Type-alpha DPLs

This paper introduces Denotational Proof Languages (DPLs). DPLs are languages for presenting, discovering, and checking formal proofs. In particular, in this paper we discus type-alpha DPLs---a simple class of DPLs for which termination is guaranteed and proof checking can be performed in time linear in the size of the proof. Type-alpha DPLs allow for lucid proof presentation and for efficient proof checking, but not for proof search. Type-omega DPLs allow for search as well as simple presentation and checking, but termination is no longer guaranteed and proof checking may diverge. We do not study type-omega DPLs here. We start by listing some common characteristics of DPLs. We then illustrate with a particularly simple example: a toy type-alpha DPL called PAR, for deducing parities. We present the abstract syntax of PAR, followed by two different kinds of formal semantics: evaluation and denotational. We then relate the two semantics and show how proof checking becomes tantamount to evaluation. We proceed to develop the proof theory of PAR, formulating and studying certain key notions such as observational equivalence that pervade all DPLs. We then present NDL, a type-alpha DPL for classical zero-order natural deduction. Our presentation of NDL mirrors that of PAR, showing how every basic concept that was introduced in PAR resurfaces in NDL. We present sample proofs of several well-known tautologies of propositional logic that demonstrate our thesis that DPL proofs are readable, writable, and concise. Next we contrast DPLs to typed logics based on the Curry-Howard isomorphism, and discuss the distinction between pure and augmented DPLs. Finally we consider the issue of implementing DPLs, presenting an implementation of PAR in SML and one in Athena, and end with some concluding remarks.

Simplifying transformations for type-alpha certificates

This paper presents an algorithm for simplifying NDL deductions. An array of simplifying transformations are rigorously defined. They are shown to be terminating, and to respect the formal semantis of the language. We also show that the transformations never increase the size or complexity of a deduction---in the worst case, they produce deductions of the same size and complexity as the original. We present several examples of proofs containing various types of "detours", and explain how our procedure eliminates them, resulting in smaller and cleaner deductions. All of the given transformations are fully implemented in SML-NJ. The complete code listing is presented, along with explanatory comments. Finally, although the transformations given here are defined for NDL, we point out that they can be applied to any type-alpha DPL that satisfies a few simple conditions.

Cálculo analítico de parâmetros estruturais de proteínas usando a teoria Alpha Shapes.

2007

Ethylene rapidly up-regulates the activities of both monomeric GTP-binding proteins and protein kinase(s) in epicotyls of pea

Igor E. Moshkov, Galina V. Novikova, Luis A.J. Mur, Aileen R. Smith, and Michael A. Hall. (2003). Ethylene rapidly up-regulates the activities of both monomeric GTP-binding proteins and protein kinase(s) in epicotyls of pea. Plant Physiology, 131(4), 1718-1726 RAE2008

Protein kinase C regulation of cell spreading in the molluscan Biomphalaria glabrata embryonic (Bge) cell line

Judith E. Humphries, Leah Elizondo and Timothy P. Yoshino (2001). Protein kinase C regulation of cell spreading in the molluscan Biomphalaria glabrata embryonic (Bge) cell line. Biochimica et Biophysica Acta - Molecular Cell Research, 1540(3), 243-252. Sponsorship: National Institutes of Health AI 15503 RAE2008

A Catalogue of the Alpha Chapter Boston University Graduates of the School of Theology 1850-1905

A listing of graduate of Boston University School of Theology and predecessor school. Arranged by class year, alphabetical by last name and geographically by region.

