Radiotherapy with 6-megavolt photons for early glottic carcinoma: potential impact of extension to the posterior vocal cord.

PURPOSE: Radiation therapy has a high success rate in the treatment of early glottic carcinoma. Excellent outcomes have been reported from centers using cobalt-60 or relatively low-energy (< or = 4 MV) radiation therapy to achieve these results. Whether similar outcomes can be achieved with a 6 MV linear accelerator has been less rigorously evaluated. This study assesses the efficacy of 6 MV radiation therapy for early stage glottic cancer and identifies prognostic factors for local control and overall survival in this common disease. MATERIALS AND METHODS: One hundred twenty-eight consecutive cases of Tis, T1, and T2 squamous cell carcinomas of the glottis from 1982 to 1996 were retrospectively analyzed with regard to local control and survival. All patients were treated with definitive radiation therapy with a 6-MV linear accelerator. Potential prognostic factors for local control and survival were evaluated with univariate and multivariate models. Median follow-up of locally controlled patients was 65 months. RESULTS: The overall 3-year actuarial local control rates for T1 and T2 carcinomas were 86% and 68%, respectively. Patients with lesions involving the posterior third of the vocal cord had significantly worse 3-year local control (76% vs. 86%, P =.038). Radiation therapy technique and overall treatment time did not significantly affect local control. For patients with Tis and T1 lesions, factors associated with significantly worse local control included cordectomy-ineligible disease (P =.024), dose less than 6,600 cGy (P =.024), and lesions limited to the posterior third of the vocal cord (P =.004). Three-year local control was 76%, with doses less than 6,600 cGy and 90% with higher doses. High rates of second primary malignancies were observed and represented the major cause of death. Five-year overall survival was 84%. CONCLUSIONS: The use of 6-MV photons for treatment of early glottic cancer seems to achieve local control similar to that reported with lower-energy photons. However, patients with posterior third involvement had a poorer local control rate with standard radiation therapy, thereby suggesting that alternative approaches be considered.

DNA as an important target in radiation-induced apoptosis of MYC and MYC plus RAS transfected rat embryo fibroblasts.

PURPOSE: This study uses a radiation chemistry approach to determine if DNA is an important target for radiation-induced apoptosis of myc (MR4) and myc plus ras (3.7) transfected rat embryo fibroblast cell lines. MATERIALS AND METHODS: The radiation protection efficiency of four thiols was compared with net molecular charge ranging from -1 to +2: mercaptopropionic acid (Z= -1), mercaptoethanol (Z=0), cysteamine (Z= +1), N(2-mercaptoethyl)-1,3-diaminopropane (Z= +2). Protection factors were determined for these thiols against radiation-induced apoptosis (Apoalert assay), mitotic cell death (clonogenic assay) and double-strand break (dsb) induction (pulse field gel electrophoresis) in MR4 and 3.7 cells. Theoretical protection factors for these thiols against dsb induction were also calculated from second-order chemical repair constants for single-strand breaks (ssb) and the concentration of added thiols in MR4 and 3.7 cell lines. RESULTS: The charge-dependent increases observed for measured protection factors against radiation-induced apoptosis did not differ significantly between the two cell lines, nor did they differ significantly from the corresponding increases observed for radiation-induced mitotic cell killing and for induction of dsb. The calculated protection factor for dsb also showed a thiol charge-dependent increase similar to the measured protection factors for all of the other parameters studied. CONCLUSIONS: These results are consistent with the hypothesis that DNA is an important target for radiation-induced apoptosis.

Simulation of the cellular oxygen effect with an SV40 DNA model system using DNA strand breaks as an end point.

