Notes on the epigenetic (transplacental and
transgenerational) origins of childhood cancer
ERNESTO BURGIO
ECERI - European Cancer and
Environment Research Institute
ISDE Scientific Committee
FOREWORD 1
(1) Cancer continuous increase
Cancer is generally associated with old age and its
continuous increase, observed throughout the 20th
century in all the industrialized countries, is often
explained
through the theory of the progressive accumulation of
oxidative and stochastic genetic damage (SMT)
and through the continuous improvement of our
diagnostic capacities.
The fact that this increase, from the end of the 80s to
2000, has involved individuals of all ages, young
people included, has been too often underestimated
1
the significant increase in the Less Developed Countries & in young people all over
the world demonstrates the limits of the SMT (necessary link between aging &CA)
Thousands per Annum
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LessDeveloped
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(2) Child cancer increase
Child cancers are generally considered as a rare
disease. But it is worth remembering
• that, statistically, about 1 in 5-600 children will
develop cancer before the age of 15;
• that, in spite of the decisive improvement in
diagnosis and therapy in the last decades, cancer
is the leading cause of death due to diseases
among children over the first year of age;
• that, even at this age, a continuous and
significant increase has been seen during
the last decades.
It is generally argued
that childhood
cancers are a rare
condition.
But it should be
reminded
that CANCER is the
main cause of
death by disease
in childhood
that there is a constant
and significant
increase of tumors
in the world for
this age group
that 1 : 5-600 children
falls ill with
cancer
That more than
13 000 children fall ill
with cancer each
year in the U.S.
Bleyer A, O’Leary M, Barr R, Ries LA,
editors. Cancer epidemiology in older
adolescents and young adults 15-29
ears of age, including SEER incidence
and survival: 1975-2000. NIH Pub. No.
06-5767. Bethesda (MD): National
Cancer Institute; 2006. Jemal A, Siegel
R, Ward E, et al. Cancer statistics,
2008. CA Cancer J Clin 2008;58:71 – 6.
Incidenza di tumori (anno/100.000)
Poumon
Sein
Colon
Estomac
Lymphôme
Rein
Cerveau
Leucémie
lymphati
que
80
60
Alberto Tommasini,
Laboratorio Immunologia
Pediatrica, IRCCS
Burlo Garofolo
is the leading cause of death due to
diseases among children over the first year
of age
40
20
10
5
4
2
1,5
1
<1
Leucémie lymphatique
Encéphale
Lymphômes
Neuroblastoma-Retinoblastoma
Tessuti molli, rene (Wilms), gonadi
(3) ACCIS
• The recent IARC reports of a significant
increase in childhood cancer in Europe and
especially in Italy has caused concern,
2
• forcing us to reconsider critically the
dominant model of carcinogenesis.
As we may easily argue from the recent project ACCIS (Automated Childhood Cancer Information System) - a
comprehensive monitoring conducted by a team of epidemiologists IARC on 63 cancer registries from 19 European
countries, for a total of over 130 thousand tumors of all types (113 thousand children and 18 thousand
teenagers)
http://www-dep.iarc.fr/accis.htm
Cancer incidence in childhood and adolescence - EUROPE
( 1970-1999)
latency
mother
A first draft of the report, published on the Lancet in 2004, demonstrates an annual
increase of 1-1,5% for all cancers (with more marked increases in lymphomas, soft tissue
sarcomas, tumors of the nervous system…) .
Steliarova-Foucher E, Stiller C, Kaatsch P, Berrino F, Coebergh
JW, Lacour B, Parkin M. Geographical patterns and time trends
of cancer incidence and survival among children and
adolescents in Europe since the 1970s (the ACCISproject): an
epidemiological study. Lancet. 2004 Dec 11-17;364(9451):2097105
..in the last 20 years (1978 e il 1997) the overall
incidence rate has increased significantly with an
average annual percentage change (AAPC) of 1,1%
(> 2% in the first year; 1,3 % during adolescence).
These data
should not be
underestimated
for at least
4 reasons:
• the large size of the study sample (63 cancer registries from 19
European countries, for a total of more than 130000 cancers of all
kinds: 113000 strictly paediatric and 18000 teenager cancers);
• a sufficiently prolonged period of observation (20 years);
• the maximum increase in the first year of age, which
suggests a transplacental (from maternal and fetal exposure
to pro-carcinogenic agents) or even a transgenerational
(epigenetic/gametic) origin;
• the concomitant increase in the whole northern
hemisphere of a variety of chronic degenerative diseases
(endocrine-metabolic: obesity, type 2 diabetes;
immune-mediated: allergies, autoimmune diseases,
neuro-development and neuro-degenerative diseases:
autism, ADHD, Alzheimer's disease.. ) for all of which
a significant pathogenic role of the mechanisms
of early epigenetic dysregulation (fetal programming)
on various organs and tissues has recently been suggested
(DOHaD-Developmental Origins of Health and Diseases).
Gluckman PD, Hanson MA. Developmental origins of disease paradigm: a
mechanistic and evolutionary perspective. Pediatr. Res. ( 2004); 56:311–17
What is Cancer ?
