Steven Kurtzman

Mechanism of radiosensitization by halogenated pyrimidines: effect of BrdU on radiation induction of DNA and chromosome damage and its correlation with cell killing

The effect of BrdU incorporation on cell radiosensitivity as well as on the induction of DNA double-strand breaks (DSB) and chromosome damage by radiation was studied in CHO cells. Induction of DNA DSB was measured by the nonunwinding filter elution technique and damage at the chromosome level was visualized and scored in G1 cells using the technique of premature chromosome condensation. The results indicated an increase in the radiosensitivity of cells grown in the presence of BrdU. Although sensitization was observed both in cells irradiated in the exponential phase and in cells irradiated in the plateau phase of growth, the degree of sensitization was greater in exponentially growing cells for the same degree of thymidine replacement by BrdU in the DNA. It is hypothesized that this indicates the possible importance of chromatin structure at the time of irradiation and/or the importance of chromatin conformation changes after irradiation in the expression of radiation-induced potentially lethal damage in cells containing BrdU. Incorporation of BrdU affected both the slope and the width of the shoulder of the survival curve and increased the induction of DNA and chromosome damage per unit absorbed dose. The increase observed in the slope of the survival curve was quantitatively similar to the increase observed in damage induction at the DNA and the chromosome level, suggesting a cause-effect relationship between these phenomena. Reduction in the width of the shoulder did not correlate with the increase in the induction of DNA and chromosome damage, suggesting that different phenomena, probably related to enhanced fixation of radiation-induced potentially lethal damage in cells containing BrdU, underlie its modulation.

Mechanism of Radiosensitization by Halogenated Pyrimidines: Effect of BrdU on Cell Killing and Interphase Chromosome Breakage in Radiation-Sensitive Cells

The effect of BrdU incorporation on cell radiosensitivity as well as on the induction of chromosome damage by radiation was studied in plateau-phase xrs-5 cells using the premature chromosome condensation (PCC) method. It is well known that xrs-5 cells are sensitive to ionizing radiation and defective in the repair of radiation-induced DNA double-strand breaks, chromosome damage, and potentially lethal damage (PLD). Compared to repair-proficient CHO 10B cells, a reduction was observed in the overall BrdU-mediated radiosensitization in plateau-phase xrs-5 cells for the same degree of thymidine replacement. This finding is interpreted with a model for BrdU-induced radiosensitization advanced previously, in which two distinct components act to produce the overall radiosensitization observed. One component involves processes associated with the increase in initial damage (DNA and chromosome) production per unit absorbed dose and causes an increase in the slope of the survival curve, while the second component involves enhanced fixation of radiation-induced damage (PLD) and causes a reduction in the width of the shoulder of the survival curve. It is suggested that in plateau-phase xrs-5 cells, the deficiency in the repair of radiation-induced damage compromises BrdU-mediated radiosensitization by leaving active only the radiosensitization component that is associated with an increase in damage induction. Enhancement of cell killing by BrdU in plateau-phase xrs-5 cells resulted in a decrease in D0, the relative value of which was similar to the relative increase in the production of chromosome damage as measured by the PCC method. The relative values for the change in D0 and the production of chromosome aberrations were similar in plateau-phase CHO 10B and xrs-5 cells, suggesting that the physicochemical and/or biochemical processes associated with this phenomenon are the same in the two cell lines. Radiosensitization of a magnitude similar to that observed in exponentially growing CHO 10B cells was induced by BrdU in exponentially growing xrs-5 cells. This effect is attributed to a partial expression of the repair gene (transiently during S phase in all cells, or throughout the cycle in a fraction of cells) that permits some repair of radiation-induced damage and which is compromised by BrdU.

Keynote address: Application of non-hypoxic cell sensitizers in radiobiology and radiotherapy: rationale and future prospects

The effects of commonly used non-hypoxic cell radiosensitizers are briefly reviewed. Emphasis is placed on the effects and the mechanism of action of halogenated pyrimidines, since recent clinical trials indicated the potential importance of these compounds in the treatment of certain types of human tumors. Evidence is presented suggesting that halogenated pyrimidines sensitize cells to radiation by increasing induction of DNA and chromosome damage per cell per unit absorbed dose, as well as by increasing the susceptibility to fixation of radiation induced PLD. The former mode of action correlates with an increase in survival curve slope, whereas the latter probably causes the reduction observed in shoulder width. The effects of repair inhibitors such as the nucleoside analogs are briefly reviewed and their possible clinical importance discussed. Results are presented indicating that combined treatment with halogenated pyrimidines and nucleoside analogs may enhance the radiosensitizing effect of the former and the specificity on tumor cells of the latter. Finally, the effects of other radiation sensitizers such as 3-aminobenzamide and diamide are briefly summarized.

