A global perspective of radiation-induced signal transduction pathways in cancer therapeutics

Radiation is a well established therapeutic modality for the treatment of solid tumors. By merging molecular biological approaches with radiation biology, a significant number of signaling events elicited by ionizing radiation have been delineated. These signaling pathways include events leading to cell cycle arrest, apoptosis or cell survival. There are two major signaling events that affect radiation response. One is the intrinsic/constitutive pro-survival signaling event that is present in proliferating tumor cells while the other is "induced pro-survival event" in response to radiation, both of these events confer resistance to the killing effects of radiation. In this review, signaling pathways that lead to either apoptosis or survival of cells following ionizing radiation are discussed in detail. In addition, mechanisms of action for gene/drug based inhibitors that modulate the expression and function of various genes and gene products involved in pro-survival signaling pathways are described. Further, novel strategies to abrogate the "induced radiation resistance" leading to enhanced therapeutic efficacy of ionizing radiation have been proposed. These novel strategies include the use of radio-gene therapy, low dose fractionated radiation therapy as a chemopotentiator and therapeutic utility of high radiation dose induced bystander effect. The complete understanding of the molecular pathways leading to apoptosis/survival of cells following ionizing radiation will help in tailoring more effective novel strategies and treatment modalities for complete eradication of cancer.     

Energy deposition spectra of negative pions and their application to radiation therapy

Summary Treatment planning for pion radiation therapy must take into account changes in radiation quality within the patient. At the biomedical channelE3 of SIN (Swiss Institute for Nuclear Research) microdosimetric measurements have been performed to investigate radiation quality within pion irradiated phantoms. Results are presented in terms of microdosimetric spectra and derived quantities. As expected marked differences are observed between dose peak and plateau for narrow pion beams. The influence of simulated site diameter on measured spectra has been found to be more pronounced in the plateau region than in the peak. Investigation of the influence of peak width on radiation quality revealed a dilution of the high-LET dose fraction for broader peaks.     

Interrelationships amongst radiation-induced genomic instability, bystander effects, and the adaptive response

Over the past two decades, our understanding of radiation biology has undergone a fundamental shift in paradigms away from deterministic "hit-effect" relationships and towards complex ongoing "cellular responses". These responses include now familiar, but still poorly understood, phenomena associated with radiation exposure such as bystander effects, genomic instability, and adaptive responses. All three have been observed at very low doses, and at time points far removed from the initial radiation exposure, and are extremely relevant for linear extrapolation to low doses; the adaptive response is particularly relevant when exposure is spread over a period of time. These are precisely the circumstances that are most relevant to understanding cancer risk associated with environmental and occupational radiation exposures. This review will provide a synthesis of the known, and proposed, interrelationships amongst low-dose cellular responses to radiation. It also will examine the potential importance of non-targeted cellular responses to ionizing radiation in setting acceptable exposure limits especially to low-LET radiations.     

Ultraviolet-B radiation leads to a reduction in free polyamines in Phaseolus vulgaris L.

Increasing levels of UV-B radiation (UV-B) at the Earth's surfaceare among a suite of anthropomorphic stresses that influence plant growth anddevelopment. Several stress reactions have been shown to occur in response toUV-B, including a stimulation of antioxidant pathways and altered or increasedpigmentation. Polyamines, plant growth regulators that have been shown toincrease during stress, have also been implicated in the UV-B response. Weinvestigated the effects of UV-B radiation on the UV-B sensitive legumePhaseolus vulgaris L. Top Crop over atwo-week period. Total free polyamines showed marked decreases in response toUV-B radiation, primarily due to a decrease in putrescine. The reduction infreepolyamines was correlated with UV-B induced chlorophyll loss.     

