DNA core-ionization and cell inactivation
Arnaud Boissière a, b, c, Christophe Champion d, Alain Touati b, Marie-Anne Hervé du Penhoat b,
Laure Sabatier c, Aloke Chatterjee a and Annie Chetioui b ,1.
a
Life Sciences Division, Lawrence Berkeley National Laboratory, University of California,
Berkeley, California 94720, USA.
b
Institut de Minéralogie et Physique des Milieux Condensés, Université Paris 6, CNRS UMR
7590, Campus Boucicaut, 140 rue de Lourmel, 75015 Paris Cedex, France.
c
Laboratoire de Radiobiologie et Oncologie, CEA/DSV/DRR, B.P. N°6, 92265 Fontenay aux
Roses, France.
d
Laboratoire de Physique Moléculaire et des Collisions, Université de Metz, 1 boulevard Arago,
Technopôle, 57070 Metz, France.
Number of Figures: 2.
Number of Tables: 7.
Corresponding author’s full mailing address: Institut de Minéralogie et Physique des Milieux
Condensés, Université Paris 6, CNRS UMR 7590, Campus Boucicaut, 140 rue de Lourmel,
75015 Paris Cedex, France.
Phone: +33(0)1 44 27 73 08
Fax: +33(0)1 43 54 28 78
E-mail: [email protected]
1
Address for correspondence: Institut de Minéralogie et Physique des Milieux Condensés, Université Paris 6, CNRS
UMR 7590, Campus Boucicaut, 140 rue de Lourmel, 75015 Paris Cedex, France; email: [email protected]
Boissière et al. DNA core ionization and cell inactivation
Boissière, A., Touati, A., Champion, C., Hervé du Penhoat, M. A., Sabatier, L., Chatterjee, A
and Chetioui, A. DNA core-ionization and cell inactivation. Radiat. Res.
The possibility for inner shell ionizations of DNA atoms, called core ionizations, to be
the critical events for cell inactivation by ionizing radiations such as 100-keV electrons and
gamma rays, is studied. The number of core ionizations in DNA atoms per Gray of the two types
of radiations is calculated from various Monte-Carlo track simulations and the probability that a
core ionization lead to cell inactivation is deduced from experimental values of the RBEs of ultra
soft X-rays. The contribution to V79 cell inactivation solely due to the core ionizations in DNA
is found to be equal to (75±27)%. This surprisingly large contribution strongly suggests new
mechanisms associated with critical lesions for cell inactivation.
2
Boissière et al. DNA core ionization and cell inactivation
INTRODUCTION
Ionizing radiation is known to cause damages to DNA molecules that can lead to cellular
death or cell inactivation. It is well understood that energy deposition by ionizing radiation at
various DNA sites, such as sugars and bases, lead to permanent molecular changes and those are
known to be the initiating events for the inactivation process. Probing with different qualities of
ions at different initial energies it was demonstrated experimentally that the inactivation crosssection increased with the increase of the linear energy transfer (LET) and the cross-section
reached a maximum at a certain LET value and then decreased continuously with further increase
in LET (1). This phenomenon could not be explained on the basis of damage to overall sugar and
base molecules.
In order to explain the maxima in the cross-section we had suggested that the damages needed to
be examined at the atomic levels that constitute the sugar and base molecules (2). Indeed for a
large range of energetic ions we had demonstrated a striking correlation between the inactivation
of various cells and inner shell ionization cross-sections in carbon, nitrogen, oxygen of the DNA
molecule. We had called this as “core ionization”. It is to be noted that the phenomena of core
ionization should also include comparatively less frequent L-shell events in very heavy atoms
such as phosphorous and sodium that are located in the backbone of DNA. It is well known that
K-ionization (or L-shell) cross-sections reach a maximum for ion velocities near the K-shell
electron velocities and hence at velocities below or above this critical parameter the cross-section
is expected to be lower. The core events are characterized by multiple ionizations at the atomic
site of interaction along with the emission of two electrons, one is the secondary electron ejected
from the K-shell and the other is the Auger electron. These electrons individually have hundreds
3
Boissière et al. DNA core ionization and cell inactivation
of electron volts and can be quite damaging as they deposit very high amount of energy within a
small volume as they undergo multiple scattering.
The phenomenon of core ionization has not gained prominence due to the fact that compared to
outer shell ionizations these are much lower frequency events. For example, compared to fast
ions, fast electrons and ejected electrons from gamma rays and X-rays, the core ionization
phenomenon may only amount to several thousandths of all the rest of the events but their
biological consequences, such as cell inactivation can be many orders of magnitude higher. In the
past in order to explain certain experimental observations for ultra-soft X-rays both the outer
shell ionizations as well as the core ionizations on DNA were included collectively which failed
to explain the biological efficiencies of these types of X-rays. This method of analysis perhaps
led to overlook the independent contribution of the core events (3).
In this study, we will present the results of our investigation in the past regarding the plausibility of
the core ionization model of lethality for ultra-soft X-rays and then its possible extension to fast
electrons and  rays.
