Antiproliferative Activity of Cephalotaxus Esters: Overcoming Chemoresistance.

Introduction: Omacetaxine, a semisynthetic form of Homoharringtonine (HHT), was approved for the treatment of Chronic Myeloid Leukemia (CML). Previously, we have published the synthesis of this natural alkaloid and three of its derivatives: Deoxyharringtonine (DHT), Deoxyhomoharringtonine (DHHT), and Bis(demethyl)-deoxyharringtonine (BDHT), and reported its refractory activity against the HL-60/RV+ cells over-expressing P-glycoprotein 1 (MDR1).

Methods: In this study, we have explored the extent of this resistance by first expanding the panel of established cell lines and using a panel of 21 leukemia patient-derived primary cells.

A Genome-Wide RNAi Screen Reveals Common Host-Virus Gene Signatures: Implication for Dengue Antiviral Drug Discovery.

Dengue is the most common mosquito-borne viral disease that in recent years has become a major international public health concern. Dengue is a tropical neglected disease with increasing global incidences, affecting millions of people worldwide, and without the availability of specific treatments to combat it. The identification of host-target genes essential for the virus life cycle, for which effective modulators may already exist, would provide an alternative path to a rapid drug development of the much needed antidengue agents.

Dual Fluorescence Isogenic Synthetic Lethal Kinase Screen and High-Content Secondary Screening for MUC16/CA125-Selective Agents.

Significant strides have been made in the development of precision therapeutics for cancer. Aberrantly expressed glycoproteins represent a potential avenue for therapeutic development. The MUC16/CA125 glycoprotein serves as a biomarker of disease and a driver of malignant transformation in epithelial ovarian cancer.

Characterization of a small molecule inhibitor of disulfide reductases that induces oxidative stress and lethality in lung cancer cells.

Phenotype-based screening can identify small molecules that elicit a desired cellular response, but additional approaches are required to characterize their targets and mechanisms of action. Here, we show that a compound termed LCS3, which selectively impairs the growth of human lung adenocarcinoma (LUAD) cells, induces oxidative stress. To identify the target that mediates this effect, we use thermal proteome profiling (TPP) and uncover the disulfide reductases GSR and TXNRD1 as targets.

A High-Content Screen for C/EBPα Expression Identifies Novel Therapeutic Agents in Dedifferentiated Liposarcoma.

Purpose: Dedifferentiated liposarcoma (DDLS), one of the most common and aggressive sarcomas, infrequently responds to chemotherapy. DDLS survival and growth depend on underexpression of C/EBPα, a tumor suppressor and transcriptional regulator controlling adipogenesis. We sought to screen and prioritize candidate drugs that increase C/EBPα expression and may therefore serve as differentiation-based therapies for DDLS.

Hepatitis B virus and hepatitis delta virus subtypes circulating in Algeria and seroprevalence of HDV infection.

Background: Little is known about molecular characteristics of HBV strains circulating in Algeria and there are few data regarding HDV infection.

Objectives: The aim of this study is to describe the genetic diversity of HBV and HDV strains existing in Algeria and to determine the seroprevalence of HDV infection.

Study Design: Plasma samples from 134 patients were analyzed by enzyme immunoassay method for HBV and HDV serological markers.

BET Inhibition-Induced GSK3β Feedback Enhances Lymphoma Vulnerability to PI3K Inhibitors.

The phosphatidylinositol 3 kinase (PI3K)-glycogen synthase kinase β (GSK3β) axis plays a central role in MYC-driven lymphomagenesis, and MYC targeting with bromodomain and extraterminal protein family inhibitors (BETi) is a promising treatment strategy in lymphoma. In a high-throughput combinatorial drug screening experiment, BETi enhance the antiproliferative effects of PI3K inhibitors in a panel of diffuse large B cell lymphoma (DLBCL) and Burkitt lymphoma cell lines. BETi or MYC silencing upregulates several PI3K pathway genes and induces GSK3β S9 inhibitory phosphorylation, resulting in increased β-catenin protein abundance.

