pohľad genetika na diagnostiku a liečbu...link between genetics and infertility disorders •...
TRANSCRIPT
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Černáková I., Petrovič r., Fischerová M.
Pohľad genetika na diagnostiku a liečbu neplodnosti
Diskusné sústredenie genetikov 2017, SZU, Bratislava, 8.6.2017
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(In)fertility
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Inferility frequency
1 : 6 couples
or
15% of couples
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Link between genetics and infertility disorders
• 1940s – gynecomastia, aspermatogenesis and increased secretion of FSH (Harry Klinefelter) – Klinefelter syndrome
• 1956 – determination of karyotype in KS patients: 47,XXY
• 1938 – the first description of Turner syndrome by Henry Turner
• 1964 – karyotype 45,X determination in patient with Turner syndrome
3 000 – 5 000 genes directly or indirectly influencing human fertility:
- genes expressed just in germ cells
- genes expressed just or also in gonads
- genes expressed in ontogenesis
Laboratory tests available:
- Genetic screening:
- basic panethnic: CF, SMA, hemoglobinopathies
- expanded population specific –Middle Eastern, Askenazi Jewish, Mediterranean
- expanded panethnic - few hunderds of autosomal recessive and X-linked diseases
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Link between genetics and infertility disorders
- Genetic diagnosis of monogene disease in the family
- Genetic examination for some infertility-related mutations
(FSHR, Fragile X, FMR I, AR gene)
- Whole genome testing of infertility
(Atypical ovarian response, premature ovarian insufficiency, male factor infertility, disorders of sexual development, recurrent pregnancy loss, polycystic ovarian syndrome...)
- Preimplantation genetic diagnosis /screening of chromosomal anomalies
(prevention of genetic diseases)
- Prenatal genetic diagnosis
(managment of delivery, treatment and care about child with genetic disease)
Clinical genetic counseling
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Why clinical genetics in IVF treatment of infertility?
Infertile and sterile patients are a source of genetic pathologies - occurence of genetic pathology in infertile patients - occurence of particular genetic disease in family - transmission of genetic disease and infertility to the child by IVF treatment
Laboratory test of: - most offen occuring genetic pathologies in infertility - genetic diseases potentially passing to the next generation (karyotype + genetic-based infertility conditions + genetic screening – CF, SMA, hemoglobinopathies...)
Genetic counseling of: - results of genetic tests - risk of transmission of genetic disease to the next generation - explanation of reproductive option of examined couple - clinical genetic examination and laboratory genetic test of relatives at risk of diagnosed disease
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Chromosomal abnormalities Preimplantation
period
Implantation I. trimester
II. trimester
III. trimester Term
Inherited birth defects
Early miscarriage 50 to 60 % when gross structural abn. is present Late miscarriage 12 % 15% 0,5 % 4 – 5 %
50% loss before implantation at 5 to 6 days postconception
Spontanneous miscarriages - 15% of clinically recognized pregnancies before 12th weeks ´gestation
The frequency of chromosomal abnormalities after birth
General population 1 : 200 0,5 %
Infertile couples (spontaneous miscarriages, stillbirths)
1 : 48 2 %
Sterile couples 1 : 10 10 %
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Prenatal/Postnatal
genetic diagnostics
Preimplantation
genetic diagnostics
and PGS + Mitoscore
Genet. dg PB Genet. dg of sperm
ET of good embryos
8 cell stage
Infertility treatment
- Endometrial receptivity assay
Zygota ET
REPRODUCTION
OF COUPLE
GENETIC EXAMINATION OF INFERTILE COUPLE
Embryo biopsy –
Day 3 or Day 5
Genetic examination of couple:
- Karyotype - Genetic tests of infertility
- Cystic fibrosis - Carrier genetic test
- Trombophilia mutations - Pharmacogenetics in infertility treatment
- Y-chromosome microdeletion
Genet. dg of spont. miscarriage
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The reasons of male infertility
Frequency of infertility world-wide 15 – 20%
Gynecological reasons 40 %
Andrological reasons 40 %
Idiopathic infertility 20 %
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WHO GUIDELINES AND
DEVELOPMENT OF SPERM QUANTITY
0
10
20
30
40
50
60
70
80
90
1951 1964 1987 2023
2010
15 mil/ml
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The reasons of male infertility
Endogenneous factors
Disorders of sperm production
Disorders of seminal ductus
Genetic factors (translocations, numerical chromosomal abnormalities, Y-deletions, CFTR
gene mutations and mutations in other genes)
Other (anatomical, hormonal, age of man ...)
