Biological effects of radiation in combination with other
physical, chemical or biological agents 

Annex L from Ionizing Radiation: Sources and Biological Effects 

United Nations Scientific Committee on the Effects of Atomic Radiation (UNSCEAR) 1982 Report to the General Assembly

United Nations, New York, 1982

IV. Biological Agents — VI Research Needs

     

    IV. BIOLOGICAL AGENTS 

    ANNEX L   CONTENTS

    							Paragraphs 
INTRODUCTION 						1-20
I. MODES OF INTERACTION 				21-72
	General approach 				21-41
	Surface of response and isobolic diagrams 	42-49
	Probabilistic assessment of the interaction 	50-61
	Theory and practice 				62-72
II. PHYSICAL AGENTS 					73-113
 A. Combinations of various types of ionizing radiation 73-78
 B. UV and ionizing radiation 				79-85
 C. Electromagnetic and ionizing radiation 		86-93
	Experimental data 				86-91
	Epidemiological evidence 			92-93
 D. Suboptimal temperature and ionizing radiation 	94-103
	High temperature 				94-99
	Low temperature 				100-103
 E. Magnetic fields and ultrasound 			104-107
 F. Dusts and fibres 					108-113
III. CHEMICAL AGENTS 					114-199
 A. Inorganic compounds 				114-120
 B. Organic radiosensitizing compounds 			121-136
 C. Carcinogenic chemicals 				137-157
 D. The special case of tobacco smoke 			158-183
	General 					158-159
	Experimental data 				160-168
	Epidemiological evidence 			169-183
 E. Other drugs 					184-199
IV. BIOLOGICAL AGENTS 					200-217
	General 					200-201
	Hormones 					202-213
	Infectious agents 				214-217
	Viral infections 				214-215
	Bacterial infections 				216-217
V. CONCLUSIONS 						218-237
VI. RESEARCH NEEDS 					238-244

							Page 
    References 					        765
     

    A. GENERAL

  1. Many biological conditions may influence the state of health of a human population. Viral and bacterial infections, eating habits and the state of nutrition, the conditions of living and working, the use of biologically active substances or drugs are known to affect to various degrees the incidence and pattern of diseases in humans and therefore to alter the actuarial characteristics of populations. There seems to be little hard evidence that conditions adversely influencing the survival and disease incidence could also substantially change man's sensitivity with respect to late radiation effects. This notion cannot however be dismissed or excluded because the complex pathogenesis of the effects of major interest in human radiation biology (tumour induction, genetic changes, developmental abnormalities) leaves scope for combined actions in both directions.
  2. The Committee wishes to stress that the above general notion is much easier to be entertained academically than to be experimentally demonstrated. The agents that may be considered as possible candidates for interaction with radiation are very many and diversified; their influence, already in the absence of radiation, is often little known in respect to the effects described above; and the data available are fragmentary. Therefore, to attempt a systematic discussion is almost impossible. In spite of such limitations, the Committee has decided to gather the available evidence on the effects of hormones and the effects of infections. Reference to existing epidemiological studies will also be made.

