infertility, in vitro fertilization (ivf) and genetic testing michele evans, m.d....
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Infertility, In Vitro Infertility, In Vitro Fertilization (IVF) Fertilization (IVF)
and Genetic and Genetic TestingTesting
Michele Evans, M.D.Michele Evans, [email protected]
Do you have any friends or family members that have struggled with infertility?
a.Yesb.No
Do you think that you are fertile?
a.Yesb.No
What percentage of the population is subfertile or infertile?
a. 1-2%b. 5-10%c. 10-15%d. 15-20%e. 20-25%
OutlineOutline InfertilityInfertility Treatment OptionsTreatment Options
In Vitro FertilizationIn Vitro Fertilization Egg FreezingEgg Freezing SurrogacySurrogacy
Genetic TestingGenetic Testing PreconceptionPreconception PreimplantationPreimplantation Prenatal Prenatal
ControversiesControversies
Normal FertilityNormal Fertility
0
0.2
0.4
0.6
0.8
1
1 2 3 4 5 6 7 8 9 10 11 12
Months of Follow Up
Cumulative Pregnancy
Rates
Hull, et al: Br Med J 1985:291;1693
Overview of InfertilityOverview of Infertility
Definition: 1 year of well-timed, Definition: 1 year of well-timed, unprotected intercourse without a unprotected intercourse without a pregnancypregnancy
10-15% of population is infertile 10-15% of population is infertile (subfertile)(subfertile)
What factor might cause problems with fertility?
a. boxer shortsb. drinking a glass of wine every dayc. eating disorderd. hiking for 30 minutes per daye. history of yeast infections
Causes of InfertilityCauses of Infertility
Female Reproductive Female Reproductive OrgansOrgans
FSH (+)
FSH=Follicle Stimulating Hormone
PhysiologyPhysiology
E2 (-)
E2=Estradiol
Developing follicle
Causes of Female Causes of Female InfertilityInfertility
OvaryOvary TubesTubes UterusUterus CervixCervix HormonesHormones ChromosomesChromosomes
Causes of Female Infertility Causes of Female Infertility - Ovary- Ovary
AGEAGE
Problems with ovulation Problems with ovulation
Premature ovarian failurePremature ovarian failure
Do women continue to produce eggs throughout their life (from puberty until death)?
a.Yesb. No
Ovary - Female AgeOvary - Female Age
Women are born with their lifetime Women are born with their lifetime egg supplyegg supply 4 million at 20 weeks gestation4 million at 20 weeks gestation 400,000 at birth400,000 at birth
100,000 eggs left at time of puberty100,000 eggs left at time of puberty Fertility initially declines at age Fertility initially declines at age 2727 Significant decline at age 37-38Significant decline at age 37-38 Rare pregnancies after age 44Rare pregnancies after age 44
Percentage of Married Percentage of Married Women Who are InfertileWomen Who are InfertileFrom 3 national U.S. surveysFrom 3 national U.S. surveys
Age (years)Age (years) InfertileInfertile
20-2420-24 7.07.0
25-2925-29 8.98.9
30-3430-34 14.614.6
35-3935-39 21.921.9
40-4440-44 28.728.7
Menken et al, Science 1989;23:1389
Prevalence of genetically Prevalence of genetically abnormal oocytes in abnormal oocytes in
infertile womeninfertile womenA
bn
orm
al
(%)
Ovary - OvulationOvary - Ovulation
Ovary - Causes of Ovary - Causes of AnovulationAnovulation
Hormone imbalanceHormone imbalance ObesityObesity AnorexiaAnorexia Significant stressSignificant stress Patients display:Patients display:
Irregular menstrual cyclesIrregular menstrual cycles Skipped cyclesSkipped cycles Minimal or absent premenstrual Minimal or absent premenstrual
symptomssymptoms
Ovary – Premature Ovarian Ovary – Premature Ovarian FailureFailure
Menopause prior to age 40Menopause prior to age 40 Decreased EstrogenDecreased Estrogen Increased FSHIncreased FSH
CausesCauses AutoimmuneAutoimmune GeneticGenetic Idiopathic Idiopathic
1-2% pregnancy rate1-2% pregnancy rate
What factor would not cause a woman's fallopian tubes to be blocked?