Most in vitro studies of the modification of radiation damage to DNA by oxygen and reduced thiol compounds have excluded non-thiol .OH scavengers. However, the non-thiol .OH scavengers are known to be very effective radiation protectors and are present in cells in near molar concentrations. To better relate data obtained from an in vitro system to data at the intracellular level, we have carried out simultaneous measurements of single and double-strand breaks (SSBs and DSBs) in a model system using closed-circular supercoiled SV40 DNA (25 micrograms/ml) in aqueous solution. Observations were made in the presence and absence of oxygen and over wide concentration ranges encompassing intracellular conditions of both glutathione (GSH), the dominant soluble intracellular reduced thiol compound, and glycerol, a widely used poly-alcohol non-thiol .OH scavenger. The dose-response curves for both SSBs and DSBs in either air or nitrogen are predominantly linear when 75 or 750 mM glycerol is combined with 0-20 mM GSH. Glutathione together with glycerol shows a concentration-dependent preferential protection in nitrogen compared to air against radiation-induced SSBs and DSBs, qualitatively similar to our prior observations at comparable concentrations of GSH alone (Ayene et al., Radiat. Res. 144, 1-8, 1995). With GSH alone, we observed peak oxygen enhancement ratios (OERs) of 6.5 and 8.0 for SSBs and DSBs, respectively, at 5 mM GSH. In contrast, we observed previously that glycerol alone showed a preferential protection against radiation-induced SSBs and DSBs in air compared to nitrogen (Ayene et al., Radiat. Res. 142, 133-143, 1995). In the presence of 750 mM glycerol, approximately equivalent to the total effective concentration of non-thiol .OH scavengers in the cell, we now observe that the OER for radiation-induced DSBs increases from 1.2 at 0.5 mM GSH to 3.3 at 5.0 mM. This roughly parallels the dependence of the OER on [GSH] reported for cell survival. A possible mechanism for the lower OER at higher .OH scavenging efficiency is discussed.

Modification of radiation-induced strand breaks by glutathione: comparison of single- and double-strand breaks in SV40 DNA.

A number of investigations have suggested that the widely observed oxygen enhancement of radiation-induced cell killing or intracellular DNA damage requires the presence of glutathione (GSH) or other thiols. We have adapted an in vitro model system to investigate the effects of GSH on radiation-induced DNA double-strand breaks (DSBs), lesions felt to be critical to cell death. Superhelical SV40 DNA, 25 micrograms/ml, was irradiated in air or nitrogen in the presence of 0-20 mM GSH and single-strand breaks (SSBs) and DSBs were measured using neutral gel electrophoresis/ethidium bromide fluorescence. Control experiments demonstrated that a substantial concentration of free SH was still present after irradiation. Dose-response curves for SSBs and DSBs in air or nitrogen were predominantly linear at all GSH concentrations tested from 0-20 mM, except for 20 mM GSH in nitrogen, indicating that both SSB and DSB formation are predominantly by one-hit mechanisms under these conditions. Dose-response curves for both SSBs and DSBs in nitrogen at 20 mM GSH closely tracked the corresponding linear curves in air for doses up to about 200 Gy, then reached a plateau at higher doses. Induction efficiencies in 20 mM GSH, calculated from these initial slopes for both SSBs and DSBs in nitrogen, were unexpectedly higher than the corresponding efficiencies in 5 mM GSH, suggesting additional damage, rather than the expected additional protection. The possible mechanism for a damaging effect from GSH is discussed. Oxygen enhancement ratios (OERs) were calculated from the slopes of dose-response curves. The OERs for SSBs did not differ substantially from those for DSBs at the same [GSH], contrary to the observations of Prise et al. (Radiat. Res. 134, 102-106, 1993). The OERs for SSBs and DSBs peaked at 6.5 and 8, respectively, at 5 mM GSH. These similarities suggest that the much lower OERs (2.5-3.0) generally reported for radiation killing of cells, which also typically contain about 5 mM GSH, cannot be accounted for by differences in OER between lethal lesions, represented by DSBs, and nonlethal lesions, represented by SSBs. In view of the present results, another possible explanation, that intracellular compounds other than reduced thiols are important in the chemical modification of the response of DNA to radiation, seems to be much more likely.

Role of scavenger-derived radicals in the induction of double-strand and single-strand breaks in irradiated DNA.

