Journal: Journal of Virology

Article Title: Cellular DNA Topoisomerases Are Required for the Synthesis of Hepatitis B Virus Covalently Closed Circular DNA

doi: 10.1128/JVI.02230-18

Figure Lengend Snippet: Effects of topoisomerase poisons on cccDNA synthesis

Article Snippet: Topotecan (HY-13768A) and idarubicin (HY-17381) were purchased from MedChemExpress (MCE).

Techniques: Control

Journal: Journal of Virology

Article Title: Cellular DNA Topoisomerases Are Required for the Synthesis of Hepatitis B Virus Covalently Closed Circular DNA

doi: 10.1128/JVI.02230-18

Figure Lengend Snippet: TOP1 and TOP2 inhibitors block cccDNA synthesis. (A) Schematic presentation of the experimental schedule. HepAD38 cells were cultured in the absence of Tet, and 2 mM PFA was added to the culture medium 2 days after Tet removal to arrest viral DNA synthesis. Four days later, while PFA was withdrawn, Tet was added back to the culture medium to stop viral pgRNA transcription from the transgene. Cells were left untreated or treated with topotecan (TPT; 1 μM) or doxorubicin (Doxo; 1 μM) at 16 h after PFA removal and harvested at the indicated time points. (B) Hirt DNA after heat denaturalization at 88°C for 8 min and EcoRI digestion was resolved by agarose gel electrophoresis and HBV DNA species were detected by Southern blot hybridization with a riboprobe specifically hybridizing to negative-strand DNA. mtDNA served as a loading control. (C) The amounts of cccDNA were quantified by phosphorimager, normalized to the amount of mtDNA, and plotted as the percentage of that in the mock-treated (UT) cells harvested at 16 h post-PFA removal. Means and standard deviations (n = 4) are presented. *DP-rc, denatured deproteinized rc DNA; ccc*, EcoRI-linearized cccDNA.

Article Snippet: Topotecan (HY-13768A) and idarubicin (HY-17381) were purchased from MedChemExpress (MCE).

Techniques: Blocking Assay, Cell Culture, DNA Synthesis, Agarose Gel Electrophoresis, Southern Blot, Hybridization, Control

Journal: Journal of Virology

Article Title: Cellular DNA Topoisomerases Are Required for the Synthesis of Hepatitis B Virus Covalently Closed Circular DNA

doi: 10.1128/JVI.02230-18

Figure Lengend Snippet: TOP1 and TOP2 inhibitors inhibited HBV cccDNA synthesis in de novo infection. (A) Schematic representation of the experimental schedule. C3AhNTCP cells were infected with HBV at an MOI of 250 genome equivalents for 24 h. The cells were mock treated or treated with 200 nM doxorubicin (Doxo), 200 nM topotecan (TPT), 200 nM doxorubicin and 200 nM topotecan (Doxo + TPT), 200 nM aclarubicin (Acla), or 1 μg/ml myrcludex B (Myr-B) starting from HBV infection for a total of 36 h. (B) Hirt DNA was resolved by agarose gel electrophoresis after heat denaturalization at 88°C for 8 min and EcoRI digestion. HBV DNA species were detected by Southern blot hybridization with a riboprobe specifically hybridizing to negative-strand DNA. mtDNA served as a loading control of Hirt DNA analysis. (C) cccDNAs were quantified by a phosphorimager. The data from three independent experiments are presented. P values calculated by Student's t test are presented.

Article Snippet: Topotecan (HY-13768A) and idarubicin (HY-17381) were purchased from MedChemExpress (MCE).

Techniques: Infection, Agarose Gel Electrophoresis, Southern Blot, Hybridization, Control

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: IC50 Values for small molecule inhibitors and topotecan in neuroblastoma cell lines

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques:

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: A. Shows a representative bar graph for the combination effects of inhibitors and topotecan on neuroblastoma cells growth treated with fixed one concentration of each inhibitor at sub-IC50 concentration for 72 hours. For combination treatments, SH-SY-5Y cells were treated with 13-197, BI2536, vismodegib and topotecan at 5 μM, 5 nM, 50 μM, and 5 nM concentrations, respectively; IMR-32 cells were treated with 13-197, BI2536, vismodegib and topotecan at 5 μM, 5 nM, 50 μM, and 10 nM concentrations, respectively; and SK-N-BE(2) were treated with 13-197, BI2536, vismodegib and topotecan at 5 μM, 5 nM, 50 μM and 50 nM concentrations, respectively. Following treatment, the cells were subjected to growth analyses using MTT assay. B-D. MTT assay showing the combination effects of indicated small molecule inhibitors and topotecan on neuroblastoma cells growth at 72 hours in a dose-dependent manner. The values represent the means ± SD from four wells of 96-well plates. *, p<0.05.

