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![unidentified] gene products results in a range of susceptibilities to ionizing radiation. The notion of individual susceptibilities
cannot be translated into practical action plans and policies in large terrestrial populations at this time. However, a small cohort
of individuals, such as International Space Station astronauts, could be evaluated for individual risk prediction by assessing indi-
vidual response to radiation exposure.
7.09.6.1 Risks Effect Definition
Deterministic effects are those effects that occur only above dose thresholds, for example, acute radiation sickness, damage to the
central nervous system (CNS), and cataracts. The severity of the effect increases with increasing dose. The acceptable risk of a deter-
ministic effect occurring within a radiation worker is zero, and occupational dose limits ensure that these thresholds are not
exceeded.
Stochastic effects are those effects whose probability of occurrence, but not their severity, is a function of the dose. They occur
without a dose threshold. The dose to radiation-induced cancer risk conversion factors is highly dependent on gender, age at expo-
sure, and tissue type. Stochastic effects include cancer, hereditary effects, or neurological disorders.
With regards to hereditary effects, the National Academy of Sciences estimates of the probability that radiation will increase
genetic defects, for a dose of 0.1 Sv, are as follows: first generation of exposedd0.15–0.4Â10À 3
; all generations subsequent to
exposured1.15–2.15Â10À3
(the background rate, for reference, is 3.6–4.6Â10À3
). The risk rate is mainly due to dominant
gene mutations and chromosomal diseases; however, the risks for malformation and growth/developmental retardation are also
high, with most sensitive period being between 8 and 25 weeks gestation. The probability of mental retardation with exposure
during this time period is 0.04 for each dose of 0.1 Sv.
Another means of applying dose limitations is based on the risk of “radiation detriment” (Table 6). This more modern approach
to dose limitation uses the concept of detriment to measure total risk to an individual. Components of overall radiation exposure
detriment are (1) risk of fatal cancer, (2) allowance for relative years of life lost for specific fatal cancers, (3) allowance for morbidity,
and (4) allowance for the risk of hereditary diseases.
Upton estimates of the lifetime risk of cancer incidence and fatality from 1.0 Sv exposure to 10, 000 persons found the highest
incidence in breast (McKenna et al., 1990), lung (ICRP, 1991), and thyroid (Hallahan et al., 1994), but the highest fatality in lung
(Heidenreich et al., 1999) and breast (Fornace et al., 1992; Upton et al., 1986).
7.09.7 Projecting Lifetime Carcinogenesis Risk
Radiation-induced (excess) cancer risks and uncertainties for carcinogenesis can be based, in part, on results of follow-up studies of
Japanese atomic bomb survivors (Petersen and Abrahamson, 1998). The standard linear quadratic exponential (LQE) excess cancer
relative risk (ERR) model based on cell initiation and killing was modified to add a third element, radiation-induced carcinogenesis
at high doses, based on stem-cell repopulation kinetics. (Brenner, 2011) Wheldon and Lindsay, as well as Dasu, et al, developed 2-
stage mutational models which produced a typical bell shaped dose-response curve with diminishing cancer risk at doses 20 Gy,
presumably due to cell sterilization versus transformations (the dose response in a competition model, which takes into account the
probability of cell survival in addition to the probability of induced DNA mutations, after irradiation). A similar curve bending into
a bell-shape was observed by Schneider and Walsh when extending the A-bomb epidemiological data to include exposure dose cate-
gories 4–6 Sv and 6–13 Sv. (Schneider, 2011) A cohort population where this comparison technique can be applied includes
manned spaceflight activities. In the case of space-based radiation exposure, projected risks based on the Japanese data are
Table 6 Contribution to detriment for average worker (average worker dose of 0.1 Sv)
Tissue Fatal cancer (%) Hereditary Aggregated detriment (%)
Bladder 0.024 0.024
Bone marrow 0.04 0.083
Breast 0.016 0.029
Colon 0.068 0.082
Liver 0.012 0.013
Lung 0.068 0.064
Esophagus 0.024 0.019
Ovary 0.008 0.012
Skin 0.002 0.003
Stomach 0.088 0.080
Thyroid 0.006 0.012
Gonads 0.06 0.080
Other 0.045 0.053
Total 0.4 0.56
212 Ionizing Radiation as a Carcinogen
Comprehensive Toxicology, Third Edition, 2018, 183–225
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![Where xD is a quantile for bias correction of random and systematic errors in DS86, xT is a quantile for bias correction in transfer
of risk from Japan to the United States, xP is a quantile for bias correction of uncertainty in projection over a lifetime, xQ is a quantile
for bias correction of unknown uncertainty, and xE is the uncertainty distribution for the dose and dose-rate effective factor. After
5000 iterations are performed, the various quantiles of xmix are ranked in ascending order. Central estimates of risk are then based
on the median, m(xmix), with 90% subjective confidence limits based on the 5th and 95th percentiles of the ranked quantiles. The
process of randomly selecting either the additive, NIH-multiplicative, or multiplicative model for life table calculations during each
interval is the basis for the “mixture model” of risk.
The probability of causation (PC), at various ages, has been estimated for leukemic and nonleukemic cancers (all cancers except
leukemia) for males exposed to 1 Sv of radiation, accounting for age of exposure. From the estimates, the greatest excess risk of
5.59% occurs at age 90 for males exposed to 1 Sv at age 20. Overall, risk decreases with increasing age at exposure and approaches
1% for males exposed at age 90. The median risk of leukemia among males showed a similar trend but was greatest for exposure at
age 40, where it approached 4.3Â10- 3
at age 90. The effect of the short 40-year plateau period during which leukemia risk is
accumulated can be noticed in the curve for males exposed at age 20. When exposed at age 20, risk accumulation starts at age
22 (20 þ 2-year latency) and continues until age 62 (20 þ 2 þ 40). Cumulative risk of leukemia from exposure at older ages is
higher than risk from exposure at younger ages because the excess absolute leukemia risk is added to baseline rates. Among US
males, baseline rates of leukemia at ages 60–64 are one-third the baseline rates at ages 75–79 and less than a fourth of the rates
beyond age 85. If plateau periods for leukemia were longer than 40 years, a different picture would emerge (Aguiar, 1998; Preston,
2005; Preston et al., 1994).
The highest PC for nonleukemic cancers was 31.3%, and this occurred at age 40 among those exposed at age 20. For leukemia,
the PC at age 30 among those exposed at age 20 was 95.4%. While PCs for nonleukemic cancers decrease uniformly with increasing
age at exposure, PCs for leukemia follow the same trend as exposure age increases and remain fixed after the end of the plateau
period in which risk is accumulated. For females (Fig. 8), the trends in nonleukemic malignancy risk at various ages, accounting
for age at exposure, were similar to those observed for males. Leukemia risks for females, based on the sum of excess absolute risks
and baseline risk, are generally lower than those for males because the female risk coefficients are lower (see Fig. 4 of Preston et al.,
1994) and female baseline rates are about half of those for males [at all ages].
Some general conclusions on the risks presented from the PC and ERR estimates are as follows: For males, the highest nonleu-
kemic cancer risks observed for exposure at ages 20, 30, 40, 50, 60, and 70 were 5.6% at age 90, 4.3% at age 90, 3.3% at age 100,
2.4% at age 100, 1.6% at age 100, and 1.1% at age 100, respectively. For females, the highest nonleukemic cancer risks observed for
exposure at ages 20, 30, 40, 50, 60, and 70 were 4.8% at age 80, 5.4% at age 100, 4.1% at age 90, 2.9% at age 90, 2.0% at age 90, and
1.3% at age 100, respectively. PCs for leukemia approached 95% for males exposed at age 20 and 90% for females exposed at age
20; for nonleukemic cancer, PCs approached 30% for exposure at age 20. Lifetime cancer incidence risks approaching 3% for expo-
sure to 1 Sv could be expected for males exposed at age 40 and females exposed at age 50.
7.09.8 Radiation Protection from Ionizing Radiation
7.09.8.1 Shielding
Radiation protection aims to mitigate or eliminate the hazardous effects of ionizing radiation on human health and well-being. In
principle, this can be achieved by physical shielding, by setting acceptable exposure limits, and by radioprotective agents and/or
therapy. There are three means to reduce exposure to ionizing radiation: (1) increasing distance from the radiation source, (2) mini-
mizing the time of exposure, and (3) using shielding.
Fig. 8 Site-specific excess relative risk (ERR) and 90% confidence intervals, expressed in terms of risk for females aged 30 at exposure.
214 Ionizing Radiation as a Carcinogen
Comprehensive Toxicology, Third Edition, 2018, 183–225
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Comprehensive toxicology: Ionized Radiation as Carcinogen.
This document provides the full text of a chapter from the book "Comprehensive Toxicology" on ionizing radiation as a carcinogen. The chapter is copyrighted material provided for non-commercial educational use. It discusses the mechanisms of radiation damage at cellular and molecular levels, evidence of radiation-induced cancer from human populations and animal/in vitro studies, and models for assessing radiation cancer risk.
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- 1. Provided for non-commercialresearch and educational use. Not for reproduction, distribution or commercial use. This article was originally published in Comprehensive Toxicology, 3e, published by Elsevier, and the attached copy is provided by Elsevier for the author's benefit and for the benefit of the author's institution, for non-commercial research and educational use including without limitation use in instruction at your institution, sending it to specific colleagues who you know, and providing a copy to your institution's administrator. All other uses, reproduction and distribution, including without limitation commercial reprints, selling or licensing copies or access, or posting on open internet sites, your personal or institution’s website or repository, are prohibited. For exceptions, permission may be sought for such use through Elsevier’s permissions site at: https://www.elsevier.com/about/our-business/policies/copyright/permissions From Jones, J. A., Karouia, F., Cristea, O., Casey, R. C., Popov, D., & Maliev, V. (2018) Ionizing Radiation as a Carcinogen. In: McQueen, C. A., Comprehensive Toxicology, Third Edition. Vol. 7, pp. 183–225. Oxford: Elsevier Ltd. ISBN: 9780081006016 Copyright © 2018 Elsevier Ltd. All rights reserved. Elsevier Science Author's personal copy
- 2. 7.09 Ionizing Radiationas a Carcinogenq JA Jones, Baylor College of Medicine- Center for Space Medicine, Urology, Houston, Texas, USA F Karouia, NASA Ames Research Center, Moffitt Field, California, USA O Cristea, University of Ottawa, Ottawa, Ontario, Canada RC Casey, University Space Research Associates, NASA- Johnson Space Center, Houston, TX, USA D Popov, Advanced Medical Technologies & Systems, Richmond Hill, Ontario, Canada V Maliev, Vladicaucasian Scientific Center of Russian Academy of Sciences, Vladicaucas, North Ossetia–Alania, Russia © 2018 Elsevier Ltd. All rights reserved. 7.09.1 Introduction 190 7.09.2 Radiation Spectrum 191 7.09.2.1 Units 192 7.09.2.2 Dose and Dose Rate 192 7.09.2.3 Biophysical Damage Mechanisms 193 7.09.2.4 Atomic Ionization 195 7.09.3 Radiobiology 196 7.09.3.1 Primary Versus Secondary Damage 196 7.09.3.2 Mechanisms of Damage From Ionizing Radiation 196 7.09.3.3 Direct DNA Damage 196 7.09.3.4 Indirect DNA Damage or Oxidative Injury 198 7.09.3.5 Epigenetic Effects 198 7.09.3.6 Bystander Effects 198 7.09.3.7 Adaptive Response 199 7.09.4 Evidence for Radiation Carcinogenesis 200 7.09.4.1 Human Data 200 7.09.4.2 Atomic/Nuclear Weapon Survivors 200 7.09.4.3 Industrial Accidents 201 7.09.4.3.1 Chernobyl and JCO nuclear criticality accident 201 7.09.4.3.2 Techa River Basin 202 7.09.4.4 Medical Radiation Therapy and Cancer Risk 202 7.09.4.5 Medical Conditions Predisposing to Radiation-Induced Cancer 203 7.09.4.6 Radiation Carcinogenesis in Animal Models 203 7.09.4.7 In Vitro Evidence for Radiation Carcinogenesis 204 7.09.4.7.1 Chromosome Aberrations 204 7.09.4.7.2 Cell Transformation 205 7.09.4.7.3 Mutagenesis in Somatic Cells 206 7.09.4.7.4 Gene Expression 206 7.09.4.7.5 Biological Effects Occurring in Nonirradiated Cells 207 7.09.5 Molecular Biology of Radiation Carcinogenesis 207 7.09.5.1 Cell Cycle Delay and Gene Induction 207 7.09.5.2 Oncogenes in Radiation Carcinogenesis 208 7.09.5.2.1 Ras genes 209 7.09.5.2.2 c-myc gene 209 7.09.5.2.3 bcl-2 genes 209 7.09.5.2.4 erbB2 genes 210 7.09.5.2.5 Oncogenes in Radiation Resistance 210 7.09.5.3 Tumor Suppressor Genes 210 7.09.5.3.1 BRCA1/2 genes 210 7.09.5.3.2 p53 genes 211 7.09.5.4 Interactions Between Oncogenes and Tumor Suppressor Genes in Radiation-Induced Cancer 211 7.09.5.5 Genomic Instability 211 7.09.6 Radiation Carcinogenesis Risk Modeling 211 7.09.6.1 Risks Effect Definition 212 7.09.7 Projecting Lifetime Carcinogenesis Risk 212 q Change History: February 2017, JA Jones and F Karouia updated the text and the references. This is an update of JA Jones, RC Casey, F Karouia, Ionizing Radiation as a Carcinogen, Comprehensive Toxicology, Second Edition, edited by Charlene A. McQueen, Elsevier, Oxford, 2010, Volume 14, Pages 181–228. 183 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 3. 7.09.8 Radiation Protectionfrom Ionizing Radiation 214 7.09.8.1 Shielding 214 7.09.8.2 Dose Limitation 215 7.09.8.3 Radioprotectants/Chemoprevention 215 7.09.8.4 Immunoprotection Strategies 216 7.09.9 Conclusions 216 References 219 Glossary Absorbed dose Mean energy imparted by ionizing radiation to an irradiated object per unit mass. Units: gray, rad. Thus, absorbed dose is the amount of energy deposited by a radiation in a quantity of material (e.g., tissue) through ionization and excitation. Units of measurement are rad/mrad or Gy/mGy (SI unit). 1 Gy¼100 rad or 10 mGy¼1 mrad. Absorbed dose is a useful parameter for determining the likelihood and severity of acute radiation effects. A typical chest X-ray yields $10 mrad of dose. Acute effects of radiation Effects that occur shortly after exposure to radiation, usually within a week. These acute effects result from exposure to radiation at relatively high doses, usually greater than 1 Gy. They are usually due to the killing cell in critical tissues in the body. Alpha radiation Type of particulate radiation that is released from a radioactive atom. Alpha particles are helium nuclei. Alpha particle radiation is densely ionizing (high LET) radiation and can be very damaging, but it is very limited in its ability to penetrate tissue. For example, alpha radiation will not penetrate the outer layers of the skin. Background radiation Radiation received by the entire human population due to natural occurring radiation and radioactive materials in the environment. The three major sources of natural background radiation are (1) cosmic radiation, (2) radiation from naturally occurring radioactive elements in the Earth’s surface, and (3) internal radiation arising from radioactive atoms normally present in foodstuffs or in the air. Cell transformation A process by which cells in vitro, which have a limited ability to divide, are altered by radiation or chemicals so as to have an unlimited division potential. Comet assay Comet assay, also called single cell gel electrophoresis (SCGE), is a sensitive and rapid technique for quantifying and analyzing DNA damage in individual cells. As such, this is one of the techniques used in the area of cancer research for the evaluation of genotoxicity and effectiveness of chemoprevention. Swedish researchers Östling and Johansson developed this technique in 1984. Singh et al. later modified this technique, in 1988, as the alkaline comet assay. The resulting image that is obtained resembles a “comet” with a distinct head and tail. The head is composed of intact DNA, whereas the tail consists of damaged (single-strand or double-strand breaks) or broken pieces of DNA. While most of the applications of the comet assay have been to study animal eukaryotes, there have been reports of successful application in the study of plant cells. Individual cells are embedded in a thin agarose gel on a microscope slide. All cellular proteins are then removed from the cells by lysing. The DNA is allowed to unwind under alkaline/neutral conditions. Following the unwinding, the DNA undergoes electro- phoresis, allowing the broken DNA fragments or damaged DNA to migrate away from the nucleus. After staining with a DNA- specific fluorescent dye such as ethidium bromide or propidium iodide, the gel is read for amount of fluorescence in head and tail and length of tail. The extent of DNA liberated from the head of the comet is directly proportional to the amount of DNA damage. Cross section Probability of a nuclear interaction within a defined region of a more stable material. Likewise, the chloride atom has seven electrons in the outermost electron orbit. Gaining one electron fills the orbit with eight electrons, which makes the atom more stable. Thus, the give and take of selected electrons is ordinarily involved in the formation of ions. Curie Unit of measurement of radioactivity. In modern nuclear physics, the curie is precisely defined as the amount of a substance in which 37 billion atoms per second undergo radioactive disintegration. A curie was originally defined as the amount of radon in equilibrium with 1 g of radium, or the amount of any radioactive substance that undergoes the same number of radioactive disintegrations in the same time as 1 g of radium. When it is defined this way, the unit is still used as a measure of radiation dosage in medicine. The Becquerel is the preferred unit of measure for radioactivity in SI units. One curie equals 3.7Â10À10 Bq. Deterministic effects Effects formally known as nonstochastic effects that may appear early or late after irradiation. There is a threshold dose above which both the probability of occurring and the severity of the effects increase. Most deterministic effects involve cell killing. DNA fingerprint Some minisatellites share a repeating unit similar in base sequences. Such minisatellites can be examined using a common probe and 10–20 minisatellite bands of different sizes are detected on a Southern filter. The band pattern is extremely variable among different individuals and is quite useful for molecular identification of individuals, thus the term DNA fingerprints. Dose See absorbed dose. 184 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 4. Dose effect (dose–response)model Mathematical formulation used to predict the magnitude of an effect that would be produced by a given dose of radiation. Dose equivalent Amount of biologically effective dose deposited by an ionizing radiation. Dose equivalent is only valid for assessing the risk of cancer or genetic effects in humans as a result of radiation exposure. It is computed by multiplying the absorbed dose by a radiation species-dependent quality factor. Quality factors can range from 1 to 20. Units of measurement are rem/mrem or Sv (SI unit). 1 Sv¼100 rem or 10 mSv¼1 mrem. A typical chest X-ray yields $10 mrem of dose equivalent. Dose rate Quantity of absorbed dose delivered per unit of time. Dose rate effectiveness factor (DREF) Factor by which the effect caused by a specific dose of radiation changes at low dose rates as compared with high dose rates. Energetic solar particle event Events characterized by >100 MeV proton flux greater than or equal to 1 p.f.u. at geosynchronous orbit as measured by a GOES satellite. Energetic SPE flux can range from $1 to 550 p.f.u. (or higher). Energetic SPEs can persist from several hours (very small events) to several days. Energetic SPEs are believed to result from the propagation of fast interplanetary shocks formed by CMEs. eV Kinetic energy of charged particles, which is measured in units of eV. Multiples of eV are frequently used. These include: keV ¼ 1000 eV; MeV ¼ 1000000 eV; GeV ¼ 1000000000 eV, Excess risk Refers to the excess cases of a particular health effect associated with exposure to radiation. Excess risk can be described in various ways including the (1) difference in the rate of occurrence of a particular health effect in an exposed population and an equivalent population with no radiation exposure (rate difference or absolute risk, the unit for which is excess number of cases per person-year-sievert); (2) ratio of the rates in exposed and unexposed populations (relative risk, a unit of no dimension); and (3) ratio of the rate difference to the rate in an unexposed population (excess relative risk, i.e., relative risk minus 1). Excess risk can depend on various factors including radiation dose, age at exposure, time since exposure, current age, and sex. Assessments of the risk among the atomic bomb survivors are carried out using regression methods, and risk estimates are usually reported for a specific dose (often 1 Gy or 1 Sv) for a specified combination of other important factors. In general, it is not possible to summarize the excess risk associated with a single factor. Fluence Rate at which particles are incoming, measured in number of particles per second. Flux Density at which particles are incoming, measured in number of particles per square centimeter. Gamma rays Short-wavelength EMR of nuclear origin with an energy range of about 10 keV–9 MeV. Gray (Gy) Quantity of energy imparted by ionizing radiation to a unit mass of matter. A gray (abbreviated as Gy) is the amount of energy deposited in tissues; technically, one joule of energy per kilogram of tissue. When ionizing radiation passes through a body, some of its energy is imparted to the surrounding tissue. The average cumulative radiation doses from natural, medical, and occupational sources are estimated to be about 2 mGy (or 0.002 Gy) per year for humans. Gray is used in the SI system, the term “rad” used in the USA, is replaced by gray (Gy) and is defined as: 1 Gy ¼ 1 J kgÀ1 Since 1 rad ¼ 10À2 J kgÀ1 ; 1 Gy ¼ 100 rad High-Z, high energy (HZE) particles Heavy (high atomic number), high energy particles, such as carbon or iron nuclei, with an energy range in cosmic rays between approximately 102 and 103 MeV per nucleon. Ionization An ion is an atom or a molecule that has an unequal positive and negative charge. The negative electrical charge is derived from electrons and the positive charge is derived from protons in the nucleus. An example of an ion is sodium chloride (NaCl). When NaCl is dissolved in water, sodium and chloride separate to form two kinds of ions: Naþ and ClÀ . The sodium loses one electron and the chloride gains one electron. Why does this happen? Because the sodium atom has a single electron in the outermost electron orbit, losing it makes the atom more stable. Likewise, the chloride atom has seven electrons in the outermost electron orbit. Gaining one electron fills the orbit with eight electrons, which makes the atom more stable. Thus, the give and take of selected electrons is involved in the formation of ions ordinarily. Isotropic Radiation with the same magnitude (values) from all axes (directions). JCO On Sep. 30, 1999, a criticality accident occurred at the Tokai nuclear fuel plant in Japan. The plant operated by JCO Co. Ltd, a 100% subsidiary of Sumitomo Metal Mining Co. Ltd, converts enriched uranium hexafluoride (UF6) to uranium dioxide (UO2) for use in nuclear fuel. Criticality accidents involve a self-sustaining chain reaction caused