Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2018 Jun 27:13:3713-3728.
doi: 10.2147/IJN.S162939. eCollection 2018.

Doxorubicin and anti-VEGF siRNA co-delivery via nano-graphene oxide for enhanced cancer therapy in vitro and in vivo

Qi Sun  1   2   3 , Xiaoli Wang  1   2   3 , Chunying Cui  1   2   3 , Jing Li  1   2   3 , Yifan Wang  1   2   3
Affiliations

Doxorubicin and anti-VEGF siRNA co-delivery via nano-graphene oxide for enhanced cancer therapy in vitro and in vivo

Qi Sun et al. Int J Nanomedicine. .

Abstract

Background: Graphene oxide (GO) has attracted intensive interest in biological and medical fields in recent years due to its unique physical, chemical, and biological properties. In our previous work, we proved that GO could deliver small interfering RNA (siRNA) into cells and downregulate the expression of the desired gene.

Methods: This study investigated the potential of a modified GO nanocarrier for co-delivery of siRNA and doxorubicin (DOX) for enhanced cancer therapy. Fourier transform infrared spectroscopy, laser particle size analyzer, UV-visible spectroscopy, gel electrophoresis retardation, and in vitro release assay were studied.

Results: The results of real-time polymerase chain reaction revealed that the expression of vascular endothelial growth factor (VEGF) mRNA was decreased 46.84%±3.72% (mean ± SD). Enzyme-linked immunosorbent assay indicated that the expression of VEGF protein was down-regulated to 52.86%±1.10% (mean ± SD) in vitro. In vivo tumor growth assay GO-poly-l-lysine hydrobromide/folic acid (GPF)/DOX/siRNA exhibited gene silencing and tumor inhibition (66.95%±2.35%, mean ± SD) compared with naked siRNA (1.62%±1.47%, mean ± SD) and DOX (33.63%±5.85%, mean ± SD). GPF/DOX/siRNA exhibited no testable cytotoxicity.

Conclusion: The results indicated that co-delivery of siRNA and DOX by GPF could be a promising application in tumor clinical therapy.

Keywords: cervical carcinoma; co-delivery carrier; graphene oxide; siRNA delivery.

PubMed Disclaimer

Conflict of interest statement

Disclosure The authors report no conflicts of interest in this work.

