The polyamines spermidine and spermine and their diamine precursor putrescine are naturally occurring, polycationic alkylamines that are essential for eukaryotic cell growth. The requirement for and ...the metabolism of polyamines are frequently dysregulated in cancer and other hyperproliferative diseases, thus making polyamine function and metabolism attractive targets for therapeutic intervention. Recent advances in our understanding of polyamine function, metabolic regulation, and differences between normal cells and tumour cells with respect to polyamine biology, have reinforced the interest in this target-rich pathway for drug development.
In cancer drug discovery, it is important to investigate the genetic determinants of response or resistance to cancer therapy as well as factors that contribute to adverse events in the course of ...clinical trials. Despite the emergence of new technologies and the ability to measure more diverse analytes (e.g., circulating tumor cell (CTC), circulating tumor DNA (ctDNA), etc.), tumor tissue is still the most common and reliable source for biomarker investigation. Because of its worldwide use and ability to preserve samples for many decades at ambient temperature, formalin-fixed, paraffin-embedded tumor tissue (FFPE) is likely to be the preferred choice for tissue preservation in clinical practice for the foreseeable future. Multiple analyses are routinely performed on the same FFPE samples (such as
(IHC), in situ hybridization, RNAseq, DNAseq, TILseq, Methyl-Seq, etc.). Thus, specimen prioritization and optimization of the isolation of analytes is critical to ensure successful completion of each assay. FFPE is notorious for producing suboptimal DNA quality and low DNA yield. However, commercial vendors tend to request higher DNA sample mass than what is actually required for downstream assays, which restricts the breadth of biomarker work that can be performed. We evaluated multiple genomics service laboratories to assess the current state of NGS pre-analytical processing of FFPE. Significant differences in pre-analytical capabilities were observed. Key aspects are highlighted and recommendations are made to improve the current practice in translational research.
Over the past decade, next-generation sequencing (NGS) technology has experienced meteoric growth in the aspects of platform, technology, and supporting bioinformatics development allowing its ...widespread and rapid uptake in research settings. More recently, NGS-based genomic data have been exploited to better understand disease development and patient characteristics that influence response to a given therapeutic intervention. Cancer, as a disease characterized by and driven by the tumor genetic landscape, is particularly amenable to NGS-based diagnostic (Dx) approaches. NGS-based technologies are particularly well suited to studying cancer disease development, progression and emergence of resistance, all key factors in the development of next-generation cancer Dxs. Yet, to achieve the promise of NGS-based patient treatment, drug developers will need to overcome a number of operational, technical, regulatory, and strategic challenges. Here, we provide a succinct overview of the state of the clinical NGS field in terms of the available clinically targeted platforms and sequencing technologies. We discuss the various operational and practical aspects of clinical NGS testing that will facilitate or limit the uptake of such assays in routine clinical care. We examine the current strategies for analytical validation and Food and Drug Administration (FDA)-approval of NGS-based assays and ongoing efforts to standardize clinical NGS and build quality control standards for the same. The rapidly evolving companion diagnostic (CDx) landscape for NGS-based assays will be reviewed, highlighting the key areas of concern and suggesting strategies to mitigate risk. The review will conclude with a series of strategic questions that face drug developers and a discussion of the likely future course of NGS-based CDx development efforts.
The hadronic width of the ground state of pionic hydrogen has been redetermined by X-ray spectroscopy to be
Γ
1
s
π
H
=
(
856
±
16
stat
±
22
sys
)
meV. The experiment was performed at the ...high-intensity low-energy pion beam of the Paul Scherrer Institute by using the cyclotron trap and a high-resolution Bragg spectrometer with spherically bent crystals. Coulomb de-excitation was studied in detail by comparing its influence on the line shape by measuring the three different transitions K
α
, K
β
, and K
γ
at various hydrogen densities. The pion-nucleon scattering lengths and other physical quantities extracted from pionic-atom data are in good agreement with the results obtained from pion-nucleon and nucleon-nucleon scattering experiments and confirm that a consistent picture is achieved for the low-energy pion-nucleon sector with respect to the expectations of chiral perturbation theory.
Abnormal DNA CpG island hypermethylation and transcriptionally repressive histone modifications are associated with the aberrant silencing of tumor suppressor genes. Lysine methylation is a dynamic, ...enzymatically controlled process. Lysine-specific demethylase 1 (LSD1) has recently been identified as a histone lysine demethylase. LSD1 specifically catalyzes demethylation of mono- and dimethyl-lysine 4 of histone 3 (H3K4), key positive chromatin marks associated with transcriptional activation. We hypothesized that a novel class of oligoamine analogues would effectively inhibit LSD1 and thus cause the reexpression of aberrantly silenced genes.
