A Meta-analysis on the Effectiveness of Extracellular Vesicles as Nanosystems for Targeted Delivery of Anticancer Drugs

While the efficacy of anticancer drugs is hampered by low bioavailability and systemic toxicity, the uncertainty remains whether encapsulation of these drugs into natural nanovesicles such as extracellular vesicles (EVs) could improve controlled drug release and efficacy for targeted tumor therapy. Thus, we performed a meta-analysis for studies reporting the efficacy of EVs as nanosystems to deliver drugs and nucleic acid, protein, and virus (NPV) to tumors using the random-effects model. The electronic search of articles was conducted through Cochrane, PubMed, Scopus, Science Direct, and Clinical Trials Registry from inception up till September 2022. The pooled summary estimate and 95% confidence interval of tumor growth inhibition, survival, and tumor targeting were obtained to assess the efficacy. The search yielded a total of 119 studies that met the inclusion criteria having only 1 clinical study. It was observed that the drug-loaded EV was more efficacious than the free drug in reducing tumor volume and weight with the standardized mean difference (SMD) of −1.99 (95% CI: −2.36, −1.63; p < 0.00001) and −2.12 (95% CI: −2.48, −1.77; p < 0.00001). Similarly, the mean estimate of tumor volume and weight for NPV were the following: SMD: −2.30, 95% CI: −3.03, −1.58; p < 0.00001 and SMD: −2.05, 95% CI: −2.79, −1.30; p < 0.00001. Treatment of tumors with EV-loaded anticancer agents also prolonged survival (HR: 0.15, 95% CI: 0.10, 0.22, p < 0.00001). Furthermore, EVs significantly delivered drugs to tumors as revealed by the higher concentration at the tumor site (SMD: −2.73, 95% CI: −3.77, −1.69; p < 0.00001). This meta-analysis revealed that EV-loaded drugs and NPV performed significantly better in tumor growth inhibition with improved survival than the free anticancer agents, suggesting EVs as safe nanoplatforms for targeted tumor therapy

Acridine orange (AO, green) and propidium iodide (PI, red) double staining fluorescent micrographs of differentiated neuronal cells.

<p>(a) 4 h H₂O₂ treated cells, (b) 72 h myrosinase pre-treated plus 4 h H₂O₂ exposed cells, (c) 72 h 1.25 μg/ml GMG-ITC pre-treated plus 4 h H₂O₂ exposed cells, (d) untreated cells (normal control). The images were captured in multiple times and x20 magnification was used.</p

Ultrastructural analysis of differentiated neuronal cells by transmission electron microscopy.

<p>(a) 4 h H₂O₂ treated cells, (b) 72 h myrosinase pre-treated plus 4 h H₂O₂ exposure cells, (c) 72 h GMG-ITC pre-treated plus 4 h H₂O₂ exposure cells, (d) untreated (normal control) cells. CM = chromatin margination, IN = intact nucleus, LD = lipid droplet, NC = nuclei convolution. Magnification (x 3000).</p

Cytotoxicity of GMG-ITC on differentiated neuronal cells at different concentrations (0.313 to 10) μg/ml.

<p>(A) display 24 h, (B) 48 h and (C) 72 h of treatment. Whereas (D) is a cytotoxic analysis result of H₂O₂ used in this study with IC₅₀ = 300 μM. Values are presented in means ± SD of triplicate experiments and means with different letters varies significantly (p<0.05).</p

Surface morphological analysis of differentiated neuronal cells by scanning electron microscopy.

<p>(a) 4 h H₂O₂ treated cells, (b) 72 h myrosinase pre-treated plus 4 h H₂O₂ exposure cells, (c) 72 h GMG-ITC pre-treated plus 4 h H₂O₂ exposure cells, (d) untreated (normal control) cells. AB = apoptotic body, IDVC = intact differentiated viable cells, FN = folded neurites, MB = membrane blabbing, NDAC = neurite disrupted apoptotic cells. Magnification (x 5000).</p

Micrographs of neuronal cells differentiation by 10 μM all trans retinoic acid (ATRA).

<p>(a) Undifferentiated cells cultured in 10% complete growth media for seven (7) days and viewed under phase contrast, (b) Differentiated cells cultured in 3% heat-inactivated FBS complete growth media containing 10 μM ATRA for seven (7) days and viewed under phase contrast, and (d) expressed tuj-1 in both cytoplasm and neurites. SN = short neurites, EN = extended neurites, CYP = cytoplasm,. Magnification (x 20).</p

Annexin V-FITC assay of differentiated neuronal cells analysed by flow cytometry.

<p>Where (a) 4 h H₂O₂ treated cells, (b) 72 h myrosinase pre-treated plus 4 h H₂O₂ exposure cells, (c) untreated (normal control) cells, (d) GMG-ITC pre-treated for 24 h plus 4 h H₂O₂ exposure, (e) GMG-ITC pre-treated for 48 h plus 4 h H₂O₂ exposure and (f) GMG-ITC pre-treated for 72 h plus 4 h H₂O₂ exposure. Whereas (g) represent distribution of cells at death. Values are presented in means ± SD of triplicate experiments and means of viable cells with different letters varies significantly (p<0.05).</p