Genomic instability, bystander effects and radiation risks: implications for development of protection strategies for man and the environment

The last few years has seen what people are now referring to as a "shifting Paradigm" in our way of thinking about radiation effects on biological systems. The concept of the central role of DNA damage due to double strand breaks induced by a radiation "hit" has been itself hit by many studies showing persistent effects in the distant progeny of radiation exposed cells. This phenomenon is known as radiation induced genomic instability. More recently evidence has been accumulating that not even the parent cell need be exposed to radiation (the bystander effect). The new paradigm suggests that cellular stress responses or damage signalling through a range of signal transduction pathways are involved and that cell-cell contact or secretion of damage signalling molecules can induce responses in undamaged and unirradiated cells. Are these new effects relevant to risk assessment, or does it matter HOW radiation affects cells if we have good epidemiological evidence of which to base our risk estimates? The aim of this paper is to introduce the new concepts and to consider reasons why they might alter our methods of risk estimation. The paper also considers the impact of the new concepts on environmental protection and discusses the need for research in the field of comparative radiobiology if we are to develop policies which can adequately protect biodiversity.     

Non-targeted bystander effects induced by ionizing radiation

Radiation-induced bystander effects refer to those responses occurring in cells that were not subject to energy deposition events following ionizing radiation. These bystander cells may have been neighbors of irradiated cells, or physically separated but subject to soluble secreted signals from irradiated cells. Bystander effects have been observed in vitro and in vivo and for various radiation qualities. In tribute to an old friend and colleague, Anthony V. Carrano, who would have said "well what are the critical questions that should be addressed, and so what?", we review the evidence for non-targeted radiation-induced bystander effects with emphasis on prevailing questions in this rapidly developing research field, and the potential significance of bystander effects in evaluating the detrimental health effects of radiation exposure.     

Adenoviral gene therapy, radiation, and prostate cancer

Viral gene therapy has exceptional potential as a specifically tailored cancer treatment. However, enthusiasm for cancer gene therapy has varied over the years, partly owing to safety concerns after the death of a young volunteer in a clinical trial for a genetic disease. Since this singular tragedy, results from numerous clinical trials over the past 10 years have restored the excellent safety profile of adenoviral vectors. These vectors have been extensively studied in phase I and II trials as intraprostatically administered agents for patients with locally recurrent and high-risk local prostate cancer. Promising therapeutic responses have been reported in several studies with both oncolytic and suicide gene therapy strategies. The additional benefit of combining gene therapy with radiation therapy has also been realized; replicating adenoviruses inhibit DNA repair pathways, resulting in a synergistic sensitization to radiation. Other, nonreplicating suicide gene therapy strategies are also significantly enhanced with radiation. Combined radiation/gene therapy is currently being studied in phase I and II clinical trials and will likely be the first adenoviral gene therapy mechanism to become available to urologists in the clinic. Systemic gene therapy for metastatic disease is also a major goal of the field, and clinical trials are currently under way for hormone-resistant metastatic prostate cancer. Second- and third-generation "re-targeted" viral vectors, currently being developed in the laboratory, are likely to further improve these systemic trials.     

Are chromosomal instabilities induced by exposure of cultured normal human cells to low- or high-LET radiation?

Radiation-induced genomic instability has been proposed as a very early, if not an initiating, step in radiation carcinogenesis. Numerous studies have established the occurrence of radiation-induced chromosomal instability in various cells of both human and rodent origin. In many of these studies, however, the cells were not "normal" initially, and in many cases they involved tumor-derived cell lines. The phenomenon clearly would be of even greater interest if it were shown to occur generally in cells that are normal at the outset, rather than cells that may have been "selected" because of a pre-existing susceptibility to induced instability. As a test of the generality of the phenomenon, we studied low-passage normal diploid human fibroblasts (AG1521A) to determine whether they are susceptible to the induction of chromosomal instability in the progeny of surviving cells after exposure in G(0) to low- and high-LET radiation. Cytogenetic assays for instability were performed on both mixed populations of cells and clones of cells surviving exposure. We found no evidence for the induction of such instability as a result of radiation exposure, though we observed a senescence-related chromosomal instability in the progeny of both irradiated and unirradiated cell populations.     

ultraviolet radiation from F and K stars and implications for planetary habitability