from handling of too large amounts of enriched uranium. The chain reaction continued for around 20 h, before it could be stopped.The uranium pro- cessed was enriched to 18.8% of uranium-235, rather than the 3%–5% used for commercial light water reactor fuel. Material of this high enrichment grade was being produced for the experimental Joyo Fast Breeder reactor.The plant has an annual production capacity of 715 tonnes of uranium for light water reactors and 3 tonnes of uranium for fast breeder reactors.The chain reaction caused heavy releases of gamma and neutron radiation. Three workers were exposed to doses of up to 17 Sv (sieverts), causing severe radiation sickness. The worker exposed to the highest dose died on Dec. 21, 1999. The worker exposed Ionizing Radiation as a Carcinogen 185 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 5. to the secondhighest dose of 6–10 Sv died on Apr. 27, 2000. Sixty-eight other persons were irradiated at lower levels. Among them were the workers who stopped the chain reaction: they were exposed to doses of up to 119.79 mSv, exceeding the 100 mSv limit for emergency situations. The annual dose limit for workers is 50 mSv (while ICRP currently recommends 20 mSv). As of Oct. 7, 1999, radiation levels remained high inside the plant building, preventing inspection of the damage inside the plant.On Nov. 4, 1999, the Science and Technology Agency submitted a report to the Nuclear Safety Commission, containing estimates of radiation exposures during the criticality accident, based on an analysis of uranium solution sampled from a tank inside the fuel processing plant and the neutron levels monitored around the plant. The report says that1. the radiation dose received at a distance of 80 m from the accident site (that is the nearest boundary of the plant) was estimated at 75 mSv (this figure was revised to 30 mSv, according to Yomiuri Shimbun of Dec. 12, 1999; details are not known yet) for the first 25 min of the criticality accident and 160 mSv for the whole criticality period of 20 h; and 2. the radiation dose received at a distance of 350 m from the plant (that was the evacuation boundary) was estimated at almost equal to the annual permissible dose of 1 mSv for the first 25 min and 2 mSv for the whole event.Since the evacuation started only 5 h after the beginning of the criticality accident, residents may have received doses of more than 75 mSv, that is, 75 times the permissible annual dose of 1 mSv.The government has so far said 69 people were exposed to radiation, but the latest survey says the number of affected people could increase. LET Quantifies the amount of energy deposited per unit length of particle track. This factor increases with the square of the charge and is inversely proportional to the energy of the radiation particle. The influence of the electric force field depends on the velocity of the particle. Slower moving charged particles will produce more ionizations per unit path length than faster moving ones. LET accounts for all energy transfers along the particle path, irrespective of mechanism. Thus, LET of protons, deuterons, alpha particles, and other heavy, charged particles lose kinetic energy rapidly as they penetrate matter. Most of this energy is lost as the particles interact inelastically with electrons of the absorbing medium. Energy transferred to an electron in excess of its binding energy appears as kinetic energy of the ejected electron. An ejected electron and the residual positive ion constitute an ion pair; for example, an average energy of 33.7 eV, termed the W-quantity or W, is expended by charged particles per ion pair produced in air. The specific ionization (SI) is the number of primary and secondary ion pairs produced per unit length of path of the incident radiation. LET is the average loss in energy per unit length of path of the incident radiation. The LET is the product of the specific ionization and the W-quantity: LET ¼ SIð Þ Wð ÞLET ¼ SIW where SI is the specific ionization and W is the W-quantity of the medium.The range of ionizing particles in a particular medium is the straight line distance traversed by the particles before they are stopped completely. For heavy particles with energy E, the range in a particular medium may be estimated from the average LET: range ¼ E=LETrange ¼ E=LET The SI and LET are not constant along the entire path of monoenergetic charged particles traversing a homogeneous medium. The increase in SI near the end of the path of the particles reflects the decreased velocity of the particles as they traverse the medium. As the particles slow, the SI increases because nearby atoms are influenced for a longer period. The region of increased SI is termed the Bragg Peak. Linear dose response Relationship between dose and biological response, which is a straight line. In other words, the rate of change (slope) in the response is the same at any dose. A linear dose response is written mathematically as follows: If Y represents the expected or average response and D represents dose, then Y ¼ aDY ¼ aD where a is the slope, also called the linear coefficient. Linear quadratic dose response Relationship between dose and biological response that is curved. This implies that the rate of change in response differs at different doses. The response may change slowly at low doses, for example, but rapidly at high doses. A linear quadratic dose response is written mathematically as follows: If Y represents the expected or average response and D represents dose, then Y ¼ aD þ bD2 Y ¼ aD þ bD2 where a is the linear coefficient (or slope) and b is the quadratic coefficient (or curvature). microsatellite The genome contains sequences consisting of two to five base repeats, shorter than minisatellite core sequences, which are thus called microsatellites. In contrast to minisatellites, microsatellites distribute evenly throughout the genome and are frequently used as landmarks for gene mapping. Human hereditary diseases caused by unusual expansion of three base repeats are now known (e.g., CGG repeats in fragile X syndrome, CAG repeats in Huntington’s disease). The number of repeats varies among normal individuals, as well as among family members of the affected individuals and even among somatic cells of an individual. The inheritance pattern of the repeats does not seem to follow Mendelian rules. Patients of hereditary nonpolyposis colon cancer are known to show instability in microsatellite repeat number. Radiation effects on microsatellite instability are not known. Monte Carlo Statistical method that evaluates a probability distribution by means of random sampling. 186 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 6. Multiplicative interaction modelA model in which the relative risk (the relative excess risk plus 1) resulting in exposure to two risk factors is taken to be the product of the relative risks from the two factors taken separately. Neutron Uncharged subatomic particle capable of producing ionization in matter by collision with charged particles. Nonstochastic effect See deterministic effects. Nuclide Species of atom characterized by the constitution of its nucleus, which is specified by the atomic mass (M) and atomic number (Z). Organ dose Used interchangeably at the Radiation Effects Research Foundation (RERF) with the terms “mean organ dose” or “dose.” It is defined as the energy absorbed in a specific organ divided by its mass, that is, the average energy absorbed per unit mass. This quantity is expressed in gray or its submultiples. The organ dose calculation allows for effects of external shielding and shielding provided by other tissues. Photon Unit of EMR. One g-ray is a photon of EMR. Quality factor (Q) Function of the particle LET, as determined by the charge and energy of radiation particles. This factor, which accounts for differences in the biological effectiveness of different particles, is used to convert an absorbed dose into a dose equivalent. The Q is based on observations of RBE. ICRP, 1991 defined Q for use in risk (R) estimates for radiation protection practice as R¼k2QÂD/DREF, where k is the dose rate risk coefficient for high-rate g-rays and D is the adsorbed dose. H is the term for dose equivalent, where H¼QD. The unit for H is rem or Sv. Current values for the quality factor may range from 1 to 20. X-rays are assigned a value of unity (1). Quality factor values as high as 100 may be deemed appropriate as additional research continues. Rad (United States)/gray (international) The absorbed dose, rad, is the amount of energy absorbed from radiation per mass of material (1 rad¼100 ergs gÀ 1 ). The SI unit for absorbed dose is the gray (1 gray or Gy¼100 rad). Chemical and biological changes in tissue exposed to ionizing radiation depend upon the energy absorbed in the tissue from the radiation rather than upon the amount of ionization that the radiation produces in air. To describe the energy absorbed in a medium from any type of ionizing radiation, the quantity of radiation should be described in units of rads. The rad, which was originally an acronym for radiation absorbed dose, is a unit of absorbed dose and represents the absorption of 10À 2 Joules of energy per kilogram (or 100 ergs gÀ1 ) of absorbing material: 1 rad ¼ 10À2 J kgÀ1 ¼ 100 ergs gÀ1 The absorbed dose D in rads delivered to a small mass m in kilograms is D rad ¼ E=m Ä 10À2 J kgÀ1 where E, the absorbed energy in joules, is “the difference between the sum of the energies of all directly and indirectly ionizing particles which have entered the volume and the sum of the energies of all those which left it minus the energy equivalent of any increase in rest mass that took place in nuclear or elementary particle reactions within the volume.” This definition means that E is the total energy deposited in a small volume of irradiated medium, corrected for energy removed from the volume in any way. Radiation weighting factor (Wr) Factor, established by consensus and used in radiation protection, to weight the absorbed dose averaged over an organ to obtain the equivalent dose on a common scale for types of ionizing radiation. Radioisotope Radioactive species of an element with the same atomic number and identical chemical properties. Radionuclide Radioactive species of an atom characterized by the constituents of its nucleus (atomic number). Radiosensitivity Relative susceptibility of cells, tissues, organs, and organisms to the injurious action of radiation. Radiosensitivity and its antonym, radioresistance, are used in a comparative sense rather than an absolute sense. Relative biological effectiveness (RBE) Chemical and biological effects of irradiation depend not only on the amount of energy that is absorbed in an irradiated medium but also on the distribution of absorbed energy within the medium. For equal absorbed doses, various types of ionizing radiation often differ in the efficiency with which they illicit a particular chemical or biological response. RBE describes the effectiveness or efficiency with which a particular type of radiation evokes a certain chemical or biological effect. It is computed by comparing results obtained with the radiation in question to the results obtained with a reference radiation (e.g., medium energy X-rays): RBE ¼ dose of reference radiation=dose of experimental radiationð Þ to produce the same end point. For a particular type of radiation, the RBE may vary from one chemical or biological response to another. The RBE dose in rem is the product of the RBE and the dose in rad. RBE dose remð Þ ¼ absorbed dose radð Þ Â RBE The concept of RBE dose should be limited to the discipline of radiation biology. Relative risk Expression of risk relative to the underlying (baseline) risk. If the relative risk is 2, the excess risk equals the baseline risk. Ionizing Radiation as a Carcinogen 187 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 7. rem (United States)/sievert(international) The biological equivalent dose, rem (Roentgen equivalent man), is the absorbed dose adjusted for biological effectiveness of the particular type of radiation. It is the product of the absorbed dose and the quality factor. The SI unit for biological equivalent dose is sieverts (1 sievert or Sv¼100 rem): rem ¼ rad  quality factor sievert ¼ gray  quality factor Replication Cellular process of DNA synthesis (duplication) from DNA template. It involves unwinding of the DNA helix and separation of the strands to expose the complementary base for nucleotide addition, one at a time, and requires the enzyme DNA polymerase in eukaryotes. Risk coefficient Increase in the incidence of disease or mortality per person exposed per unit equivalent dose. The relative risk coefficient is the fractional increase in the baseline incidence or mortality rate for a unit dose. Risk estimate Number of cases (or deaths) projected to occur in a specified exposed population per unit of collective dose for a specified exposure regime and expression period, for example, number of cases per person (in gray). Roentgen One of the oldest units of radiation measurement (W. K. Roentgen discovered X-rays in 1895). The Roentgen is a measure of the quantity of ionization induced in air from radiation exposure. Sievert (Sv) In the SI system, the sievert (Sv) replaces the rem (1 Sv¼100 rem). Often, the effectiveness with which the different types of radiation produce a particular chemical or biological response varies with the LET of the radiation. The dose equivalent (DE) in rem is the product of the dose in rad and a quality factor (QF) that varies with the LET of the radiation: DE remð Þ ¼ D radð Þ Â QF The dose equivalent reflects a recognition of differences in the effectiveness of different radiations to inflict overall biological damage and is used in computations associated with radiation protection and safety. The sievert unit of radiation dose is used for radiation protection purposes. When an individual is exposed to mixed sources of radiation, the total biologically effective dose is calculated by multiplying the physical dose (expressed in units called gray) of each kind of radiation by a corresponding factor (the Q-factor) specified for the type of radiation and its energy, after which these amounts are summed. The factor for g- rays is 1; therefore, 1 Sv¼1 Gy. The factor for the neutrons in atomic bomb radiation is 10; therefore, 1 Sv¼0.1 Gy. Solar flare Sudden release of energy across the electromagnetic spectrum from a relatively small region of the Sun. Flares are important because they are believed to be a manifestation of solar processes that produce SPEs. Ground-based observatories and the GOES continuously monitor for solar flares. Flares are often characterized by brightening in an optical wavelength (H- a, a red wavelength) or soft X-rays (1–8 Å). NOAA characterizes flares by their soft X-ray intensity as measured by the GOES X- ray detector. Flares are classified on a logarithmic scale using B, C, M, and X designations for very small, small, moderate, and large X-ray flares. Solar particle event (SPE) Events characterized by the >10 MeV proton flux greater than or equal to 10 p cmÀ 2 - sÀ1 srÀ1 (p.f.u.) at geosynchronous orbit as measured by a GOES. SPE flux can range from $10 to 40 000 p.f.u. (or higher). SPEs, which can persist from several hours (very small events) to a couple of weeks, are believed to result from the propagation of moderate to fast interplanetary shocks formed by CMEs. Stochastic effects Random events leading to effects whose probability of occurrence in an exposed population of cells or individuals (rather than severity in an affected cell or individual) is a direct function of dose. These effects are commonly regarded as having no threshold. Hereditary effects are regarded as being stochastic. Some somatic effects, especially carcinogenesis, are regarded as being stochastic. Target theory (hit theory) Explains some biological effects of radiation on the basis that ionization, which occurs in a discrete volume (the target) within a cell, directly causes a lesion that later results in a physiological response to the damage at that location. One, two, or more hits (ionizing events within the target) may be necessary to elicit this response. Threshold dose Dose level below which there is no effect of radiation on the biological response. It is often difficult to distinguish between a threshold and a linear quadratic dose response, where the response changes only slightly at low doses. A threshold model postulates that radiation does not cause the effect at any level below the threshold. Radiation thresholds are generally thought to be limited to acute (short-term) effects that are called deterministic, because they require depletion of certain cells in the body to below a critical number in a given organ or tissue. These effects include radiation sickness (nausea and vomiting), infection and bleeding, and loss of hair. Transcription Cellular process of making RNA from the DNA template. This type of RNA is called messenger RNA (mRNA). Translation Cellular process of synthesizing proteins from the mRNA template in ribosomes. Transport calculation Calculation of particle distributions and energy behind a specific shield. Transportation calculation is derived from the basic nuclear cross sections for interaction and fragmentation in shielding. 188 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 8. Abbreviations AP Apurinic/apyrimidic ASA Acetylsalicylate ATAtaxia telangiectasia ATM Ataxia telangiectasia mutated bFGF Basic fibroblast growth factor CI Confidence interval CDKN1A Cyclin-dependent kinase inhibitor 1A CNS Central nervous system CSF Colony-stimulating factor DDC Diethyldithiocarbamate DMSO Dimethylsulfoxide DNA Deoxyribonucleic acid DNA-PK DNA-dependent protein kinase DREF Dose rate effectiveness factor DSB Double-strand break EGCG Epigallocatechin gallate EGF Epidermal growth factor EGR-1 Epidermal growth factor receptor EMR Electromagnetic radiation ERR Excess relative risk FGF Fibroblast growth factor FISH Fluorescence in situ hybridization GCR Galactic cosmic radiation GJIC Gap junction intercellular communication GM-CSF Granulocyte/macrophage colony-stimulating factor GPX Glutathione peroxidase Gy Gray HGF Hepatocyte growth factor HNSCC Head and neck squamous cell carcinoma HO2* Peroxy radical HPRT Hypoxanthine-guanine phosphoribosyl transferase HZE High Z, high energy HZETRN High Z, high energy transport code IAEA International Atomic Energy Agency ICRP International Commission on Radiological Protection IGF-1 Insulin-like growth factor-1 IL Interleukin IR Infrared ISS International Space Station JCO JCO Company Limited (a company operating the Tokai nuclear fuel plant in Japan, where a radiation release accident occurred in 1999) LD50 Median lethal dose (lethal for 50% of test subjects) LET Linear energy transfer LLR Long-lived organic radical LSS Life Span Study MAP Mitogen-activating protein MW Microwave NAS National Academy of Sciences NCI National Cancer Institute NCRP National Council for Radiation Protection and Measurement NHEJ Nonhomologous end joining NIH National Institutes of Health NSAIA Nonsteroidal anti-inflammatory agent Ionizing Radiation as a Carcinogen 189 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 9. OH* Hydroxyl radical OSHAOccupational Safety and Health Administration PARP Poly(ADP-ribose) polymerase PC Probability of causation PCNA Proliferating cell nuclear antigen PGE2 Platelet granule extract 2 PKC Protein kinase C PYSv Person-year sieverts rad Radiation absorbed dose RBE Relative biological effectiveness REID Risk of exposure-induced death rem Roentgen equivalent man RNA Ribonucleic acid RNS Reactive nitrogen species ROS Reactive oxygen species RR Relative risk SCE Sister chromatid exchange SI International unit of measurement SRD Specific radiation determinant SSB Single-strand break TNF Tumor necrosis factor UV Ultraviolet VEGF Vascular endothelial growth factor XRT X-ray telescope Nomenclature l Wavelength c Speed of light H Dose equivalent h Planck’s constant Hz Hertz k Dose rate risk coefficient for high rate g-rays Sv Sievert f Frequency 7.09.1 Introduction Ionizing radiation is generally considered to be any form of radiation exposure that will produce a subatomic ionization event. Ionizing radiation may be in the form of either electromagnetic waves or particles; however, it must have sufficient energy to cause ionization in the target molecule. For electromagnetic radiation (EMR), the wavelength is usually shorter and therefore the energy is higher than for nonionizing radiation; thus, ionizing radiation is more likely to produce a biological effect. Ultraviolet (UV) EMR is intermediate in wavelength and energy and is typically considered as nonionizing; however, it is clearly mutagenic, can produce ionization, and should be considered as a carcinogen. (Nonionizing radiation is discussed elsewhere in the textbook.) Although, perhaps, recognized later than its chemical counterparts, ionizing radiation is now regarded as a carcinogen and can act independently or synergistically with other carcinogens to produce neoplasia in living systems via its unique mechanisms of muta- tion and biological effect. This article discusses the carcinogenic nature of ionizing radiation. This article will first describe the physical interaction between the different forms of ionizing radiation and cellular and subcel- lular components, as well as the factors that are likely to produce an elevated risk of neoplasia. Next, the biological and molecular effects of radiation within living systems will be examined, followed by a presentation of the epidemiological evidence for radiation as a carcinogen in animals and humans. Some risk models for carcinogenesis following an exposure to ionizing radiation, as well as some strategies for protection against radiation-induced biological damage, are included. Important new avenues of research and some of the controversial issues surrounding radiation carcinogenesis are brought forth. Finally, the significance of ionizing radiation-induced carcinogenesis to the understanding and management of the broader issue of the etiology of human cancer will be addressed. 190 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 10. 7.09.2 Radiation Spectrum Radiationis a form of energy that is emitted or transmitted in the form of electromagnetic waves and/or particles. The electromag- netic spectrum is a continuum of all electromagnetic waves according to frequency and wavelength. EMR can be described in terms of a stream of photons, which are massless particles, each traveling in a wave-like pattern and moving at the speed of light. Elec- tromagnetic energy at a particular wavelength l (measured in meters) has an associated frequency f (measured in cycles per seconds, i.e., hertz) and photon energy E (measured in electron volts). Thus, the electromagnetic spectrum may be expressed in terms of any of these three quantities. They are related by the following equations: l ¼ c f and E ¼ hf or E ¼ hc l where c is the speed of light (299,792,458 m sÀ 1 ) and h is the Planck’s constant (hz6.62Â10À 34 J s). Thus, high-frequency electromagnetic waves have a short wavelength and high energy, whereas low-frequency waves have a long wavelength and low energy. Therefore, a shorter wavelength corresponds to more energetic radiation and an increased potential for biological harm. The electromagnetic spectrum is divided into different types of radiation based on wavelength ranges and encom- passes a wide range of terrestrial applications. These different types of radiation, expressed as a function of decreasing wavelength, include radio, microwave (MW), infrared (IR), visible, UV, X-ray, and g-ray (Fig. 1). Fig. 1 The electromagnetic spectrum. The diagram shows the entire spectrum of the electromagnetic waves. The scale at the bottom indicates representative objects that are equivalent to the wavelength scale and their respective energy. Reproduced from NASA (http://son.nasa.gov/tass/ content/electrospectrum.htm). Ionizing Radiation as a Carcinogen 191 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 11. Ionizing radiation iscomposed of either particles or photons that have enough energy to ionize an atom or molecule by completely removing an electron from its orbit, thus creating a more positively charged atom. The ionization of matter can be divided into two processes: indirect and direct ionization (BEIR-VII, 2006). Certain types of EMR, such as X- and g-rays, are termed indirectly ionizing because part or all the photon energy is transferred to the electrons in the cell’s molecules, which, upon release, produce the bulk of subsequent ionization events. In contrast to this, charged particles, such as high-energy electrons, protons, a-particles (a helium atom nucleus moving at a very high speed), b-particles (a high-speed electron or positron), and fast heavy ions, are termed direct ionizing radiation because while they traverse the cell, they ionize numerous molecules by direct collisions with their electrons. Nonionizing radiation includes the spectrum of UV, visible light, IR, MW, radiofrequency, and extreme low frequency (Fig. 1). Nonionizing radiation is present in a wide range of occupational settings and can pose a considerable health risk. UV is classified into near-, medium-, and far UV according to energy level, where near UV is nonionizing. However, medium- and far UV radiation may carry enough energy per quantum to ionize atoms and molecules. Thus, UV radiation can also be considered within the spec- trum of ionizing radiation. 