MATERIALS AND METHODS
Demonstration of core events by ultra-soft X-rays
In order to demonstrate the phenomenon of core events, ultra-soft X-rays have been used by
properly tuning its energies to match near the various K-shell and L-shell ionization potentials.
Energy values of these types of radiation can be selected to be either below the threshold or
above the threshold of the K-shell or the L-shell binding energies. By varying the X-ray energy
above a given K-threshold, it is possible to produce a specific type of Auger electrons and also
choose the energy of the secondary electron which allows to mimic various categories of core
4
Boissière et al. DNA core ionization and cell inactivation
events produced by energetic electrons (4). We will address reference 4 as Paper I throughout
this paper. We have demonstrated that the biological efficiencies can be manipulated by varying
the energies around the respective K-thresholds of Carbon and Oxygen. For energies of soft Xrays between 290 eV and 540 eV that are just above the K-threshold for carbon atoms and just
below the K-threshold for oxygen atoms, the number of DNA core events for a given dose in a
cell is about twice larger than outside this range. This effect is due to the Carbon abundance
about three times larger in the DNA than in the medium that contain the cells. Hence within this
energy range there will be a sharp increase in the biological effects. It is customary to express the
relative efficiencies of a given radiation by comparing its effects for a given biological end point,
such as cell survival, for that of standard 60Co-  rays. The ratio of the relative doses between the
two radiation qualities to obtain the same level of survival of irradiated cells is known as RBE.
Within the energy range mentioned above there is a sharp increase in RBE demonstrating the
phenomenon as well as the importance of core events.
Three experiments were performed at a synchrotron facility in Orsay, France, with V79 cells (Paper I
and (5)). From the data presented in Paper I it is quite evident that the RBE increased by a factor of
two as the energy of the ultra-soft X-ray varied from 250 eV to 350 eV. The results were further
reproduced in an experiment with another human cell line called B3 cells2.
An expression for mean lethal efficiencies of specific core events
In order to assess the core contribution to lethality in the case of fast electrons and  rays it is
necessary that one can extract the mean lethal efficiency of core ionizations produced by an X-
2
A. Boissière, Contribution « K » à l’effet biologique des rayonnements ionisants. PhD Thesis, Paris, 2004.
5
Boissière et al. DNA core ionization and cell inactivation
ray of any energy. We need to derive a mathematical expression that will include known
experimental data.
Let us denote a core event by a pair of energies (EA) and (ES) where EA and ES respectively refer
to Auger and secondary electron energies. Let us also define Y(E) as the mean yield of lethal
events due to YK(E) core events, where E  EA + ES and it will be equated to 2.3 lethal events for
a dose (D10) that correspond to 10% survival level. Hence
Y(E) 
2 .3
D10 ( E )
With D10 
[1]
7.2
RBE ( E )
[2]
The value of 7.2 Gy refers to the dose of 60Co- rays for 10 % survival level.
Thus Y(E) 
2.3  RBE ( E )
7.2
[3]
In the core picture of lethality, the mean lethal efficiency of core ionizations produced by X-rays
of energy E is:
 (E) 
Y ( E ) 2.3  RBE ( E )

YK ( E )
7 .2  Y K ( E )
[4]
YK (E) values have been calculated as indicated in our previous publication (Paper I) and
available experimental data on RBE are presented in Table 5.
Sometimes it has been useful to represent (E) in terms of a well-known value, (E=278eV) that
has been experimentally determined with great accuracy. It should be noted that this energy is
just below the K-threshold energy of 290 eV for Carbon-K shell and then the core ionizations are
just the L-ionizations of phosphorus atoms. One can then write the following expression:
 (E)   (278) 
RBE ( E ) YK ( 278)

RBE ( 278) YK ( E )
[5]
6
Boissière et al. DNA core ionization and cell inactivation
Equation [5] is a more desirable way of evaluating the core lethal efficiencies because it is
represented in terms of the ratio of the RBE values rather than the absolute values of RBE.
Note that when the X-ray energy is greater than 1500 eV, core ionizations occur not only as a
result of direct photo-ionizations but also from atomic interactions of the secondary electrons, a
phenomenon which requires a threshold-energy of about twice the K binding energy (6). These
secondary core events are more complex than the direct ones since they associate four ionizing
particles: the projectile electron, before and after ionization, and the ejected- and Augerelectrons. Calling Ei, Ej, Ek, E1 the energies of these particles, s(Ei, Ej, Ek, E1) may be calculated
with one or the other of the two following hypotheses (Paper I):
-a single electron is sufficient to create the kind of complex damage leading to cell inactivation.
The s efficiency is then just the sum of the efficiencies of the various electrons, considered either
individually or in pairs. Since only this last configuration may be experimentally investigated, the
following expression is used:
 S1 (E i , E j , E k , E l )   (E i , E j )   (E k , E l )
[6]
-the production of complex lethal damage requires the participation of two electrons. Then the
various electron pairs must be considered and the following expression is used:
 S 2 (E i , E j , E k , E l )   (E i , E j )   (E i , E k )   (E i , E l )   (E j , E k )   (E j , E l )   (E k , E l )
[7]
In the absence of mechanisms involved in the production of lethal damage, equation [8] is used
as a first approximation:
S 
1
( S 1   S 2 )
2
[8]
7
Boissière et al. DNA core ionization and cell inactivation
For X rays of various energies the probability of secondary core events may be found in (7). After
correction from their contribution to RBE(1500) (equation [8]), the lethal efficiency of the pure
(500, 1000) event is found to be ~ 1.7%.
