bjectives Understand Preconcep.on and Post-concep.on op.ons for gene.c tes.ng Pre-Concep.on Post-Concep.on

Transcription

1 bjectives Understand Preconcep.on and Post-concep.on op.ons for gene.c tes.ng Pre-Concep.on Review current guidelines for gene.c carrier screening (Amie) Review the in vitro fer.liza.on process (Kenan) Highlight the difference between preimplanta.on gene.c diagnos.c tes.ng and preimplanta.on gene.c screening Review the different stages of embryo development Post-Concep.on Chorionic villous sampling, amniocentesis Aneuploidy screening Diagnos.c gene.c tes.ng

2 ase 1 6 yo G0 who is interested in star.ng family. She has not family history of ny medical problems, but notes that her ancestors are of Ashkenazi Jewish escent. er husband 28 year old and is of Northern European descent and notes a mily history of Hun.ngton s disease in both his father and his paternal randfather. He does not know whether he carries the Hun.ngton gene and ould prefer not to know. hat Preconcep.on carrier screening tes.ng is recommended? hat addi.onal counseling does the woman s age necessitate? hat is their chance of having a child with Hun.ngton s Disease given that e husband s status is unknown?

3 Current Guidelines for Carrier Screening Professional guidelines (ACOG/ACMG/NSGC) Cys.c fibrosis offer to all Caucasian and Ashkenazi Jewish pa.ents offer pan-ethnically Spinal Muscular Atrophy 2008 ACMG recommended offer pan-ethnically ACOG recommends based on family history Screening offered based on stated ethnicity Hemoglobinopathies Thalassemias Ashkenazi Jewish carrier tes.ng Screening offered based on personal or family history Fragile X syndrome No society is recommending expanded carrier screening

4 urrent Guidelines for Ashkenazi Jewish Carrier Screening ACOG (2004/2009/2014)! Offer to all: Cys.c Fibrosis, Canavan disease, Familial Dysautonomia and Tay Sachs! Make informa.on available on other tests so that pa.ent can make informed decision regarding addi.onal tes.ng ACMG (2008/2013)! Offer to all: Cys.c Fibrosis, Canavan disease, Familial dysautonomia, Tay Sachs AND Fanconi Anemia type C, Niemann Pick type A, Bloom syndrome, mucolipidosis type IV and Gaucher disease! Screening for addi.onal condi.ons as follows:! natural history of the condi.on is well understood and it carries a poten.al for significant morbidity and/or mortality! >90% detec.on rate OR allele frequency >1% in the Ashkenazi Jewish popula.on

5 rogenity Ashkenazi Panel

6 Limita.ons of Current Guidelines Inadequacy of ethnicity-based screening: 2010 US Census data 32% increase in individuals repor.ng mul.ple races Preference against using racial and ethnic categoriza.on in medicine Possible unknown or unreported ancestry Lack of family history, limited knowledge, adop.on Family history based screening may not be helpful: Recessive nature of many condi.ons

7 Why not just test everything? Several expanded carrier panels are being marketed! Typically screen for ~400 muta.ons in ~100 autosomal recessive disorders! The long tail of Mendelian disease Does everyone need to know what they are carriers for? Srinivasan BS et al, RBMO, 2010

8 Experience with Expanded Carrier Screening 3,738 couples tested with Inheritest or Counsyl 25.1% had 1 posi.ve result Only 7 couples (0.19%) had findings that impacted subsequent clinical management 3 of these were CF and would have been captured on rou.ne screening Franasiak, ASRM 2014

9 untington disease Autosomal dominant disorder caused by an expansion of a CAG trinucleo.de tract in the HTT gene Progressive disorder of motor, cogni.ve, and psychiatric condi.ons Mean age of onset is years of age

10 F / B A / E? A / B C / D

11 F / B A / E AT RISK NOT AT RISK? A / B C / D A / D B / C A / C B / D B / C

12 ASE 1 Carrier screening nega.ve for both. Couple would like to proceed with preimplanta.on gene.c diagnosis (PGD) to reduce risk of having child with Hun.ngton's disease