7.09.2.1 Units Some biophysical notions are fundamental to an effective understanding of ionizing radiation carcinogenesis. In the context of bio- logical systems, ionizing radiation must be understood in terms of the radioactivity of the source, the energy of the radiation, the level of background radiation, and the level of radiation energy absorbed. The latter, from an occupational exposure point of view, is the most important parameter. By definition, the common unit of measure for energy present in ionizing radiation is the electron volt (eV). One eV equals 1.6Â10À12 ergs or 1.6Â10À 19 J. Other parameters, apart from the energy level of a particular type of radi- ation, are important to the understanding of the biological effects of ionizing radiation (Table 1). A distinction is made between sparsely ionizingdor low linear energy transfer (LET)dand densely ionizingdor high LETdradiation. The LET of an ionizing charged particle is defined as the average energy lost by the particle due to electronic interactions per unit length of its trajectory; it is expressed in kiloelectronvolts per micrometer (keV mmÀ1 ) (BEIR-VII, 2006). High-energy EMR, such as X- and g-rays, is sparsely ionizing since, in cells, it results in the release of fast electrons that have low LET. In contrast to this, a-particles, neutrons, and heavy particles (HZE) are densely ionizing because, in cells, they can release fast protons and heavier atomic nuclei that have high LET (Fig. 2). 7.09.2.2 Dose and Dose Rate The absorbed dose D, measured in gray (Gy), is the amount of energy deposited per unit mass of material (e.g., living tissue). One gray is equivalent to 1 J of radiation energy absorbed per kilogram of tissue (1 J kgÀ 1 ¼6.25Â1018 eV kgÀ 1 ), and one centigray (0.01 Gy) is equivalent to a rad. However, the types of radiation are diverse in how they deposit energy; therefore, the absorbed dose is a poor descriptor of biological effects (Durante and Cucinotta, 2008). A dose of energetic particles normally causes more damage than an equivalent dose of energetic photons (X- or g-rays). If the same biological event is induced by a dose of a standard radiation (e.g., X-rays) and by a dose of a test radiation (e.g., HZE ions), then the ratio of the standard to test radiation dose is defined as the relative biological effectiveness (RBE) of the test radiation. The RBE depends on several parameters, including Table 1 The main parameters relevant to ionizing radiation Parameter Radioactivity Absorbed dose Dose equivalent Exposure Energy Definition The spontaneous disintegration of atomic nuclei. The nucleus emits a-, b-particles, or electromagnetic rays during the process The mean of energy absorbed per unit mass of material The estimate of radiation risk that accounts for the differences in the biological effectiveness of different types of radiation that produce the absorbed dose Quantity that expresses the ability of radiation to ionize air The ability to do work Common units Curie (Ci), 1 Ci¼3.7Â1010 Bq Radiation absorbed dose (rad), 1 rad¼100 ergs gÀ1 ¼0.01 J kgÀ1 Roentgen equivalent man (rem) Roentgen (R) Joule (J) International System of unit Bequerel (Bq), 1 Bq equals 1 event of radiation emission per second Gray (Gy), 1 Gy¼100 rad Sievert (Sv), 1 Sv¼100 rem Coulomb/kilogram (C kgÀ1 ), 1R¼2.58Â10À4 C kgÀ1 of air Electronvolt (eV) 1.6Â10À19 J Modified from Rask, J., Elland, C., Vercoutere, W. (2006). Radiation biology educator guide. National Aeronautics and Space Administration (NASA). 192 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 12. LET, particle velocityand charge, dose and dose rate, biological end point, and oxygen concentration. To estimate biological effects it is customary to scale the absorbed dose by a quality factor Q(LET), which is estimated from the measured RBE values for late effects (Table 2). Current values for Q range from 1 at low LET (<10 keV mmÀ 1 ) to 30 at high LET (around 100 keV mmÀ1 ) and then decrease at very high LET values because of what is called overkill or wasted energy (Durante and Cucinotta, 2008). The dose equivalent or biologically dose equivalent, H¼DÂQ, represents the absorbed dose adjusted for the biological effectiveness of a particular type of radiation. H is measured in sievert (Sv) and the centisievert (0.01 Sv) is equivalent to a rem (rad equivalent in humans). Thus, the dose equivalent is intended to encompass all aspects of a certain radiation exposure influencing a biological effect. Finally, another important factor needs to be introduced to understand the influence of dose rate on biological effect. The dose rate effectiveness factor (DREF) measures the difference between acute exposures (a single large exposure) and chronic expo- sures (an exposure fractionated over time) of the same type of radiation at the same dose. Similar to RBE, the DREF is expressed as a ratio and is therefore an important scaling factor for the physician, allowing meaningful comparisons to be made between acute exposure events for which there is historical evidence linking outcome and dose (e.g., atomic bomb survivors) and long-term expo- sures involving low dose rates (e.g., long duration human spaceflight) (Jones et al., 2008). 7.09.2.3 Biophysical Damage Mechanisms A full description of radiation physics and the interaction between radiation and target atoms is beyond the scope of this article and can be found elsewhere (Jones et al., 2008). However, some of the relevant biophysical damage mechanisms need to be covered to appreciate the processes important in energy deposition. Incident ionizing radiation can interact with matter by being either Fig. 2 Comparison of particle tracks in human cells and nuclear emulsions. The figure emphasizes the biological impacts as a function of charged particle tracks. (A–C) The DNA double-strand break distribution in human fibroblasts is depicted in situ by g-H2AX immunofluorescence staining (every green focus corresponds to a DNA double-strand break). The different patterns of energy deposition (LET) for various particles is shown in the different distribution of DNA double-strand break in cells. (A) The cells are exposed to sparsely ionizing, that is, low LET, g-rays. DNA breaks are uniformly distributed in the nucleus. (B) The cells are exposed to densely ionizing, that is, high LET, one silicon nuclei particle. (C) The cells are exposed to densely ionizing, that is, high LET, three iron nuclei particles. Cells exposed to high energy heavy ions show DNA damage along the path traveled by the particles. Additionally, cellular biological damage (DNA double-strand break) increases as a function of LET. (D) The damage tracks of different ions, from proton to iron, are seen in nuclear emulsions and show the increasing ionization density (LET) as charge, Z, increases. Our understanding of biological knowledge decreases with increasing atomic number. A cell has been drawn to scale for comparison purposes. Reproduced from Cucinotta, F. A.; Durante, M. (2006). Lancet Oncology 7, 431–435. Ionizing Radiation as a Carcinogen 193 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 13. scattered or absorbed.As such, it can interact with one of the atomic components (i.e., nucleus or electrons), which usually produce secondary radiation following the destruction of the incident radiation, the target, or both. Additionally, it can also interact with atomic components that usually change the target atoms and lose energy via a complex chain reaction of radiation events (Jones et al., 2008). The mechanisms of absorption are of particular interest because (1) absorption in body tissue may result in physio- logical injury, (2) absorption is a phenomenon upon which the detection of ionizing radiation is based, and (3) the degree of absorption, or type of interaction, is a primary factor in determining shielding requirements (NCRP, 1993). The transfer of energy from an incident photon or particle to atoms of an absorber (e.g., tissue) may occur by excitation loss and ionization. The process of excitation involves the addition of energy to an atomic or molecular system, thereby transferring it from its ground or stable state to an excited or unstable state. Depending upon the type of interaction, either the atomic nucleus or one of its orbital electrons may absorb the excitation energy. An excited electron will not retain its energy but will tend to return to its orig- inal energy level either by emitting the excess energy in the form of a photon (e.g., X-ray) or by transferring its energy to the electrons of other atoms or molecules. As indicated previously, ionization is any process that results in the removal of an electron from an atom or molecule, thereby leaving the atom or molecule with a net positive charge. Ionization occurs if a- or b-particles, or g- photons, transfer sufficient energy to dislodge one of the electrons from the outer orbital shells of the target atom. Each ionization event produces an ion pair consisting of a free electron and the positively charged remainder of the atom. As discussed in the previous sections, photons produced from EMR play an important role in the ionizing process. In the inter- actions of photons with matter, the energy of the photons is transferred via collision; usually these collisions occur with orbital electrons in an atom of the absorbing medium. The most relevant energy transfer processes whereby photons of sufficient energy eject electrons from an atom, which can then interact with other atoms and molecules to produce a cascade of alterations that ulti- mately lead to observable biological effects, will be described. These are the photoelectric effect, Compton scattering, and pair production (BEIR-VII, 2006; Jones et al., 2008). At low energies (<0.1 MeV), the photoelectric effect dominates in tissue. A photon interacts with and ejects an electron from one of the inner shells of an atom. The photon is extinguished, and most of its energy is imparted to the ejected electron as kinetic energy. As outer electrons fill the vacancy, this energy change is balanced by the emission of a photon. In tissue, this type of photon emission has a low energy, typically 0.5 keV, and is of little biological consequence. At medium photon energies (about 0.5–3.5 MeV), Compton scattering is the most probable event. Compton scattering occurs when an incoming photon’s energy greatly exceeds the electron-binding energy of the affected atom. In this case, the energy of the incoming photon is converted into the kinetic energy of an ejected electron and a secondary “scattered” photon. Hence the products of Compton interactions are a scattered, less energetic photon of reduced wavelength, a high-speed electron, and an ionized atom. The ejected electron will travel some distance in matter, producing ionizations along its track. In the course of this travel, the photon Table 2 Quality factors associated with various types of radiation Radiation type and energy range Source/occurrence Penetration properties in human Quality factor, Q X-rays X-ray machine and accelerators, Van Allen belts, solar radiation, electromagnetic processes X- and g-rays penetrate deeply (only a fraction of the rays interact with each layer of tissue) 1 g-rays Radioisotopes decay, Van Allen belts, solar radiation, electromagnetic processes b-particles Radioisotopes decay, Van Allen belts, solar radiation, galactic cosmic radiation The level of penetration depends on the energy but is usually limited to less than 8 mm in tissue 1 Neutrons: <10 keV Nuclear reactor, accelerators, radiation therapy, atmosphere, Van Allen belts, solar radiation, galactic cosmic radiation Neutrons penetrate deeply (only a fraction of the neutrons interact with each layer tissue) 5 Neutrons:10–100 keV 10 Neutrons: 100 keV to 2 MeV 20 Neutrons:2–20 MeV 10 Neutrons: >20 MeV 5 Protons of energy > 2 MeV Accelerators, radiation therapy, Van Allen belts, solar radiation, galactic cosmic radiation The level of penetration depends on the energy 2 a-Particles, fission fragments, heavy nuclei Radioactive decay, solar radiation, galactic cosmic radiation The level of penetration depends on the energy but is limited to about the thickness of the epidermis for a-particles !20 Modified from Jones, J., Karouia, F. (2008). In: Barratt, M., Pool, S. (Eds.). Principles of clinical medicine for space flight, 1st ed. Springer, pp. 475–519. 194 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 14. may undergo additionalCompton collisions until its energy is sufficiently degraded for the photoelectric process to predominate. Thus, the photons in this energy range have their energy distributed over a relatively large volume of matter and may therefore have significant biological effects. At energies greater than 1.02 MeV, pair production can occur. A photon interacts with an atomic nucleus, and the photon energy is converted into a positron and an electron. The photon energy above 1.02 MeV is converted into the kinetic energy of the newly created particles. The electron and the positron interact with and can ionize other molecules until the excess kinetic energy is exhausted. 7.09.2.4 Atomic Ionization The so-called heavy nuclei are the nuclei of ordinary atoms of high atomic number whose electrons have been stripped away yielding a very heavy, highly charged particle. Energy from a heavy ion is deposited along the core of the track, where the ionization events produced in glancing collisions are quite dense. The core can be as wide as a few nanometers (Fig. 2D). Surrounding the core is a penumbra of low LET energetic electrons (d-rays), where the density of ionization events is much less than that in the core but extends for many microns (Cucinotta et al., 2000; Kramer and Kraft, 1994). These features allow even a single heavy ion particle to affect many cells in an irradiated tissue, which make the biological effects of heavy ions different from those of other radiation phenomena. As such, heavy ions are several times more effective than X-rays in terms of inducing radiation effects on small DNA segments (Cucinotta et al., 2000; Goodhead, 1994, Fig. 3). In addition to natural background radiation, the general population is exposed to low- and high LET radiation from man-made sources such as X-ray equipment and radioactive materials used in medicine, research, and industry. The man-made ionizing radi- ation exposure of the population of the United States has been estimated to account for 18% of the total annual US population exposure, whereas the remainder originates from background radiation (Fig. 3) (NCRP, 1987). Fig. 4 illustrates the relative contri- butions of the various man-made forms of radiation to the US population. People working in medical facilities, mining, milling, or with nuclear material are required to protect themselves from occupa- tional exposures to radiation. Therefore, the Occupational Safety and Health Administration (OSHA) strictly regulates the maximal amount of radiation that workers can receive in connection with their occupation. The limits are 50 mSv yearÀ1 for the whole body for terrestrial radiation workers and 50 cSv for astronauts (BEIR-VII, 2006; CERSSE, 2008). As we have seen earlier, predicting the risks associated with exposure of biological tissue to a given quantity of radiation is a complicated process. The current preferred measure of risk is the risk of exposure-induced death (REID) and has been imple- mented by data originated from studies of Japanese atomic bomb survivors, animals, and cell cultures. REID quantifies the risk of an exposed individual dying from a certain cancer as a function of the effective dose (Jones et al., 2008). Of note, cancer in the United States was reported to account for 23% of all deaths in 2004 (CERSSE, 2008). Fig. 3 The natural background exposure worldwide. The figure depicted here illustrates the relative contributions of natural sources to the global population exposure. The present estimate of the central value of background radiation is 2.4 mSv. More than half of the total exposure comes from exposure to radon gas and its decay products. Cosmic radiation is subsequently the next highest percentage of natural ionizing radiation exposure. Reproduced from UNSCEAR (2000). Source and effects of ionizing radiation, vol. 1. United Nations Scientific Committee on the Effects of Atomic Radiation. United Nations Publications: New York. Ionizing Radiation as a Carcinogen 195 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 15. 7.09.3 Radiobiology 7.09.3.1 PrimaryVersus Secondary Damage A cell’s sensitivity to radiation is influenced by its stage in the cell cycle, its state, and the component of the cell that was exposed. With regard to cell cycle stage, cells are generally most sensitive to reproductive death when irradiated during M phase (mitosis), to chromosomal damage and division delay when irradiated during G2, and to problems with DNA synthesis during early G1. They are most resistant during late S phase and during G0 (Prasad, 1995). The timing of irradiation also affects the progression of the cell through the cell cycle in a way that is dependent upon the normal rate of division in a given cell line; for example, low dose radiation stops slowly dividing cellsdbut not rapidly dividing onesdin G1. With regard to cell state, cells irradiated in vitro are more radio- sensitive than those irradiated in vivo (Jones et al., 2008). Amplification of the expression of specific oncogenes (e.g., ras, especially when myc is coexpressed, or raf) or the presence of radiosensitive or radioprotective genes can affect the radioresistance of cells. With regard to cellular components, the nucleus is more sensitive to both low- and high LET radiation than the cytoplasm. Redun- dancy in the number of mitochondria may confer radioresistance; for example, lymphocytes contain few mitochondria and are exquisitely sensitive to irradiation. Another factor affecting radiation-induced cell death or inactivation is the oxygen tension in the cellular environment; many cells are more sensitive to irradiation under normoxic conditions as compared to hypoxic environ- ments (Jones et al., 2008). 7.09.3.2 Mechanisms of Damage From Ionizing Radiation The main cellular effects of ionizing radiation relate to specific ionization events that produce molecular alterations. Space radia- tion, as opposed to typical terrestrial sources, contains a much greater proportion of particulate radiation. Of greatest concern are HZE particles: high LET radiation particles, which produce dense ionization tracks. Cells exposed to radiation have one of four fates: (1) complete recovery to the preradiation state; (2) partial recovery with repair of injury but with diminished functionality; (3) mutations caused by incomplete or erroneous repair; or (4) cell death (Jones et al., 2008). Incident radiation injures cells both directly and indirectly. Approximately one-third of biological damage from low LET radi- ation is thought to be from direct ionization, with the remainder incurred from indirect damage. The vast majority of damage from high LET radiation results from direct ionization. The following sections outline the mechanisms by which radiation directly and indirectly induces genetic damage (i.e., damage to a cell’s DNA), followed by a brief review of mechanisms of additional epigenetic damage (Jones et al., 2008). 