Genetics and Reproductive Options for SMA Families2018 Annual SMA ConferenceDallas, TexasFriday, June 15, 2017
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Part 1: SMA and GeneticsLouise R Simard, PhD
Part 2: SMA Carrier ScreeningMelissa Gibbons, MS CGC
Part 3: Reproductive Options for SMA Families
Harvey J Stern, MD, PhD
Genetics and Reproductive Options for SMA Families
Part 1: SMA and Genetics
Louise R. Simard, PhDUniversity of Manitoba, Faculty of Medicine
Dept. Biochemistry & Medical GeneticsWinnipeg, MB, Canada R3E 0J9
1987 - Ph.D. U Toronto – Medical Genetics. 1988-2006 – Ste. Justine Hospital Research Centre, U Montreal
Independent research program in SMADNA diagnostics Consultant, incl. SMA testing
2006-present – Professor, Department Head, U Manitoba
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Questions are welcome!
Genetics and Reproductive Options for SMA Families
Outline• SMA
Lower Motor Neuron Disease With a wide range of clinical presentation (severity)
• SMA Gene SMN1 and SMN2 SMN1 Mutations SMA severity
• SMA Genetics Chromosomes as vehicles of the DNA genomic blueprint Autosomal Recessive Inheritance
• SMA Molecular Tests Diagnosis Carrier
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Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
SMA is a lower motor neuron disease that recurs in families (inherited).
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Type 2Intermediate SMA
< 18 monthsNever stand unaided
Decreased life expectancy
Type 3Kugelberg-Welander
18 monthsStand alone but loss of mobility
Normal life expectancy
Type 4 Adult SMA≥21 years of age
Progressive muscle weaknessNormal life expectancy
Type 1Werdnig-Hoffman
< 6 monthsNever sit unaided
Death before 2 years
Type 0Prenatal SMA
In uteroRespiratory support
Death before 1 month
Disease Severity
01
23
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Wide ClinicalPresentation!
Image adapted from from“Principles of Neural Science”, ER Kandel, JH Schwartz, TM Jessell,
eds., 4th Ed. McGraw Hill.
Loss of MN cells leads to Muscle Atrophy
Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
Identifying the SMA gene 1 gene 5 SMA types
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Nature (1990) 345:823-825Nature (1990) 540-541
Nature (1990) 344:767-768 Lancet (1990) 336:271-273
Lefebvre et al., (Melki laboratory, INSERM Institut Necker)
Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
Genetic Code = Blueprint of life~20,000 genes in the human genome
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DNA = four nucleic acids (C, G, T, A)Different sequence of A-C-T-G = geneC (cytosine) pairs with G (guanine)T (thymine) pairs with A (adenine)
CGT
AT A
A
TCG GC
CG
GC
AT TA
www.photoresearchers.com
1953
Part 1: SMA and Genetics
Geno
me
Deco
ratio
n Pa
ge/N
CBI
https://autisable.com/
Genetics and Reproductive Options for SMA Families
SMN1 vs. SMN2 gene Chr 5q13
1 2 3 4 5 6 7 8Promoter
CSTOP
1 2 3 4 5 6 7 8Promoter
TSTOP
1 2 3 4 5 6 8
STOP
Start
STOP
1 2 3 4 5 6 7 8
Start
DNA (a)
RNA (b)
mRNA (c)
proteinunfunctionalprotein
functionalprotein
~10% Full-length transcripts
~90% Nonfunctional SMN protein
100% Full-length transcripts
Functional Wild Type SMN protein
> 99 % identical
SMN2 SMN17T
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C
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http://www.bbc.com/news/science-environment-27592655
Mutations in SMN1Cause 5q-SMA
Genetics and Reproductive Options for SMA Families
Deletions (missing DNA) is a common mutation in SMA
SMN1 7 8
Very large deletions are often associated with type I and type II
SMA. Type I SMAs most often cannot make ANY SMN.
In Type II & III SMA, the mutation is often confined to SMN1 exon 7.
% p
atie
nts
Simard et al., (1997) Am J Med Genet 72:51-58
0
10
20
30
40
50
60
70
80
90
100
type I type II type III
SMN null SMN E7
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Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
SMA locus is very variable
But…also find:
SMN2 SMN1
SMN2 SMN1On average:
SMN1
SMN2 SMN1SMN2
SMN1SMN1No apparent Clinical effect
SMN2
SMN2 SMN1 SMN2SMA causing
Chromosomes
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Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
SMN2 is a major modifier geneSMN2 copy number decreases with increasing severity
0
10
20
30
40
50
60
70
80
90
1 2 3 4
type I
type II
type III
% p
atie
nts
# SMN2 copies
n=188n=110n=77
Taken from Feldkotter et al., (2002) Am J Hum Genet 70:358-368
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Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
SMA is inherited as an Autosomal Recessive Disorder
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The fundamental unit of inheritance is DNA.Each cell has over 5 feet of DNA! It must be compacted to fit inside a cell in the form of chromosomes.