Further analysis of the validity of the core model of lethality for ultra-soft X-rays
For ultra-soft X-rays above the Carbon K threshold, core ionization is by far the main
interaction channel with dry DNA (~90% of all interactions (paper I)). However, below 290 eV
the relative probability of this phenomena become smaller (~50% of all interactions for instance
at 278 eV). We discuss below experiments which validate the core model even in this energy
range.
When the soft X-ray energy is 278 eV then as a result of L shell ionization in phosphorous we
have two ejected electrons, one is due to the Auger process and the other one is due to the photoionization process. Both of these electrons have the same energies, namely 120 electron volts.
This L(P)(120,120) process has a lethal efficiency which is found to be equal to (1.1±0.3)% using
the experimental RBE of 278 eV X-rays (Table 5). This value may be compared with the one
extracted from other experiments performed at X-ray energy equal to 2153 eV which is
equivalent to the K-shell binding energy in phosphorous (8). The K(P) absorption process gives
rise to the emission of a 2 keV K Auger electron along with the simultaneous emission of two
120 eV L Auger electrons. It can be argued that the whole event should have nearly the same
efficiency as that of L(P) absorption at 278 eV. The 2 keV Auger electron due to the absorption
of 2153 eV contributes very little to the local ionization density compared to 120 eV electrons.
Also the ionizations produced at a distance along its trajectory contribute very little to the overall
lethal efficiency due to the K(P) event. Indeed a  10–4 lethal efficiency is obtained for these 2
8
Boissière et al. DNA core ionization and cell inactivation
keV electrons from equation [4]. So the K(P) absorption at 2153 eV and the L(P) at 278 eV differ
in the residual charges left after the Auger emissions, respectively +4 and +2, which may have an
impact on the direct deposition of energy on the strand where the phosphorous atom is located.
However, results on the fragmentation of dry oligonucleotides at the phosphorous absorption
edge have been on the contrary (9).
Experimentally RBE values were determined for 2147 eV and 2153 eV X-ray beams. These two
beams are respectively off-resonance and on-resonance with respect to K(P) process. At both
these energies the ionization patterns are quite identical except there are ~20 additional K(P)
vacancies on DNA per Gray at 2153 eV. This number is the difference between the total number
of K vacancies per Gray at 2147 eV (12 when calculated as in Paper I) and the one at 2153 eV,
calculated from the mean ratio of the absorptions of a DNA film at the two energies (9-10). For
SHE cells used in these experiments, the mean nuclear doses for one lethal event are respectively
1.7 and 3.1 Gray so that 0.27 lethal event is associated with the 20 additional core ionizations in
P atoms. This leads to  (K(P)) = (1.4±0.4)%. Since V79 cells are about 20% less sensitive than
SHE cells (8), their (K(P)) value (~ (L(P)(120,120)) is (1.1±0.3)%.
So the value (1.1±0.3)%, obtained for (L(P)(120,120) when attributing the whole lethal effect of
the 278 eV X-rays to the sole core ionizations on DNA, is the same that the one, (1.1±0.3)%,
obtained without any assumption at 2153 eV. This means that the contribution to lethality of
interactions other than core ionizations on DNA is small, if exists, in this case. This conclusion
should be even more true for ultrasoft X-rays with energies above the Carbon and Oxygen Kthresholds when the proportion of core interactions on DNA is of the order of 90%.
Evaluation of core-event contribution to cell inactivation by low-LET radiation
9
Boissière et al. DNA core ionization and cell inactivation
Conceptually the phenomena of core ionization events are relatively simple to understand for
ultra-soft X-rays. For standard ionizing radiations, such as high- energy electrons and gamma
rays, it is somewhat complex because of the presence of many events besides core events. Outer
shell excitation and ionization phenomena have large cross-sections and they are present
simultaneously with the core events. Experimentally it is difficult to separate them and hence one
has to rely on theoretical models to evaluate the importance of core events. We now proceed to
calculate the “K” contribution to cell inactivation for low-LET radiation. Typical examples of
such radiation qualities are 100 keV electrons and gamma rays and there are several others.
For such radiation quality we will define the “K” contribution to the cell inactivation (CK) as the
ratio between the theoretical value of the number of lethal events per Gray due to the K events
only and the experimentally determined total number of lethal events per Gray due to all the
processes of ionization and excitation including the K events.
For 100 keV electrons, based on equation [3] and an average RBE of 0.9 for V79 cells (11-12),
the yield of experimental lethal events is 0.28. The mean number of lethal events per gray
coming from “K” events only on DNA can be expressed for its part by:
Ylethal.K 
 Y Ki   leti( E ) 
[9]
i
where Y Ki is the number of core events per gray for the species i, and  leti (E )  is the mean
lethal efficiency relative to core events of species i. (We call lethal efficiency of an event, his
probability to lead into cell death).
