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AN-7, a butyric acid prodrug, sensitizes cutaneous T-cell lymphoma cell lines to doxorubicin via inhibition of DNA double strand breaks repair

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Summary

We previously found that the novel histone deacetylase inhibitor (HDACI) butyroyloxymethyl diethylphosphate (AN-7) had greater selectivity against cutaneous T-cell lymphoma (CTCL) than SAHA. AN-7 synergizes with doxorubicin (Dox), an anthracycline antibiotic that induces DNA breaks. This study aimed to elucidate the mechanism underlying the effect of AN-7 on Dox-induced double-strand DNA breaks (DSBs) in CTCL, MyLa and Hut78 cell lines. The following markers/assays were employed: comet assay; western blot of γH2AX and p-KAP1; immunofluorescence of γH2AX nuclear foci; Western blot of repair protein; quantification of DSBs-repair through homologous recombination. DSB induction by Dox was evidenced by an increase in DSB markers, and DSBs-repair, by their subsequent decrease. The addition of AN-7 slightly increased Dox induction of DSBs in MyLa cells with no effect in Hut78 cells. AN-7 inhibited the repair of Dox-induced DSBs, with a more robust effect in Hut78. Treatment with AN-7 followed by Dox reduced the expression of DSB-repair proteins, with direct interference of AN-7 with the homologous recombination repair. AN-7 sensitizes CTCL cell lines to Dox, and when combined with Dox, sustains unrepaired DSBs by suppressing repair protein expression. Our data provide a mechanistic rationale for combining AN-7 with Dox or other DSB inducers as a therapeutic modality in CTCL.

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Acknowledgments

We would like to thank Robert Gniadecki for the Hut78 and the MyLa cell lines and Alexandre Sellam and Elsa Levy for helping with the data analysis. This study was funded in part by the Alexander Samidoda Foundation of the Israel Cancer Association.

Funding

The work was partially supported by Alexander Samidoda Foundation of the Israel Cancer Association.

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Authors and Affiliations

  1. Laboratory for Molecular Dermatology, Felsenstein Medical Research Center, Petach Tikva and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel

    Lilach Moyal, Neta Goldfeiz, Batia Gorovitz & Emmilia Hodak

  2. Department of Dermatology, Rabin Medical Center, Petach Tikva, and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel

    Lilach Moyal, Neta Goldfeiz, Batia Gorovitz & Emmilia Hodak

  3. Laboratory for Pharmacology and Experimental Oncology, Felsenstein Medical Research Center, Petach Tikva, and Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel

    Ada Rephaeli & Nataly Tarasenko

  4. The David and Inez Myers Laboratory for Cancer Research, Department of Human Molecular Genetics and Biochemistry, Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel

    Efrat Tal & Yael Ziv

  5. Division of Medicinal Chemistry, Department of Chemistry, Bar Ilan University, Ramat Gan, Israel

    Abraham Nudelman

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  1. Lilach Moyal
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Correspondence to Lilach Moyal.

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Conflict of interest

Lilach Moyal declares that she has no conflict of interest.

Neta Goldfeiz declares that she has no conflict of interest.

Batia Gorovitz declares that she has no conflict of interest.

Ada Rephaeli declares that she has no conflict of interest.

Efrat Tal declares that she has no conflict of interest.

Nataly Tarasenko declares that she has no conflict of interest.

Abraham Nudelman declares that he has no conflict of interest.

Yael Ziv declares that she has no conflict of interest.

Emmilia Hodak declares that she has no conflict of interest.

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Figure S1

The expression levels of pKAP1 (a,c) and γH2AX (b,d) in western blot as presented in Fig. 2a,b were normalized to actin and presented in a column curve, MyLa (a,b), Hut78 (c,d). (TIFF 3203 kb)

High Resolution Image (GIF 18 kb)

Figure S2

The expression levels of DDR proteins in western blot as presented in Fig. 3a,b were normalized to actin. The expression in MyLa-treated cells at 48 h (a) and Hut78 cells at 24 h (b) relative to the untreated cells are presented in column curves. (TIFF 3112 kb)

High Resolution Image (GIF 8 kb)

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Moyal, L., Goldfeiz, N., Gorovitz, B. et al. AN-7, a butyric acid prodrug, sensitizes cutaneous T-cell lymphoma cell lines to doxorubicin via inhibition of DNA double strand breaks repair. Invest New Drugs 36, 1–9 (2018). https://doi.org/10.1007/s10637-017-0500-x

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