Mitochondrial disorders represent the most common group of inborn errors of metabolism. Clinical manifestations can be extremely variable, ranging from single affected tissues to multisystemic syndromes. Maternally inherited mitochondrial DNA (mtDNA) mutations are a frequent cause, affecting about one in 5000 individuals. The expression of mtDNA mutations differs from nuclear gene defects. Mutations are either homoplasmic or heteroplasmic, and in the latter case disease becomes manifest when the mutation load exceeds a tissue-specific threshold. Mutation load can vary between tissues and in time, and often an exact correlation between mutation load and clinical manifestations is lacking. Because of the possible clinical severity, the lack of treatment and the high recurrence risk of affected offspring for female carriers, couples request prevention of transmission of mtDNA mutations. Previously, choices have been limited due to a segregational bottleneck, which makes the mtDNA mutation load in embryos highly variable and the consequences largely unpredictable. However, recently it was shown that preimplantation genetic diagnosis offers a fair chance of unaffected offspring to carriers of heteroplasmic mtDNA mutations. Technically and ethically challenging possibilities, such maternal spindle transfer and pronuclear transfer, are emerging and providing carriers additional prospects of giving birth to a healthy child.
Mitochondrial disorders represent the most common group of inborn errors of metabolism. Clinical manifestations can be extremely variable, ranging from single affected tissues to multisystemic syndromes. Maternally inherited mutations in the mitochondrial DNA (mtDNA) are a frequent cause, affecting about one in 5000 individuals. The expression of mtDNA mutations differs from nuclear gene defects. Mutations are either homoplasmic (100% mtDNA mutated) or heteroplasmic (mixture wild-type and mutated mtDNA) and in the latter case disease become manifest when the mutation load exceeds a tissue-specific threshold. Mutation load can vary between tissues and in time, and often an exact correlation between mutation load and clinical manifestations is lacking. Because of the possible clinical severity, the lack of treatment and the high recurrence risk of affected offspring for carrier females, couples request to prevent transmission of mtDNA mutations. For many years, choices have been limited due to a segregational bottleneck, which makes the mtDNA mutation load in embryos highly variable and the consequences largely unpredictable. However, recently it was shown that preimplantation genetic diagnosis (or embryo selection) offers carriers of heteroplasmic mtDNA mutations a fair chance of unaffected offspring. New technically and ethically challenging possibilities, such as maternal spindle transfer and pronuclear transfer, in which the nuclear DNA is transferred from an oocyte or zygote containing mutated mitochondria into an enucleated oocyte or zygote containing normal mitochondria, are emerging at the horizon and are providing carriers with additional prospects of giving birth to a healthy child.
Mitochondrial or oxidative phosphorylation disorders are complex diseases, caused by mutations in either nuclear genes or in the mitochondrial DNA (mtDNA). Nuclear gene defects segregate as Mendelian diseases, whereas mtDNA defects are transmitted maternally. The latter occurs in about 15% of the cases, affecting about one in 5000 individuals (Rotig and Munnich, 2003). Carrier frequency for pathogenic mtDNA mutations in the population is higher – from 1:400 (Manwaring et al., 2007) to even >1:200 (Elliott et al., 2008) – but in general the mutation load remains below the level of clinical expression. Still, symptoms such as hearing loss can be present and undiagnosed individuals can turn out to be oligosymptomatic upon further investigation (Manwaring et al., 2007). Mitochondrial diseases can manifest with symptoms in many different organs and vary profoundly in severity and age of onset (reviewed in McFarland et al., 2010). Clinical manifestations may present in just a single affected tissue or organ, such as the loss of vision in Leber’s hereditary optic neuropathy (LHON), but a multisystemic or multiorgan involvement is more common. The clinical spectrum (Chinnery and Hudson, 2013) involves the brain (ataxia, dementia, migraine, myoclonus, neuronal loss and stroke), the peripheral nervous system (neuropathy), the heart (cardiomyopathy, conduction disorders, Wolfe-Parkinson-White syndrome), skeletal muscle (fatigue, myopathy, weakness), the liver (hepatopathy), the pancreas (diabetes), the eyes (optic neuropathy, ophthalmoplegia, retinopathy), the ears (sensori-neuronal hearing loss), the kidney (Fanconi syndrome, glomerulopathy), the colon (pseudo-obstruction), the blood (Pearson syndrome) and the gonads (ovarian failure). Well-known neurological syndromes, caused by mitochondrial dysfunction and partly due to mtDNA mutations, are Leigh syndrome (subacute necrotizing encephalomyelopathy), mitochondrial encephalopathy, lactic acidosis and stroke-like episodes (MELAS syndrome), neuropathy, ataxia and retinitis pigmentosa (NARP syndrome) and myoclonic epilepsy with ragged red fibres (MERRF syndrome). Fatally affected newborns represent the severe end of the spectrum. A frequent symptom in paediatric patients is developmental delay and failure to thrive. When at least two organ systems unexplained by other diseases are involved in a single person or in affected (maternal) relatives, then mitochondrial disorder must be considered. Clinicians should be aware that apparently unrelated symptoms might have a common genetic cause (McFarland et al., 2010). Given the potential for severe clinical disease in a child, the ability to prevent transmission of these inherited disorders using reproductive technology is highly desirable.
