Next-Generation Sequencing-Based Mutations Scanning Strategy of the BCR-ABL Kinase Domain in Patients with PhiladelPhia-Chromosome Positive Leukemias Treated with Tyrosine Kinase Inhibitors

In chronic myeloid leukemia and Philadelphia-positive acute lymphoblastic leukemia patients resistant to tyrosine kinase inhibitors (TKIs), BCR-ABL kinase domain mutation status is an essential component of the therapeutic decision algorithm. The recent development of Ultra-Deep Sequencing approach (UDS) has opened the way to a more accurate characterization of the mutant clones surviving TKIs conjugating assay sensitivity and throughput. We decided to set-up and validated an UDS-based for BCR-ABL KD mutation screening in order to i) resolve qualitatively and quantitatively the complexity and the clonal structure of mutated populations surviving TKIs, ii) study the dynamic of expansion of mutated clones in relation to TKIs therapy, iii) assess whether UDS may allow more sensitive detection of emerging clones, harboring critical 2GTKIs-resistant mutations predicting for an impending relapse, earlier than SS. UDS was performed on a Roche GS Junior instrument, according to an amplicon sequencing design and protocol set up and validated in the framework of the IRON-II (Interlaboratory Robustness of Next-Generation Sequencing) International consortium.Samples from CML and Ph+ ALL patients who had developed resistance to one or multiple TKIs and collected at regular time-points during treatment were selected for this study. Our results indicate the technical feasibility, accuracy and robustness of our UDS-based BCR-ABL KD mutation screening approach. UDS was found to provide a more accurate picture of BCR-ABL KD mutation status, both in terms of presence/absence of mutations and in terms of clonal complexity and showed that BCR-ABL KD mutations detected by SS are only the “tip of iceberg”. In addition UDS may reliably pick 2GTKIs-resistant mutations earlier than SS in a significantly greater proportion of patients.The enhanced sensitivity as well as the possibility to identify low level mutations point the UDS-based approach as an ideal alternative to conventional sequencing for BCR-ABL KD mutation screening in TKIs-resistant Ph+ leukemia patients

Optimization and development of low environmental impact propulsion systems

The aim of the Ph.D. research project was to explore Dual Fuel combustion and hybridization. Natural gas-diesel Dual Fuel combustion was experimentally investigated on a 4-Stroke, 2.8 L, turbocharged, light-duty Diesel engine, considering four operating points in the range between low to medium-high loads at 3000 rpm. Then, a numerical analysis was carried out using a customized version of the KIVA-3V code, in order to optimize the diesel injection strategy of the highest investigated load. A second KIVA-3V model was used to analyse the interchangeability between natural gas and biogas on an intermediate operating point. Since natural gas-diesel Dual Fuel combustion suffers from poor combustion efficiency at low loads, the effects of hydrogen enriched natural gas on Dual Fuel combustion were investigated using a validated Ansys Forte model, followed by an optimization of the diesel injection strategy and a sensitivity analysis to the swirl ratio, on the lowest investigated load. Since one of the main issues of Low Temperature Combustion engines is the low power density, 2-Stroke engines, thanks to the double frequency compared to 4-Stroke engines, may be more suitable to operate in Dual Fuel mode. Therefore, the application of gasoline-diesel Dual Fuel combustion to a modern 2-Stroke Diesel engine was analysed, starting from the investigation of gasoline injection and mixture formation. As far as hybridization is concerned, a MATLAB-Simulink model was built to compare a conventional (combustion) and a parallel-hybrid powertrain applied to a Formula SAE race car.

Development and food security in africa from the 1960s to 2010s: the case of Ethiopia

Persistent food insecurity and famines have continued to significantly shape the development policies of Ethiopia for decades. Over the decades, frequent famines caused not only the death of hundreds of thousands of victims but also significantly contributed to two revolutions that swept away the Haile Selassie and Derg regimes, as well as significantly taxing the legitimacy of the incumbent regime. As a result, agriculture and food security have become increasingly the top policy priorities for all political regimes in Ethiopia. However, the development policies of the ruling elites of Ethiopia have consistently failed to transform backward agriculture and ensure food security. The failures of the development policies of the Ethiopian governments over the years were attributed to several factors. Ethiopian authoritarian politics, centralized rule with a lack of transparency and accountability; the isolation of peasants from the development and governance process, and the lack of coherent agricultural development strategies that invest in peasant agriculture and create synergy among sectors are identified as key issues that have contributed to the persistence of food insecurity in the country. The literature on the failure of Ethiopia's political regimes to address food insecurity and famine has two major gaps that this study aims to fill. First, the cumulative and path-dependent food security and agricultural development policy environment were not adequately considered. Second, the strategy of extraversion by subsequent political regimes to use external support as a relief to prevent the famine-induced political crisis. This study used a mixed approach to collect data and present the evolution of the interplays of development policies and food security in three regimes within the context of international food security discourses. This study found out how the historical patterns of approaches of Ethiopia’s regimes to development and governance led to frequent famines and persistent food insecurity.

