International Journal of Molecular Sciences

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Open AccessEditorial

DNA Damage, DNA Repair, and Cancer: Second Edition

by Kazuhiko Kuwahara

Int. J. Mol. Sci. 2023, 24(23), 16835; https://doi.org/10.3390/ijms242316835 - 28 Nov 2023

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Abstract

Following our first Special Issue, we are pleased to present this Special Issue in the International Journal of Molecular Sciences, titled ‘DNA Damage, DNA Repair, and Cancer: Second Edition’ [...] Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

Research

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23 pages, 4393 KiB

Open AccessArticle

AZD-7648, a DNA-PK Inhibitor, Induces DNA Damage, Apoptosis, and Cell Cycle Arrest in Chronic and Acute Myeloid Leukemia Cells

by Beatriz Santos Lapa, Maria Inês Costa, Diana Figueiredo, Joana Jorge, Raquel Alves, Ana Raquel Monteiro, Beatriz Serambeque, Mafalda Laranjo, Maria Filomena Botelho, Isabel Marques Carreira, Ana Bela Sarmento-Ribeiro and Ana Cristina Gonçalves

Int. J. Mol. Sci. 2023, 24(20), 15331; https://doi.org/10.3390/ijms242015331 - 18 Oct 2023

Cited by 2 | Viewed by 1725

Abstract

The non-homologous end joining pathway is vital for repairing DNA double-strand breaks (DSB), with DNA-dependent protein kinase (DNA-PK) playing a critical role. Altered DNA damage response (DDR) in chronic (CML) and acute myeloid leukemia (AML) offers potential therapeutic opportunities. We studied the therapeutic [...] Read more.

The non-homologous end joining pathway is vital for repairing DNA double-strand breaks (DSB), with DNA-dependent protein kinase (DNA-PK) playing a critical role. Altered DNA damage response (DDR) in chronic (CML) and acute myeloid leukemia (AML) offers potential therapeutic opportunities. We studied the therapeutic potential of AZD-7648 (DNA-PK inhibitor) in CML and AML cell lines. This study used two CML (K-562 and LAMA-84) and five AML (HEL, HL-60, KG-1, NB-4, and THP-1) cell lines. DDR gene mutations were obtained from the COSMIC database. The copy number and methylation profile were evaluated using MS-MLPA and DDR genes, and telomere length using qPCR. p53 protein expression was assessed using Western Blot, chromosomal damage through cytokinesis-block micronucleus assay, and γH2AX levels and DSB repair kinetics using flow cytometry. Cell density and viability were analyzed using trypan blue assay after treatment with AZD-7648 in concentrations ranging from 10 to 200 µM. Cell death, cell cycle distribution, and cell proliferation rate were assessed using flow cytometry. The cells displayed different DNA baseline damage, DDR gene expressions, mutations, genetic/epigenetic changes, and p53 expression. Only HEL cells displayed inefficient DSB repair. The LAMA-84, HEL, and KG-1 cells were the most sensitive to AZD-7648, whereas HL-60 and K-562 showed a lower effect on density and viability. Besides the reduction in cell proliferation, AZD-7648 induced apoptosis, cell cycle arrest, and DNA damage. In conclusion, these results suggest that AZD-7648 holds promise as a potential therapy for myeloid leukemias, however, with variations in drug sensitivity among tested cell lines, thus supporting further investigation to identify the specific factors influencing sensitivity to this DNA-PK inhibitor. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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20 pages, 4479 KiB

Open AccessArticle

Chemical Inhibition of RPA by HAMNO Alters Cell Cycle Dynamics by Impeding DNA Replication and G2-to-M Transition but Has Little Effect on the Radiation-Induced DNA Damage Response

by Rositsa Dueva, Lisa Marie Krieger, Fanghua Li, Daxian Luo, Huaping Xiao, Martin Stuschke, Eric Metzen and George Iliakis

Int. J. Mol. Sci. 2023, 24(19), 14941; https://doi.org/10.3390/ijms241914941 - 6 Oct 2023

Cited by 2 | Viewed by 1171

Abstract

Replication protein A (RPA) is the major single-stranded DNA (ssDNA) binding protein that is essential for DNA replication and processing of DNA double-strand breaks (DSBs) by homology-directed repair pathways. Recently, small molecule inhibitors have been developed targeting the RPA70 subunit and preventing RPA [...] Read more.