    3. B. HORMONES

  3. The influence of hormones on the radiation sensitivity of human populations with respect to cancer induction can be predicted on the general notion that many experimental and human tumours are known to be variously susceptible to the action of hormones. Mammary gland, prostate and thyroid tumours are very hormone-dependent, while for other malignancies a certain degree of dependency may be postulated, for example, on the notion of a different susceptibility between sexes or on the effect of castration. As to the practical significance of a combined action of hormones and radiation, changes of the hormonal state take place during physiological conditions (menarche, pregnancy, menopause, stress). Treatment of many diseases requires prolonged use of hormonal preparations and an increasingly large part of the female population use hormonal treatments (essentially oestrogens) for contraceptive purposes. Oestrogens are also contained in commercially available cosmetic preparations and some drugs used rather extensively (derivatives of Rauwolfia, phenothiazine, chlorinated hydrocarbons) have hormonomimetic activities. Finally, hormones themselves could be to some degree carcinogenic [C4]. There appears to be therefore sufficient ground for some analysis of their combined actions with ionizing radiations.
  4. Much general evidence about tumour induction in animals has been discussed in the 1977 report of the Committee, Annex I [Ul]. It was concluded that various radiation-induced tumours are differently affected by the animals' hormonal balance during the course of the carcinogenic process. The effects reported seemed to be tumour-, strain- and sex-specific and it appeared likely that the mechanisms of action (which are at present almost unknown at the molecular or even at the cellular level) might have been very different under the various conditions tested. Annex K to this report contains some discussion of the influence of the animal's sex on the life shortening action of ionizing radiation. This appears mainly as a higher susceptibility to sex-specific tumours, particularly the mammary neoplasms and the tumours of the genital tract in the female.
  5. Segaloff and Maxfield [S7] studied specifically the influence of oestrogens on mammary carcinogenesis in the rat. Pellets containing 5 mg diethylstilbestrol (DES) and 15 mg cholesterol were implanted subcutaneously into 8-week old A x C rats. The animals were hysterectomized to prevent fatal oestrogen-induced uterine infections. X-radiation was delivered only to the left mammary chain by shielding the opposite one. Spontaneous mammary tumours in this strain of rat are essentially nil. Radiation alone (about 8 Gy) produced only a small number of tumours (1.1 per chain at risk) appearing late (median 80 weeks at the appearance of the first tumour). DES alone gave 1.7 tumours per chain with median appearance times of 33 weeks. Combined treatments resulted in an earlier appearance of the tumours (26 weeks) and in an increased incidence (5.6 tumours/chain). Even a crude estimate based on final incidence would lead to an interaction factor of the order of 2, an estimate which (apart from its unknown statistical value) fails to take account of the appearance time which is shortened by the hormonal treatment.
  6. Shellabarger et al. [S8] irradiated with 0.43 MeV neutrons rats of the strain A x C in doses of 0.096 Gy. The carcinogenic response to irradiation was insignificant (3 adenocarcinomas in 33 rats); DES, on the other hand, produced some effect (182/25 rats). Combining the treatments led to an earlier appearance of tumours in much greater number (842/35 rats). There were therefore strong indications of a synergistic interaction and a crude estimation of is in the range of about 3. However, on Sprague-Dawley rats the same combined treatment produced a negligible incidence of tumours (2/31 rats) in comparison with the action of radiation alone (11/31 rats). DES in this case had no synergistic but rather an antagonistic effect. The experiment is a good example that, depending on the strain used, the same type of treatment may give rise to antagonistic or synergistic actions.
  7. Most recent experiments by the same group [H26] confirmed the effect of DES and showed, in addition, a synergistic action of 17-ethinyl-estradiol (EE2) in rats. The complex of these data would imply that the synergistic interaction is not with the hormones examined but rather with their oestrogenic activity. In other studies by Segaloff and Pettigrew [S5] graded doses of radiation of 0.5, 1.5 and 4.5 Gy were used. Radiation given alone increased the incidence of benign tumours in proportion to dose, but the increase of malignant tumours did not follow a statistically significant proportionality and resulted in a rather low number. Combining radiation and DES produced a synergistic interaction at 0.5 Gy ( 1.4), but the increase in tumour incidence was most pronounced at 1.5 Gy (crude = 2.0) and somewhat less at 4.5 Gy (crude = 1.6). The combined treatment led to an earlier tumour development.
  8. The role of prolactin in combination with radiation and with the chemical carcinogen N-nitroso-N-buthylurea was studied by Yokoro et al. [Y2] in W/Fu rats. Prolactin was produced by grafting a mammotropic pituitary tumour. Prolactin alone was ineffective in inducing mammary tumours. After doses of 2 Gy of x rays two fibroadenomas were seen among 27 animals with mean appearance times of about 6 months. Prolactin in combination with irradiation accelerated tumour appearance and induced tumours in 60% of the animals at the dose of 2 Gy. There were statistically significant differences in the tumour incidence between animals receiving 0.5 or 2.0 Gy and a similar synergistic interaction of prolactin and radiation was also found in respect to 14 MeV neutrons. Two interesting observations were made in these experiments. First, delaying the pituitary graft as long as seven months after irradiation still produced an enhanced effect, showing that the transforming lesions induced by radiation could remain available for hormonal interaction for a very long time. Secondly, most of the tumours produced by the interaction were adenocarcinomas, while most of the spontaneously occurring ones in this strain of rat are late appearing fibroadenomas.