Section snippets
Mitochondrial DNA
The first description of a circular DNA located in the mitochondria dates from more than 40 years ago (Nass, 1966). The mtDNA has a number of unique characteristics that discriminates it from its nuclear counterpart. First, the mtDNA is a double-stranded circle (16,569 bp) with a structure and code different from the nuclear DNA. The mtDNA contains 37 genes, of which 13 genes encode OXPHOS subunits and 22 tRNA and two rRNA genes. Approximately 6% of the mtDNA is noncoding, located predominantly
mtDNA defects and recurrence risks
Disease causing mutations in the mtDNA can be due to large rearrangements, point mutations or a reduced copy number, in some cases leading to depletion of the mtDNA. One has to be aware that a mtDNA defect can be the primary cause of disease, but that it also can be the manifestation of nuclear gene defects (e.g. defects in genes involved in mtDNA maintenance causing multiple mtDNA deletions and/or mtDNA depletion), mitotoxic drugs (e.g. nucleoside reverse transcriptase inhibitors can induce
Preventing the transmission of mtDNA diseases
A number of approaches exists to prevent the transmission of mtDNA disorders, all having their specific advantages and disadvantages and their technical and ethical constraints (Poulton et al., 2010). The remainder of this paper mainly concentrates on methods to prevent the transmission of point mutations in the mtDNA, which have been demonstrated to be prime causes of disease in affected family members and which are not secondary to nuclear or environmental factors.
Oocyte donation
The use of donor oocytes with spermatozoa of the partner is a reliable method to prevent the transmission of mitochondrial disease caused by mtDNA mutations, with the drawback that the resulting child is only genetically related to the father and not to the mother. However, the woman who carries and gives birth to a child will be recognized as its legal mother. The use of donor oocytes of close maternal relatives is not advisable in the case of familial disease since these may carry the same
Prenatal diagnosis
Conventional prenatal diagnosis (PND) during pregnancy on chorionic villi or amniotic fluid cells has its own complexity for heteroplasmic mtDNA mutations. Although the mutation load can be determined accurately in the DNA collected, it is the sampling and the interpretation which makes PND problematic. A number of criteria have been proposed to allow reliable PND in mtDNA disease (Poulton and Turnbull, 2000). First, the distribution of mutant mtDNA in all extra-embryonic and fetal tissues
Preimplantation genetic diagnosis
Preimplantation genetic diagnosis (PGD) is an alternative to PND. Oocytes are fertilized in vitro and cells, usually from the 8-cell embryo, are biopsied and tested for the presence of a genetic defect. Unaffected embryos are transferred into the uterus. PGD avoids the dilemmatic choice of a pregnancy termination, which is a major advantage compared with PND in mtDNA disorders, which often results in multiple affected or ambiguous pregnancies. A technical advantage of PGD for mtDNA diseases is
Nuclear genome transfer
Nuclear genome transfer (Figure 3) involves the transfer of the nuclear genome from an oocyte or zygote with mutated mtDNA in the cytoplasm (donor) to an enucleated acceptor oocyte or zygote of a healthy donor (acceptor) with presumably normal, mutation-free mtDNA (Craven et al., 2011). Consequently, the offspring would not carry the mtDNA mutation present in the mother and would not suffer from the familial mtDNA disease. (Immature) spindle–chromosome transfer is carried out at the level of
Conclusions and future prospects
At this moment the transmission of mtDNA disease due to heteroplasmic mutations can be effectively stopped in many cases, although this awareness is often lacking among clinical geneticists and other clinicians involved. PND is a reliable option for mothers of a child with a de-novo mutation and carriers of recurrent mutations, which fulfil established criteria. PGD is the preferred option for reliable risk reduction for most carriers of heteroplasmic mutations. The use of PGD to select
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Pathological manifestations of mtDNA mutations are highly dependent on the level of heteroplasmy with symptoms occurring if a certain threshold is passed, a phenomenon called the “threshold effect” (Saneto and Sedensky, 2013). mtDNA mutations are maternally inherited and asymmetric segregation followed by clonal expansion of mtDNA during oocyte maturation can lead to a random shift of mutational load between generations (Smeets, 2013; Stewart and Larsson, 2014). More than 150 pathogenic mtDNA mutations have been described in humans; these are associated with severe multi-systemic diseases, which currently have no treatment options (Park and Larsson, 2011; Wallace, 2010).