Studies of intact cells have generally shown a substantial enhancement by oxygen of radiation-induced cell killing or DNA breakage, with an oxygen enhancement ratio (OER) of 2.5-3. Analogous studies of radiation damage to purified DNA in aqueous solution, performed at much lower .OH scavenging efficiencies, have usually shown little or no damage enhancement by oxygen (OER approximately 1) unless reduced thiol compounds were also present during irradiation. Milligan and Ward (Radiat. Res. 137, 295-299, 1994) recently observed an excess of DNA single-strand breaks (SSBs) in nitrogen compared to air (OER < 1) for SV40 DNA irradiated in the presence of high concentrations of DMSO, and they presented evidence that this excess resulted from secondary DMSO radicals. We have here investigated the role of secondary radicals from glycerol, another non-thiol .OH scavenger, in the induction of DNA double-strand breaks (DSBs), lesions which are critical to cell death. Superhelical SV40 DNA, 25 micrograms/ml, was irradiated in air or nitrogen in the presence of 0-2500 mM glycerol. Resultant DSBs and SSBs were measured simultaneously by computer analysis of digital video images of ethidium bromide-stained gels. The OERs for DSBs and SSBs decreased from 1.3 at the lowest glycerol concentration to 0.65 and 0.45, respectively, at the highest concentration, consistent with an important role for anoxic secondary radicals in the presence of high glycerol concentrations. The dose-response curves for SSBs due to glycerol radicals are predominantly "one-hit" over the entire glycerol concentration range where they were observable (> or = 75 mM). The dose-response curves for DSBs due to glycerol radicals are predominantly one-hit at high glycerol concentrations (> or = 750 mM). Analysis of the data suggests that the dominant mechanism for the formation of glycerol radical-induced one-hit DSBs at lower glycerol concentrations (< or = 750 mM) involves a single glycerol radical resulting in two nearby SSBs on opposite DNA strands, analogous to the mechanism proposed by Siddiqi and Bothe (Radiat. Res. 112, 449-463, 1987) for the induction of a DSB by a single OH radical. Our data for the highest concentration (2500 mM) imply that it is now important to take into account a second mechanism.

Recursive partitioning analysis of prognostic factors in three Radiation Therapy Oncology Group malignant glioma trials.

BACKGROUND: Despite notable technical advances in therapy for malignant gliomas during the past decade, improved patient survival has not been clearly documented, suggesting that pretreatment prognostic factors influence outcome more than minor modifications in therapy. Age, performance status, and tumor histopathology have been identified as the pretreatment variables most predictive of survival outcome. However, an analysis of the association of survival with both pretreatment characteristics and treatment-related variables is necessary to assure reliable evaluation of new approaches for treatment of malignant glioma. PURPOSE: This study of malignant glioma patients used a non-parametric statistical technique to examine the associations of both pretreatment patient and tumor characteristics and treatment-related variables with survival duration. This technique was used to identify subgroups with survival rates sufficiently different to create improvements in the design and stratification of clinical trials. METHODS: We used a recursive partitioning technique to analyze survival in 1578 patients entered in three Radiation Therapy Oncology Group malignant glioma trials from 1974 to 1989 that used several radiation therapy (RT) regimens with and without chemotherapy or a radiation sensitizer. This approach creates a regression tree according to prognostic variables that classifies patients into homogeneous subsets by survival. Twenty-six pretreatment characteristics and six treatment-related variables were analyzed. RESULTS: The years). Patients younger than 50 years old were categorized by histology (astrocytomas with anaplastic or atypical foci [AAF] versus glioblastoma multiforme [GBM]) and subsequently by normal or abnormal mental status for AAF patients and by performance status for those with GBM. For patients aged 50 years or older, performance status was the most important variable, with normal or abnormal mental status creating the only significant split in the poorer performance status group. Treatment-related variables produced a subgroup showing significant differences only for better performance status GBM patients over age 50 (by extent of surgery and RT dose). Median survival times were 4.7-58.6 months for the 12 subgroups resulting from this analysis, which ranged in size from 32 to 256 patients. CONCLUSIONS: This approach permits examination of the interaction between prognostic variables not possible with other forms of multivariate analysis. IMPLICATIONS: The recursive partitioning technique can be employed to refine the stratification and design of malignant glioma trials.

Radiation chemical mechanisms of single- and double-strand break formation in irradiated SV40 DNA.