  9. In a more recent study by the same laboratory [Y6] fission neutrons (2 MeV mean energy) mixed with gamma rays were given to W/Fu rats. Only 2% of the animals developed mammary tumours after irradiation alone (up to 0.2 Gy) but 42% did when prolactin was given shortly after irradiation by grafting the prolactin-secreting pituitary tumour. Delaying the prolactin treatment up to 12 months produced 24% tumours, which observation supports the one previously reported [Y2]. A similar synergistic interaction of diethylstilbestrol (DES) and neutron irradiation in the production of mammary, pituitary and hepatic tumours was observed in castrated male W/Fu rats [S41].
  10. The above results suggested to Yokoro [Y2] that in the previous studies the synergism between DES and radiation [S5, S7] could act via an increased production of prolactin. At the same time, Shellabarger [S6] was able to show that A x C rats (in which interaction with hormones was found) carried prolactin-secreting pituitary tumours; on the contrary, the Sprague-Dawley rats which did not show any synergism between radiation and DES carried no such tumours. In recent experiments by the group of Shellabarger [S44, H26] on A x C rats a strong dependence was shown between the interaction factor and the dose of DES and radiation. The dependence on the DES dose appeared to be mediated via the oestrogenic stimulation of prolactin secretion. The higher and the earlier the levels of prolactin in plasma, the greater was the yield of individual and multiple mammary adenocarcinomas.
  11. Another oestrogenic hormone (polyestradiol phosphate) and a corticosteroid (methyl-prednisolone) were tested in combination with internal irradiation by Nilsson et al. [N8] on CBA mice. Three doses of 90Sr (0.925, 1.850 and 7.400 kBq/g) were applied, which led to a maximum of 2% animals with pituitary tumours. Polyestradiol alone produced 10% of such tumours. Combining the treatments resulted in 44 and 37% of animals with tumours, for the first and the second dose of 90Sr, respectively, an increase corresponding to an interaction factor of approximately 4. Combined treatment also led to a decrease of the tumour induction time with respect to the groups given the radionuclide alone, close to that of the animals receiving only the hormone. Prednisolone in combination with radiation was ineffective in increasing the incidence or decreasing the induction time in comparison with groups receiving strontium alone.
  12. Modelling of situations in animals that may operate in women taking contraceptive oestrogens was undertaken in the Netherlands [B3, B5]. The complete outline of these experiments calls for three different strains of rat (Sprague-Dawley, Wistar Wag/Rij, Brown Norway); four types of radiation (300 kV x rays and 0.5, 4 and 15 MeV neutrons); a range of different doses (from 0.1 to 2 Gy, according to the radiation employed); and various types of female animals (intact or hysterectomized, respectively with or without hestradiol-17-beta). The results of this series are still incomplete, but some preliminary conclusions may be drawn. For WAG/Rij rats the proportion of animals surviving without tumours abruptly decreased starting from nine months of age after irradiation with 4 Gy x rays and hormonal treatment. For animals receiving only irradiation or hormonal treatment a 50% decrease was observed after 22 months. The total yield of tumours for the combined treatment group was also higher. Considerable differences in the susceptibility to tumour induction were found between strains. Brown Norway rats having the lowest spontaneous incidence of mammary tumours had an intermediate susceptibility to the radiation-induced ones. Pathological data showed that malignant tumours were relatively rare in the Brown Norway and in the Sprague-Dawley strains, but were instead quite common in the Wag/Rij rats, amounting in the latter strain to nearly one-half of all tumours. A synergistic interaction of radiation with the oestrogen treatment was manifested not only through an increased proportion of rats with malignant tumours (from 0.43 to 0.83 in the Wag/Rij rats) but also through an increased absolute incidence of neoplasia in Wag/Rij and Sprague-Dawley rats. The minimum latency period in untreated control animals could be in excess of 22 months; in irradiated animals without hormones this period decreased to 10–12 months and a decreased latency in the hormone-treated rats in comparison with untreated groups was observed as a rule. The synergistic action of the hestradiol-17-beta is of the same type as the interaction between radiation and DES.
  13. Kennedy and Weichselbaum [K15] reported a synergistic interaction between cortisone and x rays for transformation of C3H 10 T 1/2 cells in culture. The end-point scored is of great significance since it relates to tumour induction in vivo and the synergistic effect was statistically significant at P < 0.001. However, the transformation mechanisms in this particular cell line are still little understood [K16] and it is not possible to quantitate the results in terms of transformation frequency per surviving cell. It seems thus more prudent to test for effects in vivo before accepting the conclusions as generally valid. 
  14. Although the most informative data on the subject of combined action of radiations and hormones can only come from epidemiological surveys, data in this area are only indirect. It is known for breast cancer induction that age at exposure is a major determinant in all series available [T8, B27, S47]. Taken together, the data suggest that when the most profound hormonal changes occur (menarche, menopause) the risk per unit dose deviates most significantly from the mean risk for the whole life.

