Thermal Enhancement Ratio Research Articles

Concurrent ferromagnetic hyperthermia and 125I brachytherapy in a rabbit choroidal melanoma model.

Ferromagnetic (FM) thermoseeds and radioactive (125I) seeds were combined in an episcleral plaque to give concurrent hyperthermia and irradiation for enhanced tumour destruction. A Greene melanoma cell line was utilized to study the interaction between these treatment modalities. We attached five FM thermoseeds (with an operating temperature of 48 degrees C) in parallel with alternating rows of 125I seeds onto the inner surface of each 14 mm Silastic plaque. Plaques were centred over a 3-6 mm (diameter) intraocular melanoma in each rabbit. Some rabbits were then placed within a heating coil, and their eye tumours were warmed rapidly to therapeutic temperatures (43.6 degrees C across the tumour base) while the temperature of normal conjunctiva across the globe did not exceed 38.5 degrees C. Analysis of 49 treated eye melanomas showed 50% local tumour control at 41.7 Gy for 125I alone, whereas only 9.5 Gy were needed to give the same local control rate after 125I with concurrent FM hyperthermia. Thus, a thermal enhancement ratio of 4.4 was obtained. Hyperthermia alone gave a 20% tumour response rate, but responses were only temporary. We conclude that FM thermoseeds can be used to deliver biologically effective hyperthermia concurrently with radiation, thereby reducing the dose of radiation needed for tumour control.

Significance of additive heat effect in the therapeutic gain factor in combined hyperthermia and radiotherapy: Murine tumor response and foot reaction

The thermal enhancement ratio (TER) and therapeutic gain factor (TGF) were evaluated for combined hyperthermia and radiation treatments of a murine fibrosarcoma, FSa-II. The TER is the ratio of the radiation dose that induces a given reaction without hyperthermia to that with hyperthermia. The TGF is defined as the ratio of TER for tumor response to TER for normal tissue response. Tumors in the subcutaneous tissue of the right foot were irradiated with graded radiation doses when they reached an average diameter of 6 mm (110 mm). Hyperthermia was given by immersing animal feet in a constant temperature water bath 10 min before or after irradiation. The tumor growth time to reach 500 mm 3 was obtained for each tumor and the median tumor growth time was calculated for each treatment group. For the normal tissue study, the non-tumor bearing murine foot was treated, as was the tumor, and the foot reaction was scored after treatment, according to our numerical score system for radiation damage, until the 35th post-treatment day and averaged. Using the fraction of animals showing a given average foot reaction score in a treatment group, the RD 50, or the radiation dose to induce the given foot reaction or greater, was calculated. A single heating at 45.5°C for 10 min and a step-down heating (first heat at 45.5°C for 10 min immediately followed by the second heat at 41.5°C for 60 min) prolonged the tumor growth time, indicating that hyperthermia per se resulted in some cell killing. The prolongation was greater following step-down heating than following single heating. These heat treatments alone induced no noticeable heat damage on the foot, but decreased the threshold dose observed on the radiation dose response curves for the foot reaction. Accordingly, TER and TGF were evaluated with or without normalizing this thermal effect. TER's for both tumor and foot responses without normalization were greater than the TER's after normalization and decreased with increasing radiation dose (between 1.9 and 7.1 or greater for tumor and between 1.3 and 4.3 or greater for foot reaction), whereas the normalized TER's were relatively constant (between 1.6 and 1.7 for tumor and between 0.7 and 1.5 for foot reaction). TGF's without normalization were greater than those obtained after normalization. The former was large at small doses and decreased with increasing radiation dose (between 1.5 and 4.0 or greater), whereas the latter was within 0.8 and 1.3 and relatively independent of radiation dose. TGF's following step-down heating were greater than the TGF following single heating (> 3.0 and between 1.5 and 2.0 for SDH and single heating given before radiation). This greater non-normalized TGF is likely due to the additive effect of hyperthermia, that is, due to the greater thermal effect on the tumor than the normal foot. Similarly, greater TGF's following step-down heating may be due to greater damage, induced by this heating, to the tumor. TGF following hyperthermia given before or after radiation appeared to depend on the heat dose applied.