Now that extrasolar planets have been found, it is timely to ask whether some of them might be suitable for life. Climatic constraints on planetary habitability indicate that a reasonably wide habitable zone exists around main sequence stars with spectral types in the early-F to mid-K range. However, it has not been demonstrated that planets orbiting such stars would be habitable when biologically-damaging energetic radiation is also considered. The large amounts of UV radiation emitted by early-type stars have been suggested to pose a problem for evolving life in their vicinity. But one might also argue that the real problem lies with late-type stars, which emit proportionally less radiation at the short wavelengths ( < 200 nm) required to split O2 and initiate ozone formation. We show here that neither of these concerns is necessarily fatal to the evolution of advanced life: Earth-like planets orbiting F and K stars may well receive less harmful UV radiation at their surfaces than does the Earth itself.     

Biological effect of 9 GeV protons and the radioprotective effect of ATP and AMP on epithelial cells of the mouse cornea

The cytogenetic and cytological effects induced in mouse cornea epithelium cells by 9 GeV protons and "standard" radiation have been studied. The RBE coefficients are different at different times of observation. ATP and AMP are shown to produce a protective effect on the frequency of formation of aberrant mitoses. DMF values for protons determined 24 and 72 h following irradiation are 1.8 +/- 0.2 and 1.7 +/- 0.2, respectively.     

<Emphasis Type="Italic">Rhizophagus irregularis</Emphasis> MUCL 41833 can colonize and improve P uptake of <Emphasis Type="Italic">Plantago lanceolata</Emphasis> after exposure to ionizing gamma radiation in root organ culture

Long-lived radionuclides such as ⁹⁰Sr and ¹³⁷Cs can be naturally or accidentally deposited in the upper soil layers where they emit β/γ radiation. Previous studies have shown that arbuscular mycorrhizal fungi (AMF) can accumulate and transfer radionuclides from soil to plant, but there have been no studies on the direct impact of ionizing radiation on AMF. In this study, root organ cultures of the AMF Rhizophagus irregularis MUCL 41833 were exposed to 15.37, 30.35, and 113.03 Gy gamma radiation from a ¹³⁷Cs source. Exposed spores were subsequently inoculated to Plantago lanceolata seedlings in pots, and root colonization and P uptake evaluated. P. lanceolata seedlings inoculated with non-irradiated AMF spores or with spores irradiated with up to 30.35 Gy gamma radiation had similar levels of root colonization. Spores irradiated with 113.03 Gy gamma radiation failed to colonize P. lanceolata roots. P content of plants inoculated with non-irradiated spores or of plants inoculated with spores irradiated with up to 30.35 Gy gamma radiation was higher than in non-mycorrhizal plants or plants inoculated with spores irradiated with 113.03 Gy gamma radiation. These results demonstrate that spores of R. irregularis MUCL 41833 are tolerant to chronic ionizing radiation at high doses.     

Risk calculations for hereditary effects of ionizing radiation in humans

Summary A prediction of the extent to which an additional dose of ionizing radiation increases the natural germ cell mutation rate, and how much such an increase will affect the health status of future human populations is part of the service that human geneticists are expected to offer to human society. However, more detailed scrutiny of the difficulties involved reveals an extremely complex set of problems. A large number of questions arises before such a prediction can be given with confidence; many such questions cannot be answered at our present state of knowledge. However, such predictions have recently been attempted. The 1988 report of the United Nations Scientific Committee for the Effects of Atomic Radiation and the fifth report of the Committee on Biological Effects of Ionizing Radiation of the US National Research Council have presented a discussion of the human genetics problems involved. Empirical data from studies on children of highly radiation-exposed parents, e.g. parents exposed to the atomic bombs of Hiroshima and Nagasaki, or parents belonging to populations living on soil with high background radiation, have been mentioned in this context. Whereas precise predictions are impossible as yet because of deficiencies in our knowledge of medical genetics at various levels, the bulk of the existing evidence points to only small effects of low or moderate radiation doses, effects that will probably be buried in the background noise of changing patterns of human morbidity and mortality.     

