Hybrid Radiobioconjugated Superparamagnetic Iron Oxide-Based Nanoparticles for Multimodal Cancer Therapy

Abstract

Superparamagnetic iron oxide nanoparticles (SPIONs) are widely used for biomedical applications for their outstanding properties such as facile functionalization and doping with different metals, high surface-to-volume ratio, superparamagnetism, and biocompatibility. This study was designed to synthesize and investigate multifunctional nanoparticle conjugate to act as both a magnetic agent, anticancer immunological drug, and radiopharmaceutic for anticancer therapy. The carrier, ¹⁶⁶Ho doped iron oxide, was coated with an Au layer, creating core-shell nanoparticles ([¹⁶⁶Ho] Fe₃O₄@Au. These nanoparticles were subsequently modified with monoclonal antibody trastuzumab (Tmab) to target HER2+ receptors. We describe the radiobioconjugate preparation involving doping of a radioactive agent and attachment of the organic linker and drug to the SPIONs' surface. The size of the SPIONs coated with an Au shell measured by transmission electron microscopy was about 15 nm. The bioconjugation of trastuzumab onto SPIONs was confirmed by thermogravimetric analysis, and the amount of two molecules per one nanoparticle was estimated with the use of radioiodinated [¹³¹I]Tmab. The synthesized bioconjugates showed that they are efficient heat mediators and also exhibit a cytotoxic effect toward SKOV-3 ovarian cancer cells expressing HER2 receptors. Prepared radiobioconjugates reveal the high potential for in vivo application of the proposed multimodal hybrid system, combined with magnetic hyperthermia and immunotherapy against cancer tissues.

Keywords: SPION; anticancer therapy; drug delivery; magnetic hyperthermia; multimodal therapy; radio-labeled nanoparticles; superparamagnetic nanoparticles; trastuzumab.

Figure 1

Scheme of precipitation synthesis and coating of [¹⁶⁶Ho]Fe₃O₄.

Figure 2

Schematic image of [¹⁶⁶Ho]Fe₃O₄ coating with Au.

Figure 3

Scheme of [¹⁶⁶Ho]Fe₃O₄ coating with Au.

Figure 4

Transmission Electron Microscopy images of NPs: (a) Citrate stabilized [Ho]Fe₃O₄@CA and [Ho]Fe₃O₄@Au; (b) [Ho]Fe₃O₄@Au coated with PEG linker, before the attachment of Tmab; (c) [Ho]Fe₃O₄@Au-Tmab radiobioconjugates (PEG linker presence is not mentioned for clarity); and (d) [Ho]Fe₃O₄@Au-Tmab bioconjugates after magnetic hyperthermia treatment. Red arrows (b,c) point to the PEG or PEG-Tmab “misty” corona surrounding the [Ho]Fe₃O₄@Au core.

Figure 5

UV-vis spectra for [¹⁶⁶Ho]Fe₃O₄ and [¹⁶⁶Ho]Fe₃O₄@Au.

Figure 6

Thermograms of [Ho]Fe₃O₄, [Ho]Fe₃O₄@CA, [Ho]Fe₃O₄@Au, and [Ho]Fe₃O₄@Au-Tmab.

Figure 7

Heating of radio-labeled [Ho]Fe₃O₄@Au-Tmab bioconjugate in various ranges of frequency of magnetic field: (a) 100 G, (b) 150 G, (c) 75–150 G, and (d) dependence of SAR for various frequencies of the magnetic field as a function of the amplitude of the magnetic field. S.D. bar represents the largest value of five independent experiments.

Figure 8

Stability studies of: (a) [¹⁶⁶Ho]Fe₃O₄@Au, (b) [¹⁶⁶Ho]Fe₃O₄@Au-Tmab in water DI, 0.9% NaCl and human serum. Data are expressed as mean ± SD (n = 3).

Figure 9

Cell viability after treatment with different concentrations of: (a) [Ho]Fe₃O₄@Au, (b) [Ho]Fe₃O₄@Au-PEG, and (c) [Ho]Fe₃O₄@Au-Tmab. SKOV-3 were incubated for 24, 48, and 72 h, after which their viability was determined by MTS assay. The results are expressed as a percentage of control cells. Data are expressed as the mean ± SD (n = 3). Statistical significance was considered if p < 0.05 (*), p < 0.01 (**), p < 0.001 (***), p < 0.0001 (****), p < 0.00001 (*****).

Figure 10

Specificity of binding on SKOV-3 (HER2+) and MDA-MB-231 (HER2−). [Ho]Fe₃O₄@Au-[¹³¹I]Tmab was incubated in the presence and absence of 100x molar fold of free trastuzumab. Data are expressed as a mean ± SD (n = 3). Statistical significance was considered if p < 0.05 (*).