We have 23 pairs of chromosomes; each set is inherited from our biological parents.
www.sciencemuseum.org.uk
SMA autosomal recessivemode of inheritance
********************************************There must be a mutation (error) in both copies of a gene (maternal and paternal) for the genetic disorder to be passed onto a child.
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https://www.biography.com/.image/t_share/MTIwNjA4NjMzNTA5MDg2NzMy/gregor-mendel-39282-1-402.jpg
Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
SMA is inherited as an Autosomal Recessive Disorder
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• Affected individuals have mutations in both copies of a recessive gene.
• Carriers have a mutation in only one copy of a recessive gene and is asymptomatic.
• Males and females can be carriers.
• Males and females can be affected.
• If both parents are carriers, the risk of having a child with the disease is 25%.
Part 1: SMA and Genetics
Having an affected child is a surprise because carrier parents are
asymptomatic. From this point onwards – genetic
counseling becomes possible.
1 in 4 Chance25%
1 in 4 Chance25%
1 in 2 Chance (50%)
Taken from ghr.nlm.nih.gov
Genetics and Reproductive Options for SMA Families
>95% of patients have deleterious mutations in the SMN1 gene(deletions, gene conversions, point mutations)
SMN1 7SMN27T C
X
point mutation
deletion
gene conversion SMN1 SMN2SMN27T T
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Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
Molecular Diagnostic Tests
• Detection of homozygous mutations presence/absence of SMN1 Ex7 = diagnosis
• Detection of heterozygotes Number of copies of exon 7 = carrier testing
• Analysis of point mutations = diagnosis Not yet routine, specialized labs only
SMN2 SMN17T
7
C
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Part 1: SMA and Genetics
Genetics and Reproductive Options for SMA Families
(164 pb)SMN2(187 pb)SMN1
Van der Steege et al. (1996) Am J Hum Genet 58:834-838.
All or None test!
Diagnostic Tests: SMN1 mutationsDiagnosis of SMA patients
Compared to EMG & muscle biopsy, the DNA test is:• non-invasive• sensitive, detects >95% of all SMAs• accurate, all lacking SMN1 exon 7 have SMA• ideal for newborn screening
Limitation: Some SMA individuals have a point mutation in the SMN1 gene. Such mutations are not detected by the deletion or quantitative PCR assay.
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Part 1: SMA and Genetics
Taken from Passon et al. (2010) Mol Cell Probes 24:310
Genetics and Reproductive Options for SMA Families
Diagnostic Tests: Carrier Detection2 vs. 1 copy of SMN1 exon 7 test (dosage)
• 95% of SMA patients are lacking SMN1 exon 7
• Because SMA is an autosomal recessive disorder, most SMA carriers should have only 1 copy of SMN1 exon 7
SMN1
SMN2SMN1:SMN2 copies
2:1 2:21:31:20:4 0:4 0:3
McAndrew et al. (1997) Am J Hum Genet 60:1411-1422 Rochette et al., (1997) Neurogenetics 1:141-147.
Detection of heterozygotesSMN quantitative test
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Part 1: SMA and Genetics
Taken from Passon et al. (2010) Mol Cell Probes 24:310
Genetics and Reproductive Options for SMA Families
Diagnostic Challenges Rarely able to give a 100% Yes or No result.
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Part 1: SMA and Genetics
These tests do not detect small mutations in the SMN1 gene.
Taken from Alías et al. (2009) Human Genetics 125:29
2 to 5% of individuals have both SMN1 genes on the same chromosome
(2+0 genotype) - so are carriers
On average:SMN2 SMN1
SMN2 SMN1
SMN2
SMN2 SMN1 SMN1
But….
Genetics and Reproductive Options for SMA Families
A Genetic Counselor is a valuable resource to help you understand the genetics of SMA, opportunities for testing (diagnosis, carrier status, prenatal) and how to interpret test
results.
