Diagnosis and treatment of
autism considering features of
the genetic background and
metabolic status
Ukrainian Institute of Clinical Genetics of KNMU
Member-correspondent of NAMS of Ukraine,
M.D., professor
E.Y. Grechanina
1
 Genetic
bases of many human diseases
are successfully studied for last 20 years.
 Confession of World Health Organization
that the basis of somatic , psychic and
reproductive health is genomic health
contributed this success.
 In opinion of H.Y. Zoghbi et al., Beaudet
(2010) studying relationship between
genotype and phenotype gives challenge
for clinicists and researchers because
some observations can’t be explained so
easily.
2
GENE INTERACTION
Gene 1
ACGTAGCTAG
Nuclear DNA
Gene 2
Mitochondrial DNA
ACGTAGCTAG
Substitution of gene
fragment
Gene 2
ACGAGСCTAG
GENOMIC
HEALTH
EPIGENETIC
FACTORS
ENVIRONMENTAL
FACTORS 3
Genomic health = somatic, psychic,
reproductive health
4

The role of the epigenome
(changes of genetic information
without changes of DNA
nucleotides sequence) in normal
as well as in pathological
physiology of the genome.
5
 Many
researchers proved that the
causes of many inherent diseases
are epigenetic mutations, which
can change DNA methylation.
6
 By
Ellis’ data, relationship between human
genome and epigenome extended type
range of molecular events, which cause
human diseases.
 They can be de novo mutations or
inherited from previous generations,
genetic or epigenetic and can be a result
of the influence of environmental factors.
7
 The
appearance of convincing information
about that environmental factors (at the
first place – nutritional pattern) change
the epigenome (DNA methylation) gave us
better understanding the pathogenesis of
human multifactorial diseases and at the
first place – neurological disorders and
psychic diseases.
8
 At
the beginning of genetics creation, the
well known psychiatrist, professor
Bocherikov asked me, beginner genetician,
to prove that psychic disorders are
material.
 Very
long professional way led to this
understanding.
9
 We
have to solve problems of thousands of
children «…we have no alternative –we
have to look at these problems, try to find
out all details, to consider the point of view
of everybody, whom this regards and out
best for agreement achievement.The
necessity to achieve the success is the
another cause, where we need to use the
antagonism between different points of
view of the problem and the sooner the
better. Let (all) voices be heart in
discussion, let they try to come to
understanding, and don’t try to shout down
each other».
10
 Autism
becomes one of the global human
problem. It influences on many sides of
physical life and spiritual life. It requires
from us emergent development and
introduction of a new paradigm of
medicine 4 х «Р» -predictive, prognostic,
preventive, partner.
11
 Parents
of children with autism and
doctors become partners. The sooner this
partnership is achieved, the sooner this
problem will be solved.
 Parents – all day persons on duty for their
children,that’s why their information is
invaluable, although it sometimes require
medical correction.
 As soon as the resonance between
partners is established, the next autistic
child will begin to speak.
12
A
doctor, who received information from
analysis and phenotype assessment of a
patient, has to be at the head of the
triangle «child-parents-doctor» with all
responsibility in the process of search of
the truth.
 Considering this I allow myself to analyze
our way to understanding autism and
desire to help.
 Everybody who will hear us, will be heard
by us.
13
Autism – heterogeneous syndrome,
which is characterized by disorders in в 3
central domains (fr. Domaine- —area):
1.Social interaction
2.Speech
3.Interests
and expressed genetic and phenotypic
heterogeneity.
14
Autism – the most severe result of disorders of
nervous system development which is referred to
autistic spectrum disorders (ASD).
The frequency of incidence of ASD 37 in 10 000
Boys are prevalent, especially in clinically severe cases.
The frequency of autism 13 in 10 000.
Women/men ratio 4:1
(in severe forms 1:1)
The frequency of Asperger syndrome 2,6:10 000
Women/men ratio 8:1
15
The main feature of modern knowledge
about ASD – its uncertainty.
Many parallel approaches are necessary to
understand genetic factors which underlie
ASD:
1.Studies of the whole genome;
2.Associative studies;
3.Revealing mutation;
4.Expansion of clinical genetic examination of
probands and their relatives
16
Established genetic basis of autism:
1.Increase of the number of publications confirming that
mutation and structural changes in any of several genes
can significantly increase the risk of this disease.
2.If the diagnosis of autism is established in a child, the
risk for the family will be 25 tomes higher.
3.Cognitive behavior features, which are similar to those
observed in probands, are more likely observed in sibs and
parents of an ill child than in controls .
4.Independent studies of twins show concordance for
monozygotic twins 70-90 %, for dizygotic twins from 0 to
10%.
17
Molecular studies of genes
identified by nowadays show that
no one molecular explanation will
be enough.
18
 Many studie indicate system course of
disorders in ASD development.
 Different influence of maternal and
paternal 15q11 in ASD is an important
confirmation of cytogenetic disorders.
 It is supposed that different molecular
events are at the level of systems.
19
In recent years greater number of
associated with autism syndromes
are found (Tab. 1).
