Richard C. Miller

The radiobiology of intravascular irradiation

PURPOSE There is increasing interest in the use of vascular irradiation, from an internally introduced radioactive source to control restenosis after balloon angioplasty. Both animal experiments and early clinical studies appear to show promising results in this regard. We consider various mechanistic interpretations of the experimental and clinical observations that doses of 12-20 Gy appear to be efficacious in preventing restenosis. We develop and investigate simple models, both experimental and theoretical, of the kinetics of radiation-induced smooth muscle cell (SMC) inactivation and regrowth, as a first step toward optimizing the design of clinical vascular irradiation. METHODS AND MATERIALS Using in vitro models of human SMCs, we investigate the relative radiosensitivity of SMCs compared with endothelial cells and measure the dose-dependent ability of SMCs to repopulate a denuded region in a confluent layer of cells. We then use quantitative information on the number, radiation sensitivity, and growth rate of the potential arterial target cells to model the time course of the SMC population after irradiation. RESULTS AND CONCLUSION Doses >20 Gy, which would be required to completely eliminate the SMC population which has the potential to cause restenosis, are too large to be practical because of the unacceptable risk of late complications. However, doses that can be practically given in vascular irradiation (<20 Gy) will certainly delay restenosis by 1-3 years, with larger doses producing longer delays. Whether such doses can avert restenosis permanently is unclear, as permanent prevention at realistic doses depends critically on the assumption that those SMCs which survive irradiation have a significantly limited capacity for proliferation. With regard to current animal model experiments, routine follow-up of <1 year, which is standard practice, is probably too short to address some of the key mechanistic question in intravascular radiation therapy.

The biological effectiveness of radon-progeny alpha particles. II: Oncogenic transformation as a function of linear energy transfer

Epidemiological studies have established an association between exposure to radon and carcinoma of the lung. However, based on data for either lung cancer in uranium miners exposed to radon or bronchial epithelial carcinomas in Japanese A-bomb survivors, it has not been possible to assign estimates of risk of lung cancer for the general population exposed to radon in their homes. Based on past success with the excellent quantitative properties of the C3H 10T1/2 in vitro oncogenic transformation assay system, the relative biological effectiveness (RBE) for radiation-induced transformation for charged particles of defined LET has been determined. As the LET of the radiation was increased, the rate of induction of oncogenic transformation increased and the RBEm approached 20. At higher LETs, RBE dropped precipitously. The rapid drop in effectiveness for alpha particles with LETs between 120 and 265 keV/microns implies a lower quality factor than the 20-25 currently considered appropriate when estimating lung cancer mortality.

The how and why of in vitro oncogenic transformation.

The techniques devised to study oncogenic transformation in vitro, involving established cell lines as well as fresh explants from hamster embryos, represent a powerful tool to study the mechanisms involved in radiation carcinogenesis. Large numbers of cells can be irradiated and transformation incidence scored over a wide range of doses down to as low as 0.01 Gy of X rays. When data with small confidence intervals are available for doses below 1 Gy, it becomes evident that the dose-response relationship has a complex shape and is not well represented by a linear relationship between transformation incidence and dose. Experiments with fractionated doses of X rays indicate that dividing the dose into two equal fractions separated by 5 hr results in a decrease of transformation incidence compared with a single exposure of the same total dose for doses above 1.5 Gy, but that at lower dose levels splitting the dose enhances transformation incidence. In a further series of experiments, it has been shown that t...