Targeting aberrant kinase activity in myeloid leukaemias

Acute myeloid leukaemia (AML) is the most common form of acute leukaemia in adults. Its treatment has remained largely unchanged for the past 30 years. Chronic myeloid leukaemia (CML) represents a tremendous success story in the era of targeted therapy but significant challenges remain including the development of drug resistance and disease persistence due to presence of CML stem cells. The Aurora family of kinases is essential for cell cycle regulation and their aberrant expression in cancer prompted the development of small molecules that selectively inhibit their activity. Chapter 2 of this thesis outlines the efficacy and mechanism of action of alisertib, a novel inhibitor of Aurora A kinase, in preclinical models of CML. Alisertib possessed equipotent activity against CML cells expressing unmutated and mutated forms of BCR-ABL. Notably, this agent retained high activity against the T315I and E255K BCR-ABL mutations, which confer the greatest degree of resistance to standard CML therapy. Chapter 3 explores the activity of alisertib in preclinical models of AML. Alisertib disrupted cell viability, diminished clonogenic survival, induced expression of the forkhead box O3 (FOXO3a) targets p27 and BCL-2 interacting mediator (BIM), and triggered apoptosis. A link between Aurora A expression and sensitivity to ara-C was established. Chapter 4 outlines the role of the proto-oncogene serine/threonine-protein (PIM) kinases in resistance to ara-C in AML. We report that the novel small molecule PIM kinase inhibitor SGI-1776 disrupted cell viability and induced apoptosis in AML. We establish a link between ara-C resistance and PIM over-expression. Finally, chapter 5 explores how the preclinical work outlined in this thesis may be translated into clinical studies that may lead to novel therapeutic approaches for patients with refractory myeloid leukaemia.

Synthesis and reactivity of alpha-thio-beta-chloroacrylamides and alpha-thio-beta-chloroenones

Development of novel synthetic methodology for selective transformation of organic compounds is a central element underpinning organic synthesis with control of chemo-, regio- and stereoselectivity a very high priority. Reactions which can be conducted under mild reaction conditions and, ideally in an environmentally attractive manner, are particularly advantageous. The principal objective of this thesis was to explore the synthesis, reactivity and synthetic utility of a series of α,β-thio-β-chloroenones. The stereochemical features of these transformations and the potential of this novel series of compounds in the synthesis of bioactive compounds were of particular interest. In exploring the reactivity of these compounds, the key transformations included nucleophilic additions and Stille cross-coupling at the β-carbon. Chapter 1 reviews the literature relevant to the research conducted, and focuses in particular on the synthesis of β-chloroenones and related unsaturated carbonyl compounds. The synthesis of chalcone compounds from various precursors is also discussed, with particular emphasis on the use of palladium cross-coupling reactions in the preparation of these compounds. The biological activity of chalcones is also summarised in this chapter. The second chapter delineates the stereoselective synthesis of the novel α-thio-β-chloroenones from the corresponding α-thioketones in a multistep reaction cascade initiated by a NCS-mediated chlorination. A range of both alkyl and aryl β-chloroenones were prepared in this work and the oxidation of these compounds to the corresponding sulfoxides and sulfones is also outlined. The electrophilicity of the β-carbon of the enones was examined in nucleophilic addition/substitution reactions with successful access to a variety of synthetically useful novel adducts including acetals and enaminoketones. Investigation of the synthetic potential of the Stille cross-coupling reaction with the novel α-thio-β-chloroenones was explored and provided an efficient route for the synthesis of a novel series of chalcones. Most importantly this new methodology provided a new and synthetically powerful approach for carbon-carbon bond formation at the β-carbon under mild neutral conditions. A preliminary investigation into the use of these β-chloroenones as dienophiles in Diels-Alder cycloaddition reactions is also discussed in this chapter. Chapter 2 also reports the nucleophilic addition of N, O, S and C nucleophiles to previously described β-chloroacrylamides and their corresponding sulfoxide derivatives. This work builds on previous research carried out in this programme and the reactivity of these β-chloroacrylamides at the sulfide and sulfoxide level is compared. Comparison of the reactivity of the β-chloroacrylamides, in nucleophilic substitution and Stille-coupling, with that of the novel β-chloroenones is of interest. Finally, the biological activity of both the β-chloroenones and the β-chloroacrylamides in terms of cytotoxicity is summarised in Chapter 2. The final chapter, Chapter 3, details the full experimental procedures, including spectroscopic and analytical data for the compounds prepared during this research.