7.09.3.3 Direct DNA Damage Ionizing radiation can penetrate the cytoplasm of a cell and interact with the molecularly rich cell nucleus, which is packed with DNA, histone proteins, and nuclear matrix. The severity of the injury depends on the track, the cross section, and the LET of the particle. When electromagnetic or particle radiation strikes the DNA and other macromolecules directly, molecular damage occurs in the form of ionization and, possibly, molecular bond breaks (Turner, 1995). The hydrogen bonds (including hydrogen– hydrogen (H–H) and sulfhydryl (SH)) are the weakest in the macromolecular structure and are therefore the most vulnerable to disruption by ionizing radiation. Breaks in these bonds lead to changes in the secondary and tertiary structure of proteins and enzymes, which in turn lead to decreases or loss of functional activity. Cellular proteins may express alterations in their viscosity, conductivity, and other physical properties. The side chains of amino acids are the most radiosensitive portions of proteins. Large Fig. 4 Relative contributions of the man-made radiation to the US population. The figure shows the relative contributions of man-made radiation. Medical X-rays and nuclear medicine account for about 79% of the man-made radiation exposure. Elements in consumer products, such as tobacco and domestic water supply, account for 16%. Occupational exposures, fallout, and nuclear fuel cycle account for the remaining. Reproduced from NCRP (1987). Ionizing radiation exposure of the population of the United States. National Council on Radiation Protection and Measurements, Report No. 93, Washington, DC. 196 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 16. macromolecules with repeatedidentical units often show disruption in the same bond, suggesting that the energy absorbed in the molecule can be transmitted down the molecular chain to the weakest bond. Histone proteins may lose their associations with DNA, and the secondary and tertiary DNA structure may be altered with disruption of the hydrogen bond linkage between base pairs; both effects can lead to errors in transcription and translation (Prasad, 1995). Molecular disruptions in the DNA molecule are characterized as strand breaks (single or double), apurination, or deamination. Strand breaks often occur between a sugar (ribose) and a phosphate, although these breaks will often rejoin if the broken end is not peroxidized by a reactive oxygen species (ROS). Radiation of energy as low as 30–40 eV can produce a break in one of the two strands of DNA (a single-strand break (SSB)), and an exposure of a cell to 0.01 Sv (1 rem) can be expected to produce 10–20 SSBs (Fig. 5). Double-strand breaks (DSBs) can occur when two SSBs are juxtaposed or when a single densely ionizing particle (HZE with >500 eV) produces a cluster of ionization within a span of about 20 degrees. High LET radiation, at a given energy, will induce more non-rejoining strand breaks than will low LET radiation, and non-rejoining strand breaks are more likely to lead to cell death. Another mechanism of DNA damage is cross-linking, an irreversible binding between chemically active loci produced in adjacent molecules or within the same molecule. Base pair dimerization, a type of cross-linking from an ionizing expo- sure, can easily produce a downstream mutation (Jones et al., 2008). Radiation-induced SSBs between a sugar and the phosphate group of the nucleotide can readily be repaired with high fidelity, since the template for the nucleotide is preserved. Occasionally, if SSBs occur in adjacent sister chromatid regions, the affected DNA segments undergo a process called sister chromatid exchange (SCE). However, when an ionization event leads to a DSB, the template is lost and errors in repair are much more likely, producing a point or segmental mutation. Such injuries or mutations can be lethal if the DNA damage is severe enough to cause the loss of function of one or several key proteins, or if repair is not possible and the chromosomal elements beyond the break are lost. A process similar to SCE that preserves the broken chromosome can sometimes remove DSBs, but such repair may place genes under different control mechanisms (as can happen with genetic recombination), which can also lead to changes in cellular activity and phenotype (Jones et al., 2008; Prasad, 1995). Single hits within chromosomes are more likely to be repairable by normal cellular mechanisms, but multiple hits in the same region of a chromosome may require more complex repair mechanisms or may not be repairable at all. Depending on the path of the ionizing particle, multiple damage sites can occur in proximity to one another. If the sites are located less than 20 degrees apart, the ionization event is usually lethal to the cell, whereas injuries to sites separated by more than 80 degrees are usually survivable but are likely to lead to mutations. HZE exposure tends to produce more complex nuclear biochemical events than those produced by low LET radiation. The complex events can lead to “unfaithful” or non-rejoining strand breaks and clusters of injury (e.g., base damage, SSBs, DSBs). Specific postexposure chromosomal aberrations observed in cytogenetic analysis of lymphocytes include inversions, dicentrics, fragments, rings, and translocations (Prasad, 1995). Fig. 5 Diagram of DNA lesion by direct effects. Reproduced from SRHWG (2001). Space radiation health project description. Houston, TX: NASA/JSC. Ionizing Radiation as a Carcinogen 197 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 17. If the cellsurvives the damage event, several downstream effects may occur. Translation errors can be seen if a DSB occurred in a coding region of the DNA, leading to mutated or truncated proteins with aberrant or lost function and subsequent alterations in phenotype. Mutations can also result in replication errors during mitosis. Errors in replication, if they occur in a sensitive region of the genome, can cause further mutations in daughter cells through rearrangements; such errors are the root of potential carcinogen- esis in these cells (Jones et al., 2008). In addition to overt injury, incidental radiation exposure can induce genomic instability. This can be produced with as little as 0.2–0.3 Gy (20–30 rem) of high LET radiation in mammary and other cell lines (Fry et al., 1983; Hall et al., 2001). In one exper- iment, transplanted bronchial epithelial cells that were irradiated with 0.3 Gy of 56Fe (<1 particle per cell) and, 6 months later, with 1 Gy of X-rays developed tumors in three of seven animals upon implantation; no tumors formed when cells had been irra- diated with either 0.3 Gy of 56Fe or 1 Gy of X-rays immediately before transplantation (Hei et al., 1998). These results imply that exposure to as little as a single HZE particle may render a cell genetically more sensitive or unstable for months and, therefore, at greater risk for subsequent neoplastic initiation events. Recent animal models of tumorigenesis have also demonstrated the extent to which high LET particle radiation is, itself, highly carcinogenic, with evidence suggesting that there is an increased risk of solid tumor formation subsequent to exposure to high LET particle radiation, as compared to g-rays or X-rays (Datta et al., 2013; Weil et al., 2014; Wang et al., 2015). In one study, groups of mice were irradiated with 28Si, 56Fe, protons, or g-rays at doses ranging from 0.01 to 0.3 Gy; mice irradiated with 28Si and 56Fe ions demonstrated a markedly higher incidence of hepatocellular carcinoma, as compared to mice irradiated with protons or g-rays administered at the same doses (Weil et al., 2014). The mechanism for high LET-induced genomic instability is not fully understood, but this phenomenon may account for the carcinogenic side effect of such irradiation. This condition seems to persist for several generations of cellular offspring after exposure. However, cells trans- formed by high LET radiation cannot be distinguished phenotypically from those transformed by low LET radiation. 7.09.3.4 Indirect DNA Damage or Oxidative Injury Ionizing radiation can interact with other parts of the cell besides the nucleus. The nucleus-to-cytoplasm ratio of cells varies from 1:5 to less than 1:1 depending on the cell type and maturity. For the majority of cells, the probability of radiation interacting with cyto- plasmic organelles and molecular species is statistically much larger than that of interacting with nuclear species. Damage to either the cytoplasm or the nucleus by ionizing radiation can result not only from direct damage but also from secondary reactive species. Radiation exposure results in energy being released into cellular materials, causing excitation of electrons or secondary ionization (Turner, 1995). In addition to the formation of ions, radiation can cause the loss of an electron from an atom or molecule, resulting in an unstable, highly reactive entity called a free radical. The unpaired outer shell electron of these electrically neutral radicals causes them to react very quickly with one another or with stable molecules (Jones et al., 2008). Since the human body consists of about 70% water, such events primarily involve aqueous products, particularly the highly reactive hydroxyl (OH) and peroxy (HO2 l ) radicals. Reactive species such as the oxidizing agents OH and HO2 l and the reducing agent H can propagate and disseminate, interacting with various parts of the cell such as cytosolic proteins and other macromolecules, membrane constituents such as lipids, and nuclear contents, including DNA. The base structures are particularly susceptible to direct damage by OH radicals, and the pyrim- idine bases are almost twice as sensitive to radiation effects as are the purines. In macromolecules, radicals can cause hydrogen bond breakage, molecular degradation or breakage, and intra- and intermolecular cross-linking (Conklin and Walker, 1987). Hydroxyl species produced by g-irradiation can induce DNA–protein cross-links, which tend to occur mostly in areas of the genome that are being actively transcribed (Xue et al., 1994). Components of these links, known as DNA adducts (e.g., 8-OHdG), can be quan- tified as an indication of the extent of DNA damage from chemical or radiation exposure. 7.09.3.5 Epigenetic Effects As noted above, reactive species can be generated anywhere in the cell and can propagate and disseminate, eventually interacting with chromosomal elements, including the DNA itself. Such interactions can create DNA adducts and (hypo- or hyper-)methylation events, which do not mutate the structure of the DNA but change the pattern of expression of the affected genes. Epigenetic effects arise from one of the three mechanisms: (1) modifications of nongenetic nuclear proteins (histones or nonhistones) that affect tran- scriptional or translational activities or prolong the activity of protein kinases; (2) binding between carcinogens and tRNA, which can change amino acid codons, or binding between carcinogens and RNA polymerases, which can increase the expression of enzy- matic proteins; and (3) the action of cocarcinogens such as hormonal transcription factors (Jones et al., 2008). 7.09.3.6 Bystander Effects The DNA-centric paradigm that the nucleus is the quintessential target for radiation damage had prevailed among radiobiologists for some time, and earlier observations of nontargeted effects were not integrated into the mainstream of their studies (Hamada et al., 2007). Subsequently, advances in nonuniform radiation fields and microbeams have allowed for the emergence of new interest in ionizing radiation-induced bystander effects: the fact that deposition of energy in a cell not only alters that individual cell but also triggers signal pathways that can result in alterations in nonhit cellsdthat is, bystander cells (Brooks, 2005). The bystander effect was first demonstrated when monolayer cell cultures were exposed to mean a-particle doses in which only 198 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 18. 0.1%–1% of thecell nuclei were traversed by a single a-particle track; this resulted in an enhanced frequency of SCEs in 20%–40% of Chinese hamster ovary cells (Nagasawa and Little, 1992). Furthermore, exposure to both particulate and EMR has demonstrated the occurrence of in vitro and in vivo bystander effects in a wide variety of cell types (Azzam and Little, 2004; Azzam et al., 2004; Hei, 2006; Kassis, 2004; Mothersill and Seymour, 2004). Bystander studies have revealed that the resulting effects are manifested as the expression of a wide variety of end points, such as SCEs (Nagasawa and Little, 1992), mutagenesis (Nagasawa and Little, 1999; Zhou et al., 2000), chromosomal and genomic instability (Hall and Hei, 2003; Lorimore et al., 2003), micronucleus formation (Belyakov et al., 2005; Prise et al., 1998; Shao et al., 2001), neoplastic transformation (Mothersill and Seymour, 2004; Sawant et al., 2001a), proliferation and differentiation (Azzam et al., 2004; Iyer et al., 2000; Mothersill and Seymour, 2004; Shao et al., 2003), decreased clonogenic survival (Liu et al., 2006; Mothersill and Seymour, 1997, 1998; Mothersill et al., 2004; Sawant et al., 2002), and apoptosis (Lyng et al., 2006). Multiple intercellular and intracellular signal transduction pathways have been implicated in the bystander response (Hamada et al., 2007). The cellular communication bringing about the radiation-induced bystander responses is thought to occur through direct physical connections between cells, such as gap junction intercellular communications (GJICs), or through the culture medium (Ballarini et al., 2002; Little, 2006). Four possible models for intercellular signal pathways capable of producing the radiation-induced bystander response have been proposed (1) through the GJIC, (2) through interactions between ligands and their specific receptors, (3) through interactions between the secreted factors and their specific receptors, and (4) directly through the plasma membrane (Hamada et al., 2007; Matsumoto et al., 2007). Irrespective of whether the cytoplasm or nucleus is targeted by ionizing radiation, irradiated cells release signals in the form of cytokines, growth factors, membrane-permeable reactive species (e.g., H2O2 and NO), and other factors that include long-lived organic radicals (LLRs) (Hamada et al., 2007). More recently, microarray analysis of irradiated and bystander fibroblasts in confluent cultures has shown different expression profiles; this implies that intercellular signaling between irradiated and bystander cells activate intracellular signaling, leading to the transcriptional stress response in bystander effects (Iwakawa et al., 2008). Finally, selective induction of DNA damage levels, global DNA methylation, cell proliferation, and apoptosis has been correlated to gender differences in exposed and bystander spleen tissue of male and female mice induced by ionizing radiation (Koturbash et al., 2008a). Interestingly, the gender specificity of radiation-induced bystander effects may be due to gender-specific microRNAs involved in gen- otoxic stress responses and may help to explain gender specificity of radiation-induced carcinogenesis (Koturbash et al., 2008b). 7.09.3.7 Adaptive Response The radiation-induced adaptive response is described as the reduced effect of radiation received as a challenging dose in instances where there has been previous induction by a low radiation dosedthe so-called priming or conditioning dose (Tapio and Jacob, 2007). The adaptive response is the only radioprotective mechanism that has been formally recognized by international organiza- tions and agencies, such as the International Atomic Energy Agency and World Health Organization (Leonard, 2007). Adaptive responses have been observed in vitro and in vivo using various end points, such as cell lethality, chromosomal aberrations, muta- tion induction, radiosensitivity, and DNA repair (Cai, 1999; Cai and Liu, 1990; Cramers et al., 2005; Gajendiran et al., 2001; Shad- ley and Wiencke, 1989; Yonezawa et al., 1996). Additionally, radioadaptive responses have been observed to both low LET (X-rays, g-rays, b-particles) (Azzam et al., 1994; Olivieri et al., 1984; Shadley and Wiencke, 1989) and high LET (neutrons, a-particles) radi- ation (Gajendiran et al., 2001; Sawant et al., 2001b). In cellular studies, values of priming doses and doses rates resulting in adaptive behavior have been found to range from 0.01 to 0.5 Gy and from 0.01 to 1 Gy minÀ1 , respectively (Tapio and Jacob, 2007). Although the mechanisms responsible for the radioadaptive response are not fully understood, this phenotype has been associated with an increase of certain cellular functions such as DNA repair (e.g., poly (ADP-ribose) polymerase (PARP), apurinic/apyrimidic (AP) endonuclease, DNA-dependent protein kinase (DNA-PK)) (Coleman et al., 2005; Iyer and Lehnert, 2002a,b; Takahashi et al., 2002; Wiencke et al., 1986), cell cycle function (e.g., M phase phosphoprotein) (Coleman et al., 2005), and transducers (e.g., ataxia telangiectasia mutated (ATM), and p53) (Coleman et al., 2005; Sasaki et al., 2002; Takahashi, 2002). Such adaptive cellular responses have been mediated either by the release of diffusible signaling molecules or by GJICs (Coates et al., 2004). In contrast to cellular studies, where the adaptive response lasts from a few hours to only one cell cycle, the adaptive response in animals, and probably in humans, can be maintained from several weeks to several months, and, in some cases, throughout the entire life span (Tapio and Jacob, 2007). The factors determining the length of the adaptive response remain unclear; however, reactive oxygen and nitrogen species (ROS and RNS) may contribute to this response by (1) directly inducing DNA damage that initiates the radioadap- tive response; (2) inducing DNA damage that brings about the transcriptional/posttranscriptional regulation of certain genes that confer radioprotective properties to cells or enhance the functions of certain proteins to promote radioadaptive responses; and (3) inducing certain proteins (e.g., transcriptional factors) that induce the cellular events necessary to conduct a radioadaptive response (Matsumoto et al., 2007). Additionally, ROS/RNS may be the link between the adaptive response and bystander effects (Matsumoto et al., 2007; Tapio and Jacob, 2007). Animal studies indicate a possible role of the radioadaptive response in the development of various cancers, as well as in the induction of radioresistance, probably via the induction of the immune activation (Ina et al., 2005). The main demonstrated in vivo effect of a low-dose exposure is a reduction in the rate at which spontaneously initiated cells prog- ress to malignancy (Mitchel et al., 2003). Persons living in high natural radiation areas have been shown to have an induced immune response (Tapio and Jacob, 2007). More recently, an Australian study found that while increased levels of solar UV expo- sure were associated with increased chromosome breakage, there was also an unexpected association with a decrease in the rate of DNA strand break misrepair (Nadir-Shaliker et al., 2012). However, the broader subject of radiation hormesisdthat is, the notion that there are health benefits that arise from exposure to low-dose ionizing radiationdremains controversial and in need of further Ionizing Radiation as a Carcinogen 199 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 19. study. In awell-publicized example, thousands of Taiwanese residents were exposed to low-dose g-radiation over a period of years when the steel used to construct a number of apartment buildings was inadvertently contaminated with radioactive cobalt-60 during the early 1980s. A subsequent investigation of this population revealed increased risks of leukemia and thyroid cancer but, surprisingly, a decreased risk of all other malignancies and solid tumors (Hwang et al., 2006). As the authors note, these results may be subject to confounders and must be interpreted with caution. Nevertheless, others have called for a reevaluation of our approach to the risk assessment of low dose radiation exposure (Chen et al., 2007; Scott, 2008). Moving forward, the role of the radioadaptive response in carcinogenesis will be more effectively understood by embracing a more systemic approach. Such an approach will take account of the potential benefit of the adaptive response to human health and may demonstrate a possible reduction in natural-occurring carcinogenic diseases (Leonard, 2007). 7.09.4 Evidence for Radiation Carcinogenesis 7.09.4.1 Human Data As discussed in many articles of this volume, there are a number of accepted criteria for determining whether a particular agent is a human carcinogen. The most convincing argument, of course, comes from empirical evidence, usually epidemiologic, that the agent has caused cancer in humans. For the case of ionizing radiation, such evidence is, unfortunately, plentiful (Table 3). Several populations have been exposed, often deliberately, to ionizing radiation, sometimes at very high doses. The number of such groups is sufficiently large to allow epidemiologists to derive fairly detailed information concerning the effects of radiation dose and quality, age at exposure, and other variables on cancer incidence in a large variety of organs. Based on data collected from exposed cohorts over several decades, it appears that ionizing radiation is a potent carcinogen with no organ or tissue specificity, other than as determined by the circumstances of exposure such as the site and manner of irradiation (Boice and Fraumeni, 1984; Burns et al., 1986; Upton et al., 1986). In addition, reports are appearing that select groups may, due to their occupational level of radiation exposure, have an increased incidence of certain types of cancer. A prominent example is the recent number of manuscripts inferring that flight crews (pilots and flight attendants) have, due to their altitude-based exposures, especially with polar flight routes, a higher incidence of solid tumors, especially breast cancer in the female crewmembers (Barr et al., 2007; Rafnsson et al., 2000). 7.09.4.2 Atomic/Nuclear Weapon Survivors Although the use of atomic weapons constitutes a grim wartime event, the information available from their use can be used to benefit those in radiation occupations or those who suffer inadvertent exposures. The Radiation Effects Research Foundation (formerly the Atomic Bomb Casualty Commission) is a binational organization formed to evaluate the medical effects of radiation on humans and on diseases affected by radiation. Laboratories in Hiroshima and Nagasaki are dedicated to studying the acute and chronic effects of the atomic detonations in those cities. Epidemiologic tracking of the survivors has allowed for the study of the relationship between estimated radiation dose and the development of leukemias and solid tumors (Thompson et al., 1994). The acute, annual, and career radiation exposure limits recommended by organizations such as the International Commission on Radiological Protection (ICRP) and the National Council for Radiation Protection and Measurement (NCRP) are largely based on findings from this cohort of acutely exposed individuals (ICRP, 1991). By examining the fate of family members who were located in the same houses during the atomic bomb explosions, it has been estimated that doses of 2.7–3.1 Gy (270–310 rad) to the bone marrow caused death within 2 months in some 50% of cases. Esti- mates of the LD50/60 (death of 50% of the exposed population within 60 days), generated by the United Nation’s Scientific Table 3 Cancer associated with exposure to ionizing radiation populations Cancer type AB AS PM TC TH RP UM RD Leukemia þ þ þ þ Thyroid þ þ Breast þ þ Lung þ þ þ þ Bone þ Stomach þ þ Esophagus þ þ Lymphoma þ þ þ Brain þ þ Liver þ Skin þ þ þ AB, atomic bomb survivors; AS, ankylosing spondylitis patients; PM, postpartum mastitis patients; TC, tinea capitis patients; TH, patients receiving thorotrast; RP, radium dial painters; UM, underground miners; RD, radiologists. 200 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 20. Committee from theinformation on atomic bomb survivors, accidental radiation exposure cases, and radiation therapy studies, suggest that the LD50/60 is 2.5–3.2 Gy (250–320 rad) to the bone marrow when little medical assistance is available, and about 5 Gy (500 rad) when extensive medical care is provided (Petersen and Abrahamson, 1998). Animal studies have shown that admin- istering various growth factors to stimulate surviving blood-forming stem cells in the bone marrow facilitates more rapid recovery from radiation injury, and the lives of 100% of the exposed population can be saved after a whole-body dose of up to about 10 Gy (1000 rad). An excess risk of leukemia was one of the earliest delayed effects of radiation exposure observed in the victims of the atomic bombs dropped on Hiroshima and Nagasaki in Aug. 1945. Now, more than 50 years after these events, this excess risk is widely seen as the most apparent long-term effect of radiation. As of 1990, 176 of the 50, 113 survivors in the Life Span Study who had significant exposures (0.005 Gy or 0.5 rad) had died of leukemia, and about 90 of these deaths were attributable to radiation exposure. This excess was especially apparent because much of these occurred during the first 10–15 years after exposure. Unlike the dose–response curves for other types of cancer, the leukemia dose–response curve seems to be nonlinear, with low doses being less effective than would be predicted by a simple linear dose response. With regard to other types of cancer, 4687 people in the Life Span Study had died of nonleukemic forms of cancer by 1990, which represents an excess of 381 deaths as compared with an esti- mated 4306 deaths in a population that had not been exposed (Pierce and Vaeth, 2003; Pierce et al., 1996). A comparison of excess deaths in the Life Span Study population between 1950 and 1990, according to radiation dose, is shown in Table 4; the number of deaths sorted by the type of cancer is shown in Table 5. 7.09.4.3 Industrial Accidents 7.09.4.3.1 Chernobyl and JCO nuclear criticality accident Nuclear power accidents, such as those of the Japan Nuclear Fuel Conversion Company (JCO) (Tokaimura, Japan, Sep. 1999) and Chernobyl, have resulted in population exposures that are being medically monitored. Acute exposure of 600 workers at the Table 5 Cancer deaths between 1950 and 1990 among life span study survivors according to cancer site Type of cancer Total number of deaths Estimated excess deaths Evidence for effect Stomach 2529 65 Strong Lung 939 67 Strong Liver 753 30 Strong Uterus 476 9 Moderate Colon 347 23 Strong Rectum 298 7 Weak Pancreas 297 3 Weak Esophagus 234 14 Strong Gallbladder 228 12 Moderate Breast (female) 211 37 Strong Ovary 120 10 Strong Bladder 118 10 Strong Prostate 80 2 Weak Bone 32 3 Moderate Other solid tumors 948 47 Strong Lymphoma 162 1 Weak Myeloma 51 6 Strong Modified from Pierce, D. A., Shimizu, Y., Preston, D. L., Vaeth, M., Mabuchi, K. (1996). Radiation Research 146, 1–27. Table 4 Cancer deaths between 1950 and 1990 among life span study survivors according to dose 0.005–0.2 Sv 0.2–0.5 Sv 0.5–1 Sv 1 Sv Number of deaths from leukemia 70 27 23 56 Estimated excess deaths 10 13 17 47 Percent attributable to radiation (%) 14 48 74 84 Number of deaths from all other cancers 3391 646 342 308 Estimated excess deaths 63 76 79 121 Percent attributable to radiation (%) 2 12 23 39 Modified from Pierce, D. A., Shimizu, Y., Preston, D. L., Vaeth, M., Mabuchi, K. (1996). Radiation Research 146, 1–27. Ionizing Radiation as a Carcinogen 201 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 21. Chernobyl site onApr. 