Enhancement of sensitivity to hyperthermia by lonidamine in human cancer cells

Human glioma (87MG) and squamous cell carcinoma of the head and neck UMSCC-1 were shown to be sensitized to hyperthermia by Lonidamine treatment before and during hyperthermia. The degree of thermal sensitization increased with increasing heating times and temperatures. In addition, the thermal sensitization by Lonidamine as well as cellular thermal sensitivity were dependent on pH and increased with the more acidic pH. Even though plateau phase cells were more thermally resistant than exponentially growing cells, Lonidamine treatment caused thermal sensitization under both conditions. These data show that Lonidamine may hold potential to enhance the effectiveness of hyperthermia in cancer treatment and that especially in tumours with low pH an enhanced therapeutic gain may be achieved.     

Lonidamine can enhance the cytotoxic effect of cisplatin in human tumour cells and rodent cells

Lonidamine combined with cisplatin treatment was tested in human glioma (U87MG) human squamous cell carcinoma (UMSCCl) and Chinese hamster (HA1) cells. The order of sensitivity to cisplatin was HA1 greater than U87MG greater than UMSCCl. Lonidamine caused little sensitization in U87MG cells, moderate sensitization in HA1 cells and large sensitization in UMSCCl cells. The degree of sensitization was correlated to the effect of lonidamine on cell growth. The degree of sensitization to cisplatin by lonidamine treatment was dependent on lonidamine exposure concentration, time and treatment sequence. Lonidamine shows potential for enhancement of cisplatin chemotherapy but this appears to be cell type specific and should be tested for cells derived from sites of clinical interest for such drug therapy.     

The effect of lonidamine (LND) on radiation and thermal responses of human and rodent cell lines

Rodent and human cells were tested for response to Lonidamine (LND) (1-(2,4 dichlorobenzyl) 1-indazol-3-carboxylic acid) combined with radiation or hyperthermia. Lonidamine exposure before, during, and after irradiation caused varying degrees of inhibition of potentially lethal damage (PLD) repair which was cell line dependent. In human glioma, melanoma, squamous cell carcinoma, and fibroblasts, LND exposure did not inhibit or only partially inhibited repair of potentially lethal damage. LND up to 100 micrograms/ml produced only a low level of toxicity in these cells and only slightly inhibited glucose consumption at the maximum concentration. In human glioma cells, LND treatment alone did not inhibit PLD repair, but when combined with hyperthermia treatment at moderate levels easily achievable in the clinic, there was complete inhibition of potentially lethal damage repair. These data suggest that LND effectiveness is cell type dependent. Combinations of LND, hyperthermia, and radiation may be effective in cancer therapy especially in tumors such as glioma in which repair of potentially lethal damage may be extensive.     

Effects of Tumor Necrosis Factor and Hyperthermia on Meth-A Tumors

The combined effects of purified human natural tumor necrosis factor (TNF) and hyperthermia were investigated in a transplanted TNF-sensitive Meth-A tumor model. We assessed the sequence and interval for the two treatments, the temperature that caused maximal heat sensitization, and the effects of pH modification on this combination therapy. Tumor response was evaluated by means of a tumor growth delay assay. TNF at a dose of 50 JRU/g caused significant tumor growth delay. A synergistic effect of TNF and hyperthermia was observed when TNF was administered 10 min before heating. This thermal enhancement of the action of TNF became more prominent with an increase in the heating temperature. Tumor growth delay was maximal when TNF was given immediately before or after hyperthermia. However, after an increase in the time interval to more than 2 h, there was no enhancement of growth delay. Injection of glucose (5 g/kg) caused a significant fall in pH at 10 and 30 min after administration. Further enhancement in tumor growth delay was seen with the tri-modality of glucose, TNF, and heat compared to combined treatment with heat and either TNF or glucose at a hyperthermia of 42°C. This effect was not obtained with heating to 40°C. TNF appears to be a potent heat sensitizer when an appropriate temperature and time interval between hyperthermia and TNF administration are used. Trimodality treatment with hyperthermia, TNF, and glucose may be a new method of anticancer therapy.     