Hominid ancestry
Exogenneous factors
Industrial exhalation
Chemicals – herbicids, pesticids, insecticids
X-ray, microwave, mobil cells
Industry – heavy metals
Life style factors – smoking, alcohol, drugs, obesity, sauna/thermal baths
Inflammation diseases, radio-/chemotherapy
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Human male germ cells
Characterization:
Poor quality exhibiting frequent morphological defects
Impaired motility
Poor chromatine compaction and condensation
High incidence of DNA fragmentation
Impaired capacity for oocyte recognition and fusion
Major source of of disease-causing mutations in humans
Infertility Dominant single gene disease
Childhood cancer
Birth defects
Immunological disorders
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Sperm analysis - basic
(WHO 2010)
Volume > 1,5 ml
Count > 15 mil/ml
Motility > 40 %
Progressive motility > 32 %
Morphology > 4% of normal sperm
pH ≥ 7,2
Liquification up to 30 min
Leucocytes < 1 mil/ml Leu
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SPERM DNA DAMAGE
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Sperm cell disorders
Paternal imprinting disorders
Chromosomal aneupoidy
Defects of morphology
Defects of vitality (mitochondria insufficiency)
Residual cytoplasma
Protamines insufficiency
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VitalSperm Vitality examination – Progressive vitality < 40%
Living immotile sperm Dead sperm - necrozoospermia
Pathological finding
of epididymis
Structural defects of tail
+
Defects/lower count of mitochondria
Urological, sperm donation DNA fragmentation
Antioxidants + ICSI
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VitalSperm
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LeucoTest
Examination
of leucocytes in ejaculate
Small round cells
> 1 mil/ml
Epitelial cells Leu > 1 mil/ml
Urological examination
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Why is important to do correct semen analysis?
To know correct diagnosis of male infertility
To suggest and realize additional genetic tests
To propose to the couple their real opportunities to have baby
To choose the appropriate method of ART
(IVF/ICSI, PICSI, MACS or TESE)
or to decise for a sperm donation
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ProtamineSperm
Examination of presence of protamines in sperm nucleus
– Morphology < 4%
– Idiopatic infertility
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Protamine test When the strands are not packed well –
long DNA strands succeptible to damage
Sperm DNA fragmentation
Low probability of fertilization of oocytes
Infertility
Counselling of couple, IVF / ICSI treatment
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Oxidative stress
Production of DNA adducts
DNA endonukleases cleavage
Imprinting disorders
DNA damage and
mutations
Assisted reproduction
Thermal stress in scrotum
DNA Reparation normal
Normal
embryogenesis
Abnormal development of
embryos
Unsuccessful implantation
Spontanneous miscarriages
Single gene disease in
children
Children tumor diseases abnormal
Infection
Disorders of spermatogenetis:
- residual of cytoplasma
- abortive apoptosis
- abnormal protamination
Disorders of DNA replication
increased by age
Xenobiotics:
- life style
- medications and oncotherapy
- professional environment
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FragmentSperm
Direct examination of DNA damage
-ssDNA a dsDNA –
by TUNEL method
(fluorescence microscopy)
INDEPENDENT PROGNOSTIC
FACTOR for measurement
of sperm quality relating to the
MALE INFERTILIY
Sperm DNA fragmentation
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Sperm DNA fragmentation
Normal sperm
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Up to 15 % - normal level
> 15 % - IUI
> 20 % - IVF/ICSI
> 30 % - small probability of
spontaneous pregnancy
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Men with OAT + Men with normal SPG
idiopatic infertility (normal SPG)
repeated unsuccessful IVF cycles: non-fertilized oocytes
no cleveage of embryos
embryo fragmentation
no clinical pregnancy
spontanneous miscarriages
men older than 40 years
exposition of toxins/medications
