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BISMUTH UPTAKE AND COMPARTMENTALIZATION IS AIDED BY GLUTATHIONE SEQUESTRATION IN A SELF-PROPELLED FEEDBACK PROCESS Bismuth (Bi) is a trivalent heavy metal that is used to treat gastrointestinal infection. Bismuth drugs display high selectivity for pathogenic cells but have low toxicity to human cells. While bismuth cellular uptake and disposal mechanisms are unknown, the Bi3+ ion has high affinity toward glutathione. Hongzhe Sun and his team studied the relationship between the two and quantitatively determined the metabolism of bismuth in human cells ((2015) Proc. Natl. Acad. Sci. U.S.A., 112, 3211−3216). The researchers found that in humans, bismuth uptake first follows a saturation pattern of passive transport. During this time, bismuth uses approximately half of the intracellular glutathione that is present. This phase is followed by exponential growth, during which glutathione levels increase over 4-fold from the original levels. The authors looked at how glutathione depletion affected bismuth metabolism and found that it dramatically reduced bismuth uptake and that bismuth-treated, glutathionedepleted cells exhibited disturbed morphology with lower survival rates. Furthermore, they discovered that glutathione is essential to bismuth subcellular compartmentalization, which protects cells from acute toxicity. The researchers found that after bismuth is passively absorbed by human cells, the vast majority of it is conjugated to cytosolic glutathione and then transported into perinuclear vesicles; in a self-sustaining positive feedback process, this glutathione sequestration activates de novo biosynthesis, which facilitates passive uptake of bismuth. Glutathione is absent from many Gram-positive bacteria which may explain bismuth drug selectivity. Abigail Druck Shudofsky
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AN IRIDIUM-CONTAINING NUCLEOSIDE CAUSES APOPTOSIS IN CANCEROUS CELLS AND PROVIDES INSIGHT INTO TRANSPORT
cells and rapidly induces apoptosis. Ir(III)−PPY can directly and noninvasively monitor and quantify the cellular activity of hENT1 in real time. Visualizing Ir(III)−PPY will allow its anticancer activity to be correlated with cellular uptake and localization and can help to identify patients who may respond favorably to therapeutic treatment with anticancer nucleoside analogues. Abigail Druck Shudofsky
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ENVIRONMENTAL STRESS CAUSES MUTAGENESIS OF TRINUCLEOTIDE REPEATS Human DNA contains microsatellites, stretches of repetitive short nucleotide sequences. These dynamic sequences mutate by expanding or contracting the number of repeats in the sequence, which can shift reading frames or alter gene expression. While DNA transactions stimulate trinucleotide repeat (TNR) mutagenesis, TNR stability is maintained by a chaperone protein that regulates double-strand break repair. As stress can impact DNA repair and increase mutagenesis in yeast and bacterial cells, John H. Wilson led a team to discover whether environmental stress can increase TNR mutagenesis in human cells ((2015) Proc. Natl. Acad. Sci. U.S.A., 112, 3764−3769). The authors exposed cells to heat, cold, hypoxia, and oxidative stress and saw that TNR mutations increased significantly several days postexposure. This same effect was not seen immediately after the stress was removed or when starvation was a stressor. Stressed human cells produce stress-response factors (SRFs), and the authors knocked down individual SRFs associated with heat, cold, hypoxia, and oxidative stress. Elimination of SRFs blocked stress-induced TNR mutagenesis, as did knockdown of a structure-specific endonuclease and a DNA helicase. This implied that stress-induced mutagenesis occurs in an SRF-dependent manner through a mechanism related to replication. The researchers found evidence that DNA rereplication, which occurs when a cell’s genome is replicated more than once per cell cycle, is involved in stress-induced TNR
Reprinted with permission from Choi et al. (2015) J. Biol. Chem., DOI: 10.1074/jbc.M114.620294. Copyright 2015 American Society for Biochemistry and Molecular Biology.
