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Extracellular vesicles (EVs) are nanosized, membrane-bound structures that have emerged as promising tools for drug delivery, especially in the treatment of lysosomal storage disorders (LSDs) with central nervous system (CNS) involvement. This review highlights the unique properties of EVs, such as their biocompatibility, capacity to cross the blood–brain barrier (BBB), and potential for therapeutic cargo loading, including that of enzymes and genetic material. Current therapies for LSDs, like enzyme replacement therapy (ERT), often fail to address neurological symptoms due to their inability to cross the BBB. EVs offer a viable alternative, allowing for targeted delivery to the CNS and improving therapeutic outcomes. We discuss recent advancements in the engineering and modification of EVs to enhance targeting, circulation time and cargo stability, and provide a detailed overview of their application in LSDs, such as Gaucher and Fabry diseases, and Sanfilippo syndrome. Despite their potential, challenges remain in scaling production, ensuring isolation purity, and meeting regulatory requirements. Future developments will focus on overcoming these barriers, paving the way for the clinical translation of EV-based therapies in LSDs and other CNS disorders. [ABSTRACT FROM AUTHOR]

Cancer maintenance, metastatic dissemination and drug resistance are sustained by cancer stem cells (CSCs). Triple negative breast cancer (TNBC) is the breast cancer subtype with the highest number of CSCs and the poorest prognosis. Here, we aimed to identify potential drugs targeting CSCs to be further employed in combination with standard chemotherapy in TNBC treatment. The anti-CSC efficacy of up to 17 small drugs was tested in TNBC cell lines using cell viability assays on differentiated cancer cells and CSCs. Then, the effect of 2 selected drugs (8-quinolinol -8Q- and niclosamide -NCS-) in the cancer stemness features were evaluated using mammosphere growth, cell invasion, migration and anchorage-independent growth assays. Changes in the expression of stemness genes after 8Q or NCS treatment were also evaluated. Moreover, the potential synergism of 8Q and NCS with PTX on CSC proliferation and stemness-related signaling pathways was evaluated using TNBC cell lines, CSC-reporter sublines, and CSC-enriched mammospheres. Finally, the efficacy of NCS in combination with PTX was analyzed in vivo using an orthotopic mouse model of MDA-MB-231 cells. Among all tested drug candidates, 8Q and NCS showed remarkable specific anti-CSC activity in terms of CSC viability, migration, invasion and anchorage independent growth reduction in vitro. Moreover, specific 8Q/PTX and NCS/PTX ratios at which both drugs displayed a synergistic effect in different TNBC cell lines were identified. The sole use of PTX increased the relative presence of CSCs in TNBC cells, whereas the combination of 8Q and NCS counteracted this pro-CSC activity of PTX while significantly reducing cell viability. In vivo, the combination of NCS with PTX reduced tumor growth and limited the dissemination of the disease by reducing circulating tumor cells and the incidence of lung metastasis. The combination of 8Q and NCS with PTX at established ratios inhibits both the proliferation of differentiated cancer cells and the viability of CSCs, paving the way for more efficacious TNBC treatments. [ABSTRACT FROM AUTHOR]

Within tumors, Cancer Stem Cell (CSC) subpopulation has an important role in maintaining growth and dissemination while preserving high resistance against current treatments. It has been shown that, when CSCs are eliminated, the surrounding Differentiated Cancer Cells (DCCs) may reverse their phenotype and gain CSC-like features to preserve tumor progression and ensure tumor survival. This strongly suggests the existence of paracrine communication within tumor cells. It is evidenced that the molecular crosstalk is at least partly mediated by Extracellular Vesicles (EVs), which are cell-derived membranous nanoparticles that contain and transport complex molecules that can affect and modify the biological behavior of distal cells and their molecular background. This ability of directional transport of small molecules prospects EVs as natural Drug Delivery Systems (DDS). EVs present inherent homing abilities and are less immunogenic than synthetic nanoparticles, in general. Currently, strong efforts are focused into the development and improvement of EV-based DDS. Even though EV-DDS have already reached early phases in clinical trials, their clinical application is still far from commercialization since protocols for EVs loading, modification and isolation need to be standardized for large-scale production. Here, we summarized recent knowledge regarding the use of EVs as natural DDS against CSCs and cancer resistance. [ABSTRACT FROM AUTHOR]