a. diabetesb. ruptured appendixc. endometriosisd. chlamydia infectione. tubal ligation
Causes of Female Infertility Causes of Female Infertility – Fallopian Tubes– Fallopian Tubes
Infection (chlamydia)Infection (chlamydia)
EndometriosisEndometriosis
Tubal ligation (female sterilization)Tubal ligation (female sterilization)
Open TubesOpen Tubes
Blocked TubesBlocked Tubes
Female Infertility - Female Infertility - UterusUterus
UterusUterus FibroidsFibroids PolypsPolyps
Mullerian (congenital) defectsMullerian (congenital) defects AbsentAbsent Bicornuate/SeptumBicornuate/Septum
Female InfertilityFemale Infertility
Uterine muscle Uterine muscle tumortumor
Benign (>95%)Benign (>95%)
25-30% of women25-30% of women
Normal Shape of UterusNormal Shape of Uterus
Fibroid UterusFibroid Uterus
Female Infertility - Female Infertility - UterusUterus
UterusUterus TumorsTumors
FibroidsFibroids PolypsPolyps
Mullerian defects (congenital)Mullerian defects (congenital) Absent uterusAbsent uterus Bicornuate/septateBicornuate/septate
Mullerian DefectMullerian Defect
Treatment with Treatment with HysteroscopyHysteroscopy
Treatment with Treatment with LaparoscopyLaparoscopy
Female Infertility - Female Infertility - CervixCervix
CervixCervix Post-surgicalPost-surgical
StenosisStenosis Mucus changesMucus changes
Female Infertility - Female Infertility - HormonesHormones
Endocrine abnormality (hormones)Endocrine abnormality (hormones) ThyroidThyroid ProlactinProlactin Polycystic ovary syndrome (PCOS)Polycystic ovary syndrome (PCOS)
Estrogen, insulinEstrogen, insulin Hypothalamic hypogonadismHypothalamic hypogonadism
StressStress Exercise (ballet dancer)Exercise (ballet dancer)
Other Causes of Female Other Causes of Female InfertilityInfertility
OthersOthers Chromosome abnormalitiesChromosome abnormalities
Turner’s syndrome (XO)Turner’s syndrome (XO) Androgen Insensitivity (XY)Androgen Insensitivity (XY)
Male pseudohermaphroditeMale pseudohermaphrodite Female phenotypeFemale phenotype Blind vaginal canalBlind vaginal canal Inguinal hernia (50%)Inguinal hernia (50%)
Sperm Are Also Sperm Are Also Required!!Required!!
Do men continue to produce sperm throughout their life (from puberty until death)?
a.Yesb.No
Causes of Male InfertilityCauses of Male Infertility
Abnormality in sperm productionAbnormality in sperm production
Abnormality in sperm functionAbnormality in sperm function
Obstruction in the ductal systemObstruction in the ductal system
Male Reproductive Male Reproductive OrgansOrgans
Sperm: Semen AnalysisSperm: Semen Analysis
Volume: Volume: >> 2 mL 2 mL Concentration: Concentration: >> 20,000,000 per mL 20,000,000 per mL Motility: Motility: >> 50% 50% Normal morphology: Normal morphology: >> 40% normal 40% normal
Krueger strict criteria: Krueger strict criteria: >> 14% normal 14% normal Best predictor of fertilizing ability Best predictor of fertilizing ability
Normal Sperm Normal Sperm MorphologyMorphology
Abnormal MorphologyAbnormal Morphology
Abnormal MorphologyAbnormal Morphology
SpermSperm
How many are needed for fertilization?How many are needed for fertilization? Natural conceptionNatural conception
20,000,00020,000,000 Intra-uterine inseminationIntra-uterine insemination
1,000,0001,000,000 In-vitro fertilization (IVF)In-vitro fertilization (IVF)
10,00010,000 Intra-cytoplasmic sperm injection (ICSI)Intra-cytoplasmic sperm injection (ICSI)
11
Causes of Male InfertilityCauses of Male Infertility
Abnormality in sperm productionAbnormality in sperm production
Abnormality in sperm functionAbnormality in sperm function
Obstruction in the ductal systemObstruction in the ductal system
Abnormalities of Sperm Abnormalities of Sperm ProductionProduction
GeneticGenetic Y chromosome microdeletionsY chromosome microdeletions
Damage to testes – anatomicalDamage to testes – anatomical CryptorchidismCryptorchidism VaricoceleVaricocele
InfectionInfection Mumps orchitisMumps orchitis
GonadotoxinsGonadotoxins
Abnormalities of Sperm Abnormalities of Sperm FunctionFunction
Antisperm antibodiesAntisperm antibodies Genital tract inflammationGenital tract inflammation
prostatitisprostatitis VaricoceleVaricocele Failure of acrosome reactionFailure of acrosome reaction Problems with sperm Problems with sperm
binding/penetrationbinding/penetration
Obstructions in Ductal Obstructions in Ductal SystemSystem
VasectomyVasectomy
Congenital bilateral absence of the Congenital bilateral absence of the vas deferensvas deferens
Epididymis/ejaculatory ductsEpididymis/ejaculatory ducts Congenital or acquiredCongenital or acquired
What percentage of men desire another pregnancy after having a vasectomy?