Cancerous cells hyperproliferate through uncontrolled DNA synthesis. Accordingly, many anticancer drugs target DNA synthesis, including antimetabolitic nucleoside analogues with sugar modifications that cause their corresponding deoxynucleoside triphosphates to act as chain terminators. To function effectively as therapeutics, nucleoside analogues must be taken up by cells and metabolized to nucleoside triphosphates. Most require an active transport system for efficient cellular uptake. It is important to accurately measure the activity of nucleoside transporters, as cellular levels can serve as positive predictive factors for individual response to anticancer analogues. Anthony J. Berdis led a team that developed a nucleoside analogue with a deoxyriboside attached to a bis-cyclometalated scaffold containing iridium ((2015) J. Biol. Chem., DOI: 10.1074/jbc.M114.620294). This nucleoside, designated Ir(III)− PPY, has a compact three-dimensional structure, is highly stable in aqueous cellular environments, and can emit visible light over a range of wavelengths. The researchers demonstrated that the nucleoside transporter hENT1 helps Ir(III)−PPY to enter cells in a matter of several hours, where it localizes to the nucleus and mitochondria in a time- and dose-dependent manner. While Ir(III)−PPY has negligible cytotoxic effects in noncancerous cell lines, it interferes with G2/M cell cycle progression in cancer © 2015 American Chemical Society
Published: May 18, 2015 829
DOI: 10.1021/acs.chemrestox.5b00151 Chem. Res. Toxicol. 2015, 28, 829−830
Chemical Research in Toxicology
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higher 2 h post UV exposure than immediately after exposure. The team demonstrated that the presence of melanin was critical for the induction of dark CPDs. The researchers identified several inhibitors of dark CPD production, including inhibitors of melanin-synthesis and nitric oxide (NO•) and scavengers of ROS and superoxide (O2•−). NOX (reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase) is an O2•− source that, along with the NO• generator iNOS, is expressed in melanocytes and is UVinducible. These two radicals produce peroxynitrite (ONOO−), which degrades melanin into fragments and electronically excites them to a short-lived high-energy triplet state, a process that continues for hours post UV exposure. The excited melanin derivatives transfer their energy to DNA, inducing formation of CPDs. Abigail Druck Shudofsky
mutagenesis. This work suggests a pathway for stress-induced TNR mutagenesis in human cells whereby environmental stress stimulates SRFs, which induce DNA rereplication. DNA rereplication then leads to genome instability and triggers TNR mutagenesis. Abigail Druck Shudofsky
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POLYMERIC NANOPARTICLES ARE EFFECTIVE GENE THERAPY VECTORS THAT CAN SUCCESSFULLY TARGET BRAIN TUMORS
Adapted from Mangraviti et al. (2015) ACS Nano, 9, 1236− 1249. Copyright 2015 American Chemical Society.
Glioblastomas are malignant brain tumors with no reliable cure. A potential treatment approach is gene therapy, which uses a vector to deliver genetic material to a targeted cellular location where it is expressed. Viral vectors are often used but may present safety concerns; nonviral vectors are less risky but less efficient. Betty Tyler and Jordan J. Green led a team to look at the efficacy of poly(β-amino ester)s (PBAEs) nanoparticles as vectors ((2015) ACS Nano, 9, 1236−1249). PBAE polymers contain primary, secondary, and tertiary amines and hydrolytically cleavable ester bonds. They can selectively transfect tumor tissue and degrade quickly under physiological conditions, minimizing potential cytotoxicity. The researchers synthesized a PBAE library and evaluated the nanoparticles for their ability to deliver herpes simplex virus thymidine kinase (HSVtk) DNA to brain tumor cells in vitro and in vivo. HSVtk phosphorylates the prodrug ganciclovir, which allows the cytotoxic nucleoside analogue to be incorporated into DNA, causing premature DNA chain termination and apoptosis. The lead polymer for optimum drug delivery was known as 447. In vitro, 447/HSVtk displayed a high transfection efficiency with no cytotoxicity and killed cancer cells completely in the presence ganciclovir. When 447/HSVtk was infused intratumorally in vivo, the nanoparticles penetrated cells throughout the tumor mass. Treated rats showed significantly improved survival with systemic ganciclovir administration. PBAE-based HSVtk gene delivery has a strong safety profile and may be an efficient and successful nanomedical glioblastoma treatment. Abigail Druck Shudofsky
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UV-INDUCED REACTIVE SPECIES COMBINE TO ELECTRONICALLY EXCITE MELANIN DERIVATIVES, CAUSING DNA PHOTOPRODUCTS Melanin protects skin cells from harmful sun damage. However, research performed by Douglas E. Brash and his team indicates that melanin also extends the DNA-damaging effects of ultraviolet (UV) radiation well past exposure time ((2015) Science, 20, 842−847). Most UV-induced melanoma mutations originate from cyclobutane pyrimidine dimers (CPDs), molecular lesions that connect neighboring pyrimidines and alter the structure of DNA. While CPDs are typically created immediately after a UV photon is absorbed, the researchers found that mouse melanocytes generated CPDs in nuclear DNA more than 3 h post UV exposure. These “dark CPDs” were also generated in human melanocytes. In vivo mouse studies showed that epidermal CPD levels were three times 830
DOI: 10.1021/acs.chemrestox.5b00151 Chem. Res. Toxicol. 2015, 28, 829−830
