VictorJuiz/thesis huggingface dataset:
Cytogenetic and epigenetic analysis of human gametes and preimplantation embryos

Despite major advances in the assisted reproductive treatments, it remains the case that most embryos transferred to the uterus fail to implant or are lost shortly after. Chromosome aneuploidy has been characterised as one of the major factors impacting embryo viability.
While the incidence of whole chromosome aneuploidies has been well characterised, little is known about segmental aneuploidy affecting fragments of chromosomes. One of the aims of this study was to investigate the frequency, origin and clinical relevance of segmental aneuploidy in human oocytes and preimplantation embryos. The incidence of segmental aneuploidy significantly increased following fertilisation, peaking at the cleavage stage and declined during the transition to the blastocyst stage. Unlike whole chromosomal aneuploidy, occurrence of segmental aneuploidy was not correlated with advancing female age. Interestingly, IVF culture conditions such as media systems and gas composition were found to significantly influence the incidence of segmental aneuploidy. In addition, the study found that embryos generated by patients suffering from recurrent implantation failure had a higher proportion of segmental aneuploidies than those generated by patients undergoing IVF treatment for advanced maternal age or recurrent miscarriage, suggesting an underlying patient factor.
Preimplantation genetic testing for aneuploidy (PGT-A), using embryo biopsies, is routinely employed during in vitro fertilisation to select against aneuploid embryos. However, it is speculated that the invasive nature of the embryo biopsy procedures can adversely impact embryo viability. Therefore, non-invasive sources of embryonic DNA, including blastocoel fluid and spent culture media, were explored during the course of this study and a non-invasive PGT-A method was successfully developed. Promising results were obtained following the application of the developed non-invasive assay to spent culture media samples, showcasing its potential for future clinical application.
Although not as well characterised as aneuploidy, epigenetics is a key emerging factor impacting embryo development. Of particular interest to the IVF community is the epigenetic phenomenon of genomic imprinting wherein a gene is monoallelically expressed in a parent of origin fashion. A few studies have shown a significantly higher rate of imprinting disorders such as Beckwith Wiedemann syndrome in children born after ART as compared to those conceived naturally. Hence, the last part of this study aimed to investigate the imprinting landscape in preimplantation human embryos. A multiplex PCR based bisulfite sequencing approach was successfully developed which was further utilised to study imprinting patterns of 23 imprinted genes simultaneously in trophectoderm and inner cell mass biopsies from euploid human blastocysts. To our knowledge, this is the first study to successfully analyse the methylation patterns of a large number of imprinted genes simultaneously in human blastocysts.

Advisor: Wells, D
Characterising the determinants of hypoxia inducible transcription factors binding to chromatin

Hypoxia regulates many hundreds of genes that play important roles in numerous physiological and pathophysiological processes. The hypoxia inducible transcription factors (HIFs) are central to the transcriptional activation of these hypoxia-regulated genes. However, to date, little is known about the determinants of HIF-1 and HIF-2 binding site selection. Both HIF-1 and HIF-2 appear to bind as HIF-α/HIF-1β heterodimers, and recognise the same core consensus DNA motif, the hypoxia response element (HRE). However, each has its own distinct but partially overlapping set of binding sites that accounts for only a subset of the total accessible HRE motifs. Here, I have utilised ChIP-seq to systematically compare pan-genomic HIF-1α, HIF-2α and HIF-1β DNA binding in multiple cell lines and have related this to RNA-seq analyses and other publically-accessible next-generation sequencing datasets. I show that endogenous HIF-α subunits exhibit a high-degree of binding site concordance with HIF-1β, consistent with largely canonical binding of intact heterodimers. Despite cell-type specific differences in HIF-1 and HIF-2 binding site occupancy, each isoform exhibits a remarkable rigidity in its preference to bind either promoter-proximal or promoter-distal sites, respectively. These specific distribution patterns are unaffected by the absence of the other HIF-α subunit, suggesting that they do not result from exclusion of one isoform by competition with the other, but rather are discrete properties of each. Furthermore, hypoxia regulated genes neighbouring sites that are shared by both HIF-1 and HIF-2 are more likely to be regulated by HIF-1 when the site is closer to the gene and by HIF-2 when further away, indicating that post-binding mechanisms of transcriptional regulation also follow a similar pattern. Comparison of sites preferentially bound by HIF-1 and HIF-2, respectively, revealed associations with distinct histone environments, distinct accessory transcription factor binding motifs and distinct patterns of transcription factor binding site occupancy, suggesting that each may be influencing specific HIF-1 and HIF-2 binding site selection. In particular, both the AP-1 motif and AP-1 binding site occupancy were enriched within HIF-2 binding sites compared to HIF-1 sites. Intervention on AP-1 DNA-binding using the dominant-negative protein, AFos, attenuated HIF binding, specifically at sites co-occupied by AP-1 and HIF. This indicates that a cooperative relationship exists between the two transcription factors. However, binding of both HIF-1α and HIF-2α were affected suggesting that while AP-1 binding may account for the ability of HIF to bind some HRE motifs but not others, it is not a determinant of differential binding between the two isoforms. Overall, this work reveals remarkably distinct and functionally relevant patterns of HIF-1 and HIF-2 binding across the genome, and provides insight into underlying mechanisms of binding.

