Apisarnthanarax, S. and Chao, K. S. C. Current Imaging Paradigms in Radiation Oncology. Radiat. Res. 163, 1–25 (2005).

The technological revolution in imaging during recent decades has transformed the way image-guided radiation therapy is performed. Anatomical imaging (plain radiography, computed tomography, magnetic resonance imaging) greatly improved the accuracy of delineating target structures and has formed the foundation of 3D-based radiation treatment. However, the treatment planning paradigm in radiation oncology is beginning to shift toward a more biological and molecular approach as advances in biochemistry, molecular biology, and technology have made functional imaging (positron emission tomography, nuclear magnetic resonance spectroscopy, optical imaging) of physiological processes in tumors more feasible and practical. This review provides an overview of the role of current imaging strategies in radiation oncology, with a focus on functional imaging modalities, as it relates to staging and molecular profiling (cellular proliferation, apoptosis, angiogenesis, hypoxia, receptor status) of tumors, defining radiation target volumes, and assessing therapeutic response. In addition, obstacles such as imaging-pathological validation, optimal timing of post-therapy scans, spatial and temporal evolution of tumors, and lack of clinical outcome studies are discussed that must be overcome before a new era of functional imaging-guided therapy becomes a clinical reality.

Anderson, R. M., Tsepenko, V. V., Gasteva, G. N., Molokanov, A. A., Sevan'kaev, A. V. and Goodhead, D. T. mFISH Analysis Reveals Complexity of Chromosome Aberrations in Individuals Occupationally Exposed to Internal Plutonium: A Pilot Study to Assess the Relevance of Complex Aberrations as Biomarkers of Exposure to High-LET α Particles. Radiat. Res. 163, 26–35 (2005).

We recently demonstrated that a significant proportion of apparently stable insertions induced after exposure to a mean of one α particle/cell, detected using three-color FISH, were part of larger unstable complexes when visualized by 24-color FISH. Interestingly, regardless of the long-term persistence capability of the cell, the complexity of each α-particle-induced complex appeared to be specific to the nuclear traversal of a single α particle. To assess whether aberrations of a similar complexity are observed in vivo and also to examine the usefulness of detecting such aberrations as a biomarker of chronic exposure to α particles, we have carried out a limited pilot study of Russian workers with large body burdens of α-particle-emitting plutonium. We found unstable cells containing non-transmissible complex aberrations in all of the plutonium-exposed subjects analyzed by mFISH. In addition, all of the complexes seen were consistent with those previously observed in vitro. Non-transmissible complex aberrations were more common than transmissible-type complexes, consistent with ongoing/chronic exposure, and insertions were dominant features of both types of complex. Accordingly, this preliminary study supports the proposal that aberration complexity and non-transmissibility are the major cytogenetic features of α-particle exposure that could potentially be exploited as a specific indicator of chronic exposures to high-LET α particles.

Hill, M. A., Ford, J. R., Clapham, P., Marsden, S. J., Stevens, D. L., Townsend, K. M. S. and Goodhead, D. T. Bound PCNA in Nuclei of Primary Rat Tracheal Epithelial Cells after Exposure to Very Low Doses of Plutonium-238 α Particles. Radiat. Res. 163, 36–44 (2005).

Bystander effects from ionizing radiation have been detailed for a number of cell systems and a number of end points. We wished to use a cell culture/ex vivo rat model of respiratory tissue to determine whether a bystander effect detected in culture could also be shown in a tissue. Examination by immunofluorescence techniques of tracheal cell cultures after exposure to very low doses of α particles revealed a large proportion of cells with proliferating cell nuclear antigen (PCNA) bound in their nuclei. PCNA was selected as an end point because it is involved in both DNA repair and the changes in cell cycle that are typical of many reported bystander effects. Maximum response can be detected in up to 28% of the cells in sub-confluent cultures with a dose of only 2 mGy. At this dose less than 2% of the cell nuclei have experienced a particle traversal and less than 6% of the cells have experienced an α-particle traversal through either their nucleus or some part of their cytoplasm. The hypothesis that this bystander response in nontargeted cells is mediated through secreted factor(s) is presented, and supporting evidence was found using partial irradiation and co-culture experiments. Examination of the effect with excised pieces of trachea demonstrated a response similar to that seen in culture.

