Formal Discussion
of: Somatic Mutations
HOWARD
Biology Department,
J.
and Carcinogenesis'
CURTIS
Brookhaven National Laboratory,
Upton, New York
SUMMARY
A great deal of evidence
is now available
indicating
that mutations
play a dominant
role in the aging process.
Because carcinogenesis is one facet of the aging process, mu
tations must play a major role here also. However, carcinogenesis certainly does not
consist
of the simple
uncontrolled
tions,
The mutation
theory
to all workers
attractive
which
theory,
argue
of a mutation
as well as a specific event,
that mutations
known
induction
before
field.
It
carcinogenesis
is old and well
is an extremely
and there are many pieces of evidence
Radiation,
which
is a very
po
tent mutagenic agent, is also a very effective carcinogen.
Most, if not all, of the chemical carcinogens are also muta
gens. The virus theory of carcinogenesis is well estab
lished
in certain
special
cases, and in a very real sense it is
a facet of mutation theory.
On the other hand, there are many puzzling phenomena
which cause one to doubt this theory.
For example, if
the theory is true, why does it take as long as 40 years
after
a person
receives
a dose of radiation
for a tumor
to
appear?
If a mutation to carcinogenicity
is produced,
one would expect it to manifest itself very soon thereafter.
There is also the question of whether mutations, especially
spontaneous mutations, can be produced in somatic cells.
A number
1 The
research
havenNational
reported
in
this
Laboratoryunder
paper
was
carried
out
at
the property
of
can take place.
It is postulated
for both conditions.
of the 1st generation (3). We then scored chromosome
aberrations in liver cells of mice by inducing cell division
by partial hepatectomy
and examining anaphase figures
(14). (Fig. 1). The percentage of cells containing aberra
tions is taken as an index of the mutations present in these
cells.
With the use of this technic a number of different situa
tions have been investigated.
First it was found that
aberrations increase steadily with age (Chart 1). Next,
following a dose of X-radiation,
there are a large increase
in aberrations and a very slow return to normal (Chart 1).
It is known that chronic X- or ‘y-radiationis only about
25 % as effective in shortening the life-span as an acute
dose of the same total size. It is found that the same is
true for the production of chromosome aberrations (Chart
2). It is also known that for neutron irradiation, chronic
irradiation is just as effective in shortening the life-span
of years ago it was noted that the life expec
as acute
tancies of animals exposed to ionizing radiation are short
ened in proportion
to the radiation dose received.
It
appears that the animals die from the same diseases and
in about the same proportion as normal animals, but they
contract the diseases sooner.
In investigating
this phe
nomenon, it seemed that a very likely explanation would
be that the radiation induced mutations in the somatic
cells which caused a lack of efficiency and, thus, deteriora
tion in organs whose cells are not capable of cell division
and cancer in organs whose cells regularly undergo mitosis.
In order to test this idea, it was necessary to develop a
method for estimating mutations in somatic cells. For
this we made an analogy with plants, where, because
somatic cells differentiate to form germ cells, chromosome
aberrations in somatic cells can be related to true muta
tions scored in the next generation.
It was found that
the mutations scored in the 2nd generation are propor
tional to chromosome aberrations scored in somatic cells
confers
There must be some general condi
in one form or another are responsible
of carcinogenesis
in the
in its favor.
in one cell which
growth on that cell and its progeny.
irradiation.
The
same
has been
production of chromosome aberrations
It is well known that some inbred
quite
found
short-lived
and
that chromosome
in one short-lived
strain
(4) (Chart
strain
found
for the
(8) (Chart 3).
strains of mice are
some quite
long-lived.
It was
aberrations
increase quite rapidly
and quite slowly in a long-lived
4).
Thus in every situation tried so far there is a direct
correlation between the degree of life shortening and the
development of somatic mutations.
It should be kept in
mind that irradiated animals die, to a very considerable
extent,
factors
from cancer, so that
which cause increased
one can equally well say that
somatic mutations
also quan
titatively increase cancer production.
One of the problems raised by this work is the long de
lay between cause and effect. Thus a dose of radiation
Brook
the auspices ofthe U. S. Atomic
Energy Commission.
given
to a young
adult
mouse
may
produce
serious
chro
mosomal damage in almost every cell in the body; yet
the mouse appears unaffected and may not show signs of
aging until many months later.
At first glance, this situa
tion seems to contradict the theory, but in the light of
1305
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Vol. 25, September 1965
Cancer Research
1306
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CHART 3.—Chromosome aberrations
AGE - MONTHS
CHART 1.—Chromosome aberrations
in liver cells as a function
of age in normal mice and in mice that had received a large dose of
X-rays.