20
Table 1: Syndromes associated with ASD
№
Syndromes
Genes which
are
associated
with
syndromes
Proportions
of patients
with
syndromes
followed by
ASD
Proportions
of patients
with ASD,
who have
these
syndromes
1
15q dup
Angelman syndrome
UBI3A
(and other)
>40
1,2%
2
16p11 del
Gene is
unknown
high
-1%
3
22q del
SHANK3
high
-1%
4
Syndrome of cortical
dysplasia of focal
epilepsy
CNTNAP2
~ 70%
rare
5
Fragile Xchromosome
FMR1
25% of men;
6% of women
1-2%
21
Table 1 (continuation):
Syndromes associated with ASD
№
Syndromes
Genes
associated
with
syndromes
Proportions of
patients with
syndromes
followed by
ASD
Proportions
of patients
with ASD,
who have
these
syndromes
6
Hobart syndrome
GOUBIRT,
Many loci
25%
rare
7
Potocki-Lupski syndrome
Chromosomes
17 р 11
~ 90%
unknown
8
Smith-Lemli-Opitz
syndrome
DHSA7
50%
rare
9
Rett syndrome
MISP2
All individuals
who have Rett
syndrome
~ 0,5%
10
Timothy syndrome
SASNAIS
60-80%
unknown
11
Tuberous sclerosis
TSC1, TSC2
20%
~1%
22
•Gene
polymorphism – a genetic
event in which building of genes
changes and this influence on
protein function.
If only one letter changes in
genetic code, this is called single
nucleotide polymorphism.
23
Genetic enzymatic polymorphism of homocysteine metabolism
(G.R. Akopyan)
Name of enzyme
GENE
COENZY
ME
MUTATIONS
Methyltetra-hydrofolate
reductase
MTHFR
Vit.B9
Vit.B2
C677T
(Ala to Val)
A1298C
DISEASES
Thromboembolia
Neural tube defects
Diabetes mellitus
(Asp to Gly)
Vit.B12
Methionine
synthase
MTR
Methionine synthase
reductase
MTRR
Сystathionine-βsynthase
CBS
Cystathionine-γ-lyase
CSE / CBL
Congenital
cystathionineuria
Methionine
adenosyltransferase
MAT I / III
Hypermethioninemia
Glycine Nmethyltransferase
deficiency
GNMT
Liver pathology
S-Adenosyl-homocysteine
hydrolase
SAHH
Psychomotor development
delay, neurological
abnormalities, hepatitis,
myopathy
A2756G
Thromboembolia
Colorectal cancer
Malignant lymphoma
A66G
Vit.B6
Ile to Thr
Gly to Ser
Cardiovascular
Homocystinuria
24
Polymorphisms of genes of folate and methionine cycle
 Hyperhomocysteinemia was found in every third from examined
patients with IHD and preeclampsia
 Genotypes with high predisposition to homocysteine-associated
thrombophilia in the case of their assessment by four
polymorphic loci MTHFR С677T_A1298C / MTR 2756 AG /
MTRR 66 AG: CT_AA/AA/GG, CT_AC/AA/GG , CС_AA/AA/GG ,
CT_AC/AA/AA , CС_AC/GG/GG, CC_AC/AA/AG, CC_AA/AA/AG
were found.
 The risk of hyperhomocysteinemia is likely associated with AA
carrier of MTR genotype and GG of MTRR genotype
 Carriers of four and more mutant alleles (MTHFR, MTR, MTRR)
need screening for homocysteine content in blood plasma.
 There is necessity of hyperhomocysteinemia verification using
methionine loading test.
25
 All
biochemical processes in a cell are
performed with the help of cycles, among
them there is folate cycle, which achieved
key positions: folate metabolism is the
basis of cellular metabolism (G.R.
Akopyan)
26
 The
-
-
following events are performed in this
cycle:
Synthesis of nucleic acids;
Synthesis of biologically active substances:
adrenaline, melatonin, creatinine,
phospholipids, polyamines (spermicides
and spermines), glutamic acid, dihydrotetrahydrobiopterin, nitric oxide ;
Epigenetic changes of DNA, DNA
(methylation), RNA, chromatin, amino
acids, proteins, lipids.
27
We supposed and confirmed that if there is
enzymatic activity of folate cycle in human
body – methylentetrahydrofolatereductase is
low, this leads to methylation disorder
(switching on and off gene activity) and then
it causes a lot of inherent and multifactorial
syndromes.
•
28
The following was underlain the basis of
this study: creation of single information
database including all levels of prevention
of inherent pathology at all levels of
ontogenesis.
29
Association of
families with
chromosomal
pathology
PRENATAL
CENTRE
Centre of
prenatal
education
ONCOGENETIC
CENTRE
REGIONAL
PULMONARY CENTRE
KHARKIV SPECIALIZED MEDICAL
GENETIC CENTRE
(practical basis)
Association of families
with cystic fibrosis
UKRAINIAN INSTITUTE OF CLINICAL
GENETICS , DEPARTMENT OF MEDICAL
GENETICS OF KhNMU
(scientific basis)
Association of families with
spinal muscle atrophy
Association of families
with phenylketonuria
REGIONAL
METABOLIC CENTRE
Service of urgent
biochemical
diagnosis
Centre of
studying
epigenetic
diseases
Centre of
connective tissue
pathology
Association of families
with cystic fibrosis
Association of families
with organic acidurias
Association of families
with mitochondrial
diseases
30
Comparative characteristics of IDD by screening data on
Kharkiv region for 2000-2008
2000
Diseases
2001
A
b.
a
m
o
u
nt
2002
In.
val
ue
A
b.