Lim, H. B., Cook, G. G., Barker, A. T. and Coulton, L. A. Effect of 900 MHz Electromagnetic Fields on Nonthermal Induction of Heat-Shock Proteins in Human Leukocytes. Radiat. Res. 163, 45–52 (2005).

Despite many studies, the evidence as to whether radiofrequency fields are detrimental to health remains controversial, and the debate continues. Cells respond to some abnormal physiological conditions by producing cytoprotective heat-shock (or stress) proteins. The aim of this study was to determine whether exposure to mobile phone-type radiation causes a nonthermal stress response in human leukocytes. Human peripheral blood was sham-exposed or exposed to 900 MHz fields (continuous-wave or GSM-modulated signal) at three average specific absorption rates (0.4, 2.0 and 3.6 W/kg) for different durations (20 min, 1 h and 4 h) in a calibrated TEM cell placed in an incubator to give well-controlled atmospheric conditions at 37°C and 95% air/5% CO₂. Positive (heat-stressed at 42°C) and negative (kept at 37°C) control groups were incubated simultaneously in the same incubator. Heat caused an increase in the number of cells expressing stress proteins (HSP70, HSP27), measured using flow cytometry, and this increase was dependent on time. However, no statistically significant difference was detected in the number of cells expressing stress proteins after RF-field exposure. These results suggest that mobile phone-type radiation is not a stressor of normal human lymphocytes and monocytes, in contrast to mild heating.

Mirzayans, R., Scott, A., Cameron, M. and Murray, D. Induction of Accelerated Senescence by γ Radiation in Human Solid Tumor-Derived Cell Lines Expressing Wild-Type TP53. Radiat. Res. 163, 53–62 (2005).

Recent studies have demonstrated that p21^WAF1 (now known as CDKN1A)-dependent and -independent accelerated senescence responses are a major determinant of the sensitivity of cancer cells to chemotherapeutic agents. The objective of the present study was to determine whether human solid tumor-derived cell lines that express wild-type TP53 can exhibit levels of CDKN1A induction after exposure to ionizing radiation that are sufficient to activate the accelerated senescence program. Exposure to ⁶⁰Co γ radiation (≤8 Gy) triggered accelerated senescence in all five TP53 wild-type tumor cell lines examined, albeit to differing degrees. Three of the TP53 wild-type tumor cell lines, HCT116, A172 and SKNSH, activated the TP53 signaling pathway similarly to normal human fibroblasts, as judged by the nuclear accumulation of TP53, magnitude and duration of induction of CDKN1A mRNA and CDKN1A protein, and propensity to undergo accelerated senescence after radiation exposure. In the clonogenic survival assay, the degree of radiosensitivity of these three tumor cell lines was also in the range displayed by normal human fibroblasts. On the other hand, two other TP53 wild-type tumor cell lines, A498 and A375, did not maintain high levels of CDKN1A mRNA and CDKN1A protein at late times postirradiation and exhibited only low levels of accelerated senescence after radiation exposure. Studies with a CDKN1A knockout cell line (HCT116CDKN1A^−/−) confirmed that the radiation-triggered accelerated senescence is dependent on CDKN1A function. We conclude that (1) clinically achievable doses of ionizing radiation can trigger CDKN1A-dependent accelerated senescence in some human tumor cell lines that express wild-type TP53; and (2) as previously documented for normal human fibroblasts, some TP53 wild-type tumor cell lines (e.g. HCT116, A172 and SKNSH) may lose their clonogenic potential in response to radiation-inflicted injury primarily through undergoing accelerated senescence.

Lu, F. and Wong, C. S. A Clonogenic Survival Assay of Neural Stem Cells in Rat Spinal Cord after Exposure to Ionizing Radiation. Radiat. Res. 163, 63–71 (2005).