Evaluation of Compound Optical Interference in High-Content Screening.

Compound optical interference remains an inherent problem in chemical screening and has been well documented for biochemical assays and less so for automated microscopy-based assays. It has also been the assumption that the latter should not suffer from such interference because of the washing steps involved in the process, thus eliminating the residual nonspecific compound effects. Instead, these compounds may have no relevance to the actual target, and as such, compound optical interference contributes to a number of false-positives, resulting in a high attrition rate during subsequent follow-up studies.

ABCG2 regulates self-renewal and stem cell marker expression but not tumorigenicity or radiation resistance of glioma cells.

Glioma cells with stem cell traits are thought to be responsible for tumor maintenance and therapeutic failure. Such cells can be enriched based on their inherent drug efflux capability mediated by the ABC transporter ABCG2 using the side population assay, and their characteristics include increased self-renewal, high stem cell marker expression and high tumorigenic capacity in vivo. Here, we show that ABCG2 can actively drive expression of stem cell markers and self-renewal in glioma cells.

Proteasome Addiction Defined in Ewing Sarcoma Is Effectively Targeted by a Novel Class of 19S Proteasome Inhibitors.

Ewing sarcoma is a primitive round cell sarcoma with a peak incidence in adolescence that is driven by a chimeric oncogene created from the fusion of the EWSR1 gene with a member of the ETS family of genes. Patients with metastatic and recurrent disease have dismal outcomes and need better therapeutic options. We screened a library of 309,989 chemical compounds for growth inhibition of Ewing sarcoma cells to provide the basis for the development of novel therapies and to discover vulnerable pathways that might broaden our understanding of the pathobiology of this aggressive sarcoma.

Nanomolar Inhibitors of Trypanosoma brucei RNA Triphosphatase.

Unlabelled: Eukaryal taxa differ with respect to the structure and mechanism of the RNA triphosphatase (RTPase) component of the mRNA capping apparatus. Protozoa, fungi, and certain DNA viruses have a metal-dependent RTPase that belongs to the triphosphate tunnel metalloenzyme (TTM) superfamily. Because the structures, active sites, and chemical mechanisms of the TTM-type RTPases differ from those of mammalian RTPases, the TTM RTPases are potential targets for antiprotozoal, antifungal, and antiviral drug discovery.

Azaphilones from an Acid Mine Extremophile Strain of a Pleurostomophora sp.

An extremophilic fungus identified as a Pleurostomophora sp. was isolated from the Berkeley Pit, an acid mine waste lake. When grown in liquid culture, the fungus produced berkchaetoazaphilones A-C (1, 2, and 5), the red pigment berkchaetorubramine (6), and the known compound 4-(hydroxymethyl)quinoline.

A High-Content Assay to Screen for Modulators of EGFR Function.

Cell-based assays have the potential and advantage to identify cell-permeable modulators of kinase function, and hence provide an alternative to the conventional enzymatic activity-driven discovery approaches that rely on purified recombinant kinase catalytic domains. Here, we describe a domain-based high-content biosensor approach to study endogenous EGFR activity whereby EGF-induced receptor activation, subsequent trafficking, and internalization are imaged and quantified using time-dependent granule formation in cells. This method can readily be used to search for EGFR modulators in both chemical and RNAi screening; with potential applicability to other receptor tyrosine kinases.

Pharmacokinetics, Safety, and Efficacy of Intravitreal Digoxin in Preclinical Models for Retinoblastoma.

Purpose: To assess in vitro cytotoxic activity and antiangiogenic effect, ocular and systemic disposition, and toxicity of digoxin in rabbits after intravitreal injection as a potential candidate for retinoblastoma treatment.

Methods: A panel of two retinoblastoma and three endothelial cell types were exposed to increasing concentrations of digoxin in a conventional (72-hour exposure) and metronomic (daily exposure) treatment scheme. Cytotoxicity was defined as the digoxin concentration that killed 50% of the cells (IC50) and was assessed with a vital dye in all cell types.