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Part 2: SMA Carrier ScreeningMelissa Gibbons, MS CGCCertified Genetic Counselor
Neuromuscular Clinic Children’s Hospital Colorado
Assistant ProfessorUniversity Colorado School of Medicine
MemberCure SMA Medical Advisory Committee
Genetics and Reproductive Options for SMA Families
Genetic Counselors are:
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• Master’s-trained health care professionals who combine their knowledge of: ―Basic science―Medical genetics―Epidemiological principles―Counseling theory
• With their skills in: ―Genetic risk assessment―Education―Interpersonal communication and counseling
• To provide services to clients and their families for a diverse set of genetic or genomic indications
Genetics and Reproductive Options for SMA Families
Genetic Counselors work in:
• Clinical Setting―Preconception―Prenatal―Pediatric―Adult―Cancer
• Genetic Testing Laboratory• Pharmaceutical Companies
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Genetics and Reproductive Options for SMA Families
Genetic Counselor can:
• Explain your/your child’s results• Discuss different genetic testing options• Assist with coordinating carrier testing for
―You―Extended Family
• Provide family education • Discuss treatment options• Discuss reproductive options
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Genetics and Reproductive Options for SMA Families
Understanding Your Results
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http://www.curesma.org/documents/support--care-documents/genetics-of-sma.pdf
Genetics and Reproductive Options for SMA Families
Understanding Your Results
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Positive Deletion Results:
Inconclusive Deletion Results:
Additional Types of Sequencing Results:
Sequencing Results:
Carrier Testing Affected Individual has
2 SMN1 Deletions
Parents and Extended Family can be tested by
Deletion Studies
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Affected Individual has1 SMN1 Deletion &
1 SMN1 Point Mutation
Parents should be tested first to determine
targeted testing for the extended family
Genetics and Reproductive Options for SMA Families
Different Types of Carriers“1 + 0”
“2 + 0”
5q
5q
SMN1
5q
5q
SMN1SMN1
5q
5q
SMN1 SMN1
5q
5q
SMN1SMN1
SMN1
*“2 + 0*”
*Point mutation or microdeletion.Adapted from PMCID: PMC3234503
“1 + 1*”*
Genetics and Reproductive Options for SMA Families
What does negative mean?
Genetics and Reproductive Options for SMA Families
What does negative mean?g.27134T>G
Genetics and Reproductive Options for SMA Families
Options for Carrier Testing • Single gene testing
―Testing for one condition―Also referred to as known familial mutation testing
• Targeted carrier screening―Testing based on your ethnicity or family history―Also referred to as ethnic-based carrier screening
• Expanded carrier screening―Many disorders are screened for using a single sample―Test selection is done without regard to race or ethnicity―Each company determines the genes on their panel ―Screening panels usually focus on severe disorders that
affect a person’s quality of life from an early age
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Genetics and Reproductive Options for SMA Families
Genetics and Reproductive Options for SMA Families
Carrier Testing
• Allows for more informative preconception counseling
− Reproductive risks for the couple− Discussion of preimplantation/prenatal testing options− Development of PGD for your family
• Provides information to family members− Who should be tested− What testing do they need
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Genetics and Reproductive Options for SMA Families
Resources
• Genetic Counselors on the Cure SMA Medical Advisory Board
− Jin Yun (Helen) Chen, Massachusetts General Hospital− Khalida Liaquat, Quest Diagnostics− Melissa Gibbons, Children’s Hospital Colorado
• National Society of Genetic Counselors―NSGC.org
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Part 3: Reproductive Options for SMA Families
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Harvey J. Stern MD, PhDDirector, Reproductive GeneticsGenetics & IVF Institute
Genetics and Reproductive Options for SMA Families
Workshop GoalsWhat are the genetic aspects of SMA especially as they relate to
reproduction?
What are the reproductive choices for families at-risk for SMA?
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RECESSIVE DISEASE
Genetics and Reproductive Options for SMA Families
Very difficult decisions about family building
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Reproductive Options for SMA Families
Genetics and Reproductive Options for SMA Families
Reproductive Options for SMA Families
• Decisions regarding childbearing are personal and reflect our own ethical, moral and religious views. This is not a “one size fits all” type of issue.
• Couples who are at-risk for a genetic disorder should be allowed to make up their own minds after considering all their reproductive options.
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Genetics and Reproductive Options for SMA Families
Reproductive Options for SMA Families
• Decisions are influenced by the couple’s experience with SMA. Those who have had a child or sib affected will react differently from a couple identified to be at-risk by genetic screening.
• Decisions are also influenced by couple’s attitudes towards prenatal diagnosis and assisted reproduction (“playing God”)
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Genetics and Reproductive Options for SMA Families
Reproductive Options for SMA Families
Most families do not wish to bring another child with SMA into the world.
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Genetics and Reproductive Options for SMA Families
• Have no or no more children (Most Common Choice)
• Adoption• Use a gamete donor (pre-tested for SMA)
Reproductive Options for SMA Families
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Genetics and Reproductive Options for SMA Families
Reproductive Options for SMA Families
Natural conception with 1st or 2nd
trimester prenatal testing.Preimplantation Genetic Diagnosis
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Genetics and Reproductive Options for SMA Families
What is PGD?