A comparison of hyperthermia cisplatin sensitization in human ovarian carcinoma and glioma cell lines sensitive and resistant to cisplatin treatment

 Two pairs of human tumor cell lines (glioma and ovarian carcinoma (OvCa)) each having a parental cell line and cisplatin-resistant variant, were evaluated for (a) cisplatin response, (b) hyperthermia response, and (c) combined hyperthermia and cisplatin response. The two resistant lines had comparable resistant responses while for the parental lines, the OvCa was more sensitive than the glioma to cisplatin doses up to 14 μg/ml. For the hyperthermia response, the OvCa parental line was more resistant than the variant line at low-temperature hyperthermia (41° C or 42°C) but became more sensitive at high temperature (45°C). For the glioma, the parental line was more sensitive to hyperthermia at all temperatures tested. Hyperthermia caused sensitization to cisplatin in all cell lines but was generally greater in the glioma cell lines. In the OvCa system, hyperthermia had a slightly greater sensitizing effect on the resistant cell lines, while in the glioma the opposite was true. The degree of sensitization increased with hyperthermia temperature. In summary, the results showed that there is no cross-resistance for hyperthermia and cisplatin, that the degree of thermal sensitization is not reduced in cisplatin-resistant cell lines, and that cisplatin thermal sensitization is cell-line and temperature dependent. Thus, hyperthermia can effectively improve tumor cell response to cisplatin and may be useful in overcoming resistance to cisplatin. Received: 10 March 1995/Accepted: 21 July 1995     

Effects of tumor necrosis factor and hyperthermia on Meth-A tumors

The combined effects of purified human natural tumor necrosis factor (TNF) and hyperthermia were investigated in a transplanted TNF-sensitive Meth-A tumor model. We assessed the sequence and interval for the two treatments, the temperature that caused maximal heat sensitization, and the effects of pH modification on this combination therapy. Tumor response was evaluated by means of a tumor growth delay assay. TNF at a dose of 50 JRU/g caused significant tumor growth delay. A synergistic effect of TNF and hyperthermia was observed when TNF was administered 10 min before heating. This thermal enhancement of the action of TNF became more prominent with an increase in the heating temperature. Tumor growth delay was maximal when TNF was given immediately before or after hyperthermia. However, after an increase in the time interval to more than 2 h, there was no enhancement of growth delay. Injection of glucose (5 g/kg) caused a significant fall in pH at 10 and 30 min after administration. Further enhancement in tumor growth delay was seen with the trimodality of glucose, TNF, and heat compared to combined treatment with heat and either TNF or glucose at a hyperthermia of 42 degrees C. This effect was not obtained with heating to 40 degrees C. TNF appears to be a potent heat sensitizer when an appropriate temperature and time interval between hyperthermia and TNF administration are used. Trimodality treatment with hyperthermia, TNF, and glucose may be a new method of anticancer therapy.     

Hyperthermia enhancement of radiation response and inhibition of recovery from radiation damage in human glioma cells

Three human glioma cell lines were tested for the effectiveness of hyperthermia and thermal radiosensitization. Thermal sensitization was evaluated from the perspective of increased radiosensitivity as well as inhibition of recovery from radiation damage. The three glioma cell lines tested showed large shoulders on the radiation survival curve and a large capacity for recovery of potentially lethal radiation damage. Hyperthermia caused radiosensitization in all three cell lines, which was primarily characterized by the reduction of the survival curve shoulder with moderate decreases in the survival curve slope. The radiosensitization was dependent on the time and temperature of the hyperthermia treatment. At 45°C for 60 min the shoulder of the radiation survival curve could be completely eliminated and the degree of enhanced cell killing at the 2 Gy level ranged from factors of 10 to 20 under the various conditions. When hyperthermia was given to cells which were irradiated and then plated immediately, or delayed for 8 h before plating to allow recovery, hyperthermia was found to cause radiosensitization under both conditions. In addition, when the hyperthermia dose was increased the difference between the immediate plating and the delayed plating survival curve decreased and for 45°C for 60 min this difference was completely eliminated, concomitantly with the elimination of the survival curve shoulder. These data indicate that hyperthermia may play a role in radiosensitization for the treatment of human glioma.     