We will have for the “K” contribution to the lethality:
CK 
Y
Ki
  leti ( E ) 
i
0.28
[10]
10
Boissière et al. DNA core ionization and cell inactivation
Theoretical evaluations of Y Ki and  leti (E )  are explained below.
Number of core events on the DNA per gray. We can simulate by Monte Carlo technique the
interactions produced by the track of a radiation in a homogenous environment. The purpose of
our calculation is to evaluate the mean number of “K” events created by 1 Gy of 100 keV
electrons in liquid water. We then deduce the number of “K” events in the DNA by a
proportionality factor reflecting the atoms involved and their capacity to be ionized in their innershell. The calculation is performed for a collection of cells irradiated at electron equilibrium
involving secondary electrons.
For a V79 cell nucleus (with mass Mnuclei=289 pg (13)), 1 Gy of 100 keV electrons correspond to
the full absorption of Ne primary electrons:
15
1
(kg)
19 ( eV / kg)  289  10
1
.
6
.
10
Ne 
 18.1
3
100  10 (eV )
[11]
In the following we call NK the total number of “K” events produced along a 100 keV electron
track in water. The yield YK of “K” events on DNA per gray of 100 keV electron absorbed in the
nucleus is extracted from the number N e  N K of “K” events produced by the 100 keV electrons
in the nucleus-equivalent volume of water, corrected for the relative abundances of the atoms in
the DNA and in the nucleus, weighted by the respective “K” ionization cross sections.
By calling nC , nN , nO , nP , nNa the numbers of atoms of the various species in the DNA and NH2O
the number of water molecules in the nucleus-equivalent volume (see Table 1) the yield of “K”
events on DNA per gray absorbed in cell is:
YK  N e  N K  
[nC   KC ( E )  nO   KO ( E )  n P  4   LP ( E )  n N   KN ( E )  n Na   KNa ( E )]

N H 2 0   KO
[12]
11
Boissière et al. DNA core ionization and cell inactivation
where  KC ,  KN ,  KO ,  LP ,  KNa are the cross sections for inner-shell ionizations of carbon,
nitrogen, oxygen, phosphorus and sodium atoms by primary and secondary electrons with energy
E. The above average is done over all the energies appearing in the slowing down of the primary
particle and of the electrons of subsequent generations.
The calculation will be derived considering that the target is DNA alone without the first
hydration layer. Indeed events in this first layer participate to direct effect (14) but they have a
low probability to yield double-strand breaks (15).
We can define YK i the number of “K” event for the species i:
YKi  N e  N K 
ni
 i (E)
  KO

N H2 0
 K (E)
[13]
<Table 1>
TABLE 1
Numbers of atoms of the various species in the DNA and number of water molecules in a V79
nucleus-equivalent volume
Element
Number of Atoms
M
Water
NH2O  nucleus  6.02.1023 = 9.66x1012
m H 2O
Carbon
Oxygen
Nitrogen
Phosphorus
Sodium
nC = 1.11x1011
nO = 7.82x1010
nN = 3.88x1010
nP = 1.04x1010
nNa = 0.57x1010
The ratios of the cross sections in the equation [12] have been determined in the linear decreasing
part of the cross section curves relative to the energy, which correspond to the range of the mean
energy of the electron in this interaction of 100 keV electrons with water molecules. We use the
Kim’s cross sections corrected for relativistic effects (16) which agree with experiment under 5%
of uncertainty. The various ratios
 Ki
are given in the next Table 2. Ratios are per electron and
 KO
12
Boissière et al. DNA core ionization and cell inactivation
per shell. For the phosphorus it is the L shell with 8 electrons, explaining thus the factor 4 in the
equation [12].
<Table 2>
TABLE 2
Average ratios of inner-shell cross sections in shell j (K or L) of the atoms i of the DNA: ji,
relative to the one in the shell K of the oxygen: KO
O
 KN
C
 LP
K


 K





O
O
O
K
K
K
 KO
2.0
1.5
4.2
1
The values of NK have been calculated by two different codes: TILDA for water vapor and
TRACELE for liquid water.
The TILDA code has been previously described (17). For electron kinetic energies below 10 keV,
it is now implemented with refined theoretical ionization cross sections (18) taking into account
the projectile-electron and target-electron interactions in the entrance and exit channels
respectively. Doubly differential
d ²
as well as singly differential cross sections are
ddE S
calculated and the last cross sections compare well with experimental data. For high-energy
electrons, Kim’s cross sections corrected for relativistic effects (16) have been used.
The TRACELE code is derived for liquid water and is described in details in (19). Ionization
cross sections are those of Rudd (20) adapted for the liquid phase and are used for electrons up to
150 keV.
Table 3 presents the results of the two codes concerning the number of core events created by a
100 keV electron along its path. Core events created by the primary electron and by the
secondary ones are here distinguished since their local ionization densities are not the same so
that different lethal efficiencies must be associated with them.