The interaction of soil micromycetes with "hot" particles in a model system]

A model system which permits observing for a long time and fixing interaction of fungi with a radiation source has been created on the basis of an isolated "hot" particle, deficient mineral medium (saccharose content 60 mg/l) and suspension of fungal conidia. Five species (six strains) of micromycetes isolated from radionuclide-contaminated soils and fifteen "hot" particles have been tested. It has been found out for the first time that Cladosporium cladosporioides and Penicillium roseo-purpureum are able actively overgrow "hot" particles whose radioactivity did not exceed 3.1-1.0(-7) Ci by gamma-spectrum and to destroy them 50-150 days later. Certain changes in morphology of fungi-destructors of "hot" particles are revealed. A problem on ecological significance of the found phenomenon is discussed.     

Bystander effects induced by direct and scattered radiation generated during penetration of medium inside a water phantom

Background

The biological effects of ionizing radiation have long been thought to results from direct targeting of the nucleus leading to DNA damage. Over the years, a number of non-targeted or epigenetic effects of radiation exposure have been reported where genetic damage occurs in cells that are not directly irradiated but respond to signals transmitted from irradiated cells, a phenomenon termed the “bystander effects”.

Aim

We compared the direct and bystander responses of human A 549, BEAS-2-B and NHDF cell lines exposed to both photon (6 MV) and electron (22 MeV) radiation inside a water phantom. The cultures were directly irradiated or exposed to scattered radiation 4 cm outside the field. In parallel, non-irradiated cells (termed bystander cells) were incubated in ICM (irradiation conditioned medium) collected from another pool of irradiated cells (termed donor cells).

Materials and methods

In directly irradiated cells as well as ICM-treated cells, the frequency of micronuclei and condensation of chromatin characteristic for the apoptotic process were estimated using the cytokinesis-block micronucleus test.

Results

In all tested cell lines, radiation induced apoptosis and formation of micronuclei. A549 and BEAS-2B cells cultured in ICM showed increased levels of micronuclei and apoptosis, whereas normal human fibroblasts (NHDF line) were resistant to bystander response. In A549 and BEAS-2B cells placed outside the radiation field and exposed to scattered radiation the formation of micronuclei and induction of apoptosis were similar to that after ICM-treatment.

Conclusion

Results suggest that the genetic damage in cells exposed to scattered radiation is caused by factors released by irradiated cells into the medium rather than by DNA damage induced directly by X rays. It seems that bystander effects may have important clinical implications for health risk after low level radiation exposure of cells lying outside the radiation field during clinical treatment.     

Ionizing radiation and genetic risks. X. The potential "disease phenotypes" of radiation-induced genetic damage in humans: perspectives from human molecular biology and radiation genetics.

