Transcript
Mitochondrial Donation: are we ready?
Lyndsey CravenWellcome Trust Centre for
Mitochondrial Research
Molecular genetic basis of mitochondrial disease
Nuclear DNA: 3,000,000,000 bp mtDNA: 16,569 bp
>300 different disease-causing mutations
paternal mitochondria eliminated after fertilisation
Mitochondrial Genetics: maternal inheritance
egg
sperm
fertilisation
nuclear DNA
mitochondria
female pronucleus
male pronucleus
fertilised egg
Fertilisation
Primordial germ cell containing mutant mtDNA
Primary oocytes
Oocyte maturation and
mtDNA amplification
Mature oocytes
High level of mutation (affected offspring)
Low level of mutation (unaffected offspring)
= normal mitochondria
= mutant mitochondria
Intermediate level of mutation (mildly
affected offspring)
Mitochondrial Genetics: the bottleneck
Mitochondrial DNA Disease
Estimated that 2473 women of child bearing age are at risk of transmitting mtDNA disease in the UK
Currently no effective treatments for mtDNA disease
Several options for women with mtDNA mutations:- genetic counselling- oocyte donation- prenatal diagnosis- preimplantation genetic diagnosis- mitochondrial donation?
14-20 weeks
Amniocentesis
11-13 weeks
Chorionic Villus Sampling (CVS)
Prenatal Diagnosis
Preimplantation Genetic Diagnosis• Genetic testing of cells removed from oocytes or
early embryos• Requires in vitro fertilisation (IVF) procedures:
ovarian stimulation
oocyte retrieval fertilisation
embryo culture
embryo transfer
Unfertilised oocyte
Pronucleate stage zygote
Cleavage stage embryo
Blastocyst stage embryo
Day 1 Day 3 Day 5
PGD in Newcastle• PGD licence obtained in 2009• 14 PGD cycles performed to date for 6 different
mtDNA mutations:m.3688G>A m.8993T>Gm.9176T>C m.3243A>Gm.10158T>C m.8344A>G
• Successful pregnancy achieved in 3 PGD cycles• Embryos with low level of mutation selected for
transfer• Low levels of mtDNA mutation confirmed in the child
Preimplantation Genetic Diagnosis
• PGD for mtDNA disease is useful but does have limitations– not suitable for women with homoplasmic mutations– not suitable for women who produce only embryos with
high mutation loads
• Urgent need to develop other IVF-based techniques to prevent mtDNA disease– Mitochondrial donation
Mitochondrial Donation: experimental strategy
donor egg
mutant mitochondrianormal mitochondria
affected egg
Mitochondrial Donation: experimental strategy
mutant mitochondrianormal mitochondria
nuclear DNA removed
nuclear DNA removed
donor egg
affected egg
Mitochondrial Donation: experimental strategy
mutant mitochondrianormal mitochondria
nuclear DNA removed
nuclear DNA removed
donor egg
affected egg
Mitochondrial Donation
• Technique that replaces mutant mtDNA in an egg or embryo
• Involves transfer of nuclear DNA to a healthy egg or embryo
• Transfer can be performed at different stages
Maternal Spindle Transfer (MST)
Pronuclear Transfer (PNT)
Polar Body Transfer
Greggains et al, Sci Rep. 2014 Jan 24;4:3844
Mitochondrial Donation
Mitochondrial Donation: pronuclear transfer
Mitochondrial Donation: pronuclear transfer
mutant mitochondrianormal mitochondria
pronuclei removed
pronuclei removed
donor embryo
affected embryo
• Last hot cycle PCR-RFLP used to determine mtDNA carryover in abnormally fertilised PNT embryos
• Average level of mtDNA carryover was <2%
• Most embryos had undetectable levels
• Suggests that PNT has the potential to prevent transmission of mtDNA disease in humans
Mitochondrial Donation: pronuclear transfer
Mitochondrial Donation: pronuclear transfer
• PNT now being optimised at Newcastle Fertility Centre using normally fertilised human embryos– oocytes from research egg donors
• Important to determine safety and efficacy PNT– embryo development – embryo quality – genetic analysis: mitochondrial DNA
nuclear DNA
Mitochondrial Donation: legal issues
• September 2005: HFEA granted a research licence to allow the development of techniques to prevent mtDNA disease in human embryos
• November 2008: Following about 2 years of debate, thorough revisions to the HFE Act were passed by a significant majority in both Houses of Parliament
• These included a specific provision to allow for regulations to be passed by Parliament to permit the clinical application of “techniques that alter the DNA of an egg or embryo to prevent the transmission of serious mitochondrial disease”
• February 2011: HFEA working group (“core panel of experts”) commissioned by Department of Health to perform a scientific review of the safety and efficacy of methods to avoid
mitochondrial disease
Mitochondrial Donation: safety issues
Mitochondrial Donation: safety issues
• The evidence currently available does not suggest that the techniques are unsafe
• Minimum set of experiments proposed before techniques considered for clinical use
• Separate scientific reviews in 2013 and 2014 reached the same conclusion
• January 2012: Government asked the HFEA to seek public views on IVF techniques designed to prevent the transmission of mitochondrial disease
• July to August 2012: public dialogue work takes place- deliberative public workshops- public representative survey
• September to December 2012: open consultation runs- open consultation questionnaire
- open consultation meetings
- patient focus group
Mitochondrial Donation: public consultation
Public Consultation • September 2012: online public consultation launched
Public Consultation • March 2013: results of public consultation
“General support for permitting mitochondrial replacement in the UK”“Overall view is that ethical concerns are outweighed by the arguments in favour of permitting mitochondrial replacement”
Mitochondrial Donation: ethical issues
“Due to the health and social benefits to individuals and families of living free from mitochondrial disorders, and where potential parents express a preference to have genetically-related children, on balance we believe that if these novel techniques are adequately proven to be acceptably safe and effective as treatments, it would be ethical for families to use them, if they wish to do so and have been offered an appropriate level of information and support.”
