Research ArticleHEALTH AND MEDICINE

Erythrocyte leveraged chemotherapy (ELeCt): Nanoparticle assembly on erythrocyte surface to combat lung metastasis

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Science Advances  13 Nov 2019:
Vol. 5, no. 11, eaax9250
DOI: 10.1126/sciadv.aax9250
  • Fig. 1 Schematic illustration of the ELeCt platform and characterization of drug (DOX)–loaded biodegradable PLGA NPs.

    (A) Schematic illustration of the composition and mechanism of the biodegradable drug NP assembling on the erythrocyte platform (ELeCt) for lung metastasis treatment. (B to D) Average size (B), zeta potential (C), and drug loading contents (D) of plain and drug-loaded NPs. (E) SEM images showing the morphological features of the NPs. Scale bars, 200 nm. (F) Size distribution of plain and drug-loaded NPs. (G) Drug release kinetics from the biodegradable NPs in a complete medium (n = 4). (H and I) Flow cytometry histogram plots (H) and CLSM images (I) showing the interaction of drug-loaded NPs with B16F10-Luc melanoma cells. In (I), cell nuclei were stained using 4′,6-diamidino-2-phenylindole (DAPI). (J and K) Dose-response curve (J) and median inhibitory concentration (IC50) values (K) of B16F10-Luc cells after being treated with different formulations for 24 hours (n = 6). n.s., not significantly different (Student’s t test).

  • Fig. 2 Doxorubicin-loaded biodegradable PLGA NPs efficiently assemble onto mouse and human erythrocytes.

    (A) Flow cytometry analysis of assembly of DOX-loaded PLGA NPs to mouse erythrocytes at different NP-to-erythrocyte ratios (left to right: 0:1, 50:1, 200:1, 400:1, and 800:1). (B) Percentage of mouse erythrocytes carrying at least one NP. (C) Nanoparticle binding efficiency and (D) drug dose on mouse erythrocytes at different NP–to–mouse erythrocyte ratios. (E) CLSM and (F) SEM images of mouse erythrocytes with drug-loaded NPs assembled on them. Scale bars in (F), 2 μm. (G) CLSM and (H) SEM images of human erythrocytes with drug-loaded NPs assembled on them. Scale bars in (H), 2 μm. (I) Flow cytometry assay of the assembly of drug-loaded NPs to human erythrocytes at different NP-to-erythrocyte ratios (left to right: 0:1, 200:1, 800:1, and 1600:1). (J) Nanoparticle binding efficiency and (K) drug dose on human erythrocytes at different NP-to-erythrocyte ratios.

  • Fig. 3 The ELeCt platform enables enhanced and targeted delivery of NP drugs to the lungs bearing metastasis.

    (A) Pharmacokinetics of intravenously administered DOX formulations. Extended blood circulation time of DOX was achieved by erythrocyte hitchhiking compared with using free drug or NPs alone (n = 3). Significantly different [one-way analysis of variance (ANOVA)]: *P < 0.05 and **P < 0.01. (B) Hitchhiked drug-loaded NPs could specifically detach from mouse and human erythrocytes under the lung-corresponding shear stress. Samples were sheared for 20 min (n = 3). Low shear indicates rotary shear (~1 Pa), while high shear was at 6 Pa. Significantly different (Student’s t test): ***P < 0.001. (C) Drug accumulation in the lungs of mice bearing B16F10-Luc lung metastasis at 20 min and 6 hours after intravenous administration of different DOX formulations (n = 3). Significantly different (one-way ANOVA): *P < 0.05 and ***P < 0.001. (D) Comparison of the drug concentration in the lungs of erythrocyte hitchhiking group to that of the free drug and NP-alone groups (n = 3). (E) Drug distribution in the diseased lungs 20 min after intravenous administration of DOX formulations. Dashed lines indicate the edge of metastasis nodules.

  • Fig. 4 The ELeCt platform inhibits lung metastasis progression and improves survival in the early-stage B16F10-Luc metastasis model.

    (A) Schematic chart of the treatment schedule. (B) Bioluminescence images of lung metastasis at different time points. EXP indicates “Expired.” (C) Lung metastasis progression curve as depicted from in vivo bioluminescence signal intensity. (D) Quantification of lung metastasis burden at different time points (n = 7). (E) Scatter plot comparing the lung metastasis burden in different treatment groups as depicted from bioluminescence signal intensity on day 16 (n = 7). Significantly different (Kruskal-Wallis test): *P < 0.05, **P < 0.01, and ****P < 0.0001. (F) Scatter plot comparison of the lung metastasis burden on day 23 (n = 7). Significantly different (Kruskal-Wallis test): *P < 0.05, **P < 0.01, and ****P < 0.0001. (G) Body weight change of mice during the treatment period (n = 7). (H) Survival of mice under different treatments as displayed by Kaplan-Meier curves (n = 7). Significantly different (log-rank test): *P < 0.05 and ***P < 0.001.

  • Fig. 5 The ELeCt platform inhibits lung metastasis progression and extends survival in the late-stage B16F10-Luc metastasis model.

    (A) Schematic illustration of the treatment schedule. (B) Bioluminescence images of lung metastasis progression at different time points. (C) Lung metastasis growth curve in mice treated with different DOX formulations. (D) Quantitative analysis of lung metastasis burden as depicted from bioluminescence signal intensity (n = 7). Significantly different (one-way ANOVA): *P < 0.05 and **P < 0.01. (E) Quantification of metastasis nodule numbers on excised lungs from mice in different treatment groups on day 16 (n = 7). Significantly different (one-way ANOVA): **P < 0.01 and ***P < 0.001. (F) Body weight change of mice during the treatment period (n = 7). (G) Kaplan-Meier survival curves of mice in different treatment groups. Significantly different (log-rank test): **P < 0.01 and ***P < 0.001.

  • Fig. 6 Other chemotherapeutic agent–loaded biodegradable NPs can efficiently bind to erythrocytes.

    The tested chemotherapeutic agents include camptothecin, paclitaxel, docetaxel, 5-fluorouracil, gemcitabine, methotrexate, and the combination of 5-fluorouracil and methotrexate. Scale bars, 1 μm.

Supplementary Materials

  • Supplementary material for this article is available at http://advances.sciencemag.org/cgi/content/full/5/11/eaax9250/DC1

    Supplementary Materials and Methods

    Fig. S1. Representative H&E staining images of lungs of mice.

    Fig. S2. Representative H&E staining images of organs of mice treated with different drug formulations.

    Fig. S3. Size distribution of different chemotherapeutic agent–loaded biodegradable PLGA NPs.

    Table S1. Physicochemical properties of different chemotherapeutic agent–loaded biodegradable PLGA NPs.

  • Supplementary Materials

    This PDF file includes:

    • Supplementary Materials and Methods
    • Fig. S1. Representative H&E staining images of lungs of mice.
    • Fig. S2. Representative H&E staining images of organs of mice treated with different drug formulations.
    • Fig. S3. Size distribution of different chemotherapeutic agent–loaded biodegradable PLGA NPs.
    • Table S1. Physicochemical properties of different chemotherapeutic agent–loaded biodegradable PLGA NPs.

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