Effects of a controlled-release cisplatin delivery system used after resection of mammary carcinoma in mice.

OBJECTIVE: To evaluate efficacy of a controlled-release cisplatin delivery system, used after marginal resection of mammary carcinoma (ie, resection of grossly evident tumor) in mice, to prevent tumor regrowth and metastasis. ANIMALS: 42 female C3H-HeJ mice. PROCEDURE: Mice were inoculated with mammary carcinoma cells. Between 2 and 6 days later, tumors were marginally resected and mice were assigned to 1 of 3 groups: no treatment (control; n = 14), cisplatin administered intraperitoneally (i.p. cisplatin; 14), and cisplatin delivered by use of an open-cell polylactic acid system placed within the tumor bed (slow-release cisplatin; 14). Tumor regrowth was measured daily. Mice were euthanatized 14 days after surgery, and complete necropsies were performed. RESULTS: Tumor regrowth was not detected in the slow-release cisplatin group; however, tumor regrowth was detected in 7 of 14 mice in the i.p. cisplatin group and 14 of 14 mice in the control group. Median (+/-SD) number of days to tumor regrowth was 13.5+/-0.64 and 7.79+/-0.87 in the i.p. cisplatin and control groups, respectively. Mice in the i.p. cisplatin group had significantly delayed tumor regrowth, compared with control mice. Metastases to lungs were detected in 8 of 14 control mice but were not detected in mice in either cisplatin treatment group. CONCLUSIONS AND CLINICAL RELEVANCE: The open-cell polylactic acid with cisplatin delivery system was successful in delaying local tumor regrowth and metastasis in mice with marginally resected mammary carcinoma. Use of a controlled-release cisplatin delivery system may be an effective adjunct treatment following excision of mammary carcinoma in humans and other animals.

Slow release cisplatin combined with radiation for the treatment of canine nasal tumors.

Thirteen dogs with malignant tumors of the nasal cavity were treated with a combination of slow release cisplatin and megavoltage radiation. Radiation was delivered on a Monday through Friday schedule using a 6 MV linear accelerator. The median total dose was 49.5 Gy (range 49.5-56 Gy). Cisplatin was given using an open-cell polylactic acid polymer, impregnated with the drug and implanted intramuscularly at a distant site, as a slow release delivery system (OPLA-Pt [THM Biomedical, Inc]). The median dose used was 60 mg/m2 (range 60-100 mg/m2). When combined with radiation, this delivery system caused no systemic drug toxicity, and a local tissue reaction was seen in only two dogs. Acute side effects to normal tissue from radiation were not enhanced, as measured by subjective assessment. When compared to a group of historical controls that received radiation without OPLA-Pt, the dogs that received combined radiation and cisplatin had longer overall survival times, with a median of 580 days. The control group had a median survival of 325 days. Previously reported median survival times for comparable megavoltage radiation treatment range from 6 to 13 months. Some dogs in both groups also received adjubant chemotherapy but this did not influence survival time. By multivariate analysis, only the use of OPLA-Pt was found to significantly influence survival, with a p value of p = 0.023. Mega-voltage radiation and slow release cisplatin appears to be a well tolerated combination that may favorably affect survival of dogs with nasal tumors.

Irradiation-enhanced binding of carboplatin to DNA.

Carboplatin (3.6 mM) and salmon sperm DNA (3 micrograms), either double-stranded (dsDNA) or denatured single-stranded) (ssDNA), were irradiated (100 Gy) under hypoxic or oxic conditions. The drug and DNA were mixed either before, during, or after irradiation, and platinum binding to DNA was measured using atomic absorption spectrometry. Irradiation significantly increased the binding of carboplatin to dsDNA under hypoxic conditions, but did not increase the binding of carboplatin to ssDNA under similar hypoxic conditions. Irradiation did not result in an increased dsDNA-platinum binding under oxic conditions. When carboplatin was added to dsDNA immediately following irradiation of dsDNA, no increase in platinum binding was observed. When carboplatin and dsDNA were irradiated simultaneously but separately and then mixed there appears to be a slight increase in platinum binding, but it was not significant. Tetra-ammineplatinum (II) chloride, a non-cytotoxic platinum compound, did not show an increase in platinum-DNA binding following irradiation under hypoxic conditions. The results suggest that radiation-enhanced carboplatin binding to DNA might be at least partly responsible for radiosensitization by carboplatin, especially in a cell line sensitive to carboplatin.

Enhanced radiation-induced cell killing by carboplatin in cells of repair-proficient and repair-deficient cell lines.