Using an electrophoresis assay system developed in our laboratory, we have simultaneously measured single- and double-strand DNA breaks (SSBs and DSBs) induced by gamma radiation in small SV40 viral DNA molecules, under conditions of greatly varying radical scavenger concentration and DNA configuration. In our experiments with aqueous solutions of SV40 DNA, we observe that SSB induction is linear with dose (one-hit response), over the entire hydroxyl scavenger efficiency range examined, from approximately 0 to 5 x 10(9) s-1, while DSB induction shifts from having a major quadratic component (two-hit response) at very low scavenger efficiencies to nearly pure linear for efficiencies greater than 10(7) s-1. The mean ratio of SSBs to one-hit DSBs remains relatively constant with increasing scavenger efficiency, decreasing from about 100:1 to 40:1 as the scavenger efficiency increases from 2 x 10(5) s-1 to 5 x 10(9) s-1, and the absolute induction efficiencies for breaks decrease by three orders of magnitude. This decrease takes place primarily at scavenger efficiencies above 1 x 10(8) s-1. Irradiation of intranuclear SV40 minichromosomes induces SSBs and DSBs at nearly the same efficiencies as does irradiation of free DNA at the highest scavenger concentrations examined, and at only about twice the efficiencies observed at -75 degrees C, where direct effects are believed to predominate. Our observations that the linear-quadratic mix of the dose-response curve for DSBs depends critically on scavenger efficiency may help to clarify the considerable confusion in the literature on the shape of such curves. Our observations of a relatively constant ratio between one-hit SSBs and DSBs at low and moderate scavenger efficiencies are in agreement with the recent hypothesis of Siddiqi and Bothe (Radiat. Res. 112, 449-463 (1987)) that, contrary to widely and long-held beliefs, the formation by indirect effects of a one-hit DSB in DNA occurs under these conditions predominantly by a mechanism involving a single OH radical, with a presumed radical transfer between complementary DNA strands. In contrast, our results for strongly protective conditions are not consistent with this hypothesis, but are consistent with the predictions of Ward's hypothesis (Radiat. Res. 86, 185-195, (1981)) that one-hit DSBs from indirect effects are produced predominantly by local clusters of OH radicals from single energy deposition events (locally multiply damaged sites) rather than by single OH radicals.(ABSTRACT TRUNCATED AT 400 WORDS)

Measurement of radiation-induced DNA damage using gel electrophoresis or neutral filter elution shows an increased frequency of DNA strand breaks after exposure to pH 9.6.

The filter elution technique using nondenaturing conditions is widely used to assay DNA double-strand break (DSB) induction and repair. It has been reported that in the measurement of strand breaks higher rates of elution and of initial rejoining are obtained at pH 9.6 compared to pH 7.2. In the present experiments neutral elution at pH 7.2 and 9.6 were compared in the assay of damage to DNA induced by X rays, 125I decay, and restriction enzyme digestion, in an effort to explain this discrepancy and to determine whether the higher rate of elution observed at pH 9.6 corresponds to a greater number of DSBs. X-ray damage to cellular DNA resulted in significantly different elution profiles at the two pH values. In contrast the elution profiles of the DSB induced by intragenomic 125I decays or restriction endonuclease were independent of the pH of the elution buffer. When gamma-irradiated SV40 DNA was exposed to pH 7.2 or 9.6 elution buffer prior to analysis by gel electrophoresis, a significantly greater number of DNA DSBs were detected in the DNA exposed to pH 9.6. We conclude that X and gamma radiation produce lesions (pH 9.6-labile lesions), in proportion to dose, that have the potential of becoming measurable DSBs following incubation under the mildly alkaline condition of pH 9.6. The data suggest that these lesions may result from single-hit events.

Further studies of the induction and intracellular repair of DNA strand breaks using intranuclear SV40 as a test system.

We have previously published the techniques and preliminary results of an SV40 viral probe assay for gamma-radiation-induced single- and double-strand DNA breaks and their intracellular repair in higher cells (Radiat. Res. 101, 356-372, 1985). Those experiments with SV40 infected CV-1 monkey kidney cells suggested that this assay technique demonstrates slow but extensive intracellular repair of single-strand breaks (SSB), and possible early repair of double-strand breaks (DSB), followed by later induction of DSB. Following up on these early observations, many additional infection-incubation experiments have now been performed with both human and simian cells. Analysis of data from these experiments involving up to 6 h of postinfection intranuclear incubation shows the same distribution of strand break damage in incubated and unincubated samples. This implies that under these experimental conditions there is neither intracellular repair nor further production of SSB or DSB in intranuclear viral DNA. We have evidence which suggests that this lack of repair or degradation occurs because the bulk of intranuclear SV40 DNA is relatively inaccessible to host cell enzymes.

A sensitive SV40 viral probe assay for DNA strand breaks and their biological repair in higher cells: techniques and preliminary results.