Fractionated thermochemotherapy in vivo of a C3H mouse mammary carcinoma

The interaction between fractionated heat treatment and fractionated drug treatment with cyclophosphamide (CTX) was investigated in a transplantable C3H mouse mammary carcinoma inoculated into the hind leg of C3D2F1/Bom mice. A tumour core temperature of 43.5 ± 0.1 °C was achieved by immersing the tumour-bearing leg into a water bath thermostatically maintained at 43.7 ± 0.1 °C. CTX was administered i.p. using the maximum tolerated dose (MTD) (LD 1 %) for single fraction treatment (100 mg/kg) as the maximum fraction dose. For combined treatment CTX was given 15 min prior to heating. The endpoint was the time to reach a tumour volume of 5 times the volume at first treatment. Specific growth delay was used as effect parameter. In dose-effect experiments using total treatment time at 43.5 C as dose parameter, drug enhancement ratio (DER) was determined as the ratio of the slope of the dose-effect curve for MTD of CTX plus heat to the slope of the curve for heat alone. In dose-effect experiments using total CTX dose as dose parameter, thermal enhancement ratio (TER) was determined as the ratio of the slope of the dose-effect curve for CTX plus 43.5 ° C for 30 min to the slope of the curve for CTX alone. The regimens investigated were single fraction treatment and 3 and 5 fractions with time intervals of 3 and 5 days. For single fraction treatment DER was 1.4 ± 0.1 and TER 2.3 ± 0.2. The drug sensitization of the effect of heat treatment tended to increase with increasing number of fractions. The thermal sensitization of the effect of drug treatment was about the same for fractionated and single fraction treatment. Both the drug sensitization and the thermal sensitization were larger for a treatment interval of 3 days than of 5 days, i.e. largest with the fraction interval at which the thermotolerance is known to be largest in this tumour. The data indicate a potential for improved therapeutic ratio by fractionation of thermochemotherapy.

The response of human and rodent cells to hyperthermia

Inherent cellular radiosensitivity in vitro has been shown to be a good predictor of human tumor response in vivo. In contrast, the importance of the intrinsic thermosensitivity of normal and neoplastic human cells as a factor in the responsiveness of human tumors to adjuvant hyperthermia has never been analyzed systematically. A comparison of thermal sensitivity and thermo-radiosensitization in four rodent and eight human-derived cell lines was made in vitro. Arrhenius plots indicated that the rodent cells were more sensitive to heat killing than the human, and the break-point was 0.5°C higher for the human than rodent cells. The relationship between thermal sensitivity and the interaction of heat with X rays at low doses was documented by thermal enhancement ratios (TER's). Cells received either a 1 hr exposure to 43°C or a 20 minute treatment at 45°C before exposure to 300 kVp X rays. Thermal enhancement ratios ranged from 1.0 to 2.7 for human cells heated at 43°C and from 2.1 to 5.3 for heat exposures at 45°C. Thermal enhancement ratios for rodent cells were generally 2 to 3 times higher than for human cells, because of the fact that the greater thermosensitivity of rodent cells results in a greater enhancement of radiation damage. Intrinsic thermosensitivity of human cells has relevance to the concept of thermal dose; intrinsic thermo-radiosensitization of a range of different tumor cells is useful in documenting the interactive effects of radiation combined with heat.

Enhancement of the thermal response of murine tumour and normal tissues by a streptococcal preparation, OK-432 (Picibanil)