smoking
infections, inflammation
Sperm donation – testing of donors
INDICATIONS – Sperm DNA fragmentation
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AneuSperm
FISH examination of sperm karyotype – chromosomal
aneuploidy
Examination of aneuploidy level
in sperm sample
- chromosomes 13, 18, 21, X a Y
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OAT gravis
Normálny SPG
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Men with
sperm aneuploidy
Infertile male patients with sperm
of pregnancy is miscarried by couples with abnormal aneuploidy rate in sperm
60%
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Embryos: aneuploidy in embryos
triploidy in embryos
Pregnancy: decreased pregnancy rate of the treatment of infertility
increased frequency of spontaneous miscarriages
Births: increased risk of aneuploidy in baby
Identification of men with low reproductive success
Personalized genetic counseling of infertile couple in relation to IVF treatment
Preimplantation genetic screening of aneuploidy – goals:
- to increase the pregnancy rate
- to decrease the risk of spontaneous miscarriage
- for a higher chance to have healthy baby
Sperm aneuploidy effects
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Impact of sperm aneuploidy on pregnancy rate/ET
in IVF treatment and the incidence of spontaneous miscarriages
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Men with OAT + Men with normal SPG
Repeated unsuccessful IVF cycles: non-fertilized oocytes
no cleavage of embryos
embryo fragmentation
no clinical pregnancy
Repeated spontanneous miscarriages with normal karyotype of couple
Chemicals /Medication exposition /Chemotherapy/Radiotherapy
Preimplantation genetic diagnostics
Donation programme
The indications - AneuSperm
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Why to do the comprehensive sperm analysis?
• To know the correct diagnosis of male infertility
• To suggest the additional tests for correct diagnosis (karyotype, CF mutation screening, Y-chromosome deletions, FSH gene and FSH receptor gene polymorphism for hormonal treatment ...)
• To explain possible causes of infertility and the risk of transmission of infertility, recurrent miscarriages and genetic diseases to their children
• To guide the treatment of male infertility: - antioxidants - hormonal treatment by low dose-long term FSH according to the genotype - to recommend the appropriate methods of IVF: ICSI, TESE for non-obstructive azoospermia, MACS / PICSI preimplantation genetic screening for aneuploidy, for CF ....)
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Folicule stimulating hormone (FSH) is produced by adenohypophysis – central hormone of human reproduction, which is essential for development of gonads and production of gametes
FSH stimulates the growth of folicules and oocytes in ovaries and Sertolli cells in testes and supports the production of sperm
Male idiopatic infertility
FSHB – gene encoding β-subunit of FSH
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FSHB gene promotor : transcription control of FSHB gene
SNP rs10835638 .... nucleotide substitution -211G>T
Standard allele G derived allele T (20 – 25% of USA and Europe population)
Function of derived allele T: significant decrease of transcriptional activity of FSHB gene
Genotypes:
-211G/-211G .... Normal homozygote -211G/-211T .... Heterozygote .... - 25% of activity of FSHB gene -211T/-211T .... Homozygote for derived allele T .... - 50% of activity of FSHB gene (lower serum level of FSH) .... Oligozoospermia, lower volume of testes, decreased
level of testosterone and higher level of LH in sera
Variants in promotor of FSHB gene
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FSHR – FSH receptor gene
2
FSHR gene promotor : SNP rs1394205 .... -29G>A Dominant , occurence independent on SNP in exon 10 Exon 10 of FSHR gene: SNP rs6165 .... c.919G>A .... 307: Thr/Ala SNP rs6166 .... c.2039A>G .... 680: Asn/Ser - genetic linkage
http://regionweb.sk/reprogen/wp-content/uploads/2011/10/FSH-receptor-gene-schema.jpg
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SNP rs6165 .... nucleotide substitution c.919G>A .... 307: Thr/Ala