<Table 3>
13
Boissière et al. DNA core ionization and cell inactivation
TABLE 3
Number of “K” events created by a 100 keV electron along its path
Code
NK NK primary NK secondary
TILDA
8
7.6
0.4
TRACELE 9.4 8.5
0.9
These codes also make it possible to calculate the average potential of ions-pair production (Wip)
for various energies. The results obtained in particular for an electron of 100 keV are in concord
with the data of the literature, which confirms the reliability of these codes.
Table 4 presents the various yields of « K » events on the DNA per gray of 100 keV electrons as
indicated in equation [13] from the predictions of the TILDA and TRACELE codes.
<Table 4>
TABLE 4
Various yields of « K » events on the DNA per gray of 100 keV electrons
Code
KC KN KO LP
KP KNa
K
TILDA
3.4
0.9
1.2
2.6
0.05 0.10 8.1
TRACELE 4.0
1.1
1.4
3.0
~0
~0
9.5
Calculation of the mean lethal efficiencies of the core events created by 100 keV electrons
along their pathways. The structure of the “K” events produced by fast electrons is represented
by the diagram of the Fig. 1.
<Figure 1>
FIG. 1. Scheme of the structure of the core events produced by fast electrons. EA: energy of the
Auger electrons. ES: energy of the secondary electrons.
14
Boissière et al. DNA core ionization and cell inactivation
Whereas the Auger electrons have well defined energies (see Fig. 1) the secondary electrons have
a continuous probability distribution approximately proportional to
1
given by the theory of
E S2
Kim (16).
The mean lethal efficiency for “core” ionization in the species i is given by:
 lét
i
(E ,E ) d dE

dE

d dE
 dE
A
s
s
s
[14]
s
s
where d are the Kim differential cross section for the mean initial electron energy.
dE s
In this formula the efficiencies  ( E A , E s ) are the one displayed in Fig. 2.
<Figure 2>
FIG. 2. Lethal efficiencies (EA, ES) of the specific C and O core ionizations produced by the Xrays of Table 5. Since  values likely depend on EA and ES, two separates curves are presented,
corresponding to (~300, ES) and (~500, ES) energy pairs, i.e. to C and O core ionizations
respectively. Note that the (EA=500, ES=310) is equivalent to the (EA~300, ES=500) so the (850)
value have been used for the two curves.
15
Boissière et al. DNA core ionization and cell inactivation
The experimental values of the Fig. 2 represent the lethal efficiencies measured for core event
created with ultra-soft X rays. They are extracted from the experimental RBE and from the
number of “K”-type event on the dry DNA, in a way similar to that detailed in Paper I and (5),
their values are reported in Table 5. Other values are impossible to be measured experimentally
because the energy required would be so much attenuated in cytoplasm that they hardly could
reach the nucleus, thus they are calculated and extrapolated from correspondence with
experimental values.
<Table 5>
TABLE 5
Experimental lethal efficiencies of « K »-type events induced by X-rays. We specify in
parenthesis for each X energy, the couple (EA; Ephoto-electron) corresponding to the energy of the
Auger electron emitted and the one of the photoelectron ejected
E
References
 (E)
M. R. Raju et al. (1987) Radiat. Res. 110, 369-412.
~278 eV
K.M.S. Townsend et al. (1990) Int. J. Radiat. Biol. 58, 499-508.
1.1%
(145 ; 115) D. T. Goodhead et al. (1990) Radiat. Prot. Dosimetry 31, 343-350.
C.M. de Lara et al. (2001) Radiat. Res. 155, 440-448.
350 eV
1.0%
B. Fayard et al. (2002) Radiat. Res. 157, 128-40.
(250 ; 60)
P. O’Neill et al. (1989) Photo-Opt. Instrum. Eng. 1140, 232-236.
~850 eV
4.3%
C.K. Hill et al. (1998) Radiat. Res. 150 513-520.
(500 ; 310)
F. Gobert (2003) PhD Thesis, Pierre et Marie Curie University.
P. O’Neill et al. (1989) Photo-Opt. Instrum. Eng. 1140, 232-236.
~850 eV
4.3%
C.K. Hill et al. (1998) Radiat. Res. 150 513-520.
(500 ; 310)
F. Gobert (2003) PhD Thesis, Pierre et Marie Curie University.
960 eV
3.2%
S.W. Botchway et al. (1997) Radiat. Res. 148 , 317-324.
(500 ; 420)
R. Cox et al. (1977) Int. J. Radiat. Biol. 31, 561-576.
1490 keV
1.7%
M. R. Raju et al. (1987) Radiat. Res. 110, 369-412.
(500 ; 960)
S.W. Botchway et al. (1997) Radiat. Res. 148, 317-324.
Table 6 summarizes the various mean lethal efficiencies of core events in the various atoms of
the DNA.
<Table 6>
TABLE 6
Lethal efficiencies of inner-shell ionizations in the atoms i of the DNA
16
Boissière et al. DNA core ionization and cell inactivation
Type of ionization (i)
shell K of C
shell K of N
shell K of L
shell L of P
EA : Energy of the Auger
electrons (eV)
250
360
500
110
ES : Mean energy of the
secondary electrons (eV)
147
205
273
77
  lét (E) 
i
3.0%
2.4%
1.7%
1.1%
A fraction of core events, those created by low energy electrons (about 3% according to the Monte
Carlo simulation) are more complex. This is the case of “K” events of second generation and those
produced along the primary electron track end, which have a higher ionizing power. The
consideration of these complex events in the way described earlier in Materials and Methods
increases the overall efficiency by ~7%.
