H-NMR spectra ( and ) and NOE difference spectra ( and ) of compounds and

<p><b>Copyright information:</b></p><p>Taken from "Synthesis and characterization of oligonucleotides containing 2′-fluorinated thymidine glycol as inhibitors of the endonuclease III reaction"</p><p>Nucleic Acids Research 2006;34(5):1540-1551.</p><p>Published online 17 Mar 2006</p><p>PMCID:PMC1409675.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> ( and ) the major isomer; ( and ) the minor isomer. The H6 resonance was saturated in the NOE experiments

Competition of the 13 bp substrate duplexes and the Tg-containing duplexes in the Endo III reaction

<p><b>Copyright information:</b></p><p>Taken from "Synthesis and characterization of oligonucleotides containing 2′-fluorinated thymidine glycol as inhibitors of the endonuclease III reaction"</p><p>Nucleic Acids Research 2006;34(5):1540-1551.</p><p>Published online 17 Mar 2006</p><p>PMCID:PMC1409675.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> The P-labeled substrates without fluorine were incubated with Endo III in the presence of the competitors containing Tg. The amounts of the nicked products (standardized to those without the competitors) were plotted against the concentrations of the competitor. Open circles, P-5-Tg + 5-Tg; filled circles, P-5-Tg + 5-Tg; open triangles, P-5-Tg + 5-Tg; filled triangles, P-5-Tg 5-Tg

Formation of the covalent complex between T4 Endo V and a CPD-containing oligonucleotide duplex fluorinated at the 2′ position

<p><b>Copyright information:</b></p><p>Taken from "Synthesis and characterization of oligonucleotides containing 2′-fluorinated thymidine glycol as inhibitors of the endonuclease III reaction"</p><p>Nucleic Acids Research 2006;34(5):1540-1551.</p><p>Published online 17 Mar 2006</p><p>PMCID:PMC1409675.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> The 5′ and 3′ ends of the CPD-containing strand were P-labeled. After hybridization with the complementary strand, the duplexes were incubated with T4 Endo V at 30°C for 30 min, and the mixtures were subjected to 15% SDS–PAGE

Analysis of the 5-Tg () and 5-Tg () 13mers by MALDI-TOF mass spectrometry

<p><b>Copyright information:</b></p><p>Taken from "Synthesis and characterization of oligonucleotides containing 2′-fluorinated thymidine glycol as inhibitors of the endonuclease III reaction"</p><p>Nucleic Acids Research 2006;34(5):1540-1551.</p><p>Published online 17 Mar 2006</p><p>PMCID:PMC1409675.</p><p>© The Author 2006. Published by Oxford University Press. All rights reserved</p> The calculated molecular weight is 3977.69

PAGE analysis of BER activities of AlkA, hMPG and HeLa CFE1s for Oxa and Oxa–Sp

<p><b>Copyright information:</b></p><p>Taken from "Repair activity of base and nucleotide excision repair enzymes for guanine lesions induced by nitrosative stress"</p><p>Nucleic Acids Research 2005;33(7):2181-2191.</p><p>Published online 14 Apr 2005</p><p>PMCID:PMC1079971.</p><p>© The Author 2005. Published by Oxford University Press. All rights reserved</p> () 25OXA, 25OXA–SP and 34MG (all 2 nM, and ) were incubated with the indicated amounts of AlkA and hMPG at 37°C for 30 min. After incubation, the reaction mixture was treated with 0.1 M NaOH to cleave AP sites, and products were separated by 16% denaturing PAGE. The nicked products due to β-elimination (upper bands) and β,δ-elimination (lower bands) are indicated by open brackets. () 25OXA and 25HX (both 2 nM, and ) were incubated in hMPG buffer with the indicated amounts of HeLa CFE1s at 37°C for 1 h. Products were analyzed as described above

Selective cytotoxicity of the anti-diabetic drug, metformin, in glucose-deprived chicken DT40 cells