Structural maintenance of chromosomes protein 2 (SMC2) is a central component of the condensin complex involved in DNA supercoiling, an essential process for embryonic stem cell survival. SMC2 over-expression has been related with tumorigenesis and cancer malignancy and its inhibition is regarded as a potential therapeutic strategy even though no drugs are currently available. Here, we propose to inhibit SMC2 by intracellular delivery of specific antibodies against the SMC2 protein. This strategy aims to reduce cancer malignancy by targeting cancer stem cells (CSC), the tumoral subpopulation responsible of tumor recurrence and metastasis. In order to prevent degradation and improve cellular internalization, anti-SMC2 antibodies (Ab-SMC2) were delivered by polymeric micelles (PM) based on Pluronic® F127 amphiphilic polymers. Importantly, scaffolding the Ab-SMC2 onto nanoparticles allowed its cellular internalization and highly increased its efficacy in terms of cytotoxicity and inhibition of tumorsphere formation in MDA-MB-231 and HCT116 breast and colon cancer cell lines, respectively. Moreover, in the case of the HCT116 cell line G1, cell-cycle arrest was also observed. In contrast, no effects from free Ab-SMC2 were detected in any case. Further, combination therapy of anti-SMC2 micelles with paclitaxel (PTX) and 5-Fluorouracil (5-FU) was also explored. For this, PTX and 5-FU were respectively loaded into an anti-SMC2 decorated PM. The efficacy of both encapsulated drugs was higher than their free forms in both the HCT116 and MDA-MB-231 cell lines. Remarkably, micelles loaded with Ab-SMC2 and PTX showed the highest efficacy in terms of inhibition of tumorsphere formation in HCT116 cells. Accordingly, our data clearly suggest an effective intracellular release of antibodies targeting SMC2 in these cell models and, further, strong cytotoxicity against CSC, alone and in combined treatments with Standard-of-Care drugs. [ABSTRACT FROM AUTHOR]

Inclusion bodies (IBs) are protein nanoclusters obtained during recombinant protein production processes, and several studies have demonstrated their potential as biomaterials for therapeutic protein delivery. Nevertheless, IBs have been, so far, exclusively sifted by their biological activity in vitro to be considered in further protein-based treatments in vivo. Matrix metalloproteinase-9 (MMP-9) protein, which has an important role facilitating the migration of immune cells, was used as model protein. The MMP-9 IBs were compared with their soluble counterpart and with MMP-9 encapsulated in polymeric-based micelles (PM) through ionic and covalent binding. The soluble MMP-9 and the MMP-9-ionic PM showed the highest activity values in vitro. IBs showed the lowest activity values in vitro, but the specific activity evolution in 50% bovine serum at room temperature proved that they were the most stable format. The data obtained with the use of an air-pouch mouse model showed that MMP-9 IBs presented the highest in vivo activity compared to the soluble MMP-9, which was associated only to a low and a transitory peak of activity. These results demonstrated that the in vivo performance is the addition of many parameters that did not always correlate with the in vitro behavior of the protein of interest, becoming especially relevant at evaluating the potential of IBs as a protein-based nanomaterial for therapeutic purposes. [ABSTRACT FROM AUTHOR]

Therapeutic resistance seen in aggressive forms of breast cancer remains challenging for current treatments. More than half of the patients suffer from a disease relapse, most of them with distant metastases. Cancer maintenance, resistance to therapy, and metastatic disease seem to be sustained by the presence of cancer stem cells (CSC) within a tumor. The difficulty in targeting this subpopulation derives from their dynamic interconversion process, where CSC can differentiate to non-CSC, which in turn de-differentiate into cells with CSC properties. Using fluorescent CSC models driven by the expression of ALDH1A 1(aldehyde dehydrogenase 1A1), we confirmed this dynamic phenotypic change in MDA-MB-231 breast cancer cells and to identify Serine/Threonine Kinase 2 (AKT2) as an important player in the process. To confirm the central role of AKT2, we silenced AKT2 expression via small interfering RNA and using a chemical inhibitor (CCT128930), in both CSC and non-CSC from different cancer cell lines. Our results revealed that AKT2 inhibition effectively prevents non-CSC reversion through mesenchymal to epithelial transition, reducing invasion and colony formation ability of both, non-CSC and CSC. Further, AKT2 inhibition reduced CSC survival in low attachment conditions. Interestingly, in orthotopic tumor mouse models, high expression levels of AKT2 were detected in circulating tumor cells (CTC). These findings suggest AKT2 as a promising target for future anti-cancer therapies at three important levels: (i) Epithelial-to-mesenchymal transition (EMT) reversion and maintenance of CSC subpopulation in primary tumors, (ii) reduction of CTC and the likelihood of metastatic spread, and (iii) prevention of tumor recurrence through inhibition of CSC tumorigenic and metastatic potential. [ABSTRACT FROM AUTHOR]