer. The development of malignant tumor is controlled by a complex biological system that depends on genetic abnormalities as well as the interplay between tumor cells and their microenvironment. Immune context of tumor microenvironment(TME) including abundant immune cells have been reported to be involved in breast cancer prognosis and are being pursued as potential therapeutic targets. Peripheral blood is the most common and most widely-accepted material used in disease diagnosis. Peripheral blood is conveniently and quickly obtained, and it is considered ideal for non-invasive molecular diagnosis. Compared to surgical tissue samples, which are usually obtained at a single point in time, peripheral blood can be sampled regularly so that dynamic changes in disease progression and/or prognosis can be easily monitored. Peripheral blood mononuclear cells(PBMCs) are composed of several types of immune cells, which are key components in the immune defense system. Immune cells in the peripheral blood are recruited to the microenvironment and become tumor infiltrating immune cells. However, no in-depth, systematic research has been reported regarding the significance of relationship and interaction between tumor immune microenvironment and the peripheral immune system in breast cancer diagnosis, personalized treatment and prognosis.Clinical bioinformatics is a new emerging science combining clinical informatics, bioinformatics, medical informatics, information technology, mathematics, and omics science together. It plays an important role in tumor research, focusing on the combination of clinical measurements and signs with human tissue-generated bioinformatics, understanding clinical symptoms and signs, disease development and therapeutic strategy, and mapping relationships that integrate discrete elements that collectively direct global function within a particular omic category. Based on m RNA expression profiling datasets in GEO, this research proposed comprehensive bioinformatics approaches to re-analysis of large data sets, to offer valuable clues for breast tumor diagnosis, treatment, and prognosis research. First, the changes of gene expression in peripheral blood mononuclear cells(PBMCs) of benign and malignant breast tumor were observed, to evaluate the clinical utility for PBMCs as a noninvasive material for detecting immune status of breast tumor patients. Then, investigated similarities and differences of genome-wide expression patterns between breast cancer stroma and peripheral blood mononuclear cells(PBMCs) of breast cancer patients, and finally established the molecular crosstalk between breast tumor tissue and PBMCs, to uncover the tumor-driven systemic perturbations.The main contributions are as follows:1. Gene expression profiles analysis identifies key genes of PBMCs in patients with benign and malignant breast tumor. Datasets of gene expression profiling were downloaded from the GEO database, including PBMCs profiling of 37 benign breast tumor samples, 57 breast cancer samples and 31 healthy controls. GEO2 R tool was used to analyze the data to identify the differentially-expressed genes(DEGs). Function of DEGs were annotated by DAVID. Protein interaction analysis and hub gene selection were then performed using STRING database. 563 and 237 DEGs in benign and malignant breast tumor respectively were identified. DEGs in breast cancer PBMCs involved in biological process of leukocyte activation, angiogenesis and leukocyte transendothelial migration. The hub genes interleukin-8, ras homolog family member B(RHOB), integrin,beta 1(ITGB1),may play a role in systemic immune of breast tumor.2. Relationship between stroma immune microenvironment and peripheral blood mononuclear cells in breast cancer patients. Microarray data of breast tumor stroma, which includes 53 cases of tumor stroma, and 31 cases of individual-matched normal adjacent stroma were available in GEO. BRB-Array Tools performed a series of preprocessing steps,to analyze the transcription profiles and to screen for DEGs. Function of DEGs were annotated by DAVID. A total of 103 overlap genes of the two datasets were identified. Functional annotation indicated that DEGs of both breast cancer PBMCs and stroma were involved in immune response. Further study on the profiles of cytokines and chemokines was performed. 81 cytokines and cytokine receptors were differentially expressed(76 up- and 5 down-regulated) in breast tumor PBMCs. 55 up-regulated and 48 down-regulated cytokines and cytokine–related genes in breast tumor stroma. The common disturbed pathways of cytokines and related genes were inflammatory reaction, immunosuppression and promoting tumorigenesis.3. Revealing potential molecular crosstalk between breast cancer and peripheral immune system. A bioinformatic analysis of three publically available microarray datasets for breast tumor stroma, PBMCs, breast tumor epithelium respectively were performed, integrating transcriptomics data, protein-protein interactions(PPIs), subcellular localization to identify genes and biological pathways that contribute to the dialogue between the tumor cells and the peripheral circulation, as well as tumor stroma and the peripheral circulation. A total of 101 interaction relationships about tumor cellsPBMCs crosstalk were identified and visualized. The genes such as FN1, ITGB1, FLT1, CXCR3, COPM, CXCL12 were hub nodes of the crosstalk network and may play an important role in response for tumor-derived chemoattractants. Similarly, a network consisted of 19 tumor stroma cell-derived differentially up-regulated ligands and 26 PBMC membrane receptors interacting with 73 intracellular proteins were constructed. The genes of the integrin family as well as CXCR4 were hub nodes of the crosstalk network and may play an important role in response for stroma-derived chemoattractants. The crosstalk molecules were mainly involved in inflammation, cell adhesion and migration. ECM receptor interaction, cytokine-cytokine receptor interaction, chemokine signaling pathway were included in the pathway lists.In summary, based on high throughput gene-expression datasets and comprehensive bioinformatics approaches, the present study systematically analyzed the PBMCs gene expression profiling, compared the expression pattern between PBMCs and stroma of breast cancer, provided further understanding of the interaction between breast tumor and the immune system. The genes, such as IL-8, RHOB, ITGB1, FN1, FLT1, COMP, CXCL12, ITGB3, ITGA2 B, CXCR4 may play an important part in formation of immune microenvironment originating from peripheral blood. Among them, ITGB1 and CXCR4, significantly up regulated in breast tumor PBMCs and simultaneously be as targets of both tumor cell and stroma, may need further explored. This study has the potential for development of therapeutic strategies that target the systemic signals travelling through the circulation and interdict tumor favorite cells recruitment.