Cell killing of melanoma B16 in vivo by hyperthermia and cytotoxins.

The aim of this study was to investigate antitumour activity of cisplatin, dacarbasine, cyclophosphamide and a new compound from the nitrosourea group--acetamido-CNU ((2-chloroethyl)-1-nitroso-3-(methylenecarboxamido)-urea)--applied with or without local hyperthermia (43.5 degrees C/60 min). The tumour model for the investigation of antitumour activity was a mouse melanoma B16 transplanted into the footpad. Dacarbasine, cyclophosphamide and acetamido-CNU applied as a single treatment had statistically significant antitumour activity, while cisplatin applied as a single agent had no effect. Local hyperthermia alone had statistically significant antitumour activity. The best therapeutic effect (synergistic) was obtained when combined treatment (cytotoxins plus local hyperthermia) was used. Synergistic therapeutic results were achieved even when cisplatin and hyperthermia were combined, although cisplatin was ineffective when given as a single agent. Therapeutic results achieved with acetamido-CNU (newly synthesized compound) applied alone were similar to the therapeutic results achieved with dacarbasine or cyclophosphamide. In combined therapy (acetamido-CNU + HT), achieved therapeutic results were significantly better (p < 0.05) than results achieved by combining cisplatin and hyperthermia or dacarbasine and hyperthermia.     

Cell killing of melanoma B16 in vivo by hyperthermia and cytotoxins

The aim of this study was to investigate antitumour activity of cisplatin, dacarbasine, cyclophosphamide and a new compound from the nitrosourea group - acetamido-CNU ((2-chloroethyl)-1-nitroso-3-(methylenecarboxamido)-urea) - applied with or without local hyperthermia (43.5°C/60 min). The tumour model for the investigation of antitumour activity was a mouse melanoma B16 transplanted into the footpad. Dacarbasine, cyclophosphamide and acetamido-CNU applied as a single treatment had statistically significant antitumour activity, while cisplatin applied as a single agent had no effect. Local hyperthermia alone had statistically significant antitumour activity. The best therapeutic effect (synergistic) was obtained when combined treatment (cytotoxins plus local hyperthermia) was used. Synergistic therapeutic results were achieved even when cisplatin and hyperthermia were combined, although cisplatin was ineffective when given as a single agent. Therapeutic results achieved with acetamido-CNU (newly synthesized compound) applied alone were similar to the therapeutic results achieved with dacarbasine or cyclophosphamide. In combined therapy (acetamido-CNU + HT), achieved therapeutic results were significantly better ( p <0.05) than results achieved by combining cisplatin and hyperthermia or dacarbasine and hyperthermia.     

Increased tumour response of a murine fibrosarcoma to low temperature hyperthermia and low dose rate brachytherapy

The present animal tumour study was carried out to determine the effectiveness of low temperature hyperthermia combined with low dose rate radiation based on the cell culture studies of our laboratory and others that demonstrated a significant radiosensitization obtained by low temperature hyperthermia and low dose rate radiation. Well-oxygenated murine fibrosarcoma Meth-A tumours growing in Balb/c mice were treated with heat (41d`C tumour temperature) by immersion of the tumour-bearing leg in a waterbath concurrently with low dose rate radiation. Radiation was delivered using <sup>192</sup>Ir interstitial implantation at absolute dose rates of 0.416–0.542 Gy/h. The effect of heat alone on tumour growth and normal tissue was minimal. Tumour growth delay following 30 Gy radiation was 4.9 days. Significant delay in tumour growth was observed with the addition of low temperature hyperthermia delivered concurrently. Enhancement in radiation response was seen with increasing duration of heat treatment; tumour growth delays were 9.5 days following 4h heat (41d`C) treatment and 16 days following 6 h treatment. Three sessions of fractionated hyperthermia 4 h/day during the course of low dose-rate radiation significantly delayed tumour growth to 18.6 days. The results indicate that fractionated heat treatment in conjunction with low dose rate radiation has potential for improving tumour response without adversely affecting normal tissue reaction. This in vivo study represents an extension of the cell culture data and provides further radiobiological basis for the combined use of low temperature hyperthermia and low dose rate radiation.     