C. INFECTIOUS AGENTS

1. Viral infections

  1. Viruses have a very important role in the pathogenesis of some radiation-induced experimental tumours like the thymic lymphoma, the myeloid leukaemia and the osteogenic tumours of different strains of mice. The Committee has reviewed the relevant evidence in Annex I of its 1977 report. It is difficult in fact in many instances to separate the action of the virus from that of radiation, because the interplay of the biological and of the physical factors is in these cases so intimate that it would not be possible to elicit the effect without the presence of the two agents combined. To think of a synergistic effect under the circumstances would be inappropriate because none of the agents alone may be active for the specific end-point. Moreover, the vertical transmission of the viruses through successive animal generations makes it a normal constituent of their genome, which is exactly the reason why some tumours are specific to some strains.
  2. Radiation enhancement of in vitro cell transformation by viruses has long been reported [P13, S45]. An example was given of a combined treatment of Wistar/ Furth rats with radiation and Gross mouse leukaemia virus [Y1]. Animals aged 7–8 weeks were intraperitoneally inoculated with a standard dose of virus (0.4 ml of leukaemic filtrate) and none of the 15 animals injected developed leukaemia. Whole-body x-irradiation (four doses of 1.5 Gy given at five days interval) produced also no tumours in 12 irradiated animals. The combination of both treatments gave rise to more than 50% leukaemias in 20 treated animals. In view of the lack of effects by the separate treatments, the inter-action factor would in this particular case be equal to infinity. It was suggested that radiation might have acted through a modification of the physiological state of the target cells by rendering them susceptible to the action of the virus or through a modification of the immunological response of the host.

    3. 2. Bacterial infections

  3. Environmental conditions have often been reported to influence the induction of specific tumour types in irradiated animals through their action on the microflora. It is conceivable that the response to any carcinogenic stimulus, including radiation, may interfere with expression of the carcinogenic damage by modifying the number, susceptibility or turnover rate of the target cells or by altering the immunological response against transformed cells. The most extreme conditions under which to test such hypotheses are provided by the study of germ-free as opposed to gnotobiotic or conventional animals.
  4. Following irradiation of RF/Un mice myeloid leukaemia is decreased in the absence of microbial flora [W6], an effect which has been attributed to the reduced myelopoietic cell proliferation in germ-free animals [W7, W8]. Radiation-induced lymphatic leukaemia is, on the contrary, unaltered by germ-free conditions in many other strains of mice [P7, WI, W6]. Gnotobiotic and conventional animals show no qualitative differences with regard to virus particles found with the electron microscope [P7]. Induction of other solid tumours in irradiated mice gives variable results [A10, W7] and radiation-induced malignant or benign tumours are unaffected in germ-free rats. Thus, the data essentially show that the pathogenesis of radiation-induced cancer is similar in conventionally reared or in gnotobiotic animals. It should be concluded that the microbial flora as such has only a minor role in the development of haemopoietic neoplasms, perhaps via a modification of the immune system.

    5.  

