Use of the vasodilator sodium nitroprusside to increase tumour temperature during loco-regional hyperthermia with a capacitive ring applicator in a rat tumour model.

The influence of sodium nitroprusside (SNP) induced hypotension (to a mean arterial pressure of 60 mmHg) on tumour and normal tissue temperature during hyperthermia (HT) was examined. Loco-regional HT was given to the calf of BD IX rats by external radiofrequency heating from a capacitive ring applicator. In experiments in rats with subcutaneous BT(4)An tumours, the mean tumour temperature increased by 0.49 degrees C from 42.36 to 42.85 degrees C, on average, during SNP-hypotension. This represented 58% of the increase in tumour temperature found in the same rats when the tumour circulation was stopped completely by sacrificing the rats. SNP-hypotension resulted in a decrease in mean muscle temperature from 41.73 to 41.23 degrees C. The temperature difference between the tumour and the underlying muscle thereby increased by approximately 1 degrees C, indicating that SNP can increase tumour temperature during HT without increasing the risk of heat-related damage to skeletal muscle. Experiments in rats without tumours were also done to further examine the effect of SNP-hypotension on muscle temperature under different treatment conditions (variation of radiofrequency energy deposition and water bolus temperature). It was found that SNP decreased the muscle temperature during HT in two experiments where the average muscle temperature was 42.1 and 42.6 degrees C, respectively. In an experiment where the muscle temperature was 43.0 degrees C, on average, before SNP infusion, the muscle temperature increased during SNP-hypotension. This finding indicates that SNP-hypotension during HT may increase the risk of skeletal muscle necrosis with muscle temperatures at this level.     

Development of an inductive, non-invasive RF applicator for studying hyperthermia in a rat brain tumour model

A non-invasive inductive RF applicator has been designed specially for local heating of 6 mm diameter tumours growing in the brains of F344 rats (144-148 MHz). It is shown that an inductive applicator is the choice for this particular case. The power deposition pattern is analysed by modelling the applicator with a three-dimensional array of magnetic dipoles. The total hyperthermia system is briefly described and results of phantom and in vivo experimental measurements are presented. These confirm the theoretical analysis.     

Development of an inductive,non-invasive RF applicator for studying hyperthermia in a rat brain tumour model

A non-invasive inductive RF applicator has been designed specially for local heating of 6 mm diameter tumours growing in the brains of F344 rats (144–148 MHz). It is shown that an inductive applicator is the choice for this particular case. The power deposition pattern is analysed by modelling the applicator with a three-dimensional array of magnetic dipoles. The total hyperthermia system is briefly described and results of phantom and in vivo experimental measurements are presented. These confirm the theoretical analysis.     

RF-power and temperature data analysis of 444 patients with primary cervical cancer: Deep hyperthermia using the Sigma-60 applicator is reproducible

Treatment reproducibility is important to guarantee reproducible treatment-outcome, a low-complication rate and efficient treatment procedures. This study evaluated the performance of loco-regional deep hyperthermia with four BSD-2000 configurations during 1990–2005 using the direct available parameters, i.e., temperature and power. Primary cervical cancer patients (n = 444) were all treated within the Sigma-60. Patients were grouped in three weight-groups: <61 kg, 61–70 kg, >70 kg. Different temperature and power indices were extensively analyzed per BSD configuration, per weight-group, and over the time-period. No substantial variations were found for temperature/power indices over the four BSD configurations or for the temperature doses in similar weight-groups. The ‘bare’ power indices were increased with weight; however, the derivative power-related (W/kg, W/cm²) and temperature indices decreased. Large variations were found in the power-related parameters during 1991–1996 (1st time-period), whereas they were much smaller during 1997–2005 (2nd time-period). The most relevant change noted was the adaptation of the treatment strategy with respect to power modulation. The average frequency of switched-off was 3.4 and 8.9 times/treatment session for the 1st and 2nd time-period, respectively, while the average duration of each switched-off time was 78.2 vs. 38.3 s. The yearly average of vagina T₅₀ was in the range of 39.3–40.2°C (1st time-period) and 40.0–40.5°C (2nd time-period). In 40% of the patients, a positive correlation was found between normalized net integrated power per pelvic area and vagina T₅₀. Good reproducible heating is achieved with the BSD-2000 Sigma-60 irrespective of the regular technological upgrades of the system and variation of trained staff-members.     