Catalyst, additive and substrate effects in intramolecular aromatic additions of alpha-diazoketones

This thesis is focused on transition metal catalysed reaction of α-diazoketones leading to aromatic addition to form azulenones, with particular emphasis on enantiocontrol through use of chiral copper catalysts. The first chapter provides an overview of the influence of variation of the substituent at the diazo carbon on the outcome of subsequent reaction pathways, focusing in particular on C-H insertion, cyclopropanation, aromatic addition and ylide formation drawing together for the first time input from a range of primary reports. Chapter two describes the synthesis of a range of novel α-diazoketones. Rhodium and copper catalysed cyclisation of these to form a range of azulenones is described. Variation of the transition metal catalyst was undertaken using both copper and rhodium based systems and ligand variation, including the design and synthesis of a novel bisoxazoline ligand. The influence of additives, especially NaBARF, on the enantiocontrol was explored in detail and displayed an interesting impact which was sensitive to substituent effects. Further exploration demonstrated that it is the sodium cation which is critical in the additive effects. For the first time, enantiocontrol in the aromatic addition of terminal diazoketones was demonstrated indicating enantiofacial control in the aromatic addition is feasible in the absence of a bridgehead substituent. Determination of the enantiopurity in these compounds was particularly challenging due to the lability of the products. A substantial portion of the work was focused on determining the stereochemical outcome of the aromatic addition processes, both the absolute stereochemistry and extent of enantiopurity. Formation of PTAD adducts was beneficial in this regard. The third chapter contains the full experimental details and spectral characterisation of all novel compounds synthesised in this project, while details of chiral stationary phase HPLC and 1H NMR analysis are included in the appendix.

The role of mitogen activated protein kinase phosphatase-1 in neurotoxic and inflammatory-induced changes in the development of midbrain dopaminergic neurons: relevance to Parkinson’s disease

Parkinson’s disease (PD) is a progressive neurodegenerative disorder characterised by the loss of midbrain dopaminergic neurons from the substantia nigra pars compacta(SNpc), which results in motor, cognitive and psychiatric symptoms. Evidence supports a role for the mitogen-activated protein kinase p38 in the demise of dopaminergic neurons, while mitogen-activated protein kinase phosphatase-1 (MKP-1), which negatively regulates p38 activity, has not yet been investigated in this context. Inflammation may also be associated with the neuropathology of PD due to evidence of increased levels of proinflammatory cytokines such as interleukin-1β (IL-1β) within the SNpc. Because of the specific loss of dopaminergic neurons in a discreet region of the brain, PD is considered a suitable candidate for cell replacement therapy but challenges remain to optimise dopaminergic cell survival and morphological development. The present thesis examined the role of MKP-1 in neurotoxic and inflammatory-induced changes in the development of midbrain dopaminergic neurons. We show that MKP-1 is expressed in dopaminergic neurons cultured from embryonic day (E) 14 rat ventral mesencephalon (VM). Inhibition of dopaminergic neurite growth induced by treatment of rat VM neurons with the dopaminergic neurotoxin 6- hydroxydopamine (6-OHDA) is mediated by p38, and is concomitant with a significant and selective decrease in MKP-1 expression in these neurons. Dopaminergic neurons transfected to overexpress MKP-1 displayed a more complex morphology and contributed to neuroprotection against the effects of 6-OHDA. Therefore, MKP-1 expression can promote the growth and elaboration of dopaminergic neuronal processes and can help protect them from the neurotoxic effects of 6-OHDA. Neural precursor cells (NPCs) have emerged as promising alternative candidates to fetal VM for cell replacement strategies in PD. Here we show that phosphorylated (and thus activated) p38 and MKP-1 are expressed at basal levels in untreated E14 rat VM NPCs (nestin, DCX, GFAP and DAT-positive cells) following proliferation as well as in their differentiated progeny (DCX, DAT, GFAP and βIII-tubulin) in vitro. Challenge with 6-OHDA or IL-1β changed the expression of endogenous phospho-p38 and MKP-1 in these cells in a time-dependent manner, and so the dynamic balance in expression may mediate the detrimental effects of neurotoxicity and inflammation in proliferating and differentiating NPCs. We demonstrate that there was an up-regulation in MKP-1 mRNA expression in adult rat midbrain tissue 4 days post lesion in two rat models of PD; the 6-OHDA medial forebrain bundle (MFB) model and the four-site 6-OHDA striatal lesion model. This was concomitant with a decrease in tyrosine hydroxylase (TH) mRNA expression at 4 and 10 days post-lesion in the MFB model and 10 and 28 days post-lesion in the striatal lesion model. There was no change in mRNA expression of the pro-apoptotic gene, bax and the anti-apoptotic gene, bcl-2 in the midbrain and striatum. These data suggest that the early and transient upregulation of MKP-1 mRNA in the midbrain at 4 days post-6-OHDA administration may be indicative of an attempt by dopaminergic neurons in the midbrain to protect against the neurotoxic effects of 6-OHDA at later time points. Collectively, these findings show that MKP-1 is expressed by developing and adult dopaminergic neurons in the midbrain, and can promote their morphological development. MKP-1 also exerts neuroprotective effects against dopaminergic neurotoxins in vitro, and its expression in dopaminergic neurons can be modulated by inflammatory and neurotoxic insults both in vitro and in vivo. Thus, these data contribute to the information needed to develop therapeutic strategies for protecting midbrain dopaminergic neurons in the context of PD.