a. 1%b. 5%c. 10%d. 25%e. 100%
Male Infertility - Male Infertility - LifestyleLifestyle
TobaccoTobacco MarijuanaMarijuana AlcoholAlcohol CocaineCocaine Steroids (can be permanent)Steroids (can be permanent) HeatHeat ExerciseExercise
Infertility: Initial Infertility: Initial EvaluationEvaluation
EggsEggs OvulationOvulation Egg qualityEgg quality
SpermSperm PresencePresence QualityQuality
Gamete transport/ImplantationGamete transport/Implantation HysterosalpingogramHysterosalpingogram
Unexplained InfertilityUnexplained Infertility
Work-up is negativeWork-up is negative
15-20 % of couples15-20 % of couples
Infertility TreatmentsInfertility Treatments
Improve Timing of IntercourseImprove Timing of Intercourse Intrauterine insemination (IUI)Intrauterine insemination (IUI)
Clomiphene citrate + IUIClomiphene citrate + IUI FSH + IUIFSH + IUI
In Vitro Fertilization (IVF)In Vitro Fertilization (IVF) ““Standard” IVFStandard” IVF Egg donation + IVF Egg donation + IVF Egg Freezing + IVFEgg Freezing + IVF
Intrauterine Intrauterine Insemination (IUI)Insemination (IUI)
Goal is to Maximize the Chance of Fertilization• Increase Number of Eggs• Position Sperm Closer to Eggs
Infertility Treatment Infertility Treatment OptionsOptions
IUI, FSH or FSH + IUIIUI, FSH or FSH + IUI Patients with unexplained infertilityPatients with unexplained infertility
Serhall et al, Fertil Steril 1988;49:602
TreatmeTreatmentnt
CyclesCycles PregnancPregnancyy
Pregnancy Pregnancy per cycleper cycle
IUIIUI 3030 11 2.7%2.7%
FSHFSH 4949 33 6.1%6.1%
FSH+IUIFSH+IUI 3434 99 26.4%26.4%
FSH (+)
FSH=Follicle Stimulating Hormone
How Does Clomid Work To Increase How Does Clomid Work To Increase Egg Number?Egg Number?
E2 (-)
E2=Estradiol
Developing follicle
Clomid
FSH (+)
FSH=Follicle Stimulating Hormone
How Does Recombinant FSH Work How Does Recombinant FSH Work To Increase Egg Number?To Increase Egg Number?
E2 (-)
E2=Estradiol
Developing follicle
In Vitro FertilizationIn Vitro Fertilization
Why In Vitro Why In Vitro Fertilization?Fertilization?
InfertilityInfertility DNA TestingDNA Testing
Genetic DisordersGenetic Disorders Gender SelectionGender Selection
Non-Traditional Life StylesNon-Traditional Life Styles
Who should decide if a couple/person can have infertility treatment?
a. state governmentsb. national fertility agencies (ASRM)c. reproductive specialistsd. psychologists/psychiatristse. the couple/person
Should a 52 year old postmenopausal woman be able to use her daughter's eggs to have a child with her new 28 year oldhusband?
a.Yesb.No
Should a woman be able to have sperm extracted from her husband's newly dead body so that she can have "their child"?
a. Yes b. No
In Vitro Fertilization - In Vitro Fertilization - HistoryHistory
1978 – First “test tube” baby was born in 1978 – First “test tube” baby was born in England England
1981 – IVF in U.S.1981 – IVF in U.S.
Started with GIFT and ZIFTStarted with GIFT and ZIFT
2008 - >98% IVF with transfer of embryo 2008 - >98% IVF with transfer of embryo to uterusto uterus
In Vitro Fertilization In Vitro Fertilization (IVF)(IVF)
IVF Statistics - 2005IVF Statistics - 2005
422 U.S. programs offer IVF422 U.S. programs offer IVF
134,260 cycles of ART treatment134,260 cycles of ART treatment
9,649 donor oocyte cycles9,649 donor oocyte cycles
38,910 deliveries (birth of 52,041 38,910 deliveries (birth of 52,041 neonates)neonates)
CDC 2005 National Report
Who Needs IVF?Who Needs IVF?