We have studied the effect of a streptococcal preparation, OK-432, given alone or in combination with hyperthermia on murine tumour and normal tissues. The experimental tumour was a spontaneous non-immunogenic fibrosarcoma FSa-II transplanted in the foot of C3Hf/Sed mice. Local hyperthermia was achieved by immersing the mouse foot into a constant temperature water bath (42-45 degrees C) for various lengths of time. Tumour response was studied by analysing the tumour growth (TG) time. Various doses of OK-432 (1-5 KE/mouse) were injected locally into the tumour. Local administration of OK-432 alone (without hyperthermia) had no effect on the TG time. Thermal enhancement ratio (TER) for combined treatment was independent of drug dose greater than or equal to 2 KE, and the mean TER was 1.48 +/- 0.27 (95% CL). The TER was greater for 6 mm tumours than for 4 mm tumours, and it was greatest if the time interval between hyperthermia and drug administration was 3 h or less. There was no effect if the drug was administrated 4 days before hyperthermia, but its application 9 days prior to hyperthermia caused a slight prolongation of the TG time of non-heated tumours, and a reduction in the TG time of heated tumours. Normal-tissue response was studied by scoring the peak foot reaction and RD50 (the treatment time to induce a specified response in one-half of the treated animals). The effect of OK-432 on the thermal response of the foot was also studied at various temperatures. The mean TER was 1.11 +/- 0.07. Local administration of OK-432 failed to modify significantly the kinetics of thermotolerance. Present experiments demonstrated that OK-432 after local administration enhanced the thermal response of murine tumour and normal tissues. This enhancement was greater for the tumour than for the normal tissue, resulting in a favourable differential effect between normal and malignant tissues (the average therapeutic gain was 1.33 +/- 0.19).

Effect of Step-down Heating on the Interaction between Heat and Radiation in a C3H Mammary Carcinomain Vivo

The effect of step-down heating (SDH) on the interaction between heat and radiation was investigated in a C3H mammary carcinoma in vivo. SDH consisted of an initial sensitizing treatment (ST) performed at 44.5 degrees C or 43.5 degrees C followed by a lower temperature test treatment (TT) in the range 41.0-43.0 degrees C. Step-up heating (SUH), i.e. TT followed by ST, and single heating were used as controls. The end-point was the radiation dose needed to control 50% of the tumours (TCD50). The results were evaluated by calculating the thermal enhancement ratio (TER) defined as TER = TCD50 (radiation alone)/TCD50 (radiation and heat). For a simultaneous application of TT and radiation a significant enhancement of direct heat radiosensitization was observed with increasing ST time or ST temperature using SDH. In contrast, only a minor increase was seen with SUH. A comparison between TCD50 values for the corresponding SUH and SDH schedules revealed that the SDH effect was largest at 41.0-42.0 degrees C and decreased with increasing TT temperature. The radiosensitizing effect of SDH also decreased if an interval was allowed between ST and TT or between TT and radiation. However, as a result of an increased cytotoxicity towards hypoxic tumour cells, the TCD50 value for SDH remained significantly smaller than for SUH, even with a sequential combination of radiation and heat.

Augmentation of mouse natural killer cell activity by combined hyperthermia andstreptococcalpreparation, OK-432 (Picibanil) treatment

It has previously been demonstrated that the local administration of OK-432 (Picibanil) enhances the response of a murine tumour and normal tissue to elevated temperatures. The experimental animals were C3Hf/Sed mice and the tumours were the fourth generation isotransplants of a spontaneous non-immunogenic fibrosarcoma, FSa-II. The thermal enhancement ratio was greater for the tumour than for normal tissue, resulting in a favourable differential effect between normal and malignant tissues. Further studies were conducted to disclose the mechanism of OK-432 induced thermal enhancement. Although OK-432 showed a slight direct cytotoxic activity against tumour cells in vitro, the in vivo antitumour effect of combined OK-432 and hyperthermia treatments was greater than the effect expected from in vitro cytotoxicity, indicating the involvement of the host-mediated mechanisms. Whole body irradiation (WBI), which suppressed the host's immune reaction, did not affect the thermal enhancement mediated by OK-432, suggesting that radiosensitive cells (sensitive to WBI) were not involved in this process. As expected, the i.v. injections of the anti-mouse T-cell serum did not have any effect on the thermal enhancement of OK-432. The administration of blockers of the reticuloendothelium system, i.e. silica or trypan blue, did not inhibit the OK-432 induced thermal enhancement. Moreover, local intratumoural injections of normal or OK-432 induced macrophages showed no effect on tumour growth. The thermal enhancement by OK-432 was eliminated following treatments with anti-(asialo GM1) globulin. Despite the fact that the cytotoxic effect of anti-(asialo GM1) globulin was not selective to natural killer (NK) cells, all the experimental results indicated that NK cells were probably attributable to thermal enhancement by OK-432 in vivo. The NK cell activity was stimulated when the OK-432 was locally administered with hyperthermia.