Replication protein A (RPA) is the major single-stranded DNA (ssDNA) binding protein that is essential for DNA replication and processing of DNA double-strand breaks (DSBs) by homology-directed repair pathways. Recently, small molecule inhibitors have been developed targeting the RPA70 subunit and preventing RPA interactions with ssDNA and various DNA repair proteins. The rationale of this development is the potential utility of such compounds as cancer therapeutics, owing to their ability to inhibit DNA replication that sustains tumor growth. Among these compounds, (1Z)-1-[(2-hydroxyanilino) methylidene] naphthalen-2-one (HAMNO) has been more extensively studied and its efficacy against tumor growth was shown to arise from the associated DNA replication stress. Here, we study the effects of HAMNO on cells exposed to ionizing radiation (IR), focusing on the effects on the DNA damage response and the processing of DSBs and explore its potential as a radiosensitizer. We show that HAMNO by itself slows down the progression of cells through the cell cycle by dramatically decreasing DNA synthesis. Notably, HAMNO also attenuates the progression of G2-phase cells into mitosis by a mechanism that remains to be elucidated. Furthermore, HAMNO increases the fraction of chromatin-bound RPA in S-phase but not in G2-phase cells and suppresses DSB repair by homologous recombination. Despite these marked effects on the cell cycle and the DNA damage response, radiosensitization could neither be detected in exponentially growing cultures, nor in cultures enriched in G2-phase cells. Our results complement existing data on RPA inhibitors, specifically HAMNO, and suggest that their antitumor activity by replication stress induction may not extend to radiosensitization. However, it may render cells more vulnerable to other forms of DNA damaging agents through synthetically lethal interactions, which requires further investigation. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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21 pages, 4558 KiB

Open AccessArticle

Transcriptomic Analysis of CRISPR/Cas9-Mediated PARP1-Knockout Cells under the Influence of Topotecan and TDP1 Inhibitor

by Nadezhda S. Dyrkheeva, Anastasia A. Malakhova, Aleksandra L. Zakharenko, Larisa S. Okorokova, Dmitriy N. Shtokalo, Sophia V. Pavlova, Sergey P. Medvedev, Suren M. Zakian, Anna A. Nushtaeva, Alexey E. Tupikin, Marsel R. Kabilov, Svetlana N. Khodyreva, Olga A. Luzina, Nariman F. Salakhutdinov and Olga I. Lavrik

Int. J. Mol. Sci. 2023, 24(6), 5148; https://doi.org/10.3390/ijms24065148 - 7 Mar 2023

Cited by 3 | Viewed by 2236

Abstract

Topoisomerase 1 (TOP1) is an enzyme that regulates DNA topology and is essential for replication, recombination, and other processes. The normal TOP1 catalytic cycle involves the formation of a short-lived covalent complex with the 3′ end of DNA (TOP1 cleavage complex, TOP1cc), which [...] Read more.

Topoisomerase 1 (TOP1) is an enzyme that regulates DNA topology and is essential for replication, recombination, and other processes. The normal TOP1 catalytic cycle involves the formation of a short-lived covalent complex with the 3′ end of DNA (TOP1 cleavage complex, TOP1cc), which can be stabilized, resulting in cell death. This fact substantiates the effectiveness of anticancer drugs—TOP1 poisons, such as topotecan, that block the relegation of DNA and fix TOP1cc. Tyrosyl-DNA phosphodiesterase 1 (TDP1) is able to eliminate TOP1cc. Thus, TDP1 interferes with the action of topotecan. Poly(ADP-ribose) polymerase 1 (PARP1) is a key regulator of many processes in the cell, such as maintaining the integrity of the genome, regulation of the cell cycle, cell death, and others. PARP1 also controls the repair of TOP1cc. We performed a transcriptomic analysis of wild type and PARP1 knockout HEK293A cells treated with topotecan and TDP1 inhibitor OL9-119 alone and in combination. The largest number of differentially expressed genes (DEGs, about 4000 both up- and down-regulated genes) was found in knockout cells. Topotecan and OL9-119 treatment elicited significantly fewer DEGs in WT cells and negligible DEGs in PARP1-KO cells. A significant part of the changes caused by PARP1-KO affected the synthesis and processing of proteins. Differences under the action of treatment with TOP1 or TDP1 inhibitors alone were found in the signaling pathways for the development of cancer, DNA repair, and the proteasome. The drug combination resulted in DEGs in the ribosome, proteasome, spliceosome, and oxidative phosphorylation pathways. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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18 pages, 3460 KiB

