Figures & data
Table I. Summary of post-treatment tissue damage immediately beneath the tumour. Tissue was evaluated for the presence of oedema, muscle necrosis and haemorrhage. Zones were 0.75 mm thick. Zone 1 began at the deep edge of the tumour. If no damage was observed, ‘normal tissue’ was reported.
Figure 1. TEM image of mNP associated with murine mammary adenocarcinoma (MTGB) tumour cells. This section was acquired 5 min following delivery of 7.5 mg Fe/cm3 directly into the MTGB tumour mass. mNP (black specs) are located between cells (interstitial space), attached to the exterior aspect of the cell plasma membrane or in membrane associated vesicles. Scale bar = 2 μm. Qualitative evaluation of tumours (n = 4), harvested at the prescribed treatment time, indicate that the mNP distribution at the initiation of AMF is dominated by extracellular mNP.
Figure 2. This figure demonstrates the techniques used to deliver the alternating magnetic field (AMF) to a tumour bearing mouse. The AMF field was generated by a water cooled, whole body circular coil which produced an AMF field of 165 KHz and 450 Oe (35.8 kA/m). The location of the flank-based tumour within the coil is indicated by the red encircled dot. This region demonstrates a near uniform, homogenous AMF. The tumour temperatures and core temperature of the mouse were measured throughout the treatment using real time fibre-optic thermometry. The thermal history of the treatment was continuously calculated and displayed for the tumour and body core throughout the experimental period.
Figure 3. The microwave applicator used for tumour treatments consist of an open-ended pair of coaxial conductors, which was driven by a 915 MHz microwave generator, cooled by circulating water and sized to fit over the flank tumour of the mice. Tissue-applicator coupling gel was placed between the applicator and the tumour surface. Tumour temperatures and core temperature of the mice were measured throughout the treatment, with thermal history (CEM) calculated in real-time.
Figure 4. This graph demonstrates the tumour heating characteristics for the mNPH and microwave treatment modalities. All tumours received a thermal dose equal to 60 min at 43 °C (CEM60). Minor tumour geometry and/or mNP biodistribution variations resulted in slightly different heating rates (relationship of heating time and temperature). These differences did not meaningfully affect treatment efficacy.
Figure 5. This figure represents the histological parameters used to assess a treated mouse tumour and associated normal tissue deep to the tumour. Zones 1, 2 and 3 were used to determine and quantify the morphological tissue response at various depths beneath the tumour following treatment. Each zone had a thickness of 0.75 mm. Zone 1 started at the deep edge of the tumour. A quantitative cross-sectional area method was used to determine the relative amount of viable versus non-viable tumour area. The presence of normal tissue/muscle necrosis, peritumoural oedema and/or haemorrhage was categorised for each of the three zones for each tumour.
Figure 6. Kaplan-Meyer curve demonstrating survival/tumour regrowth for each of the four treatment groups (no treatment, AMF, mNPH and 915 MHz microwave hyperthermia). Tumours were treated at 150 mm3 ± 40 mm3. Animals were removed from the study when tumour volumes reached a three-fold treatment volume increase. Treatment with mNP hyperthermia resulted in a slightly more effective treatment than 915 MHz microwave hyperthermia (p = 0.09, 5 days).
Figure 7. These photomicrographs acquired 24 h post mNPH treatment, demonstrate the morphological changes observed in two different tumours which received an identical thermal dose (CEM60). Both tumours show extensive uniform necrosis. The lower tumour demonstrates significant oedema in the space between the tumour and underlying muscle (zones 1 and 2). Although minor inflammation was seen, including mNP-containing macrophages, there was no haemorrhage or necrosis in the overlying skin or peritumoral tissue.
Figure 8. Photomicrographs of two mouse mammary tumours 24 h post 915 microwave treatment. These photomicrographs show similar effects consisting largely of generalised central tumour necrosis. The superficial tumour is spared, likely due to applicator cooling. The high magnification (10×) photomicrographs of muscle tissue deep to the tumour demonstrate varying degrees of muscle necrosis, haemorrhage and inflammation. The majority of the changes are seen in zones 1 and 2. See Figure 5 for information on tissue damage quantification techniques.
Figure 9. A representative MTGB tumour with histological sections taken 5 min after mNP injection. These H&E and Prussian blue photomicrographs demonstrate regional heterogeneity in mNP distribution. The 110 nm diameter mNP were injected in four tissue quadrants, with a total of 7.5 mg of Fe per cm3 tumour, 5 min prior to tumour removal and processing. Regions indicated on the low magnification images are shown in the high magnification images. H&E and Prussian blue. (A, B) 10 × magnification, (C, D, E, and F) 100 × magnification.
Figure 10. This graph demonstrates a comparison of necrotic tumour area 24 h following mNPH and microwave treatment (CEM60). Histological evaluation of tumours treated with 915 MHz microwave hyperthermia show 75% of the tumour area is necrotic in comparison to mNPH (79% necrotic). Statistical significance between groups was not found (p = 0.55). Error bars represent standard deviation. All tumours were evaluated 24 h following treatment.