Hyperthermia enhancement of radiation response and inhibition of recovery from radiation damage in human glioma cells

Three human glioma cell lines were tested for the effectiveness of hyperthermia and thermal radiosensitization. Thermal sensitization was evaluated from the perspective of increased radiosensitivity as well as inhibition of recovery from radiation damage. The three glioma cell lines tested showed large shoulders on the radiation survival curve and a large capacity for recovery of potentially lethal radiation damage. Hyperthermia caused radiosensitization in all three cell lines, which was primarily characterized by the reduction of the survival curve shoulder with moderate decreases in the survival curve slope. The radiosensitization was dependent on the time and temperature of the hyperthermia treatment. At 45 degree C for 60 min the shoulder of the radiation survival curve could be completely eliminated and the degree of enhanced cell killing at the 2 Gy level ranged from factors of 10 to 20 under the various conditions. When hyperthermia was given to cells which were irradiated and then plated immediately, or delayed for 8 h before plating to allow recovery, hyperthermia was found to cause radiosensitization under both conditions. In addition, when the hyperthermia dose was increased the difference between the immediate plating and the delayed plating survival curve decreased and for 45 degrees C for 60 min this difference was completely eliminated, concomitantly with the elimination of the survival curve shoulder. These data indicate that hyperthermia may play a role in radiosensitization for the treatment of human glioma.     

Inhibition of DNA synthesis by lonidamine and hyperthermia in human chronic myeloid leukemia cells

Lonidamine (LNA) is antitumor agent that lacks the characteristic properties of antiproliferative drugs and has narrow spectrum of antitumour effects. It is also a hyperthermia sensitizer. Hyperthermia affects on various cellular organelles, and the malignant cells are known to be more sensitive to hyperthermia. The present study examines the in vitro effects of LNA (0.01 and 0.02 mM) and hyperthermia (42 degrees C for 1 and 2 hrs) alone and in combination, on 11 human chronic myeloid leukemia cell samples. The inhibition in incorporation of 3H-thymidine was statistically significant (P less than 0.001) in human CML cells when exposed to the combination of LNA (0.01 and 0.02 mM) with hyperthermia (1 and 2 hrs) compared to the single treatment either at the same concentration of LNA at 37 degrees C or hyperthermia alone. Cytotoxicity was evaluated as the inhibition of the incorporation of 3H-thymidine in the cellular nucleic acid. The drug effects were estimated from the changes in rates of incorporation of precursor into DNA and compared with untreated control samples. Cancer cells were incubated in vitro in the presence of radioactive 3H-thymidine and drug.     

Interaction of SR-4233 with hyperthermia and radiation in the FSaIIC murine fibrosarcoma tumor system in vitro and in vivo