Neural stem cells play an important role in neurogenesis of the adult central nervous system (CNS). Inhibition of neurogenesis has been suggested to be an underlying mechanism of radiation-induced CNS damage. Here we developed an in vivo/ in vitro clonogenic assay to characterize the survival of neural stem cells after exposure to ionizing radiation. Cells were isolated from the rat cervical spinal cord and plated as single cell suspensions in defined medium containing epidermal growth factor and basic fibroblast growth factor. The survival of the proliferating cells was determined by their ability to form neurosphere colonies. The number and size of neurospheres were analyzed quantitatively at day 10, 12, 14 and 16 after plating. Plating cells from 5, 10 and 15 mm of the cervical spinal cord resulted in a linear increase in the number of neurospheres from day 10–16. Compared to the nonirradiated spinal cord, there was a significant decrease in the number and size of neurosphere colonies cultured from a 10-mm length of the rat spinal cord after a single dose of 5 Gy. When dissociated neurospheres derived from a spinal cord that had been irradiated with 5 Gy were allowed to differentiate, the percentages of neurons, oligodendrocytes and astrocytes as determined by immunocytohistochemistry were not altered compared to those from the nonirradiated spinal cord. Secondary neurospheres could be obtained from cells dissociated from primary neurospheres that had been cultured from the irradiated spinal cord. In conclusion, exposure to ionizing radiation reduces the clonogenic survival of neural stem cells cultured from the rat spinal cord. However, neural stem cells retain their pluripotent and self-renewing properties after irradiation. A neurosphere-based assay may provide a quantitative measure of the clonogenic survival of neural stem cells in the adult CNS after irradiation.

Morris, G. M., Coderre, J. A., Micca, P. L., Nawrocky, M. M., Hopewell, J. W. and Miura, M. Porphyrin-Mediated Boron Neutron Capture Therapy: A Preclinical Evaluation of the Response of the Oral Mucosa. Radiat. Res. 163, 72– 78 (2005).

Preclinical studies are in progress to determine the potential of boron neutron capture therapy (BNCT) for the treatment of carcinomas of the head and neck. Recently, it has been demonstrated that various boronated porphyrins can target a variety of tumor types. Of the porphyrins evaluated so far, copper tetracarboranylphenyl porphyrin (CuTCPH) is potentially a strong candidate for clinical use. In the present investigation, the response of the oral mucosa to CuTCPH-mediated boron neutron capture (BNC) irradiation was assessed using the ventral surface of the tongue of adult male Fischer 344 rats, a standard rodent model. CuTCPH was administered by intravenous infusion, at a dose of 200 mg/kg body weight, over a 48-h period. Three days after the end of the administration of CuTCPH, biodistribution studies indicated very low levels of boron (<2 μg/g) in the blood. Levels of boron in tongue tissue were 39.0 ± 3.8 μg/g at this time. This was the time selected for irradiation with single doses of thermal neutrons from the Brookhaven Medical Research Reactor. The estimated level of boron-10 in the oral mucosa was used in the calculation of the physical radiation doses from the ¹⁰B(n,α)⁷Li reaction. This differs from the approach using the present generation of clinical boron carriers, where boron levels in blood at the time of irradiation are used for this calculation. Dose–response curves for the incidence of mucosal ulceration were fitted using probit analysis, and the doses required to produce a 50% incidence of the effect (ED₅₀ ± SE) were calculated. Analysis of the dose–effect data for CuTCPH-mediated BNC irradiation, compared with those for X rays and thermal neutrons alone, gave a compound biological effectiveness (CBE) factor of ∼0.04. This very low CBE factor would suggest that there was relatively low accumulation of boron in the key target epithelial stem cells of the oral mucosa. As a consequence, with low levels of boron (<2 μg/g) in the blood, the response of the oral mucosa to CuTCPH-mediated BNCT will be governed primarily by the radiation effects of the thermal neutron beam and not from the boron neutron capture reaction [¹⁰B(n,α)⁷Li].

Lomax, M. E., Salje, H., Cunniffe, S. and O'Neill, P. 8-OxoA Inhibits the Incision of an AP Site by the DNA Glycosylases Fpg, Nth and the AP Endonuclease HAP1. Radiat. Res. 163, 79–84 (2005).