<div><p>Metformin is a biguanide drug that is widely used in the treatment of diabetes. Epidemiological studies have indicated that metformin exhibits anti-cancer activity. However, the molecular mechanisms underlying this activity currently remain unclear. We hypothesized that metformin is cytotoxic in a tumor-specific environment such as glucose deprivation and/or low oxygen (O₂) tension. We herein demonstrated that metformin was highly cytotoxic under glucose-depleted, but not hypoxic (2% O₂) conditions. In order to elucidate the underlying mechanisms of this selective cytotoxicity, we treated exposed DNA repair-deficient chicken DT40 cells with metformin under glucose-depleted conditions and measured cellular sensitivity. Under glucose-depleted conditions, metformin specifically killed <i>fancc</i> and <i>fancl</i> cells that were deficient in FANCC and FANCL proteins, respectively, which are involved in DNA interstrand cross-link repair. An analysis of chromosomal aberrations in mitotic chromosome spreads revealed that a clinically relevant concentration of metformin induced DNA double-strand breaks (DSBs) in <i>fancc</i> and <i>fancl</i> cells under glucose-depleted conditions. In summary, metformin induced DNA damage under glucose-depleted conditions and selectively killed cells. This metformin-mediated selective toxicity may suppress the growth of malignant tumors that are intrinsically deprived of glucose.</p></div

FA pathway-related proteins involved in removing DNA lesions induced by metformin.

<p>(A) Histograms of the IC₅₀ values of metformin in wild-type cells and cell lines deficient in various FANC-related proteins. Cells were treated with metformin under glucose-depleted conditions for 24 h and colonies formed on complete media. All data represent IC₅₀ values ± 95% confidence intervals; (B) The toxicity of metformin to cells deficient in the FANCC or FANCL protein and deficient cell lines stably expressing the indicated transgenes. Data represent the mean ± S.D.; (C) The toxicity of metformin to cells deficient in the TDP1 or PARP1 protein and cells simultaneously deficient in both TDP1 and PARP1 proteins. Data represent the means ±S.D.</p

Toxicity of metformin and comparison of cellular sensitivities to metformin among various DNA repair-deficient DT40 cell lines under glucose-depleted conditions.

<p>(A) Wild-type cells were treated with the indicated doses of metformin for 24 h in complete medium, no glucose medium, or the 2% O₂ hypoxic condition with complete medium, and colonies formed on methylcellulose-containing complete media under normal conditions for 7 days. All data represent the mean ± S.D. normalized to cells not treated with metformin from three independent experiments. In each experiment, relative viabilities were measured as N/N₀ where N is the mean number of colonies at each dose of metformin in treated cells and N₀ is the mean number of colonies in untreated controls; (B) Histograms of the IC₅₀ values of metformin in the wild-type and various DNA repair-deficient cell lines. Cells were treated with metformin under glucose-depleted conditions for 24 h and colonies formed on complete media. All data represent IC₅₀ values ± 95% confidence intervals normalized to cells not treated with metformin from three independent experiments. In each experiment, relative viabilities were measured as N/N₀ where N is the mean number of colonies at each dose in metformin-treated cells and N₀ is the mean number of colonies in untreated controls. Abbreviations: Wt, wild type; NER, nucleotide excision repair; BER, base excision repair; Topo-DNA, repair of DNA-topoisomerase (Topo) crosslinks; TLS, translation DNA synthesis; NHEJ, non-homologous end joining; HR, homologous recombination repair; FA, FA pathway (ICL repair).</p

Induction of chromosomal breakages by metformin under glucose-depleted conditions.

<p>(A) Wild-type cells, cell lines deficient in FANCC or FANCL, and reconstituted cells were incubated with or without 13 μM metformin for 24 h under glucose-depleted conditions; (B) Wild-type cells and cell lines deficient in FANCC or FANCL were incubated with the indicated doses of metformin for 24 h in glucose-depleted or -containing medium. We analyzed 50 metaphase nuclei, and quantified the number of chromosomal aberrations per cell (Y-axis). Data represent the mean ± S.E. Asterisks (*) indicate <i>p</i> < 0.05 by a multiple comparison one-way ANOVA (Tukey’s test). N.S.: not significant (<i>p</i> ≥ 0.05).</p