Over the past decade, there has been a significant increase in preclinical development and clinical use of approved nanomedicines. The nanomedicinal field represents a wide variety of products including early diagnostics such as in vivo imaging, medicinal devices, and innovative therapeutics. Nanomedicines exhibit unique physiochemical properties compared with their nonnanoformulated counterparts; however, clinical reports of moderate-to-severe allergic, anaphylaxis, and hypersensitivity reactions raise major concerns with regard to nanomedicine immunotoxicity. This review summarizes some of the main applications of nanomedicines, safety, and immunotoxicity implications and current testing strategies.

Osteoarthritis (OA) is a degenerative joint disease, causing impaired mobility. There are currently no effective therapies other than palliative treatment. Mesenchymal stromal cells (MSCs) and their secreted extracellular vesicles (MSC-EVs) have shown promise in attenuating OA progression, promoting chondral regeneration, and modulating joint inflammation. However, the precise molecular mechanism of action driving their beneficial effects has not been fully elucidated. In this study, we analyzed MSC-EV-treated human OA chondrocytes (OACs) to assess viability, proliferation, migration, cytokine and catabolic protein expression, and microRNA and mRNA profiles. We observed that MSC-EV-treated OACs displayed increased metabolic activity, proliferation, and migration compared to the controls. They produced decreased proinflammatory (Il-8 and IFN-γ) and increased anti-inflammatory (IL-13) cytokines, and lower levels of MMP13 protein coupled with reduced expression of MMP13 mRNA, as well as negative microRNA regulators of chondrogenesis (miR-145-5p and miR-21-5p). In 3D models, MSC-EV-treated OACs exhibited enhanced chondrogenesis-promoting features (elevated sGAG, ACAN, and aggrecan). MSC-EV treatment also reversed the pathological impact of IL-1β on chondrogenic gene expression and extracellular matrix component (ECM) production. Finally, MSC-EV-treated OACs demonstrated the enhanced expression of genes associated with cartilage function, collagen biosynthesis, and ECM organization and exhibited a signature of 24 differentially expressed microRNAs, associated with chondrogenesis-associated pathways and ECM interactions. In conclusion, our data provide new insights on the potential mechanism of action of MSC-EVs as a treatment option for early-stage OA, including transcriptomic analysis of MSC-EV-treated OA, which may pave the way for more targeted novel therapeutics.

Biologics, including monoclonal antibodies (mAb), have proved to be effective and successful therapeutic agents, particularly in the treatment of cancer and immune-inflammatory conditions, as well as allergies and infections. However, their use carries an inherent risk of an immune-mediated adverse drug reaction. In this study, we describe the use of a novel pre-clinical human in vitro skin explant test for predicting skin sensitization and adverse immune reactions. The skin explant test was used to investigate the effects of therapeutic antibodies, which are known to cause a limited reaction in a small number of patients or more severe reactions. Material and Methods: Immune responses were determined by T cell proliferation and multiplex cytokine analysis, as well as histopathological analysis of skin damage (grades I–IV in increasing severity), predicting a negative (grade I) or positive (grade ≥ II) response for an adverse skin sensitization effect. Results: T cell proliferation responses were significantly increased in the positive group (p < 0.004). Multiplex cytokine analysis showed significantly increased levels of IFNγ, TNFα, IL-10, IL-12, IL-13, IL-1β, and IL-4 in the positive response group compared with the negative response group (p < 0.0001, p < 0.0001, p < 0.002, p < 0.01, p < 0.04, p < 0.006, and p < 0.004, respectively). Conclusions: Overall, the skin explant test correctly predicted the clinical outcome of 13 out of 16 therapeutic monoclonal antibodies with a correlation coefficient of 0.770 (p = 0.0001). This assay therefore provides a valuable pre-clinical test for predicting adverse immune reactions, including T cell proliferation and cytokine release, both associated with skin sensitization to monoclonal antibodies.

Mesenchymal stromal cells (MSCs) have demonstrated therapeutic efficacy across numerous clinical applications, with evidence suggesting their paracrine effects, particularly through extracellular vesicles (EVs), possibly driving functional outcomes. In this study we perform the comprehensive characterization of microRNA expression profiles in human MSC-derived EVs (MSC-EV) compared to their parental cells, cultured under clinically relevant xeno-free conditions. MSCs were isolated from the bone marrows of healthy donors and characterised according to the International Society for Cellular Therapy criteria, while MSC-EVs were isolated using differential ultracentrifugation and validated according to the International Society for Extracellular Vesicle guidelines. NanoString profiling identified 590 mature microRNAs expressed across both populations, with 42 being significantly differentially expressed between MSC-EVs and parental MSCs. Five microRNAs were distinctly highly expressed in MSCs and five in MSC-EVs, while fifteen of the top twenty most abundant microRNAs showed high expression in both populations. MicroRNA expression patterns were validated in an independent cohort. Functional pathway analysis of differentially expressed microRNAs showed enrichment of key biological processes including cell proliferation, differentiation, and immune regulation. This standardised profiling approach develops our understanding of MSC/MSC-EV microRNA cargo, using a transparent methodological approach that allows for the improved comparability of datasets for the development and advancement of MSC-EV therapeutics.

Introduction: Monoclonal antibodies (mAbs) are important therapeutics. However, the enhanced potential for aggregation has become a critical quality parameter during the production of mAbs. Furthermore, mAb aggregation may also present a potential health risk in a clinical setting during the administration of mAb therapeutics to patients. While the extent of immunotoxicity in patient populations is uncertain, reports show it can lead to immune responses via cell activation and cytokine release. In this study, an autologous in vitro skin test designed to predict adverse immune events, including skin sensitization, was used as a novel assay for the assessment of immunotoxicity caused by mAb aggregation. Material and Methods: Aggregation of mAbs was induced by a heat stress protocol, followed by characterization of protein content by analytical ultra-centrifugation and transmission electron microscopy, revealing a 4% aggregation level of total protein content. Immunotoxicity and potential skin sensitization caused by the aggregates, were then tested in a skin explant assay. Results: Aggregated Herceptin and Rituximab caused skin sensitization, as shown by histopathological damage (grade II–III positive response) together with positive staining for Heat Shock Protein 70 (HSP70). Changes in T cell proliferation were not observed. Cytokine analysis revealed a significant increase of IL-10 for the most extreme condition of aggregation (65 °C at pH3) and a trend for an overall increase of IFN-γ, especially in response to Rituximab. Conclusions: The skin explant assay demonstrated that aggregated mAbs showed adverse immune reactions, as demonstrated as skin sensitization, with histopathological grades II-III. The assay may, therefore, be a novel tool for assessing immunotoxicity and skin sensitization caused by mAb aggregation.