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

New treatments for anemia, ischemia, and cancer emerging from studies of von Hippel-Lindau Disease

Detailed formal protocol with illustrations and extensive bibliographyA recording of the protocol presentation is available on UT Southwestern’s Mediasite. Note: Access to the video is restricted to authorized UT Southwestern users only.UT Southwestern--Internal Medicin

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

Effect of HIF2α shRNA Is Specifically Due to Downregulation of HIF2α

<div><p>(A) Parental 786-O cells (<i>VHL</i>[−/−]) and 786-O cells stably producing HIF2α shRNA #3 that were coinfected with an empty retrovirus (Empty) or a retrovirus encoding a HIF2α mRNA with three silent mutations in the #3 recognition site (MT*) 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 HIF2α shRNA #3 cells infected with the indicated retroviruses.</p> <p>(C) Tumor weights approximately 9 wk after subcutaneous implantation of 786-O HIF2α shRNA 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 HIF2α shRNA #3 cells in left (upper) flank and 786-O HIF2α shRNA #3 cells infected with retrovirus encoding HIF2α MT* mRNA on right (lower) flank.</p></div

Tumor Suppression by HIF2α shRNA Is Not Restricted to a Single Cell Line

<div><p>(A) Tumor weights approximately 8 wk after subcutaneous implantation of A498 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>(B) Representative histological sections after staining with hematoxylin and eosin of tumors formed by A498 cells infected with the indicated retroviruses.</p></div