The effects of SR-4233 (3-amino-1,2,4-benzotriazine-1,4-dioxide), a hypoxic cell cytotoxic agent, were assayed against the FSaIIC murine fibrosarcoma in vitro and in vivo alone and in conjunction with hyperthermia and radiation. In vitro, a concentration of 500 microM of SR-4233 upon exposure of the cells for 1 h decreased the survival of hypoxic cells by about 1 log more than euoxic cells at 37 degrees C and pH 7.40. At the same concentration at pH 6.45, this difference in cytotoxicity increased to about 3 logs. In conjunction with 42 or 43 degrees C hyperthermia at pH 7.40, the killing of both euoxic and hypoxic cells was markedly increased (hypoxic greater than oxic), and the effect of hyperthermia on SR-4233 cytotoxicity was further increased at pH 6.45. SR-4233 proved to be an effective radiosensitizer of hypoxic cells in vitro, producing an enhancement ratio of 2.6 +/- 0.2 at pH 7.40 and 2.7 +/- 0.2 at pH 6.45. In vivo, however, SR-4233 (50 mg/kg) used with single dose radiation (10, 20, or 30 Gy) did not alter the slope of the radiation dose-dependent tumor growth delay curve but did produce a significant additive increase in tumor growth delay. Local hyperthermia (43 degrees C, 30 min) plus SR-4233 (30 mg/kg) produced a tumor growth delay of 9.1 +/- 2.2 days, whereas SR-4233 alone caused a tumor growth delay of only 1.7 +/- 0.9 days and the hyperthermia, only 1.4 +/- 0.7 days. The tumor growth delay increased to 28.2 +/- 4.4 days with the addition of daily radiation (3 Gy for 5 days) to SR-4233 and hyperthermia given on treatment day 1 only. Hoechst 33342 dye-selected tumor subpopulation analysis at 24 h following treatment demonstrated that SR-4233 (30 mg/kg) was more toxic to dim (presumably hypoxic) cells by about 1.8-fold. The addition of hyperthermia to treatment with SR-4233 increased the killing of dim cells by about 5-fold but of bright cells by only 2-fold. Trimodality treatment with SR-4233, hyperthermia, and radiation increased the killing of bright cells by about 6.5-fold and of dim cells by about 16.5-fold as compared with radiation alone. These results indicate that SR-4233 might be used quite effectively with radiation and/or hyperthermia to treat tumors with significant hypoxic subpopulations.     

Cisplatin sensitization by concurrent mild hyperthermia in parental and mutant cell lines deficient in homologous recombination and non-homologous endjoining repair

The effect of mild hyperthermia on cisplatin sensitization was examined in two cell line pairs, CHO parental AA8 and irsISF, an XRCC3 mutant (deficient in homologous recombination repair), and mouse parental MEF and knockout Ku80 mutants (deficient in non-homologous endjoining repair). The results showed that mild hyperthermia 40, 41 and 42 degrees C given concurrently with cisplatin treatment caused significant sensitization. The degree of sensitization was comparable for the parental and mutant lines, indicating that these repair pathways were likely not involved in cisplatin thermal sensitization. The shorter concurrent treatments cause a larger sensitization than the longer treatments. The reasons for this are not clear, but thermotolerance may be a factor.     

Effect of hypoxia and acidosis on the cytotoxicity of four platinum complexes at normal and hyperthermic temperatures

The cytotoxicities of cis-diamminedichloroplatinum(II) (CDDP) and of three recently developed dichloro complexes of bivalent platinum with radiosensitizing ligands [(1,2-diamino-4-nitrobenzene)dichloroplatinum(II) (Plato), trans-bis(2-amino-5-nitrothiazole)dichloroplatinum(II) (Plant), and trans-bis(2-nitroimidazole)dichloroplatinum(II) (NIPt)] were evaluated at 37 degrees C, 42 degrees C, and 43 degrees C at normal pH, at pH 6.45, and under normally oxygenated and hypoxic conditions in EMT6 cells in vitro. For CDDP, marked hyperthermic sensitization to the drug was evident in normally oxygenated cells, but hypoxic cells showed essentially no sensitization to the cytotoxicity of CDDP at elevated temperature at normal pH. Low pH further increased the cytotoxicity of CDDP toward normally oxygenated but not hypoxic cells at 37 degrees C and 42 degrees C. At 43 degrees C, however, low pH increased the cytotoxicity of CDDP toward both normally oxygenated and hypoxic cells, restoring nearly the full sensitizing effect of hyperthermia on CDDP cytotoxicity in the hypoxic cells. Plato was much more cytotoxic toward hypoxic than normally oxygenated cells under all culture conditions. At normal pH, hyperthermia increased the cytotoxicity of Plato in both hypoxic and normally oxygenated cells. At low pH, however, the cytotoxicity of Plato was inhibited at all temperatures and in both normally oxygenated and hypoxic cells. Plant was also more toxic to both normally oxygenated and hypoxic cells at elevated temperatures at normal pH. In contrast to Plato, however, Plant became much more cytotoxic toward hypoxic cells and showed increased cytotoxicity in normally oxygenated cells at low pH. Hyperthermia, however, did not further increase the rate of cell killing by Plant at low pH. NIPt, at the concentrations tested, was essentially nontoxic to cells at normal pH at 37 degrees C. Hyperthermia significantly increased the killing of hypoxic cells by NIPt under both normal and low pH conditions, but little cytotoxicity was noted for NIPt in normally oxygenated cells under any culture conditions. These results demonstrate that pH and the level of oxygenation of cells significantly affect the cytotoxicity of drugs at both normal and elevated temperatures. This sort of investigation may help delineate optimum drugs for use against environmentally determined subpopulations of cells within tumors.     