    V. CONCLUSIONS

  5. The interaction between ionizing radiation and other agents represents a field of great potential importance in view of the ubiquitous nature of radiation and of the many situations of interaction that might occur in modern life with a variety of physical, chemical or biological agents. Yet, it is very difficult to define and substantiate the notion of interaction with even a moderate degree of refinement. Many reports have claimed some kind of interaction but comprehensive analysis does not show a sufficiently good conceptual basis for the nature of the interactions. There is a lack of systematic treatment of any given case, particularly with regard to the mechanisms of action. There is further a need to apply existing methodologies of analysis from other fields of the biological sciences to the study of these problems.
  6. The Committee has carried out a preliminary analysis of the combined actions in the radiobiological field, centered mainly around situations that may possibly be of importance for risk assessments in man and may therefore reflect on the present foundations of radiation protection. Available information on tumour induction, genetic defects and developmental effects was therefore scrutinized in the course of this analysis for any evidence of combined actions. The conditions of long-term exposure to low levels of the interacting agents were reviewed in detail, although in most of the reports the levels of exposure were much higher than the environmental. Where possible, the accent was on the results of epidemiological studies in humans, although the bulk of the information relates to animals.
  7. The Committee proposes that two types of inter-action may be considered. The first is one where both the ionizing radiation and the other interacting agent(s) are capable of producing some effect. Additivity, synergism and antagonism are the three possible conditions of interaction. The second type of combined action is that between ionizing radiation and other agents which are, when given alone, inactive. Protection or sensitization are the terms that apply in these cases, when reduction or enhancement, respectively, of the radiation effect are the end-results of the interactions. Such classification is not an absolute one because the doses of the interacting agents and the types of effect may influence profoundly the nature and degree of the interaction.
  8. The concepts of exposure, dose and response may be applied to the special case of the combined action with ionizing radiation. The existing methodologies of analysis (isobolic diagram, envelope of additivity, surface of response) allow the assessment, at least on a semi-quantitative basis, of the results of combined treatments. These analyses may be further extended to generalized probabilistic treatments of the experimental results, taking into account the variability of the biological systems under study and leading to a more quantitative and satisfactory description of the interaction factors.
  9. The applicability of these rather abstract notions to practical situations, particularly in the presence of complex biological effects, has been discussed. The need to define the effects with precision and to explore the full exposure-response ranges to all agents, acting separately or jointly, is a necessary prerequisite to meaningful studies. Also, pitfalls have been identified which may simulate conditions of interaction. In relation to important biological end-points such as the induction of tumours, the need to combine pathological and actuarial observations for a complete description of the phenomena has been underlined.
  10. The temporal pattern of the exposure (contemporaneous or sequential, chronic or acute, single or fractionated) as well as the order of administration appear of decisive importance in respect to the production of a given type or degree of effect and have also been examined in the Annex. All these conditions relate to practical situations, even though they may tend to blur the clearly defined notions of additivity, synergism and antagonism. A detailed knowledge of the nature of the effects, their relationships to time and to the full range of doses of the interacting agents, including the zero-dose condition, is important. In many papers these basic conditions were imperfectly described. In other cases, the statistical significance of the results was too low for a complete assessment of interaction. Thus, the present conclusions should only be considered as preliminary.
  11. An instance of interaction could be that between two different types of ionizing radiation, usually a combination of high- and low-LET radiation. Uncertainties exist as to the degree of interaction, owing to the essentially unknown nature of the primary radiation lesions and their repair systems. Even in cases where the yield of effect per unit dose of the two radiations differs by an order of magnitude, the interaction is within the limits of hetero- and iso-additivity. The study of the combined action of UV and ionizing radiation may be very valuable for the analysis of primary lesions and repair mechanisms. Experiments on survival of mammalian cells point to simple additivity. The important practical case of skin cancer induction, when tested in the animal, produced no evidence of interaction.
  12. Examples of synergistic effects have apparently been reported in workers exposed jointly to ionizing radiation and microwaves in the radiotechnical industry. Functional disturbances of the nervous system and subjective symptoms of discomfort were mainly found in these workers. The nature of the symptoms, the difficulties of their quantification, the frequently uncontrollable conditions of exposure and the unsatisfactory dosimetry, the incomplete statistical evaluation, are all reasons for which these reports should be regarded with some reservation.
  13. The combined action of suboptimal temperatures and radiation has given evidence of interaction in both directions, synergistic or antagonistic, depending perhaps on the type of effect, order of administration and level of exposure to the interacting agents. It would not be expected that any such effect would normally play any important role in higher animals, in view of their highly developed system of body temperature regulation. High altitude, metabolic or physical stress, mechanical damage, magnetic fields and ultrasound were also considered for a possible interaction with radiation: the results were variable but there was no evidence of significant synergistic interaction. In all these fields the data are very few, the effects non-specific and the mechanisms too obscure to allow any definitive statement.