13 PGD for Single Gene Disorders Putative Advantages Safer than elec.ve termina.on More ethically and psychologically acceptable than termina.on for many couples Provides the couple with more op.ons than: Adop.on, steriliza.on, donor eggs or sperm

14 Preimplantation Genetic Testing Preimplanta.on Gene.c Diagnosis (PGD) Done in individuals with KNOWN GENETIC DISORDERS (o0en fer5le) Different test for all individuals Even those with the same gene.c disorder Even those with the same muta.ons The analy.cal plaporm chosen may depend on the nature of the specific disorder

15 Single Gene Disorders Are typically the result of a single mutated gene Muta.ons Trinucleo.de repeat expansions (HD) Dele.ons Duplica.ons > 6000 human diseases due to single gene disorders; occur in ~1 in 200 births Inheritence parerns: Autosomal dominant Autosomal recessive X-linked dominant X-linked recessive Y-linked recessive Mitochondrial

16 Single Gene Disorders Autosomal Recessive Disorders β-thalassemia and/or sickle cell syndromes Cys.c fibrosis Spinal muscular atrophy Autosomal Dominant Disorders Hun.ngton Disease Myotonic distrophy type I Charcot Marie-Tooth Sex Linked Disorders Fragile X syndrome Duchenne and Becker muscular dystrophy Hemophilia A and B HLA Matching (Saviour Siblings) β-thalassemia and/or sickle cell syndromes Fanconi anemia

17 Differences Origins Expanded of between Carrier PGD for Testing: Single PGD/PGS Concerns Gene and Iden.fy Prenatal/Antenatal Genetic Pa.ents Carrier With Screening Increased Risk Disorders Testing 5 Couples with X linked disorders Adrenoleukodystrophy X linked mental retarda.on Day 3 biopsy DNA amplifica.on with PCR Y chromosome specific sequence iden.fied Only female embryos transferred 1990 Handyside, et al. Nature 1990;244:

18 GD is a 3 step process In vitro fer.liza.on (embryo crea.on) S.mula.on Egg retrieval Insemina.on/fer.liza.on Embryo culture Biopsy and test the DNA from the embryo (embryo tes.ng) Place the embryo s back into the uterus (embryo transfer)

19 VF (Stimulation process)

20 VF (egg retrieval process)

21 VF (insemination and fertilization)

22 VF (embryo culture and biopsy)

23 ay of Biopsy: Polar Body Blastomere (day 3) Trophectoderm (day 5-6)

24 Testing the DNA: PGD methodologies Types of analyses Short tandem repeat (PCR fragment size analysis) DNA sequencing Mul.plexing Nested PCR Whole genome amplifica.on

25 VF (embryo transfer)

26 reimplantation Genetic Screening Controversy False posi.ve rates Which pa.ents Scaleability Which plaporm

27 PGS for aneuploidy does not improve IVF success rates and may be detrimental

28 hat had limited the uccess of PGS? pact and timing of biopsy e embryo itself Mosaicism Self-correction of abnormalities its of fluorescent-in-situ-hybridization (FISH) Limited number of chromosomes Technical issues Fixation Hybridization Signal interpretation ntribution of aneuploidy to implantation failure may be overestimated

29 FISH for Aneuploidy Screening the interpretation of PGD objective? bridization for 13, 18, 21, X and Y

30 uilding Evidence for Benefit of CCS: etrospective Studies Lee E, et al. Hum Reprod PMID:

31 uilding Evidence for Benefit of CCS: rospective Studies Lee E, et al. Hum Reprod PMID:

32 rehensive Chromosome Screening (CCS) (24 osome) para.ve genomic hybridiza.on (CGH) y based CGH (acgh) microarrays n.ta.ve real-.me PCR (qpcr) GeneraCon Sequencing (NGS)?

