Ekkehard Pomplun

Auger electron spectra--the basic data for understanding the Auger effect.

Understanding the strong radiotoxicity of DNA-incorporated Auger electron-emitting nuclides requires a detailed knowledge of the nuclides emission spectrum. A Monte Carlo computer code was previously developed to simulate Auger cascades and to provide electron spectra of 125I. To utilize experimental data for a direct validation of these simulations, the code has been adapted for cascades in xenon, which is adjacent to iodine in the table of elements. Only minor modifications of the code were necessary to obtain a very good agreement with the experimental findings. The role of shake-off electrons and the need for energy considerations during the cascades could be demonstrated. A previously published electron spectrum of 125I was recalculated and detailed results are presented here. Furthermore, to consider implications from a molecular binding of the Auger emitter, for the first time semi-empirical quantum mechanical calculations for an iodine-labelled thymine molecule were performed showing that even in the condensed phase a Coulomb explosion cannot be excluded a priori.

Is Coulomb explosion a damaging mechanism for 125IUdR

Purpose: To test the integrity of the thymine molecule that experiences an increasing number of charges due to the loss of Auger electrons emitted by the decay of incorporated 125I. Besides the radiation action of these electrons, Coulomb explosion is suspected to be an additional mechanism responsible for the strong radiotoxic effect of decaying DNA‐incorporated 125I. The two‐step decay process initiates a first Auger cascade within 10−16 to 10−14 s resulting in the release of about 7 electrons on average and a corresponding large positive charge on the 125Te daughter atom. Being part of iododeoxyuridine (125IUdR), the analogue of the DNA base thymine, the base is suddenly confronted with this charge. Experimentally, the situation was investigated with small molecules (CH3125I and C2H5125I) resulting in ion fragmentation in agreement with a Coulomb explosion model (Carlson and White, 1963, 1966). Materials and methods: Semi‐empirical quantum mechanical calculations on the Parametric Method 3 (PM3) level (Stewart, 1989a, 1989b) were performed and geometry optimisation was applied for the identification of stable molecule conformations. Subsequently, semi‐empirical molecular dynamics simulations allowed changes in the conformations to be studied as a function of time. Results: First results show that there is no stable molecular configuration with a total charge of ≥+5e. PM3 calculations will not converge for such a charge located at the 125I/125Te position. This finding is supported by total energy considerations, which begin to favour a system of isolated atoms versus molecular bound atoms when the molecular charge is greater than +4e. The distribution of the partial charges indicates that most of the charge will remain on the tellurium atom with slight increases of charge at the other molecular partners within 125IUdR. Moreover, the molecular dynamics simulations reveal a breaking of chemical bonds between those atoms with the strongest charge increase. Conclusions: Coulomb explosion must be taken into account as a possible damaging mechanism following the decay of DNA‐incorporated Auger electron emitters. Lobachevsky and Martin (2000) have identified the same mechanism to be responsible for part of strand breakage in oligo‐deoxynucleotides. To elucidate a possible link between both damage patterns the molecular mechanics simulations have to be extended to larger parts of the DNA molecule.

Monte Carlo-simulated Auger electron spectra for nuclides of radiobiological and medical interest - a validation with noble gas ionization data.

Abstract Purpose: To further validate Monte Carlo calculation codes simulating cascades of Auger electron transitions in radionuclides that decay by electron capture or internal conversion. In particular, the need for an appropriate kinetic energy determination of the Auger electrons emitted from multiple-ionized atoms as well as the consideration of shake-off electrons would be investigated implicitly. Methods: Charge distributions of noble gases after photoionization for different photon energies were calculated and compared with experimental data from the literature. In addition, new electron emission spectra were generated for 99mTc and 123I. Results: By including strict energy book-keeping and allowing shake-off electrons, the agreement between experimentally detected charge distributions and Monte Carlo simulations was very good. On this basis, the number of emitted electrons per decay was found to be between 1 and 17 with a mean of 4.0 for 99mTc and between 1 and 26 with a mean of 7.4 for 123I. Conclusions: Because of the good agreement with the experimental findings, the validation can be considered to be successful.