Keywords: Transcription
Gene regulation
Hypoxia
Molecular biology
Advisor: Ratcliffe, P
Mole, D
Mathematical modelling of tumour evolution and radiation response: The impact of heterogeneity

This thesis seeks to use mathematical and computational models to develop measures of clinically available data to deepen our understanding, and improve our treatments, of cancer. We consider two broad characteristics of cancer: heterogeneity, in the form of differences in cellular phenotype, and the physical microenvironment; and evolution, which has become accepted as a driver of tumour progression. To ensure that the conclusions drawn are as translatable as possible, we will attempt to use data types that are clinically available. Using a hybrid discrete-cell-based model in two spatial dimensions, we focus on these fundamental aspects of cancer, with the hope of generating new understanding and useful hypotheses to benefit current patients and oncologists.
First, we model a tumour growing under the rules of the cancer stem cell hypothesis and a neutral model of evolution, and ask if we can infer the underlying biological proliferative structure. Specifically, we work toward predicting the symmetric division probability of our simulated tumours from clincally relevant observables, as this is a key driving parameter of tumour progression and therapeutic response. We focus on measures of clonal diversity, group size and shape, and a suite of statistical measures of the phylogenetic trees resulting from the tumour's evolution in different regions of parameter space. We find strikingly different patterns in these measures for changing symmetric division probability which hinge on the inclusion of spatial constraints. These results give us insight into differences between solid and liquid tumours, and also generate a number of actionable clinical and biological hypotheses.
Second, we explicitly consider the physical microenvironment of tumours invading into healthy tissue, and model oxygen transport, uptake and cellular competition. We then explore the effect of spatial organisation of blood vessels within the tumour on tumour growth kinetics and cellularity. Finding wide variability in the distribution of oxygen across tumours dependent on both vascular organisation and density, we proceed to explore the utility of spatial measures of vessels on radiation efficacy. Our results offer a novel hypothesis as to the failure of vascular normalisation therapy and radiation, and a possible clinical solution.

Keywords: cancer
radiation therapy
mathematical oncology
Advisor: Maini, P
Anderson, A
Fletcher, A
Development of cation-directed acylation reactions

This thesis describes work conducted towards the enantioselective acylation of ketones using phase-transfer catalysis. Chapter 1 introduces the field of ketone acylation, with a focus on catalytic methods accompanied by the generation of axes, planes or points of chirality during these transformations. Chapters 2 describes the synthesis and evaluation of a variety of different frameworks for enantioselective ketone O-acylation; we highlight the selectivity and reactivity challenges faced when attempting this transformation (Scheme I).
Chapter 3 focuses on the intramolecular O-acylation of enolates to generate axially chiral lactones. The development of these systems led to the discovery that the C/O-selectivity of this reaction was dependent on the steric hindrance about the axis of rotation, and C-acylation could be favoured under the correct conditions.
Chapter 4 describes the conception and development of an enantioselective C-acylation reaction, culminating in the first reported catalytic enantioselective direct C-acylation of a ketone (Scheme II). The scope and limitations of this methodology were investigated and the products were successfully elaborated to afford a range of spirocyclic architectures.
Finally, chapter 5 focuses on the kinetic analysis of the enantioselective C-acylation reaction in order to increase our understanding of the mechanisms behind phase-transfer catalysis and the mode of activation of the catalyst.

