Targeting DNA Polymerase ß for Therapeutic Intervention

Goellner, Eva M.; Svilar, David; Almeida, Karen H.; Sobol, Robert W.

doi:10.2174/1874-470211205010068

Cited by 11 publications

(7 citation statements)

References 207 publications

(264 reference statements)

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“…Furthermore, the concentration at which pro- 13 is effective in HeLa cells is comparable to or superior to even the most recent reported Pol β inhibitors. 7, 9, 13 These observations indicate that Pol β inactivation is a promising approach for potentiating DNA damaging agents.…”

Section: Discussionmentioning

confidence: 97%

“…A structurally diverse group of Pol β inhibitors have been identified. 7, 9 Some of these molecules target the polymerase active site, while others such as the molecule examined here inhibit the enzyme’s lyase activity. Comparison of inhibitors from independent studies is challenging due to different cell types, DNA damaging agents employed, and inhibition mechanism.…”

Section: Discussionmentioning

confidence: 99%

“…The most effective inhibitors exhibit IC 50 ’s in the low micromolar range, and in some instances the selectivity for Pol β over other enzymes is not known and/or whether the molecule targets the polymerase or lyase activity. 7–9 The approach described herein was inspired by the discovery that lesions produced by anti-tumor agents that oxidatively damage DNA irreversibly inhibit Pol β and Pol λ (Scheme 2). 30–33 C4-AP and/or DOB are produced by C4′- and C5′-hydrogen atom abstraction, respectively.…”

mentioning

confidence: 99%

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Synergistic Effects of an Irreversible DNA Polymerase Inhibitor and DNA Damaging Agents on HeLa Cells

2017

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DNA repair is vital to maintaining genome integrity, but thwarts the effects of cytotoxic agents that target nucleic acids. Consequently, repair enzymes are potential targets for molecules that modulate cell function and anti-cancer therapeutics. DNA polymerase β (Pol β) is an attractive target because it plays a key role in base excision repair (BER), a primary pathway that repairs the effects of many DNA damaging agents. We previously identified an irreversible inhibitor of Pol β whose design was based upon a DNA lesion that inactivates Pol β and its back up BER enzyme, DNA polymerase λ (Pol λ). Using this molecule as a starting point we characterized an irreversible inhibitor (13) of Pol β (IC50 = 0.4 μM) and Pol λ (IC50 = 1.0 μM) from a 130-member library of candidates that is ~50-fold more effective against Pol β. Pro-13 (5 μM) is only slightly cytotoxic to human cervical cancer cells (HeLa), but potentiates the cytotoxicity of methyl methanesulfonate (MMS). DNA isolated from HeLa cells treated with MMS contain a ~3-fold greater amount of abasic sites when pro-13 is present, consistent with inhibition of DNA repair. Proinhibitor pro-13 continues to induce cytotoxicity in DNA damaged cells following MMS removal. HeLa cell cytotoxicity is increased ~100-fold following a 8 h incubation with pro-13 after cells that were originally subjected to conditions under which 20% of the cells survive and reproduce. The potentiation of MMS cytotoxicity by pro-13 is greater than any previously reported BER enzyme repair inhibitor.

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Section: Discussionmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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Synergistic Effects of an Irreversible DNA Polymerase Inhibitor and DNA Damaging Agents on HeLa Cells

2017

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“…6 This repair intermediate is then processed by Polβ, which removes the 5′-DRP structure to afford a 5′-phosphate and then adds the appropriate complementary base to the 3′-terminus. 7 In the final step, DNA ligase seals the nick. While cells and animals can survive without the different DNA glycosylases, albeit with increased sensitivity to DNA damaging agents, 8–11 the genetic deletion of APE-1, which is expressed ubiquitously, is lethal in cells.…”

