Targeting HIF-activated collagen prolyl 4-hydroxylase expression disrupts collagen deposition and blocks primary and metastatic uveal melanoma growth.

Uveal melanoma (UM) is the most prevalent primary intraocular malignancy in adults, and patients that develop metastases (~50%) survive <1 year, highlighting the urgent need for new therapies. TCGA has recently revealed that a hypoxia gene signature is associated with poor UM patient prognosis. Here we show that expression of hypoxia-regulated collagen prolyl-4-hydroxylase genes P4HA1 and P4HA2 is significantly upregulated in UM patients with metastatic disease and correlates with poor prognosis, suggesting these enzymes might be key tumor drivers.

Correction: EZH2 targeting reduces medulloblastoma growth through epigenetic reactivation of the BAI1/p53 tumor suppressor pathway.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

EZH2 targeting reduces medulloblastoma growth through epigenetic reactivation of the BAI1/p53 tumor suppressor pathway.

Medulloblastoma (MB) is a malignant pediatric brain tumor for which new therapies are urgently needed. We demonstrate that treatment with EPZ-6438 (Tazemetostat), an enhancer of zeste homolog 2 (EZH2) inhibitor approved for clinical trials, blocks MB cell growth in vitro and in vivo, and prolongs survival in orthotopic xenograft models. We show that the therapeutic effect is dependent on epigenetic reactivation of adhesion G-protein-coupled receptor B1 (BAI1/ADGRB1), a tumor suppressor that controls p53 stability by blocking Mdm2.

Arylsulfonamide 64B Inhibits Hypoxia/HIF-Induced Expression of c-Met and CXCR4 and Reduces Primary Tumor Growth and Metastasis of Uveal Melanoma.

Purpose: Uveal melanoma (UM) is the most prevalent and lethal intraocular malignancy in adults. Here, we examined the importance of hypoxia in UM growth and tested the antitumor effects of arylsulfonamide 64B, an inhibitor of the hypoxia-induced factor (HIF) pathway in animal models of UM and investigated the related mechanisms.

Experimental Design: UM cells were implanted in the uvea of mice eyes and mice systemically treated with 64B.

A novel small-molecule arylsulfonamide causes energetic stress and suppresses breast and lung tumor growth and metastasis.

Neoplastic cells display reprogrammed metabolism due to the heightened energetic demands and the need for biomass synthesis of a growing tumor. Targeting metabolic vulnerabilities is thus an important goal for cancer therapy. Here, we describe a novel small-molecule arylsulfonamide (N-cyclobutyl-N-((2,2-dimethyl-2-pyrano[3,2-]pyridin-6-yl)methyl)-3,4-dimethoxybenzenesulfonamide) that exerts potent cytotoxicity and energetic stress on tumor cells while largely sparing non-cancerous human cells.

Purifying Properly Folded Cysteine-rich, Zinc Finger Containing Recombinant Proteins for Structural Drug Targeting Studies: the CH1 Domain of p300 as a Case Example.

The transcription factor Hypoxia-Inducible Factor (HIF) complexes with the coactivator p300, activating the hypoxia response pathway and allowing tumors to grow. The CH1 and CAD domains of each respective protein form the interface between p300 and HIF. Small molecule compounds are in development that target and inhibit HIF/p300 complex formation, with the goal of reducing tumor growth.

Design and synthesis of benzopyran-based inhibitors of the hypoxia-inducible factor-1 pathway with improved water solubility.

While progress has been made in treating cancer, cytotoxic chemotherapeutic agents are still the most widely used drugs and are associated with severe side-effects. Drugs that target unique molecular signalling pathways are needed for treating cancer with low or no intrinsic toxicity to normal cells. Our goal is to target hypoxic tumours and specifically the hypoxia inducible factor (HIF) pathway for the development of new cancer therapies.

A role for activated Cdc42 in glioblastoma multiforme invasion.

Cdc42 is a Rho-GTPase which plays a major role in regulating cell polarity and migration by specifying the localization of filopodia. However, the role of Cdc42 in GBM invasion has not been thoroughly investigated. We generated stable doxycycline-inducible clones expressing wild type (WT)-, constitutively active (CA)-, and dominant negative (DN)-Cdc42 in three different human glioma cell lines.

