1990, The British Journal of Radiology, 63, 752-759
Chronic anaemia, hyperbaric oxygen and tumour radiosensitivity By Mary McCormack, BSc, MSc, PhD, A. H. W. Nias, DM, FRCR, MRCPath and Eileen Smith, BSc, PhD Richard Dimbleby Department of Cancer Research, UMDS, St Thomas' Hospital, London SE1 7EH {Received January 1990 and in revised form April 1990)
Abstract. Anaemia is an important factor in the response of some human tumours to radiotherapy. The outcome is also influenced by whether the treatment is given in air or high pressure oxygen (HPO). The present study examined the relationship between anaemia and tumour response to radiation given in air or HPO in C3H mice transplanted with a mammary adenocarcinoma using a growth delay assay to assess the radiation response. Chronic anaemia was induced by the use of a low iron diet and was characterized by a significant reduction in host haematocrit and whole blood viscosity. In addition, anaemia was associated with a right shift in the oxyhaemoglobin dissociation curve and an increase in the volume doubling time of the tumour; but there was no change in the concentration of 2,3-diphosphoglycerate in the red cells. Radiation studies with these anaemic mice demonstrated that the tumour radiosensitivity was decreased when treatment was given in air. HPO was successful in overcoming the increased radioresistance associated with anaemia. This result suggested that tumours grown in anaemic mice have a higher hypoxic fraction than those grown in control mice. Changes in host physiology with chronic anaemia may contribute to the benefit seen with HPO but such alterations per se may be inadequate to maintain tumour oxygenation when treatment is given in air.
It has long been recognized that the response of tumours to radiotherapy in anaemic patients is less satisfactory than in those with normal haemoglobin (Hb) levels. Thus, in a study of 895 patients with carcinoma of the cervix, Evans and Bergsjo (1965) found that for Stage II and Stage III disease there was a significantly lower survival in those patients who were anaemic at the time of treatment compared with nonanaemic patients. The authors suggested that the normal compensatory mechanism in the presence of anaemia might fail to overcome tissue hypoxia in the abnormal vascular system of tumours. A report by Bush et al (1978) supported the thesis that hypoxia influences the local control rate with radiotherapy for patients with advanced carcinoma of the cervix. In this study of 2803 patients there was a significant difference in the local control rates, for all patients, analysed according to Hb levels. Those patients with Hb concentrations < 1 2 g % had a poorer prognosis than those with higher Hb concentrations. Relatively few studies have evaluated the usefulness of hyperbaric oxygen (HPO) in the treatment of anaemic cancer patients. In 1972, Dische and Hewitt reported on one patient with carcinoma of the cervix with severe anaemia (Hb4.3g%) who was treated by radiotherapy in HPO (six fractions in 3 weeks) without a blood transfusion, followed by a radium implant. The immediate response was satisfactory and with cessation of bleeding the Hb concentration rose to a normal value. This patient was still alive and well 16 years after treatment. A retrospective analysis, according to Hb concentrations, of patients in the Medical Research Council (MRC) trials of HPO in the radiotherapy of carcinoma of the cervix was carried out by Dische et al (1983). All patients with Hb concentrations below 10g% were 752
given a blood transfusion prior to therapy and again during therapy if required. Local tumour control rates for anaemic transfused patients treated in HPO were significantly better than those for similar patients treated in air. Sealy et al (1989) report a beneficial effect when they treated patients with carcinoma of the cervix by inducing anaemia (by exchange transfusion) followed by its correction prior to therapy in HPO. The aim of the present study was to elucidate the relationship between anaemia and tumour response to radiation, with particular regard to HPO as an adjunct to such treatment. This investigation centred on three main themes. (i) To establish a good model of chronic anaemia in the mouse, (ii) To examine the relationship between anaemia and tumour radiosensitivity in air and HPO. (iii) To examine the role, if any, of physiological adaptation to anaemia on tumour radiation response. Materials and methods
A serially transplanted C3H mouse mammary adenocarcinoma (Nias et al, 1980) was used in SPF-derived inbred C,H mice. Single doses of 250 kVp X rays (0.83 mm Cu half-value thickness) were delivered at 200 cGy/min to mice in the chamber used for hyperbaric oxygen studies (Tozer et al, 1984), with all but the tumour shielded by 2 mm of lead. Ketamine (45 mg/kg) and diazepam (0.5 mg per mouse) anaesthesia was used. Tumour volume was determined from three perpendicular measurements and growth expressed in terms of the mean (and standard error (SE) of the mean) time taken by a group of tumours to reach 3.5 times the treatment volume, which was approximately 200 mm3. The exact end-point sizes of individual tumours were related to The British Journal of Radiology, October 1990
Anaemia, HPO and radiosensitivity 501
their individual treatment volumes. At least 10 animals were used for each experimental point. Diet A pelleted diet (Tecklad, USA) with an iron content of < 5 mg/kg was fed ad libitum for 9 weeks to weaned mice (approximately 21 days old) obtained from Bantin and Kingman Ltd. Mice were usually maintained on the low iron diet for 9-10 weeks prior to treatment and were subsequently fed on a normal laboratory diet. Haematocrit determinations were made at weekly intervals from tail-vein samples during the 9-week period.
