Inhibition of vascular endothelial growth factor with a sequence-specific hypoxia response element antagonist

Vascular endothelial growth factor (VEGF) and its receptors have been implicated as key factors in tumor angiogenesis that are up-regulated by hypoxia. We evaluated the effects of DNA-binding small molecules on hypoxia-inducible transcription of VEGF. A synthetic pyrrole-imidazole polyamide designed to bind the hypoxia response element (HRE) was found to disrupt hypoxia-inducible factor (HIF) binding to HIRE. In cultured HeLa cells, this resulted in a reduction of VEGF mRNA and secreted protein levels. The observed effects were polyamide-specific and dose-dependent. Analysis of genome-wide effects of the HIRE-specific polyamide revealed that a number of hypoxia-inducible genes were down-regulated. Pathway-based regulation of hypoxia-inducible gene expression with DNA-binding small molecules may represent a new approach for targeting angiogenesis

Synthetic lethality: a framework for the development of wiser cancer therapeutics

The challenge in medical oncology has always been to identify compounds that will kill, or at least tame, cancer cells while leaving normal cells unscathed. Most chemotherapeutic agents in use today were selected primarily for their ability to kill rapidly dividing cancer cells grown in cell culture and in mice, with their selectivity determined empirically during subsequent animal and human testing. Unfortunately, most of the drugs developed in this way have relatively low therapeutic indices (low toxic dose relative to the therapeutic dose). Recent advances in genomics are leading to a more complete picture of the range of mutations, both driver and passenger, present in human cancers. Synthetic lethality provides a conceptual framework for using this information to arrive at drugs that will preferentially kill cancer cells relative to normal cells. It also provides a possible way to tackle 'undruggable' targets. Two genes are synthetically lethal if mutation of either gene alone is compatible with viability but simultaneous mutation of both genes leads to death. If one is a cancer-relevant gene, the task is to discover its synthetic lethal interactors, because targeting these would theoretically kill cancer cells mutant in the cancer-relevant gene while sparing cells with a normal copy of that gene. All cancer drugs in use today, including conventional cytotoxic agents and newer 'targeted' agents, target molecules that are present in both normal cells and cancer cells. Their therapeutic indices almost certainly relate to synthetic lethal interactions, even if those interactions are often poorly understood. Recent technical advances enable unbiased screens for synthetic lethal interactors to be undertaken in human cancer cells. These approaches will hopefully facilitate the discovery of safer, more efficacious anticancer drugs that exploit vulnerabilities that are unique to cancer cells by virtue of the mutations they have accrued during tumor progression

2-Oxoglutarate-dependent dioxygenases in cancer

Abstract 2-Oxoglutarate-dependent dioxygenases (2OGDDs) are a superfamily of enzymes that play diverse roles in many biological processes, including regulation of hypoxia-inducible factor-mediated adaptation to hypoxia, extracellular matrix formation, epigenetic regulation of gene transcription and the reprogramming of cellular metabolism. 2OGDDs all require oxygen, reduced iron and 2-oxoglutarate (also known as α-ketoglutarate) to function, although their affinities for each of these co-substrates, and hence their sensitivity to depletion of specific co-substrates, varies widely. Numerous 2OGDDs are recurrently dysregulated in cancer. Moreover, cancer-specific metabolic changes, such as those that occur subsequent to mutations in the genes encoding succinate dehydrogenase, fumarate hydratase or isocitrate dehydrogenase, can dysregulate specific 2OGDDs. This latter observation suggests that the role of 2OGDDs in cancer extends beyond cancers that harbour mutations in the genes encoding members of the 2OGDD superfamily. Herein, we review the regulation of 2OGDDs in normal cells and how that regulation is corrupted in cancer

HIF2α Overrides Tumor Suppression by pVHL

<div><p>(A) 786-O subclones that were transfected to produce wild-type pVHL (WT8) or with an empty plasmid (PRC3) cells, as well as WT8 cells infected with an empty retrovirus (Empty) or retroviruses encoding the indicated HIF2α variants [ (P→A)² = P405A;P531A and * = bHLH mutation] were grown in the presence of 21% or 1% oxygen and immunoblotted (IB) with the indicated antibodies.</p> <p>(B) In vitro proliferation of WT8 cells infected with the indicated retroviruses.</p> <p>(C) Tumor weights approximately 9 wk after subcutaneous implantation of WT8 cells infected with the indicated retroviruses in <i>nude</i> mice. Number of tumors analyzed is shown in parentheses. Error bars = one standard error.</p></div

Downregulation of HIF2α Is Sufficient to Suppress Tumor Growth by pVHL-Defective Renal Carcinoma Cells

<div><p>(A) Parental 786-O cells (<i>VHL</i>[−/−]) and 786-O cells infected with an empty retrovirus (Empty) or retroviruses encoding HIF2α shRNAs (sequence #2 or #3) were grown in the presence of 21% or 1% oxygen and immunoblotted (IB) with the indicated antibodies.</p> <p>(B) In vitro proliferation of 786-O cells infected with the indicated retroviruses.</p> <p>(C) Tumor weights approximately 9 wk after subcutaneous implantation of 786-O cells infected with the indicated retroviruses in <i>nude</i> mice. Number of tumors analyzed is shown in parentheses. Error bars = one standard error.</p> <p>(D) Representative photograph of <i>nude</i> mouse 9 wk after subcutaneous injection of 786-O cells in left (upper) flank and 786-O cells infected with HIF2α shRNA (#3) retrovirus on right (lower) flank.</p></div

Degradation of the SCF component Skp2 in cell-cycle phase G1 by the anaphase-promoting complex

Cell-cycle transitions are driven by waves of ubiquitin-dependent degradation of key cell-cycle regulators. SCF (Skp1/Cullin/F-box protein) complexes and anaphase-promoting complexes (APC) represent two major classes of ubiquitin ligases whose activities are thought to regulate primarily the G1/S and metaphase/anaphase cell-cycle transitions, respectively. The major target of the Skp1/Cul1/Skp2 (SCF(SKP2)) complex is thought to be the Cdk inhibitor p27 during S phase, whereas the principal targets for the APC are thought to be involved in chromatid separation (securin) and exit from mitosis (cyclin B). Although the role of the APC in mitosis is relatively clear, there is mounting evidence that APCs containing Cdh1 (APC(CDH1)) also have a function in the G1 phase of the cell cycle. Here, we show that the F-box protein Skp2 is polyubiquitinated, and hence earmarked for destruction, by APC(CDH1). As a result, accumulation of SCF(SKP2) requires prior inactivation of APC(CDH1). These findings provide an insight into the orchestration of SCF and APC activities during cell-cycle progression, and into the involvement of the APC in G1