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques: Concentration Assay, MTT Assay

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: Combination indexes of small molecule inhibitors and topotecan in neuroblastoma cell lines

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques:

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: A. Illustrates a representative scatter diagram for the apoptotic cell analyses following treatment as indicated above in Figure with small molecule inhibitors alone or in combination with topotecan in neuroblastoma cells. B. Quantification of the apoptotic cells (% Annexin-V/PI double positive) following small molecule inhibitors alone or in combination with topotecan treatment in SH-SY-5Y, IMR-32 and SK-N-BE(2) neuroblastoma cells. The values represent the means ± SD of three separate experiments. *, p<0.05.

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques:

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: Each neuroblastoma cell line was treated with inhibitors alone or combination with topotecan (as described in Figure ) for 24 hours and the expression of associated pathways/molecules were determined using western blot analyses. β-Actin was used as a loading control in these experiments.

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques: Expressing, Western Blot, Control

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: A. Shows a representative micrograph of small, medium and large neurospheres in 13-197 (5 μM), BI2536 (5 nM) and vismodegib (50 μM) alone or in combination with topotecan (10 nM) treated IMR-32cells. The micrographs were taken using a phase contrast microscope at 4X magnification. Scale bar; 100 μm. B. Quantification of sphere assay data in above described treated cells. The measurement of 50 to <100 μm size for small, >100 to 200 μm for medium and >200 μm size for the large neurosphere were considered. The values represent the means ± SD from three wells of 6-well plates. *, p<0.01. C. Western blot analyses for the Nestin and CD133 expression following the indicated treatments of neurospheres. β-Actin was used as a loading control in this experiment.

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques: Microscopy, Western Blot, Expressing, Control

Journal: Oncotarget

Article Title: Improved therapy for neuroblastoma using a combination approach: superior efficacy with vismodegib and topotecan

doi: 10.18632/oncotarget.7714

Figure Lengend Snippet: NSG mice bearing tumors were treated with vehicle (DMSO) or vismodegib (50 mg/kg) and topotecan (10 mg/kg) alone, or both, twice a week for four weeks. For the combination, mice were treated with each agent first (denoted as Pre-) and then next day with the other agent. A. Tumor growth analyses following treatments. B. Kaplan–Meier analyses for the survival of mice using the log-rank test. C. Histological (H&E) and immunohistochemical (Smo, Ki-67, MYCN and cleaved caspase3) analyses of tumors in mice treated as indicated. The images were scanned and captured using a digital scanner VENTANA Image software (Roche, Germany) at ×40 magnification.

Article Snippet: Hedgehog inhibitor vismodegib was purchased from LC laboratories (Woburn, MA) and PLK1 inhibitor BI2536 and chemotherapy topotecan were purchased from Selleckchem Company (Houston TX).

Techniques: Immunohistochemical staining, Software

Journal: Molecular Cancer Therapeutics

Article Title: Novel missense mutation of the DNA topoisomerase I gene in SN-38-resistant DLD-1 cells

doi: 10.1158/1535-7163.mct-05-0246

Figure Lengend Snippet: Figure 1. Cytotoxicity of camptothecin derivatives. Parental DLD-1 cells and resistant DLDSNR6 cells were cultured for 72 h in the presence of the indicated SN-38, camptothecin, and topotecan concentrations. Cell viability was determined by the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethony- phenol)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt assay. Bars, SD for three independent experiments.

Article Snippet: Drugs and Antibodies SN-38 was kindly provided by Yakult Co. Ltd. (Tokyo, Japan), topotecan was purchased from LKT Laboratories (St. Paul, MN), and other drugs were purchased from Sigma (St. Louis, MO).

Techniques: Cell Culture

Journal: bioRxiv

Article Title: Frontotemporal dementia patient-derived iPSC neurons show cell pathological hallmarks and evidence for synaptic dysfunction and DNA damage

doi: 10.1101/2024.04.12.589061

Figure Lengend Snippet: NUPR2, potentially related to the DNA damage pathway, is the only differentially expressed gene when comparing the C9-HRE and sporadic FTD neurons ( A ). Both sporadic and C9-HRE-carrying FTD neurons display nuclei, which are significantly rounder in shape as well as significantly smaller compared to healthy neuron nuclei, indicated by altered nuclear eccentricity ( B ), and a higher number of micronuclei ( C ). A representative image of a micronucleus next to the nucleus is shown (arrow). Kruskal-Wallis test followed by Dunn’s multiple comparisons test was used. n [Control] = 74; n [C9-] = 17; n [C9+] = 21. In addition, the distribution of larger and smaller sized nuclei is different in FTD neurons than that in control neurons ( D-F ). Topotecan treatment significantly increases the number of γH2A.X-positive foci in the nuclei in all neurons, indicating increased DNA damage. At baseline, C9+ neurons display a slightly higher number of γH2A.X foci compared to HC neurons, although this difference is not statistically significant ( H-J ). Kruskal-Wallis test followed by Dunn’s multiple comparisons test was used. n [Control] = 26-28; n [C9-] = 15-18; n [C9+] = 44-64. Statistically significant differences are shown as *p < 0.05, **p <0.01, and ***p <0.001.