Failed other treatmentsFailed other treatments Tubal damageTubal damage Significant male factorSignificant male factor Absent uterusAbsent uterus Carriers of genetic diseasesCarriers of genetic diseases Gender selectionGender selection Cancer patientsCancer patients Non-traditional lifestyleNon-traditional lifestyle
Ovarian Ovarian HyperstimulationHyperstimulation
Egg RetrievalEgg Retrieval
Good EggGood Egg
Bad EggBad Egg
FertilizationFertilization
2 Pronuclei (2PN)2 Pronuclei (2PN)
1 day after egg 1 day after egg retrievalretrieval
Day 3 EmbryoDay 3 Embryo
Pre-Implantation Genetic Testing Pre-Implantation Genetic Testing StageStage
Day 3 EmbryoDay 3 Embryo
Blastocyst – Day 5Blastocyst – Day 5
Source of Stem CellsSource of Stem Cells
Hatching BlastocystHatching Blastocyst
Embryo TransferEmbryo Transfer
Embryo TransferEmbryo Transfer
Special IVF ProceduresSpecial IVF Procedures
Assisted hatchingAssisted hatching Intracytoplasmic sperm injection Intracytoplasmic sperm injection
(ICSI)(ICSI) Preimplantation genetic Preimplantation genetic
diagnosis (PGD)diagnosis (PGD) FreezingFreezing Egg donationEgg donation SurrogacySurrogacy
Assisted HatchingAssisted Hatching
Empty ZonaEmpty Zona
ICSIICSI
ICSIICSI
ICSIICSI
What are the risks of IVF?
a. bleeding requiring blood transfusionb. pelvic infectionc. increased risk of congenital anomaliesd. multiple pregnanciese. all of the above
How many embryos are recommended to transfer into a 42 year old woman's uterus?
a. 1b. 2c. 3d. 4e. 5
How Many Embryos are How Many Embryos are Transferred?Transferred?
Related to age and embryo qualityRelated to age and embryo quality <35 = 2<35 = 2 35-37 = 2-335-37 = 2-3 38-40 = 3-438-40 = 3-4 >40 = up to 5>40 = up to 5
For patients with 2 or more failed IVF For patients with 2 or more failed IVF cycles, or a poor prognosis, can add more cycles, or a poor prognosis, can add more based on clinical judgementbased on clinical judgement
What Happens to the Other What Happens to the Other Embryos?Embryos?
Freeze EmbryosFreeze Embryos Donate For Research/Stem CellsDonate For Research/Stem Cells Embryo AdoptionEmbryo Adoption DiscardDiscard
What Would You Do With Your Embryos?
a.Freeze and Store Themb.Donate For Research (e.g., Stem Cells)c.Donate To Others For Adoptiond.Discard
IVF Success Rates - 2005IVF Success Rates - 2005
U.S. Fertility Centers From SART/CDC U.S. Fertility Centers From SART/CDC (HRC)(HRC)
Female ageFemale age <35 – 37% (43%)<35 – 37% (43%) 35-37 – 30% (36%)35-37 – 30% (36%) 38-40 – 20% (27%)38-40 – 20% (27%) >40 – 11% (18%)>40 – 11% (18%)
IVF Statistics - 2000IVF Statistics - 2000
65.1% singletons (16,533)65.1% singletons (16,533)
30.8% twins (7,817)30.8% twins (7,817)
3.9% triplets (1,000)3.9% triplets (1,000)
0.2% higher order multiples (44)0.2% higher order multiples (44)
Singleton PregnancySingleton Pregnancy
Twin PregnancyTwin Pregnancy
Triplet PregnancyTriplet Pregnancy
IVF and Multiple IVF and Multiple PregnancyPregnancy
Maternal complicationsMaternal complications Fetal complicationsFetal complications CostCost ““Selective reduction”Selective reduction” Single embryo transfer vs. Single embryo transfer vs.
success ratessuccess rates
Cost of IVFCost of IVF
IVF cycle + medications = $10,000-IVF cycle + medications = $10,000-15,00015,000
Assisted hatching = $500Assisted hatching = $500 ICSI = $1,500ICSI = $1,500 Freezing = $650Freezing = $650 Storage = $360 Storage = $360 Egg Donor = Minimum $5,000Egg Donor = Minimum $5,000 Surrogate = Minimum $10,000-15,000Surrogate = Minimum $10,000-15,000
Not Covered By Insurance In Most Not Covered By Insurance In Most States! States!
Egg DonationEgg Donation
Would you consider being an egg or sperm donor?
a.Yes
b.No
Have you ever been an egg or sperm donor?
a.Yesb.No
Egg Egg donationdonation
IVF for twoIVF for two
DonorDonor Standard controlled Standard controlled
ovarian ovarian hyperstimulationhyperstimulation
Egg retrievalEgg retrieval
RecipientRecipient Embryo transferEmbryo transfer
Who are candidates to be an egg donor ?