A Multiplexed Cell-Based Assay for the Identification of Modulators of Pre-Membrane Processing as a Target against Dengue Virus.

The DenV pre-membrane protein (prM) is a crucial chaperone for the viral envelope protein, preventing premature fusion with vesicles during viral export. prM molecules in immature particles are cleaved by host proteases, leading to mature fusogenic virions. Blockade of prM cleavage would restrict fusion and represents a novel druggable opportunity against DenV.

Asparagine plays a critical role in regulating cellular adaptation to glutamine depletion.

Many cancer cells consume large quantities of glutamine to maintain TCA cycle anaplerosis and support cell survival. It was therefore surprising when RNAi screening revealed that suppression of citrate synthase (CS), the first TCA cycle enzyme, prevented glutamine-withdrawal-induced apoptosis. CS suppression reduced TCA cycle activity and diverted oxaloacetate, the substrate of CS, into production of the nonessential amino acids aspartate and asparagine.

Novel imidazoline antimicrobial scaffold that inhibits DNA replication with activity against mycobacteria and drug resistant Gram-positive cocci.

Bacterial antimicrobial resistance is an escalating public health threat, yet the current antimicrobial pipeline remains alarmingly depleted, making the development of new antimicrobials an urgent need. Here, we identify a novel, potent, imidazoline antimicrobial compound, SKI-356313, with bactericidal activity against Mycobacterium tuberculosis and Gram-positive cocci, including vancomycin-resistant Enterococcus faecium (VRE) and methicillin-resistant Staphylococcus aureus (MRSA). SKI-356313 is active in murine models of Streptococcus pneumoniae and MRSA infection and is potently bactericidal for both replicating and nonreplicating M.

Small-molecule inhibitors of SETD8 with cellular activity.

SETD8/SET8/Pr-SET7/KMT5A is the sole protein lysine methyltransferase (PKMT) known to monomethylate lysine 20 of histone H4 in vivo. SETD8's methyltransferase activity has been implicated in many essential cellular processes including DNA replication, DNA damage response, transcription modulation, and cell cycle regulation. Developing SETD8 inhibitors with cellular activity is a key step toward elucidating the diverse roles of SETD8 via convenient pharmacological perturbation.

Discovery of a dicer-independent, cell-type dependent alternate targeting sequence generator: implications in gene silencing & pooled RNAi screens.

There is an acceptance that plasmid-based delivery of interfering RNA always generates the intended targeting sequences in cells, making it as specific as its synthetic counterpart. However, recent studies have reported on cellular inefficiencies of the former, especially in light of emerging gene discordance at inter-screen level and across formats. Focusing primarily on the TRC plasmid-based shRNA hairpins, we reasoned that alleged specificities were perhaps compromised due to altered processing; resulting in a multitude of random interfering sequences.

Plasmid-based shRNA lentiviral particle production for RNAi applications.

Lentiviral vectors have become mainstream gene transfer vehicles for their ability to deliver and integrate into host cells. In RNA interference (RNAi) applications, lentiviral constructs constitutively express double-stranded RNA molecules, usually as short hairpin RNA (shRNA), enabling long-term gene silencing, and, when pseudotyped with a broad host glycoprotein envelope, allow a multitude of cell types to be transduced. Their successful use ultimately relies on the production of lentiviral particles in high titer and uniformity.

A 1536-well fluorescence polarization assay to screen for modulators of the MUSASHI family of RNA-binding proteins.

RNA-binding proteins (RBPs) can act as stem cell modulators and oncogenic drivers, but have been largely ignored by the pharmaceutical industry as potential therapeutic targets for cancer. The MUSASHI (MSI) family has recently been demonstrated to be an attractive clinical target in the most aggressive cancers. Therefore, the discovery and development of small molecule inhibitors could provide a novel therapeutic strategy.