Modelling of Initial Events and Chemical Behaviour of Species Induced in DNA Units by Auger Electrons from 125I, 123I and Carbon

Auger electron spectra for 123I and 125I generated by Monte Carlo calculation and Auger electrons emitted from carbon after photoelectric effect on its K-shell as well as two DNA models (linear plasmid and nucleosome model) based on x-ray diffraction experiments have been used to simulate the behaviour of all species and radicals created during the physical and the chemical phase of the Augers transport. By introducing appropriate assumptions for the induction of strand breaks the number of these breaks can also be determined and correlated to experimentally found numbers of lethal events. Efficiency differences between the iodine nuclides themselves and in comparison with the rather monoenergetic Auger electrons from carbon are shown with regard to the direct and indirect effects on the two DNA models. The characteristic products in the physical, chemical and biochemical phase are compared with corresponding results from the literature for low-LET radiation.

On the biological efficiency of I-123 and I-125 decay on the molecular level.

Purpose: To evaluate DNA damage of Auger emitters by numerical modelling at the molecular level. Material and methods: Energy emission spectra of I-123 and I-125 were used as input data for a computer code that simulates the complete transport of electrons and photons from the physical stage up to the primary chemical stage at 10−7 s. The simulation was performed in a complex environment of liquid water, DNA structures and scavengers. Electron and photon interactions with the DNA molecules were carefully managed. Simulations were carried out with both I-123 and I-125 bound to a pBR322 plasmid or free in its vicinity. Results: The distributions of direct and indirect single strand breaks (SSB) and double strand breaks (DSB) as a function of the kinetic energy of the emitted Auger electrons show that damage is caused primarily by electrons with energies lower than 800 eV, while higher energy electrons are mainly involved in indirect effects. The yields per unit energy emitted strengthen this fact. When compared to experimental values, the calculated yields of linearization (LE) and relaxation (RE) events show good agreement as well as does the ratio LE/RE for each radionuclide and the ratio I-125/I-123 in the case of LE.

A Nucleosome Model for the Simulation of DNA Strand Break Experiments

Using a set of Monte Carlo simulation models, track structures of 125I Auger electrons generated in liquid water are superimposed on a nucleosome DNA model able to precisely localize energy deposition events on sub-molecular units of the DNA strands. After scoring direct hits taking place during the physical phase (at about 10(-15) s) the radiation chemistry of the whole system is simulated between 10(-12) and 10(-8) s, taking into account all reactions between water radio-chemical species, radicals, sub-molecular units of DNA (Ribose, Adenine, Thymine, Guanine, and Cytosine), and scavengers like Tris or Formate ions. The models possibility to distinguish between direct and indirect hits has been utilized to introduce different assumptions for strand break induction by both hit modes. The number of SSB and DSB as well as their local distribution will be given and compared with experimental and theoretical results from the literature.

Iodine-125-labeled DNA-Triplex-forming oligonucleotides reveal increased cyto- and genotoxic effectiveness compared to Phosphorus-32

Abstract Purpose: The efficacy of DNA-targeting radionuclide therapies might be strongly enhanced by employing short range particle-emitters. However, the gain of effectiveness is not yet well substantiated. We compared the Auger electron emitter I-125 to the ß−-emitter P-32 in terms of biological effectiveness per decay and radiation dose when located in the close proximity to DNA using DNA Triplex-forming oligonucleotides (TFO). The clonogenicity and the induction of DNA double-strand breaks (DSB) were investigated in SCL-II cells after exposure to P-32- or I-125-labeled TFO targeting the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene and after external homogeneous exposure to gamma-rays as reference radiation. Materials and methods: TFO were labeled with P-32 or I-125 using the primer extension method. Cell survival was analyzed by colony-forming assay and DNA damage was assessed by microscopic quantification of protein 53 binding protein 1 (53BP1) foci in SCL-II cells. Results: I-125-TFO induced a pronounced decrease of cell survival (D37 at ∼360 accumulated decays per cell, equivalent to 1.22 Gy cell nucleus dose) and a significant increase of 53BP1 foci with increasing decays. The P-32-labeled TFO induced neither a strong decrease of cell survival nor an increase of 53BP1 foci up to ∼4000 accumulated decays per cell, equivalent to ∼1 Gy cell nucleus dose. The RBE for I-125-TFO was in the range of 3–4 for both biological endpoints. Conclusions: I-125-TFO proved to be much more radiotoxic than P-32-TFO per decay and per unit dose although targeting the same sequence in the GAPDH gene. This might be well explained by the high number of low energy Auger electrons emitted by I-125 per decay, leading to a high ionization density in the immediate vicinity of the decay site, probably producing highly complex DNA lesions overcharging DNA repair mechanisms.