Sensitizing for cis-diamminedichloroplatinum(II) action by hyperthermia in resistant cells

cDDP-resistant Ehrlich ascites tumour (EAT) cells (ER cells) were tested for cellular content of total glutathione, heat sensitivity, cDDP sensitivity and synergistic effects of a combined treatment of heat and chemotherapy. In comparison with the non-resistant EAT cells (EN) the ER cells had an elevated level of glutathione. Treatment with d,l-buthionine-(S,R)-sulphoximine SO), resulting in almost complete depletion of cellular glutathione, did not cause drug sensitization. The ER cells were somewhat less heat sensitive compared with the EN cells. Heat chemosensitization was observed for the EN cells as well as for the ER cells. At 43°C (but not at 42°C) the thermal enhancement ratio (TER) for cDDP toxicity was significantly higher in the ER cells. The total number of cells killed by the combined treatment was less in the ER cells than in the EN cells. After analysing existing literature, combined with the current results, it is concluded that although cDDP-resistant cells can often considerably be chemosensitized by hyperthermia, in most cases the difference in cDDP sensitivity cannot be overcome totally. In those situations where cDDP-resistant cells are more sensitive to heat and also show a high TER, especially at clinically relevant temperatures, hyperthermia as added modality is indicated for clinical treatment.     

Acute extracellular acidification reduces intracellular pH, 42 degrees C-induction of heat shock proteins and clonal survival of human melanoma cells grown at pH 6.7.

Two human melanoma cell lines, SK-Mel-28 and DB-1, were used for in vitro studies of the mechanisms underlying heat resistance of human tumour cells adapted to growth in acidic environments. Adaptation to growth at low pH was characterized by resistance to 42 degrees C cytotoxicity and accompanied by an increase in endogenous levels of Hsp70 and/or Hsp27. Acute extracellular acidification to levels below pH 6.5 was required to sensitize the melanoma cells to 42 degrees C. Furthermore, cells grown at low pH were more resistant to sensitization by acute acidification than cells grown at pH 7.3. The intracellular pH (pHi) of cells grown at pH 6.7 was less than the pHi of cells grown at pH 7.3 both before and after acute acidification. A pHi threshold existed for melanoma cells growing at pH 7.3 below which they became sensitized to 42 degrees C. This pHi threshold differed between the SK-Mel-28 and DB-1 cells. In contrast, a pHi threshold for heat sensitization did not exist for cells growing at pH 6.7: any reduction in pHi before heating resulted in increased cell killing. Since cells grown at low pH lack a pHi threshold for heat sensitization, they are sensitized more to 42 degrees C per unit decrease in pHi than cells grown at pH 7.3. Acute acidification abrogated the 42 degrees C-induction of Hsp70 and Hsp27 in the melanoma cells. The pHi thresholds for abrogation of these HSPs are slightly higher than or comparable with the thresholds for cytoxicity for each cell line grown at pH 7.3, but abrogation occurred over a narrower range of pHi compared with cytotoxicity. Abrogation of heat-induced expression of these HSPs correlates with cytotoxicity in both cell lines with the exception of Hsp27 expression in SK-Mel-28 cells. In conclusion, strategies that reduce pHi in melanoma cells growing at low pH, such as in acidotic regions of tumours, could selectively sensitize them to hyperthermia because they lack a pHi threshold for heat sensitization.     