Advisor: Smith, M
Clark, H
Dwelling encounters: Migration, diversity and ambivalence in an Istanbul neighborhood

This thesis is about processes and lived experiences of inhabiting urban contexts deeply and continuously impacted by migration driven population changes. It builds on an ethnographic study of a neighbourhood in Istanbul known as Kumkapı, which over recent decades has emerged as a zone of arrival and initial settlement for successive new waves of migrants coming from an ever more expanding geography. Consequently, today it stands as one of Istanbul’s most diverse residential neighbourhoods, with its population differentiated on innumerable fronts including ethnicity, religion, race, gender and age composition, legal rights and statuses, migration channels and intentions, employment opportunities, and the like. In this dissertation I apply a novel theoretical approach drawing on the concept of dwelling for understanding how residents of Kumkapı relate to their environment and make sense of it. I propose three lines of argument that unfold sequentially through the organizing structure of the thesis. Firstly, I argue that comprehending urban contexts of migration driven diversity through a dwelling lens allows one to recognize the different temporalities that are at play in shaping the present moment. There are multiple pasts and futures inhabiting the present, shaping material forms, daily rhythms, systems of differentiation, and socialization patterns. Secondly, I argue that a dwelling lens positions the private sphere at the centre of diversification processes and recognizes space beyond its containing capacities. The ethnography explores the diverse reasons, conditions and temporalities of inhabiting Kumkapı today and how this diversity in turn leads to a breadth of residential practices. Thirdly, I argue that dwelling in urban contexts of migration driven diversity is very often characterized by a deep sense of ambivalence and continuous acts of balancing to cope with these conflicting factors simultaneously infringing upon people's lives. In building these arguments, the thesis draws together migration and urban research both within and outside anthropology in a novel way, while also contributing to scholarly debates on various themes including, home and housing, ethnicity, race, gender, informality and conviviality.

Advisor: Keith, M
Duvell, F
Hole extracting layers in perovskite solar cells: influence on performance and stability

As one of the most promising technologies for highly efficient and low-cost photovoltaics, hybrid organic-inorganic metal halide perovskites solar cells have received a great deal of attention over the past 5 years. However, the moisture sensitivity of this material hinders commercial deployment, and the next grand challenge is now the long-term, stable operation of such devices. This thesis presents novel hole-extraction layers for perovksite solar cells, and studies their impact on both the performance and the long-term stability of the devices.
Initially, this work focuses on the development of a polymer composite hole transporting material (HTM) based on poly(3-hexylthiophene) nanowires (P3HT-NW) embedded in an inert poly(methyl methacrylate) (PMMA) matrix. This HTM gives superior efficiency when compared to a neat P3HT HTM, and outperforms conventional HTM systems after thermal stressing in air. Next, the commonly employed and unstable lithium bis(trifluoromethanesulfonyl)imide (Li-TFSI) p-dopant for 2,2’,7,7’-tetrakis(N,N’-di-p-methoxyphenylamine)-9,9’-spirobifluorene (spiro-OMeTAD), is successfully replaced by two molybdenum tris(dithiolene)s. These devices show a substantial enhancement of the long-term stability, due to a significant enhancement of the moisture resilience of the HTM layer while maintaining high device power conversion efficiencies. Finally, the interaction of these molybdenum complexes with the perovskite is studied. This study demonstrates that the molybdenum complex modifies the treated interface in two ways; it passivates surface defects and creates an interfacial dipole. As a result, the perovskite/HTM interface shows greater hole selectivity resulting in a substantial enhancement of the device performance, underscoring the importance of the charge-selective interfaces for carrier extraction.