Figures

Figure 1
Figure 1
Zeta potential measurements of GO (A), GO-PLL (B), GPF (C), and GPF/DOX/VEGF-siRNA (D). Abbreviations: DOX, doxorubicin; GO, graphene oxide; GPF, GO-PLL/folic acid; PLL, poly-l-lysine hydrobromide; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 2
Figure 2
FTIR spectra of GO, PLL, GO-PLL, and GPF. Abbreviations: FTIR, Fourier transform infrared; GO, graphene oxide; GPF, GO-PLL/folic acid; PLL, poly-l-lysine hydrobromide.
Figure 3
Figure 3
TEM images of GO (A), GPF (B), and GPF/DOX/VEGF-siRNA (C). Abbreviations: DOX, doxorubicin; GO, graphene oxide; GPF, GO-poly-l-lysine hydrobromide/folic acid; siRNA, small interfering RNA; TEM, transmission electron microscopy; VEGF, vascular endothelial growth factor.
Figure 4
Figure 4
The dispersibility and stability of GPF (A), GO (B), and the Tyndall effect of GO and GPF (C). Abbreviations: DMEM, Dulbecco’s Modified Eagle’s Medium; GO, graphene oxide; GPF, GO-poly-l-lysine hydrobromide/folic acid.
Figure 5
Figure 5
Agarose gel electrophoresis retardation assays of VEGF-siRNA complexed with GPF (A) and GO (B). Abbreviations: GO, graphene oxide; GPF, GO-poly-l-lysine hydrobromide/folic acid; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 6
Figure 6
Release profiles of DOX at pH 7.4 (physiological) and pH 5.0 (tumor mimicking) from GPF/DOX and GO/DOX in vitro (n=3). Abbreviations: DOX, doxorubicin; GO, graphene oxide; GPF, GO-poly-l-lysine hydrobromide/folic acid.
Figure 7
Figure 7
Degradation of GPF/VEGF-siRNA with heparin and anti-RNase A. Abbreviations: GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 8
Figure 8
Confocal images of the HeLa cells. (A) Blank control, (B) naked FAM-VEGF-siRNA, (C) DOX, (D) GPF/FAM-VEGF-siRNA, (E) GPF/DOX, (F) Lipo™2000/FAM-VEGF-siRNA, and (G) GPF/DOX/FAM-VEGF-siRNA. Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 9
Figure 9
Anti-proliferation effect of GPF/DOX/VEGF-siRNA on HeLa cells (n=3). Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; NC, normal control; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 10
Figure 10
VEGF mRNA expression of HeLa cells treated with different medicines. Data are presented as the mean ± SD, n=3. Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; NC, normal control; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 11
Figure 11
VEGF protein expression of HeLa cells treated with different medicines. Data are presented as the mean ± SD, n=3. Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; NC, normal control; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 12
Figure 12
Image of tumors of blank control, positive control, naked VEGF-siRNA, DOX, GPF/VEGF-siRNA, GPF/DOX, and GPF/DOX/VEGF-siRNA (n=10). Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 13
Figure 13
The tumor weights of blank control, naked VEGF-siRNA, DOX, GPF/VEGF-siRNA, GPF/DOX, DOX (positive control), and GPF/DOX/VEGF-siRNA (n=10). Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 14
Figure 14
The tumor volume of blank control, naked VEGF-siRNA, DOX, GPF/VEGF-siRNA, GPF/DOX, and GPF/DOX/VEGF-siRNA (n=10). Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; NS, normal saline; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Figure 15
Figure 15
The expression of VEGF protein in vivo (n=5). Abbreviations: DOX, doxorubicin; GPF, graphene oxide-poly-l-lysine hydrobromide/folic acid; NS, normal saline; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
Scheme 1
Scheme 1
Scheme for GPF and GPF/DOX/VEGF-siRNA preparation. (I) KOH, 70°C, 24 hours, DI water. (II) Room temperature, 24 hours. (III) Stirred at room temperature, without light, 12 hours, DI water. (IV) Incubation at 37°C, 30 minutes, DEPC water. Abbreviations: DEPC, diethyl pyrocarbonate; DI, deionized; DOX, doxorubicin; FA, folic acid; GO, graphene oxide; GPF, GO-PLL/FA; PLL, poly-l-lysine hydrobromide; siRNA, small interfering RNA; VEGF, vascular endothelial growth factor.
See this image and copyright information in PMC

References

    1. Liu Z, Chen K, Davis C, et al. Drug delivery with carbon nanotubes for in vivo cancer treatment. Cancer Res. 2008;68(16):6652–6660. - PMC - PubMed
    1. Liu CW, Lin WJ. Systemic co-delivery of doxorubicin and siRNA using nanoparticles conjugated with EGFR-specific targeting peptide to enhance chemotherapy in ovarian tumor bearing mice. J Nanopart Res. 2013;15(10):1–14.
    1. Huang HY, Kuo WT, Chou MJ, Huang YY. Co-delivery of anti-vascular endothelial growth factor siRNA and doxorubicin by multifunctional polymeric micelle for tumor growth suppression. J Biomed Mater Res A. 2011;97(3):330–338. - PubMed
    1. Li JH, Chia M, Shi W, et al. Tumor-targeted gene therapy for nasopha-ryngeal carcinoma. Cancer Res. 2002;62(1):171–178. - PubMed
    1. Takeda T, Inaba H, Yamazaki M, et al. Tumor-specific gene therapy for undifferentiated thyroid carcinoma utilizing the telomerase reverse tran-scriptase promoter. J Clin Endocrinol Metab. 2003;88(8):3531–3538. - PubMed

MeSH terms