Abstract
Background: It is well-described that the transcriptome of peripheral blood mononuclear cells (PBMCs) can be altered in the context of many malignancies to allow them avoid the effective immune response, which leads to cancer invasiveness. Here, we used an MS-based strategy to discover biomarkers in the PBMCs of breast cancer (BC) patients and validated them at different stages of BC.Methods: PBMCs were isolated from the breast cancer patients and were cultured alone or co-cultured with breast cancer cell lines. The role of PBMC in the invasion property of breast cancer cells was explored. NF-kB activity was also measured in the co-cultured breast cancer cells. Identification of protein profiles in the secretome and proteome of the co-cultured PBMCs was performed using SWATH mass spectrometry. Pathway enrichment and gene ontology analyses were carried out to look for the molecular pathways correlated with the protein expression profile of PBMCs in the breast cancer patients. Quantitative real-time polymerase chain reaction (qPCR) was performed to validate the candidate genes in the PBMC fraction of the breast cancer patients at the primary and metastatic stages. In silico survival analysis was performed to assess the potential clinical biomarkers in these PBMC subtypes.Results: PBMCs could significantly increase the invasion property of the BC cells concomitant with a decrease in E-cadherin and an increase in both Vimentin and N-cadherin expression. The NF-kB activity in the BC cells significantly increased following co-culturing implying the role of PBMCs in EMT induction. Enrichment analysis showed that the differentially expressed proteins in PBMCs are mainly associated with IL-17, PI3K-Akt, and HIF-1 signaling pathway, in which a set of seven proteins including TMSB4X, HSPA4, S100A9, SRSF6, THBS1, CUL4A, and CANX were frequently expressed. Finally, in silico analysis confirmed that a gene set consisting of S100A9, SRSF6, THBS1, CUL4A, and CANX were found to provide an insight for the identification of metastasis in breast cancer patients.Conclusion: In conclusion, our study revealed that the protein expression profile in PBMCs is a reflection of the proteins expressed in the BC tissue itself; however, the abundance level is different due to the stage of cancer.
Highlights
Tumors that originate from the epithelial cells grow in a dynamic and intricate stroma composed of endothelial cells, blood vessels, immune cells, and a variety of associated tissue-type cells along with matrix proteins and soluble factors [1,2,3]
Since no changes in the expression levels of epithelial to mesenchymal transition (EMT) markers were detected in the two cell lines after co-culturing with the peripheral blood mononuclear cells (PBMCs) taken from healthy individuals, we decided to further examine the breast cancer cell lines co-cultured with PBMCs obtained from patients in the primary stage of breast cancer
MDA-MB-231 cells revealed a significant increase in vimentin and N-cadherin expression upon co-culturing with PBMCs obtained from triple negative patients in the mere presence of conditioned media (50%) (Figures 1E,F)
Summary
Tumors that originate from the epithelial cells grow in a dynamic and intricate stroma composed of endothelial cells, blood vessels, immune cells, and a variety of associated tissue-type cells along with matrix proteins and soluble factors [1,2,3]. Two of the most abundant immune cell populations found in the stroma of breast cancer are tumorassociated macrophages (TAMs) representing up to 50% of the tumor mass and T cells [10,11,12,13,14] Both of them might have been associated with a poor prognosis and detrimental clinical outcome in breast cancer patients, likely by promoting the epithelial to mesenchymal transition (EMT) process, metastasis, and angiogenesis. During EMT, tumor cells exhibit dynamic changes in epithelial and mesenchymal compositions, which endow tumor cells with enhanced invasive and resistance phenotypes [11, 15,16,17] It is well-described that the transcriptome of peripheral blood mononuclear cells (PBMCs) can be altered in the context of many malignancies to allow them avoid the effective immune response, which leads to cancer invasiveness. We used an MS-based strategy to discover biomarkers in the PBMCs of breast cancer (BC) patients and validated them at different stages of BC
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