21-35 years old (older if a friend or 21-35 years old (older if a friend or relative)relative)
FSH <10FSH <10 Negative donorNegative donor Good health and genetic historyGood health and genetic history Preferably prior egg donation Preferably prior egg donation
experienceexperience How many eggs were produced?How many eggs were produced? Did pregnancy result?Did pregnancy result?
Current status of ART Current status of ART in the USAin the USA
SART registry, 1998
Live birth rates per transferfor fresh embryos from own anddonor eggs, by age of recipient
Egg DonationEgg Donation
Grade A: The Market for a Yale Grade A: The Market for a Yale Woman’s EggsWoman’s Eggs When a Yale undergraduate explored When a Yale undergraduate explored becoming an egg donor for a wealthy becoming an egg donor for a wealthy couple willing to pay top dollar to the couple willing to pay top dollar to the right candidate, she didn't realize how right candidate, she didn't realize how unsettling the process of candidacy unsettling the process of candidacy would prove to bewould prove to be
by Jessica Cohen by Jessica Cohen
Egg DonationEgg Donation October 23, 1999October 23, 1999 Selling Fashion Models' Eggs Online Raises Ethics Selling Fashion Models' Eggs Online Raises Ethics
IssuesIssues By CAREY GOLDBERG By CAREY GOLDBERG
CAMBRIDGE, Mass. -- To the horror and disgust of CAMBRIDGE, Mass. -- To the horror and disgust of mainstream infertility groups, a longtime fashion mainstream infertility groups, a longtime fashion photographer has begun offering up models as egg photographer has begun offering up models as egg donors to the highest bidders, auctioning their ova donors to the highest bidders, auctioning their ova via the Internet to would-be parents willing to pay via the Internet to would-be parents willing to pay up to $150,000 in hopes of having a beautiful up to $150,000 in hopes of having a beautiful child. child.
Egg Donation - Ethical Egg Donation - Ethical IssuesIssues
Egg Donor Egg Donor Known or anonymousKnown or anonymous How many times to donate?How many times to donate?
RecipientRecipient How old is too old?How old is too old?
Pregnancy in the Sixth Pregnancy in the Sixth Decade of LifeDecade of Life
USC experience: 1991-2000
77 recipients of egg donation Mean age 52.8 + 2.9 years
Of the 77 women, 42 (54.5%) had live Of the 77 women, 42 (54.5%) had live birthsbirths
45 deliveries in 42 women45 deliveries in 42 women
Paulson, Tourgeman, Boostanfar et al, JAMA 2002:228;2320.
Pregnancy in the Sixth Pregnancy in the Sixth Decade of Life: Obstetric Decade of Life: Obstetric
ComplicationsComplications Pre-eclampsiaPre-eclampsia
35%35% Background IncidenceBackground Incidence
3-10% 3-10% Gestational DiabetesGestational Diabetes
20%20% Background IncidenceBackground Incidence
5% 5%
Is 55 a “physiological limit”?Is 55 a “physiological limit”?
Marked increase in pre-eclampsiaMarked increase in pre-eclampsia
Increase in diabetesIncrease in diabetes
How old is too old?How old is too old?
Genetic TestingGenetic Testing
PreconceptionPreconception
PreimplantationPreimplantation
PrenatalPrenatal
PostnatalPostnatal
Preconception Preconception CounselingCounseling
Offered to all womenOffered to all women Prenatal vitamins – 400 micrograms Prenatal vitamins – 400 micrograms
folic acid/dayfolic acid/day Rubella immunityRubella immunity Varicella immunityVaricella immunity Rh statusRh status HIVHIV Hepatitis B screenHepatitis B screen Cystic Fibrosis screeningCystic Fibrosis screening Spinal muscular atrophy (SMA)Spinal muscular atrophy (SMA)
Preconception Preconception CounselingCounseling
Offered to certain ethnic groupsOffered to certain ethnic groups Mediterranean – thalassemiaMediterranean – thalassemia African-American – sickle-cell anemiaAfrican-American – sickle-cell anemia Caucasian/Hispanic – cystic fibrosisCaucasian/Hispanic – cystic fibrosis Ashkenazi Jews – 7 autosomal recessive Ashkenazi Jews – 7 autosomal recessive
disordersdisorders Gaucher disease (1/13), Tay-Sachs (1/30), Gaucher disease (1/13), Tay-Sachs (1/30),
Familial dysautonomia (1/30), Canavan disease Familial dysautonomia (1/30), Canavan disease (1/40), Fanconi anemia (1/89), Niemann-Pick (1/40), Fanconi anemia (1/89), Niemann-Pick disease (1/90), Bloom syndrome (1/100)disease (1/90), Bloom syndrome (1/100)