Stereologic evaluation of the vasculature in a MX1 xenotransplanted tumour model after combinations of treatment with ifosfamide,hyperthermia and irradiation

The vascularization of tumours is a critical parameter of their growing and metastatic behaviour. However, little is known about the morphologic reactions of the microvasculature, especially the capillary bed of tumours and the adjacent tissue. In this study, the vessels in MX1 xenotransplants in athymic nu/nu nude mice were quantified and the angioarchitecture was visualized with the aim of presenting stereologic parameters of vessels based on a morphometric analysis of post mortem tissue blocks which were processed by standard histological procedures. In order to study changes of the microvasculature of MX1 tumours, the xenotransplanted nude mice were treated by different therapeutic regimens. Standardized hyperthermia, ifosfamide and irradiation therapies were applied. Special interest was focused on early changes of capillaries and of the pre- as well as post-terminal vascular bed. The stereologic evaluation of capillaries and larger vessels immediately after the therapy with ifosfamide and hyperthermia shows an increase of the mean capillary sizes. Furthermore, tumour samples after the 5th day of irradiation (5.2 Gy) and combinations of irradiation and chemotherapy treatment have been investigated. After 5 days of irradiation, a significant decrease of the vascular density was found. The results presented here clearly show that the timing and the mode of therapy influence the capillary morphology and periterminal vasculature of xenotransplanted MX1 tumours.     

An edge-element based finite element model of microwave heating in hyperthermia: application to a bolus design

Heating of superficial tumours with microwave waveguide applicators has been shown in phase III trials to significantly improve the local control of small lesions when combined with radiation therapy. This success has not yet translated to the treatment of larger tumours, due to difficulty in adequately heating the entire tumour region. Several modifications to the water bolus used with external waveguide applicators have been made in the past in order to increase the heating area. One such modification consisted of a large, microwave-absorbing patch placed inside the bolus, which flattens out the beam profile produced by the applicator. Using this bolus instead of a conventional one resulted in a 30% increase in the effective heating volume produced by the BSD MA120 applicator. This paper describes an optimization procedure for this bolus design which utilises a new finite element model of microwave heating described in an accompanying paper. The optimization procedure resulted in a further 28% increase in the effective heating volume.     

Increasing the systemic temperature during regional hyperthermia: effect of a cooling strategy on tumour temperatures and side-effects

In the application of regional hyperthermia, optimization of the temperature distribution remains necessary. One of the tools that might be used is a modest increase in the systemic temperature to diminish cooling by blood perfusion. This study investigates (1) if it is feasible to increase the systemic temperature by applying other cooling strategies, without inducing unacceptable systemic stress, and (2) whether a rise in systemic temperature results in improvement of tumour temperatures. Eleven patients with locally advanced cervical carcinoma and 12 patients with locally advanced prostate carcinoma were treated with our Coaxial TEM regional hyperthermia system. In this system, the temperature of the open water bolus can be easily adjusted. Two cooling methods were applied alternately, one with a relatively low water temperature (method A), the other with a higher water bolus temperature in combination with extensive head/chest cooling by a hand shower (method B). Method B resulted in significantly higher systemic temperatures, for both patient groups separately (0.8, respectively, 0.5°C) and for the total patient group (0.7°C). Additionally, all tumour index temperatures were higher. For the combined group (for T₅₀: 0.4°C) and for the cervix group (for T₅₀: 0.7°C), it reached statistical significance. The raise in core temperature led to a significantly higher increase in heart rate. For the group of cervix patients, higher systemic temperatures resulted in more treatment-limiting systemic stress. For the prostate patients, systemic stress was not an important issue. Since the raise in systemic temperature did not influence the overall tolerance of treatment, method B could be applied to this group. However, the increases in tumour temperatures were small, and potential hazards of systemic temperature increase should be considered.     