Is cancer a
Genetic
Disease ?
Do the somatic cells of a single tissue undergo regression for intrinsic, accidental reasons (mutations or epimutations) and de-differentiate …
The Somatic Mutation
Theory of Carcinogenesis
Over your lifetime, random gene changes are passed along
as your body cells grow and divide, so they accumulate
Nature
The Somatic Mutation
Theory of Carcinogenesis
Genes are altered, or "mutated," in various ways
as part of the mechanism by which cancer arises.
Chemicals and radiation act by damaging genes, viruses introduce their own genes into cells,
and heredity passes on alterations in genes that make a person more susceptible to cancer
What’s Cancer ? 1
We suggest that the vast catalog of cancer cell genotypes is a
manifestation of six essential alterations in cell physiology that
collectively dictate malignant growth
Tumor development proceeds via a process formally
analogous to Darwinian evolution,
in which a succession of stochastic mutations,
each conferring one or another type of growth
advantage, leads to the progressive conversion of
normal human cells into CA-cells…
CA-cells have defects in regulatory circuits that
govern normal cell proliferation and homeostasis…
the vast catalog of CA-cell genotypes is a
manifestation of six essential alterations in cell
physiology that collectively dictate malignant
growth:
1
2
3
4
5
6
2
6
5
6
2
1
4
P
P
3a
P
3
P
3b
RTK Receptor
Tyrosine Kinases
What’s Cancer ? 2
The revolution in cancer research can be summed up
in a single sentence: cancer is, in essence, a genetic disease
Alterations in three types of genes are responsible for tumorigenesis:
oncogenes, tumor-suppressor genes and stability genes
Unlike diseases such as cystic fibrosis or muscular dystrophy,
wherein mutations in one gene can cause disease, no single gene defect
'causes' cancer. Mammalian cells have multiple safeguards to protect them
against the potentially lethal effects of cancer gene mutations,
and only when several genes are defective does an invasive cancer develop
Vogelstein B, Kinzler KW. Cancer genes and the pathways they control Nat Med. 2004 Aug;10(8):789-99.
The GWAS efforts are
certainly creating
bigger haystacks …
The full genome sequence of a lung cancer cell line,
for example, yielded 22,910 point mutations,
only 134 of which were in protein-coding regions
In a recent editorial
on Nature
Heidi Ledford stated
that the millions of
genetic sequences
and SNPs
accumulated in an
attempt to decipher
the genetics of
cancer have built
giant haystacks
in which researchers
have gone lost …
towards an epigenetic paradigm
• .. there is now ample evidence that some specific epigenetic
alterations, (primarily the hypomethylation of DNA, with activation
of oncogenes and increased mobility of mobile sequences) ** are
the result of protracted genomic stress (eg chronic inflammation
and persistent oxidative stress) and generally anticipate, to some
extent preparing it, genetic modification and an overall genomic
instability
• … could these data change our way of representing cancer ?
** and hypermethylation of tumor
suppressor genes promoters
The ‘‘methylation
paradox’’
of cancer cells.
Cancer cells present a gain
of methylated streches at
regions that are usually
unmethylated
(hypermethylation)
concomitantly with
loss of methylation at
genomic loci that are
normally methylated
(global) (hypomethylation),
R Villa, F De Santis, A Gutierrez, S Minucci, PG Pelicci, L Di Croce
Epigenetic gene silencing in acute promyelocytic leukemia Biochem
Pharmacol (2004) 68: 1247-54
Retrosequences
activation
Interphase chromosome
What’s Epigenetics ?
Mitotic chromosome
Heterochromatin
Euchromatin
FOREWORD 2
2
1
We have
3
wrongly
extended the
linear theory of
the gene to the
“realm”
of the gene
management...
but the gene
management is
an entirely
different
process,
involving
interactive
cellular
processes that
display an
interactive
complexity…
which is
epigenetic in
nature
In 1997 the well known molecular biologist R. Strohman attempted an oblique attack against the central dogma of
molecular biology; the deterministic, linear, uni-directional, and encapsulated path from DNA to RNA to proteins to
phenotype..
Francis Crick's statement of the central dogma, from an early draft of Crick (1958)
available at http://profiles.nlm.nih.gov/SC/B/B/F/T/_/scbbft.pdf
©2009 by The Royal Society
Bromham L Biol. Lett. 2009;5:503-505
we now know that things are quite
different: information flow is
circular between genome and
environment
If the Central Dogma of Molecular Biology depicted
one direction-flow of genetic information………………
SYSTEMS
GENOMICS
(BIOLOGY)
REVERSE
TRANSCRIPTION
GRNetworks
TRANSPOSABLE ELEMENTS
NATURAL GENETIC ENGINEERING
TRANSCRIPTION
FACTORS
(and COFACTORS)
EPIGENETIC
CHANGES
The “Fluid
(Epi)Genome”
Shapiro JA. Revisiting the central dogma in the 21st century Ann N Y Acad Sci. (2009);1178:6-28
ENVIRONMENT
From directing the fate of stem cells to determining how.. we grow, the genes in our body
act in complex networks.. the whole Genome is a Complex and highly dynamic molecular
Network of interacting Genes and non-codifying sequences.. and proteins
….Genes Know How to Network…BUT...