The objective of this study was to determine whether a deficiency for either one of two repair processes influences the phenomenon of enhancement of radiation-induced cell killing by carboplatin which has been reported previously in one cell line (V79) and which is presumably a result of an interaction between these two therapeutic modalities. Cell killing was enhanced in cells of four cell lines when the cells were exposed to carboplatin before and during irradiation in either air or hypoxia. In cell lines proficient in both excision repair and DNA double-strand break repair (K1 and AA8), and in a cell line deficient in nucleotide excision repair (UV41), the enhancement was characterized as both a reduction in the shoulder region of the survival curves indicated by a reduced Dq and a reduction in D0 in the terminal region of the survival curves determined for cells exposed in air and under hypoxic conditions. Only the latter effect was observed in a cell line deficient in DNA double-strand break repair (xrs-5). The survival curves were fitted to the data using the repair saturation model and a computer program developed by N. Albright (Radiat. Res. 118, 112-130, 1989). In hypoxia, the reductions in Dq were as great as from 7.0 Gy to 2.1 Gy, 3.3 Gy to 0 Gy and 1.7 Gy to 0 Gy for K1, AA8 and UV41 cells, respectively. Sensitizer enhancement ratios ranged from 1.3 to 1.7 and were similar for irradiation in air and under hypoxic conditions. This enhanced cell killing by carboplatin combined with radiation required levels of the drug sufficient to produce cytotoxicity by the drug alone as exemplified by the UV41 cell line, which is intrinsically sensitive to carboplatin and in which 1/30 of the drug concentration required for the other cell lines produced an enhanced cell killing at an equitoxic dose of only 5 microM.

Irradiation enhances cellular uptake of carboplatin.

PURPOSE: Because radiation is known to damage cellular membranes, the purpose of this study was to determine whether irradiation of cultured cells might modify the cellular uptake of the chemotherapy agent carboplatin. METHODS AND MATERIALS: Total intracellular platinum was measured using atomic absorption spectrometry in cultured V79 cells and in four Chinese hamster ovary (CHO) cell lines. RESULTS: Intracellular carboplatin concentrations increased linearly with radiation dose (10-50 Gy) under both hypoxic and oxic irradiation conditions. Similar doses of radiation did not significantly increase the uptake of a nontoxic platinum compound [Pt(NH3)4Cl2.H2O] (p > 0.5). Compared to unirradiated controls, there was no increase in intracellular carboplatin concentrations when carboplatin was irradiated prior to administration to the cell cultures (p > 0.5). Within the 32.5 min or less required to deliver the radiation, a dose of 50 Gy produced approximately a 50% increase in intracellular platinum in V79 cells and approximately an increase of a factor of 1.3-1.6 in the CHO cell lines. Although the increase in drug uptake would be expected to be less than 10% for most cell lines at the doses of radiation used to investigate radiosensitization by carboplatin, this level of increase may play a significant role in the radioenhancement observed in UV41 cells because these excision-repair--deficient cells are much more sensitive to carboplatin as measured by cytotoxicity. CONCLUSION: These results suggest that some of the enhanced cell killing that results when cells are exposed to carboplatin in combination with radiation may be attributed to an increased cellular uptake. One mechanism of radiopotentiation may be an enhanced chemotoxicity resulting from a radiation-induced increase in carboplatin uptake.

Carboplatin enhances the production and persistence of radiation-induced DNA single-strand breaks.

Fluorometric analysis of DNA unwinding and alkaline elution were used to investigate the production and persistence of DNA single-strand breaks (SSBs) in Chinese hamster V79 and xrs-5 cells treated with the chemotherapeutic agent carboplatin in combination with radiation. Carboplatin was administered to cells before irradiation in hypoxic conditions, or the drug was added immediately after irradiation during the postirradiation recovery period in air. The results of DNA unwinding studies suggest that carboplatin enhances the production of radiation-induced SSBs in hypoxic V79 cells and xrs-5 cells by a factor of 1.86 and 1.83, respectively, when combined with radiation compared to the SSBs produced by irradiation alone. Carboplatin alone did not produce a measurable number of SSBs. Alkaline elution profiles also indicated that the rate of elution of SSBs was higher in cells treated with the carboplatin-radiation combination in hypoxia when compared to irradiation alone, resulting in an increased yield of radiation-induced SSBs by a factor of 1.46 in V79 cells with 20 Gy irradiation and by a factor of 2.02 in xrs-5 cells with 20 Gy irradiation. When carboplatin is present after irradiation and during the postirradiation recovery period, the rejoining of radiation-induced SSBs is inhibited during this postirradiation incubation period (radiopotentiation) with a relative inhibition factor at 1 h postirradiation of 1.25 in V79 cells and 1.15 in xrs-5 cells. An increased production and persistence of SSBs resulting from the interaction of carboplatin with radiation may be an important step in the mechanism responsible for the potentiated cell killing reported previously from studies in animal tumors and in cultured cells.