An alternative experimental approach to the investigation of intracellular repair of DNA strand breaks by mammalian higher cells has been developed using simian virus 40 (SV40), which has no known intrinsic DNA repair capacity and possesses a minichromosome structure, as an intracellular probe. In this approach unirradiated simian or human cells are infected with irradiated virus and incubated for varying periods. Nuclei are isolated, and viral DNA is extracted and assayed for residual damage. This assay involves separation of the viral DNA by agarose gel electrophoresis into three sharply demarcated bands corresponding to DNA molecules containing a double-strand break (DSB), single-strand breaks (SSB), or no breaks. Quantitative data are obtained by a combination of DNA hybridization with 32P-labeled SV40 DNA, autoradiography, and densitometry. Various experiments have been carried out to investigate the feasibility of this approach and to establish the optimal experimental conditions for its use. These experiments indicate that there is rapid and efficient cellular uptake of SV40. independent of prior radiation dose to the virus, and that this multistep experimental procedure gives excellent recovery and quantitation of the three DNA forms when compared with more direct methods of measurement. Radiation dose-response experiments with purified extracellular SV40 virus, using this approach, are quite reproducible and give results closely comparable to those obtained with techniques in current use. Initial time-course incubation experiments with SV40 infected CV-1 monkey kidney cells indicate that this approach can demonstrate slow but extensive intracellular repair of SSB; and limited presumptive early repair of DSB, followed by later and more extensive induction of DSB.

The somnolence syndrome in leukemic children following reduced daily dose fractions of cranial radiation.

A group of children with acute lymphocytic leukemia was studied to investigate if a reduction in daily dose fraction of cranial radiation would reduce the incidence of somnolence syndrome. Thirty-one evaluable patients received 100 rad X 18 cranial radiation therapy. Sixty-six similar evaluable patients were given 180 rad X 10. Both groups received the same chemotherapy including intrathecal methotrexate. Clinically detectable somnolence appeared in 58% of ech group without significant differences in the overall frequency or severity of somnolence (p greater than 0.5). This study failed to substantiate a radiation dose fraction size dependence for somnolence syndrome in children with acute lymphocytic leukemia.

The somnolence syndrome in leukemic children following reduced daily dose fractions of cranial radiation

A group of children with acute lymphocytic leukemia was studied to investigate if a reduction in daily dose fraction of cranial radiation would reduce the incidence of somnolence syndrome. Thirty-one evaluable patients received 100 rad X 18 cranial radiation therapy. Sixty-six similar evaluable patients were given 180 rad X 10. Both groups received the same chemotherapy including intrathecal methotrexate. Clinically detectable somnolence appeared in 58% of ech group without significant differences in the overall frequency or severity of somnolence (p greater than 0.5). This study failed to substantiate a radiation dose fraction size dependence for somnolence syndrome in children with acute lymphocytic leukemia.

Induction and repair of double- and single-strand DNA breaks in bacteriophage lambda superinfecting Escherichia coli.

Induction and repair of double- and single-strand DNA breaks have been measured after decays of 125I and 3H incorporated into the DNA and after external irradiation with 4 MeV electrons. For the decay experiments, cells of wild type Escherichia coli K-12 were superinfected with bacteriophage lambda DNA labelled with 5'-(125I)iodo-2'-deoxyuridine or with (methyl-3H)thymidine and frozen in liquid nitrogen. Aliquots were thawed at intervals and lysed at neutral pH, and the phage DNA was assayed for double- and single-strand breakage by neutral sucrose gradient centrifugation. The gradients used allowed measurements of both kinds of breaks in the same gradient. Decays of 125I induced 0.39 single-strand breaks per double-strand break. No repair of either break type could be detected. Each 3H disintegration caused 0.20 single-strand breaks and very few double-strand breaks. The single-strand breaks were rapidly rejoined after the cells were thawed. For irradiation with 4 MeV electrons, cells of wild type E. coli K-12 were superinfected with phage lambda and suspended in growth medium. Irradiation induced 42 single-strand breaks per double-strand break. The rates of break induction were 6.75 x 10(-14) (double-strand breaks) and 2.82 x 10(-12) (single-strand breaks) per rad and per dalton. The single-strand breaks were rapidly repaired upon incubation whereas the double-strand breaks seemed to remain unrepaired. It is concluded that double-strand breaks in superinfecting bacteriophage lambda DNA are repaired to a very small extent, if at all.

DNA breakage, repair, and lethality accompanying 125I decay in microorganisms.

