Polymorphisms in Glutathione S-Transferase (GST) Genes Modify the Effect of Exposure to Maternal Smoking Metabolites in Pregnancy and Offspring DNA Methylation.

Maternal smoking in pregnancy (MSP) affects the offspring's DNA methylation (DNAm). There is a lack of knowledge regarding individual differences in susceptibility to exposure to MSP. Glutathione S-transferase (GST) genes are involved in protection against harmful oxidants such as those found in cigarette smoke. This study aimed to test whether polymorphisms in GST genes influence the effect of MSP on offspring DNAm. Using data from the Isle of Wight birth cohort, we assessed the association of MSP and offspring DNAm in 493 mother-child dyads (251 male, 242 female) with the effect-modifying role of GST gene polymorphism (at rs506008, rs574344, rs12736389, rs3768490, rs1537234, and rs1695). MSP was assessed by levels of nicotine and its downstream metabolites (cotinine, norcotinine, and hydroxycotinine) in maternal sera. In males, associations of hydroxycotinine with DNAm at cg18473733, cg25949550, cg11647108, and cg01952185 and norcotinine with DNAm at cg09935388 were modified by GST gene polymorphisms (p-values < 0.05). In females, associations of hydroxycotinine with DNAm at cg12160087 and norcotinine with DNAm at cg18473733 were modified by GST gene polymorphisms (p-values < 0.05). Our study emphasizes the role of genetic polymorphism in GST genes in DNAm's susceptibility to MSP.

Association of prenatal acetaminophen use and acetaminophen metabolites with DNA methylation of newborns: analysis of two consecutive generations of the Isle of Wight birth cohort.

Acetaminophen is used by nearly two-thirds of pregnant women. Although considered safe, studies have demonstrated associations between prenatal acetaminophen use and adverse health outcomes in offspring. Since DNA methylation (DNAm) at birth may act as an early indicator of later health, assessments on whether DNAm of newborns is associated with gestational acetaminophen use or its metabolites are needed. Using data from three consecutive generations of the Isle of Wight cohort (F0-grandmothers, F1-mothers, and F2-offspring) we investigated associations between acetaminophen metabolites in F0 serum at delivery with epigenome-wide DNAm in F1 (Guthrie cards) and between acetaminophen use of F1 and F2-cord-serum levels with F2 cord blood DNAm. In epigenome-wide screening, we eliminated non-informative DNAm sites followed by linear regression of informative sites. Based on repeated pregnancies, indication bias analyses tested whether acetaminophen indicated maternal diseases or has a risk in its own right. Considering that individuals with similar intake process acetaminophen differently, metabolites were clustered to distinguish metabolic exposures. Finally, metabolite clusters from F1-maternal and F2-cord sera were tested for their associations with newborn DNAm (F1 and F2). Twenty-one differential DNAm sites in cord blood were associated with reported maternal acetaminophen intake in the F2 generation. For 11 of these cytosine-phosphate-guanine (CpG) sites, an indication bias was excluded and five were replicated in F2 with metabolite clusters. In addition, metabolite clusters showed associations with 25 CpGs in the F0-F1 discovery analysis, of which five CpGs were replicated in the F2-generation. Our results suggest that prenatal acetaminophen use, measured as metabolites, may influence DNAm in newborns.

Nicotine and Its Downstream Metabolites in Maternal and Cord Sera: Biomarkers of Prenatal Smoking Exposure Associated with Offspring DNA Methylation.

Nicotine is a major constituent of cigarette smoke. Its primary metabolite in maternal and cord sera, cotinine, is considered a biomarker of prenatal smoking. Nicotine and cotinine half-lives are decreased in pregnancy due to their increased rate of metabolism and conversion to downstream metabolites such as norcotinine and 3-hydroxycotinine. Hence, downstream metabolites of nicotine may provide informative biomarkers of prenatal smoking. In this study of three generations (F0-mothers, F1-offspring who became mothers, and F2-offspring), we present a biochemical assessment of prenatal smoking exposure based on maternal and cord sera levels of nicotine, cotinine, norcotinine, and 3-hydroxycotinine. As potential markers of early effects of prenatal smoking, associations with differential DNA methylation (DNAm) in the F1- and F2-offspring were assessed. All metabolites in maternal and cord sera were associated with self-reported prenatal smoking, except for nicotine. We compared maternal self-report of smoking in pregnancy to biochemical evidence of prenatal smoking exposure. Self-report of F0-mothers of F1 in 1989-1990 had more accuracy identifying prenatal smoking related to maternal metabolites in maternal serum (sensitivity = 94.6%, specificity = 86.9%) compared to self-reports of F1-mothers of F2 (2010-2016) associated with cord serum markers (sensitivity = 66.7%, specificity = 78.8%). Nicotine levels in sera showed no significant association with any DNAm site previously linked to maternal smoking. Its downstream metabolites, however, were associated with DNAm sites located on the MYO1G, AHRR, and GFI1 genes. In conclusion, cotinine, norcotinine, and 3-hydroxycotinine in maternal and cord sera provide informative biomarkers and should be considered when assessing prenatal smoking. The observed association of offspring DNAm with metabolites, except for nicotine, may imply that the toxic effects of prenatal nicotine exposure are exerted by downstream metabolites, rather than nicotine. If differential DNA methylation on the MYO1G, AHRR, and GFI1 genes transmit adverse effects of prenatal nicotine exposure to the child, there is a need to investigate whether preventing changes in DNA methylation by reducing the metabolic rate of nicotine and conversion to harmful metabolites may protect exposed children.