26,l 1986 resulted in 134 cases of radiation sickness due to high exposure and 31 deaths (28 in the first 3 months and 3 delayed), according to the 2000 UNSCEAR report to the general assembly (Holm, 2001; NEA95, 1995). Several studies have concluded that Chernobyl accident-generated radioactive iodine has resulted in an increased incidence of thyroid cancer among the exposed population, especially among children in neighboring Belarus, where the winds took the vast majority of the radioactive cloud and large amounts of 131I were concentrated in cow’s milk (Farahati et al., 2000; Jacob et al., 2000; Rybakov et al., 2000; Williams, 1994). The thyroid cancer rate in persons less than 18 years of age, in the region surrounding Chernobyl, increased from 9 cases per the 5-year interval pre-exposure (1981–1985) to 37 in the 1986–1990 interval and 177 in the 1991–1995 interval. The incidence rate was still high more than 10 years after exposure, with 116 reported cases during the 3-year interval from 1996 to 1998 (Rybakov et al., 2000). At least 1800–2000 thyroid tumors are attributed to the Chernobyl accident- associated radiation exposure (Williams, 1994). The tumors in these patients were characterized by a shorter latency period and increased aggressiveness at presentation, as measured by the extent of regional and distant metastases (Heidenreich et al., 1999; Rybakov et al., 2000). The incidence and aggressiveness of regional urinary tumors (renal and bladder) were also increased in the 13-year period after the accident, as measured by stage, histopathologic features such as grade, PCNA (proliferating cell nuclear antigen), and K-ras expression, presumably due to ingestion and renal excretion of 137Cs (Romanenko et al., 2000, 2006). Most of the radio-induced cancers occurred in individuals exposed to high dose rates of 100 mSv yearÀ1 (Masse, 2000). Germline mutations at human minisatellite loci were studied in children born in the heavily polluted areas of the Mogliev district of Belarus after the Chernobyl accident and in a control population. The frequency of mutation was found to be twice as high in the exposed families as in the control group (Dubrova et al., 1996; Weinberg, 2001). Researchers from Texas Tech University did not find increased number of birth defects, physical deformities, or germline mutations in microsatellite DNA from exposed embryos or micronucleus formation in exposed rodents versus unexposed counterparts (Baker, 2001). In the JCO exposures, three workers who were not wearing exposure badges underwent postexposure biodosimetry to quantitate their exposure by counting the number of chromosomal aberrations in peripheral blood lymphocytes. Biodosimetry is now widely used in industry to correlate exposure to a toxicant (radiation, chemical, etc.) to the risk of the biological effects, inducing human disease (Jones et al., 2008). 7.09.4.3.2 Techa River Basin The Techa River Basin region in southwestern Siberia suffered from the environmental release of a large volumeda total of 7.6Â107 m3 , equating to 1017 Bq (2.75Â106 Ci)dof radioactive waste products over more than a decade, beginning in 1948. The environmental releases from the plutonium processing facility from within 100 km of Chelyabinsk, Russia, resulted in contam- ination of the local water system and widespread distribution into multiple villages along the Techa–Iset–Tobol river network of tributaries. Approximately 95% of the release was between Mar. 1950 and Nov. 1951, with an average daily release of 1.6Â1014 Bq (4300 Ci). Subsequently, large populations of the local people and animals, across the age spectrum, were exposed to external doses up to 1220 mGy yearÀ1 and internal doses up to 2260 mGy yearÀ1 . Some village residents received both internal and external exposures. The sources were mostly g-emitters, with some b-emitters, but included 89Sr, 90Sr, 137Cs, 95Zr, 95Nb, 106Ru, and others, typically with long half-lives (Akleyev et al., 1995, 2002a,b). The cancer-specific mortality rate in the exposed population was 24.89% versus 16.17% for the controls. The cancer-specific mortality (55 cancer death cases) in the exposed patients with signs and symptoms of chronic radiation syndrome was 274.8 deaths per 100 000 person-years (206.92–357.79, 90% CI) versus 189.9 (181.16–198.82, 90% CI) in the control population (1888 cancer death cases) (Akleyev et al., 1995, 2002a,b). The older exposed age cohort had the highest cancer death rate at 602 deaths per 100 000 person-years. The largest differences in cancer incidence of the exposed population versus the controls were for leukemiad25 per 100 000 person-years (8.09–58.20) versus 3.62 (2.53–5.01), female breastd14.99 (3.09–43.77) versus 3.42 (2.37–4.78), urinary organs (kidney, ureter, bladder)d29.98 (11.0– 65.36) versus 9.25 (7.45–11.34), lymphomad5.0 (0.12–27.85) versus 1.21 (0.63–2.12), and uterusd24.98 (8.09–58.2) versus 17.50 (15.10–20.30). The cancer specific mortality rate was dose dependent, being much higher in exposure doses 1.0 Gy, at 429.18Â10À3 versus 198.22Â10À 3 at doses 0.2 Gy (Akleyev et al., 1995, 2002a,b). 7.09.4.4 Medical Radiation Therapy and Cancer Risk Medical exposuresdthe use of diagnostic and therapeutic radiation exposure in the form of 60Co, linear accelerators, injected and ingested radionuclidesdhave been ongoing for many years and millions of patients. Studies of patients with ankylosing spondylitis (a form of arthritis) treated with external beam external radiation therapy, or X-ray therapy (XRT), have shown an increased relative risk (RR) of lung and other solid tumors of 1.5–1.8 at 8–20 years posttherapy. However, in contrast to the incidence data for atomic bomb survivors, the RR began to decrease beyond 20 years postexposure. Scottish women who received pelvic irradiation for met- ropathia hemorrhagica have a RR of 3.02 of developing bladder cancer, but the vast majority of these cancers were not observed until 20 or more years after treatment (Darby and Inskip, 1995). In the 1930s, before the effects of ionizing radiation were well understood, a number of people were exposed to radiation for medical purposes (Holm, 2001; Mole, 1987; Mole and Major, 1983). In England, patients with ankylosing spondylitis were treated with ionizing radiation from 1935 until 1954 (Smith and Doll, 1982). These patients were later found to have a fivefold higher incidence of leukemia than expected. In Rochester, NY, ionizing radiation was given to women to treat postpartum mastitis, result- ing in elevated breast cancer incidence (Mettler et al., 1990; Shore et al., 1977). In the United States and Israel, ionizing radiation was used until 1960 as a treatment for children with tinea capitis or ringworm (Shore, 1990; Shore et al., 1976). Later, as adults, 202 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 22. these people developedbasal cell carcinomas of the skin and thyroid tumors. In Germany, an X-ray contrasting agent called thor- otrast, which contained 232Th, was given to patients requiring gall bladder or liver X-ray imaging. This population later developed a very high rate of liver cancer, which is otherwise quite rare in Europe (Travis et al., 1992). In Canada, during the 1930s and 1940s, tuberculosis patients received greater than 400 rads of ionizing radiation resulting, eventually, in a 15-fold increase in lung cancer (Myrden and Hiltz, 1988). There are several other examples of medical uses of high doses of ionizing radiation, each of which has resulted in a clearly evident increased incidence of cancer at the target organ (Boice et al., 1988; Diamond et al., 1973; Evans et al., 1986). All such treatments were halted in the 1950s or early 1960s after it became apparent that ionizing radiation, apart from its other biological effects, which include cell killing and effects on immune function, was also causing cancer in people exposed to the types of doses given during the course of medical treatments. This recognition came about as a result of one of the most significant exposures of humans to ionizing radiationdthe atomic bomb explosions in Japan in 1945. 7.09.4.5 Medical Conditions Predisposing to Radiation-Induced Cancer There are a number of medical conditions, especially those with a genetic basis, which can predispose the individual to ionizing radiation-induced injury (Murnane and Kapp, 1993). The most well known and well characterized of those is ataxia telangiectasia (AT), which is a rare inherited human disease that has provided some clues to the mechanism of ionizing radiation-induced carci- nogenesis. AT strikes about 1 in every 300,000 people born (Murnane and Kapp, 1993; Murnane and Painter, 1982; Taylor et al., 1975). Patients suffer from a number of neurological disorders. One of the symptoms of the disease is a strong sensitivity to ionizing radiation. Between 10% and 20% of AT patients become victims of cancer at a relatively early age (teens and early 20s). In some ways, the disease is analogous to xeroderma pigmentosum, which renders patients highly sensitive to UV irradiation. Lymphocytes and fibroblast cells from AT patients, grown in culture, exhibit a sharply enhanced sensitivity to the effects of ionizing radiation, although not to UV radiation. Results from these experiments have led to the hypothesis that AT cells have a DNA repair deficiency that is probably related to the repair of strand breaks (Painter, 1988). In addition to the increased radiosensitivity toward cell killing and transformation, certain AT cells display an abnormal characteristic related to DNA repair kinetics. Normal cells undergo a delay in DNA synthesis and cell cycle progression (see “Molecular Biology of Radiation Carcinogenesis” section) after exposure to ionizing radiation. This delay does not occur in radiation-exposed AT cells (Beamish et al., 1994). Research using AT cells grown in vitro revealed a high level of spontaneous chromosomal translocations in cells from AT patients, which is further increased after exposure to fairly low levels of ionizing radiation. It was thought that several genes were involved in the AT phenotype since a number of different complementation groups have been identified (Murnane and Painter, 1982). The gene for AT was cloned on chromosome 11q (Savitsky et al., 1995). Surprisingly, patients from all known complementation groups exhibited mutations in the same gene, strongly suggesting that despite previous evidence, all cases of AT result from a defect in this single gene. The gene, designated ATM, codes for a putative protein with homology to phosphotidy- linositol-30 kinases. These enzymes have been shown to have a critical role in a number of cellular processes, such as growth, differ- entiation, and apoptosis. This extremely important discovery will most likely lead to rapid advances in our knowledge of the disease, as well as of basic mechanisms associated with DNA damage and repair in human cells. 7.09.4.6 Radiation Carcinogenesis in Animal Models The response to radiation differs among species, as it does among cell types. Because ionizing radiation is a carcinogen in most organs in almost all species, researchers are faced with an enormous array of choices in animal models of radiation-induced carci- nogenesis. Questions about radiation-induced carcinogenesis have been studied as whole body irradiation of most laboratory animal species. Animal models used for studying the bioeffects of radiation have included rabbits, mice, rats, and other mammalian and nonmammalian species, including dogs and monkeys flown aboard the Russian Bion satellites. Many of the bioeffects are thought to be universal responses to radiation, whereas others are thought to be specific to cell type or species. Studies conducted jointly by the US Air Force and NASA from 1963 to 1969 looked at the RBE of various types of space- associated radiation exposures on rhesus monkeys and mice (Dalrymple et al., 1991). High-energy protons (138 MeV) were found to have an RBE of 1.0–1.1, similar to that of 2 MeV X- and g-rays. Long-term follow-up studies of the exposed animals showed induction of solid tumors and leukemia; however, the observed extent of life shortening and cancer induction depended on dose and not on proton energy level (Dalrymple et al., 1991). Subsequent animal tumorigenisis models evaluating the carcino- genic potential of HZE ions, however, have demonstrated that both particle type and energy can affect carcinogenesis risk, with HZE radiation generally increasing tumor incidence and aggressiveness, as compared to X- or g-rays administered at similar doses (Weil et al., 2014; Wang et al., 2015). Other animal studies indicate that long-term (chronic) exposures to penetrating low LET radiation result in less risk of cancer than acute exposures. Animal studies have also been useful for determining the RBE of various heavy ions for producing deterministic effects such as cell killing in the gut, testis, and bone marrow. Such values range from 2 to 3 for cell killing, peaking at an LET of 100–200 keV nucleonÀ 1 . Data on the peak RBE for inducing Harderian gland tumors in mice were 30 at 100 keV nucleonÀ 1 , but no decline in effect was noted beyond an LET of 100. More focused studies about specific mechanistic questions have been applied to specific organs in different animals. Given the wide range of animals available, determining the ideal animal model for radiation experiments is determined by the experimental question posed. Choices of experimental models are also affected by availability and cost. In addition, species-specific peculiarities make some animals ideal for certain studies and poor choices for others. Thoughtful analysis is required to extrapolate the results of Ionizing Radiation as a Carcinogen 203 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 23. animal studies intoinformation that is applicable to humans. Although there is considerable debate about ideal animal models to address different research questions, it is generally agreed that careful consideration is required to choose the best available animal model out of the many available choices for radiation carcinogenesis research. Some factors that must be considered include relative dosing, timing, animal life cycle, and intrinsic biological differences and/or responses to injury. Furthermore, animal size and morphology must also be taken into account, since absorbed dose distributions for a given radiation type will vary significantly between different animals and organ systems. Efforts to address these challenges can be seen in a novel technique, which aims to more accurately reproduce dose and dose distribution of different radiation types by using megavoltage electron beam radiation (Cengel et al., 2010). Early animal studies in radiation carcinogenesis performed after World War II primarily utilized rodents, particularly rats and mice (Fry and Storer, 1987). These studies elucidated the general characteristics of radiation carcinogenesis, in conjunction with epidemiological studies in humans. Data from quantitative animal tumorigenesis (UNSCEAR, 1988) and human epidemiologic studies (UNSCEAR, 1993, 1994, 2000) suggest that single acute doses of ionizing radiation produce a dose-dependent increase in cancer risk in both humans and animals. The BEIR-VII report, which assessed the biological effects of low-level ionizing radiation, determined that the radiation-induced life shortening observed in mice is largely reflective of radiation-induced cancer mortality in humans (BEIR-VII, 2006). These studies have shown that ionizing radiation is universally carcinogenic to living creatures, although there is wide variation in radiation responses according to species, organ, and radiation distribution. A recent evaluation of animal models for radiation-induced carcinogenesis agreed that there is a wide variety of appropriate animal models for research, including rodents, dogs, pigs, and primates (Augustine et al., 2005). However, significant uncertainty still exists about the accuracy of extrap- olating the carcinogenesis results of animal studies to humans. Genetically modified mice (inbred strains, knockouts, knock-ins, transgenics) are particularly useful tools to determine molec- ular and cellular mechanisms involved in radiation-induced cancers. For example, mice with epidermal-specific deletion of p53, but not ATM, showed increased papilloma number and progression to malignant invasive carcinomas as compared with wild-type littermates (Bailey et al., 2008). Mice with a truncated version of the adenomatous polyposis coli (APC) gene exhibited an increased occurrence of intestinal tumors after exposure to X-rays (Nakayama et al., 2007). Using mice with suppressed expression of the Snai2 gene, which has been shown to modulate carcinoma progression, researchers have shown that Snai2 expression enhanced UV-induced skin carcinogenesis (Newkirk et al., 2007). IL-10 knockout mice exhibited increased sensitivity to UV-induced skin cancer (Loser et al., 2007). Connexin 32-deficient knockout mice exhibited increased radiation-induced carcinogenesis of the lung and liver, compared to wild-type mice (King and Lampe, 2004). The expression of individual genes can be suppressed or acti- vated in various mouse models, which make them particularly useful for dissecting molecular and cellular factors that contribute to the initiation, progression, and treatment of radiation-induced cancers. However, a recent evaluation of animal models identified a need for the development of additional animal models to better understand the underlying mechanisms of radiation-induced carcinogenesis, particularly for the study of radioprotectants (Augustine et al., 2005). 7.09.4.7 In Vitro Evidence for Radiation Carcinogenesis We have shown in the previous section that animal models defined many of the general characteristics of radiation carcinogenesis. These findings were supported by various epidemiologic studies in human populations receiving radiation exposure from environ- mental, occupational, medical, and accident sources (Little, 2000; Pierce et al., 1996). The universal nature of radiation as a carcin- ogen relates to its ability to penetrate cells and to deposit energy within them. Subsequently, cellular systems were developed in the 1970s to study, in vitro, the malignant transformation of individual cells (Chadwick et al., 1989). By the 1980s, the general char- acteristics of how radiation-induced cellular transformation occurred in vitro were well established and only a limited amount of work has been conducted directly in these areas since that time (Little, 2000). Conversely, over the past two decades ionizing radi- ation research has focused on cellular and molecular mechanisms that may relate specifically to the induction of cancer. The following section illustrates dose–response relationships that have been obtained for chromosome aberrations, for cell transforma- tion, for mutagenesis in somatic cells, for gene expression, and for the biological effects occurring in nonirradiated cells. 7.09.4.7.1 Chromosome Aberrations As described in “Radiobiology” section, ionizing radiation can induce a broad range of DNA lesions including damage to nucleo- tides, cross-linking, and DNA SSBs and DSBs. Originally, DSBs were believed to be one of the main critical cytotoxic lesions; it is now accepted that misrepaired DSBs are the principal lesions of importance in the induction of both chromosomal abnormalities and gene mutations (Goodhead, 1994; Ward, 1995). Normal cells irradiated with ionizing radiation in G1 or G0 arrive in their first mitosis with aberrations that are virtually all of the chromosome type, whereas after irradiation in S or G2, aberrations are virtually all of the chromatid type (Bailey and Bedford, 2006). The level of cellular killing and aberration production can depend strongly on the cells and the condition of radiation exposure. Early on, it has been shown that an X-ray dose exposure of 4 Gy would produce approximately 120–160 DNA DSBs, 1000–2000 DNA SSBs, and a similar number of base damage events in each cell (Ward, 1988). Subsequently, the exposure would kill 50% of a population of normal human fibroblast cells exposed in G0 with just under a total of one chromosome-type acentric fragment-producing aberration (dicentric, centric ring, and interstitial and terminal deletions) per cell if the subculture is delayed to allow for completion of repair processes after irradiation (Cornforth and Bedford, 1987). It is expected that twice this number would be produced after the same dose of radiation exposure to human lymphocytes (Lloyd et al., 1975). Interestingly, SSBs and base damage lesions are, typically, so rapidly and efficiently repaired that only about 204 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 24. 0.01–0.02 chromatid-type aberrationsper cell might be contributed to the total aberration yield after an X-ray dose in G0 or G1 that yields some 1–2 chromosome types per cell (Ben-Hur and Elkind, 1972; Bender et al., 1973). For many years, the radiobiological research focus has been on the biophysical modeling of the dose response and LET depen- dence for chromosome aberration induction. The vast majority of studies show that the dose response for low LET radiation is curvi- linear and fits well to the linear quadratic equation (Bedford and Dewey, 2002; Hlatky et al., 1991; Lloyd et al., 1992; Moiseenko et al., 1997; Sachs et al., 1997; Wagner et al., 1983). The development of fluorescence in situ hybridization (FISH) methods of chro- mosome painting has allowed aberration complexity to be studied in detail and has revealed a dose and LET dependence (BEIR-VII, 2006). Furthermore, aberration complexity that reflects the level of DNA DSBs involved in a given chromosomal exchange event becomes increasingly apparent at high-dose low LET and at all doses of high LET exposure (Anderson et al., 2000; Finnon et al., 1995, 1999; Griffin et al., 1995). Some investigations combining FISH painting and premature chromosome condensation tech- niques have facilitated studies of the rate of formation of aberrations, which revealed rapid complete and incomplete exchanges suggesting time dependence for pairwise exchange of DNA DSBs (Alper et al., 1988; Darroudi et al., 1998). Thus, chromosomal aberration resulting from misrepair events associated with DNA DSBs is probably associated with the dominant postirradiation function of the nonhomologous end joining (NHEJ) repair processes (BEIR-VII, 2006; Cornforth, 2006). Heavy charged particles are effective at producing chromosomal exchanges with RBE values ranging from 10 to 30, depending on the cellular stage (George et al., 2003). However, lower RBE values have been reported under comparable conditions during in vivo studies (Rithidech et al., 2007; Tucker et al., 2004). Additionally, the dose response obtained following high LET exposure in vitro to both a-particles and neutrons is generally well fitted with a linear response (Edwards, 1997). Cytogenetic studies reveal a much higher level of complexity of chromosomal aberration induced by densely, as compared with sparsely, ionizing radiation (Fig. 6). Furthermore, the insults from heavy ions trigger complex rearrangements due to an increase in the number of chromo- somes and breakpoints, which include both intra- and interchromosomal exchanges (Durante et al., 2002; Hada et al., 2007; Johannes et al., 2004). However, most of these complex rearrangements induced by generating chromosome aberrations via Fe ion exposure lead to cell death (Rithidech et al., 2007). Interestingly, chromosomal aberrations, measured in the blood lymphocytes of astronauts returning from long-term mission, have been used to estimate astronauts’ dose, dose-equivalent, and cancer risk (Boffetta et al., 2007; Durante, 2005; Norppa et al., 2006). 