Poly(methacrylate citric acid) as a Dual Functional Carrier for Tumor Therapy

Abstract

Owing to its pH-sensitive property and chelating Cu²⁺ effect, poly(methacrylate citric acid) (PCA) can be utilized as a dual functional nanocarrier to construct a nanodelivery system. Negatively charged carboxyl groups can interact with positively charged antineoplastic drugs through electrostatic interaction to form stable drug nanoparticles (NPs). Through drug experimental screening, doxorubicin (DOX) was selected as the model drug, PCA/DOX NPs with a diameter of 84 nm were prepared, and the drug-loading content was 68.3%. PCA/DOX NPs maintained good stability and a sustained release profile. Cell experiments presented that PCA/DOX NPs could inhibit effectively the growth of 4T1 cells; the IC₅₀ value was decreased by approximately 15-fold after incubation for 72 h. The cytotoxicity toward H9C2 was decreased significantly. Moreover, based on its ability to efficiently adsorb copper ions, PCA showed good vascular growth inhibition effect in vitro. Furthermore, animal experiments showed that PCA/DOX NPs presented stronger anticancer effects than DOX; the tumor inhibition rate was increased by 1.5-fold. Myocardial toxicity experiments also confirmed that PCA reduced the cardiotoxicity of DOX. In summary, PCA/DOX NPs show good antitumor efficacy and low toxicity, and have good potential for clinical application.

Keywords: chelating Cu2+ effect; doxorubicin; pH sensitive; poly(methacrylate citric acid).

Figure 1

Synthesis and characterization of the prepared drug-loaded nanoparticles. (a) Images of the drug-loaded nanoparticles from left to right: RES, NIF, IBU, HCPT, CSL, HK, DOX, and POD. (b) Preparation mechanism of PCA/DOX NPs. (c) Particle size distribution curve of PCA/DOX NPs. (d) SEM image of PCA/DOX NPs. Scale bar 500 nm.

Figure 2

The stability of PCA/DOX NPs: (a) storage stability at 4 °C; (b) media stability at 37 °C, n = 3.

Figure 3

Cumulative release curves of PCA/DOX NPs and DOX in PBS (pH 7.4 and 5.5) at 37 °C (pH 7.4 imitates the normal human environment, and pH 5.5 imitates the acidic environment of the tumor site), n = 3.

Figure 4

Cytotoxicity investigation results against 4T1 tumor cell line (a) and normal H9C2 cell line (b) at 37 °C after coincubation for 48 h, n = 5.

Figure 5

Vascular growth inhibition experiment: HUVEC cells culturing with blank ECGM medium, ECGM medium containing Cu²⁺ ions (25 μmol/L), and ECGM medium containing Cu²⁺ ions (25 μmol/L) and PCA (100 μmol/L) on Matrigel for 24 h. Cells were observed using optical microscopy.

Figure 6

The result of antitumor efficacy: 4T1 tumor volume change curves (a), tumor weight and tumor inhibition rate (b), tumor tissue images of DOX (c), and PCA/DOX NPs (d), scale bar: 100 μm. *** p < 0.001, ** p < 0.01, vs. Glu group, ^# p < 0.05, vs. DOX group, n = 10.

Figure 7

The result of DOX toxicity test: (a) body weight change curves; (b) liver and spleen index. *** p < 0.001, ** p < 0.01, vs. Glu group; ## p < 0.01, vs. DOX group, n = 10.

Figure 8

Cardiotoxicity test results: (a) heart index of different groups; (b) mice biochemical index results; (c) HE images—there was obvious myocardial interstitial edema in the DOX and PCA/DOX NPs groups (marked by the black arrow). The cardiomyocytes presented obvious nucleus lysis and apoptosis in the DOX group (marked by the red arrow), scale bar: 100 μm. *** p < 0.001, ** p < 0.01, * p < 0.05, vs. Glu group; ### p < 0.001, ## p < 0.01, vs. DOX group, n = 10.