Prenatal TestingPrenatal Testing
UltrasoundsUltrasounds
Serum screensSerum screens
Chorionic villus sampling (CVS)Chorionic villus sampling (CVS)
AmniocentesisAmniocentesis
Prenatal Tests - Prenatal Tests - UltrasoundUltrasound
Nuchal translucency screeningNuchal translucency screening
Performed between 10-13 weeks Performed between 10-13 weeks gestationgestation
Screen for Down SyndromeScreen for Down Syndrome
Nuchal TranslucencyNuchal Translucency
Prenatal Tests - Prenatal Tests - UltrasoundUltrasound
Second trimester ultrasoundSecond trimester ultrasound Detailed examDetailed exam Down Syndrome, other trisomiesDown Syndrome, other trisomies Cardiac, renal, spinal, limb, brain Cardiac, renal, spinal, limb, brain
deformitiesdeformities Cleft lip/palateCleft lip/palate
Ultrasound Ultrasound AbnormalitiesAbnormalities
Ultrasound Ultrasound AbnormalitiesAbnormalities
Prenatal Tests - SerumPrenatal Tests - Serum
15-20 weeks gestation15-20 weeks gestation
Quad ScreenQuad Screen Tests for AFP, hCG, uE3 and inhibin ATests for AFP, hCG, uE3 and inhibin A Neural tube defects, Down syndrome, Neural tube defects, Down syndrome,
Trisomy 18, Abdominal wall defectsTrisomy 18, Abdominal wall defects Readjusts age-related risksReadjusts age-related risks
Prenatal TestsPrenatal Tests
Who is offered further testing?Who is offered further testing? Advanced maternal ageAdvanced maternal age Previous child or pregnancy with birth Previous child or pregnancy with birth
defectdefect Suggestive screening test resultsSuggestive screening test results Family historyFamily history
Prenatal Tests – CVSPrenatal Tests – CVS Chorionic Villus SamplingChorionic Villus Sampling
11-13 weeks gestation11-13 weeks gestation Catheter/needle biopsy of placental cellsCatheter/needle biopsy of placental cells Performed through cervix or abdomenPerformed through cervix or abdomen Can test for chromosome and gene defectsCan test for chromosome and gene defects 1-2% miscarriage rate1-2% miscarriage rate Digit/limb deficiencies (10 weeks)Digit/limb deficiencies (10 weeks)
Prenatal Tests - Prenatal Tests - AmniocentesisAmniocentesis
Performed at 15-18 weeks gestationPerformed at 15-18 weeks gestation 10 cc amniotic fluid10 cc amniotic fluid Living cells from fetus in amniotic fluidLiving cells from fetus in amniotic fluid Cells grown in lab for 1-2 weeksCells grown in lab for 1-2 weeks Results in 3 weeksResults in 3 weeks Chromosome and gene defectsChromosome and gene defects
Postnatal Testing Postnatal Testing
Most done during first day of lifeMost done during first day of life Heel stickHeel stick California (mandatory)California (mandatory)
GalactosemiaGalactosemia Hypothyroidism (congenital) Hypothyroidism (congenital) Phenylketonuria (PKU) Phenylketonuria (PKU) Sickle Cell Disease (SCD) and Sickle Cell Disease (SCD) and
Hemoglobinopathies Hemoglobinopathies + 35 Others as of July, 2005+ 35 Others as of July, 2005
Postnatal Screening – Postnatal Screening – Tandem Mass Spectrometry Tandem Mass Spectrometry
Screening ProgramScreening ProgramFatty Acid Oxidation Disorders Carnitine/Acylcarnitine Translocase Deficiency (Translocase)
Carnitine Palmitoyl Transferase Deficiency Type I (CPT-I)2
3-Hydroxy Long Chain Acyl-CoA Dehydrogenase Deficiency (LCHAD)2,4-Dienoyl-CoA Reductase Deficiency2
Medium Chain Acyl-CoA Dehydrogenase Deficiency (MCAD)Multiple Acyl-CoA Dehydrogenase Deficiency (MADD or Glutaric Acidemia-Type II)Neonatal Carnitine Palmitoyl Transferase Deficiency-Type II(CPT-II)Short Chain Acyl-CoA Dehydrogenase Deficiency (SCAD)Short Chain Hydroxy Acyl-CoA Dehydrogenase Deficiency (SCHAD)Trifunctional Protein Deficiency (TFP Deficiency)Very Long Chain Acyl-CoA Dehydrogenase Deficiency (VLCAD)
Organic Acid Disorders 3-Hydroxy-3-Methylglutaryl-CoA Lyase Deficiency (HMG)
Glutaric Acidemia-Type I (GA I)Isobutyryl-CoA Dehydrogenase DeficiencyIsovaleric Acidemia (IVA) Acute onset Chronic2-Methylbutryl-CoA Dehydrogenase Deficiency3-Methylcrotonyl-CoA Carboxylase Deficiency (3MCC Deficiency)3-Methylglutaconyl-CoA Hydratase DeficiencyMethylmalonic Acidemias Methylmalonyl-CoA Mutase Deficiency 0 Methylmalonyl-CoA Mutase Deficiency + Some Adenosylcobalamin Synthesis Defects Maternal Vitamin B12 DeficiencyMitochondrial Acetoacetyl-CoA Thiolase Deficiency (3-Ketothiolase Def.)Propionic Acidemia (PA) Acute onset Late onsetMultiple-CoA Carboxylase DeficiencyMalonic Aciduria