Comparative study of thermoradiosensitization by misonidazole and metronidazole in vivo: antitumour effect and pharmacokinetics

Tumour control by local hyperthermia (43–5°C, 30 min) and radiation (20 Gy) given in combination with misonidazole (MISO) or metronidazole (METRO) was studied using FSa-II murine fibrosarcoma. When MISO or METRO (5 mmol/kg) was given 30 min before heat and subsequently treated with radiation, tumour regression was observed for both agents. Radiation dose-response curves for MISO and METRO with heating at 43·5°C for 30 min were identical. Mouse foot reaction was used to evaluate local toxicity following combined heat, a nitroimidazole and radiation treatment. MISO enhanced the magnitude of foot reaction and prolonged the recovery time compared with heat plus radiation controls. There were no observable differences of foot reaction between animals treated with heat plus radiation and those animals treated with heat, radiation and METRO. Pharmacokinetics of the nitroimidazoles heated at 43·5°C for 30 min in FSa-II tumours were investigated as a possible mechanism of thermal sensitization. Local hyperthermia did not alter the pharmacokinetics of METRO. Tumour concentration and tumour/plasma ratio of MISO were slightly decreased during heating. Since the hypoxic metabolism of the nitroimidazoles did not increase significantly during the heat treatment, the thermal enhancement of MISO or METRO radiosensitization cannot be explained by the increase in hypoxic cytotoxicity of the nitroimidazoles at elevated temperature alone. The two nitroimidazoles also were not accumulated in the tumour after heating. Therefore, alternation of pharmacokinetics is not the major mechanism for the thermal enhancement of nitroimidazole radiosensitization. The METRO radiosensitization effect became identical to that of MISO at elevated temperatures is of particular importance in clinical radiosensitization. The very low local and systemic toxicity together with the high efficacy of METRO at elevated temperatures will make it an attractive candidate as a future clinical radiosensitizer.     

Cisplatin,hyperthermia and radiation treatment in human cisplatin-sensitive and resistant glioma cell lines

In this study, the effects of mild protracted hyperthermia, combined with prolonged exposure to cisplatin and low dose-rate irradiation (LDRI), were examined in two human cell lines. The cell lines are human glioma parental and cisplatin resistant variant cells. The results show that mild hyperthermia at 40d`C was able to sensitize both the parental and the variant cisplatin-resistant cells to cisplatin treatments (1 μg/ml for up to 20 h) when the two treatments were given concur rently. When mild hyperthermia and cisplatin were given with LDRI concurrently, additional enhanced cell killing was observed in both the parental and the cisplatin-resistant variant cells. Further analysis of the results showed that when the effects of the trimodality treatment were normalized to the effects of the combined treatment of mild hyperthermia with cisplatin, the residual cell killing was still greater than that observed for radiation alone, indicating a synergistic interaction. This synergistic interaction was greater for the parental line compared to the cisplatin-resistant line. Thus, these data show that the concurrent application of mild hyperthermia, low concentration, long duration, cisplatin and low dose-rate irradiation may be an effective form of treatment in both normally responding and cisplatin-resistant variant human tumour cell lines.     

Hyperthermic enhancement of tumour growth inhibition by accelerated carbon-ions in transplantable human esophageal cancer

The study examined the effects of combination of hyperthermia (42d`C) and 290MeV/u carbon-ion (C—) beams or 200 kVp X-rays on tumour regrowth delay of transplantable human esophageal cancer as an in vivo model for radiotherapy of cancer. The C-beams were more effective in the tumour growth inhibition than X-rays. The relative biological effectiveness (RBE) of C-beams against X-rays was 2.00. It was observed that the interactive hyperthermic (42d`C, for 30 min) enhancement of tumour regrowth delay by high—linear energy transfer (LET) C-beams was similar to that of combination of low—LET X-rays with hyperthermia. The thermal enhancement ratios (TER) were 6.10 and 5.57 for X-rays and C-beams, respectively. These results suggest that hyperthermic treatment is effective in radiotherapy not only by low-LET radiation but also by high—LET radiation such as C-beams. In conclusion, the depression of the tumour growth by the combined treatment of hyperthermia (42d`C) and the C-beams strongly suggests the available possible application of interdisciplinary cancer therapy for refractory tumours.     

Comparative study of thermoradiosensitization by misonidazole and metronidazole in vivo: antitumour effect and pharmacokinetics.

