Co-Localization Analysis Method for High Content Screening (HCS) Measurement of Radiation Induced DNA Damage Response

The cellular response to DNA damage is emerging as a promising target for cancer therapy. In the present study, the authors exploited the relationship between the level of the phosphorylated form of histone H2AX (γH2AX) and the extent of DNA damage and developed a quantitative, cell-based, high-content screening system for measuring the DNA damage response (DDR). In this system, the authors quantified the level of γH2AX by measuring DNA damage–induced γH2AX nuclear foci using an automated cell imager. They found that the total area of γH2AX foci per cell exhibited a good correlation with the concentration of DNA damage–inducing agents, including etoposide. The effects of 2 well-known inhibitors of DNA damage could be quantified using this system, suggesting the suitability of the γH2AX-foci quantification method for large-scale screening applications. This was confirmed by using this method to screen a chemical library; the resulting “hits” included compounds that inhibited early signaling events in DDR, as well those that inhibited subsequent DNA damage repair processes. Overall, this γH2AX foci-measuring system may be an effective screening method for identifying DNA damage response inhibitors that could eventually be used to develop novel anticancer drugs.

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Regulation of DNA Damage Response and Homologous Recombination Repair by microRNA in Human Cells Exposed to Ionizing Radiation

Cancers ◽

10.3390/cancers12071838 ◽

2020 ◽

Vol 12 (7) ◽

pp. 1838

Author(s):

Magdalena Szatkowska ◽

Renata Krupa

Keyword(s):

Dna Damage ◽

Homologous Recombination ◽

Ionizing Radiation ◽

Dna Damage Response ◽

Cell Response ◽

Dna Lesions ◽

Homologous Recombination Repair ◽

Recombination Repair ◽

Damage Response ◽

Radiation Induced

Ionizing radiation may be of both artificial and natural origin and causes cellular damage in living organisms. Radioactive isotopes have been used significantly in cancer therapy for many years. The formation of DNA double-strand breaks (DSBs) is the most dangerous effect of ionizing radiation on the cellular level. After irradiation, cells activate a DNA damage response, the molecular path that determines the fate of the cell. As an important element of this, homologous recombination repair is a crucial pathway for the error-free repair of DNA lesions. All components of DNA damage response are regulated by specific microRNAs. MicroRNAs are single-stranded short noncoding RNAs of 20–25 nt in length. They are directly involved in the regulation of gene expression by repressing translation or by cleaving target mRNA. In the present review, we analyze the biological mechanisms by which miRNAs regulate cell response to ionizing radiation-induced double-stranded breaks with an emphasis on DNA repair by homologous recombination, and its main component, the RAD51 recombinase. On the other hand, we discuss the ability of DNA damage response proteins to launch particular miRNA expression and modulate the course of this process. A full understanding of cell response processes to radiation-induced DNA damage will allow us to develop new and more effective methods of ionizing radiation therapy for cancers, and may help to develop methods for preventing the harmful effects of ionizing radiation on healthy organisms.

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Polymorphisms of DNA damage response genes in radiation-related and sporadic papillary thyroid carcinoma

Endocrine Related Cancer ◽

10.1677/erc-08-0336 ◽

2009 ◽

Vol 16 (2) ◽

pp. 491-503 ◽

Cited By ~ 63

Author(s):

Natallia M Akulevich ◽

Vladimir A Saenko ◽

Tatiana I Rogounovitch ◽

Valentina M Drozd ◽

Eugeny F Lushnikov ◽

...

Keyword(s):

Dna Damage ◽

Papillary Thyroid Carcinoma ◽

Thyroid Carcinoma ◽

Radiation Exposure ◽

Dna Damage Response ◽

Papillary Thyroid ◽

Retrospective Case ◽

Increased Risk ◽

Damage Response ◽

Radiation Induced

Papillary thyroid carcinoma (PTC) etiologically occurs as a radiation-induced or sporadic malignancy. Genetic factors contributing to the susceptibility to either form remain unknown. In this retrospective case–control study, we evaluated possible associations between single-nucleotide polymorphisms (SNPs) in the candidate DNA damage response genes (ATM, XRCC1, TP53, XRCC3, MTF1) and risk of radiation-induced and sporadic PTC. A total of 255 PTC cases (123 Chernobyl radiation-induced and 132 sporadic, all in Caucasians) and 596 healthy controls (198 residents of Chernobyl areas and 398 subjects without history of radiation exposure, all Caucasians) were genotyped. The risk of PTC and SNPs interactions with radiation exposure were assessed by logistic regressions. The ATM G5557A and XRCC1 Arg399Gln polymorphisms, regardless of radiation exposure, associated with a decreased risk of PTC according to the multiplicative and dominant models of inheritance (odds ratio (OR)=0.69, 95% confidence interval (CI) 0.45–0.86 and OR=0.70, 95% CI 0.59–0.93 respectively). The ATM IVS22-77 T>C and TP53 Arg72Pro SNPs interacted with radiation (P=0.04 and P=0.01 respectively). ATM IVS22-77 associated with the increased risk of sporadic PTC (OR=1.84, 95% CI 1.10–3.24) whereas TP53 Arg72Pro correlated with the higher risk of radiogenic PTC (OR=1.80, 95% CI 1.06–2.36). In the analyses of ATM/TP53 (rs1801516/rs664677/rs609429/rs1042522) combinations, the GG/TC/CG/GC genotype strongly associated with radiation-induced PTC (OR=2.10, 95% CI 1.17–3.78). The GG/CC/GG/GG genotype displayed a significantly increased risk for sporadic PTC (OR=3.32, 95% CI 1.57–6.99). The results indicate that polymorphisms of DNA damage response genes may be potential risk modifiers of ionizing radiation-induced or sporadic PTCs.

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