Standard allele G derivoved allele A
SNP rs6166 .... nucleotide substitution c.919G>A .... 680: Asn/Ser
Standard allele A derivoved allele G
Function of derived allele: - different sensitivity of receptor to FSH hormone according to genotype
- different response to exogenous injection of FSH according to genotype
Variants in exon 10 of FSHR gene
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Impact on FSH level
Expresia receptora FSHR -29G>A (promotor)
FSHR p.N680S (exon 10)
FSHB -211G>T (promotor)
GG GA AA
GG GT TT
AA GA GG Asn/Asn Ser/Asn Ser/Ser
Serum FSH
Serum FSH
Serum FSH
Receptor sensitivity
Transcription activity Treatment
by FSH - 12 weeks (prof. M. Simoni, 2014)
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Metodika
• Súbor - 72 pacientov s OAT
Kritéria pre zaradenie do štúdie:
- vek 20-50 rokov s idiopatickou mužskou sterilitou
- stav po neúspešnom IVF cykle
- negatívny urologický nález
- hormonálny profil v medziach normy (FSH, LH, TST, Prol, TSH)
- normálny karyotyp + vylúčenie mutácie v géne pre cystickú fibrózu
a mikrodeléciu chromozómu Y
• Polymorfizmy (rs6165, c.919A>G and rs6166, c.2039A>G) v exóne 10 FSHR génu a v promótore FSHR a FSHB sme vyšetrili DNA sekvenovaním
• Muži s aspoň jednou Asn680Ser alelou – MERIONAL HPLH 75 IU
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Výsledky
Úspešnosť liečby: % hodnoty “pregnancy rate“
porovnaním hodnôt SPG v jednotlivých parametroch pred a po liečbe
„Pregnancy rate“ u mužov so závažnými formami OAT
s normálnym haplotypom (wild type)….............................................................25%,
po hormonálnej liečbe u heterozygotov…......…................................................52,4%
po hormonálnej liečbe u patolog. homozygotov…............................................63,4%
Potvrdili sme tak štatisticky významné zvýšenie PR po hormonálnej liečbe podľa typu FSHR genotypu (p < 0,0001)
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Výsledky
Pred liečbou Po liečbe
Počet spermií (mil/ 1ml) 3,8 7,6
Počet motilných spermií (mil/1ml) 2 5,2
Pohyblivosť a - 15%, b - 85% a - 25%, b - 75%
Morfológia 95% 92%
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Why clinical genetics in IVF treatment of infertility?
Infertile and sterile patients is s source of genetic pathologies - occurence of particular genetic disease in family - occurence of genetic pathology in infertile patient - transmission of genetic disease and infertility to the child by IVF treatment
Laboratory test of: - most offen occuring genetic pathologies in infertility - genetic diseases potentially passing to the next generation (karyotype + genetic-based infertility conditions + genetic screening – CF, SMA, hemoglobinopathies...)
Genetic counseling of: - results of genetic tests - risk of passing of genetic disease to the next generation - explanation of reproductive option of examined couple - clinical genetic examination and laboratory genetic test of relatives at risk of
diagnosed disease
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CLINICAL GENETICIST - Genealogy, physical examination,
genetic counseling
- Karyotype
- Cystic fibrosis
- Trombophilic mutations
- Y-chromosome microdeletion
- examination of genetic diseases occuring in some population
(thalassemias, familial mediterrean fewer, Jewish specific disorders,
or in particular patients/family (monogene diseases)
Foresta C, Ferlin A, Gianaroli L, Dallapiccola B:
Guidelines for the appropriate use of genetic tests in infertile couples.
Eur J Hum Genet, 2002, 10, 5, 303 – 312.
Preconception clinical genetic service of infertile couples
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Chromosomal abnormalities and human reproduction
Disturbance of gametogenesis
- male infertility: oligoasthenoteratozoospermia azoospermia
- female infertility: disturbance of menstrual cycle
premature ovarian insufficience
Unbalanced chromosome abnormalities in gametes
- reproduction failures : spontaneous miscarriages stillbirths
birth defects
neonatal death
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Medical indications of cytogenetic examination of infertile couple