a
m
o
u
nt
2003
In.
val
ue
A
b.
a
m
o
u
nt
2004
2005
In.
val
ue
A
b.
a
m
o
u
nt
In.
val
ue
Ab.
am
oun
t
2006
2007
In.
val
ue
A
b.
a
m
o
u
nt
In.
val
ue
Ab.
am
oun
t
2008
In.
value
A
b.
a
m
o
u
nt
In.
value
Ab.
amo
unt
In.
val
ue
1.Anencephaly
2
0,9
9
3
1,5
3
3
1,4
5
3
1,3
8
1
0,4
4
2
0,9
0
1
0,4
1
1
0,39
1
0,36
2.Myelocele
7
3,4
9
5
2,5
5
6
2,9
0
9
4,1
6
1
0
4,4
0
11
4,7
5
5
2,0
7
12
4,72
8
2,9
3.Meningocele
-
-
1
0,5
3
1
0,4
8
1
0,4
6
-
-
-
-
-
-
3
1,18
2
0,72
4.Hydrocephaly
3
1,4
9
9
4,5
9
1
4
6,7
8
1
0
4,6
3
1
3
5,7
2
7
3,5
6
1
3
5,3
8
12
4,72
8
2,9
5.Microcephaly
5
2,4
9
3
1,5
3
-
-
4
1,8
5
5
2,2
0
5
2.9
7
9
3,7
2
5
1,96
5
1,8
6.Anotia
-
-
-
-
-
-
2
0,9
2
1
0,4
4
2
0,9
0
4
1,6
4
5
1,96
3
1,09
7.Anophthalmia
1
0,4
9
-
-
1
0,4
8
-
-
-
-
-
-
-
-
1
0,39
-
-
8.Microphthalmia
2
0,9
9
1
0,5
3
1
0,4
8
1
0,4
4
4
1,7
9
4
1,6
4
31
-
-
1
0,39
2
0,72
The frequency of genotypes and alleles of polymorphic gene
variants C677T MTHFR И A66G MTRR (n=4586)
Polymorphisms
С677Т
MTHFR
А66G
MTRR
Population selection
n=200, %
Patient selection
n=4586,%
Expected
genotype
frequency, %
СТ
40.7
1994/43,48
1926.12/ 42
ТТ
7.04
416/9,07
412.74/ 9
СС
52.26
2175/47,42
2247.14/ 49
Т
27.39
30.81
AG
43.0
2015/43,93
2248,05/ 49
GG
35.5
1615/35,21
1490,0/ 32,5
AA
21.5
955/20,82
847,95/ 18,5
G
57.0
57.18
Genotypes and
alleles
AG
334/34,61
341,8/ 35,4
GG
55/5,69
51,0/ 5,3
AA
552/57,20
572,2/ 59,3
G
23.00
32
Since 2008 we have been conducting the
stage of the scientific search according to
the following hypothesis.
Hypothesis:The influence of mtDNA
polymorphisms on MTChD is a result of
pathological transformation of mtDNA
polymorphisms against the background of
the changed status of methylation as the
main genome modificator and the
presence of triggers.
33
DNA methylation
Cytosine
Guanine
34
Methylation of biologically active substances
35
Folate and methionine cycle
All reactions of methionine cycle are connected
with tanssulfuration of homocysteine
MAT I/III
(B12)
!BHMT
GNMT
SAHH
(B6)
(B2)
Betaine is a donor of methyl groups in the reaction of remethylation of
homocysteine in participation of betaine-homocysteine-methyltransferase.
(G.R. Akopyan )
CBL (B6)
36
Hyperhomocysteinemia and methylation disorder
(G.R. Akopyan)
38
Homocysteine – a strong oxidant and protein modificator
Homocysteine
thiolactone
Decreases the level and
activity of thioredoxin,
superoxide dismutase,
syntase NO
Increases
NAD(P)H-oxidase activity
О2
Acts in the normal
level of
homocysteine and
in less
concentrations (10
nmol/l) !!!
LDL oxidation
Endothelium damage +
thrombogenesis
Atheromatous plague formation
Protein modification
Homocysteine thiolactonase or paraoxonase (PON 1)
can hydrolyze homocysteine thiolactone!!!
39
 Methylation
has been admitted the main
genome modificator, central pathway of all
metabolic events in organism life
 Optimization of methylation function in A.
Yasko’s opinion (2010) becomes a model
for management of genetic polymorphism,
which influences on many important
biological events in the body.
41
METHYLATION FUNCTION:
1. DNA methylation is necessary for support
of differential expression of paternal and
maternal gene copy susceptible to the
genome imprinting .
2. For stable gene silencing on inactive Xchromosome.
42
3. Stable transcriptional repression of provirus genomes
and endogen retrotransposons depends on DNA
methylation
 DNA methylation takes part in management and support
tissue-specific patterns of gene expression in
development
 Absence of DNA methylation decreases reliability of
support of chromosomes number that leads to
chromosomal aberrations
 DNA hypomethylation in consequence of influence of
DNA-methyltransferase inhibitors leads to elimination of
tumors
 Formation of other types of tumors increases in DNA
hypomethylation
43
Entirety of methylation systems determines
genome and it means psychic, physic,
reproductive health.
Studies, which explain how environmental
factors can induce epigenetic changes and
biologic effects, have appeared.
