Available online 12 August 2023, 106454

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Cisplatin, a widely prescribed chemotherapeutic agent for treating solid tumors, induces DNA adducts and activates cellular defense mechanisms, including DNA repair, cell cycle checkpoint control, and apoptosis. Considering the circadian rhythmicity displayed by most chemotherapeutic agents and their varying therapeutic efficacy based on treatment timing, our study aimed to investigate whether the circadian clock system influences the DNA damage responses triggered by cisplatin in synchronized cells. We examined the DNA damage responses in circadian-synchronized wild-type mouse embryonic fibroblasts (WT-MEF; clock-proficient cells), cryptochrome1 and 2 double knock-out MEF (CRYDKO; clock-deficient cells), and mouse hepatocarcinoma Hepa1c1c7 cells. Varying the treatment time resulted in a significant difference in the rate of platinum-DNA adduct removal specifically in circadian-synchronized WT-MEF, while CRYDKO did not exhibit such variation. Moreover, diurnal variation in other DNA damage responses, such as cell cycle checkpoint activity indicated by p53 phosphorylation status and apoptosis measured by DNA break frequency, was observed only in circadian-synchronized WT-MEF, not in CRYDKO or mouse hepatocarcinoma Hepa1c1c7 cells. These findings highlight that the DNA damage responses triggered by cisplatin are indeed governed by circadian control exclusively in clock-proficient cells. This outcome bears potential implications for enhancing or devising chronotherapy approaches for cancer patients.

Section snippetsINTRIDUCTION

Circadian rhythms, characterized by diurnal fluctuations in physiology and behavior with a period of approximately 24 hours, are regulated by the circadian clock, a molecular time-keeping system present in virtually every cell (Dibner et al., 2010). The mammalian core molecular clock consists of an autonomous transcriptional and translational feedback loop (TTFL). This loop involves the activation of CLOCK-BMAL1 complex, a heterodimeric transcription activator, which induces the transcription of

Circadian synchronization

Wild-type mouse embryonic fibroblast (WT-MEF) and CRY1/2 double knockout mouse embryonic fibroblast (CRYDKO) cell lines used in this study were provided by Dr. KJ Kim at Seoul National University (Choi et al., 2016). Hepa1c1c7 cells were purchased from ATCC. All cell lines were cultured in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin. The cells were maintained at 37°C with 5% CO2. To ensure the absence of Mycoplasma

RESULTS

To generate a circadian rhythm of gene expression in normal fibroblasts, we treated them with forskolin (FSK) to induce circadian synchronization via activation through the cAMP/PKA pathway, which has been established previously for various types of cells (Hughes et al., 2009). As shown in Fig. 1 A, FSK treatment generated rhythmic CLOCK/BMAL1 activity in wild-type mouse embryonic fibroblasts (WT-MEF), resulting in a robust circadian oscillation of the proteins of clock-controlled genes,

DISCUSSION

Recent advancements in chronobiological research have significantly enhanced our understanding of the circadian clock's functional roles and its close relationship with human health. It is now widely recognized that the timing of drug administration, guided by the circadian rhythm of both pharmacokinetics and pharmacodynamics, plays a crucial role in optimizing treatment outcomes (Levi and Schibler, 2007). Chronotherapy, a form of personalized medicine, aims to administer anti-cancer drugs at

Funding

RS-2023-00250327 and NRF-2018R1D1A3B07043817

CRediT authorship contribution statement

Jeseok Jeon: Conceptualization, Investigation, Data Curation. Sanggon Lee: Investigation, Methodology, Validation. Jeong-Min Park: Investigation, Visualization. Tae-Hee Lee: Investigation, Data Curation. Tae-Hong Kang: Writing-original Draft, Writing-Review & Editing, Funding Acquisition

Acknowledgements

We would like to thank to Dr. KJ Kim (Seoul National University) for the generous providing of WT- and CRYDKO-MEF.

Ethics approval and consent to participate

Not applicable.

Consent for publication

All authors agreed to the publication of the manuscript.

Competing interests

The authors declare that there is no competing interests.

Authors' contributions

Conceptualization, T.H.K.; methodology, J.J., J.M.P, T.H.L. and S.L.; validation, J.J.,S.L., J.M.P. and T.H.L.; formal analysis, J.J., S.L., J.M.P. and T.H.L.; investigation, T.H.K., J.J., S.L., J.M.P. and T.H.L.; manuscript writing, T.H.K.; funding

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