RESULTS AND DISCUSSION
Contribution of core event to cell inactivation by low-LET radiations.
Table 7 gives the values of Ylethal.K calculated for 100 keV electrons and for 1 MeV photons
from equation [9]. Also are presented experimental Y lethal.exp extracted from equation [3] (with RBE
= 1 for 60Co  -rays and RBE = 0.88 for 100 keV electrons). This last value is the average of the
electron RBE at 20 keV (0.85(11)) and of the one at 1 MeV (0.91(12)). For the photon case we use
the number of “K” events on the DNA published in (7).
<Table 7>
TABLE 7
Calculated number of lethal events originating from « K » events, and experimental lethal
numbers for low LET radiation
Code
Ylethal.K Ylethal.exp
TILDA (fast e 100 keV)
0.18
0.28
TRACELE (fast e- 100 keV)
0.21
0.28
PARTRACK (1 MeV photons) 0.24
0.33
17
Boissière et al. DNA core ionization and cell inactivation
We remind that the core-event contribution to cell inactivation, defined as the ratio between the
calculated and experimental Ylethal is given by CK 
Ylethal . K
(Equation [10]). For the V79 cellYlethal . exp
line the « K » contribution is of the order of 75%.
Uncertainty on the core contribution to cell inactivation.
The main contribution to Ylethal.K (Equation [9]) is the one relative to the carbon atoms. The
relative standard error on YKc, due to the theoretical errors on the many cross sections used and to
the statistical error, is evaluated to ~20%.
On the other hand the relative standard error on

C
εlet (E)  , due to the many uncertainties on the
experimental points determining the shape and the height of the  curve in Fig. 2, is evaluated to
~30%.
So the relative standard error on Ylethal.K is of the order of 36% and the absolute standard error on
the core contribution to cell-killing efficiencies (Equation [10]) is of the order of 27%.
Finally the “K” contribution to cell inactivation by usual (low LET) radiation is found equal to ~
(75±27) %.
Core model and non-linear effects.
It is interesting to note that the induction of core events is linear with dose in view of the fact
that in this model no assumption has been made with respect to linearity of the dose-effect
relationship for lethal damage. The calculation presented here is based on a typical value of RBE
18
Boissière et al. DNA core ionization and cell inactivation
at 10% survival (Paper I). Instead of 10% if the selected value of RBE is at 1/e survival, there is
no significant change in the core contribution.
Core ionization as a potent critical event.
The main conclusion of this work is that the core contribution to the lethality of usual lowLET radiations- fast electrons, γ-rays- is very large, of the order of 75%. It is interesting to note
that we previously found a similar estimation for ions (Paper I).
Originally the core model -in which the lethal effectiveness is associated with the sole coreionizations on DNA- was proposed for ultrasoft X-rays. For these particles it is now
demonstrated that this picture is basically correct since it not only qualitatively reproduces the
RBE variations, especially the abrupt rise above versus below the Carbon K-threshold, but it also
provides absolute values of core efficiencies consistent with experimental data. This conclusion
is not surprising since core ionization is the main interaction channel of ultrasoft X-rays with
DNA.
By contrast the new result presented here for low-LET radiations is noteworthy because core
ionization phenomena represent no more than 10% of the total energy deposited by these
particles (the same is true for ions).
It should be noted that the core model provides a natural explanation for the long-time puzzling
observation of an RBE increase of X rays at decreasing energy (21)): this phenomenon is nothing
but a consequence of their increasing effectiveness to trigger core events.
The critical events leading to cell inactivation have been investigated for a long time. In former
stochastic models, clusters of ionizations produced at the track ends of low-energy electrons were
19
Boissière et al. DNA core ionization and cell inactivation
thought to be the source of biological effects (22). These models are unable to reproduce the
abrupt rise of RBE at the Carbon K-threshold even when the specific atomic composition of
DNA is considered3 .The present study shows that the lethal effect is dominantly associated with
the pattern of two spatially correlated electrons and one positive hole, as achieved by core
ionizations.
Moreover, there are some indications that these events could also be deeply involved in the
production of chromosomal aberrations (23 and ²).
Therefore the study of the various end points initiated by core ionizations on DNA appears as an
essential approach to characterize the nature of the critical radiobiological damage and the
involved repair mechanisms.
Using ultra soft X-rays, the core events will allow a completely new way to view the
fundamentals of radiobiological effects.
ACKNOWLEDGMENTS
This work was supported by EDF (contract n° RB 2005-36) and by France-Berkeley Fund.
REFERENCES
1.
G. Kraft, Radiobiology of very heavy ions: inactivation, induction of chromosome
aberrations and strand breaks, Nucl. Sci. Appl., 3, 1-28 (1987).
3
B. Fayard, Cassures d’ADN induites par ionisations en couche interne et leurs conséquences biologiques. PhD
Thesis, Paris, 1999.