33 elopment of rapid, real-time, quantitative based CCS High-level of consistency with SNP ar diagnoses Rapid turn around.me results in 4- hours Can prepare dried assay plates in advance Validated on trophectoderm biopsy, n single cells

34 Chemical Clinical Delivered Pregnancy Rates P<0.03 C C Scor et al FerCl Steril 2013

35 le embryo transfer is rare in the U.S. Percentage of IVF cycles with eset 0.7% 1.3% 2.0% 2.7% 3.1% 4.1% 5.5% Year Sourc

36 Delivery Rate by Age Group for Control SET versus CCS-SET rman E et al. Hum. Reprod. 2012;27: Published by Oxford University Press on behalf of the European Society of Human Reproduction and

37

38 derm Biopsy Cell Lysis & Control Group PreamplificaPon Study Group -Transfer best two blastocysts by tradiconal morphology criteria -All good-quality blastocysts un biopsy and CCS -All other384-well good-quality blastocysts Plate vitrified in pairs -Transfer single best euploid Real Pme PCR -All other euploid blastocysts vit individually Comprehensive Chromosome Screening Results Vitrify Discard PCR Heatmap Discard Transfer Single Euploid Blastocyst Selected for transfer

39 -14.8 to +14.1% (Per Protocol) Delivery Rate Per PaPent (n=175) Single euploid blastocyst transfer (N=89) Untested 2-blastocyst transfer (N=8 61% P=0.5 65% 95% CI of Differen -18.7% to +9.9% (Intent-to-Treat

40 oportion of Ongoing Pregnancies 100% Singletons Mul.ples Chance of ongoing singleton per randomized pa.ent: 61% vs. 34% (RR 1.8, 95% CI , P<.001) P<0.001 P< % 48% 0% Single euploid blastocyst transfer Untested 2-blastocyst transfer

41 livery Follow Up Forman EJ et al, Am J Obstet Gynecol, 2014

42 Healthier babies Median Birthweight: 3,317g Single euploid 2,778g 2-Blastocyst (P<0.001) Low birthweight (<2,500g): 11% (7/62) Single Euploid 33% (30/92) 2-Blastocyst (P=0.002) Untested 2-ET Groups Euploid eset Very low birthweight (<1,500g): 0% (0/62) Single Euploid 7% (6/92) 2-Blastocyst (P=0.08) Forman EJ et al, Am J Obstet Gynecol, PMID

43 uced Risk of NICU Admission Risk of NICU admission 26% (16/62) vs. 11% (7/61) P=0 Total.me spent in NICU 479 days vs. 93 days P=0.03 Forman EJ et al, AJ

44 ng euploid eset the standard of care Transfer Order in Fresh ETs at RMA-NJ 46% SET 33% 2-E 7% 14% 22% >

45 al Experience gnoses embryos 3168 transfers gesta.ons (62.1%) 2354 ongoing / delivered (72.1%) ors etraploid onosomies risomies Mean age 38.4 years 10 errors 7 found in losses 3 found in ongoing preg. m evaluated in 4 samples 100% mosaic

46 Maternal Age (years) Franasiak JM et al, FerCl Steril, 2014

47 ingle Gene PGD & CCS t RMA-NJ & Founda.on for Embryonic Competence, CCS always cluded with single gene PGD ost couples are fer.le and may have an affected child at home lec.ve single embryo transfer (eset) is recommended in all of these ases

48 onclusions s a highly accurate and effec.ve way to prevent transmission of herita ses ncurrent CCS enables effec.ve single embryo transfer in this seyng ader applica.on of gene.c counseling and screening may iden.fy more at-risk co ely to be highly cost-effec.ve vements in gene.c technology and embryology laboratory techniques improved accuracy and safety to enable effec.ve PGS for aneuploidy phectoderm biopsy mprehensive Chromosome Screening proved cryopreserva.on (vitrifica.on)

49 onclusions.ons for PGD/PGS are rapidly evolving radigm shiz toward universal CCS with single embryo transfer