Production of DNA double-strand breaks by interactions between carboplatin and radiation: a potential mechanism for radiopotentiation.

The production of DNA double-strand breaks (DSBs) was studied in cells of four CHO cell lines under conditions where combining radiation with carboplatin enhanced cell killing (radiosensitization and radiopotentiation). The cell lines included repair-proficient (AA8 and K1), excision repair-deficient (UV41) and DSB repair-deficient (xrs-5) cells. Double-strand breaks were analyzed by neutral elution either immediately after or 4 h after a single 55-Gy radiation dose delivered under hypoxic conditions. Carboplatin (1 mM) combined with radiation produced a small increase in DSBs compared to radiation alone immediately after irradiation in AA8, UV41 and xrs-5 cells. However, the yield of DSBs in AA8, K1 and xrs-5 cells was significantly higher 4 h after carboplatin-radiation treatment; no such increase was found at 4h in UV41 cells. Strand scission factors (SSFs) were calculated as SSF = -log [percentage DNA remaining on filter at 8 h elution (treated cells)/percentage DNA remaining at 8 h elution (untreated cells)]. The ratios of the SSF at 4 h to 0 h postirradiation for carboplatin-treated cells were 13.7 for K1, 4.9 for xrs-5, 2.5 for AA8 and 1.2 for UV41 cells. These results support a possible explanation for the enhanced killing of irradiated cells by platinum chemotherapeutic agents, namely enhanced production of DSBs.

Effects of cis-diamminedichloroplatinum II released from D,L-polylactic acid implanted adjacent to cortical allografts in dogs.

This study was performed to determine the pharmacokinetics and local and systemic effects of cis-diamminedichloroplatinum II (cisplatin) released from an open-cell polylactic acid polymer when the drug delivery device was placed adjacent to a cortical allograft. Bilateral intercalary femoral allografts were implanted in six normal beagles. The polymer containing cisplatin was implanted adjacent to the allograft in one femur, and the polymer without cisplatin was implanted adjacent to the allograft in the contralateral femur. Systemic toxicity was evaluated clinically by hematologic and serum biochemistry tests and urinalysis. Healing of the allograft was monitored radiographically. The femora were evaluated biomechanically, histologically, and histomorphometrically 7.5 months after surgery. Total serum platinum levels were measured by atomic absorption spectrophotometry, and pharmacokinetic parameters were calculated. Healing was impaired slightly by the presence of the polymer with cisplatin, and systemic and local toxicity was mild and transient. After implantation of the polymer with cisplatin, the mean peak total serum platinum concentration was low (1.71 +/- 0.19 micrograms/ml). However, the area under the curve for total serum platinum concentration versus time for the first 21 days was large (27,050 +/- 3,201 micrograms.min/ml). When cisplatin was given as an intravenous bolus at a dose of 70 mg/m2 to six other beagles, the mean peak total platinum concentration was 8.80 +/- 2.1 micrograms/ml and the area under the curve was 940.3 +/- 256.7 micrograms.min/ml. These results indicate that a sustained release of cisplatin can be delivered safely from an open-cell polylactic acid polymer. This device may be useful in the treatment of solid tumors.

Effect of phase modulation on the temperature distribution of a microwave hyperthermia antenna array in vivo.

Perfused, canine skeletal muscle and the brain tumour of a cancer patient were heated with an array of four parallel, interstitial antennas placed on the corners of a 2-cm square and driven at 915 MHz. The temperature distributions along the axial and diagonal catheters were measured with equal-phase driving of the antennas and with several time-varying schemes of driving phase differences among the antennas. When equal-phase driving was replaced by a rotating scheme of 90 degrees driving phase differences, the tissue area in the junction plane heated above a normalized index temperature of 0.6 increased by a factor of about 1.25. With a rotating phase of 135 degrees, the same area increased by a factor of about 1.6. The axial temperature distribution was not affected significantly by driving phase.

Theoretical and experimental analysis of air cooling for intracavitary microwave hyperthermia applicators.

