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Item Targeting mitochondrial DNA polymerase gamma for selective inhibition of MLH1 deficient colon cancer growth(PUBLIC LIBRARY SCIENCE, 2022-01-01) Somuncu, Berna; Ekmekcioglu, Aysegul; Antmen, Fatma Merve; Ertuzun, Tugce; Deniz, Emre; Keskin, Nazli; Park, Joon; Yazici, Ilgu Ece; Simsek, Busra; Erman, Batu; Yin, Whitney; Erman, Burak; Muftuoglu, MeltemSynthetic lethality in DNA repair pathways is an important strategy for the selective treatment of cancer cells without harming healthy cells and developing cancer-specific drugs. The synthetic lethal interaction between the mismatch repair (MMR) protein, MutL homolog 1 (MLH1), and the mitochondrial base excision repair protein, DNA polymerasey (Poly) was used in this study for the selective treatment of MLH1 deficient cancers. Germline mutations in the MLH1 gene and aberrant MLH1 promoter methylation result in an increased risk of developing many cancers, including nonpolyposis colorectal and endometrial cancers. Because the inhibition of Poly in MLH1 deficient cancer cells provides the synthetic lethal selectivity, we conducted a comprehensive small molecule screening from various databases and chemical drug library molecules for novel Poly inhibitors that selectively kill MLH1 deficient cancer cells. We characterized these Poly inhibitor molecules in vitro and in vivo, and identified 3,3'-{[}(1,1'-Bipheny1)4',4'-diyl)bis(azo)]bis{[}4-amino-1-naphthalenesu lfonic acid] (congo redItem Non-muscle invasive bladder cancer tissues have increased base excision repair capacity(NATURE PORTFOLIO, 2020-01-01) Somuncu, Berna; Keskin, Selcuk; Antmen, Fatma Merve; Saglican, Yesim; Ekmekcioglu, Aysegul; Ertuzun, Tugce; Tuna, Mustafa Bilal; Obek, Can; Wilson, David M.; Ince, Umit; Kural, Ali Riza; Muftuoglu, MeltemThe molecular mechanisms underlying the development and progression of bladder cancer (BC) are complex and have not been fully elucidated. Alterations in base excision repair (BER) capacity, one of several DNA repair mechanisms assigned to preserving genome integrity, have been reported to influence cancer susceptibility, recurrence, and progression, as well as responses to chemotherapy and radiotherapy. We report herein that non-muscle invasive BC (NMIBC) tissues exhibit increased uracil incision, abasic endonuclease and gap-filling activities, as well as total BER capacity in comparison to normal bladder tissue from the same patient (p<0.05). No significant difference was detected in 8-oxoG incision activity between cancer and normal tissues. NMIBC tissues have elevated protein levels of uracil DNA glycosylase, 8-oxoguanine DNA glycosylase, AP endonuclease 1 and DNA polymerase beta protein. Moreover, the fold increase in total BER and the individual BER enzyme activities were greater in high-grade tissues than in low-grade NMIBC tissues. These findings suggest that enhanced BER activity may play a role in the etiology of NMIBC and that BER proteins could serve as biomarkers in disease prognosis, progression or response to genotoxic therapeutics, such as Bacillus Calmette-Guerin.Item Anti-proliferative effects of indomethacin, acemetacin and their tromethamine salts in HCT116 human colon cancer cells(ISTANBUL UNIV, FAC PHARMACY, 2021-01-01) Ustundag, Gokce Cihan; Somuncu, Berna; Muftuoglu, Meltem; Karali, NilgunBackground and Aims: Since 1980's, several preclinical studies have been published on the anti-colorectal cancer activity of the nonsteroidal anti-inflammatory drug indomethacin. The direct anti-proliferative effect of indomethacin seems to occur via a variety of reported COX-independent mechanisms. Acemetacin is a glycolic acid ester derivative of indomethacin and contrary to indomethacin, there is not much published research on anti-cancer effects of acemetacin. Herein, we compared the in vitro anti-proliferative properties of indomethacin, acemetacin, and their tromethamine salts in HCT116 colon cancer cells. Methods: The tromethamine salts of indomethacin and acemetacin were synthesized and the structures were established by microanalysis, IR, H-1-NMR, C-13-NMR (APT) and 2D-NMR (HSQC and HMBC) spectrometry. Cell proliferation assays were performed using xCELLigence real-time cell analysis system. Results: Indomethacin exhibited profound