Gender equality and human development in Macedonia during transition (1991-2006)

The present thesis treats the issue of gender equality in Macedonia during the period of transition from the socialist system to the one of parliamentary democracy. The main aim is to mainstream the gender perspective in the analysis of the transitional policies through the examination of the basic citizenship rights to which citizens are entitled and by the means of the evaluation of their capabilities to exercise these rights. Gender equality, as one of the main strongholds of the concept of human development is measured through the application of nine gender relevant capabilities in a Case study conducted within selected municipalities in the country. Through the analysis of the Macedonian constitutional and legal framework and the assessment of gender based inequalities, the research questions the need for the enactment of a process of engendering of citizenship, which would integrate gender based differences, contemplate the private sphere of citizens lives and pledge participation in the political life of the country. The thesis, finally, analyses the gender equality strategy of the Macedonian government with the purpose to evaluate whether it is context based, i.e. it tackles the main fields where inequalities emerge and in this context whether it envisages a process of engendering of citizenship.

Technology acquisition through licensing. Implications for firm strategy

Nowadays licensing practices have increased in importance and relevance driving the widespread diffusion of markets for technologies. Firms are shifting from a tactical to a strategic attitude towards licensing, addressing both business and corporate level objectives. The Open Innovation Paradigm has been embraced. Firms rely more and more on collaboration and external sourcing of knowledge. This new model of innovation requires firms to leverage on external technologies to unlock the potential of firms’ internal innovative efforts. In this context, firms’ competitive advantage depends both on their ability to recognize available opportunities inside and outside their boundaries and on their readiness to exploit them in order to fuel their innovation process dynamically. Licensing is one of the ways available to firm to ripe the advantages associated to an open attitude in technology strategy. From the licensee’s point view this implies challenging the so-called not-invented-here syndrome, affecting the more traditional firms that emphasize the myth of internal research and development supremacy. This also entails understanding the so-called cognitive constraints affecting the perfect functioning of markets for technologies that are associated to the costs for the assimilation, integration and exploitation of external knowledge by recipient firms. My thesis aimed at shedding light on new interesting issues associated to in-licensing activities that have been neglected by the literature on licensing and markets for technologies. The reason for this gap is associated to the “perspective bias” affecting the works within this stream of research. With very few notable exceptions, they have been generally concerned with the investigation of the so-called licensing dilemma of the licensor – whether to license out or to internally exploit the in-house developed technologies, while neglecting the licensee’s perspective. In my opinion, this has left rooms for improving the understanding of the determinants and conditions affecting licensing-in practices. From the licensee’s viewpoint, the licensing strategy deals with the search, integration, assimilation, exploitation of external technologies. As such it lies at the very hearth of firm’s technology strategy. Improving our understanding of this strategy is thus required to assess the full implications of in-licensing decisions as they shape firms’ innovation patterns and technological capabilities evolution. It also allow for understanding the so-called cognitive constraints associated to the not-invented-here syndrome. In recognition of that, the aim of my work is to contribute to the theoretical and empirical literature explaining the determinants of the licensee’s behavior, by providing a comprehensive theoretical framework as well as ad-hoc conceptual tools to understand and overcome frictions and to ease the achievement of satisfactory technology transfer agreements in the marketplace. Aiming at this, I investigate licensing-in in three different fashions developed in three research papers. In the first work, I investigate the links between licensing and the patterns of firms’ technological search diversification according to the framework of references of the Search literature, Resource-based Theory and the theory of general purpose technologies. In the second paper - that continues where the first one left off – I analyze the new concept of learning-bylicensing, in terms of development of new knowledge inside the licensee firms (e.g. new patents) some years after the acquisition of the license, according to the Dynamic Capabilities perspective. Finally, in the third study, Ideal with the determinants of the remuneration structure of patent licenses (form and amount), and in particular on the role of the upfront fee from the licensee’s perspective. Aiming at this, I combine the insights of two theoretical approaches: agency and real options theory.

Development Aid to Water Management in Mali: The Actors, ‘Global’ Paradigms, and ‘Local’ Translations

Development aid involves a complex network of numerous and extremely heterogeneous actors. Nevertheless, all actors seem to speak the same ‘development jargon’ and to display a congruence that extends from the donor over the professional consultant to the village chief. And although the ideas about what counts as ‘good’ and ‘bad’ aid have constantly changed over time —with new paradigms and policies sprouting every few years— the apparent congruence between actors more or less remains unchanged. How can this be explained? Is it a strategy of all actors to get into the pocket of the donor, or are the social dynamics in development aid more complex? When a new development paradigm appears, where does it come from and how does it gain support? Is this support really homogeneous? To answer the questions, a multi-sited ethnography was conducted in the sector of water-related development aid, with a focus on 3 paradigms that are currently hegemonic in this sector: Integrated Water Resources Management, Capacity Building, and Adaptation to Climate Change. The sites of inquiry were: the headquarters of a multilateral organization, the headquarters of a development NGO, and the Inner Niger Delta in Mali. The research shows that paradigm shifts do not happen overnight but that new paradigms have long lines of descent. Moreover, they require a lot of work from actors in order to become hegemonic; the actors need to create a tight network of support. Each actor, however, interprets the paradigms in a slightly different way, depending on the position in the network. They implant their own interests in their interpretation of the paradigm (the actors ‘translate’ their interests), regardless of whether they constitute the donor, a mediator, or the aid recipient. These translations are necessary to cement and reproduce the network.