An intracavitary microwave antenna array system has been developed and tested for the hyperthermia treatment of prostate cancer at Thayer School of Engineering and Dartmouth-Hitchcock Medical Center. The antenna array consists of a choked dipole antenna inserted into the urethra and a choked dipole antenna eccentrically embedded in a Teflon obturator inserted into the rectum. To prevent unnecessary heating of the healthy tissue that surrounds each applicator, an air cooling system has been incorporated into the rectal applicator. The air cooling system was designed and modeled theoretically using a numerical solution of heat and momentum equations within the applicator, and an analytical solution of the Pennes bioheat equation in tissue surrounding the applicator. The 3-D temperature distribution produced by the air-cooled rectal applicator was measured in a perfused canine prostate.

A coaxial microwave applicator for transurethral hyperthermia of the prostate.

Benign prostatic hyperplasia (BPH) is a common disease of elderly men. The current definitive treatment for urinary obstruction caused by this disease is surgery (transurethral resection of the prostate, or TURP). Recent evidence suggests that hyperthermia may be a useful nonsurgical alternative for treatment of symptomatic BPH. A transurethral microwave applicator has been designed around a Foley catheter for delivery of local hyperthermia to the prostate. The Foley balloon is used to maintain the antenna position within the prostatic urethra. The Foley catheter also features an antenna choke to confine power deposition to the intended region. The antenna is a coaxial dipole designed to operate at 915 MHz. Qualitative and quantitative specific absorption rate (SAR) patterns are shown for this antenna. In vivo experiments in dog prostate demonstrate that temperatures > 42 degrees C can be obtained > 1 cm away from the catheter, while maintaining a maximum urethral temperature of 47 degrees C to 48 degrees C. Histology obtained acutely after the hyperthermia treatments showed minimal damage to the periurethral tissues. We conclude from these studies that this microwave applicator is capable of providing local hyperthermia to the prostatic tissues with a predictable and well-circumscribed thermal distribution.

Techniques for intraoperative hyperthermia with ultrasound: the Dartmouth experience with 19 patients.

Over the course of 3 years, tumours of 19 patients were heated with ultrasound in the operating room during surgical resection. Immediately following intraoperative radiation therapy, thermocouples were inserted into tumour and adjacent normal structures. Patients were then given a 60-min heat treatment with ultrasound after a 10-15-min heatup period. Temperatures were measured at a total of 133 fixed locations for the 19 patient series. Temperature mapping was done in the tumour volume when logistically feasible. Treatment sites included colorectal (n = 3), portahepatus (n = 1), pancreas (n = 7), liver (n = 1), pelvis (n = 3), sacrum (n = 2), and abdomen (n = 2). A sterile, constant-volume water circulating system was utilized to control surface temperatures. Three generations of completely immersible transducers were designed over the course of this study with a 4-cm height specification. Since the ultrasound transducer was assembled on the sterile field during surgery, a 1, 2 or 3 MHz ceramic element was placed in either a 6, 8 or 10 cm diameter aluminium housing to conform the acoustic field to the tumour size. Average of the maximum temperatures attained was 46.6 degrees C. Temperature with which 90% of all measured points equalled or exceeded (T90) was 39.2 degrees C. The T50 was 42.9 degrees C. This compared favourably with T90 and T50 of 38.8 and 41.9 degrees C, respectively, in our outpatient clinic series, in which superficial tumours were treated with a similar external applicator, and patient tolerance was often a treatment limitation.

Potentiation of hyperthermia in a murine tumour by metabolic inhibitors rhodamine 123 and 2-deoxy-D-glucose or 5-thio-D-glucose.

Rhodamine 123 injected into mice on 3 days consecutively before a single hyperthermia treatment (45 degrees C for 15 min) potentiated hyperthermia as evidenced by an increased mean tumour growth delay of a transplantable murine mammary adenocarcinoma (MTG-B). Addition of three daily injections of either 2-deoxy-D-glucose, or 5-thio-D-glucose, coordinated with the rhodamine 123, and administered before hyperthermia, resulted in an additional tumour growth delay, but not large enough to suggest an additional significant interaction between the two drugs and heat. This effect was obtained using doses of the glucose analogues which did not potentiate therapeutically when combined with heat without rhodamine 123. On the third day of treatments, rhodamine 123 or 5-thio-D-glucose, or the two drugs together, 60 min before heating, produced a longer growth delay compared with each combination treatment with drugs administered 15 min before heating. However, this effect was not significant. Results of these experiments suggest that in this murine tumour thermopotentiation can be attained by combining these two classes of metabolic inhibitors with hyperthermia.