Mitochondrial diseases are very heterogeneous in their genetic cause and clinical manifestation. During the last few decades progress has been made in the diagnosis of mitochondrial diseases, but an established therapy is so far lacking. Several experimental strategies targeting different points of intervention are currently being assessed world-wide. Numerous mouse models of OXPHOS disorders have become available enabling further optimization and validation of therapeutic strategies and paving the way for future clinical trials. In this review, we provide an update on current developments towards treatment as well as the potential and status of transition into therapeutic use.
Recently, several publications have surfaced describing methods to manipulate mitochondrial genomes in tissues and embryos. With them, a somewhat sensationalistic uproar about the generation of children with ‘three parents’ has dominated the discussion in the lay media. It is important that society understands the singularities of mitochondrial genetics to judge these procedures in a rational light, so that this ongoing discussion does not preclude the helping of patients and families harboring mutated mitochondrial genomes.
As noted above, considerable heterogeneity exists in diagnostic criteria, treatment protocols, definitions of clinical variables and data collection systems, and it contributes to signal noise. Reducing this heterogeneity will enhance the ability to find true predictive signals amidst a large number of potential markers (Smeets, 2013). As the field progresses to overcoming these scientific challenges and the genetic factors associated with female infertility disorders are elucidated, it will be important to stay mindful of the host of ethical considerations that will emerge.
Significant progress has been made in several fields of medicine towards personalizing treatment recommendations based on individual patient genotype. As the number of clinical and genetic biomarkers available to physicians has increased, predictive models able to integrate the contributions of multiple variables simultaneously have become valuable tools for medical decision making. Leveraging genotype information and multivariate predictive models holds the promise of bringing greater efficiency to, and reducing the costs of, fertility treatments. This work reviews the advances that have been made in genetic biomarker discovery and predictive modelling for fertility treatment outcomes. We also discuss some of the limitations of these studies for translation to clinical diagnostics and the challenges that remain.
Personalized medicine holds the promise of allowing doctors to create ‘bespoke’ treatment recommendations for each patient based on multiple clinical variables such as age and hormone concentrations combined with the patient’s genetic sequence information. A number of challenges remain for the field of reproductive medicine to make the research discoveries necessary to usher in this new era of personalized fertility care. Here, we discuss some of these challenges and make recommendations for overcoming them.
It took the skilled eye of an experienced clinician to recognize that the inheritance pattern of this special form of diabetes was strictly via the female line. Careful pedigree analysis of the proband indicated that only affected women, and not men, transmitted the disease to all their children, both sons and daughters, but only daughters were seen to transmit the disease again to their children (Smeets, 2013; Van den Ouweland et al., 1992). The year 1992 saw 37 publications on mitochondrial diabetes.
In assisted reproduction, there is strong evidence for some things done, but no or only very weak evidence for others. There are several reasons for this. Most assisted reproduction procedures have small signal-to-noise ratios. This means that their treatment effect is sometimes only little better than the spontaneous conception rate, or the conception rate with traditional treatment. Hence, large trials are required. These demand complex multicentre logistics. The latter require substantial funding and funding for reproductive medicine in most countries is notoriously difficult to obtain (as opposed, for example, to oncology research or cardiovascular research). Apart from these funding issues, the creation of embryos specifically for research is only allowed in a limited number of European countries, thus tempting clinicians to skip preclinical studies altogether and go directly for clinical application in their patients, raising an ethical issue. Introducing new treatments into the clinic without proper evidence, however, is perhaps even more of an ethical issue. Subfertile couples are very vulnerable and should not be exploited.
In assisted reproduction, we have strong evidence for some things we do, but no or only very weak evidence for others. There are several reasons for this. Most assisted reproduction procedures have small signal-to-noise ratios. Which means that their treatment effect is sometimes only little better than the spontaneous conception rate, or the conception rate with traditional treatment. Hence large trials are required. These demand complex multicentre logistics. The latter require substantial funding and funding for reproductive medicine in most countries is notoriously difficult to obtain (as opposed to, for example, oncology research or cardiovascular research). Apart from these funding issues, the creation of embryos specifically for research is only allowed in a limited number of European countries, thus tempting clinicians to skip preclinical studies altogether and go directly for clinical application in their patients, an ethical issue. Introducing new treatments into the clinic without proper evidence, however, is perhaps even more of an ethical issue. Subfertility couples are very vulnerable, they should not be exploited.
Bert Smeets, PhD, is Professor in Clinical Genomics with a focus on mitochondrial disorders at Maastricht UMC, The Netherlands, combining research with genetic testing services. His research focuses on the genomics of mitochondrial disorders and involves identifying the genetic defect, studying the pathophysiology, characterizing new treatment options and preventing the transmission, the latter by preimplantation genetic diagnosis. He has published over 170 original research articles, reviews and book chapters (Hirsch index = 40). His group contains 60 people involved in clinical genomics research and services and exploits central genomics facilities for Maastricht UMC and adjoining universities from Belgium and Germany.
