MICA: Identification of age-related and age-independent changes to meiotic chromosome structure and their association with aneuploidy in human oocytes

It has long been known that as a woman ages, her likelihood of having a chromosomally abnormal pregnancy rises, with the risk increasing exponentially in women over the age of 35. This means that many couples who delay conception until they are financially secure suffer subfertility and repeated pregnancy loss, causing distress to the couples, a financial burden on the NHS and economic consequences through sickness absence. This is especially significant in countries such as the UK where the number of women giving birth over the age of 35 has doubled in the last 20 years (Office of National Statistics, 2009 data). Curiously, the vast majority of chromosome problems originate from errors in chromosome segregation in the mother's egg. Correct segregation of pairs of chromosomes is essential so that the embryo eventually contains a complete set. Precisely why the fidelity of chromosome segregation decreases so abruptly in eggs from older women is unclear, but mounting evidence from mouse and human studies suggests that a multitude of factors, both environmental and genetic, contribute to faulty chromosome segregation in the egg. One major reason thought to underlie the high error-rate in eggs is the relatively long time period taken to complete female meiosis; the cellular division that halves the number of chromosomes prior to fertilisation. In contrast to sperm production in males, which initiates in puberty and takes approximately 64 days to complete, the process in females is initiated in the ovary of the developing fetus and is not completed until ovulation, which may be several decades later. It is now widely believed that this extended period before completion of meiosis may account for the high error-rate in eggs ovulated in older women.

How may these time-dependent changes in chromosome segregation be explained? One popular hypothesis to explain the so-called 'maternal age effect' is that proteins in the egg, responsible for holding together chromosome pairs together, erode over time. This has been referred to as 'becoming unglued'. One set of proteins, known as Cohesins work together with exchanges of the DNA to form cross-over points that hold pairs of chromosomes together until they separate at the first meiotic division. Cohesins certainly appear to be depleted from chromosomes in aged mouse eggs (compared to younger mice), but it is yet to be seen whether the same is true for humans.

Since a subset of the missegregating chromosome pairs are in fact 'non-exchange' pairs, (having no cross-over points), deterioration of cohesin should not, in theory, affect their segregation. Instead, other mechanisms must be invoked to explain their missegregation. Synaptonemal Complex (SC) proteins have been implicated in the segregation of non-exchange chromosomes in many other organisms, so it is possible that deterioration of SC proteins may also be associated with errors in chromosome segregation in human eggs. In support of this hypothesis is the observation that SC proteins are associated with chromosomes in human sperm during both meiotic divisions, way beyond their known role in chromosome synapsis.

In this work, we plan to use 'surplus' human eggs that cannot be used for IVF treatment to investigate age-related (as well as age-independent) changes to chromosome structure- do they become 'unglued' with time? Our work will begin by examining changes to Cohesins and Synaptonemal Complex proteins, although a broader study including many more candidate proteins is planned for the long term. Through this work we hope to gain valuable insights into the chromosomal basis of human female fertility and the 'maternal age effect', which can be used to help inform women's reproductive choices.

Changes to chromosome structure during the prolonged meiotic arrest of human oocytes have been proposed to underlie age-related aneuploidy of human foetuses. Age-related aneuploidy poses major risks to reproductive health. The goal of this project is to characterise chromosomal changes directly in human oocytes from women of different ages. We focus on several important themes based upon studies in model organisms, aiming to increase knowledge of the mechanisms responsible for oocyte aneuploidy directly in human. In possession of this knowledge, future development of clinical pre-gestational tests for aneuploidy risk will become a possibility. In this work, we shall test whether meiotic chromosome structure deteriorates with advanced maternal age in human oocytes, such as precocious separation of homologous chromosomes or sister chromatids. Such structural changes would strongly predispose chromosomes to missegregate at meiosis I or II, resulting in chromosomally imbalanced oocytes. We will also study the localization pattern of several candidate protein complexes to examine whether their localisation or relative abundance correlates with such aberrant chromosome structures. These candidate complexes include cohesin, condensin, synaptonemal complex and DNA repair factors; all implicated as major factors in chromosome structure and segregation from studies from model organisms. To achieve our objectives, we have assembled a world-class cross-disciplinary team of discovery scientists, statisticians, embryologists, as well as clinicians. We propose a logical and realistic plan of work, governed by high ethical standards and rigorous methodology to address important scientific and clinical questions.

The proposed research addresses directly in human oocytes fundamental, basic biological mechanisms that have been proposed from studies in model organisms to underpin age-related and age-independent infertility in humans. This is an area with substantial implications and impact to screen women and inform clinical practice. For example, recurrent miscarriage (three or more) accounts for 1-4% of infertile couples. Observational studies have shown that in couples with recurrent miscarriage about half the pregnancy losses are from karyotypical abnormalities. Understanding whether the depletion of cohesin or synaptonemal complex components correlate with age-related and/or age-independent infertility is important as they could be used as biomarkers for general chromosomal integrity in oocytes used for IVF/ICSI. Specifically, SYCP3 mutations has been correlated in some, but not all, population-wide studies of recurrent miscarriage. Evaluating the localization of SYCP3 at centromeres and correlating the presence or depleted levels with precocious sister kinetochore separation and/or univalent formation will provide a mechanistic model on which the role of SYCP3 could be evaluated directly in human oocytes.

The ultimate beneficiaries of our research will be girls and women who are considering having children, planning their families, and who will increasingly become aware of the risks associated with delaying conception. Our research will add specific knowledge about certain biomarkers that might have clinical relevance to reproductive health and can provide diagnostic tools with a view to future personalised risk assessments. The increasing knowledge we will provide, of the underpinning molecular mechanisms that are responsible for the age-dependent deterioration of oocyte quality, will inform women's options. In the medium term, measurement of markers in an ovarian biopsy or at IVF may allow an assessment of oocyte competence or 'oocyte age' for an individual woman. In the far future, it might become possible to ameliorate the age effect, allowing women to conceive when they want to, without increased age-associated risks, throughout their whole reproductive lifespan, however, this will not be possible without a detailed understanding of the molecular mechanisms involved. A thorough discussion of the consequences of such a profound biological and social change would also be essential, and would trigger extensive public debate and expert opinion.

Affluent countries such as UK, already have significant age-associated infertility and associated reproductive morbidity such as miscarriage and increased maternal risks. Moreover, the NHS burden of prenatal screening and sometimes termination of aneuploid pregnancies (such as Trisomy 21 Down Syndrome) is expensive and potentially traumatic. Developing nations are tending to experience the same demographic trends, showing increasing maternal age to be a significant global issue. Contributions to improving knowledge of age-related changes in human oocytes have potential for impact on a global scale.
Our research therefore has potential to increase future health and wealth, and to change current ways of life. Given the large ongoing research effort in this area, our project's specific impact is likely to be as a defined contribution focused on translation of the animal-based molecular markers towards assessing human relevance and promoting clinical application.