Vascular disrupting agent for neovascular age related macular degeneration: a pilot study of the safety and efficacy of intravenous combretastatin A-4 phosphate

BACKGROUND: This study was designed to assess the safety, tolerability, and efficacy of intravenous infusion of CA4P in patients with neovascular age-related macular degeneration (AMD). METHODS: Prospective, interventional, dose-escalation clinical trial. Eight patients with neovascular AMD refractory to at least 2 sessions of photodynamic therapy received CA4P at a dose of 27 or 36 mg/m2 as weekly intravenous infusion for 4 consecutive weeks. Safety was monitored by vital signs, ocular and physical examinations, electrocardiogram, routine laboratory tests, and collection of adverse events. Efficacy was assessed using retinal fluorescein angiography, optical coherence tomography, and best corrected visual acuity (BCVA). RESULTS: The most common adverse events were elevated blood pressure (46.7%), QTc prolongation (23.3%), elevated temperature (13.3%), and headache (10%), followed by nausea and eye injection (6.7%). There were no adverse events that were considered severe in intensity and none resulted in discontinuation of treatment. There was reduction of the excess foveal thickness by 24.15% at end of treatment period and by 43.75% at end of the two-month follow-up (p = 0.674 and 0.161, respectively). BCVA remained stable throughout the treatment and follow-up periods. CONCLUSIONS: The safety profile of intravenous CA4P was consistent with that reported in oncology trials of CA4P and with the class effects of vascular disruptive agents; however, the frequency of adverse events was different. There are evidences to suggest potential efficacy of CA4P in neovascular AMD. However, the level of systemic safety and efficacy indicates that systemic CA4P may not be suitable as an alternative monotherapy to current standard-of-care therapy

Midlands Cadences: Narrative Voices in the Work of Alan Sillitoe

This paper will examine excerpts from a range of Alan Sillitoe’s prose fiction, most notably Saturday Night and Sunday Morning (1958) and short stories from the collection The Loneliness of the Long-Distance Runner (1958), via a comparative exploration of the texts’ representations of Midlands English demotic. Both texts enact Bakhtin’s notion of novelistic dialogism and find much expressive capital in the tension between discourses: between the oral and the written. Indeed, it could be argued that much of Sillitoe’s work functions as a direct challenge to dominant notions of the literary. The narrative discourse attempts to trace a link between the quotidian experience of the Midlands English working classes represented and the demotic language which they speak. His technique also explores the link between language and sensibility; i.e. verbal articulacy need not be a limit to expression of a character’s distinctive identity. In contrast to the more radical techniques of novelists like James Kelman and Irvine Welsh, all instances of phonetically-rendered demotic remain imprisoned by what Joyce called ‘perverted commas’ – as direct speech. However, the diegetic narrative discourse itself is redolent of registers rooted in 1950s English working class life. The texts also contain different methods of representing their protagonists’ consciousness through their own idiolect. In Saturday Night and Sunday Morning, this is evidenced by the use of the second person ‘you’. It functions simultaneously as a representation of Seaton’s consciousness in the oral register which he might choose to articulate it, and as a dialogic ‘sideways glance’ at the reader and assumed shared experience. The second is more redolent of internal monologue, using the first-person form (as seen in the homodiegetic narration of the second novel); crucially, though, it remains in Standard English, if explicitly orientated towards oral register. Sillitoe’s is a novelistic discourse which refuses to normalise itself to accord with the conventions of classic realism, and as such prefigures the ambitions of many contemporary writers who incline their narrative voices towards the oral – asserting the right of a character’s dialect/idiolect to be the principal register of the narrative. The paper will demonstrate this thesis through the ideas of Bakhtin, and through an analytical taxonomy derived from literary stylistics. It aims to propose a model which can be used to analyse and explore any fiction which has been labelled as ‘working class’, and asserts that such an approach leads to a more principled characterisation of working class fiction (based on its use of language) than current literary-critical discussions based simply on cultural/social context and biography

Detection of chromosome aberrations in the human interphase nucleus by visualization of specific target DNAs with radioactive and non-radioactive in situ hybridization techniques: diagnosis of trisomy 18 with probe L1.84

The localization of chromosome 18 in human interphase nuclei is demonstrated by use of radioactive and nonradioactive in situ hybridization techniques with a DNA clone designated L1.84. This clone represents a distinct subpopulation of the repetitive human alphoid DNA family, located in the centric region of chromosome 18. Under stringent hybridization conditions hybridization of L1.84 is restricted to chromosome 18 and reflects the number of these chromosomes present in the nuclei, namely, two in normal diploid human cells and three in nuclei from cells with trisomy 18. Under conditions of low stringency, cross-hybridization with other subpopulations of the alphoid DNA family occurs in the centromeric regions of the whole chromosome complement, and numerous hybridization sites are detected over interphase nuclei. Detection of chromosome-specific target DNAs by non-radioactive in situ hybridization with appropriate DNA probes cloned from individual chromosomal subregions presents a rapid means of identifying directly numerical or even structural chromosome aberrations in the interphase nucleus. Present limitations and future applications of interphase cytogenetics are discussed