Enhancement by hyperthermia of the 'early delayed' and 'late delayed' radiation response of the rat cervical spinal cord.

The cervical spinal cord (C5-T5) of female Wistar WU rats was irradiated with 250 kV X-rays (15-32 Gy). Heat was applied at approximately the same site 7 +/- 1 min after X-rays. 'Early delayed' paralysis of the forelegs was observed 5-10 months after treatment. The ED50 (+/- SE) after single-dose irradiation alone was 25.8 +/- 0.4 Gy. 'Late delayed' paralysis and paresis were observed 11-21 months after irradiation with an ED50 (X-rays alone) of 22.7 +/- 0.6 Gy. The data for late paralysis, late paresis and minor neurological symptoms were pooled resulting in an ED50 (+/- SE) of 20.6 +/- 0.7 Gy. Hyperthermia enhanced the radiation response. Thermal enhancement ratios (TER) in the 'early delayed' response after a 30 min treatment with 41.1 +/- 0.4 degrees C 42.1 +/- 0.4 degrees C and 42.9 +/- 0.4 degrees C were 1.07 +/- 0.08, 1.17 +/- 0.08 and 1.12 +/- 0.04, respectively. For the 'late delayed' radiation response concerning paralysis and paresis the TER after 30 min at 41.1 degrees C and 42.1 degrees C were 1.25 +/- 0.10 and 1.31 +/- 0.07, respectively. The latent period for paralysis was not significantly affected. Pathological examination of the spinal cord after combined treatment of X-rays and hyperthermia showed focal demyelination with white matter necrosis and vascular injury in animals as an indication of 'early delayed' and 'late delayed' paralysis, respectively. This was not different from histopathological changes observed after irradiation alone.

Optimisation of intraperitoneal cisplatin therapy with regional hyperthermia in rats

The purpose of this study was to optimise intraperitoneal chemotherapy by combining this modality with regional hyperthermia. In vitro data demonstrated that both the uptake of cisplatin into CC531 tumour cells and cytotoxicity were increased at temperatures of 40°C (factor 4) and 43°C (factor 6) compared to 37°C. The increase of intracellular platinum concentration correlated well with the decrease in survival of these cells. In vivo, rats were treated intraperitoneally with cisplatin (5 mg/kg) in combination with regional hyperthermia of the abdomen (41.5°C, 1 h). The mean (S.D.) temperature in the peritoneal cavity was 41.5 (0.3)°C and outside the peritoneal cavity 40.5 (0.3)°C. Enhanced platinum concentrations were found in peritoneal tumours (factor 4.1) and kidney, liver, spleen and lung (all around a factor 2.0), after combined cisplatin-hyperthermia treatment. The platinum distribution in peritoneal tumours was more homogeneous after the combined treatment than after cisplatin alone, possibly due to increased penetration of cisplatin into peritoneal tumours. Pharmacokinetic data demonstrated an increased tumour exposure for unfiltered platinum in the peritoneal cavity (area under the curve [AUC] increased from 339 μmol/l/min to 486 μmol/l/min at 37°C and 41.5°C, respectively), and for total and ultrafiltered platinum in the blood. The AUC for total platinum increased from 97.9 to 325.8 μmol/min and for ultrafiltered platinum from 22.2 to 107 μmol/l/min at 37°C and 41.5°C respectively. The latter might be due to a slower elimination of platinum from the blood. The combined treatment, intraperitoneal cisplatin and regional hyperthermia, also increased toxicity. The thermal enhancement ratio (TER) using lethality as endpoint was 1.8.

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.