Mechanism of radiosensitization by halogenated pyrimidines : bromodeoxyuridine and β-arabinofuranosyladenine affect similar subsets of radiation-induced potentially lethal lesions in plateau-phase Chinese hamster ovary cells

Chinese hamster ovary (CHO) cells were grown to plateau phase in the presence of various amounts of bromodeoxyuridine (BrdU) and treated after irradiation with beta-arabinofuranosyladenine (ara-A), an inhibitor of DNA and potentially lethal damage (PLD) repair, in order to investigate the importance of repair reactions in general and of PLD repair, in particular, on the mechanism of radiosensitization by halogenated pyrimidines. The degree of BrdU-mediated radiosensitization observed in ara-A-treated cells was compared to that of cells incubated after irradiation in the absence of ara-A. A substantial reduction in BrdU-mediated radiosensitization was observed in cells treated with ara-A at concentrations that, when given alone, produced maximum potentiation in cell killing (500-1500 microM). The residual BrdU-mediated radiosensitization observed at high levels of thymidine replacement could be explained by a BrdU-mediated increase in DNA and chromosome damage induction per gray. These findings are similar to those reported previously for a repair-deficient mutant of CHO cells, the xrs-5 cell line, and consistent with the hypothesis that BrdU-mediated radiosensitization has two distinct components, one that derives from an increase in damage induction per gray, and a second one that derives from an effect of BrdU on the repair of radiation-induced damage. It is proposed that the reduction in BrdU-mediated radiosensitization observed in ara-A-treated cells is the result of ara-A-mediated expression of radiation damage, the repair of which would have been otherwise modulated by BrdU. Since ara-A is known to act by fixing a form of radiation-induced PLD (alpha-PLD), we further propose that BrdU acts by fixing alpha-PLD. A synergistic effect in the potentiation of cell killing was observed between ara-A and BrdU when ara-A was given at concentrations below 100 microM. This result suggests that a benefit may be expected in the clinic from the combined application of halogenated pyrimidines with repair inhibitors, if administered at a carefully screened range of concentrations.

Hydrogel Spacer Application Technique, Patient Tolerance and Impact on Prostate Intensity Modulated Radiation Therapy: Results from a Prospective, Multicenter, Pivotal Randomized Controlled Trial

Introduction We evaluate the safety, tolerability and impact on therapy of an absorbable hydrogel perirectal spacer (SpaceOAR® system) designed to reduce the rectal radiation dose during prostate cancer radiotherapy. Methods A multicenter, pivotal, randomized controlled trial was conducted in 222 men with stage T1 or T2 prostate cancer treated to 79.2 Gy with image guided intensity modulated radiation therapy in 44 fractions. Patients were randomized 2:1 to receive fiducial markers and perirectal spacer injection (spacer group) or fiducial markers alone (control group). Spacer placement, tolerability, perirectal space creation, impact on rectal dose and impact on quality of life were assessed. Results Most spacer procedures were conducted with the patient under general or local anesthesia. Procedures were rated easy or very easy in 98.7% of cases with a 99.3% success rate. Mild transient rectal events were noted in 10% of patients in the spacer group (eg pain, discomfort). Mean perirectal space was 12.6 mm after implant and 10.9 mm at 12.4 weeks with absorption at 12 months. A 25% or greater reduction in rectal V70 dose was produced in 97.3% of patients in the spacer group. The spacer group experienced a significant reduction in late rectal toxicity severity (p=0.044) as well as lower rates of decrease in bowel quality of life at 6, 12 and 15 months compared to the control group. There were no unanticipated adverse spacer effects or spacer related adverse events. Conclusions Hydrogel spacer application was straightforward and repeatable, resulting in consistent perirectal space creation and rectal dose reduction. Spacer application has the potential to improve prostate radiotherapy outcomes and enable advanced radiotherapy protocols.