Effect of external pH on heat sensitization by local anesthetics

The sensitization of asynchronous Chinese hamster ovary (CHO) cells to 43 degrees C by procaine, lidocaine, and tetracaine was examined, with the pH of the medium carefully controlled at approximately pH7 and 8. Thermal enhancement factors for 43 degrees C were calculated for the surviving fraction of 0.01. The thermal enhancement factors of the local anesthetics were increased at approximately pH8 by factors of 2-12, depending on the local anesthetic and its concentration. The concentration of the uncharged free base form of the local anesthetic in the culture medium correlated positively with the thermal enhancement factors of each local anesthetic and in a near linear fashion with the thermal enhancement factors for procaine. However, the concentration of the cationic form of the local anesthetics in the growth medium did not correlate with heat sensitization. We conclude that the ability of local anesthetics to sensitize cells to heat killing is dramatically influenced by the extracellular pH, with increased sensitization at the more basic pH's. Secondly, it is the extracellular concentration of the free base form of the local anesthetics that correlates with heat sensitization.     

Effects of pH on heat sensitization of mammalian cells with procaine hydrochloride

The enhancement of heat killing of CHO cells by treatment with 7 mM procaine HCl increased when cells were treated under alkaline conditions. Below a pH of 6.9, very little heat sensitization was observed; however, as the pH was increased to 7.4 and above, considerable heat sensitization occurred. There were no changes in intracellular pH at the beginning of heating that could be responsible for this phenomenon.     

A comparison of heat and radiation sensitivity of three human glioma cell lines

Three human glioma cell lines were tested for radiation and hyperthermia sensitivity and compared to the responses of a normal human fibroblast cell line. The radiation response of the glioma cell lines exhibited a large shoulder on the radiation survival curve indicating radioresistance when compared to the more radiosensitive fibroblast cell line. The hyperthermia response for the glioma cell lines was qualitatively similar to responses reported for other cell lines. When compared to normal human fibroblasts the glioma cells were found to be more sensitive to hyperthermia than the normal fibroblasts indicating hyperthermia may be a promising method or adjunct to radiotherapy in the treatment of resistant glioma cells or tumors. The results also show that both the radiation and thermal response is influenced by cell culture conditions and growth status. Two of the cell lines grown to confluency and treated in confluency showed an increased radiation resistance at low doses and the cell lines showed decreased resistance at high doses compared to cells plated to confluency (see Methods and Materials). An increased thermal resistance, especially at the lower heating temperatures, was also observed for cells grown to confluency. Measurements of residual glucose in the culture medium at the time of irradiation was about the same for the two culture methods (55%-65%). Cell cycle analysis showed that the differences were not related to changes in cell cycle distribution.     

Interaction of hyperthermia and radiation in murine cells: hypoxia and acidosis in vitro, tumor subpopulations in vivo

To better understand the effect of the level of oxygenation and pH on the heat-radiation interaction, these factors were modeled in vitro using FSaIIC cells in monolayer and correlated with the response of Hoechst 33342 dye-defined FSaIIC tumor subpopulations treated in vivo. Exposure to both 42 degrees C and 43 degrees C for 1 h in culture prior to graded single fractions of radiation resulted in a striking decrease in the radiation oxygen enhancement ratio which was pH as well as temperature dependent. The oxygen enhancement ratio at 37 degrees C and pH 7.40 (or pH 6.45) was 2.9, but decreased to 1.4 at 42 degrees C at normal pH, 1.2 at low pH, and 1.0 at 43 degrees C at both pH values tested. This decrease in the oxygen enhancement ratio resulted from a far more marked decrease in Do values for the radiation survival curves of hypoxic cells compared to normally oxygenated cells at elevated temperatures. In addition, the shoulder region of the radiation survival curves was significantly decreased with increasing temperatures and the magnitude of the decrease was greatest in hypoxic cells at low pH. In vivo treatment followed by immediate tumor excision showed that bright cells (presumably oxygenated cells at normal pH) were approximately 2-fold more sensitive to 10 Gy of radiation than were dim cells (presumably hypoxic cells at low pH) but that dim cells were 2.5-fold more sensitive to 43 degrees C for 30 min hyperthermia. The combination of hyperthermia followed by radiation proved to be 1.8-fold more toxic to dim than to bright cells. Both hyperthermia alone and hyperthermia plus radiation, in contrast to radiation alone, were significantly more cytotoxic when tumors were left in situ for 24 h prior to excision as compared with immediate excision. These results indicate that hyperthermia markedly sensitizes hypoxic cells at low pH to the cytotoxic effects of radiation, as well as effectively killing cells in this tumor subpopulation.     