Ionizing radiation induces clustered DNA damage sites, whereby two or more individual DNA lesions are formed within one or two helical turns of DNA by a single radiation track. A subset of DNA clustered damage sites exist in which the lesions are located in tandem on the same DNA strand. Recent studies have established that two closely opposed lesions impair the repair machinery of the cell, but few studies have investigated the processing of tandem lesions. In this study, synthetic double-stranded oligonucleotides were synthesized to contain 8-oxoA and an AP site in tandem, separated by up to four bases in either a 5′ or 3′ orientation. The influence 8-oxoA has on the incision of the AP site by the E. coli glycosylases Fpg and Nth protein and the human AP endonuclease HAP1 was assessed. 8-OxoA has little or no effect on the efficiency of incision of the AP site by Nth protein; however, the efficiency of incision of the AP site by Fpg protein is reduced in the presence of 8-oxoA even up to a four-base separation in both the 5′ and 3′ orientations. 8-OxoA influences the efficiency of HAP1 incision of the AP site only when it is 3′ to the AP site and separated by up to two bases. This study demonstrates that the initial stages of base excision repair can be impaired by the presence of a second base lesion in proximity to an AP site on the same DNA strand. This impairment could have biological consequences, such as mutation induction, if the AP site is present at replication.

Roginskaya, M., Bernhard, W. A., Marion, R. T. and Razskazovskiy, Y. The Release of 5-Methylene-2-Furanone from Irradiated DNA Catalyzed by Cationic Polyamines and Divalent Metal Cations. Radiat. Res. 163, 85–89 (2005).

Release of 5-methylene-2-furanone (5-MF), a characteristic marker of DNA deoxyribose oxidative damage at the C1′ position, was observed in significant quantities from X-irradiated DNA. This observation, which held for DNA irradiated either in aqueous solution or as a film, requires postirradiation treatment at 90°C in the presence of polyamines and divalent metal cations at biological pH. The 5-MF product was quantified by using reverse-phase HPLC. The radiation chemical yield of 5-MF comprised more than 30% of the yield of total unaltered base release. Polylysine, spermine and Be(II) showed the strongest catalytic effect on 5-MF release, while Zn(II), Cu(II), Ni(II), putrescine and Mg(II) were substantially less efficient. We have hypothesized that the 5-MF release from irradiated DNA occurs through catalytic decomposition of the 2′-deoxyribonolactone (dL) precursor through two consecutive β- and δ-phosphate elimination reactions. A stepwise character of the process was indicated by the S-shaped time course of 5-MF accumulation. If dL proves to be the precursor to 5-MF formation, it would then follow that dL is a very important lesion generated in DNA by ionizing radiation.

Tsoulou, E., Kalfas, C. A. and Sideris, E. G. Conformational Properties of DNA after Exposure to Gamma Rays and Neutrons. Radiat. Res. 163, 90–97 (2005).

DNA aqueous solutions were irradiated with 0–40 Gy of ⁶⁰Co γ rays and 0–1.5 Gy of (Pu-Be) neutrons. Thermal transition spectrophotometry (TTS) was used to trace the changes in the DNA conformation at the above doses. Previous results using the perturbed angular correlation (PAC) method were used to complement to the current analysis. The TTS and PAC methods are two different approaches to the study of the effects of radiation on DNA. Both showed that neutrons are more effective than γ rays in inducing DNA damage. The TTS method showed that neutrons are 11 ± 5 times more efficient than γ rays, while the PAC method had shown this value to be 34 ± 4. From the current study we deduced that the radiation damage to DNA is not a spontaneous effect but rather is an ensemble of damaging events that occur asynchronously. Any single method selected for the study of such damages can concentrate on only a part of the damage, leading to over- or underestimation of the relative effectiveness of the neutrons.

Emfietzoglou, D. and Nikjoo, H. The Effect of Model Approximations on Single-Collision Distributions of Low-Energy Electrons in Liquid Water. Radiat. Res. 163, 98–111 (2005).