20
Boissière et al. DNA core ionization and cell inactivation
2.
A. Chetioui, I. Despiney, L. Guiraud, L. Adoui, L. Sabatier, and B. Dutrillaux, Possible role
of inner-shell ionization phenomena in cell inactivation by heavy ion, Int. J. Radiat. Biol.,
65, 511-522 (1994).
3.
D. T. Goodhead, J. Thacker and R. Cox, Is selective absorption of ultra soft X-rays
biologically important in mammalian cells?, Phys. Med. Biol., 26, 1115-1127 (1981).
4.
B. Fayard , A.Touati , F. Abel , M. A. Hervé du Penhoat, I. Despiney-Bailly, F. Gobert, M.
Ricoul, A. L’Hoir , M. F. Politis, M. A. Hill, D. L. Stevens, L. Sabatier, E. Sage, D. T.
Goodhead and A. Chetioui, Cell inactivation and double-strand breaks: the role of core
ionizations, as probed by ultrasoft X rays, Radiat. Res., 157, 128-40 (2002).
5.
M. A. Hervé du Penhoat, B. Fayard, F. Abel, A. Touati, F. Gobert, I. Despiney-Bailly, M.
Ricoul, L. Sabatier, D. L. Stevens, M. A. Hill, D. T. Goodhead and A. Chetioui, Lethal effect
of carbon K-shell photoionizations in chinese hamster V79 cell nuclei: experimental
methods and theoretical analysis, Radiat. Res., 151, 649-58 (1999).
6.
Y. K. Kim and M. E. Rudd, Binary-encounter-dipole model for electron impact ionization,
Phys. Rev. A, 50, 3954-3967 (1994).
7.
P. Bernhardt, W. Friedland and H. G. Paretzke, The role of atomic inner shell relaxation for
photon induced DNA damage, Radiat. Environ. Biophys, 43, 77-84 (2004).
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M. Watanabé, M. Suzuki, K. Watanabé, K. Suzuki, N. Usami, A. Yokoya and K. Kobayashi,
Mutagenic and transforming effects of soft-X-rays with resonance energy of phosphorus Kabsorption edge, Int. J. Radiat. Biol., 61, 161-168 (1992).
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H. Yamada, K. Kobayashi and K. Hieda, Effects of the K-shell X-ray absorption of
phosphorus on the scission of the pentadoxythymidylic acid, Int. J. Radiat. Biol., 63, 151159 (1993).
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Boissière et al. DNA core ionization and cell inactivation
10. K. Hieda, T. Ito, Radiobiological experiments in the X-ray region with synchrotron radiation.
In Handbook of Synchrotron Radiation 4, (S. Ebashi, M. Koch and E. Rubenstein, Eds), pp.
431-465. Elsevier, (1991).
11. S. Iwanami, N. Oda, Theory of relative biological effectiveness (RBE) of tritium beta rays:
bacteria killing effects of tritiated water, Phys. Med. Biol., 32, 1469-1479 (1987).
12. G. W. Barendsen, Dose-survival curves of human cells in tissue culture irradiated with
alpha-, beta-, 20 keV X- and 200 keV X-radiation, Nature, 193, 1153-1155 (1962).
13. A. I. Kassis, F. Fayad, B. M. Kinsey, K. S. Sastry, R. A. Taube, S. J. Adelstein,
Radiotoxicity of 125I in mammalian cells, Radiat. Res. 111, 305-318 (1987).
14. S. G. Swarts, M. D. Sevilla, D. Becker, C. J. Tokar and K. T, Wheeler, Radiation-induced
damage as a function of hydratation, I. Release of unaltered bases, Radiat. Res., 129, 333344 (1992).
15. A. Yokoya, S. M. T. Cuniffe and P. O’Neill, Effect of hydratation on the induction of strand
breaks and base lesions in plasmid DNA films by γ- radiation, J. Am. Chem. Soc., 124, 88598866 (2002).
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17. C. Champion, A. L'Hoir, M.F. Politis, P.D. Fainstein, R.D. Rivarola, A. Chetioui, A MonteCarlo code for the simulation of heavy-ion tracks in water, Radiat. Res., 163, 222-231
(2005).
18. C. Champion, J. Hanssen, and P. A. Hervieux, Theoretical differential cross sections of
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as tool for testing and developing biophysical models of radiation action, Radiat. Prot.
Dosimetry, 31, 343-350 (1990).
22. D. T. Goodhead, The initial physical damage produced by ionizing radiations, Int. J. Radiat.
Biol., 56, 623-634 (1989).
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Abel, M.-F. Politis, B. Fayard, J. M. Guignier, L. Martins, I. Testard, L. Sabatier and A. Chetioui,
Chromosome aberrations and cell inactivation induced in mammalian cells by ultrasoft X-rays:
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23
Boissière et al. DNA core ionization and cell inactivation
FIGURE LEGENDS
FIG. 1. Scheme of the structure of the core events produced by fast electrons. EA: energy of
the Auger electrons. ES: energy of the secondary electrons.