7.09.4.7.2 Cell Transformation Cell transformation describes the changes associated with the loss of normal homeostatic control, particularly of cell division, which ultimately results in the development of a neoplastic phenotype (UNSCEAR, 2000). The majority of radiation-related research on neoplastic transformation in vitro has been quantitative in nature. Most studies have been carried out with various rodent-derived Fig. 6 Chromosome aberrations induced by heavy ions. The figure depicted here illustrates the complex-type aberrations, that is, a minimum of two chromosomes and three breakpoints, induced by energetic Fe ions in human peripheral blood lymphocytes. (A) Multicolor FISH visualization of polycentric chromosome. From top to bottom: a quadricentric involving both chromosomes 1, chromosome 9, and chromosome 6; a dicentric of chromosome 1 and 9, with an insertion of chromosome 3; a tricentric involving both chromosomes 6 and chromosome 1. (B) Multicolor banding FISH visualization of a complex rearrangement in chromosome 5. Chromosome 5 is broken into three pieces whereas the normal chromosome 5 is visible at the bottom. Reproduced from Durante, M., Cucinotta, F. A. (2008). Nature Reviews Cancer, 8, 465–472. Ionizing Radiation as a Carcinogen 205 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 25. fibroblast cell lines,such as BALB/c3T3 and the C3H10T1/2 mouse embryonic lines, because of the refractive nature of human cells to radiation-induced neoplastic transformation in vitro (Reznikoff et al., 1973; Rhim and Dristchilo, 1991). One of the few human- derived cell-based assays that have been developed for quantitative studies is the HeLa human skin fibroblast human hybrid cell assay (Mendonca et al., 1992; Redpath et al., 1987; Sun et al., 1988). Past research using in vitro transformation assays has helped in understanding the dependence of neoplastic transformation on radiation dose fractionation and on radiation quality. In general, following exposure to low LET radiation, the dose–response relationship for cell transformation is very dependent on cell cycle kinetics; nevertheless, it follows results obtained with other cellular effects with some limitation on the intensity of the exposure (Barendsen, 1985; Han and Elkind, 1979; Miller and Hall, 1978; Miller et al., 1979; Mole and Major, 1983; UNSCEAR, 1988, 1993). For transformation by low LET, various dose–response relationships have been reported. Some have described a linear dose–response relationship (Borsa et al., 1984; Hei et al., 1988; Hill et al., 1987), whereas others have described linear quadratic or curvilinear relationships (Borsa et al., 1984; Hei et al., 1988; Hill et al., 1987). Cell line comparison, that is, BALB/c3T3 and C3H10T1/2, was carried out following exposures between 100 mGy and 3 Gy and the dose–response relationships were found to be nearly linear and linear quadratic, respectively (UNSCEAR, 2000). To address the issue of dose response at low LET radiation, an identical study has been conducted among different laboratories with the C3H10T1/2 transformation system (Mill et al., 1998). The study revealed a linear dose–response relationship for cell transformation in vitro at low dose, and therefore does little to support the concept of either a threshold dose or an enhanced supralinear response. Finally, cells exposed to very low dose rate radiation exhibited a trend toward a reduction in neoplastic transformation frequency compared to the unirradiated controls (Elmore et al., 2007). This reduction seemed to diminish with time, indicating that the dose rate, rather than accumulated dose, may be the more important factor. The very low dose rate-treated cells were less sensitive to the high challenge dose than irradiated controls, suggesting the induction of an adaptive response. As expected, the exposure to high LET radiation results in a higher transformation frequency than exposure to low-LET radiation and shows a general tendency toward a linear dose–response relationship with saturation stage, which slightly decreases at high dose (Hei et al., 1988; Hill and Zhu, 1991; Miller et al., 1989, 1995). Conversely, there is no tendency for the response per unit dose to decrease at low doses or low dose rates, although a number of studies have shown an enhanced effect (UNSCEAR, 2000). 7.09.4.7.3 Mutagenesis in Somatic Cells The principal mechanism resulting in a neoplastic initiation event is induced by a broad range of potentially mutagenic lesions in DNA, which predisposes target cells to subsequent malignant development (BEIR-VII, 2006; Little, 2000; UNSCEAR, 2000). Earlier studies with the hemizygous X-linked hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene showed that radiation could induce point mutations and deletions, the latter sometimes including the entire gene (Sankaranarayanan, 1991; Thacker, 1992). There is strong evidence that, in most cases, a DNA deletion mechanism dominates mutagenic response after ionizing radiation and the genetic context of the mutation is therefore of great importance (Sankaranarayanan, 1991; Thacker, 1992). Thus, the predominant molecular structural changes associated with radiation-induced mutations are large-scale events that may include deletions, chromosomal rearrangements, or recombinational processes (Little, 2000). Such mutational changes are a result of DSBs. The effects of radiation quality have been investigated on the induction of gene mutations and show a similar relationship between relative RBE and LET as suggested for the induction of chromosome aberrations (Cox and Masson, 1979; Thacker, 1992; Thacker et al., 1979). The maximum RBE values were usually in the range of 7–10. Furthermore, most of the molecular analyses suggest that DNA deletion mechanisms dominate for all radiation qualities (Aghamohammadi et al., 1992; Gibbs et al., 1987; Jostes et al., 1994; Thacker, 1986). The involvement of short, direct, and inverted DNA repeats at deletion breakpoints is highly suggestive of an important role for illegitimate recombination processes in mutagenesis and, as for chromosome aberration induction, the involvement of DNA DSBs and error-prone NHEJ (Miles et al., 1990; Thacker et al., 1990). Thus, molecular and cellular data suggest that the principal source of radiation-induced gene mutation derive from error-prone NHEJ repair process of DNA DSBs; therefore, a linear dose response would be anticipated at low doses (UNSCEAR, 2000). However, the technical level of resolution for the dose– response relationships for mutations is far less precise than those for chromosome aberrations and a linear quadratic relationship provides a good compromise to the dose response down to 200 mGy (Thacker, 1992). 7.09.4.7.4 Gene Expression In the mid-1990s, new methods of gene analysis were developed, allowing for a global view of transcriptional responses. The primary technique in deciphering the global gene expression profile after ionizing radiation has been cDNA and oligonucleotide-based microarrays (Snyder and Morgan, 2004). Recent research has demonstrated that cells can detect and respond with alterations in gene expression after very low doses of radiation. Additionally, gene expression changes as a function of radiation dose and radiation type (Snyder and Morgan, 2004; Yin et al., 2003). In spite of difference in array platform and experimental design, several studies show similar expression of genes involved in cell cycle checkpoints and growth control (Snyder and Morgan, 2004; Yin et al., 2003). One consistent trend among array experiments involving high and low doses of ionizing radiation is the induction of cyclin-dependent kinase inhibitor 1A (CDKN1A) after exposure (Amundson et al., 1999a,b, 2000; Balcer-Kubiczek et al., 1999; Heinloth et al., 2003a,b; Jen and Cheung, 2003; Li et al., 2001; Marko et al., 2003; Robles et al., 2001; Stassen et al., 2003; Tusher et al., 2001). Another frequent result is the upregulation of growth arrest and DNA damage inducible gadd45 after radiation treatment (Amundson et al., 1999a,b; Jen and Cheung, 2003; Marko et al., 2003; Tusher et al., 2001). While many genes induced by ionizing radiation are p53, that is, tumor suppressor, regulated, there is also a substantial p53-independent component to the transcriptional response to irradiation, with 206 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 26. NFkB, that is,nuclear factor, playing a substantial role (Amundson et al., 1999a; Chaudhry et al., 2003; Park et al., 2002). Interest- ingly, half of human cancers have a mutated p53 gene and, therefore, similar genomic studies could potentially underline the molecular pathways of cancer cells exposed to ionizing radiation. 7.09.4.7.5 Biological Effects Occurring in Nonirradiated Cells New mechanistic cell and molecular studies on the effects of low doses of radiation have resulted in three major paradigm shifts. First, the observation of bystander effects demonstrated that nonhit cells might respond alongside cells in which energy is deposited. Second, recent studies have demonstrated the existence of radiation-induced changes in gene expression at very low radiation doses. These changes can result in alterations in response pathways, some of which appear to be involved in protective or adaptive responses. Finally, early changes in the initiation phase of radiation-induced cancer were thought to be induced by gene mutation and chromosome aberrations; however, it is now understood that genomic instability leading to the loss of genetic control appears to play a major role in the development of cancer (Brooks, 2005). 7.09.5 Molecular Biology of Radiation Carcinogenesis When a positive correlation between radiation and carcinogenesis was first accepted, our ability to study the underlying cellular and molecular mechanisms was limited. In the 1940s, the effects of ionizing radiation were measured as the induction of chromosomal breaks in cells (Raffel and Muller, 1940). Advances in cellular and molecular biology technologies have enabled significant eluci- dation of the molecular effects of ionizing radiation: how chromosomal breaks affect cellular process including proliferation, cell cycle, DNA repair, and cell death, and how changes in these processes induce cellular transformation and carcinogenesis. Concur- rent advances in the concepts that define radiation biology and tumor biology have also facilitated a fuller understanding of the molecular processes in both humans and animals. The current prevailing theory about radiation-induced carcinogenesis, as stated in the BEIR-VII report, is that cancer is induced by multiple interactions of weakly expressing genes affected by radiation (BEIR-VII, 2006). The hallmark of carcinogenesis is a web of dysfunctional relationships, rather than a neat linear progression. In the past, it was thought that certain genes were critical in cellular transformation or tumorigenesis. However, no single gene or combination of genes is uniformly expressed in all cancers, or even in any single type of cancer. Instead, it appears that disruption of critical cell functions, which are regulated by multiple mechanisms, is more important than the loss or dysfunction of any single gene. Because most critical cell functions are performed by redundant mechanisms, several related genes must be affected before the net effect is great enough to commit the cell to change from normal function to carcinogenesis, consistent with Knudsen’s multi-hit hypothesis (Knudson, 1971). However, certain trends have emerged on the molecular and genetic levels, such as radiation-induced disruption of oncogenes and tumor suppressor genes that are commonly observed in many cancers (Garte et al., 1989). 7.09.5.1 Cell Cycle Delay and Gene Induction Exposure to ionizing radiation results in a delay of cell division, which is often the result of arrest of the cell cycle in the G1 or G2 phase (Maity et al., 1994; Murnane, 1995; Scholz et al., 1994). The mechanism responsible for this arrest is not known, but may involve effects on cell cycle-specific proteins, such as cyclin B1 and 34cdc2 (Kharbanda et al., 1994). The functional significance of this cell cycle delay is not known with certainty, but it is likely that cells may use the delay period to repair damaged DNA following ionizing radiation exposure. It appears that wild-type p53 as well as the gadd45 gene is involved in the cell cycle delay following ionizing radiation exposure (Kastan et al., 1992). The cell cycle delay is prolonged with increasing LET (Scholz et al., 1994). During this period, immediately after exposure to ionizing radiation, a large and complex set of phenomena has been shown to occur in the exposed cell. Fig. 7 shows some possible pathways of cellular events that are associated with the cell cycle delay caused by irradiation in cells. Research has shown that, in addition to the expected molecular consequences of ionizing radiation exposure on the genome, such as gene deletions, rearrangements, chromosomal aberrations, and other types of mutations, an important group of genes is affected by changes in the level of their expression (Fornace, 1992; Fornace et al., 1988; Wilson et al., 1993). Very soon after expo- sure to ionizing radiation, a number of growth-related genes are induced in mammalian cells. Because their induction occurs so rapidlydwithin 1–3 h after exposuredthis has been called an immediate early response, and the genes induced have come to be known as immediate early response genes. The mechanism of their induction is not entirely clear. Since other DNA- damaging agents can produce the same effect, it has been hypothesized that this pleotypic induction might result from DNA damage. The genes that are induced include several whose products themselves can act to stimulate the induction of other genes, leading to a cascade of gene induction in cells following ionizing radiation exposure. Examples of some important immediate early response genes include protein kinase C (Woloschak et al., 1990) (PKC), which has also been implicated in the pleotypic response of cells to phorbol ester; AP1/jun (Sherman et al., 1990), a transcription factor that leads to enhanced expression of c-myc; c-fos (Hollander and Fornace, 1989); TNF-a (Hallahan et al., 1989, 1991a); gadd (Fornace et al., 1992); plasminogen activator (Boothman et al., 1991); and other genes (Brach et al., 1991; Peter et al., 1993; Teale et al., 1992). It has been postulated that the production of free oxygen radicals or ROS by ionizing radiation may, in fact, be the intermediary inducer of these genes (Fuciarelli et al., 1989). Arguments against this view and in favor of the damage DNA hypothesis have been Ionizing Radiation as a Carcinogen 207 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 27. presented (Baverstock andWill, 1989; Ward, 1994). There is reason to believe that SSBs in DNA are involved in the initial signal, based on calculations of damage quantitation. The mechanisms of induction of these genes cannot involve the de novo synthesis of transcription factor, because there is not sufficient time for this to occur. Furthermore, it has been shown that inhibition of protein synthesis by cyclohexamide results in a very different pattern of gene induction and transcriptional repression than that seen with ionizing radiation (Woloschak et al., 1995). After the discovery of gene induction by ionizing radiation, it was found that other DNA-damaging agents, including certain chemical carcinogens, had a similar effect (Fornace, 1992). This response has sometimes been labeled, therefore, as a DNA damage response rather than a radiation response. However, not all DNA-damaging agents, or indeed even all ionizing radiation, produce the same effects. Woloschak and colleagues have found that the ability to induce the immediate early response is a function of radiation quality. For example, high and low LET ionizing radiation induce different patterns of gene expression (Hallahan et al., 1991a). Certain ionizing radiation-inducible genes such as c-jun, Egr-1, and TNF have been shown to be induced by ionizing radiation stimulation of PKC activity (Hallahan et al., 1991a). Other genes such as gadd45 are independent of PKC activation (Fornace et al., 1992). It is very likely that the mechanisms of induction of PKC-dependent and PKC-independent genes are completely different. The enzymatic activity of PKC is affected less than 15 seconds after ionizing radiation exposure (Hallahan et al., 1991b). Although calcium is required for the PKC responsedand it could be hypothesized that a very rapid change in calcium flux might be the initial event in triggering PKC-dependent gene activationddata suggest that ionizing radiation does not cause an increase in cellular calcium concentration (Hallahan et al., 1994). The induction of gene expression by ionizing radiation exposure is of great interest for understanding the molecular events and cascades that occur after DNA damage. The relevance of this phenomenon for radiation-induced carcinogenesis is not completely clear. Certainly, a large number of the gene products are members of signal transduction pathways, such as those mediated by protein kinase C, AP-1, and others. However, the relative importance of the activation of these pathways, which occur in a large number of target cells as compared to a rare event (such as a specific gene mutation), is not yet known. Some investigators have proposed that the earliest event in radiation-induced malignant transformation is not a rare genetic error in a single cell, but, rather, an epigenetic change in all or a majority of the cells. Evidence in favor of this view from cell culture and other experiments has been presented in the literature (Kennedy et al., 1985; Kennedy and Little, 1984). The induction of genes known to be involved in control and regulation of gene transcription and cell growth control may be the molecular mechanism by which the putative nonrare, nonrandom early events take place. Further research connecting immediate early gene induction to tumorigenesis is required before the importance of this phenomenon to mechanisms of ionizing radiation carcinogenesis is established. Several factors complicate the process of radiation-related cancer studies. There is a long latency period between initial radiation exposure and the development of cancers, usually 5–10 years, but sometimes decades (Nakashima et al., 2008). This long latent period makes it difficult to distinguish between changes caused by irradiation from those that arise from other causes or spontane- ously. Cell studies have revealed that human mammary epithelial cell subtypes exhibit varying sensitivity to carcinogenesis induced by oncogene activation (Ratsch et al., 2001). A recent analysis of the biological effects of ionizing radiation observed that people at increased risk from inherited cancers associated with a genetic defect tend to be at higher risk for radiation-induced cancer (BEIR-VII, 2006). Increasing evidence suggests that interindividual differences in radiation responses may be attributed to variant genes with functional polymorphisms. These observations have helped to shape current theories about radiation-induced carcinogenesis. 7.09.5.2 Oncogenes in Radiation Carcinogenesis Oncogenes are genes whose overexpression promotes a cell’s commitment from its normal phenotype to transformation and carci- nogenesis. Overexpression can be due to a mutation in the gene itself or in other genes that modulate its expression. The activation Fig. 7 Pathways involved in cellular response to DNA damage. Reproduced by permission of Kluwer Academic Publishers from Murnane, J. (1995). Cancer Metastasis Review, 14, 17–29. 208 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 28. of an oncogeneresults in uncontrolled cell growthdone of the early steps of carcinogenesisdby increased cell proliferation or suppression of cell death. Oncogene activation is acquired by an external stimulus, such as ionizing radiation (Bowden et al., 1993; Garte et al., 1989), rather than inherited. Several genes have been identified as oncogenes and can be divided into several groups based on normal molecular functions: (1) growth factors and growth factor receptors; (2) signal transduction molecules that facilitate communication between growth factor–receptor complexes and the nucleus; (3) regulators of cell cycle progression; and (4) inhibitors of apoptosis. The disruption of these processes results in a net increase in cell numbers, nudging the cells toward the adoption of a cancer phenotype. 7.09.5.2.1 Ras genes The Ras family of small GTPases includes three highly similar p21 proteins: h-, k-, and n-ras, which are encoded by different genes and which function as molecular switches in cell proliferation, differentiation, and apoptosis signal transduction pathways (Camp- bell et al., 1998; Crespo and Leon, 2000). Ras was the first oncogene whose activation by ionizing radiation was demonstrated via the transfection of DNA from mouse thymomas induced by ionizing radiation and an alkylating agent (Sawey et al., 1987). Shortly thereafter, activated ras oncogenes were detected in several animal cancers, including rat thyroid tumors (Lemoine et al., 1988), rat skin carcinomas (Garte et al., 1990), radiation-induced mouse thymomas (Niwa et al., 1989), and g-ray-induced canine leukemia (Gumerlock et al., 1989). The ras genes are also commonly upregulated in human cancers including adenocarcinomas of the pancreas, colon, lung, thyroid tumors, myeloid leukemia, and nonsmall lung cell cancers (Bos, 1989; Graziano et al., 1999). The frequency of each ras oncogene varies in tumors depending on the tissue of origin of the neoplasia. In addition to increased expression, point mutations of ras genes are commonly observed in many cancers. In half of the patients studied, multiple unique k-ras mutations were identified among pancreatic intraductal adenocarcinoma lesions (Moskaluk et al., 1997). The ras genes are not only directly activated by ionizing radiation but also stimulated by other carcinogenic processes, which themselves are induced by ionizing radiation. Ras expression was stimulated by erbB2 ligand overexpression and erbB2 activation in pancreatic cancer (Tobita et al., 2003; Uegaki et al., 1997). Ras mutations are frequently observed after p53 is mutated or silenced in pancreatic tumors (Barton et al., 1991). The activation or mutation of the ras gene is a common theme in radiation-induced carcinogenesis, even if it is not necessarily the proximate cause of carcinogenesis. 7.09.5.2.2 c-myc gene The c-myc gene encodes a transcription factor that regulates the expression of up to 15% of all genes (Ruf et al., 2001). The over- expression of c-myc can induce gross chromosomal aberrations and gene amplifications (Felsher and Bishop, 1999; Mai et al., 1996). It can also eliminate a safeguard mechanism for genomic stability against ionizing radiation in human mammary epithelial cells by altering regulation of the cell cycle (Sheen and Dickson, 2002). Gross chromosomal abnormalities have been identified in c- myc transgenic mice (McCormack et al., 1998). The overexpression of c-myc is also observed in some but not all mouse osteosar- comas (Merregaert et al., 1986) and mouse thymomas (Bandyopadhyay et al., 1989; Van der Rauwelaert et al., 1988), as well as some human breast cancers (Liao and Dickson, 2000). The overexpression and amplification of c-myc have been observed in mouse tumors induced by ionizing radiation (Niwa et al., 1989). Amplification of the c-myc oncogene was observed in rat skin tumors as a function of radiation dose (Felber et al., 1992). Amplification of c-myc has been observed in breast cancer tumors from atomic bomb survivors (Miura et al., 2008). Antisense c-myc oligonucleotides inhibited the growth of small cell lung carcinoma (Akie et al., 2000), hepatoma (Ebinuma et al., 1999), and breast cancer (Kang et al., 1996), which makes c-myc the current focus of many cancer treatment studies. 