Survival Fraction at 2 Gy and γH2AX Expression Kinetics in Peripheral Blood Lymphocytes From Cancer Patients: Relationship With Acute Radiation-Induced Toxicities.

PURPOSE Predictive assays for acute radiation toxicities would be clinically relevant in radiation oncology. We prospectively examined the predictive role of the survival fraction at 2 Gy (SF2) and of γH2AX (double-strand break [DSB] DNA marker) expression kinetics in peripheral blood mononuclear cells (PBMCs) from cancer patients before radiation therapy. METHODS AND MATERIALS SF2 was measured with Trypan Blue assay in the PBMCs from 89 cancer patients undergoing radiation therapy at 4 hours (SF2[4h]) and 24 hours (SF2[24h]) after ex vivo irradiation. Using Western blot analysis and band densitometry, we further assessed the expression of γH2AX in PBMC DNA at 0 hours, 30 minutes, and 4 hours (33 patients) and 0 hour, 4 hours, and 24 hours (56 patients), following ex vivo irradiation with 2 Gy. Appropriate ratios were used to characterize each patient, and these were retrospectively correlated with early radiation therapy toxicity grade. RESULTS The SF2(4h) was inversely correlated with the toxicity grade (P=.006). The γH2AX-ratio(30min) (band density of irradiated/non-irradiated cells at 30 minutes) revealed, similarly, a significant inverse association (P=.0001). The DSB DNA repair rate from 30 minutes to 4 hours, calculated as the relative RγH2AX-ratio (γH2AX-ratio(4h)/γH2AX-ratio(30min)) showed a significant direct association with high toxicity grade (P=.01). CONCLUSIONS Our results suggest that SF2 is a significant radiation sensitivity index for patients undergoing radiation therapy. γH2AX Western blot densitometry analysis provided 2 important markers of normal tissue radiation sensitivity. Low γH2AX expression at 30 minutes was linked with high toxicity grade, suggesting that poor γH2AX repair activity within a time frame of 30 minutes after irradiation predicts for poor radiation tolerance. On the other hand, rapid γH2AX content restoration at 4 hours after irradiation, compatible with efficient DSB repair ability, predicts for increased radiation tolerance.

[1]  M. Wannenmacher,et al.  Acute and late toxicity, tumour control and intrinsic radiosensitivity of primary fibroblasts in vitro of patients with advanced head and neck cancer after concomitant boost radiochemotherapy. , 1999, Radiotherapy and oncology : journal of the European Society for Therapeutic Radiology and Oncology.

[2]  M. El-Sebaie,et al.  Relationship between radiosensitivity and normal tissue complications in Saudi cancer patients treated with radiotherapy. , 2004, Journal of the Egyptian National Cancer Institute.

[3]  B. Fertil,et al.  Inherent cellular radiosensitivity as a basic concept for human tumor radiotherapy. , 1981, International journal of radiation oncology, biology, physics.

[4]  M. Koukourakis,et al.  Gamma histone 2AX (γ-H2AX)as a predictive tool in radiation oncology , 2014, Biomarkers : biochemical indicators of exposure, response, and susceptibility to chemicals.

[5]  J H Hendry,et al.  The modelled benefits of individualizing radiotherapy patients' dose using cellular radiosensitivity assays with inherent variability. , 1999, Radiotherapy and oncology : journal of the European Society for Therapeutic Radiology and Oncology.

[6]  C. Parris,et al.  Prolonged expression of the γ-H2AX DNA repair biomarker correlates with excess acute and chronic toxicity from radiotherapy treatment , 2011, International journal of cancer.

[7]  T. Pajak,et al.  Toxicity criteria of the Radiation Therapy Oncology Group (RTOG) and the European Organization for Research and Treatment of Cancer (EORTC) , 1995, International journal of radiation oncology, biology, physics.

[8]  K. Satyamoorthy,et al.  Influence of double-strand break repair on radiation therapy-induced acute skin reactions in breast cancer patients. , 2014, International journal of radiation oncology, biology, physics.

[9]  W A Brock,et al.  Cellular radiosensitivity of primary head and neck squamous cell carcinomas and local tumor control. , 1990, International journal of radiation oncology, biology, physics.

[10]  Z. Darżynkiewicz,et al.  Constitutive histone H2AX phosphorylation on Ser-139 in cells untreated by genotoxic agents is cell-cycle phase specific and attenuated by scavenging reactive oxygen species. , 2006, International journal of oncology.

[11]  D. Fernandes,et al.  DNA double-strand break analysis by γ-H2AX foci: a useful method for determining the overreactors to radiation-induced acute reactions among head-and-neck cancer patients. , 2012, International journal of radiation oncology, biology, physics.

[12]  Steven Eschrich,et al.  Prediction of radiation sensitivity using a gene expression classifier. , 2005, Cancer research.

[13]  Duo Zheng,et al.  Phosphorylation of H2AX at Ser139 and a new phosphorylation site Ser16 by RSK2 decreases H2AX ubiquitination and inhibits cell transformation. , 2011, Cancer research.

[14]  C. Peterson,et al.  Cellular machineries for chromosomal DNA repair. , 2004, Genes & development.

[15]  Michel C. Nussenzweig,et al.  Genomic Instability in Mice Lacking Histone H2AX , 2002, Science.

[16]  L. Stalpers,et al.  Reduced activity of double-strand break repair genes in prostate cancer patients with late normal tissue radiation toxicity. , 2014, International journal of radiation oncology, biology, physics.

[17]  Kristijan Ramadan,et al.  Degradation‐linked ubiquitin signal and proteasome are integral components of DNA double strand break repair: New perspectives for anti‐cancer therapy , 2011, FEBS letters.