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confidence: 99%

Identification and Characterization of Human Apurinic/Apyrimidinic Endonuclease-1 Inhibitors

et al. 2012

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The repair of abasic sites that arise in DNA from hydrolytic depurination/depyrimidination of the nitrogenous bases from the sugar-phosphate backbone and the action of DNA glycosylases on deaminated, oxidized and alkylated bases is critical to cell survival. Apurinic/Apyrimidinic Endonuclease-1/Redox Effector Factor-1 (APE-1; aka, APE1/Ref-1) is responsible for the initial removal of abasic lesions as part of the base excision repair pathway. Deletion of APE-1 activity is embryonic lethal in animals and is lethal in cells. Potential inhibitors of the repair function of APE-1 were identified based upon molecular modeling of the crystal structure of the APE-1 protein. We describe the characterization of several unique nM inhibitors using two complementary biochemical screens. The most active molecules all contain a 2-methyl-4-amino-6,7-dioxolo-quinoline structure that is predicted from the modeling to anchor the compounds in the endonuclease site of the protein. The mechanism of action of the selected compounds was probed by fluorescence and competition studies, which indicate, in a specific case, direct interaction between the inhibitor and the active site of the protein. It is demonstrated that the inhibitors induce time-dependent increases in the accumulation of abasic sites in cells at levels that correlate with their potency to inhibit APE-1 endonuclease excision. The inhibitor molecules also potentiate by 5-fold the toxicity of a DNA methylating agent that creates abasic sites. The molecules represent a new class of APE-1 inhibitors that can be used to probe the biology of this critical enzyme and to sensitize resistant tumor cells to the cytotoxicity of clinically used DNA damaging anticancer drugs.

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“…27,29,55–66 The kinetic mechanism involves HNL sequestering the polymerase–DNA–dNTP complex in an unreactive configuration that decomposes slowly. To evaluate if HNL can be utilized to chemosensitize damage DNA, which is repaired by BER, we examined two reagents TMZ and BLM.…”

Section: Discussionmentioning

confidence: 99%

Honokiol Inhibits DNA Polymerases β and λ and Increases Bleomycin Sensitivity of Human Cancer Cells

Gowda

Suo

Spratt

2017

Chem. Res. Toxicol.

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A major concept to sensitize cancer cells to DNA damaging agents is by inhibiting proteins in the DNA repair pathways. X-family DNA polymerases play critical roles in both base excision repair (BER) and nonhomologous end joining (NHEJ). In this study, we examined the effectiveness of honokiol to inhibit human DNA polymerase β (pol β), which is involved in BER, and DNA polymerase λ (pol λ), which is involved in NHEJ. Kinetic analysis with purified polymerases showed that honokiol inhibited DNA polymerase activity. The inhibition mode for the polymerases was a mixed-function noncompetitive inhibition with respect to the substrate, dCTP. The X-family polymerases, pol β and pol λ, were slightly more sensitive to inhibition by honokiol based on the Ki value of 4.0 μM for pol β, and 8.3 μM for pol λ, while the Ki values for pol η and Kf were 20 and 26 μM, respectively. Next we extended our studies to determine the effect of honokiol on the cytotoxicity of bleomycin and temozolomide in human cancer cell lines A549, MCF7, PANC-1, UACC903, and normal blood lymphocytes (GM12878). Bleomycin causes both single strand DNA damage that is repaired by BER and double strand breaks that are repaired by NHEJ, while temozolomide causes methylation damage repaired by BER and O6-alkylguanine-DNA alkyltransferase. The greatest effects were found with the honokiol and bleomycin combination in MCF7, PANC-1, and UACC903 cells, in which the EC50 values were decreased 10-fold. The temozolomide and honokiol combination was less effective; the EC50 values decreased three-fold due to the combination. It is hypothesized that the greater effect of honokiol on bleomycin is due to inhibition of the repair of the single strand and double strand damage. The synergistic activity shown by the combination of bleomycin and honokiol suggests that they can be used as combination therapy for treatment of cancer, which will decrease the therapeutic dosage and side effects of bleomycin.

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Targeting DNA Polymerase ß for Therapeutic Intervention

Cited by 11 publications

References 207 publications

Synergistic Effects of an Irreversible DNA Polymerase Inhibitor and DNA Damaging Agents on HeLa Cells

Synergistic Effects of an Irreversible DNA Polymerase Inhibitor and DNA Damaging Agents on HeLa Cells

Identification and Characterization of Human Apurinic/Apyrimidinic Endonuclease-1 Inhibitors

Honokiol Inhibits DNA Polymerases β and λ and Increases Bleomycin Sensitivity of Human Cancer Cells

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