Abstract

Purpose

The complex relationship between linear energy transfer (LET) and cellular response to radiation is not yet fully elucidated. To better characterize DNA damage after irradiations with therapeutic protons, we monitored formation and disappearance of DNA double-strand breaks (DNA DSB) as a function of LET and time. Comparisons with conventional γ-rays and high LET carbon ions were also performed.

Materials and Methods

In the present work, we performed immunofluorescence-based assay to determine the amount of DNA DSB induced by different LET values along the 62 MeV therapeutic proton Spread out Bragg peak (SOBP) in three cancer cell lines, i.e. HTB140 melanoma, MCF-7 breast adenocarcinoma and HTB177 non-small lung cancer cells. Time dependence of foci formation was followed as well. To determine irradiation positions, corresponding to the desired LET values, numerical simulations were carried out using Geant4 toolkit. We compared γ-H2AX foci persistence after irradiations with protons to that of γ-rays and carbon ions.

Results

With the rise of LET values along the therapeutic proton SOBP, the increase of γ-H2AX foci number is detected in the three cell lines up to the distal end of the SOBP, while there is a decrease on its distal fall-off part. With the prolonged incubation time, the number of foci gradually drops tending to attain the residual level. For the maximum number of DNA DSB, irradiation with protons attain higher level than that of γ-rays. Carbon ions produce more DNA DSB than protons but not substantially. The number of residual foci produced by γ-rays is significantly lower than that of protons and particularly carbon ions. Carbon ions do not produce considerably higher number of foci than protons, as it could be expected due to their physical properties.

Conclusions

In situ visualization of γ-H2AX foci reveal creation of more lesions in the three cell lines by clinically relevant proton SOBP than γ-rays. The lack of significant differences in the number of γ-H2AX foci between the proton and carbon ion-irradiated samples suggests an increased complexity of DNA lesions and slower repair kinetics after carbon ions compared to protons. For all three irradiation types, there is no major difference between the three cell lines shortly after irradiations, while later on, the formation of residual foci starts to express the inherent nature of tested cells, therefore increasing discrepancy between them.

中文翻译：

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癌细胞中 DNA 双链断裂作为质子线性能量转移及其随时间变化的函数

 抽象的

 目的

线性能量转移（LET）和细胞对辐射的反应之间的复杂关系尚未完全阐明。为了更好地表征治疗性质子辐照后的 DNA 损伤，我们监测了 DNA 双链断裂 (DNA DSB) 的形成和消失，作为 LET 和时间的函数。还与传统 γ 射线和高 LET 碳离子进行了比较。

 材料和方法

在目前的工作中，我们进行了基于免疫荧光的测定，以确定三种癌细胞系（即HTB140黑色素瘤、MCF-7乳腺癌）中不同LET值沿62 MeV治疗质子扩散布拉格峰（SOBP）诱导的DNA DSB量腺癌和HTB177非小肺癌细胞。还跟踪了病灶形成的时间依赖性。为了确定与所需 LET 值相对应的照射位置，使用 Geant4 工具包进行了数值模拟。我们将质子照射后的 γ-H2AX 焦点持久性与 γ 射线和碳离子照射后的持久性进行了比较。

 结果

随着 LET 值沿着治疗质子 SOBP 上升，在三个细胞系中检测到 SOBP 远端的 γ-H2AX 焦点数量增加，而其远端衰减部分减少。随着孵育时间的延长，病灶数量逐渐下降，趋于残留水平。对于最大数量的 DNA DSB，质子照射比 γ 射线照射达到更高的水平。碳离子比质子产生更多的 DNA DSB，但数量不多。 γ 射线产生的残余焦点的数量明显低于质子，特别是碳离子。碳离子不会产生比质子多得多的焦点，正如其物理性质所预料的那样。

 结论

γ-H2AX 病灶的原位可视化揭示了临床相关质子 SOBP 比 γ 射线在三种细胞系中产生更多损伤。质子和碳离子辐照样品之间的 γ-H2AX 焦点数量没有显着差异，这表明与质子相比，碳离子辐照后 DNA 损伤的复杂性增加，修复动力学更慢。对于所有三种照射类型，照射后不久，三种细胞系之间没有重大差异，而随后，残留病灶的形成开始表达测试细胞的固有性质，因此增加了它们之间的差异。