Membrane fission by dynamin: what we know and what we need to know

The large GTPase dynamin is the first protein shown to catalyze membrane fission. Dynamin and its related proteins are essential to many cell functions, from endocytosis to organelle division and fusion, and it plays a critical role in many physiological functions such as synaptic transmission and muscle contraction. Research of the past three decades has focused on understanding how dynamin works. In this review, we present the basis for an emerging consensus on how dynamin functions. Three properties of dynamin are strongly supported by experimental data: first, dynamin oligomerizes into a helical polymer; second, dynamin oligomer constricts in the presence of GTP; and third, dynamin catalyzes membrane fission upon GTP hydrolysis. We present the two current models for fission, essentially diverging in how GTP energy is spent. We further discuss how future research might solve the remaining open questions presently under discussion

Discrete Ladders for Parallel Transport in Transformation Groups with an Affine Connection Structure

International audienceModeling the temporal evolution of the tissues of the body is an important goal of medical image analysis, for instance for understanding the structural changes of organs affected by a pathology, or for studying the physiological growth during the life span. For such purposes we need to analyze and compare the observed anatomical differences between follow-up sequences of anatomical images of different subjects. Non-rigid registration is one of the main instruments for modeling anatomical differences from images. The aim of non-rigid registration is to encode the observed structural changes as deformation fields of the image space, which represent the warping required to match observed differences. This way, anatomical changes can be modeled and quantified by analyzing the associated deformations. The comparison of temporal evolutions thus requires the transport (or "normalization") of longitudinal deformations in a common reference frame. Normalization of longitudinal deformations can be done in different ways, depending on the feature of interest. For instance, local volume changes encoded by the scalar Jacobian determinant of longitudinal deformations can be compared by scalar resampling in a common reference frame via inter-subject registration. However, if we consider vector-valued deformation trajectories instead of scalar quantities, the transport is not uniquely defined anymore. Among the different normalization methods for deformation trajectories, the parallel transport is a powerful and promising tool which can be used within the ''diffeomorphic registration'' setting. Mathematically, parallel transporting a vector along a curve consists in translating it across the tangent spaces to the curve by preserving its parallelism according to a given derivative operation called (affine) connection. This chapter focuses on explicitly discrete algorithms for parallel transporting diffeomorphic deformations. Schild's ladder is an efficient and simple method proposed in theoretical Physics for the parallel transport of vectors along geodesics paths by iterative construction of infinitesimal geodesics parallelograms on the manifold. The base vertices of the parallelogram are given by the initial tangent vector to be transported. By iteratively building geodesic diagonals along the path, Schild's Ladder computes the missing vertex which corresponds to the transported vector. In this chapter we first show that the Schild ladder can lead to an effective computational scheme for the parallel transport of diffeomorphic deformations parameterized by tangent velocity fields. Schild's ladder may be however inefficient for transporting longitudinal deformations from image time series of multiple time points, in which the computation of the geodesic diagonals is required several times. We propose therefore a new parallel transport method based on the Schild's ladder, the "pole ladder", in which the computation of geodesics diagonals is minimized. Differently from the Schild's ladder, the pole ladder is symmetric with respect to the baseline-to-reference frame geodesic. From the theoretical point of view, we show that the pole ladder is rigorously equivalent to the Schild's ladder when transporting along geodesics. From the practical point of view, we establish the computational advantages and demonstrate the effectiveness of this very simple method by comparing with standard methods of transport on simulated images with progressing brain atrophy. Finally, we illustrate its application to a clinical problem: the measurement of the longitudinal progression in Alzheimer's disease. Results suggest that an important gain in sensitivity could be expected in group-wise comparisons

PI3Kγ is a molecular switch that controls immune suppression

Macrophages play critical, but opposite, roles in acute and chronic inflammation and cancer1,2,3,4,5. In response to pathogens or injury, inflammatory macrophages express cytokines that stimulate cytotoxic T cells, whereas macrophages in neoplastic and parasitic diseases express anti-inflammatory cytokines that induce immune suppression and may promote resistance to T cell checkpoint inhibitors1,2,3,4,5,6,7. Here we show that macrophage PI 3-kinase γ controls a critical switch between immune stimulation and suppression during inflammation and cancer. PI3Kγ signalling through Akt and mTor inhibits NFκB activation while stimulating C/EBPβ activation, thereby inducing a transcriptional program that promotes immune suppression during inflammation and tumour growth. By contrast, selective inactivation of macrophage PI3Kγ stimulates and prolongs NFκB activation and inhibits C/EBPβ activation, thus promoting an immunostimulatory transcriptional program that restores CD8+ T cell activation and cytotoxicity. PI3Kγ synergizes with checkpoint inhibitor therapy to promote tumour regression and increased survival in mouse models of cancer. In addition, PI3Kγ-directed, anti-inflammatory gene expression can predict survival probability in cancer patients. Our work thus demonstrates that therapeutic targeting of intracellular signalling pathways that regulate the switch between macrophage polarization states can control immune suppression in cancer and other disorders