Use of the 90Sr applicator for intraoperative radiation therapy in a mouse tumor model

Intraoperative radiation therapy (IORT) research is limited by the lack of small-animal models. We have implanted B16 melanoma into mouse kidneys, which we subsequently operated upon and irradiated with beta rays from a ⁹⁰Sr ophthalmic applicator. The IORT has effectively prolonged survival and produced some cures. The strategy should be applicable to other murine tumors and to other internal implantation sites.     

Phosphorus NMR study of the response of a murine tumour to hyperthermia as a function of treatment time and temperature

Evaluating the response of tumours to therapy promises to become one of the major applications of in vivo phosphorus (³¹P) nuclear magnetic resonance spectroscopy. Decreases in the levels of organic phosphates in favour of inorganic phosphate (Pi) as occur in murine tumours after hyperthermia treatment, can be quantified by the ratio ATP/Pi. In this study the relationship between the time of heating (15, 30 and 60 min) and the temperature (43 and 44°C) was investigated in mice with NU-82 tumours by considering the changes in ATP/Pi ratio as a function of both variables. After 30 min treatment at 43 °C the percentage decrease in ATP/Pi ratio was similar to that observed after 15 min at 44°C (42 ±9 vs. 48 ±9); after 60 min at 43°C the decrease was similar to that after 30 min at 44°C (75 ±7 vs. 74±4). These results give further evidence for the validity of a current working definition of thermal dose: thermal dose= ± 2^T-43 dt. In addition this study shows that in vivo ³¹P NMR spectroscopy can be a useful means for assessment of thermal dose.     

Radiofrequency ablation combined with liposomal quercetin to increase tumour destruction by modulation of heat shock protein production in a small animal model

Purpose: To investigate the effect of heat shock protein (HSP) modulation on tumour coagulation by combining radiofrequency (RF) ablation with adjuvant liposomal quercetin and/or doxorubicin in a rat tumour model.

Methods: Sixty R3230 breast adenocarcinoma tumours/animals were used in this IACUC-approved study. Initially, 60 tumours (n?=?6, each subgroup) were randomised into five groups: (1) RF alone, (2) intravenous (IV) liposomal quercetin alone (1?mg/kg), (3) IV liposomal quercetin followed 24?h later with RF, (4) RF followed 15?min later by IV liposomal doxorubicin (8?mg/kg), (5) IV liposomal quercetin 24?h before RF followed by IV liposomal doxorubicin 15?min post-ablation. Animals were sacrificed 4 or 24?h post-treatment and gross coagulation diameters were compared. Next, immunohistochemistry staining was performed for Hsp70 and cleaved caspase-3 expression. Comparisons were performed by using Student t-tests or ANOVA.

Results: Combination RF-quercetin significantly increased coagulation size compared with either RF or liposomal quercetin alone (13.1?±?0.7?mm vs. 8.8?±?1.2?mm or 2.3?±?1.3?mm, respectively, P?<?0.001 for all comparisons). Triple therapy (quercetin-RF-doxorubicin) showed larger coagulation diameter (14.5?±?1.0?mm) at 24?h than quercetin-RF (P?=?0.016) or RF-doxorubicin (13.2?±?1.3?mm, P?=?0.042). Combination quercetin-RF decreased Hsp70 expression compared with RF alone at both 4?h (percentage of stained cells/hpf 22.4?±?13.9% vs. 38.8?±?16.1%, P?<?0.03) and 24?h (45.2?±?10.5% vs. 81.1?±?3.6%, P?<?0.001). Quercetin-RF increased cleaved caspase-3 expression at both 4?h (percentage of stained cells/hpf 50.7?±?13.4% vs. 41.9?±?15.1%, P?<?0.03) and 24?h (37.4?±?7.8% vs. 33.2?±?6.5%, P?=?0.045); with, triple therapy (quercetin-RF-doxorubicin) resulting in the highest levels of apoptosis (45.1?±?10.7%) at 24?h. Similar trends were observed for rim thickness.

Conclusions: Suppression of HSP production using adjuvant liposomal quercetin can increase apoptosis and improve RF ablation-induced tumour destruction. Further increases in tumour coagulation can be seen including an additional anti-tumour adjuvant agent such as liposomal doxorubicin.     

