Yu-Dong Zhou

The marine sponge metabolite mycothiazole: A novel prototype mitochondrial complex I inhibitor

A natural product chemistry-based approach was applied to discover small-molecule inhibitors of hypoxia-inducible factor-1 (HIF-1). A Petrosaspongia mycofijiensis marine sponge extract yielded mycothiazole (1), a solid tumor selective compound with no known mechanism for its cell line-dependent cytotoxic activity. Compound 1 inhibited hypoxic HIF-1 signaling in tumor cells (IC(50) 1nM) that correlated with the suppression of hypoxia-stimulated tumor angiogenesis in vitro. However, 1 exhibited pronounced neurotoxicity in vitro. Mechanistic studies revealed that 1 selectively suppresses mitochondrial respiration at complex I (NADH-ubiquinone oxidoreductase). Unlike rotenone, MPP(+), annonaceous acetogenins, piericidin A, and other complex I inhibitors, mycothiazole is a mixed polyketide/peptide-derived compound with a central thiazole moiety. The exquisite potency and structural novelty of 1 suggest that it may serve as a valuable molecular probe for mitochondrial biology and HIF-mediated hypoxic signaling.

Mechanism Targeted Discovery of Antitumor Marine Natural Products

Antitumor drug discovery programs aim to identify chemical entities for use in the treatment of cancer. Many strategies have been used to achieve this objective. Natural products have always played a major role in anticancer medicine and the unique metabolites produced by marine organisms have increasingly become major players in antitumor drug discovery. Rapid advances have occurred in the understanding of tumor biology and molecular medicine. New insights into mechanisms responsible for neoplastic disease are significantly changing the general philosophical approach towards cancer treatment. Recently identified molecular targets have created exciting new means for disrupting tumor-specific cell signaling, cell division, energy metabolism, gene expression, drug resistance and blood supply. Such tumor-specific treatments could someday decrease our reliance on traditional cytotoxicity-based chemotherapy and provide new less toxic treatment options with significantly fewer side effects. Novel molecular targets and state-of-the-art, molecular mechanism-based screening methods have revitalized antitumor research and these changes are becoming an ever-increasing component of modern antitumor marine natural products research. This review describes marine natural products identified using tumor-specific mechanism-based assays for regulators of angiogenesis, apoptosis, cell cycle, macromolecule synthesis, mitochondrial respiration, mitosis, multidrug efflux and signal transduction. Special emphasis is placed on natural products directly discovered using molecular mechanism-based screening.

Natural Product-Based Inhibitors of Hypoxia-Inducible Factor-1 (HIF-1)

The transcription factor hypoxia-inducible factor-1 (HIF-1) regulates the expression of more than 70 genes involved in cellular adaptation and survival under hypoxic stress. Activation of HIF-1 is associated with numerous physiological and pathological processes that include tumorigenesis, vascular remodeling, inflammation, and hypoxia/ischemia-related tissue damage. Clinical studies suggested that HIF-1 activation correlates directly with advanced disease stages and treatment resistance among cancer patients. Preclinical studies support the inhibition of HIF-1 as a major molecular target for antitumor drug discovery. Considerable effort is underway, in government laboratories, industry and academia, to identify therapeutically useful small molecule HIF-1 inhibitors. Natural products (low molecular weight organic compounds produced by plants, microbes, and animals) continue to play a major role in modern antitumor drug discovery. Most of the compounds discovered to inhibit HIF-1 are natural products or synthetic compounds with structures that are based on natural product leads. Natural products have also served a vital role as molecular probes to elucidate the pathways that regulate HIF-1 activity. Natural products and natural product-derived compounds that inhibit HIF-1 are summarized in light of their biological source, chemical class, and effect on HIF-1 and HIF-mediated gene regulation. When known, the mechanism(s) of action of HIF-1 inhibitors are described. Many of the substances found to inhibit HIF-1 are non-druggable compounds that are too cytotoxic to serve as drug leads. The application of high-throughput screening methods, complementary molecular-targeted assays, and structurally diverse chemical libraries hold promise for the discovery of therapeutically useful HIF-1 inhibitors.