Combined modality treatment with ternary Cu(II) complexes and X rays.

Ternary Cu(II) complexes with bidentate malonato- and heterocyclic amine ligands were tested with regard to cytotoxicity and potentiation of x-ray induced cell killing in V79 cells. Two lead complexes were also tested in a tumor assay using the MTG-B murine adenocarcinoma model growing in the flanks of female C3H/HeJ mice. One complex, [2,2'-bipyridyl malonatoCu(II)] (RL-5077), produced sensitizer enhancement ratios (SER's) of 1.8 (hypoxic conditions) and 1.0 (oxic conditions) in vitro when irradiation followed 1 hr exposure to the drug at 100 microM. When RL-5077 was administered at doses of 1/2 (11.65 mg/kg) or 1/4 (5.25 mg/kg) the maximum tolerated dose (MTD), 15 min prior to a locally delivered dose of 20 Gy, enhancement ratios (ER's) of 1.6 and 2, respectively, resulted. The second lead complex, [1,10 phenanthroline (malonato)Cu(II)hydrate] (RL-5027), produced SER's of 1.8 and 1.2 under hypoxic and oxic conditions, respectively, at a concentration of 25 microM. Injection of RL-5027 (5 mg/kg) resulted in toxicity without enhancement in combination with radiation. Analogues of these two complexes have been synthesized in an effort to optimize the potentiation of radiation effects while minimizing toxicity to drug alone and increasing water solubility of the drug. Further studies of the structure-activity relationship of Cu(II) ternary complexes using in vitro radiosensitization as the endpoint have identified four classes of ligands with varying biological activity and have supplied information about the effects of group substitution on solubility, toxicity, and radiation potentiation. This group of complexes represents a new class of radiopotentiators that deserves further investigation into its potential for clinical use.

Experimental brain hyperthermia: techniques for heat delivery and thermometry.

An experimental canine brain model was developed to assess the effects of hyperthermia for a range of time and temperature endpoints, delivered within a specified distance of an interstitial microwave antenna in normal brain. The target temperature location was defined radially at 5.0 or 7.5 mm from the microwave source at the longitudinal location of maximum heating along the antenna in the left cerebral cortex. Temperatures were measured with fiberoptic probes in a coronal plane at this location in an orthogonal catheter at 1.0 mm intervals. Six antennas were evaluated, including dipole, modified dipole, and four shorted helical antennas with coil lengths from 0.5 to 3.9 cm. Antenna performance evaluated in tissue equivalent phantom by adjusting frequency at a fixed insertion depth of 7.8 cm or adjusting insertion depth at 915 MHz showed dipoles to be much more sensitive to insertion depth and frequency change than helical antennas. Specific absorption rate (SAR) was measured in a brain/skull phantom and isoSAR contours were plotted. In vivo temperature studies were also used to evaluate antenna performance in large and small canine brain tissues. A helical antenna with a 2.0 cm coil length driven at 915 MHz was chosen for the beagle experiments because of tip heating characteristics, well-localized heating along the coil length, and heating pattern appropriate to the smaller beagle cranial vault. Verification of lesion dimensions in 3-D was obtained by orthogonal MRI scans and histology to document the desired heat effect, which was to obtain an imagable lesion with well-defined blood-brain-barrier breakdown and necrotic zones. The desired lesion size was between 1.5 to 2.5 cm diameter radially, in the coronal plane with the greatest diameter.

The effect of air cooling on the radial temperature distribution of a single microwave hyperthermia antenna in vivo.

Canine skeletal muscle was heated with a single microwave antenna within a brachytherapy catheter driven at 2000 MHz. The radial, steady-state temperature distribution was measured with and without air cooling of the antenna, as produced by room temperature air flowing in the catheter at 7.5 l/min. The axial temperature distribution was also measured with air cooling. In the antenna junction plane the area heated to a given temperature increased by a factor of four with air cooling when the same antenna temperature was enforced. With the same maximum temperature enforced, the area would increase by a factor of 2.5 with air cooling. The axial temperature distribution was not compromised by air cooling.

Effect of step-down heating on brachytherapy in a murine tumor system.