inhibitory effects with IC50 values at low micromolar ranges. Acemetacin exhibited far lower cytotoxic activity as compared to that of indomethacin. Surprisingly, indomethacin-tromethamine salt was 2-fold and 4.4-fold more potent than indomethacin at 48 and 72 h, respectively, while maintaining its activity at 24 h. The tromethamine salt of acemetacin was more potent than acemetacin at 24 h and 48 h post-treatment. Conclusion: The anti-proliferative effect of indomethacin in HCT116 cells was found to be at low micro-molar levels. The esterification of indomethacin with glycolic acid caused a strong decrease in anti-proliferative effect. The salt formation caused a positive effect on the anti-proliferative activity of indomethacin and indomethacin-tromethamine salt may be a promising candidate for additional in vivo studies.Item 5-Hydroxyuracil Incision Activity Varies According to the Histological Grade of Non-muscle-invasive Bladder Cancer(GALENOS YAYINCILIK, 2021-01-01) Keskin, Selcuk; Antmen, Fatma Merve; Somuncu, Berna; Saglican, Yesim; Doganca, Tunkut; Obek, Can; Ince, Umit; Kural, Ali Riza; Muftuoglu, MeltemObjective: High levels of endonuclease III-like 1 (NTHL1) DNA glycosylase, which plays a role in the first step of the base excision repair pathway, has been related to cancer initiation and progression. 5-hydroxyuracil (5-OHU) oxidative base damage is a substrate for NTHL1 and endonuclease VIII-like 1 enzyme 1 (NEIL1) DNA glycosylases. This study investigates the association of 5-OHU incision activity with the risk of disease progression in patients with non-muscle-invasive bladder cancer (NMIBC) regarding grade and stage. Materials and Methods: During transurethral resection of 17 NMIBC patients, the papillary tumour before monopolar resection and healthy bladder mucosal tissue from the same person were obtained using cold cup biopsy. Both the normal mucosa and NMIBC tumour were pathologically confirmed. The histological grade and stage were also determined. The 5-OHU incision activity of all tissues was measured using a radiolabelled 5-OHU modified base containing DNA substrate. Results: 5-OHU incision activity was significantly higher in all high-grade NMIBC tissue extracts compared with the corresponding normal tissues (p=0.001). However, we found no significant difference in 5-OHU incision activity in low-grade NMIBC tissues (p=0.89). There was also a significant increase in 5-OHU incision activity at the Ta/T1 stage compared with the corresponding normal tissue (p=0.001). Conclusion: The increase in 5-OHU incision activity according to the histological grade of NMIBC tissue indicates that this activity (mainly performed by NTHL1 and NEIL1 DNA glycosylases) might play a role in NMIBC prognosis. Thus, it could be used as a potential prognostic biomarker for NMIBC.Item Bacillus Calmette-Guerin Increases Base Excision Repair in Bladder Cancer Cells(GALENOS YAYINCILIK, 2021-01-01) Keskin, Selcuk; Somuncu, Berna; Muftuoglu, MeltemObjective: Most patients with non-muscle-invasive bladder cancer (NMIBC) do not respond to intravesical Bacillus Calmette-Guerin (BCG) immunotherapy and have high risk of NMIBC recurrence and progression. In addition to its therapeutic effect which increases the local immune response, BCG also exerts an anti-tumour effect by increasing oxidative stress, and producing reactive oxygen species and oxidative DNA damage in bladder cancer (BC) cells. The oxidative DNA damage is repaired by base excision repair (BER) mechanism. Thus, BER capacity of BC cells could be an important factor in response to BCG therapy. Effects of BCG on the activity of BER in BC transitional carcinoma cell line, T24 have been investigated. Materials and Methods: The uracil-initiated total BER and BER enzyme activities were measured in whole cell extracts with or without BCG treatment using a {[}gamma-32P] adenosine triphosphate-labelled 51-mer DNA substrates. Results: BCG treatment increased the activities of uracil-initiated total BER and BER enzymes, uracil DNA glycosylase and DNA polymerase beta in 6 h and 24 h repair periods and increased the activity of 8-oxoguanine DNA glycosylase in 6 h repair in T24 BC cell line. Conclusion: The enhanced BER activity in BC cells in response to BCG treatment could be an important factor in BCG resistance.