1. Occurence of chromosomal abnormality in family
2. Congenital abnormality in development and/or mental disabbility in family
3. Infertility (minimal 2 spont. miscarriages, stillbirths, repeated implantation failures in IVF cycles)
4. Idiopathic infertility (more than 1 year or 6 months when woman is older than 35 yrs.)
5. Andrologic infertility (OAT, azoospermia)
6. Disorders of menstrual cycle (amenorhea, oligomenorhea)
7. Anatomical defects of male/female genitalia
8. Professional and medical exposition
9. Couples before PGS of aneuploidy
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Cystic fibrosis
Indication of examination of CFTR gene mutation
Male infertility - oligozoospermia ( 1< 106/ml )
- azoospermia
Men with congenital bilateral absence of vas deference (CBAVD)
Familial occurence of cystic fibrosis - based on genealogy
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Indications of examination - Trombophilic mutations
Leiden mutation (Factor V gene) Protrombin gene mutation (Factor II)
- Deep venal trombosis in personal and/or familial history
- Repeated spontaneous miscarriages
- Repeated unsuccessful IVF cycles
- Reproductive history:
stillbirth, premature delivery, abruption of placenta, preeclampsia, IUGR
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Y-chromosome microdeletions
Men with severe oligozoospermia or azoospermia and Y-chromosome microdeletion:
- 100% of probability of microdeletion
and infertility transmission to the sons
- Decreased fertilization rate of oocytes
- Less quality of embryos
Indication of examination:
- Oligozoospermia (1
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The cause of spontaneous miscarriages
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Genetic laboratory tests
- couples with repeated spontaneous miscarriages
Test Who? Material
Karyotype Both partners Peripheral blood
Annexin Both partners
Peripheral bloos / buccal cells
Trombophilia mutations FV, FII
Woman Peripheral bloos / buccal cells
Sperm DNA fragmentation + ROS
Men Sperm
Sperm chromosome aneuploidy
Men Sperm
+ Clinical genetic counseling
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Annexin 5
ANXA5 gene – chromosome 4q27 4 variants in promotor of the gene: 19G>A (rs112782763) +1A>C (rs28717001) 27T>C (rs28651243) 76G>A (rs113588187)
M2 haplotype M1 haplotype
N (WT) – standard alllele M1 haplyotype – non-pathologic, no complications in pregnancy, no increased risk of spontaneous miscarriage M2 haplotyp – defect of placental vasculature, venal tromboembolism, 4x increased risk of spontaneous miscarriage, IUGR and intrauterine death of fetus, gestational hytertension, preeklampsia M2/M2 haplotype of fetus – severe form of IUGR
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Annexin 5
Activity of ANXA5 gene promoter variants in luciferase reporter gene assays.
Nadia Bogdanova et al. Hum. Mol. Genet. 2007;16:573-578
© The Author 2007. Published by Oxford University Press. All rights reserved. For Permissions, please email: [email protected]
Genotypes: WT/WT WT/M1 M1/M1 WT/M2 alebo M1/M2 M2/M2 Incidence: of IVF patients has M2 haplotype of persons has M2/M2
The fetus has an increased risk to inherit trombophilia when minimal one M2 maternal or paternal haplotype is present
Men has no risk himself, nor woman when is not pregnant!
44%
6%
• Idiopathic infertility – 37% • Repeated miscarriages • OAT - 40% • PCOS – 35% • Premature ovarian insufficiency
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Annexin 5
The treatment – low molecular weight of heparine NO ASPIRIN!
from embryo transfer – to end of puerperium
Which material to examine? • Peripheral blood • Buccal cells
How? • Genetic test • NO IMMUNOLOGIC !
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Chorionic villi biopsy
Amniocentesis
molecular-genetic diagnostics
Invasive prenatal diagnostics
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PGD / PGS Preimplantation genetic diagnostics - analysis of genetic disorders in early human embryos before their embryotransfer in uterus as a prevention of single gene disorders
• Introduced in routine: 1990 – adrenoleukodystrophy
Preimplantation genetic diagnostics – prevention of genetic disorder
transmission to the next generation
- single gene disorders - chromosomal translocations
Preimplantation genetic screening af aneploidy – prevention of transfer of
chromosomally abnormal embryos of „poor responder“ couples
- older women (35+)
- repeated unsuccessful impalntation of embryos
- repeated spontaneous miscarrages
- male infertility
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PGD medical indications
Preimplantation genetic diagnosis
- Diagnostics of single gene disorders
- Diagnostics of translocations
- Diagnostics of late-onset genetic diseases or cancer diseases occuring
in adulthood
- HLA typization of embryos
- Mitochondrial diseases
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PGD - steps
Haplotype analysis of family
– PGD set-up
IVF/ICSI
Hormonal stimulation
Oocyte pick-up
Fertilization
Embryo biopsy – Day 3,
Day 5 or blastocentesis
Genetic analysis
IVF – Embryo transfer
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Hormonal stimulation of ovaries
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Oocytes pick-up
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Embryo culture
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Vyšetrovaný materiál
1st polar body 2nd polar body
Trophectoderm of blastocyst – Day 5 Blastomere - Day 3
Embryo biopsy
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PCR – PGD steps
A. Biopted of cell (cells) is transfered
in lysis buffer solution in eppendorf
tube
B. Its genome is released from nucleus
of cell.