IN FACT Genes need to be told to switch “off”
and “on”:
• Genes need to be told how much expression
(protein) is required and where.
• Genes need to be regulated – this regulation
is not performed by DNA but by many other
controls arranged in a complex network
• DNA has been called the Book of Life by the
Human Genome Project scientists, but many
other biologists consider DNA to be simply a
random collection of words from which a
meaningful story of life may be assembled…
• In order to assemble that meaningful story, a
living cell uses a second informational system.
(...) The key concept here is that these
dynamic-epigenetic networks have a life of
their own —they follow network-rules not
specified by DNA
Strohman R. , April 2001
Beyond genetic determinism
http://news.sciencemag.org/sciencenow/2009/04/21-03.html
..configuring a new model of genomic molecular network composed by DNA, epigenome, proteome
sequences communicating in cis and trans
and, above all, reactive /interactive (in a continuous dialogue with the environment)
VIRUSES
Hormons
Xenobiotics
EMF
Naesens, M. & Sarwal, M. M Current paradigm of molecular biology Nat. Rev. Nephrol. 6, 614–628 (2010)
CEM
We may represent the environment as a continuous
stream of information (simple: photons: individual
packages of E = M = Information) or complex (organic
molecules, viruses etc) interacting with our cells
[membrane or transmembrane receptors,
signal transduction proteins, nuclear receptors,
genome (DNA + Epigenome)] forcing them to adapt
TCDD
Viruses
1
3
“FLUID EPI-GENOME”
HERVs
4
SYNERGISM ?!
2
What’s Epi-Genetics ?
Rudolf Jaenisch- Whitehead Institute and Dept.
of Biology, MIT, Cambridge, MA
The
Histone tails
are a critical
determinant
of chromatin structure
The tails of
histones could be
regarded as the
sensory /
receptive
component
of the genome
Histone Tails are subject to
a variety of covalent
modifications
Histone Code”
hypothesis: modifications
of the Histone tails
act as marks read
by other proteins
to control the
expression
or replication of
chromosomal regions
E.g. generally,
Histone Acetylation
is associated with
transcriptionally
active genes
Deacetylation
is associated with
inactive genes
(= gene silencing)
DNA methylation
Covalent modification of the DNA is important for gene silencing human cells.
Most genes have GC rich areas of DNA in their promoter regions.
These are referred to as CpG islands.
Methylation of the C residues within the CpG islands leads to gene silencing
(highly unstable base)
The “meeting-point” between the information coming from the
environment and the information encoded in the DNA
(hardware) is the epigenome (software): mimetic molecules
(EDCs) and other pollutants or danger-signals induce the
epigenome to change
Histone Acetyltransferases;
Histone Methyltransferases
Histone Deacetylases.
Histone
Lysine
Acetylation
P
H3-K9
Nuclear Receptor
DNA Response Element
H3-S10
ATP-dependent Nucleosome
Remodeling Complex
Chromatin itself is the direct target of many toxicants *
… toxicant-induced perturbations in chromatin structure
may precipitate adverse effects.. Forcing genome to change
Many toxicants cause
rapid alterations in gene
expression by activating
protein kinase signaling
cascades.
The resulting rapid,
defensive alterations in
gene activity require the
transmission of a signal
directly to the histones
present in the chromatin
of stress response genes:
within minutes
of exposure
the phosphorylation of
serine 10 of histone H3
and the
acetylation
of lysines 9 and/or 14
take place
.. we have identified several classes of environmental chemicals that
modify epigenetic marks.. including
- metals (cadmium, arsenic, nickel, chromium, CH3-mercury),
- peroxisome proliferators (trichloroethylene,
dichloroacetic acid…),
- Air Pollutants (PM 0,1/2,5/10, black carbon, benzene),
- EDCs - Endocrine-Disrupting/reproductive toxicants
(DES, bisphenol A, persistent organic pollutants, dioxin).
Because these epigenetic changes are small, potentially
cumulative, and they may develop over time, it may be
difficult to establish the cause-effect relationships among
environmental factors, epigenetic changes, and diseases.
Polycyclic Aromatic
Hydrocarbons (PAH)
(Ultra)-fine
particles
Heavy Metals
Benzene
Dioxin and
Dioxin-like
molecules
2
3
….particular awareness has aroused a
study that documented the presence
1 of (geno) toxic and mutagenic
substances in all the umbilical
cords tested, demonstrating the
ubiquity of embryo-fetal exposure
Mothers Milk: Record levels of toxic fire
retardants found in American mothers'
breast milk. Washington, DC. Available at
http://www.ewg.org/reports/mothersmilk/
NAS (National Academy of Sciences). 2000.
Scientific Frontiers in Developmental
Toxicology and Risk Assessment.
Committee on Developmental Toxicology.
Washington, DC: National Academies Press;
EPA (U.S. Environmental Protection
Agency). 2003a. America's children and the
environment. Measures of contaminants,
body burdens, and illnesses. Available
online at
http://www.epa.gov/envirohealth/children.