The development of cross sections for the inelastic interaction of low-energy electrons with condensed tissue-like media is best accomplished within the framework of the dielectric theory. In this work we investigate the degree to which various model approximations, used in the above methodology, influence electron single-collision distributions. These distributions are of major importance to Monte Carlo track structure codes, namely, the energy-loss spectrum, the inelastic inverse mean free path, and the ionization efficiency. In particular, we make quantitative assessment of the influence of (1) the optical data set, (2) the dispersion algorithm, and (3) the perturbation and exchange Born corrections. It is shown that, although the shape and position of the energy-loss spectrum remains almost fixed, its peak height may vary by up to a factor of 1.5. Discrepancies in the calculated inelastic inverse mean free path are largely within 20–30% above 100 eV; they increase drastically, though, at lower energies. Exchange and perturbation Born corrections increase gradually below 1 keV leading to a ∼30 to 40% reduction of the inverse mean free path at 100 eV. The perturbation effect contributes more than the exchange effect to this reduction. Similar to the dispersion situation, the effect of Born corrections at lower energies is also unclear since the models examined disagree strongly below 100 eV. In comparison, the vapor data are higher than the liquid calculations by 20 to 50% as the energy decreases from 1 to 0.1 keV, respectively. The excitation contribution is the main cause of this difference, since the ionization efficiency in the liquid levels off at ∼90%, whereas the plateau value for the vapor is ∼70%. It is concluded that electron inelastic distributions for liquid water, although in some respects distinctively different from the vapor phase, have associated uncertainties that are comparable in magnitude to the phase differences. The situation below 100 eV is uncertain.

Shimada, A., Eguchi, H., Yoshinaga, S. and Shima, A. Dose-Rate Effect on Transgenerational Mutation Frequencies in Spermatogonial Stem Cells of the Medaka Fish. Radiat. Res. 163, 112–114 (2005).

The estimation of transgenerational genetic risk of radiation exposure to non-human species is crucial for the protection of ecosystems. Here we determined the frequency of specific-locus mutations at the five pigmentation loci in medaka spermatogonial stem cells after γ irradiation at 0.03 cGy/min and 95 cGy/min. At each total dose, the mutation frequency was significantly lower in the 0.03-cGy/min group than in the 95-cGy/min group, suggesting a dose-rate effect. The ratio of the induced mutation frequency at 0.03 cGy/min to that at 95 cGy/min was ∼0.42 from 0 to 1.9 Gy and ∼0.33 from 1.9 to 4.75 cGy. In the mouse, this ratio is estimated to be 0.33 (Russell and Kelly, Proc. Natl. Acad. Sci. USA 79, 542–544, 1982). It is thus possible that the magnitude of the dose-rate effect on transgenerational mutation frequencies is comparable between mouse and medaka spermatogonia, suggesting similar dose-rate effects among vertebrates.

Pellmar, T. C., Rockwell, S. and the Radiological/Nuclear Threat Countermeasures Working Group. Priority List of Research Areas for Radiological Nuclear Threat Countermeasures. Radiat. Res. 163, 115–123 (2005).

To help the nation prepare for the possibility of a terrorist attack using radiological and nuclear devices, the Office of Science and Technology Policy and the Homeland Security Council established an interagency working group. The working group deliberated on the research needs for radiological/ nuclear threat countermeasures and identified and prioritized 18 areas for further attention. The highest priorities were given to research on (1) radioprotectors for use prior to exposure; (2) therapeutic agents for postexposure treatment; (3) antimicrobial therapy for infections associated with radiation exposure; (4) cytokines and growth factors; (5) mechanisms of radiation injury at the molecular, cellular, tissue and organism levels; and (6) automation of biodosimetric assays. High priority was given to (1) developing biomarkers for biodosimetry; (2) enhancing training in the radiation sciences; (3) exploring the consequences of combined injury; (4) establishing a repository of information regarding investigational countermeasures; and (5) following the health of an exposed population to better prepare for subsequent events. The research areas that the committee felt required the attention of the radiation research community are described in this report in an effort to inform this community about the needs of the nation and to encourage researchers to address these critical issues.