Abstract
Poor local control and tumor escape are of major concern in head-and-neck cancers treated by conventional radiotherapy or hadrontherapy. Reduced glutathione (GSH) is suspected of playing an important role in mechanisms leading to radioresistance, and its depletion should enable oxidative stress insult, thereby modifying the nature of DNA lesions and the subsequent chromosomal changes that potentially lead to tumor escape.This study aimed to highlight the impact of a GSH-depletion strategy (dimethylfumarate, and l-buthionine sulfoximine association) combined with carbon ion or X-ray irradiation on types of DNA lesions (sparse or clustered) and the subsequent transmission of chromosomal changes to the progeny in a radioresistant cell line (SQ20B) expressing a high endogenous GSH content. Results are compared with those of a radiosensitive cell line (SCC61) displaying a low endogenous GSH level.DNA damage measurements (γH2AX/comet assay) demonstrated that a transient GSH depletion in resistant SQ20B cells potentiated the effects of irradiation by initially increasing sparse DNA breaks and oxidative lesions after X-ray irradiation, while carbon ion irradiation enhanced the complexity of clustered oxidative damage. Moreover, residual DNA double-strand breaks were measured whatever the radiation qualities. The nature of the initial DNA lesions and amount of residual DNA damage were similar to those observed in sensitive SCC61 cells after both types of irradiation. Misrepaired or unrepaired lesions may lead to chromosomal changes, estimated in cell progeny by the cytome assay. Both types of irradiation induced aberrations in nondepleted resistant SQ20B and sensitive SCC61 cells. The GSH-depletion strategy prevented the transmission of aberrations (complex rearrangements and chromosome break or loss) in radioresistant SQ20B only when associated with carbon ion irradiation. A GSH-depleting strategy combined with hadrontherapy may thus have considerable advantage in the care of patients, by minimizing genomic instability and improving the local control.
Highlights
Carbon ion hadrontherapy is highly effective for treating cancer located near critical organs at risk that is resistant to conventional radiotherapy, such as head-and-neck squamous cell carcinoma (HNSCC), because a more precise and powerful dose can be applied, leading to a high relative biological efficiency [1]
A 4 h DMF/buthionine sulfoximine (BSO) pretreatment combined with X-ray or carbon ion exposure enabled the stabilization of the GSH depletion and the inhibition of glutathione resynthesis induced after irradiation (Table 1B)
Comet assays were performed in alkaline conditions without (A) or in the presence of formamidopyrimidine glycosylase (Fpg) enzyme (B). n SQ20B cells, mSQ20B+irradiation, SQ20B+GSH depletion, # SQ20B+GSH depletion+irradiation, & SCC61, % SCC61+irradiation. *P,0.05. doi:10.1371/journal.pone.0044367.g003; this was performed to enable a comparison of the nature of DNA damage and the consequences on the transmission of chromosomal changes according to the type of radiation, radioresistance status, and endogenous GSH content
Summary
Carbon ion hadrontherapy is highly effective for treating cancer located near critical organs at risk that is resistant to conventional radiotherapy, such as head-and-neck squamous cell carcinoma (HNSCC), because a more precise and powerful dose can be applied, leading to a high relative biological efficiency [1]. Carbon ions induce detrimental clustered damage comprising a combination of DNA double- and single-strand breaks (DSB and SSB), and abasic sites in the close vicinity of oxidized bases. In contrast to these carbon-ion-induced clustered lesions, X-rays induce rather sparse damage [2]. The growing interest in hadrontherapy for treating highly resistant cancers requires clarifying the impact of complex DNA lesions on the higher incidence of chromosomal changes (CCs) Identifying these processes would be a major advance in the understanding of cancer recurrence, a wellknown feature of radioresistant HNSCC [7,8,9,10]
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