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Review
. 2024 Jul 26;25(15):8192.
doi: 10.3390/ijms25158192.

Accidental Encounter of Repair Intermediates in Alkylated DNA May Lead to Double-Strand Breaks in Resting Cells

Shingo Fujii  1 , Robert P Fuchs  2
Affiliations
Review

Accidental Encounter of Repair Intermediates in Alkylated DNA May Lead to Double-Strand Breaks in Resting Cells

Shingo Fujii et al. Int J Mol Sci. .

Abstract

In clinics, chemotherapy is often combined with surgery and radiation to increase the chances of curing cancers. In the case of glioblastoma (GBM), patients are treated with a combination of radiotherapy and TMZ over several weeks. Despite its common use, the mechanism of action of the alkylating agent TMZ has not been well understood when it comes to its cytotoxic effects in tumor cells that are mostly non-dividing. The cellular response to alkylating DNA damage is operated by an intricate protein network involving multiple DNA repair pathways and numerous checkpoint proteins that are dependent on the type of DNA lesion, the cell type, and the cellular proliferation state. Among the various alkylating damages, researchers have placed a special on O6-methylguanine (O6-mG). Indeed, this lesion is efficiently removed via direct reversal by O6-methylguanine-DNA methyltransferase (MGMT). As the level of MGMT expression was found to be directly correlated with TMZ efficiency, O6-mG was identified as the critical lesion for TMZ mode of action. Initially, the mode of action of TMZ was proposed as follows: when left on the genome, O6-mG lesions form O6-mG: T mispairs during replication as T is preferentially mis-inserted across O6-mG. These O6-mG: T mispairs are recognized and tentatively repaired by a post-replicative mismatched DNA correction system (i.e., the MMR system). There are two models (futile cycle and direct signaling models) to account for the cytotoxic effects of the O6-mG lesions, both depending upon the functional MMR system in replicating cells. Alternatively, to explain the cytotoxic effects of alkylating agents in non-replicating cells, we have proposed a "repair accident model" whose molecular mechanism is dependent upon crosstalk between the MMR and the base excision repair (BER) systems. The accidental encounter between these two repair systems will cause the formation of cytotoxic DNA double-strand breaks (DSBs). In this review, we summarize these non-exclusive models to explain the cytotoxic effects of alkylating agents and discuss potential strategies to improve the clinical use of alkylating agents.

Keywords: DNA damages; DNA double-strand breaks; DNA repairs; chemotherapy; repair accident model.

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Conflict of interest statement

R.P.F. and S.F. are co-founders and consultants at the company, bioHalosis. The company had no role in the design of this study; in the collection, analysis, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results. The authors declare that this research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflicts of interest.

Figures

Figure 1
Figure 1
The MMR-mediated futile cycle model is driven by the persistent presence of the O6-mG lesion on the genomic DNA. Alkylating agents such as temozolomide (TMZ). Widely used in the clinic to treat glioblastomas, alkylating agents induce a broad spectrum of adducts (lesions) on genomic DNA. In the MMR-mediated futile cycle model, the O6-mG lesion among alkylating agent-induced DNA damages is exclusively focused as the only source related to alkylating agent-induced cell death phenomenon. During DNA replication, thymine is preferentially incorporated opposite the O6-mG, forming the O6-mG: T mispair (left panel). This mismatch activates the MMR system. However, the O6-mG lesion remains persistently present in the parental strand, resulting in a novel MMR attempt that is futile by nature (right panel). Such iterative rounds of the MMR repair process could ultimately lead to cell cycle arrest and apoptosis.
Figure 2
Figure 2
The Repair Accident Model. In the case of treatment with an alkylating agent such as TMZ during chemotherapy, a variety of DNA damages appear and are repaired by multiple DNA repair pathways. Indeed, distinct DNA repair systems work independently depending upon their substrate specificities. The BER system mainly acts at the N7-mG or the N3-mA lesion, while the core MMR proteins recognize not only the O6-mG: T base pair but also the O6-mG: C base pair as shown by our work [11]. Therefore, even in non-dividing or quiescent cells treated by alkylating agents; when an N7-mG or an N3-mA lesion is closely located with the O6-mG lesion (e.g., within several hundred nucleotides), the accidental encounter of BER and MMR derived repair intermediates were shown to lead to DSB when they occur within the same time frame. In non-dividing cells treated by an alkylating agent, the “repair accident model” scenario is as follows: when the MMR system recognizes the O6-mG: C base pair, the mechanism of initiation of the MMR reaction is presently unknown. It is likely that the strand discrimination signal is provided by a BER-mediated nick equally likely to occur in either strand. This is in contrast to the situation that occurs during replication where MMR is directed toward the excision of the nascent strand. In any case, Exo1-mediated strand degradation or helicase unwinding assists the DNA gap formation. If, within the same timeframe, the MMR-mediated gap formation process encounters, in the opposite strand, a nick resulting from an intermediate, independent BER repair, a DSB will result. Red square indicates a DNA lesion that is repaired by the BER system. Green arrow shows degradation of a strand by an exonuclease.
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