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Prenatal Testing in Genetic Mutation Carrrier Parents | Jindal IVF

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Prenatal Testing in Genetic Mutation Carrrier Parents | Jindal IVF

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  1. PRENATAL TESTING FOR GENETIC MUTATION CARRIER PARENTS Dr Ritika Bajaj MBBS, MS (OBGYN), Fellowship in Maternal Fetal Medicine (AIIMS, New Delhi) Consultant Fetal Medicine Jindal IVF & Sant Memorial Hospital, Chandigarh

  2. IMPACT OF GENETIC DISEASE • CHILDHOOD • Genetic disorders account for- • 50% of childhood blindness • 50% of childhood deafness • 50% of all cases of severe learning difficulties • SPONTANEOUS MISCARRIAGES • 40-50% of all recognized 1st trimester pregnancy loss have a chromosomal abnormality • NEWBORN INFANTS • 2%-3% of neonates have at least one major congenital anomaly • 50% of these are caused exclusively or partially by genetic factors • Incidence of chromosomal abnormalities in neonates- 1 in 200 • Incidence of single gene disorders in neonates- 1 in 100 As infectious causes of perinatal mortality decline, the relative contribution of genetic causes in this subgroup has increased

  3. PRESENTING COMPLAINTS IN CLINICAL PRACTISE • Infertility- Karyotype • Recurrent pregnancy loss- Karyotype • Abnormal ultrasound findings in fetus during pregnancy • History of stillbirth/ unexplained death of previous child during infancy • History of intellectual disability/ developmental delay in previous child/ close relative • History of genetic disorder in family (mental handicap/ physical handicap/ malformation/ lethality)

  4. COMMON QUESTIONS THAT BRING COUPLES TO FETAL MEDICINE SPECIALIST/ CLINICAL GENETICIST • Why did it happen? • Can it happen again? • What can be done in next pregnancy? • Genetic diagnosis • Recurrence risk • Prenatal testing

  5. TYPES OF GENETIC DISORDERS • CLASSIC CHROMOSOME • DISORDERS • Full aneuploidies (Trisomy/ Monosomy) • Partial aneuploidies (Deletions/ Duplications of sufficient size that they can be seen on Microscope) • Phenotype practically always abnormal • MICRODELETIONS/ MICRODUPLICATIONS/ COPY NUMBER VARIANTS • Very small imbalances • Not detected on Standard Karyotype • May not always lead to clinical abnormality • MENDELIAN DISORDERS • Mutation at a single genetic locus • Autosomal Dominant/ Autosomal Recessive • X-linked dominant/ X-linked Recessive/ Y-linked

  6. APPROACH TO A COUPLE WITH HISTORY OF GENETIC DISEASE • 3 generation family history (PEDIGREE), Ascertain the pattern of inheritance • Try to make a clinical diagnosis (Role of clinical geneticist) • GENETIC TESTING OF THE INDEX CASE BASED ON PRESUMPTIVE CLINICAL DIAGNOSIS • Battery of genetic tests available, different tests give different information • Which genetic test to use- Depends upon the presumptive clinical diagnosis • NO SINGLE GENETIC TEST TO DIAGNOSE ALL GENETIC CONDITIONS • Many genetic tests have a Turn Around Time of 4-6 weeks • Ideally Molecular Genetic testing should be done in the preconception period or at first ANC visit itself • Prenatal testing possible if exact molecular diagnosis is available • Majority of genetic disorders do not have a cure • Current focus- • Prevention of genetic disorder (PGT) • Timely prenatal diagnosis

  7. TOOLS FOR PRENATAL DIAGNOSIS • INVASIVE TESTING • CVS (Chorionic Villus Sampling) • AMNIOCENTESIS

  8. AMNIOCENTESIS Process of withdrawing amniotic fluid from uterine cavity for diagnostic/ therapeutic purposes Optimal gestation – 16-17wks Early amniocentesis (<15wks) – Increased fetal loss, complication rates, culture failure COMPLICATIONS- DRY TAP – Fetal membranes tent over needle tip. More common in early amniocentesis (Incomplete fusion of amnion, chorion & decidua parietalis) FETAL LOSS – 1 in 300 to 1 in 500. Can occur upto 4 wks after procedure BLOODY TAP – < 1% cases. Blood almost always of maternal origin. Usually does not affect amniotic cell growth

  9. Amniocentesis done at 19 wks Patient had history of bleeding PV off & on USG- Subchorionic hematoma RBCs in pellet after centrifugation

  10. CVS • OPTIMAL GESTATION – After 11 weeks (Once NT, NB Scan is available) • Advantage of CVS over amniocentesis – • DNA analysis can usually be carried out directly on villi obviating the need and delay of a cell culture as required after amniocentesis  • Yield of cells and DNA from CVS much greater than 20ml of amniotic fluid • Provides a shift towards earlier diagnosis and option of termination at an earlier gestation • COMPLICATIONS – • Fetal loss – Comparable to amniocentesis in experienced hands • LIMITATIONS- • Mosaicism – 1-2% samples

