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When the clonogenic survival of mouse haemopoietic stem cells (CFU-S) and leukaemia L1210 cells growth as ascites tumours are compared after being heated in vitro and assayed in vivo by spleen-colony assay, there is no significant difference in the terminal slopes of the survival curves. The shoulders of the survival curves differ, but this may be explained by differences in cell kinetics. By contrast, L1210 leukaemic marrow cells are considerably more susceptible to the lethal effects of hyperthermia (43 degrees C) than either normal marrow stem cells or L1210 leukaemic cells grown as ascites tumours. Moreover, the killing of L1210 ascites cells by hyperthermia can be enhanced by heating L1210 ascites cells with an equal number of normal marrow cells, or as upernatant removed from heated marrow cells. Most cells in lukaemic marrow are normal, and it is postulated that the increased thermal sensitivity of L1210 cells in leukaemic marrow is caused by diffusible factors (e.g. lysosomal enzymes) released by heating normal marrow cells.

To compare the relative heat sensitivities of human normal and neoplastic cells of the same tissue type, a study was carried out of the relative sensitivities to heat of granulocyte/macrophage progenitor cells (CFU-GM) derived from the peripheral blood and bone marrow of normal subjects and patients with chronic granulocytic leukaemia (CGL). Nucleated haemopoietic cells were incubated at temperatures in the range 41.5 degrees C to 44.0 degrees C for various periods before culture in agar. The results of these experiments showed that CFU-GM from normal blood were consistently less sensitive to damage by heat than normal marrow CFU-GM. There was no comparable difference in the relative heat sensitivities of CFU-GM from blood and marrow of patients with CGL and no significant difference between the heat sensitivities of CFU-GM derived from marrow from normal individuals and patients with CGL. The observed difference in heat sensitivity of CFU-GM from normal blood and marrow accords with other data suggesting that the two progenitor cell compartments are distinct: the blood CFU-GM may represent a more primitive population of committed progenitor cells. In CGL, CFU-GM in the blood may much more closely resemble those in the marrow. The data provide no support for the hypothesis that malignant cells differ intrinsically from their normal counterparts in respect of sensitivity to damage by hyperthermia.

The colonogenic survival of L1210 leukaemic cells was assessed by spleen colony assay after cells were heated for 5, 10, 15 or 30 minutes at 43 degrees C. Similar cells which were fixed within 60 seconds of heating by glutaraldehyde were examined by scanning electron microscopy (SEM). The appearance of heated cells was compared with unheated controls which had been stored for one hour at 37 degrees C, 4 degrees C, and 22 degrees C respectively. After a heat treatment of 15 minutes at 43 degrees C the clonogenic fraction is only 0.1 but the appearance of approximately half the cells examined by SEM was unchanged. Virtually every cell heated for 30 minutes at 43 degrees C was abnormal. Progressive changes in the cell membrane were seen, such as loss of microvilli, folding and corrugation of the cell membrane and development of blebs. The lethal injury to cells is probably damage to enzyme systems in the cell membrane and elsewhere in the cell which is expressed as loss of clonogenicity. It is noteworthy that some cells appear to have been sterilised before morphological changes were seen. Two explanations are possible. The lethal injury as expressed by sterilisation may precede morphological changes or alternatively as cells had been heated for only a short time (10 or 15 minutes) and fixed within 60 seconds of heating, insufficient time had elapsed for gross changes to develop.

Thermotolerance in vivo was investigated using the rat fibrosarcoma SSBla. Hyperthermia was administered using water-bath heating, with the tumour blood supply occluded by clamping during the heating procedure. Single-dose heat treatments (10-60 min at 43.5 degrees C) yielded linear dose-response curves, with response evaluated as tumour re-growth delay. In thermotolerance studies split-dose treatments were given in which a single 'priming' treatment of 30 min at 43.5 degrees C was followed 24 or 48 hr later by a variable treatment (10-90 min), again at 43.5 degrees C. Split-dose responses produced generally less uniform results, though the dose-response curves were still approximately linear. Comparison of the slopes of the dose-response curves for single-dose and split-dose treatments gave a 'thermotolerance ratio' (TTR) of 4.09 and 3.45 at 24 and 48-hr intervals respectively. Data analysis using Monte Carlo simulation techniques also suggests that the distribution of tumour sensitivities to a second heat treatment may be considerably broader than the distribution of sensitivities to a first treatment. Uniformity of tumour response to a single heat treatment may therefore conceal a spectrum of capacities for development of thermotolerance amongst individual tumours.

The regrowth rate of the rat sarcoma SSB1a was assessed, following treatment with either X-rays or hyperthermia. The growth rate of untreated tumours implanted in pre-irradiated or pre-heated normal tissue was also measured. For treatments giving similar levels of tumour regrowth delay, the regrowth of irradiated tumours was markedly slower than that of untreated tumours, whilst the regrowth rate of tumours subjected to hyperthermia did not differ from that of untreated tumours. The growth rate of untreated tumours implanted in pre-irradiated skin was also slower than that of control tumours, though the growth-rate reduction was less dramatic than for regrowth following tumour irradiation in situ. Tumours implanted in pre-heated skin grew at the same rate as control tumours. Slower regrowth in situ after X-rays than after hyperthermia would produce a larger growth delay for X-rays than for hyperthermia, for equal levels of tumour-cell kill or probability of tumour sterilization. This effect could lead to a systematic underestimation of the efficacy of hyperthermia relative to X-rays, when efficacy is assessed by growth delay.