The effects of step-down heating combined with low-dose-rate irradiation (brachytherapy) were studied using a murine mammary adenocarcinoma (MTG-B) grown in the flanks of C3H mice. Treatment was initiated when tumors reached 0.9 to 1.1 cm in diameter. Step-down heating consisted of 7.5 min at 45 degrees C immediately followed by 7.5 min at 42 degrees C. Step-up heating consisted of 7.5 min at 42 degrees C immediately followed by 7.5 min at 45 degrees C. Step-down heating and step-up heating were compared to a single 45 degrees C, 15-min hyperthermia treatment. These hyperthermia protocols were combined before, in the middle of, or after brachytherapy. There were 4 untreated controls, 6 sham controls, and 11 treated animals in each of the brachytherapy-alone and combined treatment groups. The entire experiment was repeated at brachytherapy doses of 988, 1273, and 1603 cGy. In addition, the effects of step-down heating, step-up heating, and single-temperature hyperthermia were tested alone and in combination with sham treatment for each sequence. Based on daily measurements of tumor diameter, the growth delay to doubling volume was used as the biological end point. To compare the various treatment protocols, an isoeffect thermal enhancement ratio (TERiso) was calculated. Step-down heating after 988 cGy brachytherapy had a TERiso of 2.0 +/- 0.04, while step-up heating after 988 cGy brachytherapy had a TERiso of 1.7 +/- 0.05. Overall, the thermal enhancement ratios calculated from these growth delays indicate that step-down heating caused significantly greater hyperthermic radiosensitization than step-up heating when combined with brachytherapy.

The effect of in vivo GSH depletion on thermosensitivity, radiosensitivity and thermal radiosensitization.

The effect of glutathione (GSH) depletion on the interaction of hyperthermia (HT) and single-dose external-beam radiation delivered at conventional dose rates (RT) and low-dose-rate brachytherapy (BT) was examined in a murine mammary adenocarcinoma (MTG-B). Tumour volume growth delay from the day of treatment to double the treatment volume (GDDV) was used as the experimental end-point. The growth delay for radiation alone was greater when combined with GSH depletion, while this level of GSH depletion produced no effect on the BT alone or HT alone response. The synergism ratio (SR) was calculated for all HT and RT or HT and BT protocols at each dose. Although depletion did not increase the SR for external-beam treatments, the SR for HT and BT treatments was increased with GSH depletion.

Intraoperative radiation therapy and hyperthermia. Morbidity and mortality from this combined treatment modality for unresectable intra-abdominal carcinomas.

This is a report of a phase I trial of intraoperative radiation therapy in combination with intraoperative hyperthermia for the treatment of locally advanced, unresectable intra-abdominal carcinomas. Using an ultrasound transducer system specifically designed for intraoperative applications, 19 patients have been successfully treated, demonstrating the feasibility of this combination modality. The morbidity (58%) and mortality (11%) rates reported in this series are comparable to rates reported in series of similar patients receiving intraoperative radiation therapy alone. There is still a great need for considerable technological development to enable the use of intraoperative hyperthermia to treat large, complexly shaped intra-abdominal tumors, and phase II and III trials of this combination treatment modality should be performed.

Thermal enhancement of low dose rate irradiation in a murine tumour system.

The effects of localized hyperthermia (HT) in combination with low dose rate irradiation (brachytherapy) have been investigated in vivo using a murine mammary adenocarcinoma. Flank tumours were grown to 0.45-0.70 cm3 in volume, at which time their treatment course was initiated. Tumours were locally heated in a water bath for 15 min at either 44 or 45 degrees C. For tumour irradiations a non-invasive cap was devised to permanently house three iodine-125 sealed sources located at 120 degree intervals around the circumference of the hemispherical cap. During treatment, mice were secured in a modified syringe tube allowing mobility while restricting access to the cap which was placed over the tumour. Calculated dose rates ranged from 15 to 40 cGy/h. Brachytherapy (BT) was delivered for 48 or 72 h to obtain a dose range of 830-2378 cGy. Mice were randomized into one of 10 treatment protocols: BT alone, HT-BT, BT-HT, HT-BT-HT, 1/2BT-HT-1/2BT, four control groups of HT alone and a sham treatment group. Normalized tumour doubling volume growth delays (GDDv) were used to calculate the thermal enhancement ratios (TER). In the 44 degrees C experiments, HT before BT (TER = 1.33 +/- 0.071) was more efficacious than HT after BT (TER = 1.07 +/- 0.042). Two HT treatments, one given before and one after BT (TER = 1.38 +/- 0.152), were not different from a single HT treatment given before BT. However, a single HT treatment given in the middle of an interrupted course of BT resulted in the greatest thermal enhancement (TER = 1.64 +/- 0.072) compared to any other treatment sequence. These data suggest that potentiation of low dose rate irradiation by a single heat treatment may be maximized if the HT is given either in the middle of, or simultaneously with, the BT.