  14. The combined action of radiation, given internally or externally, with various types of dust shows under repeated testing, particularly with regard to tumour induction in the respiratory system, synergistic, additive or antagonistic effects. Considering the uncertainties and limitations of the data, the synergistic effect of the combined treatment did not exceed a factor of about two and the inhibitory effects a factor of about four, compared to situations where radiation was administered alone.
  15. A variety of inorganic chemical compounds containing lead, silver, cadmium, calcium, beryllium, platinum, chlorine and fluorine, were also tested in experimental animals in conjunction with radiation for their carcinogenic, developmental or generally toxic properties. The results were once more extremely variable. In many cases the experience was so superficial, the effects so varied and the biological systems so different that no conclusions could be offered. Some of these interactions may be of significance in working situations and could profitably be explored further.
  16. In this review, radioprotective and radiosensitizing substances were not examined in detail, since conditions relevant to the exposure of the population were the main object of the Annex. High levels of radiation and nearly toxic levels of these substances have been used in the relevant studies. A great variety of underlying mechanisms, complex relationships to the dose, to the radiation type, to the presence of oxygen were described for these chemical compounds. Since these substances are only utilized in the clinical field, none of them would be expected to pose significant problems of public or occupational health.
  17. The possible combined action of radiation with compounds known for their carcinogenic properties has been the object of special attention. The substances examined include many initiators and promoters but the systematic information collected for each one of these substances is very incomplete. The evidence reviewed is conflicting and no final statement may be offered in regard to any substance or to any class of tumours before the dose, the schedule of administration and the treatment modalities are analysed to a greater depth, which is seldom the case in the experiments available.
  18. Regarding benzo(a)pyrene and dimethylnitrosamine, two compounds having a widespread diffusion in the environment, experiments on lung tumour induction provided some evidence of a synergistic interaction (expressed mostly through a shorter latency time) for the former, but not for the latter substance. Fairly elaborate experiments in the hamster on the combined effect of radiation, uranium ore dust and diesel oil exhaust fumes yielded no evidence of synergistic effects, but the animal tested could be rather refractory to lung tumour induction. These studies should be extended in view of their practical implications.
  19. Experimental data in animals and epidemiological experience on occupationally exposed human populations is available concerning the combined action of radiation and tobacco smoke. Tumours and inflammatory diseases of the respiratory system have been studied in this respect. In humans it appears that smoke may act by shortening the time of appearance of the radiation-induced lung tumours. It is not yet clear if such an action may be the result of promotion by some component of the tobacco smoke or due to a non-specific effect of the smoke on the respiratory epithelia. The experience in animals is still insufficient for a firm conclusion.
  20. The precise evaluation of an interaction factor in humans critically depends on the length of the observation period as well as on the age structure and exposure history of the populations under study. It is impossible to say if the displacement in time of the tumour appearance will eventually result in an increased final yield of tumours in the smoking as compared to the non-smoking irradiated population. However, even if the final incidence of tumours between smoking and non-smoking irradiated individuals were the same, the effect should still be regarded as a synergistic one, since it would effectively lead to a reduction of the tumour-free life of the smokers developing tumours. This appears to be the only well documented case of a synergistic interaction in humans and in this sense it is a special case.
  21. Antibiotics and other drugs were also considered for their possible interaction with radiation. Variable degrees of synergistic interaction were described for effects ranging from cell survival in vitro to tumour induction in animals. The relevance of these findings to individuals outside the clinical field is however difficult to evaluate, particularly in view of the limited diffusion of these substances in the general environment and of the high doses usually involved in the above interactions.
  22. Possible cases of interactions with biological agents which were considered included those with hormones and with infectious agents. Regarding hormones, there is evidence that a variety of tumours of the experimental animal may be sensitive to their action. Diethylstilbestrol and oestradiol-17-beta were shown to have synergistic interaction for the production of mammary tumours in various strains of rat, with interaction factors in the range of 1.5 to 4. This type of synergism is also expressed through a shortening of the time for tumour induction. There is a large variability between strains, such that the same treatment schedule could produce potentiation in some strains and inhibition in others. There is also variability in relation to the tumour type. Epidemiological information in the human species is scanty and only indirect.
  23. It is difficult for many animal tumours which are known to have a viral etiology (thymic lymphoma, myeloid leukaemia, osteogenic tumours) to consider their induction as the result of a synergistic interaction, because the effect could not be elicited in the absence of either the virus or radiation. There is also no evidence that bacterial infection may play a major role in combination with ionizing radiation in modifying the yield of tumors.
  24. For humans in environmental circumstances the Committee has been unable to document any clear case of synergistic interaction between radiation and other agents, which could lead to substantial modifications of the risk estimates for significant sections of the population. Presumably this is due to the fact that most of the agents likely to act synergistically with radiation, as judged by the results of animal experiments, are not found in sufficient concentration in nature, A specific exception is the case of tobacco smoke, which raises essentially problems of industrial hygiene in some working environments. Further research in the field of the combined effects is desirable because this area of study is still in an early stage of development and could profitably be pursued in a systematic way.