Tyr phosphorylation of PDP1 toggles recruitment between ACAT1 and SIRT3 to regulate the pyruvate dehydrogenase complex.

Mitochondrial pyruvate dehydrogenase complex (PDC) is crucial for glucose homeostasis in mammalian cells. The current understanding of PDC regulation involves inhibitory serine phosphorylation of pyruvate dehydrogenase (PDH) by PDH kinase (PDK), whereas dephosphorylation of PDH by PDH phosphatase (PDP) activates PDC. Here, we report that lysine acetylation of PDHA1 and PDP1 is common in epidermal growth factor (EGF)-stimulated cells and diverse human cancer cells.

Hypoxia inducible factor pathway inhibitors as anticancer therapeutics.

Hypoxia is a significant feature of solid tumor cancers. Hypoxia leads to a more malignant phenotype that is resistant to chemotherapy and radiation, is more invasive and has greater metastatic potential. Hypoxia activates the hypoxia inducible factor (HIF) pathway, which mediates the biological effects of hypoxia in tissues.

Arylsulfonamide KCN1 inhibits in vivo glioma growth and interferes with HIF signaling by disrupting HIF-1α interaction with cofactors p300/CBP.

Purpose: The hypoxia-inducible factor-1 (HIF-1) plays a critical role in tumor adaptation to hypoxia, and its elevated expression correlates with poor prognosis and treatment failure in patients with cancer. In this study, we determined whether 3,4-dimethoxy-N-[(2,2-dimethyl-2H-chromen-6-yl)methyl]-N-phenylbenzenesulfonamide, KCN1, the lead inhibitor in a novel class of arylsulfonamide inhibitors of the HIF-1 pathway, had antitumorigenic properties in vivo and further defined its mechanism of action.

Experimental Design: We studied the inhibitory effect of systemic KCN1 delivery on the growth of human brain tumors in mice.

Binding Model for the Interaction of Anticancer Arylsulfonamides with the p300 Transcription Cofactor.

Hypoxia inducible factors (HIFs) are transcription factors that activate expression of multiple gene products and promote tumor adaptation to a hypoxic environment. To become transcriptionally active, HIFs associate with cofactors p300 or CBP. Previously, we found that arylsulfonamides can antagonize HIF transcription in a bioassay, block the p300/HIF-1α interaction, and exert potent anticancer activity in several animal models.

Design and synthesis of novel small-molecule inhibitors of the hypoxia inducible factor pathway.

Hypoxia, a reduction in partial oxygen pressure, is a salient property of solid tumors. Hypoxia drives malignant progression and metastasis in tumors and participates in tumor resistance to radio- and chemotherapies. Hypoxia activates the hypoxia-inducible factor (HIF) family of transcription factors, which induce target genes that regulate adaptive biological processes such as anaerobic metabolism, cell motility, and angiogenesis.

Sulfonamides as a new scaffold for hypoxia inducible factor pathway inhibitors.

Solid tumors generally grow under hypoxic conditions, a pathophysiological change, which activates the expression of genes responsible for malignant, aggressive, and treatment-refractory properties. Hypoxia inducible factor (HIF) is the chief transcription factor regulating hypoxia-driven gene expression. Therefore, the HIF pathway has become a critical target for cancer therapeutics development.

Mitogen-activated protein/extracellular signal-regulated kinase kinase 1act/tubulin interaction is an important determinant of mitotic stability in cultured HT1080 human fibrosarcoma cells.

Activation of the mitogen-activated protein kinase (MAPK) pathway plays a major role in neoplastic cell transformation. Using a proteomics approach, we identified alpha tubulin and beta tubulin as proteins that interact with activated MAP/extracellular signal-regulated kinase kinase 1 (MEK1), a central MAPK regulatory kinase. Confocal analysis revealed spatiotemporal control of MEK1-tubulin colocalization that was most prominent in the mitotic spindle apparatus in variant HT1080 human fibrosarcoma cells.