7.09.5.2.3 bcl-2 genes The bcl-2 gene family suppresses the initiation of death processes in the cell (Hockenbery, 1992; McDonnell et al., 1989). It is significantly overexpressed in many cancers; for example, bcl-2 is detected 20%–50% of squamous cell carcinomas (Bartkova et al., 1995) and is overexpressed in over 60% of squamous cell carcinoma tumors (Adelaide et al., 1995). Expression of bcl-2 protein has also been detected in the majority of small cell lung carcinoma tumors (Jiang et al., 1996). Coexpression of bcl-2- and ras-induced synergistic malignant transformation was reported in the skin of UV-treated ras transgenic mice (Lee et al., 2007). Patients undergoing radical prostatectomy after radiotherapy had a significantly higher rate of bcl-2 overexpression than did patients who underwent surgery as the initial treatment (Rosser et al., 2003). The bcl-2 gene itself is subject to rearrangement, as observed in many diffuse B-cell lymphomas (Aisenberg et al., 1988; Pezzella et al., 1990). Interestingly, while ionizing radiation can induce the overexpression of bcl-2 in many cell types, increased bcl-2 levels confer resistance to radiation- induced cell killing in some cells. Bcl-2 protected murine erythroleukemia cells from p53-dependent radiation-induced apoptotic cell death and attenuates p53-independent radiation-induced cell death (Fukunaga-Johnson et al., 1995). Bcl-2 inhibited chemotherapy- and radiation-induced apoptosis by regulating the intrinsic, mitochondrial-mediated pathway of apoptosis (Cheng et al., 2001; Danial and Korsmeyer, 2004). Bcl-Xl is overexpressed in a large majority of head and neck squamous cell carcinoma (HNSCC) and is associated with chemoresistance in this disease (Trask et al., 2002). Targeted downregulation of bcl-2 induced apoptosis signaling and death in head and neck squamous cell carcinoma cells (Li et al., 2007). The survival times for mice with nasopharyngeal tumors treated with both bcl-2 antisense oligonucleotides and radiation increased significantly over radiation treatment alone (Yip et al., 2005). Currently, there is considerable interest in the potential therapeutic benefit of targeting antiapoptotic members of the bcl-2 family in cancer treatment. Ionizing Radiation as a Carcinogen 209 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 29. 7.09.5.2.4 erbB2 genes ErbB2,also known as her-2/neu, is a member of the epidermal growth factor receptor family. When complexed with epidermal growth factor, it promotes cell proliferation in normal cells and, when overexpressed, uncontrolled cell growth results. The erbB2 gene is overexpressed in cancer of the bladder (Jalali Nadoushan et al., 2007), breast (Harari and Yarden, 2000; Zaczek et al., 2008), ovary (Vermeij et al., 2008), and many other cancers. Genomic activation of the erbB2 genes was also observed in a significant proportion of invasive epithelial ovarian cancers (Vermeij et al., 2008). In addition, a majority of breast cancer tumors contained abnormal copies of at least one erbB2 oncogene (Zaczek et al., 2008). The expression of erbB2 induced cell immortalization and transformation in human cells (Labudda et al., 1995). Expression of erbB2 enhanced radiation-induced expression of NF-kB, an inhibitor of apoptotic processes, in breast cancer (Guo et al., 2004). UV-induced responses to skin cells were suppressed by the inhibition or knockdown of erbB2 (Madson et al., 2006). Like bcl-2, erbB2 also confers radioresistance in some cases, demonstrating that the relationship between oncogenes and radiation is complex. 7.09.5.2.5 Oncogenes in Radiation Resistance Resistance of cancer cells to radiation-induced killing is a common problem in the treatment of malignant tumors. There are increasing reports that several oncogenes are involved in the development of radiation-resistant phenotypes (Kasid et al., 1993). The addition of an erbB2 antibody to chemotherapy significantly inhibited DNA damage repair, promoting drug-induced killing in target cells (Pietras et al., 1994). Overexpression of ras induced radioresistance in rat cells (McKenna et al., 1990; Miller et al., 1993), while inhibition of ras induced increased radiosensitivity in rodent cells (Bernhard et al., 1996) and human tumor cell lines (Bernhard et al., 1998). Downregulation of bcl-2 enhanced the efficacy or radiation therapy in mice (Yip et al., 2005). The expression of erbB2 induced tumor resistance to radiation therapies in human breast cancer cell lines (Bowers et al., 2001; Cogswell et al., 2000). Blocking antibodies against erbB2 increased radioresistance in clinical trials (Pegram and Slamon, 2000; Pegram et al., 1998). By contrast, the transfection of ras and myc conferred radioresistance to cells (Lemoine et al., 1988; Niwa et al., 1989). A multitude of molecular techniques continue to be studied to improve the efficacy of radiotherapies via the modulation of oncogenes. 7.09.5.3 Tumor Suppressor Genes Carcinogenic processes in cells are promoted by the altered expression of both tumor suppressor genes and oncogenes; however, there are several critical differences between the two types of genes. Oncogenes are normal genes that are overexpressed, either through mutations of the genes themselves or through mutations of other genes that modulate their expression, while tumor suppressor genes are inactivated through inactivation of the affected gene. Both oncogenes and tumor suppressor genes can develop from acquired mutations, while inactivated tumor suppressor genes can also be inherited (BEIR-VII, 2006). While activated onco- genes tend to promote carcinogenesis through unchecked cellular proliferation, deactivated tumor suppressor genes promote carci- nogenesis through impaired DNA damage repair, loss of control of cell division, or decreased commitment to apoptosis (Kinzler and Vogelstein, 1997). The net result is either unchecked cell growth or the replication of cells with unrepaired DNA damage. Onco- genes act as dominant genes, in that the activation of only a single copy of the affected gene is required for effect. The loss of all functional alleles that encode tumor suppressor genes is required before the gene is deactivated (Knudson, 1971). Despite these differences, both genes are similar in that they are both sensitive to damage by ionizing radiation, which can promote carcinogenic responses by either mechanism. The importance of tumor suppressor genes in carcinogenesis was established with the identification of the Rb gene, which prevents cells with damaged DNA from reproducing, as the gene responsible for retinoblastoma (Murphree and Benedict, 1984). Since then, several other tumor suppressor genes that are deactivated by ionizing radiation have been identified. 7.09.5.3.1 BRCA1/2 genes The multifactorial BRCA1 and BRCA2 proteins regulate multiple cellular functions, including DNA damage repair, ubiquitination, and transcriptional regulation (Wang et al., 2000). Mutations in the breast and ovarian cancer susceptibility genes BRCA1 and BRCA2 are found in a high proportion of multiple case families with breast cancer, especially if they also include one or more case patients with ovarian cancer (Ford et al., 1995). BRCA1 mutation carriers have 50%–80% risk to develop breast cancer by the age of 70 (Easton et al., 1995). One defective copy of BRCA1 or BRCA2 in the germline is sufficient for cancer predisposition, but the loss of the second allele is required for cancer development (Friedman et al., 1994; Miki et al., 1994). As a consequence of this defect in homologous recombination, tumors that arise in BRCA carriers are likely to be more sensitive to ionizing radiation (Powell and Kachnic, 2003). Others have shown that BRCA1/2 gene products are essential to the prevention of a group of leukemias and lymphomas (Friedenson, 2007). Consistent with this extensive pattern of interaction, loss-of-function mutations of BRCA1 result in pleiotrophic phenotypes, including growth retardation, increased apoptosis, defective DNA damage repair, abnormal centrosome duplication, defective G2/M cell cycle checkpoint, impaired spindle checkpoint, and chromosome damage and aneu- ploidy (Brodie and Deng, 2001; Venkitaraman, 2002). These phenotypes are not compatible, at least on the surface, with the tumor suppressor functions assigned to BRCA1. It has therefore proposed that mutations in BRCA1 do not directly result in tumor forma- tion, but instead cause genetic instability, subjecting cells to a high risk of malignant transformation (Deng, 2002; Kinzler and Vogelstein, 1997; Scully and Livingston, 2000). Recent investigations in a mouse cell model have revealed that BRCA1 and 210 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 30. BRCA2 control homologousrecombination at stalled replication forks (Willis et al., 2014). The mutated gene products thus lead to an aberrant repair process, which ultimately promotes genomic instability and increases the risk of carcinogenesis. 7.09.5.3.2 p53 genes The gene product, p53, also known as TP53 and protein 53, is a transcription factor that regulates cell control. It can hold the cell cycle in check to facilitate DNA repair or commit the cell to apoptosis if DNA damage is too severe. The p53 tumor suppressor gene is the most commonly mutated gene in human cancer, present in more than 50% of human tumors (Hollstein et al., 1991; Levine et al., 1991; Vogelstein and Kinzler, 1992). Most mutations that deactivate p53 destroy its ability to bind to DNA sequences, result- ing in the accumulation of mutations, loss of cell cycle control, and unchecked cell growth. Radiation-induced p53 mutation has been observed in murine intestinal epithelium (Wilson et al., 1998), human thyroid cells (Namba et al., 1995), human colorectal cancer (Rodrigues et al., 1990), and many other cancerous cells (Lowe et al., 1994). In addition to mutating the p53 gene, ionizing radiation increased p53 expression in human mammary epithelial cells (Sheen and Dickson, 2002), suggesting a mechanism by which ionizing radiation may amplify the effects of mutated p53. The inactivation of p53 results in a significant decrease in radi- ation sensitivity in glioblastoma cells (Yount et al., 1996). In addition to its more direct contributions to carcinogenic processes, tumor suppressor p53 is a regulator of bcl-2 and bax gene expression in vitro and in vivo in murine leukemia (Miyashita et al., 1994), suggesting that modulation of p53 can affect carcinogenesis indirectly through altered regulation of other genes that promote carcinogenesis. 7.09.5.4 Interactions Between Oncogenes and Tumor Suppressor Genes in Radiation-Induced Cancer Although oncogenes and tumor suppressor genes promote carcinogenesis by different mechanisms, there is significant overlap in their mechanisms of effect; furthermore, there is a prevailing theory that multiple interactions of weakly expressing genes modulate radiation-induced tumorigenesis (BEIR-VII, 2006). Interactions between multiple oncogenes, tumor suppressor genes, or both have been implicated in carcinogenesis. Myc suppresses bcl-2 expression in hematopoietic cells (Eischen et al., 2001; Vaux et al., 1988) and precancerous B cells of myc transgenic mice (Maclean et al., 2003). Radiosensitivity of cancer cells also relies on the interaction of multiple oncogenes and tumor suppressor genes. Radiosensitivity of head and neck cancer patients depends on the ratio of p53, bcl-2, and bax protein levels (Csuka et al., 1997). Oncogenic k-ras is signaled through erbB2 (Cengel et al., 2007), suggesting that radiosensitivity is mediated by multiple interactions of oncogenes. The loss of erbB2 expression did not affect radiosensitivity in a panel of tumor cell lines, but radiosensitivity dramatically decreased when both erbB2 and erbB3 expression were inhibited (Dote et al., 2005). These studies suggest that the modulation of radiation-induced responses in the pathogenesis and treatment of cancer cells is quite complex and involve the interaction of several interactive genes. A schematic summary of the interaction of ionizing radiation with DNA and downstream oncogene effects within the cell cycle is shown in Fig. 7. 7.09.5.5 Genomic Instability The genomic integrity of normal cells, as they operate and proliferate, is maintained by complex metabolic systems; however, genetic mutations accumulate over time, contributing to both aging and cancer (Busuttil et al., 2007; Szilard, 1959). Genomic insta- bility refers to abnormally high rates of genetic changes that occur spontaneously and are passed onto subsequent generations. The loss of genetic integrity is associated with tumorigenesis (Kinzler and Vogelstein, 1997; Lengauer et al., 1998). Acquisition of genomic instability is generally attributed to accumulated mutations in genes that regulate DNA damage repair, such as ATM and BRCA1/2 (BEIR-VII, 2006). There is also evidence that the instability of telomeresdDNA sequences that protect the end of the chromosome from destruction during replicationdis a common feature of carcinogenic development (Bacchetti, 1996; Murnane and Sabatier, 2004). Shortened telomeres have been associated with accelerated aging and increased lymphoma in mice (Espejel et al., 2004). It has also reported that the loss of a single telomere resulted in instability of multiple chromosomes in a human tumor cell line (Sabatier et al., 2005). A recent analysis of the biological effects of radiation observed that individuals at increased risk from inherited cancers associated with a genetic defect tend to also be at higher risk for radiation-induced cancer (BEIR-VII, 2006). In addition, the same study proposed that multiple genetic dysfunctions promote carcinogenesis, which is consis- tent with the vulnerable genetic state induced by genomic instability. 7.09.6 Radiation Carcinogenesis Risk Modeling It is the biological response to radiation exposure that determines the long-term risk of cancer in the exposed individuals. Yet, only physical measurements of the exposure environment are typically used to derive estimates of cancer risk. This approach assumes that a physical measurement can be equated to a biologically meaningful dose. The risk coefficient that is applied to a personal dose assumes that all individuals exposed respond to radiation in the same way as a theoretically “average” person exposed to typical radiation sources. It is now established that individual radiation response will be governed by genetic makeup. As described above, there are multiple gene products interacting to respond to damage induced by radiation. The genetic heterogeneity in many of these [yet Ionizing Radiation as a Carcinogen 211 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 31. unidentified] gene productsresults in a range of susceptibilities to ionizing radiation. The notion of individual susceptibilities cannot be translated into practical action plans and policies in large terrestrial populations at this time. However, a small cohort of individuals, such as International Space Station astronauts, could be evaluated for individual risk prediction by assessing indi- vidual response to radiation exposure. 7.09.6.1 Risks Effect Definition Deterministic effects are those effects that occur only above dose thresholds, for example, acute radiation sickness, damage to the central nervous system (CNS), and cataracts. The severity of the effect increases with increasing dose. The acceptable risk of a deter- ministic effect occurring within a radiation worker is zero, and occupational dose limits ensure that these thresholds are not exceeded. Stochastic effects are those effects whose probability of occurrence, but not their severity, is a function of the dose. They occur without a dose threshold. The dose to radiation-induced cancer risk conversion factors is highly dependent on gender, age at expo- sure, and tissue type. Stochastic effects include cancer, hereditary effects, or neurological disorders. With regards to hereditary effects, the National Academy of Sciences estimates of the probability that radiation will increase genetic defects, for a dose of 0.1 Sv, are as follows: first generation of exposedd0.15–0.4Â10À 3 ; all generations subsequent to exposured1.15–2.15Â10À3 (the background rate, for reference, is 3.6–4.6Â10À3 ). The risk rate is mainly due to dominant gene mutations and chromosomal diseases; however, the risks for malformation and growth/developmental retardation are also high, with most sensitive period being between 8 and 25 weeks gestation. The probability of mental retardation with exposure during this time period is 0.04 for each dose of 0.1 Sv. Another means of applying dose limitations is based on the risk of “radiation detriment” (Table 6). This more modern approach to dose limitation uses the concept of detriment to measure total risk to an individual. Components of overall radiation exposure detriment are (1) risk of fatal cancer, (2) allowance for relative years of life lost for specific fatal cancers, (3) allowance for morbidity, and (4) allowance for the risk of hereditary diseases. Upton estimates of the lifetime risk of cancer incidence and fatality from 1.0 Sv exposure to 10, 000 persons found the highest incidence in breast (McKenna et al., 1990), lung (ICRP, 1991), and thyroid (Hallahan et al., 1994), but the highest fatality in lung (Heidenreich et al., 1999) and breast (Fornace et al., 1992; Upton et al., 1986). 7.09.7 Projecting Lifetime Carcinogenesis Risk Radiation-induced (excess) cancer risks and uncertainties for carcinogenesis can be based, in part, on results of follow-up studies of Japanese atomic bomb survivors (Petersen and Abrahamson, 1998). The standard linear quadratic exponential (LQE) excess cancer relative risk (ERR) model based on cell initiation and killing was modified to add a third element, radiation-induced carcinogenesis at high doses, based on stem-cell repopulation kinetics. (Brenner, 2011) Wheldon and Lindsay, as well as Dasu, et al, developed 2- stage mutational models which produced a typical bell shaped dose-response curve with diminishing cancer risk at doses 20 Gy, presumably due to cell sterilization versus transformations (the dose response in a competition model, which takes into account the probability of cell survival in addition to the probability of induced DNA mutations, after irradiation). A similar curve bending into a bell-shape was observed by Schneider and Walsh when extending the A-bomb epidemiological data to include exposure dose cate- gories 4–6 Sv and 6–13 Sv. (Schneider, 2011) A cohort population where this comparison technique can be applied includes manned spaceflight activities. In the case of space-based radiation exposure, projected risks based on the Japanese data are Table 6 Contribution to detriment for average worker (average worker dose of 0.1 Sv) Tissue Fatal cancer (%) Hereditary Aggregated detriment (%) Bladder 0.024 0.024 Bone marrow 0.04 0.083 Breast 0.016 0.029 Colon 0.068 0.082 Liver 0.012 0.013 Lung 0.068 0.064 Esophagus 0.024 0.019 Ovary 0.008 0.012 Skin 0.002 0.003 Stomach 0.088 0.080 Thyroid 0.006 0.012 Gonads 0.06 0.080 Other 0.045 0.053 Total 0.4 0.56 212 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 32. appropriate for thefraction of total risk attributable to low LET geomagnetically trapped protons, since the Japanese experience was primarily determined by low LET radiation exposure. The assessment of risk from energetic ion exposure is compounded by addi- tional requirements for empirical and subjective “lack-of-knowledge” uncertainties in charge, energy, LET, and end points from the galactic cosmic and energetic solar source particulate radiation of higher LET. Information regarding these radiation sources is incomplete and will require a number of years of development before a more complete picture evolves. However, low LET risk coefficients can be obtained from fitted dose–response models of cancer incidence among the Japanese survivors (Preston et al., 1994; Thompson et al., 1994). The two statistical models used most frequently for fitting cancer incidence dose–response relationships are the additive and multiplicative Poisson regression models. Excess relative risk is obtained by fitting multiplicative models and is defined as the ratio of cumulative risk at a given age among the exposed to cumulative risk among the nonexposed. Absolute risk is obtained by fitting additive models and represents the difference between cumulative risk among the exposed and the nonexposed groups. Fitted risks are used for projecting cumulative risks among exposed populations. Uncertainty in projected cumulative risk is assessed by use of iterative Monte Carlo analysis to “fold” together probability density functions for several risk projection models and bias correction factors. What follows describes life table methods used during each iteration to obtain “quantiles” of excess lifetime risk, xI,r. Methods discussed in the following sections are described in detail else- where (Peterson and Cucinotta, 1999). Let ai represent the age at exposure for the ith exposure (i¼1, 2, ., n). As an example, for exposure at ages 32, 36, and 54, the notation is a1¼32, a2¼36, a3¼54, and n¼3. Each iteration starts by randomly selecting the risk projection model to be used for generating a double-decrement life table during the iteration. This is accomplished by obtaining a pseudorandom uniform variate U(0,1) from the computer and selecting the projection model according to the criteria xr ai ; t À Á ¼ x˛ ai ; t À Á Additive 0:0 U 0; 1ð Þ 0:33 xp-NIH ai; tð Þxl;c tð Þ NIH multiplicative 0:33 U 0; 1ð Þ 0:66 xp ai; tð Þxl;c tð Þ Multiplicative 0:66 U 0; 1ð Þ 1:0 8 : where x˛(ai,t) is the quantile for excess absolute risk coefficient used in the additive projection model, xr-NIH(ai ,t) is the quantile for excess relative risk used in the National Institutes of Health (NIH) multiplicative projection model, and xr(ai ,t) is the quantile for excess relative risk used in the multiplicative projection model. Excess absolute risk coefficients in cases/104PYSv are from Table X of Thompson et al. (1994). Excess relative risk coefficients for the NIH multiplicative model (ICRP, 1991), xr-NIH(ai ,t), are calculated by dividing cumulative absolute risk by cumulative baseline risk for the 29-year follow-up period (1958–1987), which began 13 years after the bombings in 1945 (i.e., j¼13, 14, ., 42), given as xrÀNIH ai; tð Þ ¼ P42 j¼13 À x;˛ ai þ jð Þxs ai þ jð Þ P42 j¼13 À xl;c ai þ jð Þxs ai þ jð Þ where xl,c(ai þj) is the quantile for rate of spontaneously occurring cancer among a US population at age ai þj, and xS(ai þj) is the quantile for survivorship function at age aþj in the single-decrement life table for a nonexposed US population. Excess relative risk coefficients in percent per Sv are from Table X of Thompson et al. (1994) (solid cancer excess relative risks in Table X of Thompson et al. (1994) are age adjusted and are not presented as a function of attained age (t); however, excess absolute risks for leukemia in Preston et al. (1994) are given as a function of age at exposure and attained age). During life table calculations, the total excess risk in life table interval t from multiple exposures at different ages is xL;r tð Þ ¼ Xn i¼1 xH aið Þxr ai; tð Þ; a1 þ L t min an þ L þ P; 100f g where xr(ai,t) is the risk coefficient specified above and xH(ai) is the quantile for dose equivalent received at age ai given as xH aið Þ ¼ X j ð Sj Eð Þfj E; x; yð ÞQ Sj Eð Þ À Á dE where Sj(E) is the stopping power of ion j or energy E(MeV mÀ1 ), f(E,x,y) is the particle flux at shielding depth x and tissue depth y, and Q(Sj(E)) is the quality factor. Once total risk at each life table interval is obtained, that is, xL,r(t), the quantile of cumulative risk during each iteration is xI;r ¼ ðmin anþLþP; 100f g a1þL xL;r tð ÞxS t; dð Þdt where a1 is the first age at exposure, L is the latency period during which risk is not applied, an