Delivery of temozolomide to the tumor bed via biodegradable gel matrices in a novel model of intracranial glioma with resection

Introduction We have completed in vivo safety and efficacy studies of the use of a novel drug delivery system, a gel matrix-temozolomide formulation that is injected intracranially into the post-resection cavity, as a candidate for glioma therapy. Methods A rat intracranial resection model of C6-GFP intracranial glioma was used for safety and toxicity studies. Biodistribution studies were performed using gel matrix-gallocyanine formulations and were evaluated at various time intervals using real-time analysis of dye distribution. Additionally, the resection model was used to determine the efficacy of gel matrix-temozolomide as compared to blank gel matrix. A subcutaneous human xenograft glioma model was used to further assess the efficacy of gel matrix-temozolomide in reducing the overall tumor load. Results Gel matrix-temozolomide exhibited minimal cytotoxicity toward normal brain tissue while displaying high levels of oncolytic activity toward glioma cells. In the intracranial glioma resection and subcutaneous glioma model, administration of gel matrix-temozolomide directly to the tumor bed was well tolerated and effective at reducing the tumor load. A significant reduction of tumor load was observed (P < 0.0001) in the 30% temozolomide group (~95%) as compared to blank control. There was little morbidity and no mortality associated with gel matrix treatment. Conclusions Gel matrix-temozolomide appears to be safe and effective when used in vivo to treat intracranial glioma and warrants further development as a potential adjuvant therapy.     

Stereologic evaluation of the vasculature in a MX1 xenotransplanted tumour model after combinations of treatment with ifosfamide, hyperthermia and irradiation.

The vascularization of tumours is a critical parameter of their growing and metastatic behaviour. However, little is known about the morphologic reactions of the microvasculature, especially the capillary bed of tumours and the adjacent tissue. In this study, the vessels in MX1 xenotransplants in athymic nu/nu nude mice were quantified and the angioarchitecture was visualized with the aim of presenting stereologic parameters of vessels based on a morphometric analysis of post mortem tissue blocks which were processed by standard histological procedures. In order to study changes of the microvasculature of MX1 tumours, the xenotransplanted nude mice were treated by different therapeutic regimens. Standardized hyperthermia, ifosfamide and irradiation therapies were applied. Special interest was focused on early changes of capillaries and of the pre- as well as post-terminal vascular bed. The stereologic evaluation of capillaries and larger vessels immediately after the therapy with ifosfamide and hyperthermia shows an increase of the mean capillary sizes. Furthermore, tumour samples after the 5th day of irradiation (5 x 2 Gy) and combinations of irradiation and chemotherapy treatment have been investigated. After 5 days of irradiation, a significant decrease of the vascular density was found. The results presented here clearly show that the timing and the mode of therapy influence the capillary morphology and periterminal vasculature of xenotransplanted MX1 tumours.     

Comparison of intra-luminal versus intra-tumoural temperature measurements in patients with locally advanced prostate cancer treated with the coaxial TEM system: report of a feasibility study

A study was performed on regional hyperthermia for patients with locally advanced prostate carcinoma. The primary objective was to analyse the thermometry data with an emphasis on the possibility of replacing invasive thermometry by tumour-related intra-luminal thermometry. Fourteen patients were treated with a combination of conformal external beam radiotherapy (70 Gy) and hyperthermia. Hyperthermia was delivered using the Coaxial TEM system, one treatment per week, to a total of five treatments. Thermometry was performed in bladder, urethra, rectum and esophagus. Invasive thermometry in the prostate was carried out during one or two treatments for each patient by placing transperineally a central and a peripheral catheter. Heterogeneous temperature distributions were measured in the prostate. The mean average invasive temperature range was 1.1°C. Due to the temperature heterogeneity and a limited number of thermometry sensors (mean 7, range 2-13), large variability between treatments and patients existed regarding achieved temperatures and dose. The mean invasive T 90 was 40.2 &#45 0.6°C and T 50 was 40.8 &#45 0.6°C. The mean Cum min T 90 >40.5°C per treatment was 22 (range 0-50). Importantly, intra-luminal temperatures did not reliably predict invasively measured temperatures. Invasive thermometry, therefore, remains compulsory to calculate a thermal dose for an individual patient. Changes in temperature during treatment, measured by the urethral sensors, corresponded well with changes in temperature measured by the individual invasive sensors. Similar comparison of rectal temperature changes with intra-prostatic temperature changes was not as predictive. The similarity in temperature changes between the urethral and interstial sites, suggests that urethral temperatures are sufficient for treatment optimization. The SAR profile did not correspond with the temperature profile indicating heterogeneous perfusion. Although regional hyperthermia in combination with external beam radiotherapy for locally advanced prostate carcinoma is clinically feasible, the question on the importance of invasive thermometry remains.     