Cellular Differentiation: an Epigentic process
Differentiation occurs numerous times
during the development of a
multicellular organism as the organism
changes from a single zygote to a
complex system of tissues and 200 cell
types (genetically identical.. each with
its own epigenetic and morphofunctional characteristics)..
1
Nature 447, 425-432 (24 May 2007)
Differentiation
Fetal Programming
Decreasing
Plasticity
2
Gametogenesis. Maturation of germ cells is characterized
by an impressive degree of cellular restructuring and gene
regulation that involves remarkable genomic reorganization.
These events are finely tuned, but are also susceptible to the
introduction of various types of “error”
epi-mutations
Epigenetic differences in homozygotic twins
… although twins
are epigenetically
indistinguishable
during the early
years of life,
… older monozygous
twins exhibited
remarkable
differences in their
overall content
and genomic
distribution of
5-methylcytosine DNA
and histone acetylation,
affecting their
gene-expression portrait.
3-year-old twins
Epigenetic differences arise during the
lifetime of monozygotic twins
50-year-old twins
Fraga et al., PNAS. Jul 26 (2005);102(30):10604-9..
Exposition Windows
XXI e siècle: transformation spectaculaire de
l'environnement et du microenvironnement utérine
The gift our mothers
never wanted to give us
XENOBIOTICS
HEAVY METALS
X rays
+ EMFs
http://www.ewg.org/reports/generations/
ENDOCRINE DISRUPTORS
The Environmental Toxic
Burden and the Developmental
Origins of Health and Diseases
Le fardeau de l'environnement toxique et les origines
développementales de la santé et des maladies
ULTRAFINE
PARTICLES
What is the Global Chemical Burden..
Industrial chemicals in mothers and daughters:
the pollution we share and inherit
Is it true that these pollutants are present in blood and tissues of all men and women living in urban
and industrial environments and even in the cord blood and placental and fetal tissues in more and
more significant amounts year after year ?
Is it true that metals, dioxins and other lipophilic pollutants, accumulated in maternal tissue, may pass,
even many years after their absorption, into the blood and reach the fetus?
Obesity/Metabolic Syndrome
Cardiovascular
Diseases
Obesogens
DOHAD
Multiorgan Effects of
Endocrine Disruptors
Pesticides
Ipertension
In Vitro Fertilization
Materno Fetal Stress
Placenta: Prediction
of Future Health
Developmental Time Windows of
Vulnerability
Reproductive
Diseases/Dysfunctions
Semen Abnormalities
Asthma and allergies
Lung Development
Neurobehavioral Deficits
and Diseases
CANCER
Psychiatric Diseases
Ce sont des quantités minimales de molécules (epi)génotoxiques, induisant
des transformations continuelles de la chromatine, qui constituent le véritable problème. C’est un
processus très lent pouvant démarrer lors des premières
étapes du développement fœtal. Et, même dans les gamètes.
Si les tissus du fœtus sont mal programmés au début et s’il y a un stress épigénétique progressif, les
mutations génétiques et chromosomiques vont davantage se produire
Cheryl Lyn Walker UT MD Anderson Cancer Center
(4) Tomatis legacy
Renzo Tomatis - for over 10 years Director of IARC, for nearly 20 years
Scientific Director of ISDE - had already warned us since 1979,
on the Journal of the National Cancer Institute Monograph,
• that the exposure of pregnant animals to chemical carcinogens can
induce tumours in the offspring;
• supporting his thesis with a copious scientific literature concerning
38 different (pro)carcinogenic agents;
• reminding us that, unfortunately, even then, there was
epidemiological evidence concerning human beings, since dozens
of "DES daughters" had developed an adenocarcinoma of the
vagina (moreover some studies on mice and rats exposed to
carcinogens during pregnancy had shown a high incidence of
tumours not only in the litter of first generation, but also in the
offspring of the second and third generation, providing a strong
evidence of a possible transgenerational transmission of cancer)
Tomatis L. Prenatal exposure to chemical carcinogens and its effect on
subsequent generations. Natl Cancer Inst Monogr. (1979); (51):159-84.
1
2
3
MOREOVER
DES Daughters, were
NOT ONLY at an
increased risk for clear
cell adenocarcinoma
(CCA) of the vagina
and cervix, and
breast cancer
(2.5 fold increase
in after age 40)
BUT ALSO of
reproductive tract
structural differences,
pregnancy
complications
infertility and autoimmune disorders…
MOREOVER
some effects of DES
could be
transgenerational
Diethylstilbestrol
J Pediatr Hematol Oncol 2003; 25:635-636
The DES-tragedy could contribute to clarify the
problem: endocrine-mimetic molecules and many
other pollutants may act by altering
the epigenetic fetal programming ..
.. DES exposure induces changes in the expression of several uterine genes
involved in tissue patterning, such as Wnt7a, Hoxa9, Hoxa10, and Hoxa11,
contributing to changes in tissue architecture and morphology …
Cancers in adults predominantly arise in (epithelial)
tissues chronically exposed to environmental stress
and in cells and tissues continually urged to respond/react to it
While almost all childhood cancers belong to three major groups:
45% oncohaematologic tumors (leukemias and lymphomas)
25% brain tumors
25% neoplastic degeneration of embryonal residuals
The increase particularly
affects children in their first
life year (the incidence rate
increased by> 2%)
(5) Infant Cancer
• As for cancers in infants we should point out that
at least the first stages of the malignant process
5
are already present at time of birth.