March 2013: Mitochondrial Replacement Consultation, the Scientific Update, HFEA’s Advice and a Report on Practical Implications are all published and submitted to Government
June 2013: The UK Government (via the CMO) announces their decision to draft regulations to amend the HFE Act to permit the clinical application of mitochondrial donation
Mitochondrial Donation: legal issues
Public Consultation • February 2014: public consultation on draft regulations launched
• July 2014: results of public consultation published• Government made the decision to proceed with putting
regulations before Parliament• December 2014: regulations were laid before Parliament
Mitochondrial Donation: legal issues
• 3rd February 2015: debate and vote in the House of Commons to decide whether to approve the draft Regulations to allow mitochondrial donation
The House of Commons voted by
382 to 128 in support of the Regulations
Mitochondrial Donation: legal issues
• 24th February 2015: debate and vote in the House of Lords to decide whether to approve the draft Regulations to allow mitochondrial donation
The House of Lords voted by 280 to 48 in
support of the Regulations
Mitochondrial donation into law in the UK on the 4th March 2015
Mitochondrial Donation: legal issues
Next Steps
• Detailed regulations now need to be agreed and adopted by the HFEA• Licence applications to treat “patients” need to be submitted and approved. Approval will only be granted if the HFEA are satisfied that the science continues to suggest that the methods are safe and efficient• Clinics applying for a licence must comply with the regulations in terms of scientific and clinical practice, including patient support • Long term follow-up of children
AcknowledgementsMitochondrial Research GroupDr Helen TuppenDr Julie MurphyDr Bobby McFarlandProf Rob TaylorProf Doug TurnbullNewcastle Fertility CentreSam ByerleyLouise HyslopProf Alison MurdochProf Mary Herbert
Newcastle Fertility Centre @
Diabetes
ThyroidMuscle – pain and weakness
Nerves
Hearing
Brain and Central Nervous system
Respiratory muscles Eye and optic nerve
Heart
Bone marrow,
blood cells
Liver
NeurologicalNon-Neurological
Mitochondrial Genetics: multicopy genome
Mitochondrial Genetics: maternal inheritance
Mitochondrial Replacement: House of Commons debate
• September 2014: debate on mitochondrial replacement techniques and public safety
Mitochondrial Replacement: evidence hearing
• October 2014: Science and Technology Committee hold a one-off evidence session on mitochondrial replacement
• Science and Technology Committee considered the science of the new techniques and subsequently urged the Government to proceed with regulations
• December 2014: regulations were laid before Parliament
Mitochondria are the powerhouses of the cell
Mitochondrial Donation — How Many Women Could Benefit?
Mitochondrial Donation — How Many Women Could Benefit?
• We estimated that 2473 women of child bearing age are at risk for transmitting mtDNA disease nationally
• The average number of births per year among women at risk for transmitting mtDNA disease is 152 in the United Kingdom
• The average number of births per year among women at risk for transmitting mtDNA disease is 191 in Germany
14-20 weeks
Amniocentesis
11-13 weeks
Chorionic Villus Sampling (CVS)
Prenatal Diagnosis
Prenatal Diagnosis
Homoplasmic wild-type
Homoplasmic mutant
HETEROPLASMIC
Mitochondrial Genetics: heteroplasmy
wild-typephenotype
mutantphenotype
Mitochondrial Genetics: threshold effect
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