Effects of 125I decay in DNA were investigated by measurements of strand breaks and lethal efficiencies of the decays. In bacteriophages T1 and T4, local decay effects were compared with effects of the emitted electrons by induction of both single (ssb) and double strand breaks (dsb) in the intact phage head and in extended free state DNA. Most dsbs were found to result from local decay effects whereas most real ssbs are caused by the electrons. A simple one-to-one relationship seems to exist in the phages between the decays of 125I, numbers of dsbs and lethal effects. In E. coli rec+ and recA repair of dsbs was studied in addition to lethal decay efficiencies. In rec+ more than 70% of the dsbs were repaired within 1 h at 37 degrees C. No repair was observed in recA. The probability of lethality per 125I decay per completed genome was found to be 0.37 for rec+ and 0.93 for recA cells. The number of lethal events per unrepaired dsb was found to be practically equal to unity. Unrepaired dsbs thus seem to be the primary mechanism of lethality caused by 125I decay, and all unrepaired dsbs seem to be lethal.

Lethality and double-strand scissions from 14C decay in the DNA of micro-organisms.

14C-2-thymidine was incorporated into the DNA of E. coli B/r and of coliphage T4. The labelled organisms were stored for several years at -196 degrees C. Both were periodically assayed for loss of viability, and the coliphage also for the appearance of double-strand breaks (DSBs) in DNA. E. coli B/r exhibited a survival curve with a substantial initial shoulder, extrapolation number 5-2 +/- 2-3, and a final exponential portion corresponding to a lethal efficiency per 14C decay per 2-5 X 10(9) daltons of DNA, of 0-009 +/- 0-002. For coliphage T4, our best estimate for the lethal efficiency per 14C decay is 0-03 +/- 0-04, and that for the DNA breakage efficiency is -0-002 +/- 0-004. The large standard errors result from the very small number of 14C decays occurring in each phage. These results suggest that 14C decay in the DNA of micro-organisms does not cause DSBs but does cause potentially lethal damage to the thymine bases in which decay occurs, and that wild-type E. coli can repair a large number of such DNA lesions.

DNA breakage, repair and lethality after 125I decay in rec+ and recA strains of Escherichia coli.

Iodine-125 decays by electron capture and is known to cause extensive molecular fragmentation via the Augur effect. 125I was incorporated into the DNA of exponentially-growing E. coli K12 AB2487, a recA mutant, and E. coli K12 AB2497, the corresponding rec+ strain, as 5-iododeoxyuridine (IUdR), an analogue of thymidine. Radioactive bacteria were stored at - 196 degrees C, and samples were periodically assayed for loss of viability and for the induction of double-strand breaks (DSBs) in DNA. Each 125I decay in the DNA of either strain induces one DSB, i.e. alpha(DSB) = 1.0. For the recA strain, alpha(lethal) = 0.9 and for the rec+ strain, 0.4. Assays for biological repair of DSBs, involving incubation of thawed samples in growth-medium at 37 degrees C before the extraction of DNA, demonstrate significant repair of 125I-induced DSBs by rec+ cells but none by recA cells. For small numbers of decays, there is approximately a 1:1 correlation, for either strain, between lethal decays and post-incubation residual DSBs. Comparison with data for larger numbers of decays indicates that a typical rec+ cell can repair no more than three to four DSBs per completed genome (2.5 x 10(9) daltons).

Further studies of DNA damage and lethality from the decay of iodine-125 in bacteriophages.

The DNA of coliphages T4 and T1 was labelled with 125I-iododeoxyuridine. 125I decay is known to cause severe molecular damage via vacancy cascades (the Auger effect). We have compared the induction of both single- and double-strand breaks (SSBs and DSBs) in 125I-labelled T4 DNA stored at - 196 degrees C during decay, either as intact phage or as free DNA. These comparative experiments indicate that, in addition to one DSB which apparently results directly from the Auger effect, each decay in an intact phage also give rise to an additional 0-05 DSBs, as well as 1-6 SSBs, as a result of ionizing radiation absorbed in the same phage particle where the decay occurs. An examination of T4-killing by 125I decay reveals a two-phase survival curve, whose initial slope corresponds to a lethal efficency per 125I decay of 0-95 +/- 0-05, which is considerably higher than values previously determined. The results for phage T4, and of a more limited comparison of 125I suicide and DNA damage in phage T1, support the hypothesis that the vacancy cascades which accompany each 125I decay in DNA result in a double-strand break at the decay site and that each such break is a lethal event.