• The importance of genetic events in utero has
been suspected, for many years, on the basis of the
correlation studies on twins with leukemia.
• That some cases of leukemia originate in utero is also
the result of genetic studies that have found in blood
samples (Guthrie cards), taken from infants who
subsequently would develop leukemias,
translocations and gene sequences corresponding
to the fusion genes later found in leukemic blasts.
Greaves MF, Maia AT, Wiemels JL, Ford AM. Leukemia in twins: lessons in natural history Blood (2003) 1;102(7):232133; Mori H, Colman SM, Xiao Z, Ford AM, Healy LE, Donaldson C, et al. Chromosome translocations and covert
leukemic clones are generated during normal fetal development. Proc Natl Acad Sci U S A (2002);99:8242-7
Le processus de cancérogénèse est généralement très long…
Donc, le véritable problème est l’exposition des parents (gamètes) aux facteurs de risque.
Le processus de cancérogénèse a débuté bien avant la naissance de l’enfant *.
Prenatal origin of certain childhood leukaemias
On peut aussi dire que la phase d’initiation *
tumorale (1° Hit de Knudson) survient lors
du développement ontogénétique du fœtus
Il n’y a que deux possibilités:
1) l’exposition du fœtus à des agents physiques (X-rays), chimiques ou biologiques (virus) (transmis par
transmission trans-placentaire ) qui puissent endamager directement le foetus
2) la transmission trans-generationelle d’une ou plusieurs lesions genètiques ou epi-génetiques
Trasmissione
transgenerazionale
2
3
1
(epi)mutations
in gametes
Transgenerational
Carcinogenesis
The real question is:
are these
(epi) mutations
stochastic
or provoked by
environmentally
induced stress ?!?
(5b) Pro-leukemic translocations in foetuses
• Moreover, pro-leukemic translocations
are found in foetuses with a much higher
frequency than the incidence of leukemias:
• the common leukemic fusion genes:
- TEL-AML1 t (12; 21) (p12; q22) and
- AML1-ETO t (8; 21) (q22q22),
• are found in the cord blood with a
frequency 100 times greater
6
than the corresponding risk of leukemia
Greaves M. Pre-natal origins of childhood leukaemia Rev Clin Exp Hematol. (2003);7(3):233-45
.. the first unambiguous evidence for a
prenatal origin of leukaemia was derived
from studies in identical twins with
leukaemia. A case of identical
(monozygotic) infant twins with
leukaemia was recorded in 1882, and,
since that time, more than 70 pairs have
been published albeit in variable detail ...
1
2
Chromosomal translocations and preleukaemic clones arise at a
substantially higher frequency (~100 X) before birth than the cumulative
incidence or risk of disease, reflecting the requirement for complementary
and secondary genetic events that occur postnatally. A consequence of the
latter is a very variable and occasionally protracted postnatal latency of
disease (1—15 years).
3
The concordance rate of leukaemia varies
according to subtype and age.
For infants with ALL, the rate is
exceedingly high (> 50%), for
“COMMON” child-ALL, is ~10%.
Adult leukaemia (ALL/ AML),
in contrast, has a very low rate
of concordance (< 1%).
~1% of newborns had TEL-AML1 positive B lineage clones…
which represents 100 times the incidence of TEL-AML1 positive ALL (~1 in 12,000).
(5c) Pro-leukemic translocations as an active
potentially positive, adaptive genomic change
• This fact is usually interpreted as the evidence that
translocations do not necessarily determine the
onset of leukemia, which would require additional
genetic events during the postnatal period.
• An equally interesting interpretation consists in
assuming that if less than 1% of children who have
"produced" a translocation have developed leukemia,
it could be because the translocation is an active
7
potentially positive, adaptive genomic change,
which could be responsive to toxic exposures in utero
In such a context: should TRANSLOCATIONS
be considered as chromosomal aberrations
or as (re)active rearrangements ??
(6)… Pro-leukemic translocations as
active/adaptive genomic changes
It may be useful to remember something very
interesting occurring in areas with persisting
pollution, as in Seveso, where
decades after the accident which caused the
dioxin contamination of a wide territory,
many individuals have been showing a high
number of translocations 14:18 (typical of
follicular lymphoma), although without
developing lymphoma.
Baccarelli A, Hirt C, Pesatori AC, Consonni D, Patterson DG Jr, Bertazzi PA, Dölken G, Landi MT. t(14;18) translocations in
lymphocytes of healthy dioxin-exposed individuals from Seveso, Italy Carcinogenesis (2006); 27(10):2001-7
Exposure to NHL-associated carcinogens,
such as dioxin or pesticides, may cause
expansion of t(14;18)-positive clones.