• Involves the use of assisted reproduction technologies (IVF) to provide a method of prenatal diagnosis.
• Offers an alternative to traditional methods of prenatal diagnosis including chorionic villous sampling and amniocentesis, with the option of termination of affected pregnancies.
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Genetics and Reproductive Options for SMA Families
Development of PGD
• This year represents the 28th anniversary of the first application of PGD in humans.―(Handyside et al. Nature 244 1990)
• Since 1990, approximately 400,000 PGD cycles have been performed worldwide.
• This has resulted in over 100,000 PGD babies born after the procedure.
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Transfer unaffected embryos to the patient
affected affectedaffected
PGD for Genetic Disease
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In-Vitro Fertilization (IVF)
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Genetics and Reproductive Options for SMA Families
Questions About PGD
Why do I need to do IVF, I have no trouble getting pregnant?
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Genetics and Reproductive Options for SMA Families
Questions About PGD
• IVF is used to increase the odds of producing a healthy child.
• There is no change in the genetic material (egg or sperm).
• By creating multiple embryos, one increases the chance that good quality embryos which are not affected with SMA will be present and available to be returned to the uterus.
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IVF Procedure
Genetics and Reproductive Options for SMA Families
IVF/PGD Procedure
Components of IVF/PGD Cycle:1. Down-regulation (preparation)2. Ovarian Stimulation - FSH3. Egg Retrieval4. Fertilization (ICSI)5. Embryo biopsy and testing6. Embryo transfer to uterus
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Genetics and Reproductive Options for SMA Families
Parts Of An IVF Cycle
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Genetics and Reproductive Options for SMA Families
Normal Ovulation
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Embryo DevelopmentDay 2Day 3Day 4Day 5
Fertilization
2 cell4 cell
8 cell
Morula
Blastocyst
Adapted from Moore and Persaud, Saunders, 1993
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IVF: Monitoring Visit
0
500
1000
1500
2000
2500
3000
0 2 4 6 8 10 12 14
Estradiol levels
Follicle measurement
Days of FSH Stimulation
Genetics and Reproductive Options for SMA Families
Stimulated Ovary
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Egg Retrieval
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Egg Aspiration
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Egg Ready For Fertilization
Fertilization by IntracytoplasmicSperm Injection
ICSI
ICSI60
Embryo Development
2 cell
3 cell
4 cell
8 cell
61From: Veeck LL, An Atlas of Human Gametes and Conceptuses, Parthenon Publishing, 1999
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Embryo Development
Morula stageembryos
Blastocyst stage embryos
From: Veeck LL, An Atlas of Human Gametes and Conceptuses, Parthenon Publishing, 1999
Embryo Transfer
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Genetics and Reproductive Options for SMA Families
PGD for Single Gene Disorders
• Analysis involves whole genome amplification with multiplex PCR amplification of the mutation along with 3-4 linked polymorphic markers which generate a chromosomal haplotype.
• At the same time, the embryos are also tested by a chromosomal microarray for abnormalities (ie. Down syndrome) which can lead to failed implantation or pregnancy loss.
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Genetics and Reproductive Options for SMA Families
PGD for SMA
EmbryoBiopsy
SMN gene
SMN 1 & 2 = unaffected
SMN 2 only = affected
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NO SMN1Affected
SMN1 PresentSMN 1 & 2
Genetics and Reproductive Options for SMA Families 70
24 Chromosome Analysis by MicroarrayPerformed concurrently with SMA testing to identify the embryos with the best potential to make a baby as well as being free of SMA.
Genetics and Reproductive Options for SMA Families
Accuracy of PGD
• Using the combination of mutational analysis and linked markers the accuracy of the test 98-99%.
• Contamination with external DNA is a major concern.
• Analyses done in surgical clothing in biohazard hood.
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Genetics and Reproductive Options for SMA Families
Specific Issues with IVF/PGD
• Patient Discomfort: Medications are given by injection (subQ).
• Abdominal distention and discomfort and some nausea are common.
• Egg retrieval done under anesthesia.
• Ovarian hyperstimulation syndrome 1-2%.
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Genetics and Reproductive Options for SMA Families
Questions about IVF/PGD
• Is IVF Safe?• What will I feel?• Does it increase cancer risk?• Are the babies born normal?• What are the costs?• What about insurance?
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Genetics and Reproductive Options for SMA Families
Questions About IVF/PGD
What is the success rate of IVF/PGD?• Varies, particularly with maternal age, but in
large series 50-60% of patients became pregnant per IVF/PGD cycle.
• Improved with concurrent testing for chromosome abnormalities.
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Thank you!!!
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