Amino Acid Disorders Argininemia
Argininosuccinic Aciduria (ASA Lyase Deficiency) Acute onset Late onsetCarbamoylphosphate Synthetase Deficiency (CPS Def.)2
Citrullinemia (ASA Synthetase Deficiency) Acute onset Late onsetHomocystinuriaHypermethioninemiaHyperammonemia, Hyperornithinemia, Homocitrullinemia Syndrome (HHH)2
Hyperornithinemia with Gyral Atrophy2
Maple Syrup Urine Disease (MSUD) Classical MSUD Intermediate MSUD5-Oxoprolinuria (pyroglutamic Aciduria)2
Phenylketonuria (PKU) Classical PKU Hyperphenylalaninemia Biopterin Cofactor Deficiencies (4)TyrosinemiaTransient Neonatal Tyrosinemia Tyrosinemia Type I (Tyr I)2
Tyrosinemia Type II (Tyr II) Tyrosimenia Type III (Tyr III)
Other Abnormal Profiles Hyperalimentation
Liver DiseaseMedium Chain Triglyceride (MCT) Oil AdministrationPresence of EDTA Antigoagulants in blood specimenTreatment with Benzoate, Pyvalic Acid, or Valproic AcidCarnitine Uptake Deficiency2
Preimplantation Genetic Preimplantation Genetic Screening (PGS)Screening (PGS)
Can test embryos for genetic Can test embryos for genetic abnormalities prior to implantationabnormalities prior to implantation
Uses single cell (blastomere) at 8-Uses single cell (blastomere) at 8-cell stagecell stage
Which Embryo is Which Embryo is Disease-Free?Disease-Free?
PGS – Clinical PGS – Clinical IndicationsIndications
Single gene defectsSingle gene defects Balanced translocationsBalanced translocations Advanced maternal age (aneuploidy)Advanced maternal age (aneuploidy) Repetitive IVF failureRepetitive IVF failure Recurrent pregnancy lossRecurrent pregnancy loss Embryo selectionEmbryo selection
Fluorescence in situ hybridization Fluorescence in situ hybridization (FISH)(FISH) Aneuploidy/translocations and sexing Aneuploidy/translocations and sexing
(5-9 chromosomes)(5-9 chromosomes) PCRPCR
specific single gene disordersspecific single gene disorders Gene ChipsGene Chips
many gene disorders & chromosomal many gene disorders & chromosomal abnormalities at one timeabnormalities at one time
PGS
•Achondroplasia •ADPKD1 •ADPKD2 •Adrenoleukodystroph •Age-related aneuploidies •Alpha-thalassemia •Alpha-1-antitrypsin •Alport disease •Amyloid precursor protein (APP) mutation •ARPKD •Becker muscular dystrophy •Beta-thalassemia •Charcot Marie Tooth disease •Chromosomal translocations •Congenital adrenal hyperplasia •Cystic fibrosis •Down syndrome •Duchenne muscular dystophy •Dystonia •Epidermolysis bullosa •Familial dysautonomia •Fanconi anemia •FAP •Fragile X syndrome •Gaucher disease •Hemophilia A and B •HLA genotyping •HSNF5 mutation
•Huntington disease •Hypophosphatasia •Incontinentia pigmenti •Kell disease •Klinefelter syndrome •LCHAD •Lesch Nyhan syndrome •Marfan syndrome •Multiple epiphysial dysplasia •Myotonic dystophy •Myotubular myopathy •NF1 and NF2 •Norrie disease •Osteogenesis imperfecta •OTC deficiency •P53 mutations •PKU •Retinitis pigmentosa •SCA6 •Sickle cell anemia •Sonic hedgehog mutations •Spinal muscular atrophy (SMA) •Tay-Sachs disease •Tuberous sclerosis •Turner syndrome •Von Hippel Lindau •X-linked hydrocephaly •X-linked hyper IgM syndrome
PGS for Single Gene PGS for Single Gene Disorders - AdvantagesDisorders - Advantages
Safer than elective terminationSafer than elective termination More psychologically acceptable for More psychologically acceptable for
couplescouples Provides couples with another optionProvides couples with another option
AdoptionAdoption SterilizationSterilization Donor gametesDonor gametes
PGSPGS Pre-implantation Pre-implantation
genetic screening genetic screening (PGS)(PGS) has been has been successfully used in successfully used in diagnosing and diagnosing and preventing inherited preventing inherited genetic diseases like genetic diseases like Cystic Fibrosis, Tay Cystic Fibrosis, Tay Sach’s, Thalassemia, Sach’s, Thalassemia, Sickle Cell Anemia Sickle Cell Anemia and may be potentially and may be potentially used to screen for used to screen for cancer mutations.cancer mutations.