Tumour control by local hyperthermia (43.5 degrees C, 30 min) and radiation (20 Gy) given in combination with misonidazole (MISO) or metronidazole (METRO) was studied using FSa-II murine fibrosarcoma. When MISO or METRO (5 mmol/kg) was given 30 min before heat and subsequently treated with radiation, tumour regression was observed for both agents. Radiation dose-response curves for MISO and METRO with heating at 43.5 degrees C for 30 min were identical. Mouse foot reaction was used to evaluate local toxicity following combined heat, a nitroimidazole and radiation treatment. MISO enhanced the magnitude of foot reaction and prolonged the recovery time compared with heat plus radiation controls. There were no observable differences of foot reaction between animals treated with heat plus radiation and those animals treated with heat, radiation and METRO. Pharmacokinetics of the nitroimidazoles heated at 43.5 degrees C for 30 min in FSa-II tumours were investigated as a possible mechanism of thermal sensitization. Local hyperthermia did not alter the pharmacokinetics of METRO. Tumour concentration and tumour/plasma ratio of MISO were slightly decreased during heating. Since the hypoxic metabolism of the nitroimidazoles did not increase significantly during the heat treatment, the thermal enhancement of MISO or METRO radiosensitization cannot be explained by the increase in hypoxic cytotoxicity of the nitroimidazoles at elevated temperature alone. The two nitroimidazoles also were not accumulated in the tumour after heating. Therefore, alternation of pharmacokinetics is not the major mechanism for the thermal enhancement of nitroimidazole radiosensitization. The METRO radiosensitization effect became identical to that of MISO at elevated temperatures is of particular importance in clinical radiosensitization. The very low local and systemic toxicity together with the high efficacy of METRO at elevated temperatures will make it an attractive candidate as a future clinical radiosensitizer.     

Optimization of power deposition and the rate of heating of tissue during RF capacitive hyperthermia

The toxicity of the radiosensitizers misonidazole (MISO), demethylmisonidazole (DEMISO) and pimonidazole (PIM) in mice can be affected differently when combined with local hyperthermia at 43°C for 30 min. At a dose of 1 mg/g, only MISO plus heat resulted in 50% lethality in animals over a period of 7 days post-treatment, whereas 100% survival was observed in the case of DEMISO and PIM. The enhanced lethality may be associated with the production of toxic intermediates of MISO. Heat did not affect the levels of DEMISO in the tissues studied (plasma, brain and tumour), whereas those of PIM were markedly lowered in tumour but not affected in brain for up to 4 h after combined treatment. MISO was found to be decreased in the tumour at all times but affected differently in brain after 1 and 2 h. initially decreasing and then increasing significantly. In all cases the treatment sequence, i.e. sensitizer plus heat or vice-versa, did not affect the rate of survival. At a dose of 2 mg/g, DEMISO plus heat was found to be more toxic when DEMISO was given first (25% survival) compared to 58% on reversal. However, the levels of DEMISO in the tissues were not affected by heat. Thus, it would appear that there is no correlation between parent drug levels measured in plasma, tumour or brain and hyperthermia-induced drug lethality.     

Effects of local hyperthermia on the tissue levels and toxicity of three radiosensitizers in mice.

The toxicity of the radiosensitizers misonidazole (MISO), demethylmisonidazole (DEMISO) and pimonidazole (PIM) in mice can be affected differently when combined with local hyperthermia at 43 degrees C for 30 min. At a dose of 1 mg/g, only MISO plus heat resulted in 50% lethality in animals over a period of 7 days post-treatment, whereas 100% survival was observed in the case of DEMISO and PIM. The enhanced lethality may be associated with the production of toxic intermediates of MISO. Heat did not affect the levels of DEMISO in the tissues studied (plasma, brain and tumour), whereas those of PIM were markedly lowered in tumour but not affected in brain for up to 4 h after combined treatment. MISO was found to be decreased in the tumour at all times but affected differently in brain after 1 and 2 h, initially decreasing and then increasing significantly. In all cases the treatment sequence, i.e. sensitizer plus heat or vice-versa, did not affect the rate of survival. At a dose of 2 mg/g, DEMISO plus heat was found to be more toxic when DEMISO was given first (25% survival) compared to 58% on reversal. However, the levels of DEMISO in the tissues were not affected by heat. Thus, it would appear that there is no correlation between parent drug levels measured in plasma, tumour or brain and hyperthermia-induced drug lethality.     