  11. CASE NO. 1 CHROMOSOMAL ABNORMALITYBalanced Chromosomal Translocation In Either Parent Karyotype- 46,XX,t(8;10)(p21;q22) Balanced reciprocal translocation involving breakage at specific breakpoints on ch- 8 & 10 and exchange of fragments

  12. REPRODUCTIVE IMPLICATIONS OF BALANCED RECIPROCAL TRANSLOCATION IN EITHER PARENT • Behavior of Balanced reciprocal translocation at meiosis- • Problems arise at meiosis because chromosomes involved in translocation cannot pair normally to form Bivalents • They form a cluster known as Pachytene Quadrivalent • Each chromosome aligns with homologous material in quadrivalent

  13. RISKS IN RECIPROCAL TRANSLOCATION • When counseling a carrier of balanced translocation, risk of birth of an abnormal baby depends upon the particular rearrangement • This risk is usually somewhere between 1% and 10% Karyotype of child- 46,XX,der(8)t(8;10)(p21;g22)mat Additional material on short arm of ch- 8 due to maternal balanced translocation

  14. CASE NO. 2 • Achieved normal milestones & was attending regular school till 13 yrs of age • At 13 yrs of age- • Eating difficulty with mild drooling • Gait disturbances • Tremors • Speech difficulty • Dystonia • Ceruloplasmin levels, 24hr urinary copper excretion, Slit lamp examination of eyes, MRI findings s/o of Wilson’s disease • When couple presented to us at 12 wks POG, eldest child was already on treatment for Wilson/s disease • However, NO Genetic evaluation had been done

  15. DILEMMAS IN THIS CASE • Couple was from a remote area and had not brought the affected child • Already 12 wks of gestation & no genetic diagnosis available • In view of clinical diagnosis of Wilson’s disease, 25% recurrence risk in each pregnancy • Role of Clinical geneticist very important • Sample of affected child collected from home in this case as complete evaluation and clinical diagnosis was available and logistic issues • Couple counselled that prenatal diagnosis can be provided only if genetic testing of previous child reveals pathogenic variation in ATP7B gene • Coordination with lab in view of urgency of genetic diagnosis in prenatal testing

  16. Amniocentesis done at 16+4 weeks of gestation after pathogenic mutation identified in ATP7B gene FETUS UNAFFECTED

  17. Clinical suspicion of Osteopetrosis Baby died at 7.5 months of age Genetic testing of baby could not be done DNA of baby not available CASE NO. 3 30 ROLE OF TARGETED CARRIER SCREENING OF PARENTS

  18. Because clinical evaluation of previous child had been done, clinical diagnosis (Osteopetrosis) made by Geneticist was available • Based on clinical diagnosis, Targeted Carrier screeningwas able to identify mutation in parents • Both parents heterozygous for c.674G>A mutation in exon 3 of TCIRG1 gene • 60% cases of infantile osteopetrosis are due to mutations in TCIRG1 gene • Because pathogenic mutation was already identified when couple presented to us, CVS was done for prenatal diagnosis CVS- FETUS UNAFFECTED

  19. CASE NO. 4 • Both parents Beta Thalassemia Minor • WIFE- • Hb- 11gm%, HbA2- 4.9% • HUSBAND • Hb- 13.4gm%, HbA2- 4.8% • Late presentation- 18+6 wks • Counselled regarding 25% risk of Thalassemia major • Amniocentesis done • Trio sequencing done to save time • Pathogenic variant identified in parents and fetal DNA evaluated for the same mutation

  20. On an average, 10% of Indians are carriers for Beta Thalassemia gene mutation • HPLC/ Hb electrophoresis should be offered to all women who come for pre-conceptional counselling or at the First ANC visit • Hb should not be used as a criteria for offering Hb electrophoresis/ HPLC as B Thalassemia carriers can have Hb in the range of 10-11 gm% • In patients who present late for prenatal diagnosis- We counsel the parents and take a written consent explaining the need for legal route if report becomes available after legal limit for MTP and fetus is found to be affected FETUS- BETA THALASSEMIA MAJOR

  21. CASE NO. 5 • FTLSCS, B Wt.- 3.0 kg • Age-appropriate milestones till 6 months of age • H/O Recurrent LRTI • Abdominal distension, Hepatomegaly, jaundice at 6 months of age • Hepatorenal failure • Died at 7 months of age • H/0 5 abortions at 3 months POG • Cause for abortions- Not identified Died at 7 months Age d/t Hepatic failure

