Japanese Journal of Clinical Oncology 31:142-146 (2001)
© 2001 Foundation for Promotion of Cancer Research
Influence of Thoracic Radiotherapy on Exhaled Nitric Oxide Levels in Patients with Lung Cancer
1Department of Radiation Oncology, Osaka Medical Center for Cancer and Cardiovascular Diseases, Osaka, Departments of 2Radiology and 4Radiation Oncology, Biomedical Research Center, Osaka University Graduate School of Medicine, Osaka and 3Department of Radiology, Toneyama National Hospital, Osaka, Japan
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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Background: To determine the physiological role of exhaled nitric oxide (NO) in patients with lung cancer.
Methods: We investigated changes in exhaled NO levels in 29 patients undergoing thoracic radiation therapy with or without chemotherapy. The exhaled NO level was assessed using a chemiluminescence analyzer.
Results: The level of exhaled NO was higher in patients with lung cancer before treatment than in controls. With radiotherapy, the exhaled NO level decreased for patients undergoing 40 Gy irradiation and post-radiotherapy. However, five patients showed elevated levels of exhaled NO three times or more than that before radiotherapy. Three of these patients showed signs of radiation pneumonitis. However, none of the other patients showed signs of radiation pneumonitis (p = 0.002).
Conclusion: Radiation therapy can lower exhaled levels of NO and the levels of exhaled NO may be a useful index for the early prediction of radiation pneumonitis.
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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For the treatment of thoracic malignancies, especially lung cancer, thoracic irradiation is widely used (1). However, radiation pneumonitis may be a serious reaction to this treatment with sometimes fatal consequences (2). Nitric oxide (NO) can be produced by a family of enzymes known as nitric oxide synthases (NOS) and is a multifunctional biological mediator that has been implicated as an active participant in both proinflammatory and bronchodilator biology (3,4). In the airway system, epithelial cells and type II epithelial cells contain cNOS and iNOS, nNOS in the nerve in the tracheal wall and iNOS in the bronchial smooth muscle. As a proinflammatory indicator, NO can mediate airway epithelial tissue damage, probably by virtue of its oxidation to peroxynitrite, OONO, which has a highly toxic action. Increased permeability in airway vessels is one of the main characteristics of radiation-induced lung injury that could be mediated by NO. NO may also function in a homeostatic role with both NO and stabilized forms of NO, such as nitrosothiols, mediating airway relaxation and inhibiting bronchovascular leakage. It has been established that the mixed expired air recovered from patients with asthma contains elevated levels of NO compared with that recovered from non-asthmatic individuals (4). Asthma treatment with glucocorticoid, but not with bronchodilators or bronchoconstrictors, results in a decrease in the NO levels of mixed expired air (4). In lung cancer patients, owing to tumor-related inflammation and the tumor itself, several kinds of cytokines (IL-1ß, IL-6, IF-
, TNF-
, TGF-ß, etc.) are produced to enhance the production of NO (5). In addition, NO production is reportedly elevated in cases of infection (6). Although radiation stimulation is a potential enhancer of NO production (7), few reports mention the role of the exhaled NO level in radiotherapy. The examination of NO levels has several merits as it is a convenient and non-invasive examination. We therefore conducted an examination of exhaled NO levels before, during and after thoracic radiation therapy for lung cancer patients in order to determine the physiological changes in exhaled NO levels resulting from thoracic irradiation. The validity of the level of exhaled NO for prognosis of radiation pneumonitis was also examined. |
MATERIALS AND METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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Between 1996 and 1997, we analyzed exhaled NO levels in 29 patients (24 males and five females) with previously untreated lung cancer who were treated with radiotherapy at the Toneyama National Hospital. The cancer were histologically identified as 11 adenocarcinomas, 10 squamous cell carcinomas, six large cell carcinomas and two small cell carcinomas. Table 1 shows details of the patients and treatment characteristics. The patients’ age ranged from 38 to 84 years (median: 61 years) and three of them were at stage I, one at stage II, 18 at stage III and seven at stage IV. Twenty-one patients underwent combined treatment with radiation and chemotherapy, while eight received radiation therapy only. Details of the chemotherapy are also shown in Table 1. Radiation was administered with a 10 MV X-ray linear accelerator and the size of the irradiated areas is given in Table 1. At a dose level of 40 Gy, an oblique field was used to avoid spinal cord injury. To examine the relationship between a known inflammatory indicator and exhaled NO level, we compared the latter with serum C-reactive protein (CRP). Treatment results were assessed at the completion of radiotherapy. A complete response (CR) was defined as disappearance of the tumor shadow. A partial response (PR) was defined as a
50% reduction in tumor size (the product of the two longest diameters of the tumor) in comparison with the corresponding value before treatment. When the reduction in tumor size was <50%, the classification was no change (NC).
