Breast cancers (BCs) are solid tumors composed of heterogeneous tissues consisting of cancer cells and an ever-changing tumor microenvironment (TME). The TME includes, among other non-cancer cell types, immune cells influencing the immune context of cancer tissues. In particular, the cross talk of immune cells and their interactions with cancer cells dramatically influence BC dissemination, immunoediting, and the outcomes of cancer therapies. Tumor-infiltrating lymphocytes (TILs), tumor-associated macrophages (TAMs), and myeloid-derived suppressor cells (MDSCs) represent prominent immune cell populations of breast TMEs, and they have important roles in cancer immunoescape and dissemination. Therefore, in this article we review the features of TILs, TAMs, and MDSCs in BCs. Moreover, we highlight the mechanisms by which these immune cells remodel the immune TME and lead to breast cancer metastasis.

Breast cancer is the leading cause of cancer-related death in women worldwide. It is well known that breast cancer shows significant alterations in the tumor microenvironment (TME), which is composed of a variety of immune cells, including natural killer (NK) cells, that have a key role in tumor development or anti-tumor responses in breast cancer patients. Luminal B (BT474) and triple-negative breast cancer (HS578T) cell lines were cultured in 2D and 3D model systems. PMBCs from healthy donors were isolated and treated with extracellular vesicles (EVs) from monolayer and spheroids of BT474 and HS578T and analyzed using cytofluorimetric approaches. We observed that EVs can alter the activation and presence of CD335+/CD11b+ NK cells. EVs derived from BT474 and HS578T cells trigger the activation and, simultaneously, a reduction in the percentage of CD335+/CD11b+ NK cells. In addition, EVs derived from BT474 also significantly reduce CD39+ T-regulatory (T-reg) cells. Our preliminary data suggest that using EVs to treat tumors could potentially alter components of the immune system, which causes hyperactivation of specific cell types and can lead to aggressive growth. These data will guide the designing of new personalized diagnostic approaches based on in-depth study of the TME.

Goat milk is a complex biological fluid, which in addition to having a high nutritional value, it is an interesting source of extracellular vesicles (EVs). Despite the countless potential applications that they offer in many biological fields, is not easy to compare the different proposed systems, and this is a major limitation for the real translatability of these natural nanoplatforms for theragnostic purposes. Thus, it is useful to further investigate reproducible methods to separate goat milk EVs. The choice of methods but also the preprocessing of milk has an immense impact on the separation, quality, and yield of EVs. Here, we tested four protocols to separate EVs from unpasteurised goat milk: two based on differential ultracentrifugation (DUC) and two on size-exclusion chromatography (SEC). Moreover, we assessed two different approaches of pre-treatment (acidification and precipitation) to facilitate milk protein discharge. To the best of our knowledge, a similar comparison of all performed protocols on raw goat milk has never been published before. Therefore, enriched EV samples were successfully obtained from goat milk using both DUC and SEC. Taken together, our results may be helpful to obtain natural carriers for future theragnostic applications in personalised medicine.

Breast cancer is the most common cancer amongst women worldwide. Recently, owing to screening programs and new technologies, the survival rate has increased significantly. Breast cancer can potentially develop metastases, and, despite them, lung metastases generally occur within five years of breast cancer diagnosis. In this study, the objective was to analyze the effect of breast cancer-derived EVs on a lung epithelial cell line. BEAS-2B cells were treated with extracellular vesicles (EVs) derived from triple-negative breast cancer cells (TNBCs), e.g., MDA-MB-231 and HS578T, separated using differential ultracentrifugation. We observed an increased growth, migration, and invasiveness of normal epithelial lung cells over time in the presence of TNBC EVs compared to the control. Therefore, these data suggest that EVs released by tumor cells contain biological molecules capable of influencing the pro-tumorigenic activity of normal cells. Exploring the role of EVs in oncology research and their potential cargo may be novel biomarkers for early cancer detection and further diagnosis.

Prostate cancer is the fourth leading cause of death in men worldwide. Currently, the gold standard for PCa diagnosis remains the Prostate-Specific Antigen (PSA); therefore, there is a growing demand for novel diagnostic approaches for early prediction of aggressive types of prostate cancer. The complexity of tumors is further enhanced by the presence of extracellular vesicles (EVs). These membrane-enclosed spherical particles are directly derived from their parent cells with their information (mRNAs, non-coding RNAs, and proteins) and play a critical role in intercellular communication. However, the role of PCa EVs in the crosstalk between cancer cells and resident cells in the microenvironment is still under investigation. Different prostate cancer cell lines were cultured for 48 h in medium supplemented with FBS exo-free medium. Next, secreted EVs were separated using differential ultracentrifugation, and further EVs characterization was performed by nanoparticle tracking analysis, immunoblotting, and scanning electron microscopy. The effect of prostate cancer EVs on the immune cell population was analyzed after staining PBMCs using flow cytometry. Additionally, cytokines released into the conditioned media were evaluated using an ELISA assay. We illustrated how EVs from prostate cancer cells of different background variously affected immune cells and modulated the expression of several cytokines after 24 h of treatment. We observed that the number of rare monocytes, CD14 + CD16 + , was significantly reduced in PBMCs treated with PC-3 and DU 145 EVs compared with LNCaP EVs. Furthermore, CD4 + and CD8 + activated cells were significantly more responsive when PBMCs were treated with PC-3 EVs compared to LNCaP EVs. Subsequently, pro- and anti-inflammatory cytokines were found to be more abundant in samples treated with PC-3 and DU 145 EVs. This suggests a potential role of EVs in prostate cancer progression and immune system evasion. Our study provides preliminary evidence that prostate cancer EVs exert distinct immunomodulatory effects, depending on their cellular origin. Ultimately, the differences observed between PC-3 EVs, DU 145 EVs and LNCaP EVs highlight the potential use of immune cell markers as reliable indicators of disease progression and as a non-invasive tool to complement prostate cancer risk stratification strategies.