Tumour-specific enhancement of thermoradiotherapy at mild temperatures by the vascular targeting agent 5,6-dimethylxanthenone-4-acetic acid.

The effect of combining the vascular targeting agent 5,6-dimethylxanthenone-4-acetic acid (DMXAA) with both radiation and hyperthermia treatments was investigated in a transplanted C3H mouse mammary carcinoma and a normal mouse tissue. Tumours were grown on the right rear foot of female CDF1 mice and treated when sized 200 mm3. The foot skin of non-tumour-bearing CDF1 mice was used to assess normal tissue damage. Radiation and hyperthermia were given locally to the tumour/skin of restrained non-anaesthetized animals. DMXAA (20 mg/kg) was dissolved in saline and injected intraperitoneally 1 h after irradiating and then heating started 3 h later. The endpoints were local tumour control within 90 days or the development of moist desquamation in skin between 11 and 23 days after treatment. The radiation dose (+/- 95% confidence intervals) producing local tumour control in 50% of treated animals was 53 (51-55) Gy for radiation alone. This value was significantly (Chi-squared test; p < 0.05) decreased to 47 (42-52) Gy by DMXAA and to 47 (44-51) Gy by heating (41.5 degrees C/60 min) 4 h after irradiation. Combining both DMXAA and heating further reduced this to 30 (26-35) Gy. When the heating temperature was decreased to 40.5 degrees C, the effect of the triple combination was decreased but was still significant compared with radiation + DMXAA or radiation + hyperthermia. However, this enhancement disappeared at 39.5 degrees C. Radiation damage of normal foot skin was not enhanced by combining DMXAA and hyperthermia at 41.5 degrees C. In conclusion, adding DMXAA to thermoradiotherapy at 40.5-41.5 degrees C significantly improved local tumour control without enhancing normal tissue damage. Thus, including a vascular targeting agent in a mild thermoradiotherapy treatment regimen is a useful approach that may lead to a re-evaluation of the use of hyperthermia in cancer treatment.     

Whole-body hyperthermia and lonidamine as adjuvant therapy to treatment with cisplatin with or without local radiation in mouse bearing the Lewis lung carcinoma

The Lewis lung carcinoma implanted subcutaneously in the hind leg of a C57BL mouse metastasizes avidly to the lungs of the host. This tumour model system thus allows assessment of both primary and metastatic disease to treatment. Lonidamine (50 mg/kg) administered once or twice daily produced approximately additive tumour growth delay with whole-body hyperthermia (60 min to 42°C and 60 min at 42°C). The addition of lonidamine to treatment with cisplatin (10 mg/kg) and whole-body hyperthermia continued to produce increased tumour growth delay of up to 14·7 days compared with 10·8 days for cisplatin/whole-body hyperthermia. The response of the metastatic disease paralleled that of the primary tumour with a reduction in the number and percent of large metastases (> 3 mm) on day 20 post-tumour implantation. The addition of local fractionated radiation therapy (3 Gy × 5) to the primary tumour produced a very effective treatment regimen resulting in 37·5 days of tumour growth delay along with twice daily lonidamine/cisplatin whole-body hyperthermia. With this treatment regimen there was also a reduction to 50% of control of the number of lung metastases as well as the percent of large metastases on day 20. Further investigation of these treatment combinations is warranted.     

Tumour-specific enhancement of thermoradiotherapy at mild temperatures by the vascular targeting agent 5,6-dimethylxanthenone-4-acetic acid

The effect of combining the vascular targeting agent 5,6-dimethylxanthenone-4-acetic acid (DMXAA) with both radiation and hyperthermia treatments was investigated in a transplanted C3H mouse mammary carcinoma and a normal mouse tissue. Tumours were grown on the right rear foot of female CDF1 mice and treated when sized 200?mm³. The foot skin of non-tumour-bearing CDF1 mice was used to assess normal tissue damage. Radiation and hyperthermia were given locally to the tumour/skin of restrained non-anaesthetized animals. DMXAA (20?mg/kg) was dissolved in saline and injected intraperitoneally 1?h after irradiating and then heating started 3?h later. The endpoints were local tumour control within 90 days or the development of moist desquamation in skin between 11 and 23 days after treatment. The radiation dose (±95% confidence intervals) producing local tumour control in 50% of treated animals was 53 (51–55) Gy for radiation alone. This value was significantly (Chi-squared test; p < 0.05) decreased to 47 (42–52) Gy by DMXAA and to 47 (44–51) Gy by heating (41.5°C/60?min) 4?h after irradiation. Combining both DMXAA and heating further reduced this to 30 (26–35) Gy. When the heating temperature was decreased to 40.5°C, the effect of the triple combination was decreased but was still significant compared with radiation + DMXAA or radiation + hyperthermia. However, this enhancement disappeared at 39.5°C. Radiation damage of normal foot skin was not enhanced by combining DMXAA and hyperthermia at 41.5°C. In conclusion, adding DMXAA to thermoradiotherapy at 40.5–41.5°C significantly improved local tumour control without enhancing normal tissue damage. Thus, including a vascular targeting agent in a mild thermoradiotherapy treatment regimen is a useful approach that may lead to a re-evaluation of the use of hyperthermia in cancer treatment.     