En Li, Adrian Bira (2010)
44
DNA methylation and chromosomal
instability
 Ehrlich, 2003; Dobge et al, 2005 established that
DNMT3B mutations in patients with ICF
syndrome or Dnmt 3b inactivation in mice lead to
various chromosomal aberrations (structural and
quantitative)
 There is the hypothesis that DNA methylation
contributes exact chromosomal disjunction and
in its absence more often there is leading to
chromosomal disorders nondisjunction
(hypomethylation, demethylation)
45
Alternative possibility means that DNA
methylation can inhibit expression
and recombination of
retrotransposons in animals’ genome,
thus defending chromosomes from
harmful recombinations.
46
Identification of folate cycle disorders
include:
1. Inherent malabsorption of folic acid
caused by mutations in the gene which
encodes folic acid transporter.
2. Deficiency of formiminotransferase
caused by the mutation in FTCD gene.
3. Deficiency of methylentetrahydrofolate
reductase caused by the mutation in
MTHFR gene.
47
4-5. Deficiency of functional methionine synthase
as a result of mutations in MTR gene affecting
methionine synthase (cblG) or mutations
affecting methionine synthase reductase (cblE
due to the mutation in MTRР gene).
6. Cerebral deficiency of folic acid caused by
mutations in FOLR1 gene.
7. Deficiency of thrifunctional enzyme containing
methylenetetragydrofolate cyclohydrogenase
and formyltetrahydrofolate synthase caused by
mutations in MTHFD1 gene (Mac Gill, Rosenblatt
et al.).
48
It is necessary to note that homozygotous
pattern of the polymorphism means more
expressed level of enzymatic activity decrease.
If a human is a carrier of a specific mutation, it not
always means that function activity certainly will
decease, SNP are indicators of potential problem
areas which can manifest independently or
under the influence of triggers or gene
interaction

49
 Defects
in 5-methyltetrahydrofolat
homocysteine methyltransferase can
disturb detoxification process, meanwhile
toxic substances, for example, mercurous
can worse the effect because of decreased
activity methionione synthase (MTR) and
decreased detoxification effectiveness
50
There is summary of the genes that are included in a complex panel of
methylation analysis (Amy Yasko, 2010)
Mutations
or
single
nucleotide
polymorphisms:
Gene mutations - changes affecting the sequence of a single gene.
Mutations vary in size from one affecting base pair to large
segments of chromosomes. Single nucleotide polymorphisms are
small genetic changes or variations that may occur in the DNA
sequence. The genetic code is denoted by 4 "letters": A, C, G and T.
SNP variation is due to the replacement of one nucleotide for
another.
The presence of mutations in genes encoding enzymes affects their
productivity. Homozygous mutations are those mutations that affect
both copies of the gene, heterozygous mutations are those
mutations that affect only one of the copies of the gene. Each of us
has two copies of each gene obtained from each parent. Some
mutations enhance the activity of enzymes (such as CBS) while
others may decrease the activity (such as MTHFR 677 1298 COMT)
51
COMT V158M, H62H, 61
The main function of this gene is involved in the breakdown of dopamine.
Dopamine - a neurotransmitter that is involved in the formation of
behavioral reactions and attention. Dopamine contributes to the
appearance of good feeling, because it causes a feeling of pleasure
influencing the processes of motivation and learning. Dopamine is
produced during positive thinking. COMT exposed cleavage leads to
the formation of another neurotransmitter - norepinephrine. The
correspondence between the level of epinephrine and dopamine levels
is involved in ADD / ADHD; dopamine levels is important in the
development of diseases such as Parkinson's disease. COMT is also
involved in transformation of the corresponding estrogen in the body.
COMT activity is often associated with sensitivity to pain. COMT
homozygotes may be more sensitive to pain.
52
VDR/Taq and VDR/Fok (vitamin D receptor)
The panel contains some receptors of vitamin D, Taq and Fok sites. While
Fok change was due to the regulation of blood sugar, modified Taq
may affect the level of dopamine. For this reason it is important to
watch the status of COMT VDR / Taq and draw conclusions based on
the totality of the results of these two sites. Focus on changes part of
the VDR in the Fok against supplements that support the pancreas
and assist in the maintenance of blood sugar in the normal healthy
range.
53
MAO A R297R (monamine oxidase A):
Mao is involved in the cleavage of serotonin in the body. Like dopamine,
serotonin - neurotransmitter. It is associated with mood, an imbalance of
serotonin levels is associated with depression, aggression, anxiety and
OCD behavior. MAO A is localized on chromosome X and is considered
X-linked trait that does not appear in men. Because the X chromosome in
a man can come only from the mother, it means that Mao mutations of
father (or their absence) plays no role in the son. For women, as one
chromosome is inherited from each parent, geneticians, tend to reflect
the
status
of
Mao
in
both
parents.
54
ACAT 102 (acetyl coenzyme A acetyltransferase):
ACAT plays a role in lipid metabolism, helps to prevent the accumulation of
excess cholesterol in certain parts of a cell in the body. ACAT is also
involved in the production of energy in the body. Contributes to the
breakdown of proteins, fats and carbohydrates from food, energy and then
will be used in life. Furthermore, the absence of ACAT may also lead to
depletion of B12, which is required in methylation cycle.