    25. VI. RESEARCH NEEDS
  25. An eminently practical research need is that of modelling experimentally situations encountered in living or working environments to test for undesirable effects. A second important and more basic research need is the identification of interaction mechanisms. The first need is essentially descriptive, the second essentially interpretative and both may interrelate to mutual advantage. There is also a third research need for the monitoring of possible effects in human populations by epidemiological studies. This latter is the most valuable for risk estimates in man.
  26. Experiments of the first type are usually to study in experimental animals situations of practical interest for humans. It should be recalled that results obtained in a given animal species are not easily extrapolated to other species. In designing these experiments, exposure levels should be kept as similar as possible to the modelled situation. In combined action work, the assumption that effects showing at a given dose may exist to a lower degree at lower doses may not be true. Numerous examples of changes in the interaction with changing dose levels of the combining agents exist. The order and rate of administration of the agents should ideally mimic the real situation, although this may be impossible for chronic exposures of interest in practice. Long-term chronic rather than acute end-points should be focused upon. Tumour induction, effects on pre- and post-natal development after exposure in utero and genetic effects are the most significant classes of radio-biological end-points for further studies.
  27. In the more basic studies, frequently involving experiments at the cellular and subcellular levels, there is considerably more latitude for research because the range of end-points is wider and the experiments financially less demanding. Good planning requires the careful choice of experimental end-points and of exposure level.
  28. Epidemiological studies should have high priority under the existing circumstances. The inherent lack of control over many of the exposure variables should be compensated by the best possible definition of the exposure conditions, by the quantitation of the responses and by adequate statistical treatment of the observations. A conceptual and practical distinction should be made between interactions of relevance under special working environments involving possible problems of occupational medicine and large-scale exposure situations which could change risk estimates and could pose therefore more difficult problems of public health.
  29. The use of a standardized nomenclature in the field of combined effects is highly desirable, because too often misconceptions are made possible by inaccurate terminology.
  30. Considering the main technical requirements for experimental investigation of combined effects:
  1. Efforts should be made to report biological data as some function of the exposures in the target structures. For radiation, this problem is relatively simple and studies of energy deposition are reasonably advanced. In other cases (physical agents) this may simply require development of better dosimetric techniques and apparatus, but in most cases (particularly for chemical substances) it will imply detailed studies of the intake, metabolism, concentration and excretion of the inter-acting substances when a direct measure of their concentration at the level of the target structures is not possible;
  2. There is a need to define clearly and specifically the effects to be studied, especially when they are complex ones. For example, overall tumour induction may not in itself be a sufficient indication of a combined action because, even in the absence of significant changes in the overall interaction factor, changes in the spectrum of different tumour classes could take place. In the case of tumours it is important to study the rate of appearance, together with the final incidence, because shifts of the occurrence in time might reveal synergistic actions which would not be apparent otherwise. Also, actuarial and pathological observations should be combined and data corrections for competing risks should be applied;
  3. The variable "time" in the combined actions should be given proper attention, in the sense that contemporaneous and sequential treatments and reversal in the order of application should be examined. These studies are particularly important when the agents under examination have initiating or promoting characteristics and the sequence of their action is therefore decisive. Fractionated and chronic treatments could also be profitably examined, depending on the specific model situation and on the time characteristics of the agents combining;
  4. It is essential that appropriate methodologies of analysis of the interactions be used to avoid mistakes in the interpretation or inaccurate reports of the data. It is only through such objective analyses that precise statements and quantitative evaluations may be drawn. There is, more specifically, a need to refer any given interaction to conditions of iso- and hetero-addition;
  5. Appropriate control series should be set up to test exposure-response curves not only around the exposure levels of interest for the particular experiment but also for an extended range of exposures, including the zero values. Different combinations of exposures of the interacting agents should also be tested; 
  6. It is important that the nature of the interaction should be as much as possible resolved through an analysis of the effects at various levels of biological complexity, from the population level, through the whole-body, tissue, cellular and molecular levels. These studies allow generalizations and avoid misrepresentations of the interaction.