Semisynthesis of an Anticancer DPAGT1 Inhibitor from a Muraymycin Biosynthetic Intermediate.

We have explored a method to convert a muraymycin biosynthetic intermediate 3 to an anticancer drug lead 2 for in vivo and thorough preclinical studies. Cu(OAc)2 forms a stable complex with the amide 4 and prevents electrophilic reactions at the 2-((3-aminopropyl)amino)acetamide moiety. Under the present conditions, the desired 5″-primary amine was selectively protected with (Boc)2O to yield 6. The intermediate 6 was converted to 2 in two steps with 90% yield.

An antimycobacterial pleuromutilin analogue effective against dormant bacilli.

Pleuromutilin is a promising pharmacophore to design new antibacterial agents for Gram-positive bacteria. However, there are limited studies on the development of pleuromutilin analogues that inhibit growth of Mycobacterium tuberculosis (Mtb). In screening of our library of pleuromutilin derivatives, UT-800 (1) was identified to kill replicating- and non-replicating Mtb with the MIC values of 0.83 and 1.20 µg/mL, respectively. UT-800 also kills intracellular Mtb faster than rifampicin at 2× MIC concentrations. Pharmacokinetic studies indicate that 1 has an oral bioavailability with an average F-value of 27.6%. Pleuromutilin may have the potential to be developed into an orally administered anti-TB drug.

Synthesis of 3-(3-hydroxyphenyl)pyrrolidine dopamine D3 receptor ligands with extended functionality for probing the secondary binding pocket.

A series of 3-(3-hydroxyphenyl)pyrrolidine analogues which incorporate N-alkyl groups and N-butylamide-linked benzamide functionality have been synthesized and their in vitro binding affinities at human dopamine receptors have been evaluated. Our ligand design strategy was to take the 3-(3-hydroxyphenyl)pyrrolidine scaffold and extend functionality from the orthosteric binding site to the secondary binding pocket for enhancing affinity and selectivity for the D3 receptor. The N-alkyl analogues constitute a homologous series from N-pentyl to N-decyl to probe the length/bulk tolerance of the secondary binding pocket of the D3 receptor. Enantiomeric 3-(3-hydroxyphenyl)pyrrolidine analogues were also prepared in order to test the chirality preference of the orthosteric binding site for this scaffold. Benzamide analogues were prepared to enhance affinity and/or selectivity based upon the results of the homologous series.

Novel FR-900493 Analogues That Inhibit the Outgrowth of Clostridium difficile Spores.

The spectrum of antibacterial activity for the nucleoside antibiotic FR-900493 (1) can be extended by chemical modifications. We have generated a small focused library based on the structure of 1 and identified UT-17415 (9), UT-17455 (10), UT-17460 (11), and UT-17465 (12), which exhibit anti-Clostridium difficile growth inhibitory activity. These analogues also inhibit the outgrowth of C. difficile spores at 2× minimum inhibitory concentration. One of these analogues, 11, relative to 1 exhibits over 180-fold and 15-fold greater activity against the enzymes, phospho-MurNAc-pentapeptide translocase (MraY) and polyprenyl phosphate-GlcNAc-1-phosphate transferase (WecA), respectively. The phosphotransferase inhibitor 11 displays antimicrobial activity against several tested bacteria including Bacillus subtilis, Clostridium spp., and Mycobacterium smegmatis, but no growth inhibitory activity is observed against the other Gram-positive and Gram-negative bacteria. The selectivity index (Vero cell cytotoxicity/C. difficileantimicrobial activity) of 11 is approximately 17, and 11 does not induce hemolysis even at a 100 µM concentration.

A New Combination of a Pleuromutilin Derivative and Doxycycline for Treatment of Multidrug-Resistant Acinetobacter baumannii.

Multidrug-resistant (MDR) Acinetobacter baumannii is one of the most difficult Gram-negative bacteria to treat and eradicate. In a cell-based screening of pleuromutilin derivatives against a drug sensitive A. baumannii strain, new molecules (2-4) exhibit bacteriostatic activity with 3.13 µg/mL concentration and 1 shows bactericidal activity with an MBC of 6.25 µg/mL. The pleuromutilin derivative 1 displays strong synergistic effects with doxycycline in a wide range of concentrations. A 35/1 ratio of 1 and doxycycline (1-Dox 35/1) kills drug susceptible A. baumannii with the MBC of 2.0 µg/mL and an MDR A. baumannii with the MBC of 3.13 µg/mL. In vitro anti-Acinetobacter activity of 1-Dox 35/1 is superior to that of clinical drugs such as tobramycin, tigecycline, and colistin. The efficacy of 1-Dox 35/1 is evaluated in a mouse septicemia model; treatment of the infected C57BL/6 mice with 1-Dox 35/1 protects from lethal infection of A. baumannii with an ED50 value of <2.0 mg/kg.