The role of presenilin 1 and presenilin 2 in IL-1β-, LPS- and TNFα- mediated signalling events

The importance of γ-secretase protease activities in development, neurogenesis and the immune system are highlighted by the diversity of its substrates and phenotypic characterization of Presenilin (PS)-deficient transgenic animals. Since the discovery of Amyloid precursor protein (APP) and it’s cleavage by γ-secretase complexes, over 90 other type I membrane proteins have been identified as γ-secretase substrates. We have identified interleukin-1 (IL-1) receptor type I (IL-1R1), toll-like receptor 4 (TLR4) and tumour necrosis factor-α (TNFα) receptor-1 (TNFR1) as novel substrates for - secretase cleavage, which play an important role in innate immunity. In this study, using PS-deficient cells and PS-knockout animal models we examined the role of PS proteins, PS1 and PS2, in IL-1R1-, TLR4- and TNFR1- mediated inflammatory responses. Data presented show that in response to IL- 1β, lipopolysaccharide (LPS) or TNFα, immortalised fibroblasts from PS2- deficient animals have diminished production of specific cytokines and chemokine, with differential reduction in nuclear factor-κB (NF-κB) and (mitogen activated protein kinase) MAPK activities. In contrast, no defect in the response to IL-1β, LPS or TNFα was observed in PS1-deficient immortalised fibroblasts. These observations were confirmed using bone marrow-derived macrophages from PS2-null mice, which also display impaired responsiveness to IL-1β- and LPS, with decreased production of inflammatory cytokines. Furthermore, in whole animal in vivo responses, we show that PS2-deficient animals display ligand (IL-1β, LPS and TNFα)-dependent alterations in the production of specific pro-inflammatory cytokines or chemokines. Importantly, this reduced responsiveness to IL-1β, LPS or TNFα is independent of γ- secretase protease activity and γ-secretase cleavage of TNFR1, IL-1R1 or TLR4. These observations suggest a novel γ-secretase-independent role of PS2 in the regulation of innate immune responsiveness and challenge current concepts regarding the regulation of IL-1β-, LPS- and TNFα-mediated immune signalling.

