"Small extracellular vesicles: messengers at the service of breast cancer agenda in the primary and distant microenvironments"

Abstract

Breast cancer (BC) remains a leading cause of cancer-related mortality in women, with complex mechanisms driving its initiation, progression, and resistance to therapy. In recent years, the tumor microenvironment (TME) has gained attention for its critical role in shaping tumor behavior, where small extracellular vesicles (small EVs) have emerged as key mediators of intercellular communication. These vesicles carry a diverse cargo of proteins, lipids, DNA, and various non-coding RNAs-such as miR-21, miR-155, and miR-1246-mirroring the molecular status of their originating cells. This review highlights the roles of small EVs in immune modulation, stromal remodelling, and metastatic niche formation, emphasizing their contribution to therapy resistance and immune evasion. We discuss recent updates on EV biogenesis, characterisation and isolation techniques, such as ultracentrifugation, immunoaffinity and microfluidic systems. We also critically evaluate their potential for clinical application and how well they conform to the MISEV2023 guidelines. Furthermore, we examine small EVs as diagnostic tools in liquid biopsies and compare them with conventional methods such as mammography and tissue biopsies. We also discuss organotropism mediated by small EV cargo (e.g., integrins α6β4, αvβ5) and the diagnostic potential of protein and lipid signatures (e.g., PD-L1, CD63, and exosomal lipidomics). Therapeutically, we explore engineered small EVs for drug delivery, gene modulation, and immune activation, addressing challenges of targeting efficiency, in vivo stability, immunogenicity, and clinical scalability. The review discusses ongoing clinical trials involving small EVs in BC and highlights key translational gaps between preclinical advances and clinical implementation. Finally, we explores how integrating artificial intelligence, single-cell transcriptomics, and multi-omics approaches can help overcome major challenges such as small EV heterogeneity and tracking limitations. Crucially, this integration enables a more tailored understanding of each patient's tumor biology, reducing therapeutic failures by guiding more personalized and effective treatment strategies. Overall, small EVs represent a transformative tool in precision oncology, contingent on resolving key challenges in their clinical translation.

Keywords: Breast cancer; Immune cells; MicroRNAs; Microenvironment; Small extracellular vesicles; Stroma cells.

Fig. 1

Role of small EVs in breast cancer (BC) progression and metastasis. The involvement of small EVs in the metastatic cascade of BC. Within the primary tumor microenvironment, tumor-derived small EVs mediate crosstalk with both immune (macrophages, dendritic cells, T lymphocytes, myeloid-derived suppressor cells) and non-immune stromal cells (fibroblasts, endothelial cells, adipocytes), as well as the extracellular matrix. Small EVs facilitate tumor progression by promoting invasion and intravasation of cancer cells into the circulatory system. These vesicles also contribute to the formation of pre-metastatic niches at distant sites (lung, bone, brain) by transferring oncogenic cargo—miRNAs, proteins, circular RNAs, lncRNAs, and lipids—to resident cells, thus supporting extravasation and colonization. This highlights the central role of small EVs in preparing metastatic sites and modulating the tumor microenvironment at both primary and distant locations (Created with BioRender.com)

Fig. 2

Extracellular vesicles (EVs) as liquid biopsy tools and therapeutic vehicles in BC. The panel illustrates the structural and functional features of EVs, highlighting their diverse cargo—nucleic acids, proteins, and lipids—and surface components including tetraspanins, adhesion molecules, and antigens. EVs can be isolated from patient blood samples along with other liquid biopsy components such as circulating tumor cells (CTCs), circulating free DNA (cfDNA), proteins, metabolites, and microRNAs (miRNAs). Tumor-derived EVs carry bioactive molecules, including specific proteins (e.g., CD47, CD24, CD9, EpCAM), lipids, and non-coding RNAs (miRNAs, circRNAs, lncRNAs), which can serve as diagnostic and prognostic biomarkers. Additionally, EVs hold promise as therapeutic tools in oncology, acting as drug delivery vehicles, immunotherapy enhancers, and agents for overcoming drug resistance or targeting metastasis. EVs represent versatile components of liquid biopsy with potential applications in early diagnosis, prognostic assessment, longitudinal disease monitoring, and evaluation of therapeutic efficacy (Created with BioRender.com)