Temperature-dependency of the expression of heat shock proteins in rat hepatoma cells with a different degree of heat sensitivity

Of two rat hepatoma cell lines, Reuber H35 and HTC, with a different degree of differentiation and heat sensitivity, treated at 41–44°C for various periods of time, the expression of thermotolerance, recovery of protein synthesis and rate of hsp synthesis were studied in relation to heat-induced cell killing. The temperature dependency of the induction of thermotolerance, either chronic or acute, was different between the cell lines; tolerance of H35 cells being induced at lower temperature than that of HTC cells. Induction of thermotolerance was related to survival decrement and the correlation was similar for both cell lines. Protein syntheses of H35 and HTC cells are equally sensitive for incubation at increased temperature, but recovery of protein synthesis after heat treatment was slightly different between the cell lines; H35 being more delayed than HTC. A strong but different correlation between total amount of protein synthesized after heat treatment and cell survival was found for the two cell lines. The increased heat sensitivity of H35 cells parallels the increased capacity of selective hsp synthesis as compared with that of HTC cells.     

Bioluminescence imaging of metabolites in a human tumour xenograft after treatment with hyperthermia and/or the radiosensitizer pimonidazole

The levels and distribution of ATP, glucose and lactate in human tumour xenografts following either single or combined treatment with hyperthermia (43°C for 30 min) and/or pimonidazole (1mg/gb.w) were determined by bioluminescence and compared with the mean 'global' levels obtained from the same tumours using conventional biochemical analysis. In general, the levels of ATP, glucose and lactate measured with both methods were in good agreement although the latter was consistently lower after bioluminescence determination. Compared with controls, neither the levels of ATP nor glucose were greatly affected in this tumour following treatment with the various modalities, whereas those of lactate were considerably increased as determined by both methods. The spatial distribution of ATP and glucose from controls and treated tumours was largely confined to the periphery and generally remained unchanged irrespective of treatment without any apparent alterations in the shape of the distribution curve. However, the increased lactate levels tended to accumulate towards the central region of the tumour after hyperthermia and/or sensitizer, showing an almost Gaussian-like distribution compared with controls. These results are in agreement with previous studies of global tumour metabolism showing an enhanced glycolytic activity with increased lactate levels after single or combined modality treatment.     

Colorectal carcinoma in a rat model: suppression of tumour development and altered host immune status following treatment with anti B-lymphocyte serum

The rate of colonic tumour development and immune capability in rats whose B-lymphocyte function was suppressed by injections of rabbit anti-rat IgM and also given the carcinogen dimethylhydrazine (DMH) were studied. Four rat groups were arranged to receive either DMH + anti-IgM, DMH + normal rabbit serum (NRS), saline + anti-IgM, or saline + NRS. Tumour weight, blood and mesenteric lymph node B-lymphocyte numbers, in vivo allograft response, in vitro lymphocytotoxicity, and leucocyte migration inhibition response (LMI) were recorded fortnightly. Tumour induction was delayed in the DMH + anti-IgM (treated tumour) group, which developed less tumour than the DMH + NRS (untreated tumour) group (p less than 0.001). Spleen cell lymphocytotoxicities were depressed in treated rats when compared to either the saline + anti-IgM (treated control) rats or the untreated rats (P less than 0.02), whereas anti-IgM treatment suppressed lymphocytotoxicity responses in control rats (p less than 0.05). The untreated tumour rats were tumour immune by LMI; however, the treated tumour rats did not express this in vitro tumour immunity. The B-lymphocyte levels in the mesenteric lymph nodes of untreated tumour rats increased with tumour induction (p less than 0.05), whereas in the treated tumour rats B-lymphocyte levels were not similarly stimulated.