Thermal enhancement of radiation-induced leg contracture

Early and late damage in the normal tissues of the legs of mice was compared following treatment with radiation alone or radiation followed immediately by hyperthermia. Hyperthermia was given by immersing the hind leg in a water bath at 43.0°, 43.3°, or 43.5°C for 1 hr. Damage was assayed by measuring leg contracture at various intervals from 5 to 365 days after treatment. At 5 days after treatment, only hyperthermia-induced contracture was observed. At 10 and 20 days, contracture increased with radiation dose in heated legs, but little contracture had developed in mice treated with radiation alone. By 45 through 365 days, however, contracture correlated with radiation dose both in mice treated with radiation alone as well as in those treated with radiation and hyperthermia. The greatest differential in the slopes of the dose response curves, suggesting hyperthermic radiosensitization, was seen 20 days after treatment. Nevertheless, at 365 days, contracture was still significantly greater in the mice treated with radiation and hyperthermia (43.5° bath) than in the irradiated controls. Thermal enhancement ratios (TERs) were calculated from LCD50 values (LCD50 = radiation dose that would give a stated level of leg contracture in 50% of the mice). For ≥3 mm contracture, TERs were 4.1 to 7.9 at 30 days, depending on bath temperature, but only 1.1 to 1.5 at 365 days. For an isoeffect of ≥ mm contracture, TERs were 1.9 to 5.3 at 30 days, and 0.8 to 1.8 at 365 days. Thus, contracture was enhanced more at 20 to 30 days after treatment with radiation and hyperthermia than at 120 through 365 days. Radiation damage not only appeared earlier in mice treated with hyperthermia than in those treated with radiation alone, but after the highest temperature tested (43.5° bath), contracture was greater from 5 through 365 days after treatment than in controls treated with radiation alone.

Thermal enhancement of tetraplatin and carboplatin in human leukaemic cells

The thermal enhancement of tetraplatin (racemic, d-trans and l-trans isomers) and carboplatin was studied as a function of temperature in vitro in JM, a human acute lymphoblastic leukaemia cell line. Exponentially growing JM cells were exposed to tetraplatin (0-10 micrograms/ml) or carboplatin (0-45 micrograms/ml) for 1 h at 37-43 degrees C in 1 degree C increments. Graphic analysis demonstrated a breakpoint for the onset of thermal enhancement for tetraplatin at 40 degrees C. Thermal enhancement was maximal at 42 degrees C with no significant increase at 43 degrees C. The tetraplatin thermal enhancement ratio (TER) was 2.7 at 42 degrees C. The TERs for d and l isomers at 41.8 degrees C were not significantly different. The relationship of TER to temperature for carboplatin closely paralleled that of tetraplatin. The implications of these results are discussed in the context of tetraplatin's unique properties.

A comparison of thermal enhancement of cis-diamminedichloro-platinum (II) induced renal and intestinal toxicities by whole body hyperthermia in the rat

Thermal enhancement of cis-diamminedichloroplatinum (II) (DDP) induced renal and intestinal toxicities by whole body hyperthermia (WBH) were compared using a F344 rat model. Thermal enhancement ratios (TER) for DDP-induced nephrotoxicity were calculated using renal functional assays and morphological techniques. TER values for gastrointestinal (G.I.) toxicity were calculated using “severity of diarrhea” and jejunal crypt cell survival as assays. TER's for renal damage varied between 3 and 3.4, whereas the TER measured for G.I.-toxicity was 1.8. Physiological changes caused by WBH or intrinsic differences in the sensitivities of normal tissues to DDP ± WBH may be responsible for the differences in thermal enhancement of DDP-induced renal and intestinal toxicities.

Tumoricidal interactions of hyperthermia with carboplatin, cisplatin and etoposide

Interactions of hyperthermia with carboplatin, cisplatin and etoposide were investigated in vitro in JM, a human, T cell, lymphoblastic cell line. Thermal enhancement ratios (TER) for carboplatin killing increased with temperature (2.3 at 40.5°C, 3.2 at 41.8°C) and were similar to those for cisplatin killing (2.6 at 40.5°C, 3.6 at 41.8°C). In a separate experiment, cytotoxicity was additive when carboplatin and cisplatin were given simultaneously at 37°C and at 41.8°C. Etoposide, which synergizes with platinum agents, did not have supra-additive cytotoxic interactions with hyperthermia. The data presented are relevant to the conduct of clinical hyperthermia trials.