Neutron-energy-dependent oncogenic transformation of C3H 10T1/2 mouse cells

The relative biological effectiveness (RBE) of a range of neutron energies relative to 250-kVp X rays has been determined for oncogenic transformation and cell survival in the mouse C3H 10T 1/2 cell line. Monoenergetic neutrons at 0.23, 0.35, 0.45, 0.70, 0.96, 1.96, 5.90, and 13.7 MeV were generated at the Radiological Research Accelerator Facility of the Radiological Research Laboratories, Columbia University, and were used to irradiate asynchronous cells at low absorbed doses from 0.05 to 1.47 Gy. X irradiations covered the range 0.5 to 8 Gy. Over the more than 2-year period of this study, the 31 experiments provided comprehensive information, indicating minimal variability in control material, assuring the validity of comparisons over time. For both survival and transformation, a curvilinear dose response for X rays was contrasted with linear or nearly linear dose responses for the various neutron energies. RBE increased as dose decreased for both end points. Maximal RBE values for transformation ranged from 13 for cells exposed to 5.9-MeV neutrons to 35 for 0.35-MeV neutrons. This study clearly shows that over the range of neutron energies typically seen by nuclear power plant workers and individuals exposed to the atomic bombs in Japan, a wide range of RBE values needs to be considered when evaluating the neutron component of the effective dose. These results are in concordance with the recent proposals in ICRU 40 both to change upward and to vary the quality factor for neutron irradiations.

Oncogenic Transformation by Fractionated Doses of Neutrons

Oncogenic transformation was assayed after C3H 10T1/2 cells were irradiated with monoenergetic neutrons; cells were exposed to 0.23-, 0.35-, 0.45-, 5.9-, and 13.7-MeV neutrons given singly or in five equal fractions over 8 h. At the biologically effective neutron energy of 0.45 MeV, enhancement of transformation was evident with some small fractionated doses (below 1 Gy). When transformation was examined as a function of neutron energy at 0.5 Gy, enhancement was seen for cells exposed to three of the five energies (0.35, 0.45, and 5.9 MeV). Enhancement was greatest for cells irradiated with 5.9-MeV neutrons. Of the neutron energies examined, 5.9-MeV neutrons had the lowest dose-averaged lineal energy and linear energy transfer. This suggests that enhancement of transformation by fractionated low doses of neutrons may be radiation-quality dependent.

Long-Term Efficacy of Intracoronary Irradiation in Inhibiting In-Stent Restenosis

Background — Intracoronary irradiation is a promising modality for inhibition of in-stent restenosis. Results of randomized clinical trials at 6 months after gamma ray irradiation are highly encouraging. The first results at 3 years after irradiation, while still showing benefit, have shown significant late loss. The probable mechanism of the radiation is to inactivate (prevent from dividing) most cells that otherwise could proliferate to produce neointimal formation. We measured the proportion of cells that survive with their clonogenic potential intact after the doses and dose rates used in the randomized trials, and we then modeled the subsequent repopulation of the surviving cells that might cause late restenosis. Methods and Results — Human aortic smooth muscle cells were irradiated with gamma rays, including the doses and dose rates used in current trials, and clonogenic surviving fractions were measured. The subsequent repopulation of the surviving cells was modeled with the assumption that the repopulation kinetics were similar to those in unirradiated cells. The radiation is expected to delay the time to restenosis by factors of ≈6 to 8, depending on the dose, shifting the delay from a median of 6 months (for no irradiation) to median values from 36 months (for a nominal 13 Gy) to 43 months (for a nominal 15 Gy). Conclusions — These results and predictions are quantitatively consistent with clinical results and suggest that clonogenic inactivation (prevention of cellular division) is the dominant mechanism of radiation action in the delay of restenosis. Intracoronary radiotherapy is a very promising modality for significantly delaying, although probably not preventing, in-stent restenosis.

Interaction of hyperthermia and chemotherapy agents; cell lethality and oncogenic potential

Hyperthermia was combined with bleomycin, melphalan and cis-platinum in order to examine cell lethality and oncogenic transformation in C3H10T1/2 cells from the adjuvant use of heat with chemotherapy agents. When cells were exposed concurrently to 42.5 degrees C and each of the three chemotherapy agents, heat enhanced both the cytotoxic and oncogenic potential of the drugs. Hyperthermia-enhanced ratios were largest for bleomycin-treated cells. Examination of transformation incidences expressed as a function of surviving fraction, i.e. the cytotoxicity of treatment and therefore drug-heat efficacy, showed that for a given level of cell killing the combination of heat and cis-platinum resulted in fewer transformants per surviving cell than for cis-platinum alone. Hyperthermia appears to reduce the oncogenic potential of low concentrations of melphalan but has no effect on bleomycin-induced oncogenic transformation.