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Chronic anaemia, hyperbaric oxygen and tumour radiosensitivity By Mary McCormack, BSc, MSc, PhD, A. H. W. Nias, DM, FRCR, MRCPath and Eileen Smith, BSc, PhD Richard Dimbleby Department of Cancer Research, UMDS, St Thomas' Hospital, London SE1 7EH {Received January 1990 and in revised form April 1990)
Abstract. Anaemia is an important factor in the response of some human tumours to radiotherapy. The outcome is also influenced by whether the treatment is given in air or high pressure oxygen (HPO). The present study examined the relationship between anaemia and tumour response to radiation given in air or HPO in C3H mice transplanted with a mammary adenocarcinoma using a growth delay assay to assess the radiation response. Chronic anaemia was induced by the use of a low iron diet and was characterized by a significant reduction in host haematocrit and whole blood viscosity. In addition, anaemia was associated with a right shift in the oxyhaemoglobin dissociation curve and an increase in the volume doubling time of the tumour; but there was no change in the concentration of 2,3-diphosphoglycerate in the red cells. Radiation studies with these anaemic mice demonstrated that the tumour radiosensitivity was decreased when treatment was given in air. HPO was successful in overcoming the increased radioresistance associated with anaemia. This result suggested that tumours grown in anaemic mice have a higher hypoxic fraction than those grown in control mice. Changes in host physiology with chronic anaemia may contribute to the benefit seen with HPO but such alterations per se may be inadequate to maintain tumour oxygenation when treatment is given in air.
It has long been recognized that the response of tumours to radiotherapy in anaemic patients is less satisfactory than in those with normal haemoglobin (Hb) levels. Thus, in a study of 895 patients with carcinoma of the cervix, Evans and Bergsjo (1965) found that for Stage II and Stage III disease there was a significantly lower survival in those patients who were anaemic at the time of treatment compared with nonanaemic patients. The authors suggested that the normal compensatory mechanism in the presence of anaemia might fail to overcome tissue hypoxia in the abnormal vascular system of tumours. A report by Bush et al (1978) supported the thesis that hypoxia influences the local control rate with radiotherapy for patients with advanced carcinoma of the cervix. In this study of 2803 patients there was a significant difference in the local control rates, for all patients, analysed according to Hb levels. Those patients with Hb concentrations < 1 2 g % had a poorer prognosis than those with higher Hb concentrations. Relatively few studies have evaluated the usefulness of hyperbaric oxygen (HPO) in the treatment of anaemic cancer patients. In 1972, Dische and Hewitt reported on one patient with carcinoma of the cervix with severe anaemia (Hb4.3g%) who was treated by radiotherapy in HPO (six fractions in 3 weeks) without a blood transfusion, followed by a radium implant. The immediate response was satisfactory and with cessation of bleeding the Hb concentration rose to a normal value. This patient was still alive and well 16 years after treatment. A retrospective analysis, according to Hb concentrations, of patients in the Medical Research Council (MRC) trials of HPO in the radiotherapy of carcinoma of the cervix was carried out by Dische et al (1983). All patients with Hb concentrations below 10g% were 752
given a blood transfusion prior to therapy and again during therapy if required. Local tumour control rates for anaemic transfused patients treated in HPO were significantly better than those for similar patients treated in air. Sealy et al (1989) report a beneficial effect when they treated patients with carcinoma of the cervix by inducing anaemia (by exchange transfusion) followed by its correction prior to therapy in HPO. The aim of the present study was to elucidate the relationship between anaemia and tumour response to radiation, with particular regard to HPO as an adjunct to such treatment. This investigation centred on three main themes. (i) To establish a good model of chronic anaemia in the mouse, (ii) To examine the relationship between anaemia and tumour radiosensitivity in air and HPO. (iii) To examine the role, if any, of physiological adaptation to anaemia on tumour radiation response. Materials and methods
A serially transplanted C3H mouse mammary adenocarcinoma (Nias et al, 1980) was used in SPF-derived inbred C,H mice. Single doses of 250 kVp X rays (0.83 mm Cu half-value thickness) were delivered at 200 cGy/min to mice in the chamber used for hyperbaric oxygen studies (Tozer et al, 1984), with all but the tumour shielded by 2 mm of lead. Ketamine (45 mg/kg) and diazepam (0.5 mg per mouse) anaesthesia was used. Tumour volume was determined from three perpendicular measurements and growth expressed in terms of the mean (and standard error (SE) of the mean) time taken by a group of tumours to reach 3.5 times the treatment volume, which was approximately 200 mm3. The exact end-point sizes of individual tumours were related to The British Journal of Radiology, October 1990
Anaemia, HPO and radiosensitivity 501
their individual treatment volumes. At least 10 animals were used for each experimental point. Diet A pelleted diet (Tecklad, USA) with an iron content of < 5 mg/kg was fed ad libitum for 9 weeks to weaned mice (approximately 21 days old) obtained from Bantin and Kingman Ltd. Mice were usually maintained on the low iron diet for 9-10 weeks prior to treatment and were subsequently fed on a normal laboratory diet. Haematocrit determinations were made at weekly intervals from tail-vein samples during the 9-week period.
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