6b)… pro-leukemic translocations as
active/adaptive genomic changes
It is important to note that the same
translocation is frequent in subjects long
exposed to pesticides for professional
reasons, which is a clear evidence that
it is not the single chemical agent to determine
the specificity of the mutation, but rather,
the active reaction of the genome itself.
Agopian J, Navarro JM, Gac AC, Lecluse Y, Briand M, Grenot P, Gauduchon P, Ruminy P, Lebailly P, Nadel B, Roulland S.
Agricultural pesticide exposure and the molecular connection to lymphomagenesis J Exp Med. (2009) 6;206(7):1473-83
Figure 2. t(14;18)+ cells in HI are actively transcribing BCL2 from the translocated allele
We can find exactly
the same (reactive)
translocation (++
expression of the antiapoptotic gene BCL-2)
in many subjects
chronically exposed
to pesticides ..
Agopian et al. Journal of Experimental Medicine 2009:206:1473-1483
IN THE CANCEROUS B CELLS, THE PORTION OF CHROMOSOME 18 CONTAINING THE BCL-2 LOCUS HAS UNDERGONE A
RECIPROCAL TRANSLOCATION WITH THE PORTION OF CHROMOSOME 14 CONTAINING THE ANTIBODY HEAVY CHAIN
LOCUS. THIS T(14;18) TRANSLOCATION PLACES THE BCL-2 GENE CLOSE TO THE HEAVY CHAIN GENE ENHANCER.
H Chain-enhancer is very active in B cells...
t(14;18)(q32;q21) (IgH/BCL2)
If we remember that in this, as in other pro-leukemic
or pro-lymphoma translocations, an oncogene
(BCL-2: an anti-apoptotic gene, physiologically
activated in memory lymphocytes populations and in
B lymphocytes "immortalized" by Epstein Barr virus)
is constantly activated
because of the juxtaposed enhancer sequence of the
gene coding for the immunoglobulin heavy chain,
the hypothesis that these translocations are genomic
8b
active, adaptive (potentially defensive) changes
rather than simple "chromosomal aberrations"
acquires a meaning and a greater value …
Kurtulus S, Tripathi P, Moreno-Fernandez ME, Sholl A, Katz JD, Grimes HL, Hildeman DA.Bcl-2 allows effector and
memory CD8+ T cells to tolerate higher expression of Bim J Immunol.(2011) 15;186(10):5729-37; Finke J, Fritzen R,
Ternes P, Trivedi P, Bross KJ, Lange W, Mertelsmann R, Dölken G.Expression of bcl-2 in Burkitt's lymphoma cell lines:
induction by latent Epstein-Barr virus genes Blood. (1992); 15;80 (2):459-69
(7) Infant leukemias
In conclusion, we must remember that
infant leukemias
– which, as mentioned, have registered the highest
increase and, according to this dynamic model,
may represent the most emblematic
consequence of a distorted fetal programming
(an “evolutionary process gone awry”) 10
show, even at a molecular level, some peculiar
characteristics
Soto AM, Maffini MV, Sonnenschein C. Neoplasia as development gone awry: the role of
endocrine disruptors. Int J Androl.(2008);31(2):288-93
(7b) MLL1
As a matter of fact it is significant that in these forms
the MLL1 gene
- a gene encoding a histone methyltransferase - an enzyme
playing a key role in the implementation of epigenetic
and chromatin modifications which are fundamental in the
early stages of fetal development and differentiation
of tissues (and especially in emo-lymphopoiesis) - is very often involved and implicated in dozens of different
translocations that express fusion proteins capable
of interfering with the differentiation of pluripotent
hematopoietic stem cells, dysregulating the expression
patterns of HOX genes
Ernst P, Wang J, Korsmeyer SJ. The role of MLL in hematopoiesis and leukaemia Curr Opin Hematol.(2002); 9(4):282-7;
Popovic R, Zeleznik-Le NJ. MLL: how complex does it get? J Cell Biochem. (2005) 15;95(2):234-42; De Braekeleer M,
Morel F, Le Bris MJ, Herry A, Douet-Guilbert N. The MLL gene and translocations involving chromosomal
band 11q23 in acute leukaemia Anticancer Res. (2005); 25(3B):1931-44
IN ALL AND AML, THE ALL1
(ALSO NAMED MLL) GENE
CAN FUSE WITH 1 OF MORE
THAN 50 GENES. ALL1 IS
PART OF A MULTIPROTEIN
COMPLEX. MOST OF THE
PROTEINS IN THE COMPLEX
ARE COMPONENTS OF
TRANSCRIPTION COMPLEXES;
OTHERS ARE INVOLVED IN
HISTONE METHYLATION AND
RNA PROCESSING. THE
ENTIRE COMPLEX REMODELS,
ACETYLATES, DEACETYLATES,
AND METHYLATES
NUCLEOSOMES AND
HISTONES. THE FUSION OF
ALL1 WITH 1 OF these 50
PROTEINS RESULTS IN THE
FORMATION OF THE
CHIMERIC PROTEINS THAT
UNDERLIE ALL AND AML.
ALL1 (MLL) FUSION PROTEINS
DEREGULATE HOMEOBOX
GENES (WHICH ENCODE
TRANSCRIPTIONS
FACTORS)..and microRNAs
GENES SUCH AS MIR191.