The curves show the steady increase in the number of
chromosome aberrations (mutations) with natural aging and the
dramatic increase and slow return to normal following X-rays
[from Stevenson and Curtis (14)].
ducing aberrations as acute is, and it will be seen that the experi
mental points fall closely on this line. The data show that there is
no chromosomal healing following even small doses of neutrons
[from Curtis et at. (8)1.
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X-rays, (c) normal mice. The dashed line shows the rate of
build-up which would be expected for mice of Group a if chronic
irradiation were as effective in producing chromosome aberrations
as acute irradiation is. Since the experimental curve has a much
smaller slope, it is concluded that chromosome healing takes place
following small doses of radiation [from Curtis and Crowley (7)].
of each, and more than 1000 protein molecules of each.
Thus if a mutation occurs which destroys a single DNA
molecule, the cell can still function for a long time on
RNA, ribonucleic acid.
are:
DNA,
100
deoxyribonucleic
acid;
200
300
400
CHART4.—Chromosome aberrations
500
600
700
inbred strains
in liver cells of normal mice
as a function
of age.
The median
life
span of each strain is indicated by the arrows. In each case the
solid lines represent animals 8 weeks of age at the start of the ex
periment, and the broken lines, old breeding animals about 1 year
old at the start of the experiment [from Crowley and Curtis (4)].
(DNA)1
—ø'(RNA)1-—e@(Protein)1
(DNA)2 —@(RNA)2 —@(Protein)2
(DNA)3 —@(RNA)3
A schematic
representation
of the control of cellular
function is given in Chart 5. It is believed that there is
1 DNA2 molecule of each species, 10—100RNA molecules
used
<@T@7TI I
AGE - DAYS
modern theory of the role of the nucleus in cell function,
it may actually strengthen it.
abbreviations
-
of 2 different
CHART 2.—Chromosome aberrations in liver cells of (a) mice
subjected to chronic 7-irradiation,
(b) mice given a single dose of
2 The
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in liver cells of (a) mice
subjected to chronic neutron irradiation, (b) mice subjected to
acute neutron irradiation, and (c) control mice. The dashed line
gives the rate of build-up of aberrations in Group a mice which
would be expected if chronic irradiation were as effective in pro
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CHART5.—Schematic representation of the control of cell func
tion. The chromosomes contain the different individual DNA
molecules, each of which synthesizes the corresponding RNA
molecules,
which
in turn
synthesize
the corresponding
proteins.
A mutation can be considered as the elimination of 1 or more DNA
molecules, which eventually eliminates that molecular line.
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CURTIS—Formal
Discuss-ion
of Somatic
Mutations
and Carcinogenesis
1307
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FROM START OF EXPERIMENT
CHART 6.—Chromosome aberrations in liver cells of both irradiated and control mice, and
with and without carbon tetrachloride
injections every 45 days (arrows) , which destroyed
about half the liver. The standard error of each point is indicated. This induced mitosis
caused a relatively slow return to control levels [from Curtis et al. (9)].
stored RNA and proteins.
Indeed this seems to be the
case, and one can cite a number of instances in which
cells have been observed to function and even to undergo
a number of mitoses before dying from a lethal mutation
plant cells, and it now seems even more potent for animal
cells. Thus there are 2 mechanisms within the cell for
getting rid of mutations; only one can operate in the fixed
postmitotic cells, but both can operate in cells in active
(5).
turnover.
This
would
then
seem
to adequately
explain
long delay between the induction of chromosomal
and its manifestation.
Another problem, already mentioned, is that
mosome
healing.
One
can
imagine
that
the
damage
of chro
as a group
of
cells undergoes cell division, the cells containing muta
tions are gradually eliminated and the cell line thereby
kept pure. Indeed, this undoubtedly
happens in the
mammal in such tissues as bone marrow.
However,
there certainly must be more to it than that to account
for the observed chromosomal stability, especially in such
nondividing tissues as the liver. To investigate this, we
produced a high percentage of chromosome aberrations in
the livers of mice by a dose of X-rays and, following this,
gave the mice a dose of carbon tetrachloride
to destroy
about half the liver every 45 days (9) (Chart 6). It will
be noted that the aberrations were eliminated faster with
active cell division than without it, but not nearly fast
enough to allow us to say that the 1st cell division elimi
nates a mutation.
On the contrary, it must take an aver
age of more than 4 divisions to eliminate an aberration.
In the normal liver there is very little cell division, but
the aberrations produced by X-rays are eliminated never
theless.
This, combined with the evidence from chronic
-y-ray irradiation,
leads to the conclusion that chromo
somes
are capable
process
of a great
deal of self-repair.
which has long been known
This is a
to be operative
in
The importance of this result for the present discussion
lies in the fact that it gives a great hope for future cancer
research.
It means that chromosome structure is by no
means a fixed entity.
