Iron Oxide Nanoparticles as Carriers for DOX and Magnetic Hyperthermia after Intratumoral Application into Breast Cancer in Mice: Impact and Future Perspectives

Figure 1. Characterization of the heating potential of the MF66-DOX (A) and the bi-functionalized MF66-DOX-N6L MNP formulations (B). MNPs were suspended in water or immobilized in 10% polyvinyl alcohol (PVA) as well as in BT474 cells. Mean and standard deviation of the mean of n = 3.

Figure 2. DOX-functionalized MNPs and magnetic hyperthermia significantly reduces tumor volumes in vivo. Tumor volumes (volume percentage to day 0 before treatment), mean ± standard deviation (n ≥ 4 mice per group), (*) p ≤ 0.05, (**) p ≤ 0.01 and (***) p ≤ 0.001.

Figure 3. MNPs functionalized with DOX exert a good blood biocompatibility. White blood cells (A), red blood cells (B) and hemoglobin concentration (C). Mean ± standard deviation (n ≥ 4 mice per group). Dashed lines refer to reference values (Harlan Laboratories, Venray, The Netherlands; http://www.harlan.com).

Figure 4. The intratumorally injected MNPs remain within the tumor after hyperthermia treatment. Iron concentration in selected organs 28 days after magnetic hyperthermia treatment. Mean and standard deviation of n ≥ 4 animals per group.

Figure 5. The additional ligand N6L on the surface of DOX-functionalized MNPs increases their internalization into BT474 cells and further inhibits tumor cell growth in combination with magnetic hyperthermia in vitro. (A) Intracellular iron content measured in the cells at 24 h after incubation with MNP formulations. Mean ± standard deviation of n ≥ 6 samples/group, p ≤ 0.01 (**) and p ≤ 0.001 (***). (B) The relative dehydrogenase activity was measured 24 h or 48 h after incubation with MNP formulations and the alternating magnetic field (AMF) treatment, (C) MNP application without AMF and (D) treatment with the free ligands. AMF settings: H = 23.9 kA/m, f = 410 kHz, 43 °C for 60 min. Means and standard deviation of three individual experiments with three parallels each, (*) p ≤ 0.05 and (***) p ≤ 0.001.

Figure 6. Magnetic hyperthermia in combination with ((DOX and/or N6L) MNP formulations induced morphological changes and detachment of BT474 cells. Representative microscopic images of BT474 cells, which were incubated with the different MF66 formulations (100 µg Fe/mL, 24 h); 24 h or 48 h post magnetic hyperthermia (+MNPs/+AMF), MNPs control (+MNPs/−AMF), free agent controls (DOX, N6L or both) as well as untreated controls (native BT474). Scale bar = 100 µm.