Since radiation therapy remains a primary treatment modality for gliomas, the radioresistance of glioma cells and targets to modify their radiation tolerance are of significant interest. Human apurinic endonuclease 1 (Ape1, Ref-1, APEX, HAP1, AP endo) is a multifunctional protein involved in base excision repair of DNA and a redox-dependent transcriptional co-activator. This study investigated whether there is a direct relationship between Ape1 and radioresistance in glioma cells, employing the human U87 and U251 cell lines. U87 is intrinsically more radioresistant than U251, which is partly attributable to more cycling U251 cells found in G2/M, the most radiosensitive cell stage, while more U87 cells are found in S and G1, the more radioresistant cell stages. But observed radioresistance is also related to Ape1 activity. U87 has higher levels of Ape1 than does U251, as assessed by Western blot and enzyme activity assays (approximately 1.5-2 fold higher in cycling cells, and approximately 10 fold higher at G2/M). A direct relationship was seen in cells transfected with CMV-Ape1 constructs; there was a dose-dependent relationship between increasing Ape1 overexpression and increasing radioresistance. Conversely, knock down by siRNA or by pharmacological down regulation of Ape1 resulted in decreased radioresistance. The inhibitors lucanthone and CRT004876 were employed, the former a thioxanthene previously under clinical evaluation as a radiosensitizer for brain tumors and the latter a more specific Ape1 inhibitor. These data suggest that Ape1 may be a useful target for modifying radiation tolerance.

IL-4-induced gene-1 (Il4i1 or Fig1) initially isolated as a gene of unknown function from mouse B lymphocytes, is limited in expression to primarily immune tissues and genetically maps to a region of susceptibility to autoimmune disease. The predicted Il4i1 protein (IL4I1) sequence is most similar to apoptosis-inducing protein and Apoxin I, both l-amino acid oxidases (LAAO; Enzyme Commission 1.4.3.2). We demonstrate that IL4I1 has unique LAAO properties. IL4I1 has preference for aromatic amino acid substrates, having highest specific activity with phenylalanine. In support of this selectivity, IL4I1 is inhibited by aromatic competitors (benzoic acid and para-aminobenzoic acid), but not by nonaromatic LAAO inhibitors. Il4i1 protein and enzyme activity is found in the insoluble fraction of transient transfections, implying an association with cell membrane and possibly intracellular organelles. Indeed, IL4I1 has the unique property of being most active at acidic pH (pH 4), suggesting it may reside preferentially in lysosomes. IL4I1 is N-linked glycosylated, a requirement for lysosomal localization. Confocal microscopy of cells expressing IL4I1 translationally fused to red fluorescent protein demonstrated that IL4I1 colocalized with GFP targeted to lysosomes and with acriflavine, a green fluorescent dye that is taken up into lysosomes. Thus, IL4I1 is a unique mammalian LAAO targeted to lysosomes, an important subcellular compartment involved in Ag processing.

We report use of PEG-DSPE coated oxidized graphene nanoribbons (O-GNR-PEG-DSPE) as agent for delivery of anti-tumor drug Lucanthone (Luc) into Glioblastoma Multiformae (GBM) cells targeting base excision repair enzyme APE-1 (Apurinic endonuclease-1). Lucanthone, an endonuclease inhibitor of APE-1, was loaded onto O-GNR-PEG-DSPEs using a simple non-covalent method. We found its uptake by GBM cell line U251 exceeding 67% and 60% in APE-1-overexpressing U251, post 24 hours (h). However, their uptake was ~38% and 29% by MCF-7 and rat glial progenitor cells (CG-4), respectively. TEM analysis of U251 showed large aggregates of O-GNR-PEG-DSPE in vesicles. Luc-O-GNR-PEG-DSPE was significantly toxic to U251 but showed little / no toxicity when exposed to MCF-7/CG-4 cells. This differential uptake effect can be exploited to use O-GNR-PEG-DSPEs as a vehicle for Luc delivery to GBM, while reducing nonspecific cytotoxicity to the surrounding healthy tissue. Cell death in U251 was necrotic, probably due to oxidative degradation of APE-1.