Influence of the Stability of a Fused Protein and Its Distance to the Amyloidogenic Segment on Fibril Formation

Conversion of native proteins into amyloid fibrils is irreversible and therefore it is difficult to study the interdependence of conformational stability and fibrillation by thermodynamic analyses. Here we approached this problem by fusing amyloidogenic poly-alanine segments derived from the N-terminal domain of the nuclear poly (A) binding protein PABPN1 with a well studied, reversibly unfolding protein, CspB from Bacillus subtilis. Earlier studies had indicated that CspB could maintain its folded structure in fibrils, when it was separated from the amyloidogenic segment by a long linker. When CspB is directly fused with the amyloidogenic segment, it unfolds because its N-terminal chain region becomes integrated into the fibrillar core, as shown by protease mapping experiments. Spacers of either 3 or 16 residues between CspB and the amyloidogenic segment were not sufficient to prevent this loss of CspB structure. Since the low thermodynamic stability of CspB (ΔGD = 12.4 kJ/mol) might be responsible for unfolding and integration of CspB into fibrils, fusions with a CspB mutant with enhanced thermodynamic stability (ΔGD = 26.9 kJ/mol) were studied. This strongly stabilized CspB remained folded and prevented fibril formation in all fusions. Our data show that the conformational stability of a linked, independently structured protein domain can control fibril formation

From DNA sequence to application: possibilities and complications

The development of sophisticated genetic tools during the past 15 years have facilitated a tremendous increase of fundamental and application-oriented knowledge of lactic acid bacteria (LAB) and their bacteriophages. This knowledge relates both to the assignments of open reading frames (ORF’s) and the function of non-coding DNA sequences. Comparison of the complete nucleotide sequences of several LAB bacteriophages has revealed that their chromosomes have a fixed, modular structure, each module having a set of genes involved in a specific phase of the bacteriophage life cycle. LAB bacteriophage genes and DNA sequences have been used for the construction of temperature-inducible gene expression systems, gene-integration systems, and bacteriophage defence systems. The function of several LAB open reading frames and transcriptional units have been identified and characterized in detail. Many of these could find practical applications, such as induced lysis of LAB to enhance cheese ripening and re-routing of carbon fluxes for the production of a specific amino acid enantiomer. More knowledge has also become available concerning the function and structure of non-coding DNA positioned at or in the vicinity of promoters. In several cases the mRNA produced from this DNA contains a transcriptional terminator-antiterminator pair, in which the antiterminator can be stabilized either by uncharged tRNA or by interaction with a regulatory protein, thus preventing formation of the terminator so that mRNA elongation can proceed. Evidence has accumulated showing that also in LAB carbon catabolite repression in LAB is mediated by specific DNA elements in the vicinity of promoters governing the transcription of catabolic operons. Although some biological barriers have yet to be solved, the vast body of scientific information presently available allows the construction of tailor-made genetically modified LAB. Today, it appears that societal constraints rather than biological hurdles impede the use of genetically modified LAB.

Expression of the RNA helicase DDX3 and the hypoxia response in breast cancer

<p>Aims: DDX3 is an RNA helicase that has antiapoptotic properties, and promotes proliferation and transformation. In addition, DDX3 was shown to be a direct downstream target of HIF-1α (the master regulatory of the hypoxia response) in breast cancer cell lines. However, the relation between DDX3 and hypoxia has not been addressed in human tumors. In this paper, we studied the relation between DDX3 and the hypoxic responsive proteins in human breast cancer.</p> <p>Methods and Results: DDX3 expression was investigated by immunohistochemistry in breast cancer in comparison with hypoxia related proteins HIF-1α, GLUT1, CAIX, EGFR, HER2, Akt1, FOXO4, p53, ERα, COMMD1, FER kinase, PIN1, E-cadherin, p21, p27, Transferrin receptor, FOXO3A, c-Met and Notch1. DDX3 was overexpressed in 127 of 366 breast cancer patients, and was correlated with overexpression of HIF-1α and its downstream genes CAIX and GLUT1. Moreover, DDX3 expression correlated with hypoxia-related proteins EGFR, HER2, FOXO4, ERα and c-Met in a HIF-1α dependent fashion, and with COMMD1, FER kinase, Akt1, E-cadherin, TfR and FOXO3A independent of HIF-1α.</p> <p>Conclusions: In invasive breast cancer, expression of DDX3 was correlated with overexpression of HIF-1α and many other hypoxia related proteins, pointing to a distinct role for DDX3 under hypoxic conditions and supporting the oncogenic role of DDX3 which could have clinical implication for current development of DDX3 inhibitors.</p&gt