ACE (angiotensin converting enzyme): Considered for all - No longer testing
Various factors, including diet can affect the activity of ACE gene, changes
which can lead to high blood pressure. The connection between gene
activity disorders with increased anxiety, memory loss and learning
decrease has been revealed in animal studies. Increased activity of ACE
may also lead to the removal of minerals in the body by decreasing
excretion of sodium in urine and increased elimination of potassium. This
reaction is also associated with stress in a situation of chronic stress can
lead to additional sodium accumulation and an increase excretion of
potassium. This excess of potassium is in body if the kidneys function
properly. In the case of impaired renal function, it may result in retention of
potassium in the body.
55
MTHFR A1298C, C677T, 3 (methylenetetrahydrofolate reductase):
MTHFR gene product is at a critical point in the methylation cycle.
Participates in the normalization of the level of homocysteine. Some
mutations in MTHFR were well characterized, and are associated with
the risk of cardiovascular diseases and cancer and may play a role in
the level of neurotransmitters of serotonin and dopamine
MTR A2756G/MTRR A66G, H595Y, K350A, R415T, S257T,
(methionine synthase/ methionine synthase reductase):
11
These two gene product work together and are involved in the conversion
of homocysteine to methionine. Elevated homocysteine levels are risk
factors in a number of pathologies including heart disease, Alzheimer's
disease, etc. As in the case of COMT and VDR / Taq, MTR and MTRR
should be studied together. Mutations in the MTR can increase the
activity of the gene product in a way that leads to greater consumability
of B12 as the enzyme. On the other hand, recent publications show that
MTRR A66G mutation reduces the activity of the enzyme. Regardless of
which theory is correct breaking B12 cycle or methylation function
activity disorder at this point, B12 is used as an additive in all the cases.
56
BHMT 1,2,4,8 (betaine homocysteine methyltransferase):
The product of this gene is central in the short pathway of methylation,
performs remethylation of homocysteine to methionine. This gene
product activity may influence on stress, cortisol level and may play
the role in the ADD / ADHD, affecting the levels of norepinephrine.
AHCY 1,2,19 (S adenosylhomocysteine hydrolase):
Different mutations in the AHCY can affect the levels of homocysteine
and ammonia in the body.
CBS C699T, A360A, N212N (cystathionine-beta-synthase):
CBS enzyme basically acts as a gateway between homocysteine and
lower part of the path which generates ammonia in the body. There
are some positive end-products that are generated by the lower part
of methylation pathway, such as glutathione and taurine, there are
also negative side-products such as ammonia and excess sulfite.
Because of increased activity of CBS, sulfites that are toxic for the
body present an additional upload for SUOX gene product.
57
SHMT C1420T (serine hydroxymethyltransferase):
The product of this gene is involved in the setting blocks needed for
synthesis of new DNA and transformation of homocysteine to
methionine. While DNA blocks are important, mutations that affect
the ability to regulate the gene product and thereby affecting the
methylation process can cause the accumulation of homocysteine
and imbalance in the other intermediate compounds in the body.
NOS D298E (nitric oxide synthase):
NOS enzyme plays an important role in the detoxification of ammonia in
the urea cycle. Individuals who are homozygotous for NOS have the
enzyme with decreased activity. NOS mutations can affect the
regulation of CBS until increase of ammonia, which is generated by
CBS.
58
SUOX S370S (sulfite oxidase):
The product of this gene promotes detoxification of sulfites in the body.
Sulfites are generated as a natural by-product of the methylation
cycle, and enter the body with food. Sulfites, sulfur-based
preservatives that are used to prevent or reduce discoloration of
light colored fruits and vegetables to prevent the appearance of
black spots on the shrimps and lobsters, inhibit the growth of
microorganisms in fermented foods (e.g., wine), and are able to
maintain the activity of certain medications. Sulfites may also be
used for bleaching edible starch, rust and scale prevention in boilers
used for steam cooking food, and even in the production of
cellophane, for packing food products. FDA considers that one from
hundred sulfites is sensitive, approximately 5% of individuals suffer
from asthma. A person can face the problem of sulfite sensitivity at
any time of life.
59
Many cases of sulfite sensitivity have been registered, and therefore
the FDA requires that the labels have information about product
content of these substances. Scientists don’t note the smallest
concentration of sulfites needed to cause a reaction. Shortness of
breath is the most common symptom. Sulfites release sulfur dioxide
gas, which can cause irritation in the lungs and cause severe
asthma attack for those who suffer from asthma. Sulfites can cause
chest tightness, nausea, urticaria, and in rare cases, more severe
allergic reactions. Mutations in SUOX may be a risk factor for
developing certain types of cancer, including leukemia.
08.04.2015
EPIGENETICS
 Epigenetics (επί-over) – the section of medical
biological science studying principles of
changes of gene expressions or cell phenotype
caused by mechanisms not associated with
DNA sequence.
 Epigenetics characterizes the process of body
and environment interaction in genotype
formation.
К. Uodington 1947
61
61
Factors which lead to switching on
epigenesis:
- Nutrition;
- Infection;
- Smoking;
- Stress;
- Trauma;
- Operation;
- Alcohol.
62
 Triggers
(provocateurs) of switching on
epigenesis: nutrition, infection, smoking,
stress, alcohol
 The presence of gene predispositions
(mediators)
 Methylation
is the main epigenetic mark
and key reaction of epigenesis.
63
 Cryshtof

Bokk generalized scientific facts
about epigenetic regulation occurrence
and how it influence on human diseases.