It is apparently
quite unstable,
and it achieves
what
stability
it has by a very considerable
ability for self-repair.
Experiments
with labeled thymi
dine show that this compound is rather readily incorpo
rated into the DNA of interphase cell nuclei, and as readily
lost (10). This indicates an exchange reaction, which in
turn indicates that the DNA structure must be quite
labile.
With our increasing knowledge of protein struc
ture and DNA structure,
one can easily imagine that
methods will be found, perhaps in the very near future,
for stabilizing chromosome structure in the mammal.
One is very tempted to regard aging and carcinogenesis
as 2 separate
processes,
as if cancer
were
some
unhappy
accident.
Indeed it does appear that, in general, tissues
which accumulate mutations do not develop cancer, and
vice versa. However, many lines of evidence seem to mdi
cate that carcinogenesis is merely one form of the aging
process.
Jones (13) has collected data from a great many
different populations
and on many diseases.
He finds
that, in general, in a population which has a short life
expectancy the causes of death are accelerated as compared
to a long-lived population.
It is very interesting that the recent studies on the rela
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1308
Cancer Research
tion between longevity and smoking have shown that
cigarette smoking tends to accelerate a great many causes
of death, such as cancer of the pancreas and coronary
thrombosis,
smoking.
which would hardly seem to be related
to
There seems to be some general aging process
initiated by smoking which affects the organism in such a
way that it is more susceptible to all diseases.
In a recent study by Johnson
rats kept at a cold teniperature
et al. (12) it is shown that
increase their metabolism
considerably in order to maintain their body temperature,
and their life expectancy is considerably decreased.
The
interest
ing thing
is that,
with a minor
exception,
all forms
of death including cancer are accelerated.
As has already been mentioned, ionizing radiation tends
to accelerate all causes of death.
It has been known for many years that if rats or mice
are restricted in their caloric intake, their life expectancy
will be greatly prolonged.
In a recent study, Berg (2)
has shown that all causes of death are delayed in the re
stricted animals.
All these pieces of evidence indicate that when the gen
eral condition of an animal deteriorates (ages), it becomes
more susceptible to all diseases, and that until this condi
tion is met, the induction of these diseases, including can
cer, is unlikely.
On the other hand, one cannot ignore the apparent
specific causes
of cancer,
such as smoking
and lung cancer.
It then seems evident that we must consider both general
and specific causes of cancer, and perhaps both factors
must be present for cancer initiation to take place. Beren
blum and Trainin (1) have ably presented evidence for a
2-factor theory of carcinogenesis,
indicating that it is
necessary
to have
an initiator
achieve cancer induction.
discussing
substances
and a promoter
in order
to
It could well be that they are
which create
the general
and specific
condit ions for carcinogenesis.
Returning now to the radiation problem, it is apparent
that radiation may play 2 roles in the carcinogenic story.
The production of mutations in the cells of the body may
very well create the basis for the general conditions neces
sary for carcinogenesis
and at the same time produce
to account
for this 2-step
process,
and whereas
FIG. 1.—Photomicrograph
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on the Mecha
In: R. J. C. Harris (ed.),
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2. Berg, B. N., and Simms, H. S. Nutrition
and Longevity in the
Rat. II. Longevity and Onset of Disease with Different Levels
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1963.
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the
his
1965
may be correct, one can imagine other specific mechanisms
which would be equally plausible.
Recent work of Wald et al. (15) has shown that at least
some, if not all, radiation-induced
mouse leukemia is
caused by either the creation or the activation of a virus,
which then incorporates itself into a particular chromo
some of some lymphocytes.
This is an important con
cept, and it causes one to wonder just what a mutation is.
11. Failla,
mutation necessary for the production of the specific can
cer. The present experiments certainly show that the
general conditions for cancer induction can be delayed
for a large fraction of the life-span of the animal after
they are created by radiation, and the specific mutation
can wait for an equal time until the general conditions
are met. Failla (1 1) has proposed a more specific mecha
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Vol. 25, September
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of the Aging Process,
I)isease
and Life Expectancy.
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K. G., and Curtis, H. J. Chromosomal
Aberrations
in Irradiated and Nitrogen Mustard
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N., Upton,
A. C., Jenkens,
Treated Mice. Radiation
V. R., and Borges,
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Radiation Induced Mouse Leukemia: Consistent Occurrence
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of an abnormal
dividing liver cell
exhibiting 3 bridges and 3 fragments [from Curtis (6)].
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Downloaded from cancerres.aacrjournals.org on June 14, 2017. © 1965 American Association for Cancer Research.
Formal Discussion of: Somatic Mutations and Carcinogenesis
Howard J. Curtis
Cancer Res 1965;25:1305-1309.
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