Novel Allosteric Mechanism of Dual p53/MDM2 and p53/MDM4 Inhibition by a Small Molecule

Restoration of the p53 tumor suppressor for personalised cancer therapy is a promising treatment strategy. However, several high-affinity MDM2 inhibitors have shown substantial side effects in clinical trials. Thus, elucidation of the molecular mechanisms of action of p53 reactivating molecules with alternative functional principle is of the utmost importance. Here, we report a discovery of a novel allosteric mechanism of p53 reactivation through targeting the p53 N-terminus which promotes inhibition of both p53/MDM2 (murine double minute 2) and p53/MDM4 interactions. Using biochemical assays and molecular docking, we identified the binding site of two p53 reactivating molecules, RITA (reactivation of p53 and induction of tumor cell apoptosis) and protoporphyrin IX (PpIX). Ion mobility-mass spectrometry revealed that the binding of RITA to serine 33 and serine 37 is responsible for inducing the allosteric shift in p53, which shields the MDM2 binding residues of p53 and prevents its interactions with MDM2 and MDM4. Our results point to an alternative mechanism of blocking p53 interaction with MDM2 and MDM4 and may pave the way for the development of novel allosteric inhibitors of p53/MDM2 and p53/MDM4 interactions

Molecular mechanisms of growth suppression by pharmacologically activated p53

The tumor suppressor p53 is a transcription factor that is crucial for protecting cells from cancer development. The importance of p53 tumor suppression function is highlighted by the fact that the p53 pathway is inactivated in most, if not all cancers. Mutation of the p53 gene occurs in about 50% of all tumors, whereas in the tumors which retain wild-type p53, the function of p53 is abolished due to deregulation of the p53 pathway. Due to the potency of p53 in suppressing tumors, p53 is considered to be an attractive therapeutic target. This is further strengthened by the in vivo mouse models which demonstrated regression of already established tumors upon reinstatement of p53. The aims of this thesis were to identify new p53 reactivating compounds and to investigate the mechanisms of action of the previously identified p53-reactivating molecule RITA. The NCI library of low molecular weight compounds was screened for molecules that suppressed cell growth in a p53-dependent manner. We identified the small molecule MITA, which induces p53-dependent cell death in a variety of human tumor cells. The p53/MDM2 interaction is blocked by MITA which results in accumulation of p53 in cells. The expression of p53 target genes MDM2, BAX, PUMA and GADD45 is induced upon activation of p53 by MITA. Importantly, MITA does not induce p53 or its target genes in normal human diploid fibroblasts (NHDF), which correlates with the absence of growth suppression. The low molecular weight compound RITA was previously identified in a cell-based screen. RITA has been shown to bind p53 and subsequently inhibit its interaction with MDM2. RITA induces p53-dependent apoptosis in cancer cells and in vivo in human tumor xenografts in mice. In the studies presented in this thesis we further elucidated the mechanisms of RITA action. Oligonucleotide microarray analysis of gene expression profiles revealed that RITA targets p53 with high specificity and that it mainly induces pro-apoptotic targets of p53. In line with these results, p53 wild-type expressing cell lines of different origin revealed a predominant apoptotic response. We demonstrate that MDM2 released from p53 by RITA promotes degradation of p21 and the p53 cofactor hnRNP K, required for p21 transcription. This leads to downregulation of p21 on both transcriptional and protein level. Consequently, MDM2 acts as a switch between growth arrest and apoptosis upon p53 activation. The preferential induction of apoptotic response by RITA is further facilitated by the targeting of the p53 regulator HDMX. RITA inhibits the p53/HDMX interaction and induces p53-dependent downregulation of HDMX on protein and on mRNA level. This ensures a robust activation of p53 leading to induction of apoptosis. Additionally, a new cellular target of RITA was identified, namely the redox protein TrxR1. We demonstrated that RITA binds purified TrxR1 protein directly and inhibits its activity in vitro and in cells. Notably, the inhibition of TrxR by RITA appeared to be p53-depedent and correlated with the induction of a covalently linked TrxR1 dimer and induction of ROS. We believe that inhibition of TrxR1 in a tumor-dependent and p53-dependent manner might contribute to the tumor-specific cell killing properties of RITA. In conclusion, we believe that compounds such as RITA and MITA may not only be used as lead compounds for anti-cancer therapy, but may also be useful tools to study of p53 functional activity