Article Snippet: To study the DNA damage response, iPSC-derived neurons were treated with 10 μl topotecan (Topotecan hydrochloride, Tocris, 4562) or DMSO vehicle for 1h.

Techniques: Control

Journal: Drug Delivery

Article Title: Inhalation delivery dramatically improves the efficacy of topotecan for the treatment of local and distant lung cancer

doi: 10.1080/10717544.2021.1912209

Figure Lengend Snippet: Efficacy of inhaled vs. IV topotecan against orthotopic lung tumors in rats. (A) Gross and histological pictures of lungs from an age-matched normal (left) and H358-derived lung tumors from an untreated control (right) animals. (B) The tumor burden in the three treatment groups.

Article Snippet: The spray-dried powder formulation of topotecan was manufactured from (S)-topotecan (Toronto Research Chemicals, Inc., Toronto, Canada) and its physical, chemical, and aerosol characteristics were determined as described (Kuehl et al., ).

Techniques: Derivative Assay

Journal: Drug Delivery

Article Title: Inhalation delivery dramatically improves the efficacy of topotecan for the treatment of local and distant lung cancer

doi: 10.1080/10717544.2021.1912209

Figure Lengend Snippet: Inhalation delivery of topotecan is more effective in treating H358-derived orthotopic lung tumors compared to two times higher IV dose.

Article Snippet: The spray-dried powder formulation of topotecan was manufactured from (S)-topotecan (Toronto Research Chemicals, Inc., Toronto, Canada) and its physical, chemical, and aerosol characteristics were determined as described (Kuehl et al., ).

Techniques:

Journal: Drug Delivery

Article Title: Inhalation delivery dramatically improves the efficacy of topotecan for the treatment of local and distant lung cancer

doi: 10.1080/10717544.2021.1912209

Figure Lengend Snippet: Pharmacokinetic analysis of topotecan following IV or inhalation delivery in mice. The mean levels of topotecan (ng/mL) detected in the (A) plasma, (B) lung, (C) liver, and (D) brain tissues of mice at various time points following 5 mg/kg IV or 1 mg/kg inhalation delivery of the drug.

Article Snippet: The spray-dried powder formulation of topotecan was manufactured from (S)-topotecan (Toronto Research Chemicals, Inc., Toronto, Canada) and its physical, chemical, and aerosol characteristics were determined as described (Kuehl et al., ).

Techniques:

Journal: Drug Delivery

Article Title: Inhalation delivery dramatically improves the efficacy of topotecan for the treatment of local and distant lung cancer

doi: 10.1080/10717544.2021.1912209

Figure Lengend Snippet: Pharmacokinetics profile of topotecan following inhalation or IV delivery.

Article Snippet: The spray-dried powder formulation of topotecan was manufactured from (S)-topotecan (Toronto Research Chemicals, Inc., Toronto, Canada) and its physical, chemical, and aerosol characteristics were determined as described (Kuehl et al., ).

Techniques:

Journal: Drug Delivery

Article Title: Inhalation delivery dramatically improves the efficacy of topotecan for the treatment of local and distant lung cancer

doi: 10.1080/10717544.2021.1912209

Figure Lengend Snippet: Inhalation delivery of topotecan leads to superior efficacy against lung tumors at distant sites than IV delivery. The growth of tumors derived from (A) the KRAS mutant A549, (B) the KRAS and EGFR wildtype H358, and (C) the EGFR mutant H1975 human NSCLC cell lines revealed a superior efficacy of 1 mg/kg inhaled topotecan compared to 5 mg/kg IV topotecan. (D, E) The average weights of these tumors obtained at the end of the study further confirmed the tumor measurement data. Significant differences ( p <.05) from vehicle control and IV topotecan treated groups are shown as * and ϕ, respectively.

Article Snippet: The spray-dried powder formulation of topotecan was manufactured from (S)-topotecan (Toronto Research Chemicals, Inc., Toronto, Canada) and its physical, chemical, and aerosol characteristics were determined as described (Kuehl et al., ).

Techniques: Derivative Assay, Mutagenesis