C. Whole genome amplification (WGA)
– amplification of all genome of cell
D. Fragmentation analysis of examined
gene using capillary electrophoresis
– analysis of genotype of embryo
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Haplotype analysis
Allelic drop-out risk
Analysis of 5 fully informative DNA
markers linked to the mutation
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Haplotype analysis of the family
Marker –IVS10CA
proband - sick
mother
father
healthy daughter
IVS10CA Alela 1 Alela 2
Proband - sick 318 330
Mother 320 330
Father 318 320
Healthy daughter 320 320
Analysed
12 DNA markers
before, inside and
after CFTR gene
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Transfer of healthy embryo
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PGS medical indications
Preimplantation genetic screening of aneuploidy
FISH: 8 chromosomes
aCGH / NGS: all chromosomes
- Women older than 35 - 37 years
- Repeated unsuccessfull IVF cycles
- Repeated spontaneous miscarriages
- Trisomic fetus in previous pregnancy (Down syndrome)
- Male infertility (OAT gravis, TESE/MESA)
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Chromosomal abnormalities in oocytes and maternal age
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0,001,002,003,004,005,006,007,008,009,0010,0011,0012,0013,0014,0015,00
16 X 212215 131814 9 2 4 7 8 2010 1217 6 5 11 3 19 Y
Chromozóm
Pe
rce
nto
%
Frequency of chromosomal abnormalities
in spontaneously miscarriages – Ist trimester
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Frequency of aneuploiy in preimplanattion period – abnormalities detected by CGH method
Munné et al. , 2010
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PGD/PGS schematic representation Day 0 Oocyte pick-up
Day 1 Fertilisation
Day 3 Biopsy of blastomere
Day 3 + Day 4 results, report FISH, aCGH, PCR-PGD
Day 5 Embryo transfer Biopsy of vitrification of biopted embryos trophoectoderm aCGH, NGS ET in next menstrual cycle
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arrayCGH
arrayCGH
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Examples of aCGH results
-10 -22
-5p parc. +19
blastoméra
6denné embryo
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Spontaneous miscarriages I. Aneuploidy
Natural conception: 15 %
ART: 23 – 37 %
PGS: 12 - 15 %
2 x comparing to ICSI
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Spontaneous miscarriages II. Translocations
Spontaneous miscarriages:
Natural conception 87 %
PGD 18 %
5 x
Take-home-baby rate:
Natural conception: 11,5 %
PGD: 81,4 %
7 x
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Trisomy child delivery
Natural conception:
2,6 % trisomy 13, 18 a 21 (chorionic villi examinations)
PGD cycle:
0,6 % trisomy 13, 18 a 21
independent on type of hormonal stimulation
4 x
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Mitochondria during embryogenesis
Mitoskóre Implantation rate %
< 18 81%
18-24 65%
24-50 50%
> 50 18%
> 160 none
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Importance of PGD / PGS
Diagnosis - diagnosis of chromosomal anomaly/single gene disease according to the patient´s medical indication
Prevention - decreased risk of spontaneous miscarriage in IVF couples to 15%
PREVENTION OF DELIVERY OF CHILD WITH GENETIC DISORDER !