Estimates of genetic risks of radiation exposure of humans are traditionally expressed as expected increases in the frequencies of genetic diseases (single-gene, chromosomal and multifactorial) over and above those of naturally-occurring ones in the population. An important assumption in expressing risks in this manner is that gonadal radiation exposures can cause an increase in the frequency of mutations and that this would result in an increase in the frequency of genetic diseases under study. However, despite compelling evidence for radiation-induced mutations in experimental systems, no increases in the frequencies of genetic diseases of concern or other adverse effects (i.e., those which are not formally classified as genetic diseases), have been found in human studies involving parents who have sustained radiation exposures. The known differences between spontaneous mutations that underlie naturally-occurring single-gene diseases and radiation-induced mutations studied in experimental systems now permit us to address and resolve these issues to some extent. The fact that spontaneous mutations (among which are point mutations and DNA deletions generally restricted to the gene) originate through a number of different mechanisms and that the latter are intimately related to the DNA organization of the genes, are now well-documented. Further, spontaneous mutations include those that cause diseases through loss of function as well as gain of function of genes. In contrast, most radiation-induced mutations studied in experimental systems (although identified through the phenotypes of the marker genes) are predominantly multigene deletions which cause loss of function; the recoverability of an induced deletion in a livebirth seems dependent on whether the gene and the genomic region in which it is located can tolerate heterozygosity for the deletion and yet be compatible with viability. In retrospect, the successful mutation test systems (such as the mouse specific locus test) used in radiation studies have involved genes which are non-essential for survival and are also located in genomic regions, likewise non-essential for survival. In contrast, most of the human genes at which induced mutations have been looked for, do not seem to have these attributes. The inference therefore is that the failure to find induced germline mutations in humans is not due to the resistance of human genes to induced mutations but due to the structural and functional constraints associated with their recoverability in livebirths. Since the risk of inducible genetic diseases in humans is estimated using rates of "recovered" mutations in mice, there is a need to introduce appropriate correction factors to bridge the gap between these rates and the rates at which mutations causing diseases are potentially recoverable in humans. Since the whole genome is the "target" for radiation-induced genetic damage, the failure to find increases in the frequencies of specific single-gene diseases of societal concern does not imply that there are no genetic risks of radiation exposures: the problem lies in delineating the phenotypes of recoverable genetic damage that are recognizable in livebirths. Data from studies of naturally-occurring microdeletion syndromes in humans and those from mouse radiation studies are instructive in this regard. They (i) support the view that growth retardation, mental retardation and multisystem developmental abnormalities are likely to be among the quantitatively more important adverse effects of radiation-induced genetic damage than mutations in a few selected genes and (ii) underscore the need to expand the focus in risk estimation from known genetic diseases (as has been the case thus far) to include these induced adverse developmental effects although most of these are not formally classified as "genetic diseases". (ABSTRACT TRUNCATED)     

Effects of ionizing radiations on human costal cartilage and exploration of the procedures to protect the tissue from radiation damage

Synoposis The chemical changes produced following⁶⁰Co--irradiation of human costal cartilage have been monitored using critical electrolyte concentration (CEC) measurements utilizing Alcian Blue-magnesium chloride and Toluidine Blue-sodium chloride systems. The decrease in the CEC with radiation can be related to the decrease in hexose and hexosamine contents of the cartilage. Such changes arise as a result of degradation of the glycosaminoglycan component of the tissue. The state of the collagen fibres after irradiation has been examined using the Van Giesson stain and by biochemical assay of the hydroxyproline moeity. Energy transfer methods have been utilized to protect the glycosaminoglycan component of the cartilage from radiation damage.     

Neutron microscopy. The low-damage imaging of specialized organic materials

It is shown that, insofar as radiation damage is concerned, transmission neutron microscopy using neutrons in the energy range approximately 0.0001-1.0 eV is extremely attractive for the imaging of specialized organic materials. By "specialized organic materials" is meant organic specimens composed entirely of specific isotopes that have been selected on the basis of their favorable properties with regard to radiation damage. In connection with such specimens, it is demonstrated that at a resolution of, for example, 100 A, neutrons will have an advantage over soft X-rays in terms of radiation damage, provided that the inherent (neutron) bright field image contrast turns out to be greater than 10(-5). Suggestions relating to (a) the comprehensive calculation of the radiation damage sustained by specialized organic specimens under slow neutron irradiation, (b) the construction of a theory of image formation in the neutron microscope, (c) the development of neutron lenses/focusing devices, and (d) the development of a brighter neutron source (essential for neutron microscopy) are outlined in some detail. The paper concludes with two appendices, which provide important background material.     

Four mutants of Micrococcus radiodurans defective in the ability to repair DNA damaged by mitomycin-C, two of which have wild-type resistance to ultraviolet radiation

