Abstract
Cisplatin is the most commonly used chemotherapeutic drug for managing solid tumors. However, toxicity and the innate or acquired resistance of cancer cells to the drug limit its usefulness. Cisplatin kills cells by forming cisplatin-DNA adducts, most commonly the Pt-d(GpG) diadduct. We recently showed that, in mice, repair of this adduct 2 h following injection is controlled by two circadian programs. 1) The circadian clock controls transcription of 2000 genes in liver and, via transcription-directed repair, controls repair of the transcribed strand (TS) of these genes in a rhythmic fashion unique to each gene's phase of transcription. 2) The excision repair activity itself is controlled by the circadian clock with a single phase at which the repair of the nontranscribed strand (NTS) and the rest of the genome takes place. Here, we followed the repair kinetic for long periods genome-wide both globally and at single nucleotide resolution by the Excision Repair-sequencing (XR-seq) method to better understand cisplatin DNA damage and repair. We find that transcription-driven repair is nearly complete after 2 days, whereas weeks are required for repair of the NTS and the rest of the genome. TS repair oscillates in rhythmically expressed genes up to 2 days post injection, and in all expressed genes, we see a trend in TS repair with time from the 5' to 3' end. These findings help to understand the circadian- and transcription-dependent and -independent control of repair in response to cisplatin, and should aid in designing cisplatin chemotherapy regimens with improved therapeutic indexes.
Highlights
Cisplatin is the most commonly used chemotherapeutic drug for managing solid tumors
It is possible that platinating anticancer drugs exhibit circadian variations in therapeutic ratio because these drugs kill cells by binding to DNA, excision repair is the sole mechanism for repairing platinated DNA (4 –6), and excision repair of DNA is controlled by the circadian clock
To follow the Pt-d(GpG) repair pattern over a 10-day period, 59 mice were injected with cisplatin (10 mg/kg) at ZT20 (3:00 a.m.) on day 1 (Fig. 1), and for the first 2 days afterward 3 mice were sacrificed at 4-h intervals, their livers were harvested, and Pt-d(GpG) repair was quantified by slot blot and XR-seq
Summary
Cisplatin is the most commonly used chemotherapeutic drug for managing solid tumors. toxicity and the innate or acquired resistance of cancer cells to the drug limit its usefulness. It is possible that platinating anticancer drugs exhibit circadian variations in therapeutic ratio because these drugs kill cells by binding to DNA, excision repair is the sole mechanism for repairing platinated DNA (4 –6), and excision repair of DNA is controlled by the circadian clock To investigate this possibility, we have been using the XR-seq method [7,8,9] which allows us to map, across an entire genome, at single nucleotide resolution, where excision repair has occurred among a population of cells. Basal nucleotide excision repair activity exhibits a single peak at about ZT 8 –10 (ZT ϭ 0 is time of lights on and ZT ϭ 12 is lights off under 12 h light: h dark conditions) which is associated with a corresponding rhythmic oscillation of the XPA excision repair factor [2, 3] This program controls repair of the nontranscribed strand (NTS) of genes, weakly and nontranscribed genes, and intergenic regions. Circadian-controlled genes exhibit peak and trough TS repair levels that occur at times of day specific to each gene, most rhythmic genes exhibit
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