The first results from mitochondrial replacement, which some refer to as ‘3-person IVF’, in normally fertilized human embryos has been reported in Nature. To remove disease-causing mitochondria, UK researchers transplanted nuclear DNA from a fertilized egg cell into a donor cell with healthy mitochondria via a technique called ‘pronuclear transfer’. The embryos, which were not allowed to develop, retained some mutant mitochondrial DNA, but the researchers improved their technique to reduce mutant DNA to levels at which a child would be unlikely to develop mitochondrial disease.
Dr. Carlos Moraes, Lichtenstein Professor of Neurology, Cell Biology and Anatomy, University of Miami Miller School of Medicine (webpage):
Expertise: The molecular basis of mitochondrial disorders; pharmacological and genetic approaches to treat mitochondrial diseases.
“Pronuclear transfer to prevent transmission of mitochondrial DNA (mtDNA) with disease-causing mutations has become a proxy for a new era, when genetic manipulation of human embryos is not only possible but allowed. The approach is relatively simple and the benefits obvious. The pronucleus of a fertilized egg cell is removed and placed into a donor cell, where the nucleus had been removed and discarded. However, this relatively simple approach, even if already performed in several mammals (e.g. mouse, monkeys, and to a lesser extent in human embryos) is fraught with technical issues that have major consequences on the final outcome.
“In this recent report in Nature, Hyslop and colleagues went through the nuts and bolts of the procedure to maximize production of healthy and viable embryos after pronuclear transfer. They found that transplanting the pronucleus at an early stage resulted in improved survival of the embryo. They also optimized removal of the pronucleus and embryo culture procedures which yielded better quality blastocysts (later embryos) whose gene expression and incidence of chromosome abnormalities did not differ from controls. Another important practical finding is that donor egg cells should not be preserved by a process known as vitrification, as it increased the carryover of mtDNA attached to the transplanted pronucleus.
“These improvements suggest that early pronuclear transfer (ePNT) has the potential to reduce risk of mitochondrial disease in a child born through this procedure. However, it cannot exclude the possibility that a few reconstituted embryos will carry some levels of the mutant mtDNA. This word of caution has also been given in a recent paper in Cell Stem Cell* by the group of Dieter Egli. Therefore, the authors suggest prenatal screening should continue in ePNT, until more is known about the outcomes. Although incremental, these technical enhancements led to the production of healthy embryos, and likely healthy children in the not so distant future.”
Dr. Marni Falk, Assistant Professor of Pediatrics, Division of Human Genetics, The Children’s Hospital of Philadelphia (webpage):
Expertise: Mitochondrial disease; the genetic basis of mendelian disease; models of mitochondrial disease and therapy.
“This work is important to the field as it moves closer toward having safe methods to perform pronuclear transfer (PNT) with the ultimate goal of implantation in mothers at risk of transmitting mitochondrial disease. It reports results of rigorous methodological analyses to determine how to optimize the safety and efficacy of PNT.
“Whereas prior studies were done with abnormal embryos, the use of normal embryos revealed some key issues that had to be resolved, providing critical information to begin to work toward clinical use of mitochondrial replacement therapy. The investigators learned that PNT must be performed much earlier than anticipated (8 hours after fertilization rather than 24 hours). Further, many of the technical manipulations and basic culture conditions had to be substantially modified to improve their ability to consistently obtain good-quality embryos with low-levels of mutant mitochondrial DNA (mtDNA) carried over with the transplanted nuclear DNA from the intended parents.
“The levels of carryover mtDNA with their modified technique were consistently below 2% in the majority (79%) of the blastocyst-stage embryos they generated. However, when human embryonic stem cells were derived from those blastocyst-stage embryos, one of five cell lines developed much higher levels of mutant carryover mtDNA, much like results seen in the recent Cell Stem Cell paper* that performed similar studies.
“Prior studies have suggested that a child is unlikely to have mitochondrial disease if there is less than 5% of the mutant mtDNA present in the early embryo. Since the investigators ultimately achieved that goal with their revised methodology in all of the blastocyst-stage embryos they tested, they concluded that PNT has the potential to reduce the risk of mitochondrial disease. However, as they correctly acknowledged it may not guarantee prevention, since there is the chance that even small levels of mutant mtDNA can become unpredictably enriched in the developing person over time.”
Dr. Gerald Shadel, Professor of Pathology and of Genetics, Yale School of Medicine (webpage):
Expertise: The role of mitochondria in disease, aging and the immune system.
“I am not surprised by the need to tweak the precise timing and mode of the pronuclear transfers to obtain optimal results. The authors show that, even with this change in protocol, there is a small amount of mitochondrial DNA carryover with the transfers, which is one of the key remaining issues with the technique. If inherited, in rare cases this could still potentially lead to a disease outcome by genetic drift mechanisms, as proposed in the recent study* by Hirano, Egli and colleagues.
“Based on the results of the current study, it does seem the carryover can routinely be reduced to less than 2%, making the chances of disease occurring substantially less, but unfortunately still not zero. However, with the risk being very low, I would liken the situation to a potential parent carrying a rare recessive nuclear disease gene who has to be counseled that there is a higher risk that their child could inherit the disease if the other parent is also a carrier.
“Thus, I view this current report as a positive step forward in bringing this mitochondrial replacement technique into the clinic, but better understanding is needed of the precise probabilities of mitochondrial disease inheritance if carryover cannot be eliminated.”
*‘Genetic Drift Can Compromise Mitochondrial Replacement by Nuclear Transfer in Human Oocytes’. GENeS published comments on this paper which can be viewed here.
Declared interests (see GENeS register of interests policy):
‘Towards clinical application of pronuclear transfer to prevent mitochondrial DNA disease‘ by Hyslop et al, published in Nature on Wednesday, June 8, 2016.