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The dramatic increase in the prevalence of antibiotic-resistant bacteria has necessitated a search for new antibacterial agents against novel targets. Moiramide B is a natural product, broad-spectrum antibiotic that inhibits the carboxyltransferase component of acetyl-CoA carboxylase, which catalyzes the first committed step in fatty acid synthesis. Herein, we report the 2.6 Å resolution crystal structure of moiramide B bound to carboxyltransferase. An unanticipated but significant finding was that moiramide B bound as the enol/enolate. Crystallographic studies demonstrate that the (4S)-methyl succinimide moiety interacts with the oxyanion holes of the enzyme, supporting the notion that an anionic enolate is the active form of the antibacterial agent. Structure–activity studies demonstrate that the unsaturated fatty acid tail of moiramide B is needed only for entry into the bacterial cell. These results will allow the design of new antibacterial agents against the bacterial form of carboxyltransferase.

Pteris vittata exhibits enhanced arsenic uptake, but the corresponding mechanisms are not well known. The prevalent form of arsenic in most soils is arsenate, which is a phosphate analog and a substrate for Phosphate transporter 1 (Pht1) transporters. Herein we identify and characterize three P. vittata Pht1 transporters. Pteris vittata Pht1 cDNAs were isolated and characterized via heterologous expression in Saccharomyces cerevisiae (yeast) and Nicotiana benthamiana leaves. Expression of the PvPht1 loci in P. vittata gametophytes was also examined in response to phosphate deficiency and arsenate exposure. Expression of each of the PvPht1 cDNAs complemented the phosphate uptake defect of a yeast mutant. Compared with yeast cells expressing Arabidopsis thaliana Pht1;5, cells expressing PvPht1;3 were more sensitive to arsenate, and accumulated more arsenic. Uptake assays with yeast cells and radiolabeled (32)P revealed that PvPht1;3 and AtPht1;5 have similar affinities for phosphate, but the affinity of PvPht1;3 for arsenate is much greater. In P. vittata gametophytes, PvPht1;3 transcript levels increased in response to phosphate (Pi) deficiency and arsenate exposure. PvPht1;3 is induced by Pi deficiency and arsenate, and encodes a phosphate transporter that has a high affinity for arsenate. PvPht1;3 probably contributes to the enhanced arsenate uptake capacity and affinity exhibited by P. vittata.

Many cancer treatments, such as those for managing recalcitrant tumors like pancreatic ductal adenocarcinoma, cause off-target toxicities in normal, healthy tissue, highlighting the need for more tumor-selective chemotherapies. β-Lapachone is bioactivated by NAD(P)H:quinone oxidoreductase 1 (NQO1). This enzyme exhibits elevated expression in most solid cancers and therefore is a potential cancer-specific target. β-Lapachone's therapeutic efficacy partially stems from the drug's induction of a futile NQO1-mediated redox cycle that causes high levels of superoxide and then peroxide formation, which damages DNA and causes hyperactivation of poly(ADP-ribose) polymerase, resulting in extensive NAD+/ATP depletion. However, the effects of this drug on energy metabolism due to NAD+ depletion were never described. The futile redox cycle rapidly consumes O2, rendering standard assays of Krebs cycle turnover unusable. In this study, a multimodal analysis, including metabolic imaging using hyperpolarized pyruvate, points to reduced oxidative flux due to NAD+ depletion after β-lapachone treatment of NQO1+ human pancreatic cancer cells. NAD+-sensitive pathways, such as glycolysis, flux through lactate dehydrogenase, and the citric acid cycle (as inferred by flux through pyruvate dehydrogenase), were down-regulated by β-lapachone treatment. Changes in flux through these pathways should generate biomarkers useful for in vivo dose responses of β-lapachone treatment in humans, avoiding toxic side effects. Targeting the enzymes in these pathways for therapeutic treatment may have the potential to synergize with β-lapachone treatment, creating unique NQO1-selective combinatorial therapies for specific cancers. These findings warrant future studies of intermediary metabolism in patients treated with β-lapachone.

Selenium-containing quinone-based 1,2,3-triazoles were synthesized using click chemistry, the copper catalyzed azide-alkyne 1,3-dipolar cycloaddition, and evaluated against six types of cancer cell lines: HL-60 (human promyelocytic leukemia cells), HCT-116 (human colon carcinoma cells), PC3 (human prostate cells), SF295 (human glioblastoma cells), MDA-MB-435 (melanoma cells) and OVCAR-8 (human ovarian carcinoma cells). Some compounds showed IC50 values < 0.3 mu M. The cytotoxic potential of the quinones evaluated was also assayed using non-tumor cells, exemplified by peripheral blood mononuclear (PBMC), V79 and L929 cells. Mechanistic role for NAD(P)H:Quinone Oxidoreductase 1 (NQO1) was also elucidated. These compounds could provide promising new lead derivatives for more potent anticancer drug development and delivery, and represent one of the most active classes of lapachones reported. (C) 2016 Elsevier Masson SAS. All rights reserved.

Inhibitors of cancer-specific pathways can selectively kill off tumor cells. However, heterogeneity of neoplastic tissue often allows other cancer cells to repopulate the tissue area, leading to regeneration of resistant disease. β-Lapachone is a novel bioactivatable drug that relies specifically on tumor-directed upregulated levels of NAD(P)H:quinone oxidoreductase 1 (NQO1) to kill most solid cancers, such as 90 % of pancreatic and non-small cell lung, 60 % of breast, colon, and prostate, as well as 50 % of head and neck cancers. Once β-lapachone is bioactivated by the NQO1 enzyme, massive levels of hydrogen peroxide are produced that, in turn, damage the DNA of cancer cells, while associated normal tissues, which lack NQO1, are protected by high levels of catalase. If tumors are irradiated prior to applying β-lapachone, the drug (clinical form, ARQ761) can work in combination with the vast spectrum of DNA lesions created by ionizing radiation, particularly DNA base lesions, single and double strand breaks (SSBs and DSBs), in addition to the massive hydrogen peroxide-based lesions created by β-lapachone, to cause tumor-dependent poly(ADP-ribose) polymerase 1 (PARP1) hyperactivation. Once tumor-selective PARP hyperactivation is induced in cancer cells, they die due to low concomitant catalase levels. In contrast, associated normal tissue, as well as other normal tissue, lack elevated levels of NQO1 and have high catalase levels. Cancer cell death ultimately occurs by NAD+-depletion, where resistance to NQO1 bioactivatable drugs has not been noted to date. Current studies are focused on pancreatic and non-small cell lung cancers, as NQO1 is elevated in nearly all of these cancers.

There is an urgent demand for the development of new antibiotics due to the increase in drug-resistant pathogenic bacteria. A novel target is the multifunctional enzyme acetyl-CoA carboxylase (ACC), which catalyzes the first committed step in fatty acid synthesis and consists of two enzymes: biotin carboxylase and carboxyltransferase. Covalently attaching known inhibitors against these enzymes with saturated hydrocarbon linkers of different lengths generated dual-ligand inhibitors. Kinetic results revealed that the dual-ligands inhibited the ACC complex in the nanomolar range. Microbiology assays showed that the dual-ligand with a 15-carbon linker did not exhibit any antibacterial activity, while the dual-ligand with a 7-carbon linker displayed broad-spectrum antibacterial activity as well as a decreased susceptibility in the development of bacterial resistance. These results suggest that the properties of the linker are vital for antibacterial activity and show how inhibiting two different enzymes with the same compound increases the overall potency while also impeding the development of resistance.