Chromosome aberrations induced by the Auger electron emitter 125I

DNA-associated Auger electron emitters (AEE) cause cellular damage leading to high-LET type cell survival curves indicating an enhanced relative biological effectiveness. Double strand breaks (DSBs) induced by Iodine-125-deoxyuridine ((125)I-UdR) decays are claimed to be very complex. To elucidate the assumed genotoxic potential of (125)I-UdR, chromatid aberrations were analysed in exposed human peripheral blood lymphocytes (PBL). PBL were stimulated with medium containing phytohaemagglutinin (PHA). After 24h, cultures were labelled with (125)I-UdR for 18h (activity concentration 1-45 kBq) during the S-phase. Following standard cytogenetic procedure, at least 100 metaphases were analysed microscopically for each activity concentration. Cell death was measured by apoptosis assay using flow cytometry. Radiation doses were determined by using point kernel calculations. After 18h labelling with (125)I-UdR the cell cycle distribution is severely disturbed. About 40% of PBL are fully labelled and 20% show a moderate labelling of (125)I-UdR, whereas 40% of cells remain un-labelled. The dose-response relationship fits to a polynomial curve in the low dose range, whereas a linear fit supplies a better estimation in the high dose range. Even the lowest dose of 0.2Gy leads to a 13-fold increase of aberrations compared to the controls. On average every fifth (125)I-decay produces a single chromatid aberration in PBL. Additionally, a dose-dependent increase of cell death is observed. (125)I-UdR has a very strong genotoxic capacity in human PBL, even at 0.2Gy. Efficiently labelled cells displaying a prolonged cell cycle compared to moderately labelled cells and cell death contribute substantially to the desynchronisation of the cell cycle. Our data, showing for the first time, that one (125)I-decay induces ∼ 0.2 chromatid aberrations, are in very good accordance to DSB data, stating that ∼0.26 DSB are induced per decay, indicating that it takes on average 250 decays to induce one chromosome aberration (CA). [Corrected]

Cellular response on Auger- and Beta-emitting nuclides: Human embryonic stem cells (hESC) vs. keratinocytes

Abstract Purpose: We studied the response of human embryonic stem cells (hESC) to the β-emitter 131I, which affects the entire cell and to the Auger electron emitter 125I-deoxyuridine (125I-dU), primarily affecting the deoxyribonuleic acid (DNA). The effects were also studied in keratinocytes as a prototype for somatic cells. Methods: HESC (H1) and human keratinocytes (HaCaT, human) were exposed to 125I-dU (5 × 10−5 – 5 MBq/ml) and 131I-iodide (5 × 10−5 – 12.5 MBq/ml) and apoptosis was measured by DNA-fragmentation. Cell morphology was studied by light microscopy and electron microscopy. Transcriptional profiling was done on the Agilent oligonucleotide microarray platform. Results: Auger-process induced no apoptosis but a strong transcriptional response in hESC. In contrast, HaCaT cells showed a pronounced induction of apoptosis but only a moderate transcriptional response. Transcriptional response of hESC was similar after 125I-dU and 131I treatments, whereas HaCaT cells expressed a much more pronounced response to 125I-dU than to 131I. A striking radiation-induced down-regulation of pluripotency genes was observed in hESC whereas in keratinocytes the enriched gene annotations were related primarily to apoptosis, cell division and proliferation. Conclusions: Human embryonic stem cells respond to ionizing radiation by 125I-dU and 131I in a different way compared to keratinocytes. Transcriptional response and gene expression appear to facilitate an escape from programmed cell death by striking a new path which probably leads to cell differentiation.

Charge build-up during decay of DNA-incorporated 123/125 I: Consequences for labeled molecular structures

Abstract Purpose: To further elucidate the mechanisms behind the strong biological effectiveness of DNA-incorporated Auger electron emitters 123I and 125I, which are mostly attributed to the shower of low-energy electrons released during the decay. A second, frequently mentioned cause can be seen in the charges accumulated during the Auger cascade on the decaying nuclide and its subsequent intra-molecular redistribution leading to a Coulomb explosion. Methods: To assess the size of the charge and the dimensions of DNA damage thus determined, the first Auger cascade was simulated by Monte Carlo methods. The consequences of intra-molecular charge transfer in terms of structural molecular alterations were estimated by density functional theory (DFT) calculations and folding with the results of the Monte Carlo studies. Results: Charge distributions of 123I and 125I were found to be very similar with values between + 1 and + 15 and a mean value of + 6.4. The molecules could tolerate charges up to + 5 (base), + 2 (nucleoside) and + 7 (nucleotide) without being destroyed. Conclusions: The strong molecular DNA damage after 123I and 125I decay depends very much on the size of the DNA molecule involved in the calculation. In general, not every decay can be expected to lead to a Coulomb explosion.