Restraining small extracellular vesicles: Dawn of a new era in nanomedicine

In a recent paper published in Nature Materials, Gong et al. identified the tumor cell-derived small extracellular vesicles (sEVs) as a defense system that impedes selective delivery of nanoparticles to tumors. The authors also discovered that this defense system could be a prospective target for enhancing the efficacy of nanoparticle-based tumor therapies (Figure 1).¹

In the past 30 years, research in the field of cancer nanomedicine has seen tremendous expansion. Various types of nanoparticles, including lipid-based nanoparticles, polymeric nanoparticles, and inorganic nanoparticles, have been developed for targeted delivery to kill tumor cells and/or regulate the tumor microenvironment. These nanoparticles can encapsulate a variety of therapeutic payloads, including small molecules, biologics, and nucleic acids.² However, tumoritropic accumulation of cancer nanomedicines vary widely from tumor to tumor and from patient to patient.² Nanomedicines primarily accumulate passively in solid tumors through the enhanced permeability and retention (EPR) effect; however, factors such as tumor etiology, type, location, size, stage, microenvironment, vascular density, and blood perfusion status can largely cause the heterogeneity of the EPR effect. For instance, hepatocellular carcinoma and renal cell carcinoma exhibit higher vascular density, resulting in a more pronounced EPR effect compared to pancreatic cancer and prostate cancer.² Achieving high concentrations of nanomedicine at the tumor site remains a critical research focus in the field.

To increase the concentration of nanoparticles accumulated inside the tumor, solid stress, dense extracellular matrix, and abnormal vascular structures within the tumor microenvironment have been considered. Despite the considerable efforts of numerous researchers, on average, only a small fraction of injected nanoparticles reach tumors.³

Recent studies have found that tumor cells secrete numerous exotic proteins into the tumor microenvironment, which can mediate tumor cell communication, induce immunosuppression, or promote metastasis. These effects are all mediated by nucleic acids or proteins wrapped inside the exosomes. However, the physicochemical function of high-concentration exosomes at the tumor site is a long-neglected direction of research. Especially in the field of drug delivery, how the nanoscale of exosomes, composition, physical interactions, etc., affect drug delivery is an unexplored field. The size of exosomes directly influences their permeability and cellular uptake within the tumor microenvironment. Furthermore, the composition of exosomes determines their circulation time in the bloodstream and their interactions with cells. Additionally, there may be physical interactions between exosomes and nanoparticles, such as van der Waals forces. This association could lead to a reduced accumulation of nanoparticles in tumor tissues, making them more readily delivered to the Kupffer cells in the liver for uptake and degradation, rather than being taken up by tumor cells.

As a class of small vesicles secreted by cells (primarily as exosomes), sEVs are widely present in various tissue environments, particularly at higher concentrations in the tumor microenvironment, playing potential roles in tumor growth, invasion, angiogenesis, metastasis, immune response, and chemotherapy drug resistance.⁴ Elevated levels of sEVs in solid tumors may affect nanoparticle accumulation by creating a steep sEV gradient between tumors and normal tissues, which can form a biological barrier that limits nanoparticle penetration and accumulation. However, this phenomenon remains under-studied.

To overcome the complex microenvironment constituted by the dense extracellular matrix of tumors, solid stress, and abnormal vascular structures, the authors initially utilized CRISPR-Cas9 technology to knock out the key gene Rab27a, which regulates sEVs secretion, in a mouse model. They found Rab27a knockout can significantly reduce the secretion of sEVs in mouse tumor cells and greatly increase the accumulation of lipid nanoparticles (LNPs) in the tumors. Further investigation found that, through interactions such as van der Waals forces, sEVs can bind to nanoparticles and physically transport them to the Kupffer cells in the liver for degradation, consequently diminishing the accumulation of nanoparticles in the tumors. Similarly, relevant in vitro cell experiments have shown that sEVs can influence the cellular uptake of LNPs. By blocking the adhesion molecules, intercellular cell adhesion molecule-1 (ICAM-1), on the surface of sEVs with corresponding antibodies, the uptake of LNPs by Kupffer cells was greatly decreased. Moreover, Kupffer cells expressed higher levels of macrophage-1 antigen (Mac-1, the receptor for ICAM-1) compared to other cell subsets in the liver. This finding elucidates the mechanism behind the uptake of LNPs by Kupffer cells: LNP binds to tumor-derived sEVs, forming a LNP-sEV complex, which is then specifically taken up via the ICAM-1-Mac-1 interaction.¹