  22. Genetic evaluation of child had been done • Variant c.82G>T identified in SLC25A20 gene • Variant was classified as VOUS • (VOUS- Variant Of Uncertain Significance) • SLC25A20 gene is associated with Carnitine-acylcarnitine translocase deficiency • Carnitine-acylcarnitine translocase deficiency is AR metabolic disorder of long-chain fatty acid oxidation • Clinical features- Neurologic abnormalities, hypotonia, Cardiomyopathy, skeletal muscle damage, LIVER DYSFUNCTION

  23. VOUS (VARIANTS OF UNCERTAIN SIGNIFICANCE) • ACMG recommends a 5-tier system of classification of Variants • Pathogenic (2) Likely pathogenic (3) Uncertain significance (4) Likely benign (5) Benign • Counseling for VOUS should preferably be undertaken by Clinical geneticists • VARIANT REANALYSIS • Knowledge of variants is continuously evolving so an unreported variant or a VOUS at the time of initial Exome sequencing may be labelled pathogenic in future • Testing additional family members may result in re-classification of variants • Testing should be ordered only by specialists who are comfortable with the interpretation and explanation of results • Information should include options for reproductive decision making, pregnancy and perinatal management

  24. Evaluation by clinical geneticist Both parents underwent testing for the variant identified in SLC25A20 gene Sanger sequencing confirmed the presence of variant in SLC25A20 gene in both parents As parents were Heterozygous carriers for SLC25A20 gene, VOUS variant was reclassified as Likely pathogenic Genetic counselling done regarding 25% risk of recurrence Parents opted for prenatal testing by CVS FETUS – HETEROZYGOUS (UNAFFECTED)

  25. Husband- Myotonic Dystrophy Type I • Mild DM1 (Mild myotonia) • People with mild DM1 may have fully active lives & a normal or minimally shortened lifespan • Parents counselled • Myotonic Dystrophy is Autosomal Dominant Triplet CTG repeat disorder with 50% risk of transmission to children • ANTICIPATION – • Fetus may inherit repeat lengths considerably longer than transmitting parent • May lead to increasing disease severity & decreasing age of onset in successive generations CASE NO. 6 Myotonic Dystrophy Type I

  26. Molecular genetic diagnosis was available in father Couple opted for prenatal testing CVS done at 12 weeks FETUS UNAFFECTED

  27. 5th pregnancy- Unexplained IUD @ 28 wks • Genetic testing of fetus done • Fetus Heterozygous for mutation in CFTR & BTD gene • Genetic counselling done • Need to rule out carrier status of parents as both diseases (Cystic Fibrosis & Biotinidase deficiency) cause significant morbidity • Both parents found carriers for mutation in BTD gene (Biotinidase deficiency) CASE NO. 7 POG 12 wks

  28. In Present pregnancy- CVS done for both- • Fragile X syndrome • Biotinidasedefeciency • TAKEAWAY- • Whenever we find a genetic variant, we need to evaluate fully (even with very rare conditions) • What is rare in literature or in western world may not be very rare in our settings FETUS AFFECTED (HOMOZYGOUS FOR BTD GENE MUTATION)

  29. 10 yrs old Male child with DMD (Duchenne Muscular Dystrophy) GENTIC ANALYSIS- Deletion of exon 48-50 in Dystrophin gene Molecular Genetic testing of Mother- No Deletion or Duplication in 79 Exons of Dystrophin gene CASE NO. 8 DMD

  30. DMD pathogenic variant identified in affected Male is not detectable in Maternal leukocyte DNA • It could be a ‘de novo’ pathogenic variant • It could be because of Maternal germline mosaicism • What is Germline Mosaicism? • Some of the mother’s egg cells carry the Dystrophin gene mutation while other egg cells donot • Likelihood of maternal germline mosaicism in DMD is 15% to 20% • Even when the mother is not a carrier, it is possible that she has Germline mosaicism for the mutation and is at risk of having a second son with DMD • Prenatal testing should be offered to mother in all subsequent pregnancies

  31. CASE NO. 9 Present pregnancy- NT/ NB Scan- CRL- 54mm NT- 3.5mm (>99th centile) Unossified NB

  32. FETUS- TRISOMY 18

  33. THANK YOU Prenatal testing in genetic mutation carrier parents is much more than CVS or Amniocentesis In fact, that might be the easiest part Knowledge of Clinical Genetics is the backbone of prenatal testing in Single Gene Disorders

  34. PRACTICAL CONSIDERATIONS- • R/O Down syndrome if either parent carries Robertsonian translocation involving ch- 21

  35. ROBERTSONIAN TRANSLOCATION IN EITHER PARENT CASE NO. 2 • Robertsonian translocation- Breakage of two acrocentric chromosomes (13, 14, 15, 21 and 22) at or close to their centromeres with subsequent fusion of their long arms • Short arms of each chromosome are lost (no clinical importance as they contain genes only for ribosomal RNA for which there are multiple copies on other acrocentric chromosomes) • Total chromosome number is reduced to 45 • This is a functionally balanced rearrangement

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