The interaction of cisplatin plus etoposide with radiation +/- hyperthermia

The addition of concurrent etoposide and cisplatin to radiation +/- hyperthermia was evaluated in the murine FSaIIC fibrosarcoma tumor system. Tumor growth delay (TGD) demonstrated that when the drugs were tested with radiation (3 Gy daily X 5) plus (43 degrees X 30 min) local hyperthermia, cisplatin/hyperthermia/radiation (TGD approximately 25 days) was significantly more effective than etoposide/hyperthermia/radiation (TGD approximately 14 days). The addition of etoposide to cisplatin/hyperthermia/radiation, however, yielded a significantly longer growth delay (approximately 34 days). Tumor cell survival studies demonstrated that hyperthermia (43 degrees C, 30 minutes) was dose modifying for etoposide cytotoxicity (dose modifying factor approximately 2.0 as determined by comparisons of the slopes of the curves). The addition of etoposide to cisplatin modified cisplatin killing only slightly at 37 degrees C or 43 degrees C. Considerable additional cell kill was observed over a range of radiation doses with cisplatin, hyperthermia, and etoposide added singly or in combination, especially at the lowest radiation dose tested (5 Gy), but essentially no dose modification was observed. Evaluation of Hoechst 33342 dye-selected tumor subpopulations demonstrated that cisplatin, etoposide, radiation (10 Gy), etoposide plus radiation, and cisplatin plus radiation killed significantly fewer dim (presumably hypoxic) cells than bright (presumably normally oxygenated) cells. Hyperthermia killed more dim than bright cells. The combination of hyperthermia with cisplatin and radiation, however, resulted in approximately 5-fold lesser kill in dim cells, and the addition of etoposide increased this differential to 6.4-fold. These results indicate that etoposide adds small but measurable antitumor effects when used with cisplatin alone or with cisplatin in combination with radiation +/- hyperthermia (especially at lower radiation fraction sizes).     

Localized thermo-cisplatin therapy: a pilot study in spontaneous canine and feline tumours.

Local hyperthermia combined with intralesional cisplatin chemotherapy is a logical and potentially effective therapeutic approach for localized cancers. A trial using outbred animals with spontaneously occurring tumours was initiated to evaluate the toxicity and efficacy of this approach. Treatment consisted of injection of a colloidal suspension of cisplatin into the tumour prior to hyperthermia once a week for 4 weeks. Immediately after intratumoral injection of a mixture of cisplatin and collagen, thermotherapy was given. The goal temperature was 42 +/- 1 degrees C for 30 min. Ten animals (nine dogs and one cat) with soft tissue neoplasms were treated with one to four hyperthermia and cisplatin sessions for a total of 30 treatment sessions. Complete responses occurred in 4/10 cases (one carcinoma, two sarcomas, one melanoma). One dog with haemangiopericytoma had partial response. The lack of systemic toxicity and the minimal local normal tissue reactions indicate that the treatments were well tolerated. These data provide preliminary evidence that a combination of local hyperthermia and intratumoral cisplatin chemotherapy is a safe and effective method for the treatment of selected localized neoplasms.     