Development and characterization of micromachined devices for separation techniques

Nowadays microfluidic is becoming an important technology in many chemical and biological processes and analysis applications. The potential to replace large-scale conventional laboratory instrumentation with miniaturized and self-contained systems, (called lab-on-a-chip (LOC) or point-of-care-testing (POCT)), offers a variety of advantages such as low reagent consumption, faster analysis speeds, and the capability of operating in a massively parallel scale in order to achieve high-throughput. Micro-electro-mechanical-systems (MEMS) technologies enable both the fabrication of miniaturized system and the possibility of developing compact and portable systems. The work described in this dissertation is towards the development of micromachined separation devices for both high-speed gas chromatography (HSGC) and gravitational field-flow fractionation (GrFFF) using MEMS technologies. Concerning the HSGC, a complete platform of three MEMS-based GC core components (injector, separation column and detector) is designed, fabricated and characterized. The microinjector consists of a set of pneumatically driven microvalves, based on a polymeric actuating membrane. Experimental results demonstrate that the microinjector is able to guarantee low dead volumes, fast actuation time, a wide operating temperature range and high chemical inertness. The microcolumn consists of an all-silicon microcolumn having a nearly circular cross-section channel. The extensive characterization has produced separation performances very close to the theoretical ideal expectations. A thermal conductivity detector (TCD) is chosen as most proper detector to be miniaturized since the volume reduction of the detector chamber results in increased mass and reduced dead volumes. The microTDC shows a good sensitivity and a very wide dynamic range. Finally a feasibility study for miniaturizing a channel suited for GrFFF is performed. The proposed GrFFF microchannel is at early stage of development, but represents a first step for the realization of a highly portable and potentially low-cost POCT device for biomedical applications.

Genome-wide analysis of G-type lectin genes in Fragaria vesca and functional characterization of FaMBL1 gene in defense response of F. × ananassa to fungal pathogens

Strawberry (Fragaria × ananassa) is an important soft fruit but easily to be infected by pathogens. Anthracnose and gray mold are two of the most destructive diseases of strawberry which lead to serious fruit rot. The first chapter introduced strawberry anthracnose caused by Colletotrichum acutatum. The infection strategy, disease cycle and management of C. acutatum on strawberry were reported. Likewise, the second chapter summarized the infection strategy of Botrytis cinerea and the defense responses of strawberry. As we already know white unripe strawberry fruits are more resistant to C. acutatum than red ripe fruits. During the interaction between strawberry white/red fruit and C. acutaum, a mannose binding lectin gene, FaMBL1, was found to be the most up-regulated gene and induced exclusively in white fruit. FaMBL1 belongs to the G-type lectin family which has important roles in plant development and defense process. To get insight into the role of FaMBL1, genome-wide identification was carried out on G-type lectin gene family in Fragaria vesca and the results were showed in chapter 3. G-type lectin genes make up a large family in F. vesca. Active expression upon biotic/abiotic stresses suggested a potential role of G-lectin genes in strawberry defenses. Hence, stable transgenic strawberry plants with FaMBL1 gene overexpressed were generated. Transformed strawberry plants were screened and identified. The results were showed in chapter 4, content of disease-related phytohormone, jasmonic acid, was found decreased in overexpressing lines compared with wild type (WT). Petioles inoculated by C. fioriniae of overexpressing lines had lower disease incidence than WT. Leaves of overexpressing lines challenged by B. cinerea showed remarkably smaller lesion diameters compared with WT. The chitinase 2-1 (FaChi2-1) showed higher expression in overexpressing lines than in WT during the interaction with B. cinerea, which could be related with the lower susceptibility of overexpressing lines.

Orchestration of multiscale model for computational oncology

Cancer is a challenging disease that involves multiple types of biological interactions in different time and space scales. Often computational modelling has been facing problems that, in the current technology level, is impracticable to represent in a single space-time continuum. To handle this sort of problems, complex orchestrations of multiscale models is frequently done. PRIMAGE is a large EU project that aims to support personalized childhood cancer diagnosis and prognosis. The goal is to do so predicting the growth of the solid tumour using multiscale in-silico technologies. The project proposes an open cloud-based platform to support decision making in the clinical management of paediatric cancers. The orchestration of predictive models is in general complex and would require a software framework that support and facilitate such task. The present work, proposes the development of an updated framework, referred herein as the VPH-HFv3, as a part of the PRIMAGE project. This framework, a complete re-writing with respect to the previous versions, aims to orchestrate several models, which are in concurrent development, using an architecture as simple as possible, easy to maintain and with high reusability. This sort of problem generally requires unfeasible execution times. To overcome this problem was developed a strategy of particularisation, which maps the upper-scale model results into a smaller number and homogenisation which does the inverse way and analysed the accuracy of this approach.