Transcriptional control of the tumor- and hypoxia-marker carbonic anhydrase 9: A one transcription factor (HIF-1) show?

Transcriptional activation by hypoxia is mediated by the hypoxia-inducible factor (HIF) via binding to the hypoxia-responsive element (HRE). Hypoxia in solid tumors associates with poorer outcome of the disease and reliable cellular markers of tumor hypoxia would represent a valuable diagnostic marker and a potential therapeutic target. In this category, carbonic anhydrase IX (CAIX) is one of the most promising candidates.

PI3K/Akt activity has variable cell-specific effects on expression of HIF target genes, CA9 and VEGF, in human cancer cell lines.

The phosphatidylinositol 3-kinase/Akt (PI3K) pathway regulates hypoxia-inducible factor (HIF) activity. Higher expression of HIF-1alpha and carbonic anhydrase IX (CAIX), a hypoxia-inducible gene, in HT10806TG fibrosarcoma cells (mutant N-ras allele), compared to derivative MCH603 cells (deleted mutant N-ras allele), correlated with increased PI3K activity. Constitutive activation of the PI3K pathway in MCH603/PI3K(act) cells increased HIF-1alpha but, surprisingly, decreased CAIX levels.

Regulation of gene expression by hypoxia: integration of the HIF-transduced hypoxic signal at the hypoxia-responsive element.

Cells experiencing lowered O(2) levels (hypoxia) undergo a variety of biological responses in order to adapt to these unfavorable conditions. The master switch, orchestrating the cellular response to low O(2) levels, is the transcription factor, termed hypoxia-inducible factor (HIF). The alpha subunits of HIF are regulated by 2-oxoglutarate-dependent oxygenases that, in the presence of O(2), hydroxylate specific prolyl and asparaginyl residues of HIF-alpha, inducing its proteasome-dependent degradation and repression of transcriptional activity, respectively.

Rational design of minimal hypoxia-inducible enhancers.

The hypoxia-inducible factor (HIF) activates transcription via binding to the highly variable hypoxia-responsive elements (HREs). All hypoxia-inducible constructs described to date utilize multimers of naturally occurring HREs. Here, we describe the rational design of minimal hypoxia-inducible enhancers, conceptually equivalent to using an optimized HIF-binding site (HBS) as the building block.

Does inhibition of degradation of hypoxia-inducible factor (HIF) alpha always lead to activation of HIF? Lessons learnt from the effect of proteasomal inhibition on HIF activity.

At the cellular level hypoxia induces transcriptional response that is mediated by the transcription factor hypoxia-inducible factor (HIF). HIF is regulated at the level of its alpha subunit by 2-oxoglutarate (2OG)-dependent oxygenases that hydroxylate specific prolyl and asparaginyl residues of HIF-alpha, affecting its stability and activity, respectively. In the presence of O(2), the alpha subunit is degraded in a complex process with several distinct steps.

Proteasomal inhibition attenuates transcriptional activity of hypoxia-inducible factor 1 (HIF-1) via specific effect on the HIF-1alpha C-terminal activation domain.

The ubiquitin-proteasome pathway (UPP) is involved in regulation of multiple cellular processes. Hypoxia-inducible factor 1 alpha (HIF-1 alpha) is a prototypic target of the UPP and, as such, is stabilized under conditions of proteasomal inhibition. Using carbonic anhydrase IX (CAIX) and vascular endothelial growth factor (VEGF) expression as paradigmatic markers of HIF-1 activity, we found that proteasomal inhibitors (PI) abrogated hypoxia-induced CAIX expression in all cell lines tested and VEGF expression in two out of three.

The role of extracellular signal-regulated protein kinase in transcriptional regulation of the hypoxia marker carbonic anhydrase IX.

In the present study, we investigated the role of the extracellular signal-regulated protein kinase (ERK) in regulation of the hypoxia marker, carbonic anhydrase IX (CAIX). U0126, a specific inhibitor of MEK1/2, downregulated CAIX expression induced by true hypoxia and cell density. CA9 promoter activity was similarly affected.