T. Kouzarides thinks that such epigenetic
mechanisms how DNA methylation and
histone modifications (acetylation),
regulate gene expression by DNA
modulation in cellular nuclei.
64
 Such
environmental factors as nutrition
and stress influence are able to cause
changes of epigenetic status (Yeijmans
B.T. et al., 2007).
 These circumstances form opinion of many
scientists about that human epigenome
can be considered as the biochemical
record of life events, accumulated changes
throughout life.
65
Effects of epigenetics
 Genome imprinting (and its disorders)
 Cellular differentiation
 Transgenerative epigenetic effects
 Mutation process
 Blastemas
 Organism ageing
 Conservatism of genetic information
66
66
Mechanisms of epigenetics
DNA methylation
Chromatine remodeling
RNA-mediated modifications
Protein preionization
X-chromosome inactivation
67
67
 It
is established that many epigenetic
changes may not followed by phenotypic
changes, meanwhile some changes
caused by environment factor action
modulate gene activity (expression) (Herst
M, Marra M.A.,2009; Feinberg A.P., 2007;
Bijornson H.T., 2004) that’s why abnormal
epigenetic status can be associated with a
number of diseases (e.g. rheumatoid
arthritis, SLE).
68
 It
is shown that neural activity in the brain
is regulated epigenetically, and potential
relevance of epigenetic changes in
schizophrenia, biopolar disorders and
alcoholism allow us to see the problem in
a different way (Esteller M., 2007; Jones
P.H, Baylin S.B., 2007; Feinberg A.P. et al.
2006).
69
EPIGENETIC DISEASES INCLUDE
(HUDS Y.ZOGHBI, ARTHUR L. BEANDET):
1.Genome imprinting disorder.
1.1. Sister syndromes; Prader-Willi syndrome.
1.2. Beckwith-Wiedemann syndrome
1.3. Silver-Russell syndrome
1.4. Pseudohypoparathyroidism.
2. Disorders influencing on chromatin structure in transconfiguration:
2.1. Rubinshtein-Taybi syndrome
2.2. Rett syndrome
2.3. X-linked ά-thalassemia followed by mental delay. The syndrome of
immunodeficiency, instability of the centromeric region and facial
anomalies
2.4. Spondyloepiphyseal dysplasia of Schimke.
2.5. Methylenetetrahydrofolate reductase deficiency.
3. Disorders influencing on chromatin structure in cis-configuration.
3.1. άδβ-δβ-thalassemia
3.2. X-fragile syndrome
3.3. Facioscapulohumeral muscular dysrtrophy
70
Classification of epigenetic human diseases
(S.А. Nazarenko, 2004)
Epigenetic status disorder of
separate regions of the genome
(locate effect)
Disorder of epigenetic status of
the whole genome (global effect)
1. Diseases caused by inherited
mutations disturbing monoallele gene
expression – diseases of genome
imprinting (Beckwith-Wiedemann
syndrome, Prader-Willi syndrome,
Engelman syndrome)
1. Diseases caused by inherited
mutations of genes, products of which
are involved in the support of DNA
methylation level or modification of
methionine structure - ICF syndrome,
Rett syndrome, ATR-X syndrome,
Rubinshtein-Taybi syndrome, CoffinLowry syndrome
2. Diseases caused by methylation
status disorder of separate genes in
the result of de novo mutations in
somatic cells - a)cancer areas
connected with imprinting loss leading
to inactive gene activation or inhibition
of active gene expression;
b)cancer diseases caused by
hypermethylation of promoters of
tumor suppressor gene
2. Diseases caused by global disorders
of genome methylation in the result of
de novo mutations in somatic cells –
cancer disease connected with the
global genome hypomethylation
leading to activation of oncogenes,
retrotransposons and chromosomal
instability
71
Methionine






Methionine – an essential amino acid, is contained in proteins.
Is methyl group donor (in composition of S-adenosylmethionine) in synthesis of choline, adrenaline and other;
Source of sulfur in cysteine synthesis.
Has 52 biochemical synonyms.
Chemical name of methionine – (2S)-2-amino-4-methylsulfanylbutanoic acid.
Chemical formula- C5H11NO2S.
72
Unique functions of methionine

Takes part in trasmethylation reactions;

Is a donor of methyl groups;

In synthesis of biologically active substances;

Takes part in synthesis of nucleic acids;

Is an acceptor of methyl for 5-methylenehydrofolatehomocysteine methyltransferase (methionine
syntase).
73
Metabolism
74
74
 Methionine
– cysteine precursor
which gives it sulfur.
 Has
52 biochemical synonyms.
 Chemical
name of methionine – (2S)2-amino-4-methylsulfanyl-butanoic
acid.
 Chemical
formula - C5H11NO2S.
75
Biological function of methionine







An essential acid
A component of aminoacyl tRNA biosyntase
A component of glycine metabolism, serine and
trianine
A component of histidine metabolism
A component of methionine metabolism
A component of selenium amino acid
metabolism
A component of tyrosine metabolism
76
Enzymes of methionine metabolism are presented
by









Methionine syntase
Thyrosine amino transferase
S-adenosyl methionine synthetase isoform II type
Arsenit methyltransferase
Indomethylamine N-methyltransferase
S-adenosyl methionine synthetase isoform I type
Betaine homocysteine S-methyltransferase I.