Simultaneous and sequential hyperthermia and radiation treatment of an experimental tumor and its surrounding normal tissue in vivo

In order to optimize the therapeutic effect of a combined hyperthermia-radiation treatment, the influence of sequence and interval between the two modalities on local tumor control and normal tissue damage was studied during variation of heating time and temperature. A C3H mouse mammary carcinoma transplanted into the feet of C3D2F1 mice was used as a model. Local hyperthermia was given to unanesthetized mice by immersion of the tumor-bearing foot into a water bath. Radiation was given either before, during or after heating. After simultaneous treatment an increasing thermal enhancement ratio (TER) was observed with increasing temperatures and/or increasing heating time with TER values ranging from 1.2 at 41° C to about 5 at 43.5° c after a one hour heating. In all simultaneous treatment schedules, the TER values were almost similar in tumor and normal tissue; no improvement in therapeutic gain was observed. Different results were obtained by using an interval between the two modalities. Hyperthermic treatment (42.5° C/60 min) given with intervals up to 24 hours before radiation showed no definitive improvement in therapeutic ratio. However, if radiation was given before the heating, the normal tissue completely recovered from thermal sensitization within 4 hours, whereas a marked thermal enhancement was persistent in the tumor for more than 24 hours. Thus, an increased therapeutic ratio could be obtained if radiation was given before heat and an interval of 4 hours or more was allowed. This improved therapeutic ratio was dependent on the temperature and ranged from about 1.1 at 41.5° C to 2.1 at 43.5° C given for one hour. These data indicate that if both tumor and normal tissue are heated, the optimal tumor effect may not be a hyperthermic radiosensitization, but rather a direct heat killing of radioresistant tumor cells. This special heat sensitivity of radioresistant tumor cells may be explained by the characteristic environmental conditions (e.g. chronic hypoxia and acidity) influencing such cells.     

The effect of the drug lonidamine on Chinese hamster ovary cells in vitro and on experimental tumors

Hypoxic cells in solid tumors are known to be resistant to radiation, and may also be resistant to some anti-cancer drugs. Biochemical properties of hypoxic cells, such as their dependence on anaerobic glycolysis leading to production of lactate and low pH might have potential for inhibition by drugs with selective activity against hypoxic cells. Such drugs might improve the Therapeutic Index when used with radiation or some conventional anti-cancer drugs. Preliminary studies have shown that the combination of hypoxia and low pH (pH 6.5-6.0) was cytotoxic to Chinese Hamster Ovary (CHO) cells incubated in vitro for up to 6 hours, although neither factor alone reduced plating efficiency. Lonidamine, an inhibitor of mitochondrially-bound hexokinase and lactate transport, was cytotoxic to CHO cells at low pH, but had no effect at physiological pH under aerobic or hypoxic conditions. Lonidamine has also been tested for in vivo effects against three murine tumors: the KHT fibrosarcoma, 16/C mammary carcinoma and the Lewis Lung Tumor. The drug was tested either alone, or with radiation or Adriamycin to kill aerobic cells, and/or with glucose and insulin to lower intra-tumor pH. No major therapeutic effects have been demonstrated.