Tyrosyl DNA phosphodiesterase 2 (TDP2) facilitates the repair of topoisomerase II (TOP2)-linked DNA double-strand breaks and, as a consequence, is required for cellular resistance to TOP2 "poisons". Recently, a deazaflavin series of compounds were identified as potent inhibitors of TDP2, in vitro. Here, however, we show that while some deazaflavins can induce cellular sensitivity to the TOP2 poison etoposide, they do so independently of TDP2 status. Consistent with this, both the cellular level of etoposide-induced TOP2 cleavage complexes and the intracellular concentration of etoposide was increased by incubation with deazaflavin, suggesting an impact of these compounds on etoposide uptake/efflux. In addition, deazaflavin failed to increase the level of TOP2 cleavage complexes or sensitivity induced by m-AMSA, which is a different class of TOP2 poison to which TDP2-defective cells are also sensitive. In conclusion, while deazaflavins are potent inhibitors of TDP2 in vitro, their limited cell permeability and likely interference with etoposide influx/efflux limits their utility in cells.

Department of Genome Dynamics Institute of Molecular Genetics of the Czech Academy of Sciences 142 20 Prague 4 Czech Republic

Genome Damage and Stability Centre School of Life Sciences University of Sussex Falmer Brighton BN1 9RQ United Kingdom

Sussex Drug Discovery Centre School of Life Sciences University of Sussex Falmer Brighton BN1 9RQ United Kingdom

See more in PubMed

Cortes Ledesma F, El-Khamisy SF, Zuma MC, Osborn K, Caldecott KW. A human 5’-tyrosyl DNA phosphodiesterase that repairs topoisomerase-mediated DNA damage. Nature. 2009;461:674–678. PubMed

Zeng Z, Cortes Ledesma F, El-Khamisy SF, Caldecott KW. TDP2/TTRAP is the major 5’-tyrosyl DNA phosphodiesterase activity in vertebrate cells and is critical for cellular resistance to topoisomerase II-induced DNA damage. J Biol Chem. 2011;286:403–409. PubMed PMC

Gómez-Herreros F, Romero-Granados R, Zeng Z, Alvarez-Quilón A, Quintero C, Ju L, Umans L, Vermeire L, Huylebroeck D, Caldecott KW, Cortes Ledesma F. TDP2-Dependent Non-Homologous End-Joining Protects against Topoisomerase II-Induced DNA Breaks and Genome Instability in Cells and In Vivo. PLoS Genet. 2013;9:e1003226. PubMed PMC

Cuya SM, Bjornsti M-A, van Waardenburg RCAM. DNA topoisomerase-targeting chemotherapeutics: what’s new? Cancer Chemother Pharmacol. 2017;80:1–14. PubMed PMC

Nitiss JL. Targeting DNA topoisomerase II in cancer chemotherapy. Nat Rev Cancer. 2009;9:338–350. PubMed PMC

Virgen-Slane R, Rozovics JM, Fitzgerald KD, Ngo T, Chou W, van der Heden van Noort GJ, Filippov DV, Gershon PD, Semler BL. An RNA virus hijacks an incognito function of a DNA repair enzyme. Proc Natl Acad Sci USA. 2012;109:14634–14639. PubMed PMC

Maciejewski S, Nguyen JHC, Gómez-Herreros F, Cortes Ledesma F, Caldecott KW, Semler BL. Divergent Requirement for a DNA Repair Enzyme during Enterovirus Infections. MBio. 2015;7:e01931-15. PubMed PMC

Cui X, McAllister R, Boregowda R, Sohn JA, Ledesma FC, Caldecott KW, Seeger C, Hu J. Does Tyrosyl DNA Phosphodiesterase-2 Play a Role in Hepatitis B Virus Genome Repair? PLoS ONE. 2015;10:e0128401. PubMed PMC

Königer C, Wingert I, Marsmann M, Rösler C, Beck J, Nassal M. Involvement of the host DNA-repair enzyme TDP2 in formation of the covalently closed circular DNA persistence reservoir of hepatitis B viruses. Proc Natl Acad Sci USA. 2014:201409986 PubMed PMC

Raoof A, Depledge P, Hamilton NM, Hamilton NS, Hitchin JR, Hopkins GV, Jordan AM, Maguire LA, McGonagle AE, Mould DP, Rushbrooke M, et al. Toxoflavins and Deazaflavins as the First Reported Selective Small Molecule Inhibitors of Tyrosyl-DNA Phosphodiesterase II. J Med Chem. 2013;56:6352–6370. PubMed

Hornyak P, Askwith T, Walker S, Komulainen E, Paradowski M, Pennicott LE, Bartlett EJ, Brissett NC, Raoof A, Watson M, Jordan AM, et al. Mode of action of DNA-competitive small molecule inhibitors of tyrosyl DNA phosphodiesterase 2. Biochem J. 2016:BCJ20160180 PubMed PMC

Marchand C, Abdelmalak M, Kankanala J, Huang S-Y, Kiselev E, Fesen K, Kurahashi K, Sasanuma H, Takeda S, Aihara H, Wang Z, et al. Deazaflavin Inhibitors of Tyrosyl-DNA Phosphodiesterase 2 (TDP2) Specific for the Human Enzyme and Active against Cellular TDP2. ACS Chem Biol. 2016:acschembio.5b01047 PubMed PMC

Caldecott K, Banks G, Jeggo P. DNA double-strand break repair pathways and cellular tolerance to inhibitors of topoisomerase II. Cancer Res. 1990;50:5778–5783. PubMed

Jeggo PA, Caldecott K, Pidsley S, Banks GR. Sensitivity of Chinese hamster ovary mutants defective in DNA double strand break repair to topoisomerase II inhibitors. Cancer Res. 1989;49:7057–7063. PubMed

Liu LF, Rowe TC, Yang L, Tewey KM, Chen GL. Cleavage of DNA by mammalian DNA topoisomerase II. J Biol Chem. 1983;258:15365–15370. PubMed