Depletion of tyrosyl DNA phosphodiesterase 2 activity enhances etoposide-mediated double-strand break formation and cell killing

Kont, Yasemin Saygideğer; Dutta, Arijit; Mallisetty, Apurva; Mathew, Jeena; Minas, Tsion Z.; Kraus, Christina N.; Dhopeshwarkar, Priyanka; Kallakury, Bhaskar; Mitra, Sankar; Üren, Aykut; Adhikari, Sanjay

doi:10.1016/j.dnarep.2016.04.009

Cited by 25 publications

(15 citation statements)

References 40 publications

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“…TDP2 is primarily involved in catalyzing the hydrolysis of 5′-phosphotyrosyl bonds thereby releasing trapped Top II or fragments thereof from the 5′ end of aberrant Top II-induced DSBs (Figures 6A and 7D ). Consistent with this, TDP2-depleted human A549 cells show hypersensitivity to etoposide and a higher number of etoposide-induced γ-H2AX foci whereas overexpression of wild-type TDP2 but not catalytically inactive mutants (TDP2 D262A or TDP2 E152A ) in yeast ISE2 cells leads to a significant increase in resistance to etoposide ( 97 , 106 ). Similarly, in DT40 chicken cells, knocking out all three alleles of TDP2 results in a profound hypersensitivity to etoposide as well as other structurally diverse Top II poisons, doxorubicin and amsacrine (m-AMSA) which can be complemented by expressing recombinant WT hTDP2.…”

Section: Tyrosyl–dna Phosphodiesterase 2 (Tdp2)supporting

confidence: 60%

“…The discovery of NSC111041 as a TDP2 inhibitor that inhibits TDP2-DNA binding without intercalating in the DNA is promising. NSC111041 also enhanced the etoposide-induced cytotoxicity in a TDP2-dependent manner and led to increased etoposide-induced γ-H2AX foci by affecting DSB repair, suggesting that this small molecule inhibitor can enhance the efficacy of Top II poisons ( 106 ). However, similar to inhibitors of TDP1, efforts at development of TDP2 inhibitors are in the preliminary stage.…”

Section: Tyrosyl–dna Phosphodiesterase 2 (Tdp2)mentioning

confidence: 99%

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Tyrosyl–DNA phosphodiesterases: rescuing the genome from the risks of relaxation

Kawale

Povirk

2017

Nucleic Acids Research

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Tyrosyl–DNA Phosphodiesterases 1 (TDP1) and 2 (TDP2) are eukaryotic enzymes that clean-up after aberrant topoisomerase activity. While TDP1 hydrolyzes phosphotyrosyl peptides emanating from trapped topoisomerase I (Top I) from the 3′ DNA ends, topoisomerase 2 (Top II)-induced 5′-phosphotyrosyl residues are processed by TDP2. Even though the canonical functions of TDP1 and TDP2 are complementary, they exhibit little structural or sequence similarity. Homozygous mutations in genes encoding these enzymes lead to the development of severe neurodegenerative conditions due to the accumulation of transcription-dependent topoisomerase cleavage complexes underscoring the biological significance of these enzymes in the repair of topoisomerase–DNA lesions in the nervous system. TDP1 can promiscuously process several blocked 3′ ends generated by DNA damaging agents and nucleoside analogs in addition to hydrolyzing 3′-phosphotyrosyl residues. In addition, deficiency of these enzymes causes hypersensitivity to anti-tumor topoisomerase poisons. Thus, TDP1 and TDP2 are promising therapeutic targets and their inhibitors are expected to significantly synergize the effects of current anti-tumor therapies including topoisomerase poisons and other DNA damaging agents. This review covers the structural aspects, biology and regulation of these enzymes, along with ongoing developments in the process of discovering safe and effective TDP inhibitors.

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Section: Tyrosyl–dna Phosphodiesterase 2 (Tdp2)supporting

confidence: 60%

Section: Tyrosyl–dna Phosphodiesterase 2 (Tdp2)mentioning

confidence: 99%

Tyrosyl–DNA phosphodiesterases: rescuing the genome from the risks of relaxation

Kawale

Povirk

2017

Nucleic Acids Research

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“…On the other hand, amine intermediates 21 was synthesized via an S N Ar reaction of phenolic alcohol 22 with 4-nitrophenyl fluoride 23 followed by the reduction of the nitro group. To advance the synthesis, amines 20 or 21 were reacted with 6-chlorouracil (13) to give intermediate 25 which upon condensation with 4cyano-2-fluorobenzaldehyde (16) furnished subtype 11 (Scheme 1B).…”

