Oral Cancer Theranostic Application of FeAu Bimetallic Nanoparticles Conjugated with MMP-1 Antibody

Abstract

Metastatic oral squamous cell carcinoma (SCC) displays a poor disease prognosis with a 5-year survival rate of 39%. Chemotherapy has emerged as the mainstream treatment against small clusters of cancer cells but poses more risks than benefits for metastatic cells due to the non-specificity and cytotoxicity. To overcome these obstacles, we conjugated antibodies specific for matrix metalloproteinase-1 (MMP-1), a prognostic biomarker of SCC, to iron-gold bimetallic nanoparticles (FeAu NPs) and explored the capability of this complex to target and limit SSC cell growth via magnetic field-induced hyperthermia. Our results showed that 4.32 ± 0.79 nm sized FeAu NPs were superparamagnetic in nature with a saturation magnetization (Ms) of 5.8 emu/g and elevated the media temperature to 45 °C, confirming the prospect to deliver hyperthermia. Furthermore, conjugation with MMP-1 antibodies resulted in a 3.07-fold higher uptake in HSC-3 (human tongue squamous cell carcinoma) cells as compared to L929 (fibroblast) cells, which translated to a 5-fold decrease in cell viability, confirming SCC targeting. Finally, upon magnetic stimulation, MMP-1-FeAu NPs conjugate triggered 89% HSC-3 cellular death, confirming the efficacy of antibody-conjugated nanoparticles in limiting SCC growth. The synergistic effect of biomarker-specific antibodies and magnetic nanoparticle-induced hyperthermia may open new doors towards SCC targeting for improved disease prognosis.

Keywords: cancer theranostic; iron–gold bimetallic nanoparticles; magnetic hyperthermia; matrix metalloproteinase-1; oral squamous cell carcinoma.

Figure 1

The characterization of FeAu and MMP-1 antibody-conjugated FeAu NPs. (a) TEM micrograph of FeAu NPs, (b) TEM micrograph of MMP-1 antibody-conjugated FeAu NPs, (c) size distribution of FeAu NPs, (d) size distribution of MMP-1 antibody-conjugated FeAu NPs, and (e) elemental composition of FeAu NPs analyzed using EDS analysis.

Figure 2

Confirmation of MMP-1 conjugation to FeAu nanoparticles. (a) X-ray diffraction pattern of FeAu NPs, (b) FTIR pattern of FeAu and FeAu-Cys, (c) Raman spectra before and after FeAu NPs modification, and (d) UV-Vis spectrum of FeAu NPs before and after MMP-1 antibody modification.

Figure 3

Confocal images of FeAu in different filters under 515 nm laser. (a) Null, (b) 600 ± 37 nm, (c) 530 ± 43 nm, (d) 440 ± 40 nm.

Figure 4

Magnetic properties of FeAu NPs as analyzed using SQUID. (a) ZFC/FC curve of FeAu NPs and MMP1-FeAu NPs (M-T curve), (b) hysteresis curves of FeAu NPs and MMP1-FeAu NPs (M–H curve) and (c) temperature elevation of the solution with the addition of different concentrations of FeAu NPs and stimulation with AMF over 10 min.

Figure 5

Cell responses of FeAu NPs and MMP-1 antibody-conjugated FeAu NPs. (a) Cellular viability of L929 and HSC-3 by the MTT assay (n = 3); (b) bio-TEM micrographs of human oral squamous carcinoma incubated with FeAu and MMP1-FeAu over different time periods; (c) quantification of cellular uptake of FeAu NPs and MMP1-FeAu NPs in HSC-3 cells and L929 cells analyzed by ICP-AES, (d) FeAu NPs and MMP1-FeAu NPs stimulated magnetically.

Figure 5

Cell responses of FeAu NPs and MMP-1 antibody-conjugated FeAu NPs. (a) Cellular viability of L929 and HSC-3 by the MTT assay (n = 3); (b) bio-TEM micrographs of human oral squamous carcinoma incubated with FeAu and MMP1-FeAu over different time periods; (c) quantification of cellular uptake of FeAu NPs and MMP1-FeAu NPs in HSC-3 cells and L929 cells analyzed by ICP-AES, (d) FeAu NPs and MMP1-FeAu NPs stimulated magnetically.

Figure 6

The in vivo anti-cancer effects magnetic heat after 30 days. (a) The tumor volume % of original; (b) the tumor volume % of PBS (control groups). Statistical analysis was performed using two-tailed student’s t-test and the level of significance was set at 0.05. * represents p-value ≤ 0.05 and ** represents p-value ≤ 0.01.