Interaction of hyperthermia and cis-diamminedichloroplatinum(II) alone or combined with radiation in a C3H mammary carcinoma in vivo

The interaction between hyperthermia and cis-diamminedichloroplatinum(II) (c-DDP) given in various schedules as an adjuvant to radiation treatment was investigated in a C3H mouse mammary carcinoma in vivo. Both hyperthermia (43.5 degrees C for 60 min) and c-DDP (6 mg/kg i.p.) caused a delay in tumor growth when given individually. When c-DDP was given 4 h prior to hyperthermia, the increase in tumor growth time corresponded to an additive effect, but when the interval was reduced to 15 min, the tumor growth delay was significantly greater than additive. The modifying effect of these schedules on radiation was studied using local tumor control (50% tumor control dose) as the endpoint. c-DDP alone did not result in any enhancement of tumor control, irrespective of whether it was given 15 min or 4 h after irradiation. In contrast, heat treatment at 43.5 degrees C for 60 min given 4 h after irradiation resulted in a significant reduction in the 50% tumor control dose, with an enhancement ratio of 1.8. From a clamped local tumor control assay, it was found that c-DDP selectively killed aerobic cells, whereas hyperthermia was primarily directed toward the hypoxic clonogenic cells in the tumors. Combining the two modalities (simultaneously) resulted in a significant additional increase in the killing of well-oxygenated clonogenic cells, but the destruction of hypoxic cells was not different from that obtained after heat alone.     

Augmented antitumour effects of combined treatment with hyperthermia and tumour necrosis factor on human gastric cancer xenotransplanted into nude mice

Hyperthermia combined with recombinant human tumour necrosis factor (rH-TNF) was evaluated for antitumour efficacy in vivo. Use was made of human gastric cancer tissues xenografted into nude mice. When 100, 300, 600, and 1200 units of rH-TNF (2.4 x 10(6) units/mg protein) were given twice intraperitoneally, tumour regression did not occur in any animal. In contrast, a remarkable suppression of tumour growth was observed when 600 and 1200 units of rH-TNF was given in combination with hyperthermia at 43.5 +/- 0.1 degrees C. No effects were evident with the regimen of 100 and 300 units of rH-TNF plus hyperthermia at the same temperature, as compared with evidence obtained with hyperthermia alone. The tumoral blood flow, determined by the hydrogen diffusion method, decreased immediately after hyperthermia alone or hyperthermia plus 1200 units of rH-TNF, whereas a slight decrease was seen after rH-TNF alone. When hyperthermia plus 1200 units of rH-TNF were given, there was a remarkable delay in reversion to pretreatment values of tumoral blood flow, as compared to findings with rH-TNF only or heat only. These results are discussed in relation to the antitumour and side-effects of rH-TNF.     

The in vivo interaction between flavone acetic acid and hyperthermia

The in vivo interaction between flavone acetic acid (FAA) and hyperthermia was studied in a C3H mammary carcinoma grown in the feet of female CDFl mice and in normal foot skin. FAA was intraperitoneally injected prior to local tissue heating in restrained non-anaesthetized animals. Alone, FAA at doses of 100 mg/kg and above, inhibited tumour growth in a dose-dependent fashion. FAA also enhanced the tumour response to heat, the effect being dependent on both the time interval between the two modalities and the FAA dose, the greatest effect occurring when FAA doses of ≥ 150 mg/kg preceeded heat by 3–48h. These effects of FAA correlated with the drug's ability to decrease tumour blood perfusion measured using the RbCl extraction procedure. Injecting 150 mg/kg FAA 3h before heating (42.7d`C) resulted in a 2–2-fold increase in tumour heat damage, but had little effect on the response of normal foot skin in non-tumour-bearing mice. However, this treatment gave a 20-fold increase in normal tissue damage when the skin experiments were repeated in tumour-bearing animals. These effects in skin occurred in the absence of any blood perfusion changes, but appeared to be associated with FAA-induced TNF-α production.     

Localized thermo-cisplatin therapy: A pilot study in spontaneous canine and feline tumours

Local hyperthermia combined with intralesional cisplatin chemotherapy is a logical and potentially effective therapeutic approach for localized cancers. A trial using out bred animals with spontaneously occurring tumours was initiated to evaluate the toxicity and efficacy of this approach. Treatment consisted of injection of a colloidal suspension of cisplatin into the tumour prior to hyperthermia once a week for 4 weeks. Immediately after intratumoral injection of a mixture of cisplatin and collagen, thermotherapy was given. The goal temperature was 42 ± 1° for 30 min. Ten animals (nine dogs and one cat) with soft tissue neoplasms were treated with one to four hyperthermia and cisplatin sessions for a total of 30 treatment sessions. Complete responses occurred in 4/10 cases (one carcinoma, two sarcomas, one melanoma). One dog with haem-angiopericytoma had partial response. The lack of systemic toxicity and the minimal local normal tissue reactions indicate that the treatments were well tolerated. These data provide preliminary evidence that a combination of local hyperthermia and intratumoral cisplatin chemotherapy is a safe and effective method for the treatment of selected localized neoplasms.     