FIG. 2. Lethal efficiencies  ( E A , E s ) of the specific C and O core ionizations produced by
the X-rays of Table 5. Since  values likely depend on EA and ES, two separates curves are
presented, corresponding to (~300, ES) and (~500, ES) energy pairs, i.e. to C and O core
ionizations respectively. Note that the (EA=500, ES=310) is equivalent to the (EA~300, ES=500)
so the  (850) value have been used for the two curves.
24
Boissière et al. DNA core ionization and cell inactivation
FIG. 1.
25
Boissière et al. DNA core ionization and cell inactivation
FIG. 2.
26
Boissière et al. DNA core ionization and cell inactivation
FOOTNOTES
1
Address for correspondence: Institut de Minéralogie et Physique des Milieux Condensés,
Université Paris 6, CNRS UMR 7590, Campus Boucicaut, 140 rue de Lourmel, 75015 Paris
Cedex, France; email: [email protected]
2
A. Boissière, Contribution « K » à l’effet biologique des rayonnements ionisants. PhD
Thesis, Paris, 2004.
3
B. Fayard, Cassures d’ADN induites par ionisations en couche interne et leurs conséquences
biologiques. PhD Thesis, Paris, 1999.
27
Boissière et al. DNA core ionization and cell inactivation
TABLES
TABLE 1
Numbers of atoms of the various species in the DNA and number of water molecules in a V79
nucleus-equivalent volume
Element
Number of Atoms
M
Water
NH2O  nucleus  6.02.1023 = 9.66x1012
m H 2O
Carbon
Oxygen
Nitrogen
Phosphorus
Sodium
nC = 1.11x1011
nO = 7.82x1010
nN = 3.88x1010
nP = 1.04x1010
nNa = 0.57x1010
TABLE 2
Average ratios of inner-shell cross sections in shell j (K or L) of the atoms i of the DNA: ji,
relative to the one in the shell K of the oxygen: KO
O
 KN
C
 LP
K


 K





O
O
O
O
K
K
K
K
2.0
1.5
4.2
1
TABLE 3
Number of “K” events created by a 100 keV electron along its path
Code
NK NK primary NK secondary
TILDA
8
7.6
0.4
TRACELE 9.4 8.5
0.9
TABLE 4
Various yields of « K » events on the DNA per gray of 100 keV electrons
Code
KC KN KO LP
KP KNa
K
TILDA
3.4
0.9
1.2
2.6
0.05 0.10 8.1
TRACELE 4.0
1.1
1.4
3.0
~0
~0
9.5
TABLE 5
Experimental lethal efficiencies of « K »-type events induced by X-rays. We specify in
parenthesis for each X energy, the couple (EA; Ephoto-electron) corresponding to the energy of the
Auger electron emitted and the one of the photoelectron ejected
E
References
 (E)
M. R. Raju et al. (1987) Radiat. Res. 110, 369-412.
~278 eV
K.M.S. Townsend et al. (1990) Int. J. Radiat. Biol. 58, 499-508.
1.1%
(145 ; 115) D. T. Goodhead et al. (1990) Radiat. Prot. Dosimetry 31, 343-350.
C.M. de Lara et al. (2001) Radiat. Res. 155, 440-448.
350 eV
1.0%
B. Fayard et al. (2002) Radiat. Res. 157, 128-40.
(250 ; 60)
~850 eV
4.3%
P. O’Neill et al. (1989) Photo-Opt. Instrum. Eng. 1140, 232-236.
28
Boissière et al. DNA core ionization and cell inactivation
(500 ; 310)
~850 eV
(500 ; 310)
C.K. Hill et al. (1998) Radiat. Res. 150 513-520.
F. Gobert (2003) PhD Thesis, Pierre et Marie Curie University.
P. O’Neill et al. (1989) Photo-Opt. Instrum. Eng. 1140, 232-236.
C.K. Hill et al. (1998) Radiat. Res. 150 513-520.
F. Gobert (2003) PhD Thesis, Pierre et Marie Curie University.
4.3%
960 eV
(500 ; 420)
S.W. Botchway et al. (1997) Radiat. Res. 148 , 317-324.
3.2%
1490 keV
(500 ; 960)
R. Cox et al. (1977) Int. J. Radiat. Biol. 31, 561-576.
M. R. Raju et al. (1987) Radiat. Res. 110, 369-412.
S.W. Botchway et al. (1997) Radiat. Res. 148, 317-324.
1.7%
TABLE 6
lethal efficiencies of inner-shell ionizations in the atoms i of the DNA
i
Type of ionization (i)
EA : Energy of the Auger
ES : Mean energy of the
  lét (E) 
electrons (eV)
secondary electrons (eV)
shell K of C
250
147
3.0%
shell K of N
360
205
2.4%
shell K of L
500
273
1.7%
shell L of P
110
77
1.1%
TABLE 7
Calculated number of lethal events originating from « K » events, and experimental lethal
numbers for low LET radiation
Code
Ylethal.K Ylethal.exp
TILDA (fast e- 100 keV)
0.18
0.28
TRACELE (fast e 100 keV)
0.21
0.28
PARTRACK (1 MeV photons) 0.24
0.33
29