Advisor: Snaith, H
Clinical characteristics, virology and host genetic markers of Hand, Foot and Mouth Disease in Southern Vietnam

Hand, foot and mouth disease (HFMD) has been associated with large outbreaks among young children, including severe illness and fatality, in the Asia-Pacific Region since 1997. Severe illness is most often associated with enterovirus A71 (EVA71). Vietnam experienced a large sustained outbreak of almost 200.000 hospitalised cases and over 200 deaths in 2011-12, the large majority occurring in southern Vietnam.
Retrospective data from 2009 to 2015 showed that Vietnam experienced 3 phases of HFMD outbreak time: before the outbreak (2009-2010), during the outbreak (2011-2012) and after the outbreak (2013-2015). Clinical features were slightly different between phases, partly reflected the effect of national intervention strategies.
Between July 2013 and July 2015 1547 children were enrolled. Four serotypes of EV-A71, Coxsackie virus (CV) A6, A10 and A16 were responsible for 1005/1327 (75.7%) of diagnosed cases. We found an unexpected dominance of EV-A71 among both in- and outpatients, and a strong association with severe illness. CV-A6 and A10 emerged in Vietnam during our study period and replaced CV-A16. CV-A10 was associated with different clinical and laboratory characteristics.
The relative genetic diversity of subgenogroup B5 increased sharply in 2012, leading to subgenogroup switch from C4 to B5 and then remained stable from 2013 to 2015. Subgenogroup B5 dominantly circulated in Vietnam during the study time and might import to Vietnam from two independent sources.
Three potential single nucleotide polymorphism (SNPs), rs10235866 represents gene NPTX2/TMEM130, rs817854 and rs10739235 represent gene RAD23B (RAD23 homolog B), were detected to be associated with severe disease. Results from my study, covering demography, clinical features, virology and host genetic factors, provides a full view of HFMD in Southern Vietnam and therefore may have great impact for disease management and control.

Advisor: van Doorn, H
Thwaites, L
Le Van Tan
Robust portfolio optimisation with filtering uncertainty

This thesis focuses on how portfolio optimisation can be carried out under different types of uncertainty, which we often measure through the use of filters. Chapter 1 motivates the problem, gives an overview of the thesis and covers some necessary background material. Chapter 2 deals with uncertainty in the covariance matrix and how by identifying different regimes we can solve optimisation problems of interest to practitioners.
Chapter 3 focuses on the uncertainty over tail events and how we can not only extract relevant information by filtering the data but also how we can use that information to construct a portfolio optimisation problem that acts on it. In Chapter 4 we address the lack of tractability for general relative robust portfolio optimisation problems and how one can overcome this so as to make it a viable tool.
Chapter 5 considers the problem of uncertainty in the filter itself and how this uncertainty can be fully incorporated in the portfolio optimisation problem. Finally in Chapter 6 we conclude and propose topics for future research.

Advisor: Hauser, R
Targeting interstrand crosslink repair to improve chemotherapeutic strategies

Many clinically effective chemotherapeutic agents exert their toxicity through formation of DNA interstrand cross-links (ICL). These prevent the replication and transcription of DNA and if unrepaired lead to cell death.
Despite the success of ICL inducing agents in treating cancers, issues remain with a lack of sensitivity in some tumours, therapy-limiting toxicity and acquired resistance and tumour recurrence. The modulation of ICL repair could improve chemotherapy outcomes and the specific targeting of ICL repair in quiescent cells may selectively target cancer stem cells and ameliorate issues with recurrence. This thesis seeks to identify potential druggable targets that may sensitise cancer cells to ICL inducing agents and elucidate the role of ICL repair in quiescence.
Previous studies have suggested ICL repair occurs chiefly in S-phase when DNA replication forks collide with ICLs. However, recent research in budding yeast, using Xenopus cell-free extracts and using genetic reporter assays in mammalian cells suggests that ICL repair can occur in the G1 phase of the cell cycle via the nucleotide excision repair (NER) and translesional synthesis (TLS) pathways. Previously this G1 pathway had not been well explored in mammalian cells.
In this thesis a model for G1 ICL repair was created using a mammalian cell line. Flow cytometry analysis (FACS) of cell cycle dynamics following ICL induction in phase G1 demonstrated that repair of ICL can occur in G1 and this appears time dependent. Further analysis by FACS, comet assays and cell survival analysis confirmed repair of ICL in G1 and suggested that the NER factors XPF, XPB and XPD and the exonucleases SNM1A and EXO1 may be required for ICL repair in G1 but not the XPG, FAN1 or CSB repair proteins.
Along-side these studies, a large-scale siRNA screen of 640 DNA damage repair genes was performed in a lung cancer cell line utilising colony formation assays to identify potential cisplatin and radiotherapy sensitising genes. Following a secondary screen, eight potential radiation and cisplatin sensitising genes were identified and examined in further detail, with the gene PIF1 finally confirmed in CRISPR/Cas9 knockout cells as a potential cisplatin sensitising gene.
The findings in this thesis confirm ICL repair can occur in G1 in mammalian cells and identify PIF1 as a potential cisplatin sensitiser in human cells. In the future, targeting of the specific endonucleases responsible for G1 ICL repair or PIF1 may offer a potential clinical cancer treatment option.