Open AccessArticle

VRK1 Kinase Activity Modulating Histone H4K16 Acetylation Inhibited by SIRT2 and VRK-IN-1

by Eva Monte-Serrano and Pedro A. Lazo

Int. J. Mol. Sci. 2023, 24(5), 4912; https://doi.org/10.3390/ijms24054912 - 3 Mar 2023

Cited by 6 | Viewed by 2528

Abstract

The accessibility of DNA to different cellular functions requires a dynamic regulation of chromatin organization that is mediated by different epigenetic modifications, which regulate chromatin accessibility and degree of compaction. These epigenetic modifications, particularly the acetylation of histone H4 in lysine 14 (H4K16ac), [...] Read more.

The accessibility of DNA to different cellular functions requires a dynamic regulation of chromatin organization that is mediated by different epigenetic modifications, which regulate chromatin accessibility and degree of compaction. These epigenetic modifications, particularly the acetylation of histone H4 in lysine 14 (H4K16ac), determine the degree of chromatin accessibility to different nuclear functions, as well as to DNA damage drugs. H4K16ac is regulated by the balance between two alternative histone modifications, acetylation and deacetylation, which are mediated by acetylases and deacetylases. Tip60/KAT5 acetylates, and SIRT2 deacetylates histone H4K16. However, the balance between these two epigenetic enzymes is unknown. VRK1 regulates the level of H4K16 acetylation by activating Tip60. We have shown that the VRK1 and SIRT2 are able to form a stable protein complex. For this work, we used in vitro interaction, pull-down and in vitro kinase assays. In cells, their interaction and colocalization were detected by immunoprecipitation and immunofluorescence. The kinase activity of VRK1 is inhibited by a direct interaction of its N-terminal kinase domain with SIRT2 in vitro. This interaction causes a loss of H4K16ac similarly to the effect of a novel VRK1 inhibitor (VRK-IN-1) or VRK1 depletion. The use of specific SIRT2 inhibitors in lung adenocarcinoma cells induces H4K16ac, contrary to the novel VRK-IN-1 inhibitor, which prevents H4K16ac and a correct DNA damage response. Therefore, the inhibition of SIRT2 can cooperate with VRK1 in the accessibility of drugs to chromatin in response to DNA damage caused by doxorubicin. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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12 pages, 3813 KiB

Open AccessArticle

The Combination of Panobinostat and Melphalan for the Treatment of Patients with Multiple Myeloma

by Maria Gkotzamanidou, Evangelos Terpos, Meletios A. Dimopoulos and Vassilis L. Souliotis

Int. J. Mol. Sci. 2022, 23(24), 15671; https://doi.org/10.3390/ijms232415671 - 10 Dec 2022

Cited by 3 | Viewed by 1665

Abstract

Histone deacetylase inhibitors show synergy with several genotoxic drugs. Herein, we investigated the biological impact of the combined treatment of panobinostat and melphalan in multiple myeloma (MM). DNA damage response (DDR) parameters and the expression of DDR-associated genes were analyzed in bone marrow [...] Read more.

Histone deacetylase inhibitors show synergy with several genotoxic drugs. Herein, we investigated the biological impact of the combined treatment of panobinostat and melphalan in multiple myeloma (MM). DNA damage response (DDR) parameters and the expression of DDR-associated genes were analyzed in bone marrow plasma cells (BMPCs) and peripheral blood mononuclear cells (PBMCs) from 26 newly diagnosed MM patients. PBMCs from 25 healthy controls (HC) were examined in parallel. Compared with the ex vivo melphalan-only treatment, combined treatment with panobinostat and melphalan significantly reduced the efficiency of nucleotide excision repair (NER) and double-strand-break repair (DSB/R), enhanced the accumulation of DNA lesions (monoadducts and DSBs), and increased the apoptosis rate only in patients’ BMPCs (all p < 0.001); marginal changes were observed in PBMCs from the same patients or HC. Accordingly, panobinostat pre-treatment decreased the expression levels of critical NER (DDB2, XPC) and DSB/R (MRE11A, PRKDC/DNAPKc, RAD50, XRCC6/Ku70) genes only in patients’ BMPCs; no significant changes were observed in PBMCs from patients or HC. Together, our findings demonstrate that panobinostat significantly increased the melphalan sensitivity of malignant BMPCs without increasing the melphalan sensitivity of PBMCs from the same patients, thus paving the way for combination therapies in MM with improved anti-myeloma efficacy and lower side effects. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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Review