Treatment - higher effectivity of infertility treatment in poor prognosis patients
Prognosis - prognosis & change of infertility treatment in next IVF cycle
Finance - next examinations & IVF treatment
Psychology - psychic status of infertile couple
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ERA® ENDOMETRIAL RECEPTIVITY ASSAY
-
ERA® ENDOMETRIAL RECEPTIVITY ASSAY
+ =
24% of infertile women with
repeated implantation failure
has changed implantation window
(delayed or advanced)
even good quality of embryos are
transfered
-
ERA® ENDOMETRIAL RECEPTIVITY ASSAY
LH peak measuren in urine/sera
E2: 6 mg/day
E4: 800 mg/day
- Molecular customer-made microarray prepared
for diagnostics of endometrial receptivity in IVF treatment
- Examination of expression of 238 genes participating
in receptivity of endometrium
- Classification of endometrium as „receptive“ or „non-receptive“
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Personalized embryo transfer
P+3 P+5 P+7
LH+5 LH+7 LH+9
ET
pET delayed
pET advanced
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Diagnostic importance of ERA®
Diagnostics of endometrium during implantation window
Precise detection and timing of embryo transfer according
to the molecular characteristics of particular woman
Personalized embryo transfer
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Single gene disorders
• Fenotyp effects are caused by the mutation in one gene
• Mendelian type of inheritance
• 7 000 single gene disorders
WHO: Prevalence 10 : 1000
20% of cases of child mortality
in developed countries
40% of medical interventions
in children hospitals
(Kanada – Scriver, 1995)
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Carrier screening of single gene diseases
single gene diseases AR a X-linked inheritance
General population – affected 1 : 100
Carriers
Cystic fibrosis 1 : 25 4 %
Spinal muscular atrophy 1 : 50 2 %
Αlpha-/Beta-hemoglobinopathy 1 : 48 2 %
-
- analysis of 550 genes, 600 single gene diseases using NGS
- high risk of transmission of disease with recessive inheritance
(AD or X-linked) to the next generation
2.1 pathogenic mutation/person
7.1 % of tested persons don´t have pathogenic mutation
8% couples carry the mutation in the same gene
( so called „genetic incompatibility“)
Carrier screening of single gene diseases
-
Preimplantation genetic diagnostics
1 : 100 1 : 100 000
Risk of the delivery of the child affected by single gene disorder
For whom? Before natural conception
Before assisted conception
Oocytebanking and spermbanking
Carrier screening of single gene diseases
-
Single gene disorders can´t be cured, but they can be prevented
Primary prevention of occurence of single gene disorders in family
- Preimplantation genetic diagnostics
- Carrier screening of single gene disorders
-
USG / age / biochemical screening
amniocentesis / CVS
QF-PCR
KARYOTYPE
Risk of miscarriage: 0.5 – 1% dg impossible before 10th week of gestation
FISH
or
Non-invasive prenatal genetic test
DNA chip FISH
RAPID PND
Invasive
prenatal genetic test
-
From 10th weeks of pregnancy
Numerical abnormalities of chromosomes – aneuploidy 1 – 22, X, Y
Structural deletion or duplications – resolution 5 - 7 Mb (one G-band in karyotype)
Sex chromosome examination
Microdeletion syndromes: Prader Willi/Angelman syndrome – deletion 15q11.2 Di George syndrome – deletion 22q11.2 Wolf Hirschhorn syndrome – deletion 4p Cri-du Chat syndrome – deletion 5p deletion 1p36
PrenatalSafe KARYO Plus
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Comparision of non-invasive prenatal tests
PrenatalSafe KARYO PLUS – 95,5%
-
It recognizes 92,6% of chromosomal abnormalities detected in prenatal
development of fetus
Analysis of each of chromosomes
Neinvasive procedure
Unbalanced translocations
Aneuploidy
Mosaics
Marker chromosomes
Microdeletion syndromes
Triploidy
Diagnostic test
Classical karyotyping
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Genetics and IVF clinic a multidiscipline dialogue in assisted reproduction
Clinical geneticist
- physical examination
- genealogy
- pre-test counseling
- indication of genet. lab. tests
- post-test counseling
IVF clinic
Genetic lab
- Molecular cytogenetics
Genetic lab
- DNA lab
Genetic lab
- Cytogenetics
Urology
Gynecology
Genetic lab
- PGD lab
Endocrinology
-
Bring genetics in your practice
TREATMENT
GUIDE treatment, such as gonadotropin dosage and prescription of blood thinners
MANAGE treatment risks, such as ovarian hyperstimulation syndrome
DECISION-MAKING
INFORM decisions about reproductive options, plan and treatment protocol
RECOMMEND preimplantation genetic diagnosis/screening (PGD/PGS)
GUIDANCE
PREDICT and prepare patients for possible outcomes of infertility treatment
EXPLAIN possible causes of infertility or recurrent miscarriage
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Thank you for your attention
Genetic lab ReproGen
Bratislava, Slovakia
Tel: 0948 230 661
www.reprogen.sk
http://www.reprogen.sk/