Advisor: McHugh, P
Higgins, G
Novel and selective small molecule inhibitors and activators for the prolyl hydroxylase domain enzyme

Hypoxia Inducible Factors (HIF) functions are master regulators of oxygen homeostasis and have a key role in the physiological responses to hypoxia including angiogenesis and erythropoiesis. Under hypoxia, levels of HIF-α subunits increase, they hetereodimerise with HIF-1β sub unit and promote the initiation of transcription of target genes. Under normoxia, oxygen dependent HIF-α degradation is promoted by hydroxylation of either of two proline residues (Pro402 and Pro564). The interaction of prolylhydroxylated HIF-α with the Von Hippel -Lindau protein (pVHL) promotes hydrolytic degradation of HIFα through an E3 ubiquitin ligase proteasomal pathway. HIF prolyl hydroxylation is catalysed by three 2-oxoglutarate (2OG)-dependent oxygenases known as prolyl hydroxylase domain (PHD 1-3) proteins, through an Fe(II) mediated catalytic process using 2OG, and oxygen. The PHDs are part of the family of Fe(II) bound 2OG dependent oxygenases. There are approximately 70 human 2OG oxygenases many of which have biologically important roles. Small-molecule inhibitors have reached advanced clinical trials; however, many clinical candidates inhibit other structurally similar 2OG oxygenases (OGFOD1 and vCPH) potentially altering the therapeutic effect. This thesis describes the design and synthesis of potent and 2OG oxygenase selective inhibitors for the PHDs.
The 1,3,8-triazaspiro[4.5]decane-2,4-dione and 4-hydroxy-2-(pyrazole)pyrimidine-5-amide series were chosen as initial 'hits' (reported in the patent literature). The main analogues of the series were characterised in vitro and in cells as potent and selective PHD inhibitors over structurally similar 2OG oxygenases (Chapter 2). Broad structure activity relationship (SAR) of both initial series demonstrated the sensitivity for PHD2 inhibition (Chapter 3). Combination of SAR work described in Chapters 2 and 3 lead to the development of the novel 4-hydroxy pyridine series. In-depth SAR resulted in optimised analogues including 1 (IC50 69 nM) and highly selective over structurally similar 2OG oxygenases including OGFOD1. The completed SAR work led to the development of two novel pharmacophores 2 and 3. Both pharmacophores displayed potent PHD inhibition and selectivity over OGFOD1. Analogues including 1 and 3 displayed on target cellular activity stabilising HIF-1α at 20 μM (Chapter 4).
The 4-dimethylamine pyridine analogue displayed an increase in substrate hydroxylation on PHD2 in contrast to the DMSO control (Chapter 3). SAR and cellular characterisation indicated that the effect observed was not an assay artifact (Chapter 5).
Fenofibrate was used as a starting point for the development of novel inhibitors of the oxygen consumption rate (OCR) via mitochondrial inhibition (Chapter 6). Analogues were synthesised in order to conduct broad SAR and on-target cellular activity was observed in a Seahorse XF assay (50% reduction in the OCR at 1 μM). A selection of amino and amide analogues warrant further investigation.

Keywords: HIF
PHD2
Advisor: Schofield, C