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PGSPGS
After a cycle of After a cycle of in- vitro in- vitro fertilization, fertilization, biopsy of a single biopsy of a single cell can be cell can be performed from performed from an 8 cell embryo an 8 cell embryo obtained after 3 obtained after 3 days of culture in days of culture in the laboratory.the laboratory.
PGS – Timing of BiopsyPGS – Timing of Biopsy
<67 hours post-<67 hours post-retrievalretrieval
Implantation rates Implantation rates significantly lower if significantly lower if >70 hours>70 hours
Probably represents Probably represents technical issues with technical issues with compacting embryocompacting embryo
PGSPGS
The genetic The genetic material of this material of this single cell can be single cell can be amplified by PCR amplified by PCR and the and the chromosomal chromosomal mutation or an mutation or an aneuploidy can be aneuploidy can be identified in the identified in the embryo that embryo that underwent a biopsy.underwent a biopsy.
PGSPGS The embryos would The embryos would
continue to grow for continue to grow for 2 more days in the 2 more days in the laboratory, awaiting laboratory, awaiting genetic analysis, genetic analysis, and confirmation of and confirmation of which embryos were which embryos were unaffected with the unaffected with the mutation or mutation or aneuploidy.aneuploidy.
PGSPGS
The unaffected The unaffected embryos are then embryos are then transferred to the transferred to the uterus at the uterus at the blastocyst stage on blastocyst stage on day 5 of embryo day 5 of embryo culture and culture and subsequently a child subsequently a child would be born would be born unaffected from the unaffected from the screened genetic screened genetic disease.disease.
Five Chromosome PGDFive Chromosome PGD
PGS – Female EmbryoPGS – Female Embryo
Uses fluorescence Uses fluorescence in-situ in-situ hybridization hybridization (FISH) technique (FISH) technique to identify XXto identify XX
Sex-linked diseasesSex-linked diseases
““Family balancing”Family balancing”
PGS – Male EmbryoPGS – Male Embryo
Uses FISH to Uses FISH to identify XY embryoidentify XY embryo
If you had one or more children of the same sex, would youlike to be able to choose the gender of your next child?
a.Yesb.No
Prenatal vs. Prenatal vs. Preimplantation DiagnosisPreimplantation Diagnosis
PNDPND PGS PGS
CellsCells >100,000 >100,000 11
Time Time 2 weeks2 weeks 6-10 hrs6-10 hrs
AccuracyAccuracy 99%99% 99%99%
CostCost Covered Covered ~$5,000 ~$5,000
Who Would Benefit Who Would Benefit From PGS?From PGS?
Couples with a history of --Couples with a history of -- Abnormal numbers of Abnormal numbers of
chromosomeschromosomes Single gene disordersSingle gene disorders Balanced translocationsBalanced translocations
Couples who --Couples who -- Desire an offspring of a certain sex Desire an offspring of a certain sex
Future considerationsFuture considerations
Oocyte cryopreservationOocyte cryopreservation ““Pausing the biological clock”Pausing the biological clock”
Cytoplasmic transferCytoplasmic transfer Donation of enucleated oocytesDonation of enucleated oocytes
Reproduction without gametesReproduction without gametes Use of nuclear material from somatic Use of nuclear material from somatic
cellscells Donated or synthetic cytoplasmDonated or synthetic cytoplasm Reconstituted oocytesReconstituted oocytes
Questions??Questions??
Sperm donorSperm donor Female coupleFemale couple Huntington’s diseaseHuntington’s disease Single womanSingle woman Sex-linked diseaseSex-linked disease Family balancingFamily balancing ““Wrongful death” of discarded Wrongful death” of discarded
embryoembryo Implantation of the wrong embryoImplantation of the wrong embryo
Thank Thank youyou