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Exhaled NO was determined with a chemiluminescence analyzer (CLM-500, Shimadzu, Kyoto, Japan) which was connected to an on-line recorder for NO. Subjects wore nose clips and were either sitting or standing during the measurement of the exhaled NO level. Results were displayed on a chart recorder. Sampling was performed once a week before and during radiation therapy. At each examination, the patient exhaled three times into a wide-bore Teflon tube. The average of three successive peak values was identified as the exhaled NO level. As controls, 11 normal volunteers and 10 patients with bronchial asthma were also analyzed.
Chest radiographs were examined every week during radiation therapy and every month afterwards to assess lung injury until 12 months after treatment. When abnormal shadows appeared, CT images were used to identify conditions other than radiation pneumonitis, such as recurrence. We defined radiation pneumonitis as the appearance of clinical symptoms of dyspnea, chest pain, cough and fever concurrent with abnormal shadows on the CT images.
The procedure employed in this study was reviewed and approved by the local ethics committee and all patients gave their informed consent to be included in this protocol.
For statistical analysis, Student’s t-test for normally distributed data and the Mann–Whitney U-test for skewed data were used. Pearson’s correlation coefficient was computed. Percentages were determined using the chi-squared test. All analyses used the conventional p < 0.05 levels of significance.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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First, we examined the background deviation among patients with lung cancer and asthma and normal controls. Significant differences were found in the exhaled NO levels for patients with lung cancer (n = 29, 0.077 ± 0.047 p.p.m., range 0.020–0.165 p.p.m.) and normal controls (n = 11, 0.044 ± 0.013 p.p.m., range 0.025–0.065 p.p.m., p = 0.03) and asthmatic patients showed a higher elevation (n = 10, 0.355 ± 0.248 p.p.m., range 0.051–0.771 p.p.m., p < 0.0001) (Fig. 1). There was no significant relationship between the levels of exhaled NO and of CRP (Fig. 2). Therefore, the exhaled NO level does not seem to reflect inflammatory status of the whole body if the inflammation does not involve the respiratory organs.
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A reduction in exhaled NO level was observed in patients undergoing radiation therapy at 40 Gy (0.050 ± 0.030 p.p.m., range 0.016–0.124 p.p.m., p < 0.05) and after radiation therapy (0.049 ± 0.033 p.p.m., range 0.005–0.165 p.p.m., p < 0.05) compared with pre-therapy values (0.078 ± 0.047, range 0.015–0.165 p.p.m.) (Fig. 3).
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We compared the exhaled NO level at the completion of radiotherapy in terms of initial tumor response. No significant relationship was found in exhaled NO levels between patients who showed tumor regression (CR and PR; n = 24, 0.105 ± 0.084 p.p.m.) and the others (NC; n = 5, 0.058 ± 0.032 p.p.m.) (n.s.). In addition, we compared the exhaled NO level in patients who underwent combined chemotherapy or not. No significant relationship was found in exhaled NO levels between patients who underwent chemotherapy (n = 21) and the others (n = 8) (n.s.) (Fig. 4).
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The elevation of the exhaled NO level of five patients was three times or more that seen earlier in radiotherapy. Three of these patients (ID 2, 13 and 18) showed signs of radiation pneumonitis requiring steroid administration (Fig. 5). In contrast, no patient who did not have a three times or more elevation of exhaled NO showed radiation pneumonitis (p = 0.002). None of the patients died of radiation pneumonitis.
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A 56-year-old female (ID 2) registered a slight decrease in the exhaled NO level at 20 Gy (0.05 p.p.m.). However, she showed a high exhaled NO level (0.241 p.p.m. at 30 Gy, 0.134 p.p.m. at 50 Gy) which was substantially higher than the previous value (0.05 p.p.m.). She complained of worsening cough and an increase in sputum 2 months after radiotherapy. An abnormal increase in density (a few pathy opacities and grand grass shadow) was found on a chest roentgenogram obtained 3 months after radiotherapy. Although the patient recovered from her symptoms with medication, she developed multiple intrapulmonary metastasis 4 months after treatment and died 7 months after treatment.
The second case was a 70-year-old male (ID 13) who showed a steady elevation of the exhaled NO level, which increased from 0.022 p.p.m. before radiotherapy to 0.035 p.p.m. at 20 Gy, 0.039 p.p.m. at 30 Gy and 0.068 p.p.m. at 40 Gy. We stopped radiotherapy at 44 Gy because an abnormal shadow appeared on his chest roentgenogram and simultaneously his CRP increased to 20. His exhaled NO level decreased to 0.035 p.p.m. 1 week after and to 0.021 p.p.m. 2 weeks after treatment interruption and symptoms and the abnormal shadow on the roentgenogram also disappeared in 2 weeks in response to medication.