Summary Four genes concerned with the resistance of wild-type Micrococcus radiodurans to the lethal action of mitomycin-C (MTC), mtcA, mtcB, uvsA and uvsB, have been identified by isolating mutants sensitive to MTC.Two strains of M. radiodurans, 302 and 262 carrying mutations in mtcA and mtcB respectively, are between forty and sixty times as sensitive as the wild-type to MTC, only slightly more sensitive than the wild-type to ionizing () radiation and have the same resistance as the wild-type to ultraviolet (u.v.) radiation. Strain 302 can be transformed at a high frequency to wild-type resistance to MTC with DNA from strain 262, and vice versa, indicating that mtcA and mtcB have different genetic locations.Two further strains of M. radiodurans, 303 and 263 having mutations in uvsA and uvsB respectively are only from four to eight times as sensitive as the wild-type to MTC, seven to thirteen times as sensitive to -radiation but between twenty to thirty-three times as sensitive to u.v. radiation. Strain 303 can be transformed with DNA from strain 263, or vice versa, to wild-type resistance to u.v. radiation, implying that uvsA and uvsB also have different genetic locations. M. radiodurans strain 301 which is mutant in both mtcA and uvsA, and strain 261 which is mutant in mtcB and uvsB are twenty to forty times as sensitive as the wild-type to both MTC and u.v. radiation and seven to ten times as sensitive to radiation. Neither mtcA and uvsA nor mtcB and uvsB are closely linked.None of the mutant strains is deficient in recombination, as measured by transformation. The repair of MTC-induced DNA damage in M. radiodurans must be different from that described for Escherichia coli.     

Is increased UV-B a threat to crop photosynthesis and productivity?

It has been suggested that increases in ground-level UV-B, as a result of stratospheric ozone depletion, may have major deleterious effects on crop photosynthesis and productivity. The direct consequences of such effects have been projected by some as a world-wide decrease in crop yields of 20–25%. Further losses, or unrealized gains, have also been suggested as a result of increased UV-B counteracting the beneficial effects of elevated atmospheric CO₂. Deleterious UV-B effects may be largely partitioned between damage to the plant genome and damage to the photosynthetic machinery. Direct damage to DNA is a common result of absorption of high energy UV-B photons. However, most plants possess repair mechanisms adequate to deal with the levels of damage expected from projected increases in ground-level UV-B. In addition, most plants have the ability to increase production of UV-absorbing compounds in their leaves as a result of exposure to UV-B, UV-A and visible radiation. These compounds contribute substantially to reducing UV-B damage in situ. It has also been shown that in some plants, under the proper conditions, almost every facet of the photosynthetic machinery can be damaged directly by very high UV-B exposures. However, electron transport, mediated by Photosystem II (PS II) appears to be the most sensitive part of the system. Various laboratories have reported damage to virtually all parts of the PS II complex from the Mn binding site to the plastoquinone acceptor sites on the opposite surface of the thylakoid membrane. However, a critical review of the literature with emphasis on exposure protocols and characterization of the radiation environment, revealed that most growth chamber and greenhouse experiments and very many field experiments have been conducted at unrealistic or indeterminate UV-B exposure levels, especially with regard to the spectral balance of their normal radiation environment. Thus, these experiments have led directly to large overestimates of the potential for damage to crop photosynthesis and yield within the context of 100 year projections for stratospheric ozone depletion. Indeed, given the massive UV-B exposures necessary to produce many of these effects, we suggest it is unlikely that they would occur in a natural setting and urge reconsideration of the purported impacts of projected increases of UV-B on crop productivity.Abbreviations C_i leaf internal CO₂ partial pressure - CPD cyclobutane pyrimidine dimer - CVY cultivar-year, one crop cultivar grown for one season - F_V/F_M variable chlorophyll fluorescence ratio - kJ m^–2 d^–1 daily radiation energy flux - PAR photosynthetically active radiation - PAS300 UV-B_BE weighted by the generalized plant action spectrum normalized to 300 nm - TOMS total ozone mapping spectrometer instrument mounted aboard the National Aeronautics and Space Administration's Nimbus-7 satellite - UV-A ultraviolet-A radiation (400 nm>320 nm) - UV-B ultraviolet-B radiation (320 nm280 nm) - UV-B_BE biologically effective UV-B (in this paper, irradiance weighted by the generalized plant action spectrum) The U.S. Government right to retain a non-exclusive, royalty free licence in and to any copyright is acknowledged.