Soft X-ray Dosimetry and RBE for Survival of Chinese Hamster V79 Cells

Dosimetry and biological effects of 40 and 50 keV low-energy X-rays generated by a SOFTEX Model CMBW-2 apparatus were studied. Doses were measured using a thin-window parallel-plate ionization chamber; beam quality was assessed using aluminum absorbers; exposure rates per unit current were determined according to the X-ray tube current and exposure times; and thermoluminescent (BeO chip) dosimeters were used to ascertain dose distributions in the irradiation field. The average correction factors for nonuniformity were calculated from the measured dose distributions. The means for ascertaining accurate exposures and doses using these methods are discussed. The dose-survival relationship of Chinese hamster V79 cells were assessed by irradiating them with 40 and 50 kV soft X-rays, 180 kV X-rays, and 60 Co gamma rays. Soft X-rays with three distinct effective energies were tested by changing the tube voltage kV and aluminium filter thicknesses; namely (1) 40 kV without filter, (2) 40 kV with a 0.2 mm thick aluminium filter and (3) 50 kV with a 0.7 mm thick aluminium filter. The effective energies obtained according to attenuation measurements using aluminium for these soft X-rays were 8.1, 11.7 and 18.5 kV, respectively. In this study the relative biological effectiveness (RBE) at 10 per cent survival compared with 60Co gamma rays ranged from 1.5 to 1.6. The RBE of 180 kV X-rays relative to 60Co gamma rays was 1.29. This study provided experimental data for the RBE of V79 cells in the intermediate energy range between hard and ultrasoft X-rays, data for which were previously reported by Goodhead and co-workers (1977, 1979, 1981).

The Inverse Dose-Rate Effect for Oncogenic Transformation by Charged Particles Is Dependent on Linear Energy Transfer

Mouse C3H 10T1/2 cells were exposed to single or fractionated doses of charged particles of defined linear energy transfer (LET) from 25 to 200 keV/microns. Dose fractionation with prolonged time intervals enhanced the yield of transformed foci compared with a single acute dose for a range of LET values between 40 and 120 keV/microns. Radiations of lower or higher LET did not show the enhancement that is commonly referred to as the inverse dose-rate effect. The fractionation scheme that was used consisted of three dose fractions; the maximum enhancement of transformation occurred with an interval of 150 min between dose fractions. This inverse dose-rate effect, demonstrated for cycling cells in log phase, was not seen for cells in plateau phase.

Mechanistic considerations on the dose-rate/LET dependence of oncogenic transformation by ionizing radiations

When exposure to densely ionizing radiation is protracted, the resulting biological effect is sometimes, but not always, enhanced for transformational end points, relative to acute exposure. A pattern has emerged as to the dependence of this effect on dose, dose rate, and radiation quality. Previous calculations indicated that the dose and dose-rate trends can be predicted by a model in which there is a period within the cell cycle of very high sensitivity to oncogenesis. Recent experiments indicate that the inverse dose-rate effect is significant over a very limited range of LETs--from about 30 to 130 keV/microns. We discuss such LET effects in the context of cell cycle-dependent models, and suggest that the effects are understandable on the basis of such models. In essence, the inverse dose-rate effect disappears at high LET because of a reduction in the number of cells being hit, and disappears at LETs below about 30 keV/microns because most of the dose is deposited at low specific energies, insufficient to produce the saturation effect which is central to the phenomenon. At even lower LETs, damage repair yields the familiar sparing associated with protraction of X- or gamma-ray doses.