Overall, this paper describes detailed experiments and in-depth mechanistic analyses to reveal how tumor-derived sEVs hinder the delivery of nanoparticles. Traditionally, in cell biology, exosomes are believed to transport intracellular waste to the extracellular space, serving to recycle cell surface materials and facilitate intercellular communication or material transfer through secretion and re-endocytosis. This paper reveals a novel cellular biological function of exosomes, demonstrating their ability to mediate intercellular material transport, extending beyond the traditional properties of nanoparticles themselves.

It was also discovered that sEVs secreted by tumor cells act as an “active defense system.” These sEVs can bind to various therapeutic agents entering the tumor and transport them to hepatic Kupffer cells for degradation, thereby hindering selective delivery of nanoparticles to the tumor. Traditional theories of cellular defense suggest that cellular defense comprises intracellular pattern recognition receptors and corresponding defense signals, immune cells, antibodies, and complements. This paper discovers that exosomes can also function as a form of physical defense—a part of the cellular physical defense mechanisms. On the other hand, exosomes are closely linked to the occurrence and progression of cancer. Within the tumor microenvironment, exosomes can transfer bioactive molecules between tumor cells, immune cells, and stromal cells, aiding cancer cells in evading immune surveillance and inducing immune tolerance.

Furthermore, research has also found that exosomes expressing PD-L1 from tumor cells, immune cells, mesenchymal stem cells, or other cells outside the tumor microenvironment can promote tumor evasion. In addition to PD-L1, sEVs may contain other proteins that could interact with nanoparticles, affecting their stability, targeting, or cellular uptake. For instance, other immune regulatory molecules present on sEVs may influence the behavior of nanoparticles within the tumor microenvironment. By modifying the surface of nanoparticles with specific ligands, such as antibodies or peptides targeting tumor cell-specific receptors, the specificity of nanoparticles binding to tumor cells can be enhanced, while reducing nonspecific interactions with sEVs. Developing nanoparticles whose surface characteristics can be altered in response to specific stimuli (such as pH changes, enzymatic activity, or temperature variations) to release drugs within the tumor microenvironment can minimize interactions with sEVs. In addition to directly modifying nanoparticles, developing drugs that can inhibit sEV secretion or neutralize key proteins on sEVs (such as PD-L1) could serve as a combinatorial therapeutic strategy to enhance the efficacy of nanoparticles. Understanding how sEVs influence the behavior of nanoparticles and developing strategies to overcome these effects is crucial for improving the therapeutic efficacy of nanoparticles in cancer treatment. This may require interdisciplinary collaboration, bringing together researchers from fields such as materials science, immunology, and oncology.⁵

In summary, this study offers a potential strategy to enhance nanoparticle-based cancer therapies by overcoming the defense mechanisms of tumor cell sEVs. The findings not only deepen the understanding of sEVs' role in the tumor microenvironment but also provide valuable insights for developing new cancer treatment strategies.

Ming Yang: Conceptualization (equal); investigation (equal); methodology (equal); resources (equal); visualization (equal); writing—original draft (equal). Lin-Zhu Zhang: Conceptualization (equal); formal analysis (equal); methodology (equal); writing—original draft (equal); writing—review and editing (equal). Hai-Dong Zhu: Conceptualization (lead); funding acquisition (lead); project administration (lead); supervision (lead); writing—review and editing (lead). All authors have read and approved the final manuscript.

The authors declare no conflicts of interest.

Not applicable.