Hyperthermia treatment of tumors by mesenchymal stem cell-delivered superparamagnetic iron oxide nanoparticles

Abstract

Magnetic hyperthermia - a potential cancer treatment in which superparamagnetic iron oxide nanoparticles (SPIONs) are made to resonantly respond to an alternating magnetic field (AMF) and thereby produce heat - is of significant current interest. We have previously shown that mesenchymal stem cells (MSCs) can be labeled with SPIONs with no effect on cell proliferation or survival and that within an hour of systemic administration, they migrate to and integrate into tumors in vivo. Here, we report on some longer term (up to 3 weeks) post-integration characteristics of magnetically labeled human MSCs in an immunocompromized mouse model. We initially assessed how the size and coating of SPIONs dictated the loading capacity and cellular heating of MSCs. Ferucarbotran(®) was the best of those tested, having the best like-for-like heating capability and being the only one to retain that capability after cell internalization. A mouse model was created by subcutaneous flank injection of a combination of 0.5 million Ferucarbotran-loaded MSCs and 1.0 million OVCAR-3 ovarian tumor cells. After 2 weeks, the tumors reached ~100 µL in volume and then entered a rapid growth phase over the third week to reach ~300 µL. In the control mice that received no AMF treatment, magnetic resonance imaging (MRI) data showed that the labeled MSCs were both incorporated into and retained within the tumors over the entire 3-week period. In the AMF-treated mice, heat increases of ~4°C were observed during the first application, after which MRI indicated a loss of negative contrast, suggesting that the MSCs had died and been cleared from the tumor. This post-AMF removal of cells was confirmed by histological examination and also by a reduced level of subsequent magnetic heating effect. Despite this evidence for an AMF-elicited response in the SPION-loaded MSCs, and in contrast to previous reports on tumor remission in immunocompetent mouse models, in this case, no significant differences were measured regarding the overall tumor size or growth characteristics. We discuss the implications of these results on the clinical delivery of hyperthermia therapy to tumors and on the possibility that a preferred therapeutic route may involve AMF as an adjuvant to an autologous immune response.

Keywords: MRI; SPIONs; hyperthermia; mesenchymal stem cells; tumor therapy.

Figure 1

SPION uptake in MSCs and their corresponding magnetic heating characteristics. Notes: (A) SQUID measurements of the concentration of iron oxide per mesenchymal stem cell (MSC) after overnight incubation of 0.5 mg/mL of FluidMAG-CT (citric acid – 50 nm), FluidMAG-CMX (carboxylmethyldextran), FluidMAG-DX (dextran), FluidMAG-D (starch), and Ferucarbotran (carboxydextran). (B) MSCs take up SPIONs without affecting their phenotype; (i) MSCs in culture; (ii) Perl’s Prussian blue staining of MSCs after overnight culture with Ferucarbotran nanoparticles; (iii) differentiation to osteoblasts, Alizarin Red S staining; (iv) differentiation to adipocytes, Oil Red O staining. (C) Representative fiber-optic thermometry measurements of Ferucarbotran and FluidMAG-CT (50 nm- and 100 nm-sized SPIO particles) at 1 mg/mL. (D) Representative fiber-optic thermometry measurements of cell heating of a 5×10⁵ MSC pellet after overnight incubation of 0.5 mg/mL of FluidMAG-CT 50 nm or Ferucarbotran (representative of three experiments). Abbreviations: SQUID, superconducting quantum interference device; SPION, superparamagnetic iron oxide nanoparticle; sec, seconds.

Figure 2

Murine model of OVCAR-3 tumor growth, with and without magnetic heating. Notes: (A) Experimental plan of MRI and AMF heating. (B) T2 weighted MR images of a heated and nonheated OVCAR-3 tumor coinjected with Ferucarbotran-labeled MSCs, at days 14 (prior to heating), 18, and 22 (postheating) (white arrow indicates signal recovery). (C) Thermocamera image of a heated tumor (T) on the flank of a nude mouse (F) with C indicating the MACH coil and P the fiber-optic thermometry probe. (D) Fiber-optic thermometry measurements of a heated tumor over days 15, 17, 19, and 21 and a representative nonheated tumor and core body temperature. (E) Post-innoculation OVCAR-3 tumor growth for both heated and nonheated tumors. Abbreviations: MRI, magnetic resonance imaging; AMF, alternating magnetic field; MR, magnetic resonance; MACH, magnetic alternating current hyperthermia; sec, seconds; MSCs, mesenchymal stem cells.

Figure 3

Immunohistochemistry data from a human OVCAR-3 tumor model, with and without magnetic heating. Notes: (A) Sections from a representative tumor stained with DiI cell tracker dye, Perl’s Prussian blue iron stain, and H&E on day 22, derived from a coinjection, on day 1, of Ferucarbotran-labeled MSCs and OVCAR-3 tumor cells that were subsequently either heated or nonheated. (B) Percentage stained areas of representative slices throughout both heated and nonheated tumors for both Perl’s and DiI staining (error bars are SEM and **P<0.01). Abbreviations: H&E, hematoxylin and eosin; MSCs, mesenchymal stem cells; SEM, standard error of the mean.