Influence of 2-Nitroimidazoles in the Response of FaDu Cells to Ionizing Radiation and Hypoxia/Reoxygenation Stress

Abstract

Cellular adaptations to hypoxia promote resistance to ionizing radiation (IR). This presents a challenge for treatment of head and neck cancer (HNC) that relies heavily on radiotherapy. Standard radiosensitizers often fail to reach diffusion-restricted hypoxic cells, whereas nitroimidazoles (NIs) [such as iodoazomycin arabinofuranoside (IAZA) and fluoroazomycin arabinofuranoside (FAZA)] can preferentially accumulate in hypoxic tumours. Here, we explored if the hypoxia-selective uptake of IAZA and FAZA could be harnessed to make HNC cells (FaDu) susceptible to radiation therapy. Cellular response to treatment was assessed through clonogenic survival assays and by monitoring DNA damage (immunofluorescence staining of DNA damage markers, γ-H2AX and p-53BP1, and by alkaline comet assay). The effects of reoxygenation were studied using the following assays: estimation of nucleoside incorporation to assess DNA synthesis rates, immunofluorescent imaging of chromatin-associated replication protein A as a marker of replication stress, and quantification of reactive oxygen species (ROS). Both IAZA and FAZA sensitized hypoxic HNC cells to IR, albeit the former is a better radiosensitizer. Radiosensitization by these compounds was restricted only to hypoxic cells, with no visible effects under normoxia. IAZA and FAZA impaired cellular adaptation to reoxygenation; high levels of ROS, reduced DNA synthesis capacity, and signs of replication stress were observed in reoxygenated cells. Overall, our data highlight the therapeutic potentials of IAZA and FAZA for targeting hypoxic HNC cells and provide rationale for future preclinical studies.

Optimized Carbohydrate-Based Nanogel Formulation to Sensitize Hypoxic Tumors

“…Worth mentioning here is that Rashed et al have previously reported a higher SER value for IAZA in FaDu cells under hypoxia. 27,28 The discrepancy can be explained by the shorter incubation used in this study (3 h vs 24 h). Additional in vivo studies are required to corroborate the radiosensitization effects seen with nanoIAZA in vitro and to extend the potential use of this drug-delivery system as a hypoxic radiosensitizer.…”

“…A hypoxic tumor model based on FaDu tumor cellsan established human hypopharyngeal squamous cell carcinomawas used according to the study reported by Vaupel et al In a previous study, it was concluded that the oxygenation status of FaDu tumors is heterogeneous and independent of the tumor volume . Chemotoxicity evaluation of IAZA and nano IAZA was assessed in male NU/NU mice bearing FaDu tumors.…”

“…A hypoxic tumor model based on FaDu tumor cells� an established human hypopharyngeal squamous cell carcino-ma�was used according to the study reported by Vaupel et al 70 In a previous study, it was concluded that the oxygenation status of FaDu tumors is heterogeneous and independent of the tumor volume. 28 Chemotoxicity evaluation of IAZA and nanoIAZA was assessed in male NU/NU mice bearing FaDu tumors. We chose the NU/NU strain of mice because (i) it is a well-characterized immune-deficient strain for engraftment of human tumor cell lines, (ii) the hairless phenotype enables more accessible assessment of tumor growth, and (iii) it does not show the radiation hypersensitive phenotype displayed by NSG immunocompromised mice for future radiosensitization studies.…”

“…HAPs, such as 2-nitroimidazoles, offer a molecular approach to manage and target therapy-resistant hypoxic solid tumors. Radiolabeled 2-NIs 17 have shown success as hypoxia-selective radiotracers in clinical settings, and some widely used radiopharmaceuticals for hypoxia imaging include 18 F-fluoromisonidazole ( 18 F-FMISO), 13,17,18 18 F-fluoroazomycin arabinofuranoside ( 18 F-FAZA), 19−22 addition to its diagnostic capacity, 19,20,26 IAZA has shown potential as a hypoxia-directed therapeutic in cell-based assays, either as a radiosensitizer 27,28 to enhance therapeutic effects of external X-ray beam radiotherapy or for delivering in situ molecular radiotherapy (MRT) when IAZA is labeled with a therapeutic radionuclide ( 131 I). Most recently Rashed et al…”

“…HAPs, such as 2-nitroimidazoles, offer a molecular approach to manage and target therapy-resistant hypoxic solid tumors. Radiolabeled 2-NIs have shown success as hypoxia-selective radiotracers in clinical settings, and some widely used radiopharmaceuticals for hypoxia imaging include 18 F-fluoromisonidazole ( 18 F-FMISO), ,, 18 F-fluoroazomycin arabinofuranoside ( 18 F-FAZA), − and 123 I-iodoazomycin arabinofuranoside ( 123 I-IAZA). − In addition to its diagnostic capacity, ,, IAZA has shown potential as a hypoxia-directed therapeutic in cell-based assays, either as a radiosensitizer , to enhance therapeutic effects of external X-ray beam radiotherapy or for delivering in situ molecular radiotherapy (MRT) when IAZA is labeled with a therapeutic radionuclide ( 131 I). Most recently Rashed et al examined the effects of IAZA on tumor hypoxia, with a particular emphasis on identifying the cellular phenotype induced by this drug as well as analyzing their effects on target cellular macromolecules. , Cytostasis was observed in a head and neck (FaDu) cancer model, where hypoxic cells displayed higher sensitivity to IAZA exhibiting an altered cell morphology, compromised DNA replication, slower cell cycle progression, and induction of replication stress .…”