Human colorectal cancer cells were treated with the oligoamines and changes in mono- and dimethyl-H3K4 and other chromatin marks were monitored. In addition, treated cells were evaluated for the reexpression of the aberrantly silenced secreted frizzled-related proteins (SFRP) Wnt signaling pathway antagonist genes. Finally, the effects of the LSD1 inhibitors were evaluated in an in vivo xenograft model.
Treatment of HCT116 human colon adenocarcinoma cells in vitro resulted in increased H3K4 methylation and reexpression of silenced SFRP genes. This reexpression is also accompanied by a decrease in H3K9me2 repressive mark. Importantly, cotreatment with low doses of oligoamines and a DNA methyltransferase inhibitor highly induces the reexpression of the aberrantly silenced SFRP2 gene and results in significant inhibition of the growth of established tumors in a human colon tumor model in vivo.
The use of LSD1-inhibiting oligoamine analogues in combination with DNA methyltransferase inhibitors represents a highly promising and novel approach for epigenetic therapy of cancer.
Genome-wide transcript profiling was used to monitor signal transduction during yeast pheromone response. Genetic manipulations allowed analysis of changes in gene expression underlying pheromone ...signaling, cell cycle control, and polarized morphogenesis. A two-dimensional hierarchical clustered matrix, covering 383 of the most highly regulated genes, was constructed from 46 diverse experimental conditions. Diagnostic subsets of coexpressed genes reflected signaling activity, cross talk, and overlap of multiple mitogen-activated protein kinase (MAPK) pathways. Analysis of the profiles specified by two different MAPKs-Fus3p and Kss1p-revealed functional overlap of the filamentous growth and mating responses. Global transcript analysis reflects biological responses associated with the activation and perturbation of signal transduction pathways.
The K¯N system at threshold is a sensitive testing ground for low energy QCD, especially for the explicit chiral symmetry breaking. Therefore, we have measured the K-series X-rays of kaonic hydrogen ...atoms at the DAΦNE electron–positron collider of Laboratori Nazionali di Frascati, and have determined the most precise values of the strong-interaction energy-level shift and width of the 1s atomic state. As X-ray detectors, we used large-area silicon drift detectors having excellent energy and timing resolution, which were developed especially for the SIDDHARTA experiment. The shift and width were determined to be ϵ1s=−283±36(stat)±6(syst) eV and Γ1s=541±89(stat)±22(syst) eV, respectively. The new values will provide vital constraints on the theoretical description of the low-energy K¯N interaction.
Synthesizing polycationic polymers directly from existing drugs overcomes the drug-loading limitations often associated with pharmacologically inert nanocarriers. We recently described nanocarriers ...formed from a first-generation polyamine analogue, bis(ethyl)norspermine (BENSpm), that could simultaneously target polyamine metabolism while delivering therapeutic nucleic acids. In the current study, we describe the synthesis and evaluation of self-immolative nanocarriers derived from the second-generation polyamine analogue PG-11047. Polyamines are absolutely essential for proliferation and their metabolism is frequently dysregulated in cancer. Through its effects on polyamine metabolism, PG-11047 effectively inhibits tumor growth in cancer cell lines of multiple origins as well as in human tumor mouse xenografts. Promising clinical trials have been completed verifying the safety and tolerance of this rotationally restricted polyamine analogue. We therefore used PG-11047 as the basis for Nano11047, a biodegradable, prodrug nanocarrier capable of targeting polyamine metabolism. Following exposure of lung cancer cell lines to Nano11047, uptake and intracellular degradation into the parent compound PG-11047 was observed. The release of PG-11047 highly induced the polyamine catabolic enzyme activities of spermidine/spermine N1-acetyltransferase (SSAT) and spermine oxidase (SMOX). By contrast, the activity of ornithine decarboxylase (ODC), a rate-limiting enzyme in polyamine biosynthesis and a putative oncogene, was decreased. Consequently, intracellular levels of the natural polyamines were depleted concurrent with tumor cell growth inhibition. This availability of Nano11047 as a novel drug form and potential nucleic acid delivery vector will potentially benefit and encourage future clinical studies.
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