Section: Chemistrymentioning

confidence: 99%

Novel Deazaflavin Analogues Potently Inhibited Tyrosyl DNA Phosphodiesterase 2 (TDP2) and Strongly Sensitized Cancer Cells toward Treatment with Topoisomerase II (TOP2) Poison Etoposide

et al. 2019

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Topoisomerase II (TOP2) poisons as anticancer drugs work by trapping TOP2 cleavage complexes (TOP2cc) to generate DNA damage. Repair of such damage by tyrosyl DNA phosphodiesterase 2 (TDP2) could render cancer cells resistant to TOP2 poisons. Inhibiting TDP2 thus represents an attractive mechanism-based chemosensitization approach. Currently known TDP2 inhibitors lack cellular potency and / or permeability. We report herein two novel subtypes of the deazaflavin TDP2 inhibitor core. By introducing an additional phenyl ring to the N-10 phenyl ring (subtype 11), or to the N-3 site of the deazaflavin scaffold (subtype 12) we have generated novel analogues with considerably improved biochemical potency and / or permeability. Importantly, many analogues of both subtypes, particularly compounds 11a, 11e, 12a, 12b and 12h, exhibited much stronger cancer cell sensitizing effect than the best reported previous analogue 4a toward the treatment with etoposide (ETP), suggesting that these analogues could serve as effective cellular probes.

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“…3 ) (Kankanala et al, 2016), the triple inhibitors TOP1/TDP1/TDP2 indenoisoquinolines (e.g. 4 ) (Beck et al, 2016; Wang et al, 2017), compounds 5 (NSC375986), 6 (NSC114532) (Kossmann et al, 2016) and 7 (NSC111041) (Kont et al, 2016). Deazaflavin derivatives are the only described TDP2 inhibitors with activities in the nanomolar range.…”

Section: Introductionmentioning

confidence: 99%

New fluorescence-based high-throughput screening assay for small molecule inhibitors of tyrosyl-DNA phosphodiesterase 2 (TDP2)

Ribeiro

Kankanala

Shi

et al. 2018

European Journal of Pharmaceutical Sciences

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Tyrosyl-DNA phosphodiesterase 2 (TDP2) repairs topoisomerase II (TOP2) mediated DNA damages and causes resistance to TOP2-targeted cancer therapy. Inhibiting TDP2 could sensitize cancer cells toward TOP2 inhibitors. However, potent TDP2 inhibitors with favorable physicochemical properties are not yet reported. Therefore, there is a need to search for novel molecular scaffolds capable of inhibiting TDP2. We report herein a new simple, robust, homogenous mix-and-read fluorescence biochemical assay based using humanized zebrafish TDP2 (14M_zTDP2), which provides biochemical and molecular structure basis for TDP2 inhibitor discovery. The assay was validated by screening a preselected library of 1600 compounds (Z' ≥ 0.72) in a 384-well format, and by running in parallel gel-based assays with fluorescent DNA substrates. This library was curated via virtual high throughput screening (vHTS) of 460,000 compounds from Chembridge Library, using the crystal structure of the novel surrogate protein 14M_zTDP2. From this primary screening, we selected the best 32 compounds (2% of the library) to further assess their TDP2 inhibition potential, leading to the IC determination of 10 compounds. Based on the dose-response curve profile, pan-assay interference compounds (PAINS) structure identification, physicochemical properties and efficiency parameters, two hit compounds, 11a and 19a, were tested using a novel secondary fluorescence gel-based assay. Preliminary structure-activity relationship (SAR) studies identified guanidine derivative 12a as an improved hit with a 6.4-fold increase in potency over the original HTS hit 11a. This study highlights the importance of the development of combination approaches (biochemistry, crystallography and high throughput screening) for the discovery of TDP2 inhibitors.

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Depletion of tyrosyl DNA phosphodiesterase 2 activity enhances etoposide-mediated double-strand break formation and cell killing

Cited by 25 publications

References 40 publications

Tyrosyl–DNA phosphodiesterases: rescuing the genome from the risks of relaxation

Tyrosyl–DNA phosphodiesterases: rescuing the genome from the risks of relaxation

Novel Deazaflavin Analogues Potently Inhibited Tyrosyl DNA Phosphodiesterase 2 (TDP2) and Strongly Sensitized Cancer Cells toward Treatment with Topoisomerase II (TOP2) Poison Etoposide

New fluorescence-based high-throughput screening assay for small molecule inhibitors of tyrosyl-DNA phosphodiesterase 2 (TDP2)

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