Thermal enhancement of drug uptake and DNA adducts as a possible mechanism for the effect of sequencing hyperthermia on cisplatin-induced cytotoxicity in L1210 cells

An optimal scheduling of hyperthermia andcis-diammine-dichloroplatinum(II) (cisplatin) may increase the therapeutic gain of the combination of these two modalities. In this study, intracellular platinum accumulation, total platinum binding to DNA, and DNA interstrand crooslinks (ISC) were assayed to investigate the molecular mechanisms responsible for the effect of sequencing hyperthermia on the thermal enhancement of cisplatin-induced cytotoxicity in mouse leukemia L1210 cells. Simultaneous treatment with heat (41.5°C, 60 min) and cisplatin produced maximal cell killing with a 4-fold decrease in the 50% growth-inhibitory concentration (IC₅₀) of the platinum complex. Super-additive cell killing was also shown when cells were exposed to heat before cisplatin treatment, whereas no thermal enhancement in cisplatin-mediated cytotoxicity was observed in cells given heat after exposure to cisplatin. These results correlated with the degree of formation of ISC observed in cells following various treatments. A 2-to 3-fold increase in ISC formation was observed in cells given heat before or during cisplatin exposure, whereas heat after cisplatin treatment did not alter either the formation or the reversal of ICC as compared with cisplatin alone. The increased ISC formation was associated with an increase in intracellular platinum accumulation and total platinum binding to DNA in cells given heat before or during cisplatin exposure. These data, showing that hyperthermia potentiates cisplatin cytotoxicity by increasing drug uptake and the formation of DNA adducts without inhibiting the repair of DNA lesions, demonstrate the potential utility of sequencing hyperthermia combined with cisplatin as a clinical anticancer therapy.This work was supported by National Cancer Institute grants, R01-CA-43090, R01-CA-41581, R01-CA-50380, R01-CA-40090, and American Cancer Society-CH-324E. S. Ohno and Y. Kido are on leave from the Department of Surgery II, Faculty of Medicine, Kyushu University, Fukuoka 812, Japan     

Pilot study of local hyperthermia,radiation therapy,etanidazole, and cisplatin for advanced superficial tumours

Five patients (six hyperthermia sites) with advanced superficial tumours were treated with combined etanidazole, cisplatin, local hyperthermia, and radiation therapy as part of a Phase I pilot study. Treatment was given once weekly and consisted of etanidazole 3 gm/m² IV bolus, cisplatin 50 mg/m² IV bolus, hyperthermia for 60 min with a target temperature of 43°C, and radiation therapy 500 cGy/fraction (median total dose 3000 cGy) for a total of six weeks. Blood levels of etanidazole were taken during treatment at week 1 and week 4. Etanidazole drug exposure was calculated using the trapezoidal rule and expressed as the area under the curve (AUC) of plasma concentration X time. Five of six treatment sites had received prior irradiation. Prior chemotherapy had been given in three patients and tamoxifen therapy given in the other two patients. The median follow-up time is 34 months; 3/5 patients have died of disease. The most significant toxicity was grade I or II nausea and vomiting associated with 19/32 treatments (59%) and a second degree burn in 2/6 fields. None of the five patients experienced peripheral neuropathy, skin ulceration, or needed surgical repair. In addition, there was mild renal toxicity; pharmacokinetic analysis showed a 28–75% increase in the week 1 to week 4 AUC in three patients, all of whom had a decrease in creatinine clearance over the same time of 15–47%. This pilot study suggests this combined modality therapy can be delivered without major complications and that renal function, determined by creatinine clearance, affects clearance of etanidazole and alters the AUC. Therefore, monitoring renal function is important in patients receiving etanidazole in addition to other nephrotoxic agents such as cisplatin. The impact of etanidazole on the therapeutic index of hyperthermia, radiation therapy and cisplatin may be worthy of study, especially since a positive interaction between these modalities is found in laboratory models.