Lucanthone and hycanthone are thioxanthenone DNA intercalators used in the 1980s as antitumor agents. Lucanthone is in Phase I clinical trial, whereas hycanthone was pulled out of Phase II trials. Their potential mechanism of action includes DNA intercalation, inhibition of nucleic acid biosyntheses, and inhibition of enzymes like topoisomerases and the dual function base excision repair enzyme apurinic endonuclease 1 (APE1). Lucanthone inhibits the endonuclease activity of APE1, without affecting its redox activity. Our goal was to decipher the precise mechanism of APE1 inhibition as a prerequisite towards development of improved therapeutics that can counteract higher APE1 activity often seen in tumors. The IC50 values for inhibition of APE1 incision of depurinated plasmid DNA by lucanthone and hycanthone were 5 µM and 80 nM, respectively. The KD values (affinity constants) for APE1, as determined by BIACORE binding studies, were 89 nM for lucanthone/10 nM for hycanthone. APE1 structures reveal a hydrophobic pocket where hydrophobic small molecules like thioxanthenones can bind, and our modeling studies confirmed such docking. Circular dichroism spectra uncovered change in the helical structure of APE1 in the presence of lucanthone/hycanthone, and notably, this effect was decreased (Phe266Ala or Phe266Cys or Trp280Leu) or abolished (Phe266Ala/Trp280Ala) when hydrophobic site mutants were employed. Reduced inhibition by lucanthone of the diminished endonuclease activity of hydrophobic mutant proteins (as compared to wild type APE1) supports that binding of lucanthone to the hydrophobic pocket dictates APE1 inhibition. The DNA binding capacity of APE1 was marginally inhibited by lucanthone, and not at all by hycanthone, supporting our hypothesis that thioxanthenones inhibit APE1, predominantly, by direct interaction. Finally, lucanthone-induced degradation was drastically reduced in the presence of short and long lived free radical scavengers, e.g., TRIS and DMSO, suggesting that the mechanism of APE1 breakdown may involve free radical-induced peptide bond cleavage.

In the study cohort, children with CKD did not have increased exposure to BPA and phthalates. Longitudinal studies with repeated measures are likely to be more informative about the possible health effects of prolonged exposure to BPA and phthalates in pediatric patients with CKD.

No abstract

Robust predictive models are essential to manage the risk of radiation-induced carcinogenesis. Chronic exposure to cosmic rays in the context of the complex deep space environment may place astronauts at high cancer risk. To estimate this risk, it is critical to understand how radiation-induced cellular stress impacts cell fate decisions and how this in turn alters the risk of carcinogenesis. Exposure to the heavy ion component of cosmic rays triggers a multitude of cellular changes, depending on the rate of exposure, the type of damage incurred and individual susceptibility. Heterogeneity in dose, dose rate, radiation quality, energy and particle flux contribute to the complexity of risk assessment. To unravel the impact of each of these factors, it is critical to identify sensitive biomarkers that can serve as inputs for robust modeling of individual risk of cancer or other long-term health consequences of exposure. Limitations in sensitivity of biomarkers to dose and dose rate, and the complexity of longitudinal monitoring, are some of the factors that increase uncertainties in the output from risk prediction models. Here, we critically evaluate candidate early and late biomarkers of radiation exposure and discuss their usefulness in predicting cell fate decisions. Some of the biomarkers we have reviewed include complex clustered DNA damage, persistent DNA repair foci, reactive oxygen species, chromosome aberrations and inflammation. Other biomarkers discussed, often assayed for at longer points post exposure, include mutations, chromosome aberrations, reactive oxygen species and telomere length changes. We discuss the relationship of biomarkers to different potential cell fates, including proliferation, apoptosis, senescence, and loss of stemness, which can propagate genomic instability and alter tissue composition and the underlying mRNA signatures that contribute to cell fate decisions. Our goal is to highlight factors that are important in choosing biomarkers and to evaluate the potential for biomarkers to inform models of post exposure cancer risk. Because cellular stress response pathways to space radiation and environmental carcinogens share common nodes, biomarker-driven risk models may be broadly applicable for estimating risks for other carcinogens.

Objective: Studies have documented disparities in exposure to endocrine disrupting chemicals (EDC), but no studies have investigated potential implications for racial/ethnic disparities in chronic disease and associated costs. Our objective was to examine EDC levels in the US population according to race/ethnicity and to quantify disease burden and associated costs.Study Design and Setting: EDC exposure levels in 2007-2010 were obtained from the National Health and Nutrition Examination Surveys. The associated disease burden and costs for twelve exposure-response relationships were determined for Non-Hispanic Whites, Non-Hispanic Blacks, Mexican-Americans, Other Hispanics, and Other/Multicultural.Results: EDC exposure levels and associated burden of disease and costs were higher in Non-Hispanic Blacks ($56.8 billion; 16.5% of total costs) and Mexican-Americans ($50.1 billion; 14.6%) compared to their proportion of the total population (12.6% and 13.5%, respectively). Associated costs among Non-Hispanic Whites comprised 52.3% of total costs ($179.8 billion), though they comprise 66.1% of the US population. These disparities are driven by generally higher exposure to persistent pesticides and flame retardants among Non-Hispanic Blacks and Mexican-Americans.