The first and most striking property of MLL fusion proteins is their incredible diversity.
been found in 73 different translocations and 54 partner genes have been cloned
(http://atlasgeneticsoncology.org/Genes/MLL.html).
Nakamura T, Mori T, Tada S, et al. ALL-1 is a histone methyltransferase th
assembles a supercomplex of proteins involved in transcriptional regulat
Cell 2002;10:1119-1128.
AF9 Location 9p22
MLL Location 11q23
Several lines of evidence point to a mishap in non-homologous end joining of double strand breaks as the most likely reason for 11q23 translocations.
histone
methyltransfe
Translocations typical of myeloid leukaemia, probably due to maternal exposure to some toxic compound, were shown to
be present at birth in children who developed the disease years later (while not sufficient per se to cause the disease, they might
increase the risk for leukaemia by inducing genomic instability) Tomatis L. Identification of carcinogenic agents and primary
prevention of cancer. Ann N Y Acad Sci. 2006 Sep;1076:1-14
Translocation involving band
11q23 in AML may occur as a
result of a deletion or translocations
with a number of other
chromosomes and is usually
associatedwith M4 or M5 and a
poor prognosis
!
Even if leukaemia fusion gene formation is
spontaneous, the risk of this occurring may be
modified by other factors, including folate
!!
availability. There is dietary and genetic
evidence that folate has an impact
on the risk of infant and childhood leukaemia ..
MLL rearranged leukemias are associated
with poor prognosis and very brief latency
for MLL-AF4+ infant B ALL. This raises the
question of how this disease can evolve so
quickly,
MLL1-gene reactive rearrangements
• Even the high frequency of MLL1-gene rearrangements in
leukemias and myelodysplastic syndromes secondary to
treatment with topoisomerase II inhibitors is significant
as an argument in favour of the prenatal and (epi)genotoxic origin of infant leukemia,
• due to maternal and fetal exposure to substances
capable of interfering with the action of this enzyme,
which is essential for the unwinding of the double helix,
such as bioflavonoids (contained in many foods) and
11
widely used insecticides such as dipyrone (Baygon)
Strick R, Strissel PL, Borgers S, Smith SL and Rowley JD: Dietary bioflavonoids induce cleavage in the MLL gene and may
contribute to infant leukemia. Proc Natl Acad Sci USA (2000) 97: 4790-4795 ; Alexander FE, Patheal SL, Biondi A, Brandalise
S, Cabrera ME, Chan LC, Chen Z, Cimino G, Cordoba JC, Gu LJ, Hussein H, Ishii E, Kamel AM, Labra S, Magalhães IQ, Mizutani
S, Petridou E, de Oliveira MP, Yuen P, Wiemels JL, Greaves MF. Transplacental chemical exposure and risk
of infant leukemia with MLL gene fusion Cancer Res. (2001) 15; 61(6):2542-6
Our study has supported the
hypothesis that in utero exposure to
chemicals causes MLL* infant
leukemia and has generated specific
hypotheses that require further
testing.
Exposure to dipyrone is widespread,
particularly in Central and South
America where it is available as an
inexpensive, nonprescription drug.
Mosquitocidals are similarly in general
use in these same settings. Propoxur
(Baygon°) is also widely used against
cockroaches, fleas, and similar pests.
Therefore, it is important that the
associations observed in this study
are reevaluated in an extended casecontrol study
This is the first demonstration that
transplacental benzene exposure can induce
hepatic and hematopoietic tumors
in mice, which may be dependent on
fetal benzene metabolism capability
REMARK
• Those who adhere to the paradigm of
stochastic mutations
• and more generally to a linear and genecentric model of DNA
•
have obviously some difficulty to
accept all this…
8
The dynamic (epi)-genome
In fact only in a model of genome conceived as
a unitary and complex (at the same time, dynamic
9
and responsive) molecular network,
it is possible to suggest that all the epigenetic (global
DNA hypomethylation, hyper-methylation of promoter
sequences of tumor suppressor genes..), genetic
(genomic instability, mobilization of transposable
sequences..), and chromosomal (translocations)
mutations, determining the progression of cancer,
should be seen as steps in a (failed or distorted)
evolutionary/adaptive and defensive process
Esteller M. Cancer epigenomics: DNA methylomes and histone-modification maps Nat Rev Genet (2007);8(4):286-98;
Karpinets TV, Foy BD. Tumorigenesis: the adaptation of mammalian cells to sustained stress environment by epigenetic
alterations and succeeding matched mutations. Carcinogenesis. (2005); 26(8):1323-34; Hauptmann S., Schmitt W.D.
Transposable elements - Is there a link between evolution and cancer? Medical Hypotheses (2006), 66 (3):580-591;
In pre-neoplastic lesions we find some common epigenetic changes, produced by prolonged stress: an hypo-methylation
of the whole DNA sequence, a hyper-methylation of the promoters of onco-suppressor genes, a global genomic instability
1
2
3
2
These mechanisms may be
activated in cells by
continuing proliferative
and survival signaling
in a sustained stress
environment (SSE)…
longterm exposure
to this signaling
epigenetically reprograms
the genome of some cells..
tags = marcature
Many epigenetic tags are
Induced in stem cells during
fetal programming
Mutations match epigenetic tags
Selection of Mutants follows !