Cataloguing Church Music A study of The Düben Collection Dübensamlingen

rary of Sweden Statens musikbibliotek [51][pdf.gif] The aim of this study is to investigate how bibliographic description of music materials has improved through history. The historical aspect is given by a study of bibliographic data in different catalogues of a specific collection. The aim is also to understand what specific problems will arise in the work of cataloguing music materials and how the problems are discussed in the literature. The study is focusing of church music material and the history of churchmusic is therefore given an exposition. The origin of musical notation, which took place within the church, is also discussed. The empirical study is performed in two specific collections, mainly in The Düben Collection at the University of Uppsala but also at The Music Library in Stockholm. The catalogues in each collection are studied. In the discussion a comparison is given by the empirical material and the questions of this study. The empirical material should be seen as good examples rather than general conclusions. Churchmusic and musical notation has jointly improved over the years, which is seen in the improvement of music material. The art of cataloguing is influenced by the change of its material. By examining the two collections we can see how the bibliographic data has changed depending of the aim of the catalogues. Examples from the collections show how the problems with cataloguing music material are solved in each collection.Uppsatsnivå:

Tracing temporal conflicts in transitional Myanmar : life history diagrams as methodological tool

This article adds to the emerging ‘temporal turn’ in peace studies by addressing methodological questions about how temporality can be captured and explored in empirical studies. Developing our methodological tools for exploring time and temporality, we argue, is critical to move beyond the supposed linear temporality of peace processes, and make visible alternative temporal frameworks that shape everyday experiences and contestations around peace in conflict-affected contexts. Drawing on a study of two conflict-affected areas in Myanmar, we contribute towards this aim through a discussion of how life history diagrams helped us trace temporal conflicts between overarching narratives of political transition and everyday experiences of insecurity. This facilitated a deeper understanding of how relationships between war and peace, and between past, present and future, were manifested and made sense of in people’s everyday lives. Our use of life history diagrams revealed temporal conflicts between the dominant, linear temporality of the Myanmar transition, and more complex and cyclical temporal frameworks people used to describe their realities. Life history diagrams also facilitated narratives that troubled an events-based temporality focused on macro-political shifts such as ceasefire agreements and elections, and instead foregrounded everyday experiences of continuous insecurity and struggle

"They treat us like visitors in our own house" : relational peace and local experiences of the state in Myanmar

Between 2011 and 2021, political reforms and renewed peace efforts significantly reduced violence in many of Myanmar’s conflict-affected regions. Despite this, people living in these areas did not agree that they enjoyed peace; rather, this period is described as a continuation of the war’s many injustices, marked by discrimination, marginalization, and fear. This chapter argues that a relational analysis of peace can enable us to make sense of this gap between drastically different assessments of peace and conflict. The analysis draws on focus group discussions, interviews, and participant observation with local civilians, civil society activists, and members of non-state armed groups conducted in 2019 in two regions, Kayah and Mon States. A relational perspective uncovers the fact that the fundamental logics of key conflict relationships, between the Myanmar state and ethnic minority groups and communities, have not been transformed by the peace process but instead manifest themselves in new ways, whereby armed violence has been replaced by other forms of domination, underpinned by inequality, non-recognition, and distrust. Exploring these relational dynamics enables us to pinpoint areas and issues that prevent the emergence of a sustainable and legitimate peace, and demonstrate the importance of relational aspects for people’s experiences of everyday peace

'On the border, i learned how to advocate' : borderlands as political spaces for Burmese women’s activism

This article explores the political space of the border through the experiences of Myanmar women activists, for whom the borderlands in Thailand have provided refuge as well as a conducive environment for political mobilization. At the same time, the border renders refugee activists insecure and precarious. Drawing on life history interviews, our analysis expands conceptualizations of the border as a dynamic political space by illustrating its dual capacity to both facilitate and constrain the political agency of refugee women from Myanmar. In particular, the spatial and temporal fluidity and in-betweenness of the border is shown to foster both repression and resistance. Exploring the character and salience of the border as a space for activism over time, we demonstrate how the political space of the border is relational, constituted in interaction with other political spaces, such as politics and governance in Myanmar, transnational activist networks, and the politics of international aid. 

The politics of sexual violence in the Kachin conflict in Myanmar

Conflict-related sexual violence has been the focus of significant international activism and policy attention. International legal norms and frameworks have evolved to recognize it as a war crime, and a representation of sexual violence as a “weapon of war” is now widely endorsed. This article examines how international norms about conflict-related sexual violence are adopted and utilized in multiple ways in the armed conflict in Kachin state in northern Myanmar. Throughout decades of civil war, international norms on sexual violence have constituted key resources for international advocacy and awareness raising by local women’s rights activists. Further, activists have drawn on international norms to effect changes in gendered relations of power within their own communities. However, international norms on sexual violence in conflict have also been effectively used as tools for ethno-nationalist identity politics, rallying support behind the armed insurgency and mobilizing women’s unpaid labor in the service of war. Thus, international norms on conflict-related sexual violence have simultaneously opened up space for women’s empowerment and political agency and reproduced gendered forms of insecurity and marginalization. Exploring these contradictions and complexities, this analysis generates novel insights into the politics of international norms in contexts of armed conflict