Methionine-tRNA synthetase, cytoplasmic
Methionine adenosyltranferase II beta
77
Disorder of processes remethylation
(formation of methionine from
homocysteine), in the result of
deficiency of MTHFR и MTRR
enzymes leads to a number of
pathological conditions such as:
 atherosclerosis;
 atherothrombosis;
 Neural tube closure defect;
 infarcts;
 Chromosome disjunction defect in
oogenesis and the risk of birth of
children with Down syndrome.
78
Folate and methionine cycles
79
Classification (N. Blau et al., 1996)
Disorder —
Affected component
Tissue distribution
10.1 Methionine
adenosyltransferase (МАТ) of
the liver
Liver
10.2 Cystathionine betasynthase (CBS)
Liver, brain, lymphoblasts,
cultured fibroblasts,
amniocytes and choroidal
fibers
10.3 Gammacystatathionase(СТН)
Liver, lymphoblasts
10.4 Sulfitoxidase, isolated or
Liver, kidneys, lungs, heart,
lymphoblasts, choroidal
fibers, cultured fibroblasts
and amniocytes
molybdenum cofactor
10.4.1. Type А
10.4.2 Type В
Chromoso
me
localisatio
n
№
MIM
250850
21q22.3
236200
16
219500
272300
80
252150
Classification (N. Blau et al., 1996)
Disorder —
Affected component
10.5 5,10Methylenetetrahydrofolate
reductase (MTHFR)
10.6 Methionine synthase
(methyl cobalamin) cblE, cblG
10.7 Methylmalonyl-СоА-mutase
(adenosylcobalamine) and
methionine synthase
methylcobalamin)
Tissue distribution
Chromoso
me
localizatio
n
№
MIM
Liver, lymphocytes,
lymphoblasts, choroidal fibers,
cultured fibroblasts
1р36.3
236250
Liver, cultured
fibroblasts,
amniocytes
Liver, cultured fibroblasts,
amniocytes
236270
250940
277400
277410
277380
81
 Depending
on the frequency, separate
genotypes can compose bases for
development of common pathology,
other can be factors of development
of rare (orphan) diseases.
82
Spectrum of nosologies in combination of polymorphisms
С677Т MTHFR / А66G MTRR in patient selection (n=1938)
С677Т MTHFR / А66G MTRR
Nosology spectrum
Hmzg
Htzg/ N/
/Hmz Hmzg Htzg/ Hmz Hmz N/ Hmzg Htzg
/Htzg Htzg g
/N
g
g Htzg /N
Deficiency of folate cycle enzymes
(16%)
6
11
36
37
69
99
5
36
Deficiency of folate cycle enzymes
3
3
11
13
37
51
1
15
1
1
3
3
1
2
8
19
2
8
Homocysteine remethylation
disorder
Spina bifida
HCU
1
3
7
HCU (in relatives)
Thromboembolias/thrombosis
10
1
1
4
2
3
7
Thromboembolias/thrombosis
(in relatives )
1
3
1
2
1
Varicose vein dilatation
1
6
6
11
11
Varicose vein dilatation (in
relatives )
1
1
6
1
5
83
1
2
4
3
4
4
1
2
Spectrum of nosologies in combination of polymorphisms
С677Т MTHFR / А66G MTRR in patient selection (n=1938)
С677Т MTHFR / А66G MTRR
Nosology spectrum
IMD (7.8%)
Hmzg/H Hmzg/H Htzg/H Htzg/H
N/
mzg
tzg
tzg
mzg
Hmzg
9
IMD
71
64
71
124
10
44
2
10
11
13
23
4
13
2
2
1
3
2
1
IMD of sulfur-containing amino acids
4
1
2
1
2
2
8
5
9
17
IMD of sulfur-containing amino acids (in
relatives)
2
IMD of fatty acids
IMD of methionine
CTD
1
Htzg/
Hmzg/N
N
10
IMD (in relatives)
IMD of amino acids
N/
Htzg
1
3
1
1
3
1
2
4
2
7
8
8
3
1
3
21
19
13
33
2
1
1
3
11
CTD
(in relatives)
DMD
Disorder of tryptophan metabolism
1
1
84
Disaccharidose deficiency
1
Spectrum of nosologies in combination of polymorphisms
С677Т MTHFR / А66G MTRR in patient selection (n=1938)
С677Т MTHFR / А66G MTRR
Nosology spectrum
Hmzg
N/ Hmzg Htzg
/Hmz Hmzg Htzg/ Htzg/ N/
/Htzg Htzg Hmzg Hmzg Htzg /N
/N
g
2
Sulfite oxidase deficiency
1
3
1
1
2
4
10
2
19
1
1
Metabolism disorder in urea cycle
Maple syrup disease
2
1
Hyperprolinemia
Aminoacidemia
2
1
Aciduria
Hypothyrodism
Autism
Mitochondrial diseases
1
1
8
9
1
1
7
Mitochondrial diseases
(in relatives)
1
Kearns-Sayre syndrome
Epilepsy
1
1
3
1
1
5
10
1
85
2
Spectrum of nosologies in combination of polymorphisms С677Т MTHFR / А66G MTRR in
patient selection (n=1938)
Nosology spectrum