The current and potential role of hyperthermia in radiotherapy

Current clinical experience strongly suggests that hyperthermia will become an important modality as an adjuvant to radiotherapy in the treatment of locally advanced solid tumors. Hyperthermia must therefore be considered a topic of general interest. Biologically, hyperthermia has two different types of interactions with radiation. Firstly, heat has a radiosensitizing effect. This is most prominent with simultaneous application, but is of the same magnitude in both tumor and normal tissue and will not improve the therapeutic ratio unless the tumor is heated to a higher temperature than the normal tissue. Secondly, hyperthermia exhibits a direct cytotoxic effect, and a moderate heat treatment alone can almost selectively destroy tumor cells in a nutritionally deprived chronically hypoxic and acidic environment. Because such cells are the most radioresistant, a smaller radiation dose is needed to control the remaining more radiosensitive cells. If critical, irradiated normal tissues are also heated, the cytotoxicity is best utilised if heat is given at least 3–4 hours after irradiation. The magnitude of both the sensitizing and the cytotoxic effect depends on temperature and heating time. Clinically, heating of superficial tumors (e.g. breast, neck nodes and malignant melanoma) has confirmed the biological rationale for using hyperthermia as an adjuvant to radiotherapy. An overview of available data gives thermal enhancement ratios of approximately 1.5 in several superficial tumor sites after external heating. From a practical point of view, true simultaneous treatment is almost impossible using external heating, and the major effect of the combined treatment will have to rely on hyperthermic cytotoxicity. This makes the design of clinical schedules less complicated since only a few heat fractions may be needed to achieve an optimal effect. On this basis, several randomized clinical trials have been activated with the aim to evaluate the role of adjuvant hyperthermia in the primary treatment of advanced (superficial) tumors. In addition, studies are underway to specifically elucidate the clinical relevance of thermotolerance and other biological issues. So far, the clinical evaluation has almost solely been limited to superficial tumors, or to situations where interstitial heating is feasible. External heating of “deep” seated tumors is still preliminary, and most studies are in Phase I–II, with emphasis on toxicity and feasibility. The initial results are promising with regard to improved tumor control and acceptable toxicity.

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.

Thermochemotherapyin vivoof a C3H mouse mammary carcinoma: Thermotolerant tumours

The ability of cyclophosphamide (CTX) and mitomycin C (MMC) to modify the expression of thermotolerance in vivo at 43.5 degrees C was investigated in a transplantable C3H mouse mammary carcinoma grown s.c. in feet of C3D2F1/Bom mice. Dose-effect curves subjected to linear regression analysis were constructed for single-fraction treatment and for a second treatment 24 h after a priming heat treatment of 43.5 degrees C for 30 min. Tumour volume doubling time during regrowth showed no significant variation among treatment groups, justifying the use of tumour growth time as effect parameter. Thermotolerance ratio for heat alone was 11.6 +/- 2.3. Drug enhancement ratio in thermotolerant tumours was 6.2 +/- 1.4 for CTX (100 mg/kg) and 3.5 +/- 0.9 for MMC (3 mg/kg). These values are about 4 and 3 times larger than the corresponding enhancement ratios found for previously untreated tumours. Thermotolerance ratio for thermochemotherapy was 2.6 +/- 0.3 for CTX and 4.4 +/- 0.7 for MMC, i.e. the degree of thermotolerance was substantially reduced by both drugs, but not completely overcome. Thermal enhancement ratio for CTX and MMC was about equally large in thermotolerant and previously untreated tumours. Thermotolerant tumours showed a tendency for increased resistance to drug treatment alone. Both CTX and MMC may be used clinically to reduce the expression of thermotolerance, particularly in situations with inhomogeneous heating and short fractionation intervals.