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14 pages, 1600 KiB

Open AccessReview

The Initial Hepatitis B Virus-Hepatocyte Genomic Integrations and Their Role in Hepatocellular Oncogenesis

by Tomasz I. Michalak

Int. J. Mol. Sci. 2023, 24(19), 14849; https://doi.org/10.3390/ijms241914849 - 3 Oct 2023

Cited by 2 | Viewed by 1364

Abstract

Hepatitis B virus (HBV) remains a dominant cause of hepatocellular carcinoma (HCC). Recently, it was shown that HBV and woodchuck hepatitis virus (WHV) integrate into the hepatocyte genome minutes after invasion. Retrotransposons and transposable sequences were frequent sites of the initial insertions, suggesting [...] Read more.

Hepatitis B virus (HBV) remains a dominant cause of hepatocellular carcinoma (HCC). Recently, it was shown that HBV and woodchuck hepatitis virus (WHV) integrate into the hepatocyte genome minutes after invasion. Retrotransposons and transposable sequences were frequent sites of the initial insertions, suggesting a mechanism for spontaneous HBV DNA dispersal throughout the hepatocyte genome. Several somatic genes were also identified as early insertional targets in infected hepatocytes and woodchuck livers. Head-to-tail joints (HTJs) dominated amongst fusions, indicating their creation by non-homologous end-joining (NHEJ). Their formation coincided with the robust oxidative damage of hepatocyte DNA. This was associated with the activation of poly(ADP-ribose) polymerase 1 (PARP1)-mediated dsDNA repair, as reflected by the augmented transcription of PARP1 and XRCC1; the PARP1 binding partner OGG1, a responder to oxidative DNA damage; and increased activity of NAD⁺, a marker of PARP1 activation, and HO1, an indicator of cell oxidative stress. The engagement of the PARP1-mediated NHEJ repair pathway explains the HTJ format of the initial merges. The findings show that HBV and WHV are immediate inducers of oxidative DNA damage and hijack dsDNA repair to integrate into the hepatocyte genome, and through this mechanism, they may initiate pro-oncogenic processes. Tracking initial integrations may uncover early markers of HCC and help to explain HBV-associated oncogenesis. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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19 pages, 1294 KiB

Open AccessReview

Poly(ADP-ribose) Polyremase-1 (PARP-1) Inhibition: A Promising Therapeutic Strategy for ETS-Expressing Tumours

by Arnaud J. Legrand, Souhaila Choul-li, Vincent Villeret and Marc Aumercier

Int. J. Mol. Sci. 2023, 24(17), 13454; https://doi.org/10.3390/ijms241713454 - 30 Aug 2023

Cited by 1 | Viewed by 1409

Abstract

ETS transcription factors are a highly conserved family of proteins involved in the progression of many cancers, such as breast and prostate carcinomas, Ewing’s sarcoma, and leukaemias. This significant involvement can be explained by their roles at all stages of carcinogenesis progression. Generally, [...] Read more.

ETS transcription factors are a highly conserved family of proteins involved in the progression of many cancers, such as breast and prostate carcinomas, Ewing’s sarcoma, and leukaemias. This significant involvement can be explained by their roles at all stages of carcinogenesis progression. Generally, their expression in tumours is associated with a poor prognosis and an aggressive phenotype. Until now, no efficient therapeutic strategy had emerged to specifically target ETS-expressing tumours. Nevertheless, there is evidence that pharmacological inhibition of poly(ADP-ribose) polymerase-1 (PARP-1), a key DNA repair enzyme, specifically sensitises ETS-expressing cancer cells to DNA damage and limits tumour progression by leading some of the cancer cells to death. These effects result from a strong interplay between ETS transcription factors and the PARP-1 enzyme. This review summarises the existing knowledge of this molecular interaction and discusses the promising therapeutic applications. Full article

(This article belongs to the Special Issue DNA Damage, DNA Repair, and Cancer 2.0)

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