The third case was an 84-year-old male (ID 18) with superior vena cava syndrome. His exhaled NO level increased from 0.015 to 0.023 p.p.m. at 10 Gy, to 0.044 p.p.m. at 20 Gy and to 0.031 p.p.m. at 30 Gy. However we continued radiotherapy with reduced field because until then his clinical symptoms and laboratory findings had been improved by radiotherapy. Exhaled NO levels also decreased to 0.020 p.p.m. at 40 Gy and 0.008 p.p.m. at 50 Gy. One week after treatment was completed, the exhaled NO level elevated to 0.053 p.p.m. and a ground grass shadow was recognized on CT as well as mild dyspnea on effort. Medication produced an improvement in the clinical symptoms and no deterioration of the abnormal shadow on the roentgenogram.
The irradiated area and dose were not found to be significant predisposing factors for radiation pneumonitis in this series, because there were no differences in these two variables between the three patients with pneumonitis (48 ± 3 Gy, 148 ± 77 cm2) and the others (50 ± 7 Gy, 138 ± 63 cm2).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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There is evidence that NO may contribute to tumor control during radiotherapy by increasing nucleic acid damage and by disruption of intracellular signalling (8). Ibuki and Goto reported that whole-body irradiation with a low dose of
-radiation activates macrophages indirectly and consequently enhances NO production from these macrophages, thus enhancing self-defense systems such as tumoricidal activity (7). Activated macrophages produce high levels of NO that destroy or prevent the division of tumor cells by inhibition of DNA replication and prevention of mitochondorial respiration (9). Liu et al. reported that in patients with primary lung cancer, the production of NO from alveolar macrophages increased as a result of the up-regulation of iNOS activity and the extent of iNOS expression in alveolar macrophages was closely related to the exhaled NO level (10). Arias-Diaz et al. also reported that the NO level in the bronchoalveolar lavage fluid of lung cancer patients was higher than in that of normal controls (11). We obtained similar results in that our lung cancer patients showed higher levels of exhaled NO. In addition, we could not establish any relationship between CRP and exhaled NO level, or between the latter and body temperature (data not shown). Therefore, the exhaled NO level does not reflect the inflammatory status of the whole body but only the local airway status.
We noted suppression of exhaled NO level on radiotherapy. One explanation for this decrease can be found in tumor regression and/or suppression of tumor-associated inflammation (12–14). We therefore hope that a decline in the exhaled NO level might be an effective monitor for radiation therapy. However, we could not establish any relationships between the treatment results and exhaled NO level at the completion of radiotherapy. This is partly because an increase in the exhaled NO level as a result of radiation induces inflammatory changes. A high elevation of exhaled NO level of three times of more appeared in five patients, three of whom showed signs of radiation pneumonitis. Therefore, an increase in exhaled NO of three times or more implies a risk of respiratory tract inflammation, especially radiation pneumonitis.
Several limitations of this study need to be considered. Methods of examination require careful consideration. Lundberg and co-workers reported that almost all exhaled NO originates from the upper nasal airways with only a minor contribution from the lower airways (4,14). Therefore, contamination with nasal NO is likely to result in inaccurate data so that a better sampling method is needed to avoid contamination. However, as several authors have mentioned, the elevation of exhaled NO level is so large (in the case of asthma) that we believe that the value can be used to estimate changes in the lower airways caused by radiation stimulation.
A further reservation is the time of detecting a risk of radiation pneumonitis. Our experience shows that high exhaled NO levels are seen in the later periods of treatment, when almost all of the required radiotherapy has already been administered. Although tripled or more exhaled NO levels seem to be significant for the detection of pulmonary inflammation, further studies are required to detect earlier signs of radiation pneumonitis so that radiotherapy and treatment can be stopped earlier.
In conclusion, monitoring of exhaled NO levels appears to be a useful method for the assessment of radiation pneumonitis.
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Acknowledgments |
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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This study was supported in part by a Research Grant for Science and Cancer from the Ministry of Education, Science and Culture, Japan. We thank Professor N. Taniguchi for critical discussions.
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FOOTNOTES |
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+ For reprints and all correspondence: Hideya Yamazaki, Department of Radiology, Osaka University Graduate School of Medicine, 2–2, Yamadaoka, Suita, Osaka 565-0871, Japan. E-mail: hideya10@hotmail.com
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TOPABSTRACTINTRODUCTIONMATERIALS AND METHODSRESULTSDISCUSSIONAcknowledgmentsREFERENCES |
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Received October 3, 2000; accepted December 27, 2000.
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