Thermochemotherapy in vivo of a C3H mouse mammary carcinoma: Single fraction heat and drug treatment

The interaction between water bath hyperthermia (43.5°C) and six cancer chemotherapeutic agents in vivo was studied in a transplantable C3H mouse mammary carcinoma grown s.c. in the feet of C3D2F1/Bom mice. Due to differences in tumour regrowth rate between treatment groups, both tumour growth time (TGT) and specific growth delay (SGD) were used as effect parameters. The largest tumour response was observed when the drug was given 15 min prior to heat—this timing was used for dose-effect experiments. Enhancement ratios were the ratios of slopes of dose-effect curves subjected to linear regression analysis. The drug enhancement ratio (DER) was not significantly larger than 1.0 for LD 1% of adriamycin, 5-fluorouracil, methotrexate and vincristine. For cyclophosphamide (CTX) and mitomycin C (MMC) both DER and TER (thermal enhancement ratio) were significantly larger than 1.0. The TGT ratios (SGD ratios in parentheses) were: DER (LD 1%): CTX 1.4 ± 0.1 (2.1 ± 0.1), MMC 1.3 ± 0.1 (1.4 ± 0.1); TER (43.5°C 30 min): CTX 1.6 ± 0.1 (2.7 ± 0.2), MMC 2.8 ± 0.5 (3.3 ± 0.7). The data support the choice of CTX and MMC in preference to the other drugs investigated for clinical thermochemotherapy studies.

The Role of Low Intracellular or Extracellular pH in Sensitization to Hyperthermic Radiosensitization

Previous work showed that intracellular pH (pHi) and not extracellular pH (pHe) was the determinant in the low pH sensitization of hyperthermic killing. The present studies show that the same is true for heat-induced radiosensitization and loss of cellular DNA polymerase activities. Chinese hamster ovary cells after they had adapted to low pH (6.7) had an increase in pHi which rendered cells partially resistant to the low pH sensitization of heat-induced cell killing, radiosensitization, and loss of cellular DNA polymerase activities. These results were quantified by plotting versus pHe, both the thermal enhancement ratio (TER), defined as the ratio of the X-ray dose without heat to the X-ray dose with heat to give an isosurvival value of 0.01, and the thermal enhancement factor (TEF), defined as the ratio of the D0 of the radiation survival curve to the D0 of the radiation survival curve for heat plus radiation. Both the TER and TEF were higher for the unadapted cells than for the adapted cells, i.e., 1.3-1.4 fold higher at a pHe of 6.3. However, the TER or TEF plotted versus pHi was identical for the two cell types. Finally, heat-induced loss of cellular DNA polymerase activities correlated with pHi and not pHe. Therefore, we conclude that pHi and not pHe is responsible for the increase by acid in heat-induced radio-sensitization and loss of cellular DNA polymerase activities.

The effect of hyperthermia on the early and late appearing mouse foot reactions and on the radiation carcinogenesis: Effect on the early and late appearing reactions

The effect of hyperthermia on radiation-induced early- and late-appearing foot reactions was studied in C3Hf/Sed mice derived from our defined flora mouse colony. The animal foot was irradiated with 137Cs gamma-rays under hypoxic, air, or hyperbaric oxygen (O 2 30 psi) conditions. Hyperthermia of 43.5°C for 45 min was given locally in a water bath where a constant temperature ± 0.1°C was maintained. Treatment intervals between the 2 treatments were 20 min and 2 days. For the early-appearing reactions scores taken between the 14th and 35th post-irradiation days were averaged. Late-appearing reactions became apparent after approximately the 200th post-treatment day and increased with time. The foot reaction was enhanced by hyperthermia given 20 min before or after irradiation. Dose response curves for radiation given 20 min after hyperthermia for acute-appearing reactions lacked shoulders, whereas those following the same treatment schedule for late-appearing reactions showed significant shoulders. The thermal enhancement ratios (TER) for score 2.0 (complete epilation) early- and late-appearing reactions depended on the treatment interval and sequence. The TER values were greater for a short treatment interval (20 min.) than for a long treatment interval (2 days). Thermal enhancement was greater for hyperthermia given before irradiation compared to the reverse sequence. The TER values were always smaller for the late-appearing reactions than for the acute-appearing reactions. The relationships between early reaction scores and late reaction scores showed that the late reactions following combined heat and radiation are less extensive than those following radiation alone if they were compared at radiation doses which induced an equal level of early reactions. This difference was most significant at low early reaction scores and decreased with increasing score level.