Transposable elements can be seen as a natural genetic engineering system capable of acting
on the genome as a whole .. This dynamic view of the genome has been illustrated most
impressively by Shapiro who stated that the genome is composed of modular units arranged
in a “Lego-like” manner that can be altered under certain circumstances
Stressors, which cannot be compensated for with the
usual cell possibilities might arouse evolutionary
mechanisms intended to create new protein variants.
One of these is the activation of transposable elements
which leads to a reformatting of the genome.
2
3
The hypothesis developed
in this article is that chronic
stressors can induce
1
adaptive rearrangements
of the genome which might
result in new proteins
helping survival in a harmful
environment or trigger the
development of a
completely ‘‘new’’ organ –
the cancer – which also
survives – even though its
survival is at the expenses of
its host.
4
Retrosequences
activation…
Our study demonstrates
the strong link between
hypomethylation of
transposable elements
with genomic instability
in non-small cell lung
cancer and provides
early evidence for
a potential active role
of these elements
in lung neoplasia.
MOLECULAR WEIGHT OF DIFFERENT PROTEINS PRESENT IN ZEBRAFISH EMBRYO
DURING GASTRULATION PERIOD
..placing tumor cells into a "normal"
1) 37% molecular weight of about 97KDa
morphogenetic field - like that of an
2) 14.6% molecular weight of about 45 KDa
embryonic tissue - one can reverse malignant
3+4) 27.4% molecular weight of about 30-25 KDa phenotype, "reprogramming" tumor into
5) 4% molecular weight of about 20 KDa
normal cells.
6+7) 14% molecular weight of about 14 KDa
Biava M. et al Embryonic morphogenetic field induces phenotypic reversion in cancer cells.
Current Pharmaceutical Biotechnology Volume 12, Issue 2, 2011, Pages 243-253
Macroevolution
A
Symbiogenesis
Horizontal gene transfer
Neodarwinistic Paradigm
EMERGENT
Master
Genes
PROPERTIES
(microevolution)
GRNs
Environmental
crisis
NEW DEVELOPMENTAL PATTERNS
Genes
Natural Genetic
Engineering
Random Mutations
Mobile
Sequences
(Fluid) Epigenome
Reverese
Transcriptase
Phenotype
HATs
Natural Selection
B
(Environment)
HDACs
DMTs
NRs
(ecosystems)
(Fluid) Genome
Environment
Ribotype
Proteome
Phenotype
Organisms
Cells
Rs
microevolution
Rs
TFs
Molecules
Neo-Lamarckian-Constructive
Paradigm
Natural Selection
Rs: Receptors
Conclusions (a)
This was just a brief introduction to an extremely complex
and delicate issue, concerning the meaning to be given to
the increase in childhood cancers, in recent decades,
together with the (even more pronounced) increase and the
progressive anticipation of the age of the onset of many
chronic degenerative and inflammatory diseases.
It is not possible so far to prove with absolute certainty the
embryo-fetal (epigenetic) origins of these diseases
But decades after the first "prophetic" analysis made
by Renzo Tomatis
and the first explicit formulation of a theory about the foetal
origin of some cancers
D. Trichopoulos Hypothesis: does breast cancer originate in utero? Lancet (1990), 335, pp. 939–940; Sanderson ML et
al. Perinatal factors and risk of breast cancer Epidemiology (1996) 7, 34-37; Michels KB Birthweight as a risk factor for
breast cancer Lancet, (1996)348, 1542–1546; Vatten L Can prenatal factors influence future breast cancer risk? The
Lancet, (1996) 348, 9041, 1531
Conclusions (b)
… in a world increasingly characterized by ubiquitous distribution of
thousands of potentially pro-carcinogenic molecules in food
chains and even in the cord blood
• it is necessary to recall, once again, the great lesson of Tomatis,
who claimed that in order to reverse the trend of the continuous
increase in tumors there is an obligatory path to take, that of the
primary prevention id est a reduction in the overall chemical
burden, concerning
• not only those substances for which there is sufficient evidence in
experimental studies of chemical carcinogenesis,
• but also for the numerous "possible" carcinogens (IARC group 2B),
for many of which it is highly unlikely that any future research will 12
give a definitive answer about their carcinogenicity
Christiani DC Combating Environmental Causes of Cancer N Engl J Med. (2011) 3; 364(9):791-3
Tomatis L. Role of experimental and epidemiological evidence of carcinogenicity in the primary prevention of
cancer. Ann Ist Super Sanita. (2006);42(2):113-7; Tomatis L. Identification of carcinogenic agents and primary
prevention of cancer Ann NY Acad Sci. (2006);1076, 1-14
Fetal programming
Ontogenesis
3
5
The 7 keys
Devo-Evo
Mismatch
4
Developmental Plasticity
7
2
Environment
1
From Genetics
to Epigenetics
Phylogenesis
6
XX Century
Epidemiologic
Transition