244. The specific areas of work identified by the Committee as particularly important for their basic or practical implications are:

  1. At the molecular and chromosomal levels, studies on the interaction of chemical, physical and viral agents on constitutive and induced processes related to DNA replication and repair of radiation damage (in simple as well as complex organisms) and relevant to the understanding of the mechanism of mutagenesis and to the estimations of genetic risks to man. These studies should concentrate whenever possible on low doses of radiation and of exposure to other agents and be correlated to relevant biological end-points like gene mutations and chromosome abnormalities as well as to cell differentiation (e.g., the immune system, developing organisms) or carcinogenesis;
  2. Studies of the interaction of different types of radiation, particularly for end-points which are of significance for practical purposes;
  3. At the systemic and whole-body level studies of combinations of tobacco smoke, fibres and dusts, organic and inorganic carcinogens and pollutants with radiation would be of great value;
  4. In human populations, further surveys of smoking and non-smoking workers professionally exposed to internal lung irradiations should be pursued. Under special working conditions the study of interaction of radiation with chemicals and microwaves would also be appropriate;
  5. For the population at large, the possible inter-action of hormones and radiation, particularly in human females, should be tested, provided suitable groups might be identified. The increasingly widespread use of contraceptive hormones is of particular importance. 
  6. Studies of combined effects in the treatment of patients (for cancer and other diseases) by combined treatment with radiation and chemo-therapy and hormones, leading to carcinogenesis and to non-stochastic effects which may be "recalled".

 

Table 1 Lung tumours following neutron irradiation and crysotile treatment [L16]

				Number of rats with lung tumours
Group          Number of rats 	Carcinomas	Mesotheliomas
Irradiated           20		    1        	     0
Irradiated +
  Crysotile           9		    4		     3

 

Table 2  Effects on tumour development of prenatal exposure of mice to x rays and ethylnitrosourea (ENU) or to either treatment alone [S42]

			    Number of affected animals a/       	Interaction
Type of tumour          x-irra-      ENU 				factor
			diation   treatment   x rays            	
			(3 x 1 Gy) (0.5 mM/kg) + ENU     Control       	(when applicable)
Leukaemia               3 ( 5.3)   3 ( 2.4)  10 (12.6)   2 ( 2.3)       3.4
Lung tumours            8 (14.3)  22 (17.8)    6 ( 7.6)   11 (12.8)
Hepatomas               2 ( 3.5)   6 ( 4.9)   2 ( 2.5)   1 ( 1.1)       0.23
Pancreatic adenomas     0 ( 0 )    1 ( 0.8)   2 ( 2.5)   0 ( 0 )
Intestinal tumours      0 ( 0 )    2 ( 1.6)   0 ( 0 )    0 ( 0 )
Ovarian tumours         6 ( 7.0)   0 ( 0 )   10 (11.2)   0 ( 0   )      1.6
Total tumour 
  incidence            19 (33.9) 34 (27.6) 30 (38.0) 14 (16.6)          0.44 
Tumour multiplicity
  (affected organs        1.0        1.0        1.5         1.0 
  per animal)
Number of tumours 
  standardized            34         28         57          17          1.1 

a/ The percentage of affected animals is shown in parentheses.

 

Table 3 Respiratory cancer deaths (RCD) in Colorado plateau uranium miners in relation to smoking  [L2] 

		Person-	   
Smoking 	years at     Observed RCD   Expected RCD  0/     0 - E
category 	risk (PYR) 	(0) 	       (E) 	   E     PYR
Smokers 	26392 		60 	      15.5 	  3.9	1.7 10-3
Non-smokers 	 9047 		 2 	       0.5 	  4.0	1.7 10-4

 

Table 4 Interaction factors and probabilities (x 10-4) of respiratory cancer deaths per one person-year at risk for single and combined action of smoking and irradiation 

 

Table 5  Interaction factors and probabilities (x 10-4) of respiratory cancer deaths per one person-year at risk for uranium miners of different smoking categories  [L6]

 

Table 6 Dose modifying factors (DMF) for combined treatment by some chemotherapeutic drugs and radiation   [P5]

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