Vascular pathology(8%)
С677Т MTHFR / А66G MTRR
Hmzg/H Hmzg/H Htzg/H Htzg/H
N/
mzg
tzg
tzg
mzg Hmzg
2
5
13
13
N/
Htzg
Hmzg/N
Htzg/
N
46
43
2
22
5
Inborn heart defect
2
1
Inborn heart defect
(in relatives)
1
3
Cardiopathy
1
Myocardial infarction
1
1
1
2
2
3
Myocardial infarction (in relatives)
1
1
Ischemic heart disease
Ischemic heart disease (in relatives)
Vascular pathology
1
1
Insults
2
2
1
3
4
2
1
1
Vascular pathology (in relatives)
Insults (in relatives)
1
1
1
1
1
1
11
8
8
9
6
2
3
13
7
86
4
Spectrum of nosologies in combination of polymorphisms
С677Т MTHFR / А66G MTRR in patient selection (n=1938)
Nosology spectrum
С677Т MTHFR / А66G MTRR
Hmzg/ Hmzg Htzg/ Htzg/ N/
N/ Hmzg Htzg
Hmzg /Htzg Htzg Hmzg Hmzg Htzg /N
/N
Monogene pathology (5%)
2
4
11
11
20
21
Ehlers-Danlos syndrome
1
2
1
4
5
6
Prader-Willy syndrome
1
1
1
1
1
1
Louis-Bar syndrome
Klippel-Trenaunay syndrome
1
1
Tuberous sclerosis
2
2
Rendu-Osler syndrome
4
8
2
1
1
1
Silver-Russell syndrome
1
Rubinstein-Taybi syndrome
1
Hyperirritability syndrome
1
Arnold-Kiari syndrome
1
IDD syndrome
1
AGS
1
Anonichia-ectodactyly syndrome
1
McCune-Albright syndrome
1
Lesch-Nyhan syndrome
1
2
1
87
Spectrum of nosologies in combination of polymorphisms С677Т
А66G MTRR in patient selection (n=1938)
MTHFR /
С677Т MTHFR / А66G MTRR
Nosology spectrum
Hemorrhagic syndrome
Erb-Rott myopathy
Myopathy syndrome
Duchenne muscular dystrophy
PKU
Cystic fibrosis
Reiter syndrome
Marfan syndrome
Dandy-Walker syndrome
Fridreich’s ataxia
Polyneuropathy
Pertas disease
Gilbert’s syndrome
DHD syndrome
Hypophyseal nanism
Hmzg
N/ Hmzg Htzg
/Hmz Hmzg Htzg/ Htzg/ N/
/Htzg Htzg Hmzg Hmzg Htzg /N
/N
g
1
1
1
1
2
1
3
1
1
1
1
1
1
1
1
1
1
2
1
2
2
1
1
2
88
1
Spectrum of nosologies in combination of polymorphisms
С677Т MTHFR / А66G MTRR in patient selection (n=1938)
С677Т MTHFR / А66G MTRR
Nosology spectrum
Chromosomal pathology(6%)
Down syndrome
Down syndrome
(in relatives)
Hmzg
N/ Hmzg Htzg
/Hmz Hmzg Htzg/ Htzg/ N/
/Htzg Htzg Hmzg Hmzg Htzg /N
/N
g
2
1
Shereshevsky-Turner syndrome
Various chromosomal
pathologies/polymorphisms
1
16
5
10
1
29
5
27
14
2
1
14
2
6
2
3
14
5
6
4
1
6
4
3
6
5
3
2
89
Epigenetic disease (hypomethylation,
chromosomal polymorphism (46,ХУ, 9 phqh )
and polymorphic gene variants of folate cycle
(677 С-Т, А222 V mutation in heterozygotous
state).
Mild homocystinuria.
Syndromal epilepsy.
90
Rendu-Osler disease.
Polymorphic variant of 677 C/T MTHFR
gene in homozygotous state
91
Epigenetic disease?
Mosaic form of Shereshevsky-Turner syndrome.
Disorders of active enzymes of folate cycle.
Polymorphic variant of 677 С/Т MTHFR gene
was found in heterozygotous state, gene of
endothelial NO-synthase 4a/4b вin
homozygotous state).
Energy metabolism disorder (MNGIE?).
92
Familial case of epigenetic disease ?
DNA hypomethylation, folate cycle deficiency ,
methionine metabolism disorder (mosaic form of
trisomy 21, chromosomal polymorphism of
chromosome 1 ). Polymorphic variants of MTHFR
677 C/T gene in heterozygotous state, MTRR 66 G
gene in homozygotous state
93
Epigenetic disease?: glycoprotein metabolism
disorder (defect of posttranslation of lysosomal
enzymes).
Disorder of folate cycle metabolism (66A→G
(122М) polymorphism in MTRR gene in
heterozygotous state).
Chromosomal polymorphism:
46, ХY, 14 рs+.
94
Saethre-Chotzen syndrome,
secondary mitochondriopathy,
folate cycle deficiency
95
McCune-Albright syndrome in mother.
Polymorphic variants of 677ТТ
MTHFR/66A/G MTRR genes.
Healthy children
96
Robinow syndrome.
Multiple harmatose growth in the liver
Polymorphic variants of 677ТТ
MTHFR/66GG MTRR genes
97
Ukrainian Institute of Clinical Genetics
KhNMU
Kharkiv-22, Pravdu avenue, 13
Е-mail: mgc@ukr.net
98