Lyndsey Craven Convegno Mitocon 2015

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


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



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



• 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


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


• 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


• 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


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

Lyndsey Craven Convegno Mitocon 2015

Health & Medicine

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