Biomarkers for Biodosimetry and Their Role in Predicting Radiation Injury.

Radiation-related normal tissue injury sustained during cancer radiotherapy or in a radiological or mass casualty nuclear incident is a major health concern. Reducing the risk and mitigating consequences of radiation injury could have a broad impact on cancer patients and citizens. Efforts to discover biomarkers that can determine radiation dose, predict tissue damage, and aid medical triage are underway. Exposure to ionizing radiation causes changes in gene, protein, and metabolite expression that needs to be understood to provide a holistic picture for treating acute and chronic radiation-induced toxicities. We present evidence that both RNA (mRNA, microRNA, long noncoding RNA) and metabolomic assays may provide useful biomarkers of radiation injury. RNA markers may provide information on early pathway alterations after radiation injury that can predict damage and implicate downstream targets for mitigation. In contrast, metabolomics is impacted by changes in epigenetics, genetics, and proteomics and can be considered a downstream marker that incorporates all these changes to provide an assessment of what is currently happening within an organ. We highlight research from the past 10 years to understand how biomarkers may be used to improve personalized medicine in cancer therapy and medical decision-making in mass casualty scenarios.

Microarray analysis of hub genes, non-coding RNAs and pathways in lung after whole body irradiation in a mouse model.

PURPOSE: Previous research has highlighted the impact of radiation damage, with cancer patients developing acute disorders including radiation induced pneumonitis or chronic disorders including pulmonary fibrosis months after radiation therapy ends. We sought to discover biomarkers that predict these injuries and develop treatments that mitigate this damage and improve quality of life. MATERIALS AND METHODS: Six- to eight-week-old female C57BL/6 mice received 1, 2, 4, 8, 12 Gy or sham whole body irradiation. Animals were euthanized 48 h post exposure and lungs removed, snap frozen and underwent RNA isolation. Microarray analysis was performed to determine dysregulation of messenger RNA (mRNA), microRNA (miRNA), and long non-coding RNA (lncRNA) after radiation injury. RESULTS: We observed sustained dysregulation of specific RNA markers including: mRNAs, lncRNAs, and miRNAs across all doses. We also identified significantly upregulated genes that can indicate high dose exposure, including Cpt1c, Pdk4, Gdf15, and Eda2r, which are markers of senescence and fibrosis. Only three miRNAs were significantly dysregulated across all radiation doses: miRNA-142-3p and miRNA-142-5p were downregulated and miRNA-34a-5p was upregulated. IPA analysis predicted inhibition of several molecular pathways with increasing doses of radiation, including: T cell development, Quantity of leukocytes, Quantity of lymphocytes, and Cell viability. CONCLUSIONS: These RNA biomarkers might be highly relevant in the development of treatments and in predicting normal tissue injury in patients undergoing radiation treatment. We are conducting further experiments in our laboratory, which includes a human lung-on-a-chip model, to develop a decision tree model using RNA biomarkers.

Microarray analysis identifies coding and non-coding RNA markers of liver injury in whole body irradiated mice.

Radiation injury from medical, accidental, or intentional sources can induce acute and long-term hepatic dysregulation, fibrosis, and cancer. This long-term hepatic dysregulation decreases quality of life and may lead to death. Our goal in this study is to determine acute changes in biological pathways and discover potential RNA biomarkers predictive of radiation injury. We performed whole transcriptome microarray analysis of mouse liver tissue (C57BL/6 J) 48 h after whole-body irradiation with 1, 2, 4, 8, and 12 Gray to identify significant expression changes in mRNAs, lncRNAs, and miRNAs, We also validated changes in specific RNAs through qRT-PCR. We used Ingenuity Pathway Analysis (IPA) to identify pathways associated with gene expression changes. We observed significant dysregulation of multiple mRNAs across all doses. In contrast, miRNA dysregulation was observed upwards of 2 Gray. The most significantly upregulated mRNAs function as tumor suppressors: Cdkn1a, Phlda3, and Eda2r. The most significantly downregulated mRNAs were involved in hemoglobin synthesis, inflammation, and mitochondrial function including multiple members of Hbb and Hba. The most significantly upregulated miRNA included: miR-34a-5p, miR-3102-5p, and miR-3960, while miR-342-3p, miR-142a-3p, and miR-223-3p were most significantly downregulated. IPA predicted activation of cell cycle checkpoint control pathways and inhibition of pathways relevant to inflammation and erythropoietin. Clarifying expression of mRNA, miRNA and lncRNA at a short time point (48 h) offers insight into potential biomarkers, including radiation markers shared across organs and animal models. This information, once validated in human models, can aid in development of bio-dosimetry biomarkers, and furthers our understanding of acute pathway dysregulation.

Profiling mRNA, miRNA and lncRNA expression changes in endothelial cells in response to increasing doses of ionizing radiation.

Recent and past research have highlighted the importance of the endothelium in the manifestation of radiation injury. Our primary focus is on medical triage and management following whole body or partial-body irradiation. Here we investigated the usability of endothelial cells' radiation response for biodosimetry applications. We profiled the transcriptome in cultured human endothelial cells treated with increasing doses of X-rays. mRNA expression changes were useful 24 h and 72 h post-radiation, microRNA and lncRNA expression changes were useful 72 h after radiation. More mRNA expressions were repressed than induced while more miRNA and lncRNA expressions were induced than repressed. These novel observations imply distinct radiation responsive regulatory mechanisms for coding and non-coding transcripts. It also follows how different RNA species should be explored as biomarkers for different time-points. Radiation-responsive markers which could classify no radiation (i.e., '0 Gy') and dose-differentiating markers were also predicted. IPA analysis showed growth arrest-related processes at 24 h but immune response coordination at the 72 h post-radiation. Collectively, these observations suggest that endothelial cells have a precise dose and time-dependent response to radiation. Further studies in the laboratory are examining if these differences could be captured in the extracellular vesicles released by irradiated endothelial cells.