Latrunculin A and Its C-17-O-Carbamates Inhibit Prostate Tumor Cell Invasion and HIF-1 Activation in Breast Tumor Cells

The marine-derived macrolides latrunculins A ( 1) and B, from the Red Sea sponge Negombata magnifica, have been found to reversibly bind actin monomers, forming a 1:1 complex with G-actin and disrupting its polymerization. The microfilament protein actin is responsible for several essential functions within the cell such as cytokinesis and cell migration. One of the main binding pharmacophores of 1 to G-actin was identified as the C-17 lactol hydroxyl moiety that binds arginine 210 NH. Latrunculin A-17- O-carbamates 2- 6 were prepared by reaction with the corresponding isocyanates. Latrunculin A ( 1) and carbamates 4- 6 displayed potent anti-invasive activity against the human highly metastatic human prostate cancer PC-3M cells in a Matrigel assay at a concentration range of 50 nM to 1 microM. Latrunculin A ( 1, 500 nM) decreased the disaggregation and cell migration of PC-3M-CT+ spheroids by 3-fold. Carbamates 4 and 5 were 2.5- and 5-fold more active than 1, respectively, in this assay with less actin binding affinity. Latrunculin A ( 1, IC 50 6.7 microM) and its 17- O-[ N-(benzyl)carbamate ( 6, IC 50 29 microM) suppress hypoxia-induced HIF-1 activation in T47D breast tumor cells.

Methylalpinumisoflavone Inhibits Hypoxia-inducible Factor-1 (HIF-1) Activation by Simultaneously Targeting Multiple Pathways

Hypoxia is a common feature of solid tumors, and the extent of tumor hypoxia correlates with advanced disease stages and treatment resistance. The transcription factor hypoxia-inducible factor-1 (HIF-1) represents an important tumor-selective molecular target for anticancer drug discovery directed at tumor hypoxia. A natural product chemistry-based approach was employed to discover small molecule inhibitors of HIF-1. Bioassay-guided isolation of an active lipid extract of the tropical legumaceous plant Lonchocarpus glabrescens and structure elucidation afforded two new HIF-1 inhibitors: alpinumisoflavone (compound 1) and 4′-O-methylalpinumisoflavone (compound 2). In human breast tumor T47D cells, compounds 1 and 2 inhibited hypoxia-induced HIF-1 activation with IC50 values of 5 and 0.6 μm, respectively. At the concentrations that in hibited HIF-1 activation, compound 2 inhibited hypoxic induction of HIF-1 target genes (CDKN1A, GLUT-1, and VEGF), tumor angiogenesis in vitro, cell migration, and chemotaxis. Compound 2 inhibits HIF-1 activation by blocking the induction of nuclear HIF-1α protein, the oxygen-regulated subunit that controls HIF-1 activity. Mechanistic studies indicate that, unlike rotenone and other mitochondrial inhibitors, compound 2 represents the first small molecule that inhibits HIF-1 activation by simultaneously suppressing mitochondrial respiration and disrupting protein translation in vitro. This unique mechanism distinguishes compound 2 from other small molecule HIF-1 inhibitors that are simple mitochondrial inhibitors or flavanoid-based protein kinase inhibitors.

Natural Product-Derived Small Molecule Activators of Hypoxia-Inducible Factor-1 (HIF-1)

Hypoxia-inducible factor-1 (HIF-1) is a key mediator of oxygen homeostasis that was first identified as a transcription factor that is induced and activated by decreased oxygen tension. Upon activation, HIF-1 upregulates the transcription of genes that promote adaptation and survival under hypoxic conditions. HIF-1 is a heterodimer composed of an oxygen-regulated subunit known as HIF-1alpha and a constitutively expressed HIF-1beta subunit. In general, the availability and activity of the HIF-1alpha subunit determines the activity of HIF-1. Subsequent studies have revealed that HIF-1 is also activated by environmental and physiological stimuli that range from iron chelators to hormones. Preclinical studies suggest that HIF-1 activation may be a valuable therapeutic approach to treat tissue ischemia and other ischemia/hypoxia-related disorders. The focus of this review is natural product-derived small molecule HIF-1 activators. Natural products, relatively low molecular weight organic compounds produced by plants, animals, and microbes, have been and continue to be a major source of new drugs and molecular probes. The majority of known natural product-derived HIF-1 activators were discovered through the pharmacological evaluation of specifically selected individual compounds. On the other hand, the combination of natural products chemistry with appropriate high-throughput screening bioassays may yield novel natural product-derived HIF-1 activators. Potent natural product-derived HIF-1 activators that exhibit a low level of toxicity and side effects hold promise as new treatment options for diseases such as myocardial and peripheral ischemia, and as chemopreventative agents that could be used to reduce the level of ischemia/reperfusion injury following heart attack and stroke.