is the last age at exposure, P is the plateau period over which risk is applied, and xS(t,d) is the quantile for the age-specific survivorship function from the double- decrement life table. Cumulative risk at each iteration is then folded together with the NCRP (1997, 2000) bias correction factors in the form xmix ¼ xI; r xD xP xT xQ =xE Ionizing Radiation as a Carcinogen 213 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 33. Where xD isa quantile for bias correction of random and systematic errors in DS86, xT is a quantile for bias correction in transfer of risk from Japan to the United States, xP is a quantile for bias correction of uncertainty in projection over a lifetime, xQ is a quantile for bias correction of unknown uncertainty, and xE is the uncertainty distribution for the dose and dose-rate effective factor. After 5000 iterations are performed, the various quantiles of xmix are ranked in ascending order. Central estimates of risk are then based on the median, m(xmix), with 90% subjective confidence limits based on the 5th and 95th percentiles of the ranked quantiles. The process of randomly selecting either the additive, NIH-multiplicative, or multiplicative model for life table calculations during each interval is the basis for the “mixture model” of risk. The probability of causation (PC), at various ages, has been estimated for leukemic and nonleukemic cancers (all cancers except leukemia) for males exposed to 1 Sv of radiation, accounting for age of exposure. From the estimates, the greatest excess risk of 5.59% occurs at age 90 for males exposed to 1 Sv at age 20. Overall, risk decreases with increasing age at exposure and approaches 1% for males exposed at age 90. The median risk of leukemia among males showed a similar trend but was greatest for exposure at age 40, where it approached 4.3Â10- 3 at age 90. The effect of the short 40-year plateau period during which leukemia risk is accumulated can be noticed in the curve for males exposed at age 20. When exposed at age 20, risk accumulation starts at age 22 (20 þ 2-year latency) and continues until age 62 (20 þ 2 þ 40). Cumulative risk of leukemia from exposure at older ages is higher than risk from exposure at younger ages because the excess absolute leukemia risk is added to baseline rates. Among US males, baseline rates of leukemia at ages 60–64 are one-third the baseline rates at ages 75–79 and less than a fourth of the rates beyond age 85. If plateau periods for leukemia were longer than 40 years, a different picture would emerge (Aguiar, 1998; Preston, 2005; Preston et al., 1994). The highest PC for nonleukemic cancers was 31.3%, and this occurred at age 40 among those exposed at age 20. For leukemia, the PC at age 30 among those exposed at age 20 was 95.4%. While PCs for nonleukemic cancers decrease uniformly with increasing age at exposure, PCs for leukemia follow the same trend as exposure age increases and remain fixed after the end of the plateau period in which risk is accumulated. For females (Fig. 8), the trends in nonleukemic malignancy risk at various ages, accounting for age at exposure, were similar to those observed for males. Leukemia risks for females, based on the sum of excess absolute risks and baseline risk, are generally lower than those for males because the female risk coefficients are lower (see Fig. 4 of Preston et al., 1994) and female baseline rates are about half of those for males [at all ages]. Some general conclusions on the risks presented from the PC and ERR estimates are as follows: For males, the highest nonleu- kemic cancer risks observed for exposure at ages 20, 30, 40, 50, 60, and 70 were 5.6% at age 90, 4.3% at age 90, 3.3% at age 100, 2.4% at age 100, 1.6% at age 100, and 1.1% at age 100, respectively. For females, the highest nonleukemic cancer risks observed for exposure at ages 20, 30, 40, 50, 60, and 70 were 4.8% at age 80, 5.4% at age 100, 4.1% at age 90, 2.9% at age 90, 2.0% at age 90, and 1.3% at age 100, respectively. PCs for leukemia approached 95% for males exposed at age 20 and 90% for females exposed at age 20; for nonleukemic cancer, PCs approached 30% for exposure at age 20. Lifetime cancer incidence risks approaching 3% for expo- sure to 1 Sv could be expected for males exposed at age 40 and females exposed at age 50. 7.09.8 Radiation Protection from Ionizing Radiation 7.09.8.1 Shielding Radiation protection aims to mitigate or eliminate the hazardous effects of ionizing radiation on human health and well-being. In principle, this can be achieved by physical shielding, by setting acceptable exposure limits, and by radioprotective agents and/or therapy. There are three means to reduce exposure to ionizing radiation: (1) increasing distance from the radiation source, (2) mini- mizing the time of exposure, and (3) using shielding. Fig. 8 Site-specific excess relative risk (ERR) and 90% confidence intervals, expressed in terms of risk for females aged 30 at exposure. 214 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 34. The first twomeans to reduce exposure to radiation are the simplest to implement for terrestrial radiation protection. However, these means are not suitable for human spaceflight activities, which embody the most exposed population to ionizing radiation: astronauts (Durante and Cucinotta, 2008; NASA, 2004). The most commonly used means of protection for terrestrial radiation workers is shielding; that is, the placement of material between the human and the radiation source to reduce the exposure. By defi- nition, shielding is the use of materials to mitigate the effects of the incident radiation by (1) reducing the intensity of the radiation (e.g., attenuation of X-rays by absorption of photons in a lead curtain), (2) changing the incoming properties of primary radiation, which could produce secondary particles and nucleic fragments, or (3) both. Thus, in principle, shielding alone should be able to reduce exposure by attenuating the radiation and reducing the dose rates. The composition and the thickness of a material will affect its ability to shield radiation. Shielding against low LET ionizing radiation can be efficiently achieved for terrestrial radiation workers by use of thick and dense materials (e.g., lead). However, high LET and high energy radiation is very penetrating and therefore prob- lematic. This form of radiation characterizes one of the principal health threats to astronauts venturing beyond low earth orbit (Chancellor et al, 2014). The dose equivalent can be reduced by thin or moderate shielding; however, as the material thickness increases, shielding effectiveness drops due to the production of a large number of nuclear by-products, that is, secondary particles (neutrons and atomic nuclei) caused by nuclear interactions of the high LET particles with the shielding material (CERSSE, 2008Thus, heavier elements, such as aluminum, will produce more secondary radiation than lighter elements, such as hydrogen and carbon. The secondary radiation particles are of significant concern due to their potentially higher quality factor (Q) as compared to the incident radiation. Thus, these secondary particles, resulting from interaction with the shielding material, may be, in fact, more biologically harmful than the primary radiation. Recent investigations with cell cultures indicate that this may, indeed, be the case (Hu et al., 2014). Consequently, a great deal of research has been conducted in the area of shielding properties, mainly for human spaceflight purposes. Computational models, as well as experiments carried out in space analogues or during spaceflight missions, have shown that space shielding effectiveness per unit mass is the highest for hydrogen and decreases with increasing atomic number (Durante et al., 2005; Lobascio et al., 2008; Miller et al., 2003; Simonsen et al., 2000; Vana et al., 2006; Wilson et al., 1995b). Although liquid hydrogen would offer maximum performance as a shield material, it is not practical for spaceflight applications (Durante and Cucinotta, 2008; Wilson et al., 1995a). Polyethylene is known to have excellent shielding properties due to its low density, coupled with its high hydrogen content, and has been installed in the sleeping quarters of the International Space Station (ISS) to reduce the average astronaut dose and dose equivalent (Miller et al., 2003; Shavers et al., 2004). Additionally, polyethylene-fiber reinforced epoxy matrix composite is 50% better at shielding against solar flare radiation and 15% better at shielding against galactic cosmic rays (GCR), as compared to aluminum (Kaul et al., 2004). However, optimum shielding would only reduce effective CGR dose by no more than 35% (Cucinotta and Durante, 2006). Consequently, high energy GCR will still produce biologically dangerous secondary radiation in tissue and, therefore, other types of countermeasures will have to be implemented to reduce risks associated with space exploration class missions. 7.09.8.2 Dose Limitation Human beings are exposed to a small amount of daily natural background radiation from a variety of sources, such as air, soil, rocks, water, and, indeed, the rest of the universe. The average worldwide annual exposures to natural radiation sources, which include both high and low LET radiation, is generally expected to be in the range of 1–10 mSv, with 2.4 mSv being the present estimate of the central value (UNSCEAR, 2000). In the United States, the majority of exposure to background ionizing radiation comes from exposure to radon gas and its decay products; as a result, the annual background exposures are slightly higher, that is, 3 mSv (BEIR-VII, 2006). Fig. 3 summarizes the approximate sources and relative amounts of high and low LET radiation that comprise the natural background exposure. 7.09.8.3 Radioprotectants/Chemoprevention Pharmacologic agents that may lessen the cellular damage associated with ionizing radiation exposure are being developed for use, alone or in combination. These protective agents can be broadly classified into three different types: radio-modulators, radio- protectors, and radio-mitigators (Vasin, 2014). Radio-modulators act in a prophylactic manner to increase the baseline resistance of an organism to radiation exposure. These agents may include antioxidants (e.g., N-acetyl cysteine), desmutagens (e.g., polyphenols), and others. Researchers have shown that combination agent formulas may have promise in diminishing multiple pathways of radiation injury, while lessening the side effects often found with single agent use, mainly by reducing the dose of any single agent required (Jones et al., 2007). The radio-modulator formulas that are being developed take advantage of chemoprevention agents, which have shown promise in reducing oxidative damage from a variety of environmental exposures. Two specific examples for such agents are selenium and omega-3 fatty acids. Selenium, in the form of selenocysteine, is the catalytic center of glutathione peroxidase (GPX), which detoxifies hydrogen peroxide and lipoperoxides generated by free radicals and ROS. However, selenium may have antitumorigenic actions without GPX (Mayne et al., 1998). Omega-3 fatty acids (e.g., DHA and EPA), along with soluble fiber, may be protective against radiation-induced colon cancers, by protecting DNA and by upregulation of apoptotic gene expression in exposed cells (Lupton, 2001). The protective effect of certain classes of radio-modulators, such as antioxidants, has been demon- strated in cell cultures as well as in animal models of radiation-induced toxicity and carcinogenesis (Kennedy, 2014). Some possible examples of chemoprevention agents and their mechanism of protection are listed in Tables 7 and 8 (Stanford and Jones, 1999). Ionizing Radiation as a Carcinogen 215 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 35. Radio-protectors are agentsthat are administered shortly before an exposure event and act directly to protect cellular compo- nents and/or to oppose the action of radiation-induced free radicals. One family of compounds that comprise this group is thiols, also known as mercaptans. Amifostine (WR-2721) is one such compound and is, at present, the only systemic agent that is FDA approved specifically for use in radiation exposure, being used clinically for the prevention of mucositis in radiotherapy patients. Radionuclide eliminators are a separate group of compounds that work to bind or block the uptake radioactive isotopes that find their way into the body; these include potassium iodine, Prussian blue, and calcium/zinc DTPA and are used for iodine-131/125, cesium-137, and transuranics (e.g., plutonium), respectively. While available for use, as indicated, the scope of these compounds is fairly narrow. A number of radio-protectors are currently in various phases of development and FDA approval, with a general focus on the prevention or amelioration of acute radiation syndromes (Singh et al., 2015). A detailed discussion of the spectrum of acute radiation syndrome, along with its management, is beyond the scope of this article. However, the development of countermeasures in this area is of great interest, as the benefits of efficacious compounds may extend beyond the treatment of acute symptoms and may have an impact on the long-term consequences of radiation exposure, including carcinogenesis. Finally, radio-mitigators are compounds that act on a systemic level to accelerate postradiation recovery and prevent radiation- related complications. These may include compounds such as steroids, growth factors, antibiotics, and immune adjuvants. In addi- tion to radiation-specific countermeasures, the management of radiation exposure would also include general supportive measures such as intravenous fluid administration, analgesics, and blood product replacement. A detailed list of radiation countermeasures, both approved and under investigation, is provided in Table 9 (Singh et al., 2015). 7.09.8.4 Immunoprotection Strategies Some researchers contend that many aspects of radiation toxicity, including diffuse systemic bioeffects, occur due to secondary even- tsdpossibly bystander effectsdas a result of the liberation of cellular components that occur during nonprogrammed cell break- down (i.e., radiation-induced cellular necrosis). The underlying assumption is that some of these cellular components may have inherent toxic properties, which may account for a variety of observed systemic effects (Maliev et al., 2007a,b). It has previously been hypothesized that, given such a mechanism of toxicity, it may be possible to develop an immunological approach to preven- tion of radiation-induced cellular injury via administration of a hyperimmune serum or, perhaps, even a vaccine against the toxic breakdown species (Maliev et al., 2007a,b). Subsequent research has been successful in identifying specific radiation determinant (SRD) toxins, which appear to be breakdown products of radiation necrosis, consisting primarily of glycoproteins with high enzy- matic activity (Jones et al., 2013). Of note, these SRDs, when administered to nonirradiated animals, appeared to produce many of the symptoms of radiation toxicity, thus lending weight to the aforementioned proposed mechanism. During subsequent investi- gations, SRDs were isolated from the lymph of irradiated animals and were used to develop both a hyperimmune serum and vaccine; these were administered to different groups of animals, including pigs and cattle, which were subsequently irradiated with an LD100 dose of total body g-radiation. Administration of hyperimmune serum and/or an SRD radiation vaccine signifi- cantly improved overall survival, as compared to placebo-treated controls, for up to 360 days postexposure (Jones et al., 2013). These findings demonstrate an evolving understanding of the biological mechanisms underlying the deleterious effects of ionizing radiation and suggest promising new approaches toward mitigating them. 7.09.9 Conclusions This article presents substantial evidence to support the notion of ionizing radiation as a carcinogen, and it is hopefully convincing that it should be regarded as such. Environmental exposures to ionizing radiation are now commonplace, especially among those whose occupations require them to work with radioactive material, radiation-producing devices, or to leave the planetary surface and protective geomagnetosphere. Although ionizing radiation possesses, in high doses and dose rates, cell-killing proper- tiesdhence its use as a form of cancer therapydit undoubtedly has the potential to be carcinogenic at lower doses and dose rates. Table 7 Natural sources of chemoprevention agents Allium and N-acetyl cysteine (diallyl sulfide) Onions, garlic, chives, scallions Sulphoranes, indoles, and isothiocyanates (dithiolthiones, indole-3-carbinol) Cruciferous vegetables (e.g., broccoli, cauliflower, kale, cabbage) Isoflavones and phytoestrogens (also protease inhibitor Bowman–Birk) Soybeans (including tofu, soy milk) Terpenes and ascorbic acid (perillyl alcohol, limonene) Citrus fruits (especially lemon peels), cherries, tomatoes Curcumins Tumeric Carotinoids, lycopene, lutein, antioxidants Yellow vegetables, fruits (e.g., carrots, tomatoes, squash) Polyphenols and flavonoids (EGCG, thearubigens, theaflavins, resveratrol) Green and black teas, fruits, wine (Phenolic acidsdellagic acid, ferulic acid) Whole grains, nuts, tomatoes, carrots, citrus fruits Modified from Stanford, M., Jones, J.A. (1999). Acta Astronautica 45, 39–47. 216 Ionizing Radiation as a Carcinogen Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
- 36. Table 8 Agentsthat may provide protection from ionizing radiation-induced bioeffects Agent Activity WR-33278 (symmetric disulfide of WR-1065) Binds DNA, protects from ionizing action WR-151327 Increased survival in mice radiated by neutrons Lazaroid Protects neural tissue for linear accelerator runs Diethyldithiocarbamate (DDC) Raised XRT LD50 in mice Other agents Mechanism of action Allylic sulfides, N-acetyl cysteine Enhance GSH activity, thereby deactivating carcinogens Ellagic acid Decrease DNA methylation, scavenge eÀ Polyphenols Scavenge peroxy radicals, arachidomic acid, and carcinogen metabolism modulation Flavonoids Inhibit arachidomic acid metabolism, tyrosine kinases Monoterpenes (limonene, per alc) Inhibit ras farnesylation and oncogene activation Isothiocyanates Enhance GST activity, thereby directly block carcinogen damage Oltipraz (dithiolthione) Prevent carcinogen-binding DNA, via reduced glutathione DFMO Inhibit polyamine metabolism (ODC) NSAIA, ASA Decrease inflammation, arachidomic acid metabolism via COX-1,2 inhibition, decreased growth factor activation 3-Indole carbonyl Multiple pathways of action Genistein Inhibits ormithine decarboxylase (ODC), tyrosine kinases Quercetin Vitamin E Fat-soluble antioxidant of peroxidation Vitamin C Water-soluble antioxidant, direct-acting reducing agent Vitamin D3/Ca2 þ Differentiation agents, growth inhibition Carotinoids/retinoids Water-soluble antioxidants, quench singlet oxygen, apoptosis, differentiation, inhibit angiogenesis, immunologic modulation Phytoestrogens Inhibit oxidative DNA reactions Isoflavones Inhibit oxidative DNA reactions Selenium Water-soluble antioxidant via glutathione peroxidase enzyme component Glutathione Enhance clearance of carcinogens, toxins (via glutathione peroxidase, glutathione reductase) Agent classification Specific action Bioantimutagensdvanillin Enhance postreplication recombinational repair Protease inhibitors (e.g., Bowman–Birk inhibitor) Inhibit error-prone repair induced by proteases Desmutagensdinhibit carcinogen formation Polyphenols Deactivate/detoxify carcinogens Isothiocyanates Affect nitrosamine metabolism Antioxidantsdscavenge reactive electrophiles NAC Electrophile scavenger, Increases level of DNA repair Ellagic acid Glutathione-S-hydrogenase activity enhancement Allylic sulfides GSH enhancer Scavenge/clear oxygen radicals (Continued) IonizingRadiationasaCarcinogen217 ComprehensiveToxicology,ThirdEdition,2018,183–225 Author's personal copy
- 37. Table 8 Agentsthat may provide protection from ionizing radiation-induced bioeffectsdcont'd Thiol-containing molecules React with hydroxyl radicals Carotenoids: b-carotene, lycopen, a-tocopherol Interacts with/quenches singlet oxygen Scavenge peroxy, singlet oxygen, superoxides Cu–Zn superoxide dismutase, dimethylsulfoxide (DMSO) Destroys superoxide radicals GSH, GST enhancers GSH reacts with alkyl-peroxy radicals flavonoids, polyphenols Alter arachidonic acid metabolism Aspirin Cyclooxygenase inhibition, prevent reactive species Nonsteroidal anti-inflammation (sulindac, celecoxib (COX-2)) Cyclooxygenase inhibition, inhibit phospholipase C Flavonoids Cyclooxygenase inhibition, inhibit lipoxygenase and free radical production Curcumin Inhibit lipoxygenase and free radical production Polyphenols Inhibit lipoxygenase and free radical production Antiproliferatives/antiprogression modulate signal transduction Flavonoids Inhibit protein kinase C (PKC) Glycyrrhetinic acid Inhibit PKC Modulate growth factor (GF) activity Tamoxifen Increases transforming growth factor b, decreases IGF Diminish oncogene activity Genestein, quercetin Inhibit tyrosine kinases Monoterpenes Inhibit ras, G-protein farnesylation Stimulate or restore tumor suppressor function No proven agents currently Inhibit polyamine synthesis Difluoromethylornithine (DFMO) Inhibit ODC Retinoids, tamoxifen Inhibit ODC induction Correct DNA methylation problems Folic acid Regulates intracellular methyl metabolism Induce terminal differentiation Retinoids Affect binding proteins Calcium þ vitamin D3 Act on nuclear receptor Protease inhibitors Improve DNA repair fidelity Immune modulators (GM-CSF) Enhance tumor cell surveillance, clearance Restore immune response Selenium Increased NK cell killing of tumor cells Vitamin E, retinoids Immune stimulants NSAIAs Inhibit CyOx and PGE2 production Increase intercellular communication Retinoids, b-carotene Affect gap junctions Induce apoptosis Genestein, retinoids, tamoxifen Initiate programmed cell death in transformed cells Antiapoptosis IGF-1, c-Akt, IL-1, HGF, TNF-a, VEGF, bFGF, MAP kinase Inhibit cell death, enhance normal cell survival Modified from Stanford, M., Jones, J. A. (1999). Acta Astronautica 45, 39–47. 218IonizingRadiationasaCarcinogen ComprehensiveToxicology,ThirdEdition,2018,183–225 Author's personal copy
- 38. Tissues exposed toradiation, intentionally or inadvertently, during medical therapy or industrial employment, are vulnerable to carcinogenesis and should be under close surveillance for many years postexposure. Where possible, shielding materials must be used vigilantly and judiciously and ongoing studies support the emerging role of pharmacologic and/or immunologic radioprotec- tive agents. The existence of numerous exposed human populations with radiation-induced human cancers, alongside the availability of a myriad of homologous animal models, holds great promise for the eventual understanding of the basic mechanisms, dose– response relationships (at all dose levels), and the human risk of carcinogenesis arising from exposure to ionizing radiation. Under- standing the biochemical and biophysical mechanisms for the induction of radiation carcinogenesis will require a multidisciplinary investigative approach. This understanding will, in turn, allow for the formulation of effective and evidence-based preventive strat- egies, which may be our best approach to the control and mitigation of environmentally induced human cancer. See also: 7.08. Ultraviolet Radiation as a Carcinogen. References Adelaide, J., Monges, G., Derderian, C., Seitz, J. F., Birnbaum, D. (1995). British Journal of Cancer, 71, 64–68. Aghamohammadi, S. Z., Morris, T., Stevens, D. L., Thacker, J. (1992). Mutation Research, 269, 1–7. Aguiar, R. C. (1998). Leukemia and Lymphoma, 29, 17–26. Table 9 Radiation Countermeasures Ionizing Radiation as a Carcinogen 219 Comprehensive Toxicology, Third Edition, 2018, 183–225 Author's personal copy
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