Molecular mechanisms underpinning IFN-gamma and TNF-alpha synergy in intestinal epithelial cells

Cytokine-driven signalling shapes immune homeostasis and guides inflammatory responses mainly through induction of specific gene expression programmes both within and outside the immune cell compartment. These transcriptional outputs are often amplified via cytokine synergy, which sets a stimulatory threshold that safeguards from exacerbated inflammation and immunopathology. In this study, we investigated the molecular mechanisms underpinning synergy between two pivotal Th1 cytokines, IFN-γ and TNF-α, in human intestinal epithelial cells. These two proinflammatory mediators induce a unique state of signalling and transcriptional synergy implicated in processes such as antiviral and antitumour immunity, intestinal barrier and pancreatic β-cell dysfunction. Since its discovery more than 30 years ago, this biological phenomenon remains, however, only partially defined. Here, using a functional genomics approach including RNAi perturbation screens and small-molecule inhibitors, we identified two new regulators of IFN-γ/TNF-α-induced chemokine and antiviral gene and protein expression, a Bcl-2 protein BCL-G and a histone demethylase UTX. We also discovered that IFN-γ/TNF-α synergise to trigger a coordinated shutdown of major receptor tyrosine kinases expression in colon cancer cells. Together, these findings extend our current understanding of how IFN-γ/TNF-α synergy elicits qualitatively and quantitatively distinct outputs in the intestinal epithelium. Given the well-documented role of this synergistic state in immunopathology of various disorders, our results may help to inform the identification of high quality and biologically relevant druggable targets for diseases characterised by an IFN-γ/TNF-α high immune signature

Evaluation of HER2, p53, bcl-2, topoisomerase II-alpha, heat shock proteins 27 and 70 in primary breast cancer and metastatic ipsilateral axillary lymph nodes.

BACKGROUND: Breast cancer is a heterogeneous disease. Predictive biological markers (BM) of responsiveness to therapy need to be identified. Evaluation of BM is mainly done at the primary site. However, in the adjuvant therapy of breast cancer, the main goal is control of micrometastases. It is still unknown whether heterogeneity in the expression of BM between the primary site and its micrometastases exists. OBJECTIVE: To evaluate the expression of some BM with potential predictive value from the primary breast cancer site and metastatic ipsilateral axillary lymph nodes. PATIENTS AND METHODS: Focality (percentage of positive cells) and intensity staining scores were evaluated for each marker. Freshly cut sections (4 microm) from embedded blocks of breast cancer fixed in formalin or bouin were put onto superfrost slides (Menzel-Gläser). Protein expression was evaluated immunohistochemically (IHC) using monoclonal antibodies against: topo II-alpha (clone KiS1, 1 microg/ml, Roche) with a trypsine pre-treatment (P); HSP27 (clone G3.1, 1/60, Biogenex), HSP70 (clone BRM.22, 1/80, Biogenex) and HER2 (clone CB11, 1/40, Novocastra; without P); p53 (clone D07, 1/750, Dako) and bcl-2 (clone 124, 1/60, Dako) with citrate buffer as P. RESULTS: Overall, the percentage of discordant marker status in the primary tumour and its metastatic lymph nodes was 2% for HER2, 6% for p53, 15% for bcl-2, 19% for topoisomerase II-alpha, 24% for HSP27 and 30% for HSP70. For the subgroup of patients with positive BM in the primary tumour, the percentage of discordance was 6% for HER2, 7% for p53, 14% for bcl-2, 19% for HSP70, 21% for topoisomerase II-alpha and 36% for HSP27. For the subgroup of patients with positive BM in the lymph nodes, the percentage of discordance was 9% for bcl-2, 15% for HER2 and p53, 21% for topoisomerase II-alpha, 22% for HSP27 and 25% for HSP70. CONCLUSIONS: 1) No biological marker had 100% concordant results. 2) Although some discordant cases might be explained by the limitations of the IHC technique, future studies aiming to evaluate the predictive value of BM in the adjuvant therapy of breast cancer should take into account a possible difference in BM expression between the primary and the metastatic sites.