Marine natural products as inhibitors of hypoxic signaling in tumors

Marine natural products have become a major source of new chemical entities in the discovery of potential anticancer agents that potently suppress various antitumor molecular targets. As a consequence of insufficient vascularization, hypoxic regions form within rapidly growing solid tumor masses. Specific alterations of gene expression in these hypoxic tumor cells help facilitate the survival and metastatic spread of solid tumors. The transcriptional response to cellular hypoxia is primarily mediated by the transcription factor hypoxia-inducible factor-1 (HIF-1) that regulates the expression of more than 100 genes involved in cellular adaptation and survival under hypoxic stress. Clinical studies in cancer patients indicate that HIF-1 activation is directly correlated with advanced disease stages and treatment resistance. HIF-1 has emerged as an important tumor-selective molecular target for anticancer drug discovery. As a result, natural product-based inhibitors of HIF-1 activation have been identified from plants and microorganisms. Recently, structurally unique natural products from marine sponges, crinoids, and algae have been identified as HIF-1 activation inhibitors. The US National Cancer Institute’s Open Repository of marine invertebrate and algae extracts has proven to be a valuable source of natural product HIF-1 inhibitors. Among the active compounds identified, certain marine natural products have also been shown to suppress the hypoxic induction of HIF-1 target genes such as vascular endothelial growth factor (VEGF). Some of these marine HIF-1 inhibitors act by interfering with the generation of mitochondrial signaling molecules in hypoxic cells. However, the precise mechanisms of action for many newly identified marine natural product HIF-1 inhibitors remain unresolved.

The Caulerpa pigment caulerpin inhibits HIF-1 activation and mitochondrial respiration.

The transcription factor hypoxia-inducible factor-1 (HIF-1) represents an important molecular target for anticancer drug discovery. In a T47D cell-based reporter assay, the Caulerpa spp. algal pigment caulerpin (1) inhibited hypoxia-induced as well as 1,10-phenanthroline-induced HIF-1 activation. The angiogenic factor vascular endothelial growth factor (VEGF) is regulated by HIF-1. Caulerpin (10 microM) suppressed hypoxic induction of secreted VEGF protein and the ability of hypoxic T47D cell-conditioned media to promote tumor angiogenesis in vitro. Under hypoxic conditions, 1 (10 microM) blocked the induction of HIF-1alpha protein, the oxygen-regulated subunit that controls HIF-1 activity. Reactive oxygen species produced by mitochondrial complex III are believed to act as a signal of cellular hypoxia that leads to HIF-1alpha protein induction and activation. Further mechanistic studies revealed that 1 inhibits mitochondrial respiration at electron transport chain (ETC) complex I (NADH-ubiquinone oxidoreductase). Under hypoxic conditions, it is proposed that 1 may disrupt mitochondrial ROS-regulated HIF-1 activation and HIF-1 downstream target gene expression by inhibiting the transport or delivery of electrons to complex III.

Inducers of hypoxic response: marine sesquiterpene quinones activate HIF-1.

The hypoxia-inducible factor-1 (HIF-1) transcription factor regulates cellular oxygen homeostasis. Agents that activate HIF-1 and downstream HIF targets represent potential drug leads for the prevention and/or treatment of ischemic disorders. In a search for small-molecule HIF-1 activators, 1936 marine invertebrate and algal extract samples (U.S. National Cancer Institutes Open Repository) were evaluated for HIF-1 activation activity in a cell-based reporter assay. Bioassay-guided fractionation of two active extracts of the sponge Dactylospongia elegans afforded four new sesquiterpene quinones (2-5), one new sesquiterpene phenol (6), the known Golgi disruptor ilimaquinone (1), and three previously reported ilimaquinone analogues (7-9). While antiproliferative activity was observed at higher concentrations, the sesquiterpene quinones (1-3) possessing a 2-hydroxy-5-methoxy-1,4-benzoquinone moiety activated HIF-1 and increased the expression of HIF-1 target gene vascular endothelial growth factor (VEGF) in T47D cells.

The Alternative Medicine Pawpaw and Its Acetogenin Constituents Suppress Tumor Angiogenesis via the HIF-1/VEGF Pathway

Products that contain twig extracts of pawpaw (Asimina triloba) are widely consumed anticancer alternative medicines. Pawpaw crude extract (CE) and purified acetogenins inhibited hypoxia-inducible factor-1 (HIF-1)-mediated hypoxic signaling pathways in tumor cells. In T47D cells, pawpaw CE and the acetogenins 10-hydroxyglaucanetin (1), annonacin (2), and annonacin A (3) inhibited hypoxia-induced HIF-1 activation with IC(50) values of 0.02 microg/mL, 12 nM, 13 nM, and 31 nM, respectively. This inhibition correlates with the suppression of the hypoxic induction of HIF-1 target genes VEGF and GLUT-1. The induction of secreted VEGF protein represents a key event in hypoxia-induced tumor angiogenesis. Both the extract and the purified acetogenins blocked the angiogenesis-stimulating activity of hypoxic T47D cells in vitro. Pawpaw extract and acetogenins inhibited HIF-1 activation by blocking the hypoxic induction of nuclear HIF-1alpha protein. The inhibition of HIF-1 activation was associated with the suppression of mitochondrial respiration at complex I. Thus, the inhibition of HIF-1 activation and hypoxic tumor angiogenesis constitutes a novel mechanism of action for these anticancer alternative medicines.