Polyethylene Glycol-Coated Graphene Oxide Loaded with Erlotinib as an Effective Therapeutic Agent for Treating Nasopharyngeal Cancer Cells

Abstract

Introduction: Nasopharyngeal carcinoma (NPC) is a common cancer in southern China and Taiwan, and radiation therapy combined with or without chemotherapy is its mainstay treatment. Although it is highly sensitive to radiotherapy, local recurrence and distant metastasis remain difficult unsolved problems. In recent years, graphene oxide (GO) has been found to be a promising novel anticancer drug carrier. Here, we present our designed functionalized GO, polyethylene glycol-coated GO (GO-PEG), as a drug carrier, which was loaded with erlotinib and showed promising anticancer effects on NPC cells.

Methods: The effects of GO-PEG-erlotinib on the proliferation, migration, and invasion of NPC cells were investigated by WST-8 assay, wound healing assay, and invasion assay, respectively. RNA sequencing was conducted and analyzed to determine the molecular mechanisms by which GO-PEG-erlotinib affects NPC cells.

Results: Our results showed that GO-PEG-erlotinib reduced NPC cell viability in a dose-dependent manner and also inhibited the migration and invasion of NPC cells. The RNA sequencing revealed several related molecular mechanisms.

Conclusion: GO-PEG-erlotinib effectively suppressed NPC cell proliferation, migration, and invasion, likely by several mechanisms. GO-PEG-erlotinib may be a potential therapeutic agent for treating NPC in the future.

Keywords: anti-cancer; drug carrier; erlotinib; graphene oxide; nasopharyngeal carcinoma.

Figure 1

Characterizations of GO and GO-PEG. (A) TEM images of GO and GO-PEG. (B) The table lists the measurements of size and zeta potential of GO and GO-PEG analyzed by DLS. (C) The intensity of D and G band of GO and GO-PEG in Raman spectra. (D) The infrared spectrum of GO and GO-PEG by FTIR shows that PEG was successfully conjugated on GO. (E) Raman spectra of GO and GO-PEG. (F) Absorbance of GO, GO-PEG, erlotinib, and GO-PEG-erlotinib analyzed by UV-Vis spectrophotometer.

Figure 2

(A) The drug loading efficiency (LE) and encapsulation efficiency (EE) of erlotinib-loaded GO-PEG. (B) The drug release test of GO-PEG-erlotinib in pH 7.4 and 5.5.

Figure 3

Confocal microscopy images of NPC TW01 cells after treatment with FITC-labeled GO-PEG for 6 hours.

Figure 4

Cell viability of NPC TW01 after treatment with erlotinib and GO-PEG-erlotinib at various concentrations. Cell viability was determined after incubating with erlotinib or GO-PEG-erlotinib for 72 hours.

Figure 5

GO-PEG-erlotinib reduces cell migration in NPC TW01 cells. More cells migrated to the denuded area of the wound in the control group (left) compared to the cells treated with 0.7 µg/mL GO-PEG-erlotinib (middle) and 2.15 µg/mL GO-PEG-erlotinib (right) at 20 hours after the creation of the wound. *p < 0.05 compared with the control group by ANOVA.

Figure 6

GO-PEG-erlotinib inhibits cell invasion in NPC TW01 cells. Matrigel invasion assays of NPC TW01 cells showed that the invasion ability of NPC cells was reduced after treatment with 2.15 μg/mL GO-PEG-erlotinib, 2.15 μg/mL erlotinib, and 6.45 μg/mL GO-PEG for 48 hours. *p < 0.05 compared with the control group by ANOVA.

Figure 7

The top-ranked network identified by IPA analysis in GO-PEG-erlotinib study. The top-ranked network, which includes 34 genes, is related to cardiovascular disease, hematological disease, and hereditary disorder. The genes shaded in red are upregulated, and genes shaded in green are downregulated. All shaded genes are statistically significant, as indicated by the statistical analysis. A dotted line indicates an indirect interaction between the two gene products, and a solid line represents a direct interaction.

Figure 8

KEGG pathway analysis of GO-PEG-erlotinib on NPC cells. There are seven branches for KEGG pathways: cellular processes, environmental information processing, genetic information processing, human disease, metabolism, and organismal systems.